FIBIC FuBio JR2 programme report

The main objective of FuBio is to establish, in Finland, globally competitive

knowledge platforms within the field of wood biorefinery R&D for the

renewal of the forest industry and creation of new business.

FuBio is focused on development of novel value chains, in which wood is

refined into especially materials and chemicals.

www.fibic.fi

13Ohjelmatunnukset

Future Biorefinery Joint Research 2

Programme Report 2011-2014

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FUBIO JR2 PROGRAMME REPORT

Copyright Finnish Bioeconomy Cluster FIBIC 2014. All rights reserved.

This publication includes materials protected under copyright law, the copyright for which is held

by FIBIC or a third party. The materials appearing in publications may not be used for commercial

purposes. The contents of publications are the opinion of the writers and do not represent the official

position of FIBIC. FIBIC bears no responsibility for any possible damages arising from their use.

The original source must be mentioned when quoting from the materials.

ISBN 978-952-67969-6-3 (paperback)

ISBN 978-952-67969-7-0 (PDF)

Layout: Brand United Ltd

Printing: Kirjapaino Lönnberg

Coverpage: TOC graphic image from Parviainen, A., King, A. W. T., Mutikainen, I., Hummel, M., Selg, C., Hauru, L. K. J., Sixta, H., Kilpeläinen, I.: Predicting cellulose solvating capabilities of acid base conjugate ionic liquids. ChemSusChem. 2013. 6. 2161-2169. Copyright Wiley-VCH Verlag GmbH & Co. KGaA. Reproduced with permission.Page 32: Photo by Metla / Erkki Oksanen

CONTENT

Foreword ........................................................................................................................................................5

World-leading knowledge platforms for wood-based biorefineries ..........................................6

Introduction ..................................................................................................................................................10

Modelling and techno-economic evaluation of biorefinery concepts .................................... 14

New solutions for biomass fractionation ........................................................................................ 32

Ionic liquids for wood fractionation .................................................................................................... 52

Hydroxy acids (and other acids) from black liquor ....................................................................... 72

Biological effects of wood-based extracts and compounds in models of

human disease ...........................................................................................................................................88

Thermoplastic lignin and reinforcing cellulose fiber composites for advanced

biocomposite applications ................................................................................................................... 104

Biorefinery products ..............................................................................................................................122

FUBIO JR2 PROGRAMME REPORT 5

FOREWORD

The bioeconomy has fast developed as one of the most relevant platforms for business and environmental sustainability today. In parallel, Finland’s centuries-rich forest industry is undergoing radical renewal. Coupling leading-edge wood and paper products expertise with milestone advances in technology, research competence and knowledge, the industry is unlocking vast potential for new products and new integrated processes – and turning its most ambitious targets and concepts into realistic business opportunities.

Along with the renaissance of wood use in construction, design and mechanical industry, environmentally friendly wood-derived chemicals are increasingly replacing conventional oil-based materials. The wood raw material for chemical pulping contains numerous components in addition to cellulose that are currently burned for energy recovery. Intelligent process solutions drawing on new knowledge and Finland’s common logistics, energy and technology platform are now enabling the separation and generation of highly value-added products from this valuable bioresource. These new products can be used to enhance existing paper and board products or applied in new alternative business areas.

The forest industry has been the backbone of the Finnish economy throughout its history. Today, the urgent need to find innovative applications for new forest-based products is two-fold: the decline in the printing and writing paper sector and the fight to find new sustainable business based on Finland’s staple renewable resource – wood. The work done in the FuBio JR2 programme brings these efforts a significant step forward by opening exciting development opportunities and lighting the path towards innovative and sustainable industry renewal.

The five-year EUR 50 million FuBio JR2 programme, a continuation of FuBio 1, represents a dedicated effort and investment by the Finnish Bioeconomy Cluster – joint research company FIBIC Ltd. The programme lays a solid foundation for building competence in bioeconomy research and for business creation. It also significantly enhances networking between industry and the research community as well as joint understanding of the research challenges and needs of the future.

Mika HyryläUPM-Kymmene OyjChairperson, FuBio Joint Research 2 Programme Management Group

FUBIO JR2 PROGRAMME REPORT6

WORLD-LEADINGKNOWLEDGE PLATFORMS

FOR WOOD-BASED BIOREFINERIES

The renewal of the forest industry and creation of new businesses are essential prerequisites for the future success of the Finnish pulp

and paper industry as part of a sustainable, bio-based economy.

In the second FuBio Joint Research programme, companies and research organizations have jointly developed globally competitive knowledge platforms within the field of wood-based biorefinery research and development, by creating new value chains for refining novel materials and chemicals.

A wide range of Finnish forest cluster companies have actively participated in FuBio JR2. In the following chapters the companies highlight the results achieved in the programme.

Great new potential in ionic liquids

In the FuBio Joint Research programmes 1 and 2 over 100 new ionic liquids were synthesized. The best ones show great potential for certain applications studied in another FIBIC programme, FuBio Cellulose.

The FuBio JR 2 research work on ionic liquids is expected to be invaluable for the future of the Finnish forest industry and will probably have a major impact for the renewal of the biorefinery sector. Ionic liquids may even be a game changer for future biorefineries.

Although the majority of the companies that

have participated in the FuBio JR2 programme regard ionic liquids as extremely attractive and promising, they admit that some aspects need more study and development before industrial-scale utilization.

The FuBio programmes have built a competence platform on ionic liquids and led to the creation of key research teams. However, new skills are still needed, and consequently different consortia will launch more focused programmes and projects in the near future, to devise industrial applications.

The development of new ionic liquids has created great potential for new production processes in wood-based value chains. The same knowledge is applicable to other areas as well.

Mikael Hannus, Stora Enso:

“Before we have industrial applications in use,

a lot of development work is needed. In general,

breakthrough technologies are not created

overnight and are developed at high risk.

The Fibic framework is a functional environment

for this kind of research and development.”


Lignin and hemicellulose – raw materials for future bio-based business

The research has given strong indications that residues from pulp and paper production, such as lignin and hemicellulose, will have the greatest impact in the production of novel bio-based products.

The Fubio JR2 programme has created new insight into wood chemistry and promoted broad competence in hemicellulose separation technologies. Specifically it has improved understanding of the phenomena of hot water extraction of hemicelluloses.The applications foreseen are now more realistic than at the beginning of the programme. However, the separation processes will become more feasible when methods have been developed for the utilization of all possible fractions.

One of the main research interests in the FuBio JR2 programme was lignin and its potential as a component in bio-based composites. The properties and production technologies of kraft lignin fibre composites were studied, and the teams were capable of developing and producing prototype products. The demo trials performed by composite manufacturers gave excellent feedback that can be utilized in further development.

Kari Saari, Kemira:

“We regard new sustainable polymeric raw

materials, such as hemicellulose, as potential

future raw materials that can be substitutes for

fossil-based ones in use today.”

Several alternatives for bio-based barrier materials

The Fubio JR2 programme evaluated the applicability of several bio-based alternatives in barrier applications. Alkali-extracted xylan has been identified as one of the new barrier materials with the greatest potential, and its performance has been successfully demonstrated at pilot scale.

As a barrier material xylan has many advantages: its bio-content is quite high, its availability is good because the pulp and paper industry is the main supplier of the raw material, and the production process is workable. The end product has also been shown to have promising converting properties.

The issues that still need to be solved are the barrier capability against oxygen, and deeper cost calculations of the production process, as well as detailed study on implementation in industrial applications.

The programme helped in understanding the possibilities, but also the challenges, in the use of biopolymers. At the same time, it created a framework for performing analyses.

Pirkko Liias, Metsä Fibre:

“The bio-barrier study has generated knowhow

about the possibilities of different derivatives,

helping to decide which ones ought to be ruled

out and which should be taken for further

development.”


Networking created more value

In the work that studied ionic liquids, lignin composites, bio-barriers, mouldable fibre products, new filters or medicinal products, networking created more value than if the research teams had been working alone. All in all, networking is a key attribute of all Fibic programmes.

One of the participants noted that the best outcomes were achieved in the teams that worked openly and dynamically. Dialogue between representatives of the research and industrial partners has been extremely important for the successful results of the programme.

The programme was divided into work packages, each with an industrial partner represented in the steering group and in the technical meetings. This structure ensured that the programme was immediately focused in the right direction. In setting the study, important issues such as regulatory compliance and scale-up of the processes were raised at an early stage.

Mika Hyrylä, UPM-Kymmene:

“The wide international and competent network

also made it possible to test end-products, such

as composites.”

The work goes on

The research of the FuBio JR2 programme was precommercial. In order to achieve industrial applications, further development work is needed. Many companies have already launched internal projects to continue the work.

The techno-commercial models created in the programme broadened the scope of the work with commercial viewpoints, and will act as an excellent tool in further research.

Apart from pulp and paper companies, several technology suppliers participated in the programme. Through the FuBio JR2 knowledge platforms they have been able to find suitable partners.

Jussi Piira, Andritz:

“The programme has provided us with an

outlook for future processes. The new processes

require intensive development of technologies,

which is why it is essential for a technology

supplier to be part of the work as early as

possible.”


INTRODUCTION

The Finnish forest industry has evolved over the centuries through several key phases, spanning from early tar production and sawmilling to modern pulp and paper making. Today, the production of wood-based chemicals – once a focal point of the industry until its eclipse by low-cost, high-performing chemicals and materials of the petrochemical boom – is seeing a resurgence. Biorefinery has now emerged as the next branch in the evolution of end uses for Finland’s abundant forest biomass.

Future Biorefinery (FuBio) is one of the strategic focus areas of the Finnish Bioeconomy Cluster FIBIC (formerly Forestcluster Ltd). The main objective of FuBio research is to establish in Finland globally competitive knowledge platforms for the renewal of the existing forest industry and the creation of new business. The FuBio research path is expected to create new value chains in which biomass-based materials and chemicals are used in substantial, global markets. The potential markets of focus are both well-known to the forest industry (e.g. fibre-based packaging) as well as essentially new, such as textiles, nonwovens, polymers, resins and thermo-formable composites.

FuBio research was initiated in March 2009 with the launch of the first two-year joint research programme. In 2011 FuBio was further split into two separate programmes: ‘Products from Dissolved Cellulose’ (FuBio Cellulose) and, the focus of this programme report, ‘Future Biorefinery Joint Research 2’ (FuBio JR2).

At the time of planning the FuBio programme, national and international attention within the field was almost exclusively focused on biofuels. It was therefore decided after careful analysis to exclude biofuels from the FuBio theme. The largest Finnish research efforts in biorefinery at the time included the BioRefine programme of the Finnish Funding Agency for Innovation (Tekes) and various Technical Research Centre of Finland (VTT) programmes. These were also taken into account and relevant programme interfaces were established.

Creating new biorefinery value chains requires deep understanding of biomass structures. New processing technologies must be developed hand-in-hand with the design of new biorefinery concepts, including their respective value chains. Understanding the markets and having the freedom to-operate in them are essential. The first steps towards future industrial partnerships must also be taken.

"The main objective of FuBio research is to establish

in Finland globally competitive knowledge platforms

for the renewal of the existing forest industry and the

creation of new business."


In addition to the main objectives defined above, FuBio also aimed at achieving a broader national impact by:

• Improvingindustryawarenessofnewwoodbiorefinery opportunities.

• Establishingnewwaysandlevelsofcooperation within R&D in Finland.

• Shorteningthetimefromideatoinnovationthrough effective collaboration.

• Establishingstrategic,internationalR&Dpartnerships.

• MaintainingandimprovingFinnishbiorefinery R&D facilities through lab and early-stage piloting/initial material demos.

• Educatinganewgenerationofresearchers(M.Sc. and D.Sc./PhD) for the biorefinery field.

The scientific publication output of the FuBio programme has increased as a result of FuBio JR2, bringing greater global visibility for the programme and promoting the open innovation strategy chosen for FuBio.

Figure 1. The six target markets of FuBio. Regenerated fibre and chemicals was targeted by the FuBio Cellulose programme, the remaining markets by FuBio JR2.

The globally competitive knowledge platforms targeted by the programme, as mentioned above, specifically include:

• Newbiomassfractionationmethods,especially based on ionic liquids and hot water extraction,

• Advancedseparationtechnologiessuitablefor separation of biomass fractions as well as hydroxy acids from black liquor,

• Chemicalandbiotechnicalmodification/up-grading of biomass fractions to natural polymers, extractives, hydroxy acids, etc.,

• Applicationofwoodfractionsasstructuralcomposites, hydroxy acid products, new board packaging concepts, reactive/active filter materials and health applications,

• Modelling,especiallytechno-economicalevaluation of immature biorefinery concepts,

• Biomasscharacterizationandanalysis,and• Initialprocesspilotingand/ormaterial

prototype production

As a summary, Figure 1 shows the key application areas of the FuBio research theme.


Programme portfolio

FuBio JR2 was divided into eight Work Packages (WPs). The use of wood fractionation alongside a pulp mill (WP1 Hot water extraction) or even to replace a pulp mill (WP2 Novel biomass fractionation) was examined. Possible future products were identified in the area of advanced biocomposites (WP3), packaging (WP4), filters (WP4) and health applications (WP5) as well as for hydroxy acids separated from black liquor (WP6). In modelling and piloting (WP7) the focus was targeted at improving models for new processes and techno-economical evaluation of selected FuBio product cases. The main goals and results of the work packages are presented in the following chapters.

Management of the programme

The FuBio JR2 programme was led by a Management Group (MG) including representatives from industry and academia. The main responsibility for day-to-day execution was with the Programme Manager (PM). Each work package had a WP Leader and one or several Industrial Tutors. The industrial tutors’ role was to bring an industrial perspective to the work packages, while the WP leaders were responsible for day-to-day management of the WP. The work packages were divided into tasks and subtasks, and some of the tasks were assigned specific Task Leaders. The PM and WP leaders formed the execution Core Team.

The main task of the management group was to supervise the progress of the programme with respect to the objectives and the programme plan. The management group followed the work progress of specified issues through a gate system and WP results and achievements through WP presentations in management meetings and through Programme Milestones and Deliverable reports. The WP steering

groups together with the industrial tutors assisted the WP leaders in focussing the work as needed.

The Management Group had the following members:

• MikaHyrylä,UPM,Chairman(EevaJernström until December 2012)

• MikaelHannus,StoraEnso• JohannesHeiskanen,Novoplastik• SeppoHiljanen,Valmet• AnnaleenaKokko,VTT(NiklasvonWeymarn

until August 2012), Programme Manager• MarkkuLeskelä,FIBIC(LarsGäddauntil

April 2012)• PirkkoLiias,MetsäFibre• RistoLilleberg,MetsäliittoGroup• ErkkiPeltonen,Myllykoski(untilSeptember

2011)• JussiPiira,Andritz(PatersonMcKeough

until May 2013)• KariSaari,Kemira• LauriVerkasalo,MetsäBoard(AriKiviranta

until September 2011)• StefanWillför,ÅboAkademiUniversity,

Scientific Coordinator• ErjaÄmmälahti,Tekes

Dissemination of the FuBio JR2 programme results has been achieved through a variety of channels. The main channel for the distribution of results, reports and meeting minutes has been the FIBIC research portal, accessible to FuBio JR2 programme partners, and the FIBIC website (http://fibic.fi/programmes/fubio-jr2-2). Additionally, results have been extensively shared in both external (public) and internal seminars, and demonstration samples and related posters have been displayed at SHOK (Finnish strategic centres for science, technology and innovation) summits and Tekes events, as well as in programme overviews presented at FIBIC seminars and international conferences.


Participants and international cooperation

The three-year joint research programme on future biorefinery had a total of 22 industrial and research partners, bringing multi-faceted knowledge and competence to the programme.

Industrial partners:

• Andritz• FIBIC• Kemira• MetsäliittoGroup• MetsäFibre• MetsäBoard• Myllykoski• Novoplastik• StoraEnso• UPM-Kymmene• Valmet

Research partners:

• AaltoUniversity• LappeenrantaUniversityofTechnology• FinnishForestResearchInstitute,Metla• TampereUniversityofTechnology• UniversityofHelsinki• UniversityofJyväskylä• UniversityofTampere• UniversityofTurku• UniversityofOulu• VTTTechnicalResearchCentreofFinland• ÅboAkademiUniversity

International cooperation in Future Biorefinery Joint Research 2 was achieved through researcher exchange, with more than 15 researchers visiting over a dozen universities and research institutes in Portugal, Venezuela, Sweden, Spain, France, Germany and Austria for more than 40 person-months.

The programme participants have also actively presented the programme results in international conferences, COST action meetings and workshops. International seminars have also been arranged by the programme, especially in WP2 in the area of ionic liquids, which attracted high international interest and was highly commended by the programme partners. In order to demonstrate the increased competence resulting from the programme the main results have been published in peer-reviewed journals, with over 80 publications to date and close to 40 currently submitted for publication. Over 20 Master’s theses and five PhD thesis have been written by students fully or partly financed by FuBio programmes, with a further 10 PhD students expected to defend their thesis within the coming year.

14

MODELLING AND TECHNO-ECONOMIC

EVALUATION OF BIOREFINERY CONCEPTS

CONTACT PERSON – Work Package 7 leader Eemeli Hytönen, [email protected]

Aalto University: Moshood Abdulwahab, Waqar Ahmad, Ville Alopaeus, Kaj Jakobsson,Susanna Kuitunen, Zheng Liu, Kaarlo Nieminen, Juha VisuriAndritz: Paterson McKeoughGloCell: Jari Aittakari, Hanna Kalanne, Juhani Lehtonen, Pirita Mikkanen, Jukka Seppänen Kemira: Marcus Lillandt, Anna-Maija Saariaho Lappeenranta University of Technology: Mari Kallioinen, Elsi Koivula, Jarno Kohonen,Maaret Paakkunainen, Satu-Pia ReinikainenMetsä Fibre: Esko TurunenPöyry Management Consulting: Carina Björnström, Henna Jääskeläinen, Jesse Kautto,Anna Saarentaus, Katja Salmenkivi, Juulia Rouhiainen, Petri VasaraStora Enso: Kalle EkmanUniversity of Oulu: Eva Pongracz, Paula SaavalainenUPM-Kymmene: Mika Hyrylä, Seppo VirtanenValmet: Seppo Hiljanen, Päivi UusitaloVTT Technical Research Centre of Finland: Tuomas Helin, Catharina Hohenthal, Timo Kaljunen, Marjo Kauppi, Marjatta Kleen, Vesa Kunnari, Juha Leppävuori, Marja Nappa, Lotta Sorsamäki

FUBIO JR2 PROGRAMME REPORT


ABSTRACT

In Future Biorefinery Joint Research 2 (FuBio JR2), modelling was used at different levels of biorefinery technology development to support research and to indicate the industrial feasibility of the novel technologies developed in the programme. The focus was on i) phenomena-based modelling and statistical experimental design of hot water extraction of hemicelluloses from wood, ii) early-stage techno-economic modelling of future biorefinery concepts based on the wood fractionation technologies and novel biorefinery products developed in the programme, and iii) process modelling and simulation, and quantitative economic and qualitative opportunity analysis of selected promising future biorefinery concepts.

Two new models were developed for pressurized hot water extraction (PHWE). The new models are more advanced and comprehensive than the previous PHWE models presented in the literature, enabling more detailed analysis of the phenomena involved and optimization of the process.

Several process concepts were designed and evaluated. Conceptual-level techno-economic screening of some 100 concepts as well as more detailed modelling of five process and product development ideas, including hot water extraction, ionic liquids wood fractionation, black liquor hydroxy acids separation, bio-barriers and bio-composites, were conducted. New methods for process integration, market entry and comprehensive risk assessment were also developed and successfully demonstrated along with the evaluation of several selected concepts in detail. The work was done in close collaboration with research and industry partners.

Keywords:biorefinery concepts, modelling, physico-chemical modelling, techno-economic modelling


1. Background

Different knowledge and data are needed at different technology development stages. For example, initial concept analysis requires a systematic approach and modelling of overall production systems, but no detailed data is needed on the process conditions or phenomena involved in the different processing steps. In contrast, when scaling up a process step and designing its process and process equipment, understanding of the specific phenomena involved in that step in the process is needed, whereas modelling of the overall production system is not necessarily required. Therefore the problem statement and scope of modelling varies significantly during the technology development.

Many modelling methods and tools related to the fields of research of the Future Biorefinery Joint Research 2 programme have been previously developed. For example, a number of existing chemical process design methods and simulation tools are applicable to the FuBio JR2 context and these have been combined in earlier FIBIC programmes as a toolbox for quantitative and qualitative concept evaluation for aiding decision making. In addition, simulation tools for modelling pulping and bleaching physico-chemical phenomena have also been developed. These state-of-the-art tools required further modifications to meet the targets of the current programme.

Various process and product ideas were studied and developed in FuBio JR2. To steer future research efforts and estimate the future business potential of these pre-commercial stage R&D ideas, production concepts were designed, screened and evaluated and tools for modelling the phenomena taking place in the studied wood fractionation processes were developed.

2. Objective

The overall objective of the modelling work was to provide more knowledge and data on the technologies developed in the programme in order to support decision making.

The specific goals set for physico-chemical modelling were: 1) develop chip and reactor scale models for pressurized hot water extraction (PHWE), and 2) develop a comprehensive model for PHWE chemistry (especially model the evolution of the hemicelluloses’ molecular weight distribution).

The specific goals set for techno-economic modelling were: 1) to create new biorefinery concepts and evaluate their sustainability, and 2) to develop modelling tools and apply them to support the technology development work.

3. Research Approach

3.1 Physico-chemical modelling of hot water extraction

Chip and reactor scale modelling of PHWEFor the modelling of simplified PHWE chemistry for chip and reactor model development, experimental data from PHWE of coarse birch (Betula pendula) sawdust in a batch reactor was collected. A high liquid-to-wood ratio was selected to minimize limitations on the solubility of the extracted wood components. Experiments were conducted at different temperatures and with different durations. After each experiment the wood residue was collected from the reactor and the yield of the components was measured. Based on this data the appropriate stoichiometry and kinetic parameters of the reactions leading to delignification and condensation of lignin as well as carbohydrate degradation were estimated. A model combining the reaction


Figure 1. Overall techno-economic analysis approach.

kinetics with the diffusion in a wood particle was constructed in Matlab. The differential equation associated with the model was solved numerically using the finite difference method and the PDE tools available in Matlab. The model enables a simulation of the reaction product evolution at chip level during PHWE. Comsol Multiphysics was chosen for simulating the progress of PHWE at the reactor level, as the software includes tools for describing flow and heat transfer in porous media and enables equations for the reaction kinetics to be added.

Comprehensive modelling of PHWE chemistry In order to be able to build the physico-chemical model, understanding was needed of which properties and chemical constituents of wood are relevant to the modelling of hot water extraction as well as how the chemical constituents react and dissolve from wood into the extract in hot water extraction conditions. Both lignin and carbohydrate reactions are considered and, since several reactions are catalysed by hydrogen ions, special attention is paid to accurate modelling of pH evolution.

In the modelling, changes in the molecular weight distribution of the hemicellulose polymers are considered by describing chain lengths as discrete size categories where the rate of change of number concentrations for various polymer chain lengths is calculated.

A high-order numerical method capable of extremely accurate prediction of integral properties of the distribution is applied. Other parameters related to physico-chemical parameters of the components included in the modelling of hot water extraction are then estimated and gathered from public sources.

The aim was to collect as many as possible models and parameters from the literature. Some of the model parameters were regressed using literature and experimental results of the programme research.

3.2 Techno-economic modelling

The overall techno-economic modelling approach used in the programme is illustrated in Figure 1.

To support R&D and to provide preliminary techno-economic performance information for industry regarding the processes and products developed in the programme, various process concepts were created, designed, and evaluated in the screening phase. A method for this early-stage concept screening was developed. The method exploits early-stage process design methods to obtain comparable results for the different overall production systems that are possible for a given concept. The method is illustrated in Figure 2. Prices of all raw materials, chemicals, utilities, and the different factors


used for fixed cost assessment were agreed, and the concepts were created together with the project partners. For capital cost estimation, methods developed by Bridgewater and Zevnik & Buchanan 1 were used.

In screening, concepts were developed from R&D ideas by specifying targeted end products, feedstocks, design capacities, process integration opportunities and main process parameters. Variable and fixed production costs were calculated using parametric models and input-output material and energy balances.

Based on the results of the screening and the progress of the research, together with project partners conceptual designs for five cases were developed. Process modelling was then conducted for these designs: block-flow diagrams were designed or further refined from those designed in the screening phase, and mass and energy balances were modelled using process simulation. The simulation models were parameterized using experimental research results, partners’ expertise and literature. If pulp mill integration was part of the concept, the interface between the processes was defined and pulp mill impacts were simulated using a steady-state process simulation model of the entire system. A pulp mill model developed in the EffFibre programme was used. The technical analysis focused on the main process (production and purification of the main product), and

1 e.g. Holland, F.A. & Wilkinson, J.K. Perry's Chemical Engineers'

Handbook, section 9 (Process Economics), McGraw-Hill, 1999

recycling of the reaction media and catalysts. If enough data was not available, short-cut models and assumptions were used. Preliminary variable production costs were assessed for sensitivity analysis purposes. The resulting process descriptions and balances were further used in quantitative economic analysis and qualitative modelling of the cases.

Net Present Value (NPV) and Return on Investment (ROI) were the main quantitative feasibility analysis instruments used to evaluate the economic feasibility of the cases. An enhanced probability simulation tool was used in the analysis to forecast the probabilities of achieving positive outcomes for NPV and ROI. This method was chosen because the probability of a given outcome can often be more informative than the actual figures themselves, especially at the early development stage.

In the qualitative opportunity assessment many variables were assessed. The variables were divided into two categories: internal and external. The variables were evaluated on a scale from 2 (high) to low (0) opportunity. In addition, different weight factors were given to the variables, reflecting their relative importance. External variables in the analysis mainly included market-related factors such as market size and growth. Economic feasibility was also a key external variable. Internal variables, in turn, mainly included technical variables such as technical feasibility, product quality feasibility and technical availability.

Figure 2. Light techno-economic analysis method developed for concept screening.


4. Results

4.1 Physico-chemical modelling of hot water extraction

Chip and reactor scale modelling of PHWEAs an example reaction definition, Figure 3 shows the reaction scheme for xylan. The data from the experimental treatments indicate that there are two sub-components of xylan (XN1 and XN2) with different reaction rate constants for degradation into oligosaccharides (XOS).

The temperature dependency of the reaction rates is assumed to follow the Arrhenius equation. Table 1 shows the related pre-exponential (frequency) factors and activation energies.

A similar reaction scheme was developed for the degradation of glucan. In that case, the monosaccharide degrades into hydroxymethylfurfural (HMF). A Matlab program and a Comsol multiphysics program, based on estimated reaction rates and values for physical parameters describing diffusion in the wood particle and flow in the porous medium, were constructed to simulate PHWE at chip

Figure 3. Reaction pathways for xylan in PHWE. Notation: XN1 – fast-degrading xylan, XN2 – slowly degrading xylan, XOS – xylo-oligosaccharides, X – xylose, F – furfural, DP1,DP2 – unspecified low molecular weight degradation products, ki – reaction rate constant.

k1 k2 k3 k4 k5

A (min-1) 3.12 ∙ 1017 7.28 ∙ 1017 6.58 ∙ 1012 3.08 ∙ 1012 5.13 ∙ 1016

Ea (kJ mol-1) 161.04 179.54 127.94 129.91 168.52

Table 1. Pre-exponential factors (A) and activation energies (Ea) for xylan reactions.

level and reactor level, respectively. The Matlab program calculates the time development of the 3D distribution over the wood chip of the various reaction products of lignin, xylan and glucan. It is possible to alter the size of the chip, the cooking time and the temperature in the simulation. Likewise, the Comsol program simulates the product distributions in the reactor. The geometry and heating circulation of the simulated reactor can be changed, as can the porosity and permeability of the chip bed.

The main results of the modelling of hot water extraction were increased knowledge of the phenomena taking place in the system, and the simulation tool that can be used in further hot water extraction research.

Comprehensive modelling of PHWE chemistryThe comprehensive PHWE chemistry modelling work provides interesting insights into the phe-nomena taking place during PHWE. A schematic illustration of the various phenomena and reac-tions included in the model is shown in Figure 4.

Considering the onset of the PHWE reactions, the uronic acids seem to be responsible for


introducing hydrogen ions into the fibre-bound liquid, which, when the temperature is raised, begin catalysing various reactions such as deacetylation and hemicellulose degradation. Metal ions, naturally present in the fibre wall, neutralize part of the acidity. In order to accurately estimate the initial hydrogen ion concentration in the fibre wall liquid, it is crucial to know the amounts of all chemical compounds influencing pH. According to our current understanding, it seems that uronic acids and metal ions are the most relevant compounds. Although, based on our modelling results and some evidence provided in the literature, the fibre-bound liquid is acidic, the water external to the wood particles is close to neutral pH. The ion exchange, i.e. the unevenness of the electrolyte composition, in these two liquid phases (fibre-bound and external liquid phase) was modelled using Donnan theory, which is widely used and acknowledged in the context of pulp bleaching and, for example, in studying semipermeable membranes.

As hemicelluloses start to dissolve from the fi-

bre wall, the uronic acids also dissolve. Dissolv-ing uronic acids weaken the ion exchange phe-nomena, and metal ions are also able to move to the external liquid phase as they are no long-er held in the fibre-bound liquid as counter ions for the dissociated uronic acids. Besides acetic acid, uronic acids and metal ions seem to have a significant influence on the evolution of pH in the external liquid phase.

Understanding the behaviour of pH during hot water extraction is essential because hydrogen ions catalyse cleavage of the glycosidic bonds, thus shortening the hemicellulose polymers. Besides predicting the evolution of pH, another novelty of the model is its capability to predict the evolution of the hemicelluloses’ molecular weight distribution during hot water extraction. Furthermore, monomer and other oligomer concentrations are also reproduced. This feature facilitates the optimization of hot water extraction performance with respect to various end products. The amounts of dissolved lignin and furfural, which are potentially problematic for the process, are also predicted by the model.

Figure 4. Schematic of the phenomena included in the comprehensive hot water extraction model.


The chemistry model for PHWE can be used together with the chip and continuous digester models developed in the EffFibre/VIC project. Furthermore, a flow-through reactor model using a simplified chemistry model was also developed. In the chemistry model, only the cleavage of the glycosidic bonds in hemicelluloses was considered. The chemistry model also reproduces the molecular weight distributions of the extracted hemicelluloses.

The developed model enables a priori simulations of the effect of different buffers and additives on pH evolution. Thus, screening work concerning optimal conditions, such as particle size, additives, liquid-to-wood ratio, temperature and duration, can be done with the simulation model.

From the academic point-of-view, this kind of phenomena-based modelling enables testing of different theories in a quantitative manner and reveals shortcomings in the present theories. For industry, the model can be used in process optimization and development.

4.2 Early-stage techno-economic modelling of concepts based on research ideas

Over 100 ideas were evaluated using the method illustrated in Figure 2 (see Table 2 for aspects varied and combined). The concepts and overall systems were defined based on discussions with researchers and industry representatives.

Based on the early-stage techno-economic modelling of all of the process and overall system ideas, the most promising future biorefinery concepts were selected by the industrial partners for further detailed modelling.

4.3 Quantitative and qualitative modelling of selected Future Biorefinery concepts

Five promising overall production systems based on the programme’s technology development ideas were identified and evaluated:

1) Hot water extraction of high molecular

Research topic Examples of concepts

Pressurized hot water extraction (PHWE)

PHWE at a sawmill, kraft pulp mill, TMP plant, or CTMP/soda plant. Extraction of high or low molecular weight hemicelluloses.Extraction of sawdust or chips.

Ionic liquids Kraft pulp to acetate-grade dissolving pulp.Wood to TMP-like pulp.Wood to kraft-like pulp.

Composites Internally, externally or unmodified (using internal plasticization, chemi-cals, enzymes).Different lignin-fibre-plasticizer ratios.

Barriers PHWE-xylan, modification of GGM, TOFA hybrid polymers, fatty acid cellu-lose esters, reactive milling, cellulose-polymer blends.

Hydroxy acids (HA) HA separation technologies (separate & combined): a) Electrodialysis, b) Ion-exchange, c) Chromatography, d) Acidification, e) Cooling crystalliza-tion.Products: a) Hot glues, b) Chelating agents.

Future biorefineries Combinations of the above-listed concepts integrated into the forest in-dustry supply chain.

Table 2. Evaluated concepts from FuBio JR2 process and product development.


weight (MW) hemicellulose from sawdust, 2) Separation of hydroxy acids from kraft black liquor, 3) Biocomposites, 4) Biobarriers, and 5) Future sawdust biorefinery based on hot water extraction.

Case 1 – Hot water extraction of high MW hemicellulose from sawdustFigure 5 illustrates the process concept and the main process parameters and balances for extracting high molecular weight hemicelluloses from spruce sawdust. The production scale was defined based on sawdust availability in Finnish sawmills. The process conditions (temperature, pressure, solid:liquid ratio, flux) and efficiencies (extraction yield, separation and purification efficiencies) were based on experimental results. The balances were simulated using the Balas® process simulator.

Two scenarios were evaluated, in which the extraction yield and extract dry content were varied (Table 3).

The mass and energy balances and the preliminary dimensioning data for the equipment were used in quantitative economic analysis, in which it was clearly seen that the production costs for the material were high, more than double the price of competing materials. One major reason for the high costs is the small design capacity which results in high fixed costs (e.g. personnel and capital costs)

The process is considered to be relatively mature (suitable process equipment exists); however, the overall concept would require further refinement. For example, the yield based on the experimental work of the programme at the time of conducting the study was very low. Moreover, the end product (extract) is not pure high-MW hemicellulose and might not be suitable for high-value applications without further processing.

Base case Optimistic case Unit

Raw extract yield (= dissolved sawdust

fraction led to membrane)

2 10 % on dry

sawdust

Dry content of extract 1.1 5.0 %

Table 3. Scenario definition for high MW hemicellulose extraction.

Figure 5. Block-flow diagram of Case 1: High MW hemicellulose extraction from sawdust.


Case 2 – Separation of hydroxy acids from kraft black liquorFrom the 12 alternative hydroxy acid separation process concepts and overall systems (alternatives listed in Table 2), two were chosen by the industrial partners for detailed analysis. In these two overall systems, the product would be either a mixture of all hydroxy acids, or two mixtures, a high and a low molecular weight mixture targeted at hot glue or hot glue and chelating agent applications, respectively. The process concept included two alternatives, which utilized different technology for liberation of acids from their salt form: ion exchange (IEX) or electrodialysis (ED). The process concept is illustrated in Figure 6.

The main assumptions for the case evaluation were:

•Pulpmillproductioncapacity700000adt/a•Hydroxyacidsproductionrate - 1/3rd of pulp mill black liquor processed

and 15% yield of HA -> production 35 000t/a - Volatile (formic and acetic) acids yield 7%

of pulping raw material•H2S handling and volatile acids recovery

excluded•Pulpmillintegration - Evaluated using WinGems pulp mill model

developed in the EffFibre programme - Cooking variables (effective alkali and

sulfidity) kept constant by NaOH makeup and fly ash purge

The results of modelling the four scenarios were used in quantitative and qualitative evaluation of the case. The economics seem highly promising if acids production is targeted exclusively at glue production, compared to the option of using part of the acids production as chelating agent. However, although the product price is quite low, the product quality currently remains somewhat uncertain.

As process technologies, all of the considered process steps are mature in different contexts. However, processing of kraft black liquor using these separation technologies (chromato-graphic separation, ion exchange, electrodialysis) has been done only at laboratory scale and therefore further technology development is needed. Furthermore, impacts on the pulp mill chemical recovery cycle (Na/S balance) should be minimized and products’ applicability further evaluated. Thus, the overall concept still needs development.

Case 3 – BiocompositesA total of 26 overall biocomposite production systems were analysed using light techno-economic analysis. Of these, two were selected for more detailed evaluation: lignin-fibre composite without (Figure 7 a) and with internal lignin plasticization (Figure 7 b).

The recipes were based on experimental results and optimization for targeted end product properties (composite mechanical properties):

Figure 6. Block-flow diagram of Case 2: Hydroxy acids separation from kraft black liquor.


lignin content 36% and 41%, and fibre content 30% and 20% for the two concepts respectively; the remainder of the composite pellet was plasticizer. Internal plasticization was assumed to be done using purchased acetic acid anhydride, and the excess acetic acid that is recovered after plasticization was assumed to be sold.

Process equipment for the composite production exists, and the composite properties can be further developed/targeted and the most suitable applications found to develop the overall concept.

The process without internal plasticization is simpler and showed better feasibility, but as the composite price range used in the analysis is wide, product quality will ultimately determine the price. Therefore, if internal plasticization enhances the properties, the economics can absorb the cost of internal plasticization. Acetic acid as a side-product also has a significant impact on the overall feasibility.

Case 4 – Biobarriers Two alternative concepts were also selected for detailed analysis of biobarrier production based on light techno-economic analysis and technology development. The two very different concepts considered were the production of hydroxypropylated xylan based and cellulose co-polymer based barriers in fast-food packaging. The process block flow diagrams are illustrated in Figure 8.

In the hydroxypropylated xylan (HPX) case, xylan extraction from kraft pulp is included in the concept to enable efficient alkali recycling. Separation and purification process parameters were based on literature and partner input.

The cellulose co-polymer case was based on the research partner’s definition and literature process parameters.

The cellulose co-polymer product consists mainly of inorganic compounds (sodium and zinc) and its suitability for food packaging

Figure 7. Block-flow diagram of Case 3: Composite concepts: a) no modification of raw materials, b) internal plasticization of lignin.

a)

b)


Figure 8. Block-flow diagram of Case 4: Barrier concepts: a) hydroxypropylated xylan, b) cellulose polymer blend.

a)

b)

applications thus needs careful examination. On the other hand, in the hydroxypropylated xylan case the impacts on kraft pulp properties should be evaluated and possible further valorisation of the pulp after xylan extraction to dissolving pulp considered instead of selling it as kraft pulp. Therefore, in both cases, further technology and concept development is needed.

As the economic feasibility of both cases is dependent on coating layer costs, cost per area is more critical than cost per tonne. Figure 9 shows how the probability of a positive ROI changes as a function of layer cost (c/m2) with different layer amounts.

Case 5 – Future sawdust biorefinery based on hot water extractionThe fifth case was constructed around several project focus areas:

•Hotwaterextractionofhemicellulosesfromsawdust to produce high and lower MW fractions; use of the high MW fraction for barrier production.

•Kraftcookingoftheextractionresiduetoobtain black liquor and pulp; separation of lignin from the black liquor, e.g., for composite production; use of the pulp as fibre for composites, or oxygen delignified and bleached as pulp.


The concept is illustrated in Figure 10.

The main assumptions of the case were:

•100000or20000bdt/asawdust•Processintegratedintoakraftpulpmill (600 000 t/a)•Washedpulptooxygendelignificationstage

of the kraft pulp mill

The concept was modelled using the Balas® process simulator and the integration impacts were simulated using the WinGems pulp mill model developed in the EffFibre programme. The main integration impacts for 20 000bdt sawdust/a capacity would be:

•Pulpfromsawdustcookingincreasespulpproduction by 1.5%

Figure 9. Probability of a positive ROI in the studied barrier cases with different coating amounts as a function of coating layer material costs (cent/m2). Cepe: cellulose co-polymer; HPX: hydroxypropylated xylan. Reference material cost illustrated on x-axis: PE&PP – polyethylene & polypropylene, PET – polyethylene terephthalate, PA – polyamide.

Figure 10. Block-flow diagram of Case 5: FuBio concept with hypothetical production capacity.


Figure 11. Operating cost breakdown of the analysed cases

•1.5%higherfibrelinecapacityrequireddownstream from oxygen delignification

•1.4%increasedwhiteliquordemand•2.2%increasedevaporationrequirementat

evaporation plant•Decreasednetelectricityproduction•Increasedflyashpurge

Four products are generated: two streams of extracted hemicellulose, sawdust-based pulp, and lignin separated from the black liquor. The raw material for the process is sawdust. In this case, the sawdust cost is essential to the economic feasibility of the case.

The overall system requires further development and, due to its small design capacity, process simplifications may be needed, e.g., related to lignin separation. The clearly improved extraction yields and better selectivity in purification compared to Case 1 obtained at the laboratory scale indicate good potential, but the final use of the other fractions requires further development.

SummaryThe cost breakdown of the evaluated cases varies somewhat, as illustrated in Figure 11.

The difference in cost distribution results partly from the selected modelling scope (e.g. whether the feedstock has a purchase cost or not), the scale of production, and the processing type (e.g. whether chemicals are needed or not). These results should not be used to compare economic feasibility between cases, as they only indicate the relative cost distribution. A preliminary cross-case comparison can, however, be made between cases 1 and 5 where PHWE was considered to be one of the main technologies: with higher hemicellulose yield the share of fixed costs decreases (feedstock flow nearly the same in these cases) while, on the other hand, the added lignin separation process requires chemicals.


Figure 12 shows the qualitative analysis results based on the present economic evaluation and assessment of other qualitative analysis variables. Based on their total scores, the studied cases were plotted (x-axis: sum of internal variables; y-axis: sum of external variables) with the level of opportunity increasing from left to right (x-axis) and upwards (y-axis).

Comparing the cases, the level of opportunity was found to be highest in external (market-related) terms for the separation of hydroxy acids from black liquor and for lignin composites. In these two cases, technical challenges related to operation and product quality were, however, found. Technically, the most promising cases were found to be two cases based on hemicellulose fractionation: barrier films based on extraction of xylan from bleached pulp, and hot water extraction of hemicelluloses from sawdust and subsequent sawdust cooking. However, these cases scored relatively low in market-related terms.

Conclusions, risks and suggestions for each of the five cases are listed in Table 4.

5. Exploitation and impact of results

The modelling methods and tools and knowledge developed focusing on hot water extraction phenomena, purification of the hot water extract, and the sustainability of the different process concepts and value chains provides crucial new information for industry decision making and for steering future research.

The thorough modelling of PHWE was based on combined knowledge of aqueous phase thermodynamics, ion exchange, reaction kinetics, and mass transfer. The resulting model can be used in optimization of PHWE conditions to produce either sugars or high molecular weight hemicelluloses, and for

Figure 12. Qualitative opportunity assessment of the studied Fubio JR2 cases.

Fubio Opportunity EvaluationHigh

High

Case 1 – Hot water extraction of high MW hemicellulose from sawdust

Case 2 – Separation of hydroxy acids from Kraft black liqour

Case 3 – Biocomposites

Case 4 – Biobarriers

Case 5 – Future sawdust biorefinery based on hot water extraction

External

InternalLow

Low


Case Conclusion Highest risks Suggestions

1 •Newconceptthatcouldgenerate

additional revenue for a sawmill.

•Verysmallproductionratesandpoor

economic potential.

•Technicalandeconomicfeasibility

(ROI negative in all cases; labour and

capital costs significant).

•Targetconceptswherethecellulose/

lignin fraction has an end-use of

higher added value (compared to

energy use) and hot water extraction

is needed to achieve this.

•Step-wiseextractionwithmultiple

end products could be another

interesting concept (poly-, oligo- and

mono-based).

2 •Targetproductmarketsareexpected

to grow annually 4-6%

•Electrodialysis-basedproduction

concept could have >10% IRR with a

product value of 1000 €/bdt.

•Bothproductionroutesandproduct

alternatives seem to have positive as

well as negative environmental and

social impacts based on preliminary

analyses.

•Technicalfeasibility–experimental

work has been carried out with

soda black liquor, the separation

processes have not been piloted

using black liquor long enough to get

data, e.g., on fouling & cleaning.

•Productquality–application

testing has not been carried out

using obtained acid mixtures as

feedstocks.

•Lookforalternativeion-exchange

system cleaning (organic acid based,

or circulation based) to replace the

high H2SO4 demand (and to remove

high cost of chemicals and fly ash

disposal).

•Conductfurtherproducttesting(hot

melt and chelating agent).

3 •Targetmarketis~2Mt/awith

expected growth of over 5% CAGR;

prices are high on average.

•Unmodifiedlignincaseseemsto

have very good economic return

and internal plasticization case

has potential assuming the quality

improvement obtained from

plasticization is worth over 500 €/t.

•Productquality–strengthproperties

of both composite types should be

tested in the application to verify

their performance.

•Technicalaspect–relativelylarge

share of the product is plasticizer,

which is potentially produced from

food-grade feedstock.

•Conductfurtherproducttestingin

target application.

•Investigateotherpossible

plasticizers and/or the possibility to

decrease its share in the composite.

4 •Thetargetmarketis~1Mt/a(Global)

and 0.4Mt/a (Europe), considering

large fast-food chains. Annual volume

growth expected to be about 5%.

•Celluloseco-polymercaseseemsto

have good economic return due to

very good properties with thin layer

and relatively simple process.

•Hydroxypropylatedxylancase

requires thinner layer thickness with

same properties to become more

attractive.

•Productquality–Celluloseco-

polymer product consists mainly

of inorganic compounds (sodium

and zinc) and the suitability in food

packaging application needs careful

inspection.

•Technicalaspect–in

hydroxypropylated xylan case the

xylan extraction system integration

impacts on pulp mill process and

cellulose product quality.

•Studythecompatibilityofthebarrier

in combinations of different barrier

materials (oxygen, water vapour

barriers).

•Inhydroxypropylatedxylancase,

integrated concept with added

value cellulose product should be

considered.

5 •Combinedsystemshowspromising

economic performance compared to

Case 1 because of improved yield and

integration into pulp mill.

•Ligninseparationsystemisvery

small compared to e.g. announced

Lignoboost projects and requires

significant investment.

•Manyproductsandtechnologies

may be challenging.

•Sawdusthasothercompetinguses.

•Compatibilityofthehotwater

extraction based high molecular

weight hemicelluloses for barrier

application.

•Furtheranalysisofthecompatibility

of the products in the end

applications (barriers, animal feed,

lignin in composite).

•Evaluatewhichproductcombination

generates most value from the

feedstock.

Table 4. Summary of case evaluations.


obtaining information that could be difficult to measure experimentally. The new models were implemented on a digester modelling platform, which can be used for simulation of industrial-scale continuous hot water extraction units. Simulation of a hot water extraction and alkaline pulping sequence for the production of dissolving pulp could be one possible case study. Furthermore, the models for hot water extraction phenomena (incl. xylan degradation kinetics, chain scission and diffusion in a chip, and digester flow) can supplement concept modelling of processes using hot water extraction.

The light techno-economic analysis revealed the potential of the technologies developed in FuBio JR2. For example, the potential of integrating the PHWE system into different host processes, the preliminary economic performance of new hydroxy acids separation

Table 5. Partner organizations and their research roles.

techniques compared to more mature technologies, or the costs of modifying the lignin for biocomposite production. The results of the sustainability analysis on the other hand highlighted the opportunities offered by the selected concepts to different players in the value chain. This new information on business potential can be exploited by companies using the concept and companies developing enabling technologies for the value chain. Moreover, the refined overall techno-economic analysis approach can be utilized for evaluating other new technologies than those developed in FuBioJR2.

6. Networking

A summary of the partners and their contributions in this context is presented in Table 5.

Work package partners Role of the participating organization

Aalto University Physico-chemical modelling of hot water extraction and

implementation of the model into a process simulator, kinetic

modelling of hot water extraction at chip and reactor level

Andritz Work Package Coordinator

GloCell Quantitative analysis in the techno-economic modelling, market

entry evaluation method development

Kemira Industrial tutor

Lappeenranta University of Technology Multivariate analysis

Metsä Fibre Industrial tutor

Pöyry Management Consulting Techno-economic assessment through investment and

production costs and qualitative opportunity assessments

Stora Enso Industrial tutor

University of Oulu Social impact assessment

UPM-Kymmene Industrial tutor

Valmet Industrial tutor

VTT WP leader. Early-stage techno-economic modelling and method

development, new biorefinery process integration method

development, process concept design and process modelling,

life cycle assessment


7. Publications and reports

Abdulwahab, M. Modelling of ionic liquids' ther-mal separation and recycling in biomass frac-tionation, M.Sc. Thesis, Aalto University, 2013.

Borrega, M., Nieminen K. and Sixta, H. Deg-radation kinetics of the main carbohydrates in birch wood during hot water extraction in a batch reactor at elevated temperatures, Biore-source Technology, 102, 2011, 10724-10732.

Hytönen, E. and Leppävuori, J. Future Biore-finery (FuBio) research into process concepts – early stage process evaluation and screening, Nordic Wood Biorefinery Conference NWBC 2014, Stockholm, Sweden, March 25-17, 2014.

Kleen, M. Statistical modelling of pressurized hot water flow-through extraction process, VTT Research Report, VTT-R-2688-14, May 2014.

Kleen, M. Statistical modelling of pressurized hot water batch extraction process, VTT Re-search Report, VTT-R-2637-14, May 2014.

Kleen, M., Pranovich, A. and Willför, S. Statis-tical modeling of pressurized hot water extrac-tion process to produce hemicellulose with de-sired properties, 4th International Conference on Biorefinery—towards Bioenergy (ICBB2013), Xiamen, China, December 3-5, 2013.

Kuitunen, S. Phase and reaction equilib-ria in the modelling of hot water extrac-tion, pulping and bleaching (http://urn.fi/URN:ISBN:978-952-60-5618-0), Doctoral dis-sertation, Aalto University, 2014.

Kuitunen, S., Vuorinen, T. and Alopaeus, V. The role of Donnan effect in kraft liquor impreg-nation and hot water extraction of wood, Holz-forschung, 67, 2013, 511-521.

Liu, Z., Ahmad, W., Kuitunen, S. and Alopaeus, V. Modeling of mass transfer and degradation of hemicelluloses in flow-through hot water ex-traction, Submitted to Industrial & Engineering Chemistry Research.

Liu, Z., Suntio, V., Kuitunen, S., Roininen, J. and Alopaeus, V. Modeling of mass transfer and reactions in anisotropic biomass particles with reduced computational load, Industrial & Engineering Chemistry Research, 53, 2014, 4096 - 4103.

Visuri, J., Song, T., Kuitunen, S., and Alopaeus, V. Model for Degradation of Galactoglucoman-nan in Hot Water Extraction Conditions, Indus-trial & Engineering Chemistry Research, 51, 2012, 10338-10344.


NEW SOLUTIONS FOR BIOMASS

FRACTIONATION

BY PRESSURIZED HOT WATER EXTRACTION, SUPERCRITICAL

WATER TREATMENT AND DELIGNIFICATION

CONTACT PERSON – Work Package 1 leader Risto Korpinen, [email protected]

Aalto University: Fanny Bardot, Marc Borrega, Herbert Sixta, Lasse Tolonen, Yuying Zhang. Andritz: Christian Järnefelt, Tiina RauhalaFinnish Forest Research Institute: Olli Byman, Sanna Hautala, Hannu Ilvesniemi, Petri Kilpeläinen,Veikko Kitunen, Kaisu Leppänen, Zhiqiang Li, Johanna Tanner, Teemu TikkanenKemira: Marcus Lillandt, Anna-Maija Saariaho Lappeenranta University of Technology: Mohammed Al-Manasrah, Mari Kallioinen, Elsi Koivula,Mika Mänttäri, Minna Nevalainen, Tuomas Nevalainen, Liis RetsjaMetsä Fibre: Ismo ReilamaStora Enso: Kalle EkmanUniversity of Helsinki: Maija TenkanenUniversity of Jyväskylä: Raimo Álen, Jarkko Kuivanen, Joni Lehto, Mika LeppäahoUPM-Kymmene: Ulf Hotanen, Mika HyryläVTT Technical Research Centre of Finland: Anne Kallioinen, Marjatta Kleen, Hanna Kyllönen, Tiina Liitiä, Marjo Määttänen, Tarja TamminenÅbo Akademi University: Ricardo Garcia de Castro Insua, Henrik Grénman, Jarl Hemming, Jens Krogell,Zhiqiang Li, Andrey Pranovich, Jan-Erik Raitanen, Jussi Rissanen, Tapio Salmi, Annika Smeds,Maunu Toivari, Stefan Willför, Chunlin Xu


ABSTRACT

Pressurized hot water extraction (PHWE) and subsequent delignification processes were examined for their ability to separate hemicelluloses, lignin and cellulose from wood biomass. In addition, the separation and purification of PHWE extracts and high-temperature hydrothermal treatment of microcrystalline cellulose were also studied.

PHW extraction parameters were successfully tailored to enable the extraction of relatively large amounts of hemicellulose of relatively high molar mass from wood. If molar mass is not considered a critical factor, nearly all hemicelluloses – constituting approximately up to one third of wood biomass – were extractable. Additionally, the pH of the PHWE process was monitored and adjusted as desired by the addition of dilute alkali.

The hemicellulose-rich extracts obtained by PHWE contain mixtures of hemicelluloses of varying chain length. A variety of impurities, such as wood extractives and lignin-derived compounds, are also present. In addition, the dry solids content of the extracts is usually relatively low. It was possible to concentrate, purify and fractionate the extracts according to molecular size by combining appropriate pre- and/or post-treatments with membrane filtration, while maintaining sufficient filtration capacity.

Crystalline cellulose residues were successfully hydrolyzed and dissolved by rapid near- and supercritical water treatments to produce narrowly distributed, low-molar-mass celluloses and cello-oligosaccharides.

The fibrous fraction after PHWE was further isolated as a cellulose-rich fraction by sulfur-free delignification processes. Hardwoods defibrated more readily than softwoods due to differences in lignin structure. The cellulose-rich fraction can be further processed into various products, such as regenerated cellulose.

Keywords:analytical methods, cellulose, delignification, hemicelluloses, lignin, pressurized hot water extraction, pulp, purification, separation, super critical water, wood biomass


1. Background

The structural building blocks of wood – hemicelluloses, lignin and cellulose – account for the vast majority of all woody biomass. Pressurized hot water extraction (PHWE) offers an environmentally sound water-based means of separating out these valuable components. The extracted hemicellulose and lignin fractions offer a source of valuable biochemicals and other bioproducts, thus contributing to reduced reliance on petrochemicals, while the cellulose-rich fibre fraction can be used in composites or further processed into new fibre products or regenerated cellulose.

In the PHWE process, wood is treated with water at elevated pressure and temperature, up to 220 °C, to separate the hemicellulose-rich fraction from the wood matrix. No organic solvents or toxic chemicals are needed. The attained fibre fraction subsequent to PHWE is further fractionated into lignin and cellulose by sulfur-free delignification processes and possible additives.

Pressurized hot water treatment in near- and supercritical conditions, in which distinct liquid and gas phases are absent, can be used to achieve rapid hydrolysis and dissolution of crystalline cellulose residue to produce low molar mass polymer without addition of cellulose solvent.

Isolation of hemicellulose, lignin and cellulose from each other by PHWE is, however, not complete and the obtained fractions thus contain a variety of impurities. In addition, the fractionation processes are relatively water intensive. Therefore, purification and dewatering processes are needed to enable utilization of these fractions in various applications.

2. Objectives

I) Investigation of the use of pressurized hot water extraction and supercritical water treatment for the fractionation of wood. II) Concentration and purification of the obtained hemicellulose- and lignin-containing fractions to enable their utilization in various applications. III) Further delignification of the extracted residues to produce novel pulps. IV) Transfer of knowledge obtained from laboratory scale experiments to the pilot scale.

3. Research approach

Spruce and birch wood were pressurized hot water extracted using different reactor setups (batch, flow-through and cascade). The reactor volumes varied from 33 ml to 300 l. An example of a batch mode setup, accelerated solvent extraction (ASE) system, used in the PHWE experiments can be seen in Figure 1. Different extraction parameters were studied, such as temperature, pressure, time and particle size.

The aim of the extractions was to produce as high-molar-mass hemicelluloses as possible at high yield. The extracted residues were delignified using sulfur-free processes. The aim was to produce pulps suitable for the production of regenerated cellulose and other products. The hemicellulose-rich extracts were fractionated, concentrated and purified using membrane filtration and different purification techniques. Crystalline cellulose residues were hydrolyzed and dissolved by rapid near- and supercritical water treatments in order to produce narrowly distributed, low molecular weight celluloses and cello-oligosaccharides at high purity. Analytical methods needed in the wood fractionation processes were mapped and method comparisons were carried out.


4. Results

4.1 Key results of PHWE

Particle size and temperature had a significant effect on the extraction of hemicelluloses, as illustrated in Figures 2 and 3. Norway spruce sapwood of different particle sizes (0.25–1.0 mm vs. 8–12 mm) was pressurized hot water extracted using different extraction times and temperatures. The smaller particle sizes resulted in a considerably higher amount of total dissolved solids in the extracts. Furthermore, higher extraction temperature resulted in higher hemicellulose yield, but the average molar mass of the hemicelluloses decreased. At Figure 1. Accelerated solvent extractor ASE 350.

Figure 2. Amount of dissolved wood substances in extract as a function of extraction temperature and time, a) sawdust and b) blocks.

a) b)

Figure 3. Average molar mass of ethanol precipitated hemicelluloses from extracts as a function of extraction temperature and time, a) sawdust and b) blocks.

a) b)


higher temperature and prolonged extraction time, more intense hydrolytic degradation of carbohydrates and lignin takes place. Additionally, larger wood particles cause mass transfer limitations, preventing molecules from migrating out from the wood matrix. Monitoring and controlling pH is essential. If the pH drops too low during extraction the hemicellulose chains start to degrade and lower molar mass is obtained. The pH values of the extracts were typically measured at room temperature after removing the samples from the reactor. This resulted in a delay in the data. To avoid this, high-temperature pH electrodes were installed and tested. As Figure 4 shows, the pH measured inside the reactor during PHWE was approximately 0.5 units higher than that measured at room temperature. A difference of 0.5 pH units corresponds to a 3.2 times lower H+ concentration. It can also be seen that by adding dilute alkali and using high-temperature pH electrodes it was possible to adjust and maintain the pH at the desired level.

4.2 Up-scaling experiences of PHWE

Two pilot scale PHWE appliances were used to demonstrate the extraction results obtained from laboratory experiments. Figure 5 shows the 300-litre reactor (flow-through mode) and the 30-litre reactor (both batch and flow-through modes). Due to the large size of these reactors compared to laboratory scale reactors, they were also used to produce sufficient amounts of extracts and PHWE-treated fibres for further processing.

It was shown that the laboratory scale extractions could be up-scaled by a factor of 300 and 6000. The results of birch wood flow-through pressurized hot water extraction at the laboratory scale and pilot scale using the same extraction conditions and reactor dimensions are shown in Table 1.

The amounts and extraction rates of hemicelluloses extracted from birch wood were similar for both the laboratory and pilot scale, as shown in Figure 6. In addition, the pH profile of the extracts was identical.

Figure 4. In-line pH control during PHWE at 170 °C.


Based on the modelling work done in the laboratory using a 100 ml reactor, pilot scale extractions of spruce wood were carried out using a 30-litre reactor (scale-up factor 300)

in batch mode. The obtained model values and the pilot scale results were comparable (Table 2) although the pilot- and laboratory scale PHWE setups had slightly different configurations.

Figure 5. Pilot scale PHWE reactors, 300 l flow-through (a) and 30 l batch and flow-through (b).

a) b)

Laboratory scale (0.05 l) Pilot scale (300 l)

Temperature 160 °C 160 °C

Diameter / length, ratio 34 mm / 59 mm, 0.6 590 mm / 1040 mm, 0.6

Flow rate 3.3 ml/min 20 l/min

Residence time 12 min 12 min

Extraction time 60 min 60 min

Table 1. Laboratory- and pilot scale extraction conditions.

Figure 6. a) pH of the extracts, b) hemicellulose yield and average molar mass, c) cumulative yield of hemicellulose, d) chemical composition of extracts after 60 min extraction. Error bars represent the relative standard deviations of three parallel extractions.

d)a) b) c)


Spruce extractions using batch mode and 30-litre reactor were performed either at 160 °C for 40 min or at 170 °C for 60 min, both with a water-to-wood (W:W) ratio of 5 or 10. After PHWE and removal of the extract, the extracted residue was washed at 50 °C for 30 min using the same W:W as in the extraction. The amount of dissolved material was doubled by using a W:W ratio of 10 instead of 5 in the milder extraction conditions, as seen in Figure 7. In the harsher conditions, about 70% more TDS was obtained when using a W:W ratio of 10. Slightly lower molar mass hemicellulose was obtained when a higher W:W ratio was used.

The amount of dissolved material in the extract could be further increased by compressing the wood material after the extraction and washing stages, as seen in Figure 8. The dissolved material

in the extract was 63 mg/g after PHWE and 118 mg/g after PHWE, washing and compression, representing an 87% increase in TDS. Furthermore, the molar masses of the different washing and compression fractions were maintained.

The pilot scale PHWE experiments showed that it was possible to obtain relatively high-molar-mass hemicelluloses at relatively high yield by using:

• Moderateextractiontemperaturerange(155–165 °C)

• Relativelyshortextractiontime(25–35min)• Highpackingdegreeofwoodinthereactor• Moderatewater-to-woodratio(5–10:1)• Washingandcompressionofthewoodafter

extraction• Secondarywallexposedwoodbymeansof

mechanical treatment

Model values(100 ml)

155 °C, 25 min, W:W 5

Pilot results(30 l)

155 °C, 25 min, W:W 5

Model values(100 ml)

170 °C, 20 min, W:W 10

Pilot results(30 l)

170 °C, 20 min, W:W 10

pH of extract 4.2 3.9 3.7 3.6

Extraction residue yield, % of orig.

94.7 93.2 86.1 82.8

Total dissolved solids (TDS), mg/g wood

67.0 56.5 138.0 161.4

Average Mw, Da 17800 13936 9600 8346

GGM content of extract, mg/g wood

23.8 27.2 67.2 90.5

Table 2. Comparison of values from a statistical model based on laboratory- and pilot scale results.

Figure 7. Total dissolved solids (a) and average molar mass (b) of PHW extracts and wash water from spruce wood.

a) b)


4.3 Near- and supercritical water treatment

Near- and supercritical water treatment rapidly hydrolyzed and dissolved the recalcitrant crystalline cellulose and produced a mixture of cellulose polymers of varying molar mass. The used reactor setup can be seen in Figure 9.

After the treatment, the dissolved products slowly precipitated as low molar mass and highly crystalline cellulose with a narrow molar mass distribution. The rest of the dissolved material remained in solution as oligo- and monosugars and a mixture of various degradation products formed via concomitantly occurring dehydration and retro-aldol reactions. The

shares of the reaction products depended on the treatment time and temperature for the two different microcrystalline celluloses, as seen in Figure 10.

Dissolution as a precipitating polymer was promoted by increasing the temperature, in particular to above 320 °C, providing that the treatment time was kept sufficiently short to prevent extensive depolymerization of the dissolved products. It was observed that wood-derived microcrystalline celluloses (MCCs) exhibited a higher velocity of dissolution than those derived from cotton, possibly due to the dimensional differences of the cellulose crystallites.

Figure 8. Total dissolved solids (a) and average molar mass (b) of PHW extract, wash water and compression water from spruce wood.

Figure 9. “MIKKI” reactor system for near- and supercritical water treatment of microcrystalline cellulose using short reaction times below one second (a). The short treatment time is achieved by rapid heating with preheated supercritical water and quenching with cold water (b).

a) b)

a) b)


Supercritical water treatment has potential for cello-oligosaccharide production with the advantage that cellulose dissolution and hydrolysis can be carried out in a single stage. Other techniques require the use of a cellulose solvent to dissolve cellulose crystallites and heterogeneous hydrolysis, e.g. by acid hydrolysis processes, mainly produces only monosaccharides. Cello-oligosaccharide production was demonstrated by treating commercially available MCC powder in supercritical water at 380 °C for reaction times of 0.2, 0.4 and 0.6 s (Table 3 and Figure 11). Up to 42% yield of cello-oligosaccharides was reached with the 0.4 s treatment.

4.4 Delignification of PHWE-treated fibres

Pressurized hot water extractions of birch wood chips at temperatures between 180 °C and 220 °C were conducted to extract hemicelluloses

and part of the lignin prior to pulping. The intensity of PHWE was described by a modified P-factor (here Log Pxs). Soda-AQ pulping experiments were then conducted at 150 °C, with 22% NaOH and 1% AQ, based on initial dry wood. The yield of main wood components from birch wood after hot water extraction and soda-AQ pulping are shown in Figure 12.

The cellulose content of the pulp remained unaffected up to a hot water extraction intensity (Log Pxs) of about 4.5 (Figure 12 b), but higher intensities led to extensive cleavage of glycosidic bonds, thus facilitating the occurrence of peeling reactions and resulting in a cellulosic pulp with low yield. Nonetheless, unbleached pulps with an acceptable yield (over 30% based on initial dry wood), and containing over 90% cellulose, less than 5% xylan, and about 2–3% lignin, were produced.

Treatment time(s)

Residue(%)

Precipitate(%)

DP2–9 Cello-oligosaccharides

(%)

Monosaccharides (%)

0.2 8 35 29 3.0

0.4 0 11 42 6.1

0.6 0 < 1 30 9.1

Table 3. Yield of undissolved residue, precipitate, cello-oligosaccharides and monosaccharides after a supercritical water treatment at 380 °C and 250 bar for 0.2–0.6 s.

Figure 10. Mass balances of two microcrystalline celluloses prepared by acid hydrolysis from prehydrolysis kraft pulp (a) and cotton linter (b). Microcrystalline celluloses were treated in sub- and supercritical water with varying temperature. Treatment time 0.20 s and pressure 250 bar.

0 %

20 %

40 %

60 %

80 %

100 %

250

260

270

280

290

300

310

320

330

340

350

360

370

380

Temperature (°C)

0 %

20 %

40 %

60 %

80 %

100 %

250

260

270

280

290

300

310

320

330

340

350

360

370

380

Temperature (°C)

OtherWater-soluble sugarsPrecipitateResidue MCC from

prehydrolysis Kraft pulpMCC fromCotton linter

a) b)


Selected wood residues after the pressurized hot water extractions were subjected to SAQ pulping (0.1% AQ) with the addition of carbohydrate stabilization agents sodium borohydride (BH) and anthraquinone-2-sulfonic acid sodium salt (AQS). The addition of 1% AQS had no clear effect on the yield of carbohydrates, whereas the addition of 1% BH resulted in an average yield increase of about 3%, similar to the yield obtained by increasing the AQ charge from 0.1 to 1%. The yield increase was mostly due to stabilization of cellulose, although xylan was also preserved to some extent. The intrinsic viscosity of unbleached pulps derived from low-intensity autohydrolysis (log Pxs < 4.25) was

similar to the viscosity of pulp produced from untreated wood. At higher extraction intensities the viscosity rapidly decreased, reaching a minimum slightly above 100 ml/g. The addition of stabilizing agents against peeling had little effect on pulp viscosity.

The soda-AQ cooks of untreated and PHWE-treated birch sawdust were conducted under varying conditions: i.e., alkali charge 18, 20, and 22% on o.d. feedstock, AQ charge 0.1% on o.d. feedstock, cooking time 90, 120, and 150 min, temperature 170 °C, and liquor-to-feedstock ratio 5 l/kg. Pulp yields of the soda-AQ-cooks are presented in Table 4.

Figure 11. (a): Oligomer concentrations in water solution after supercritical water treatment, analyzed by HPAEC-PAD in aqueous solution (PA100 column). (b): Molar mass distributions of the solid precipitate fraction. Analyzed by GPC-RI system (4 x PL-mixed A columns) after dissolution in anhydrous 90 g/L LiCl/DMAc.

a) b)

Figure 12. Cumulative yields of main birch components in wood residue after hot water extraction (a) and in the pulp after soda-AQ pulping (b), plotted as a function of hot water extraction intensity (Log Pxs).

a) b)


Clearly higher yields were achieved with the reference material (untreated birch sawdust) compared to the pressurized hot water extracted materials. On the other hand, the colour of the pulp produced from PHW extracted feedstocks was clearly lighter (visible difference). Clearly lower kappa numbers were achieved with pulps produced from the PHW extracted materials, as seen in Table 5. As the table shows, the kappa numbers were very low, especially when pulping was conducted with pressurized hot water extracted feedstocks (T, “treated”).

Cooking experiments were also conducted using oxygen-enhanced alkali cooking with untreated and PHW extracted birch and spruce sawdust. The same cooking equipment was used for the oxygen-alkali cooks as for soda-AQ cooks. The cooking conditions were: alkali charge 19% on o.d. feedstock, cooking time 30, 60, 90, 120, and 150 minutes, temperature

170 °C, and liquor-to-wood (L:W) ratio 5 l/kg. The cooking liquor was first bubbled with oxygen (for 5 minutes) and an oxygen atmosphere was then created in the reactors by an oxygen flow. The oxygen-alkali cooking yields for both feedstocks are presented in Table 6.

Again, clearly higher cooking yields were achieved with the reference material (untreated sawdust) when compared to the pressurized hot water extracted materials. In general, PHWE-treatment conducted before oxygen-alkali cooking facilitated the defibration, especially with birch sawdust. However, only very slight improvement could be observed in the case of spruce. Overall, spruce sawdust was very poorly defibrated during oxygen-alkali cooking experiments. Pulp reject after screening (material not passing a 0.2-µm sieve, % of cooking yield) is presented in Table 7.

Soda-AQ cooks of birch wood

18% NaOH 20% NaOH 22% NaOH

Time,

min

Ref PHWE* PHWE** Ref PHWE* PHWE** Ref PHWE* PHWE**

90 51.6 53.5 38.5 50.4 54.0 38.8 49.1 52.8 38.0

120 50.9 53.1 38.1 49.7 53.5 38.5 47.6 51.4 37.0

150 51.1 52.9 38.0 49.1 51.6 37.1 47.8 50.7 36.5

Table 4. Birch soda-AQ cooking yield.

*Cooking yield (% of material charged into the reactors). **Total cooking yield (% of oven-dry feedstock before pre-treatment).

Time, min 18 % NaOH 20 % NaOH 22 % NaOH

90 (NT*)

90 (T*)

14.9

6.4

11.8

5.5

10.2

5.5

120 (NT)

120 (T)

13.3

5.8

11.1

5.4

9.8

5.5

150 (NT)

150 (T)

12.2

5.4

11.0

4.6

9.3

5.6

Table 5. Kappa numbers of the birch soda-AQ pulp samples.

*NT= not treated (i.e., no PHWE), T= treated


4.5 Separation and purification of hemicellulose-rich extracts

Hemicellulose-rich wood extracts after PHWE contain relatively large amounts of water and impurities which need to be removed before further utilization. Membrane filtration is a convenient method for simultaneously concentrating and fractionating the extract. Pilot scale membrane filtration equipment (Figure 13) was used to separate high-molar-mass hemicelluloses from the PHWE extracts.

The results from several concentration filtration experiments performed at the pilot scale revealed that concentration of wood extracts to produce a high-molar-mass hemicellulose fraction can be done using a relatively high filtration capacity (flux) without any pre-treatment when a hydrophilic

Oxygen-alkali cooks

Birch Spruce

19% NaOH 19% NaOH

Time, min Ref PHWE* PHWE** Ref PHWE* PHWE**

30

60

90

120

150

59.5

56.8

55.4

54.1

51.2

59.7

54.7

52.4

52.3

50.0

42.9

39.3

37.9

37.6

36.0

67.9

63.5

60.3

58.3

57.4

74.4

70.3

66.2

63.8

61.3

56.2

53.1

50.0

47.7

46.3

Table 6. Spruce and birch oxygen-alkali cooking yields.

*Cooking yield (% of material charged into the reactor). **Total cooking yield (% of oven-dry feedstock before pre-treatment).

Time (min) Birch (Ref) Birch (PHWE) Spruce (Ref) Spruce (PHWE)

30 70.2 29.9 86.0 82.6

60 68.9 11.5 85.4 79.9

90 64.6 3.8 81.5 76.6

120 57.9 1.7 81.0 75.8

150 47.2 ND 80.7 73.4

Table 7. Pulp reject (% of cooking yield) after oxygen-alkali cooking.

Figure 13. Cross-rotational (CR)-350 ultrafiltration equipment, membrane area 1 m2.

ultrafiltration (10 kDa) regenerated cellulose membrane is used and filtration is performed using a high shear rate filter. High volumes of water and small compounds can be removed from the extract with a reasonable filtration


capacity (> 100 kg/(m2 h)) (Figure 14). The flux gradually decreased as the feed concentration increased, which could be expected. However, at a certain point the flux suddenly collapsed (Figure 14). Changes in the feed composition due to increasing concentration, i.e. increased total solids content and average molar mass of compounds present in the feed, may have led to an increase in osmotic pressure difference across the membrane and increased viscosity. In addition, the rheological properties of the feed may have changed as a result of concentration. These combined factors may have led to the considerable and rapid flux reduction. It was found that the filtration capacity (flux) can be maintained at a good level (about 95 kg/(m2h)) if the pressure is increased in line with the reduction in feed volume (i.e. increase in concentration) at least until the TDS of the concentrated fraction is about 8% (Figure 14). This was demonstrated in the experiments in which spruce extract (TDS content 0.86%) was fractionated in constant flux mode, where the filtration pressure increased from 1.3 bar to

2.5 bar (60 °C, circumferential velocity of rotors 8.6 m/s), indicating that the decrease in filtration capacity occurring at the end of concentration filtration can be compensated by increased filtration pressure. Despite the significant flux decline seen at the end of the concentrate filtrations, the membranes were not significantly fouled and filtration capacity was easy to restore with simple alkaline cleaning.

Several pre-treatments for improving the filtration capacity and increasing the purity of the resulting hemicellulose fractions were investigated. For instance, a combination of pre-treatment, ultrafiltration and post-treatment was studied at laboratory scale using a 40 cm2 RC70PP membrane with a 10 kDa cut-off. Spruce extract was first oxidized and then ultrafiltered and diafiltered at 60 °C and 2 bar. As Figure 15 shows, the combined purification and separation steps removed significant amounts of impurities from the extract, although some hemicellulose losses occurred in the process.

Figure 14. Filtration capacity in the treatment of spruce extract with the RC70PP membrane and the CR-350 filter using constant pressure and constant flux filtration modes. Both experiments were conducted at 60 °C with a rotor circumferential velocity of 8.6 m/s.

0.0

0.5

1.0

1.5

2.0

2.5

3.0

0

20

40

60

80

100

120

140

160

180

200

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5

Pres

sure

(con

stan

t flu

x m

ode)

, bar

Flux

(60°

C),

kg/(m

2 h)

Time, h

Constant pressure (2 bar) Constant flux Pressure at constant flux mode

TDS(feed) 0.86%Constant pressure modeTDS 3.85% at 2.7 h

Constant flux modeTDS 3.79% at 4.8 h


4.6 Analytical methods for compositional and structural analyses

In order to evaluate the efficiency of the developed processes, the composition of the obtained fractions must be analysed. This is important for determining the purity and mass balance of the fractions. In addition to

composition, the structural features of the biomass fractions must also be examined in order to evaluate the applicability and value of the material. The choice of analytical techniques depends on the sample matrix. In addition, appropriate pre-treatments must also be applied. The most important analytical methods for the research challenges described above are

Solid wood and fibrous samples Obtained information

Pre-treatment: grinding and sieving

Carbohydrate content and composition

- Acid methanolysis/GC Hemicelluloses and uronic acids

- Acid hydrolysis (two-stage)-HPAEC/PAD Cellulose and hemicelluloses

Extractives content and composition

- Extraction by ASE, Soxtec, Soxhlet with aceto-

ne, gravimetric detection

Total amount of (acetone soluble) extractives

- Group analysis with short GC column Quantitation of compound groups

- Analysis by GC-FID/GC-MSD Identification of individual components

Lignin amount (after extraction)

- Acid hydrolysis, gravimetric analysis of residue

+ UV detection of the hydrolysate

Total lignin as Klason lignin + acid soluble lignin

Molar mass

- Dissolution (multi-step) in DMAc/LiCl - HPSEC Molar mass distribution of cellulose (+hemicellu-

loses), calculated average values

Table 8. Analysis methods for solid wood and fibrous samples.

ASE: Accelerated Solvent Extraction, FID: Flame Ionisation Detector, GC: Gas Chromatography, HPAEC: High Performance Anion Exchange Chromatography, HPSEC: High Performance Size Exclusion Chromatography, MSD: Mass Spectroscopic Detector, PAD: Pulse Amperometric Detector, UV: Ultraviolet.

Figure 15. Chemical composition of original spruce extract before separation and purification (a) and chemical composition of the concentrate after oxidative pre-treatment, ultrafiltration and diafiltration (b).

b)a)


listed in Tables 8, 9 and 10 for different sample matrices, including a brief description of the method and the information obtained. The methods suggested for solid wood and fibrous samples are based on either standard methods or literature.

Molar mass is one of the most crucial criteria for determining hemicellulose quality. Lignin is always present in the hemicellulose fractions either as such or linked to carbohydrates. This causes problems in molar mass analysis using universal calibration with light scattering-based detection due to autofluorescence. Because of the challenges related to this critical measurement, a comparison between several methodologies was performed using hot-water extracted spruce galactoglucomannans (GGM) and birch glucuronoxylans as substrates (Xu et al. 2013). Four different size exclusion chromatography (SEC) configurations used by different partners were compared. Ethanol precipitations were carried out on the birch and spruce extracts to explore the effect of lignin content on molar mass.

The difference in Mw values of the birch samples was found to be relatively larger than that of the spruce samples, presumably because the birch extracts contained more lignin than spruce. The difference in Mw values before and after ethanol precipitation was similarly ascribed to differences in lignin content of the birch and spruce samples. In addition, the Mw values determined by RI increased during the precipitation–dissolution cycle, probably due to aggregation. It was therefore not possible to identify an optimal method for all sample types.

Another analytical challenge related to hemicelluloses is the identification and quantitation of individual oligosaccharides, either in their native form or in altered form during processing. By combining several chromatographic and mass spectroscopic

techniques, it was possible to follow the formation of cello-oligosaccharides from cellulose under supercritical water treatment conditions (Tolonen et al. 2014). The other methods mentioned in Table 9 are based on literature.

Lignin was partly extracted in the PHWE treatments along with hemicelluloses as an impurity. However, lignin is a valuable component in itself with a range of potential applications. The structure and properties of lignin depend on its origin (raw material and process). The methods listed in Table 10 provide useful information for evaluating the quality and properties of isolated lignins.

5. Exploitation plan and impact of results

The main objective was to generate a concept based on pressurized hot water extraction (PHWE) for producing high-molar-mass hemicelluloses at high yield from wood of varying particle size. Another goal was to utilize the remaining fibre fraction after PHWE for the production of new fibre products and regenerated cellulose. Because no entirely pure fractions can be separated from wood, comprehensive studies of separation and purification technologies required for these processes were also carried out. Furthermore, fundamental understanding of the factors affecting PHWE and subsequent delignification processes is needed for developing industrial-scale processes. These general objectives were met and the processes and knowledge generated can be implemented, at least in part, in industry. As a result, increased use of biomass instead of oil for materials and products will have a positive impact on society and contribute towards a sustainable future.


PHWE extracts and isolated hemicelluloses Obtained information

Solids content, gravimetric Total amount of dissolved material

Carbohydrate content and composition

- Freeze-drying-acid methanolysis/GC Hemicelluloses and uronic acids

- Weak acid hydrolysis-HPAEC/PAD Hemicelluloses

Lignin amount

- Acidic solutions: direct UV detection or freeze-drying - acetyl bromide or Klason lignin

Content of dissolved lignin

Acetic acid / formic acid, degree of acetylation

- HPLC, CE or benzylation-GC/MS Content of volatile acids

- Alkaline hydrolysis + analysis of acetic acid Degree of acetylation

Furfural-type degradation products

- HPLC or CE Content of furfural and hydroxymethylfurfural (HMF)

Other degradation products

- Silylation-GC/MS (Hydroxy)acids

Molar mass distribution of dissolved hemi and lignin

- HPSEC-various detectors* Molar mass distribution and calculated average values

Oligosaccharides

- HPAEC-PAD/MSQ** Quantitative analysis of oligosaccharides if standards available, otherwise tentative identification by MSQ (as deacetylated analogues)

- MALDI-TOF-MS, AP-MALDI-MS/MS, ESI-MS/MS** Identification including native acetylation

Table 9. Analysis methods for PHWE extracts and isolated hemicelluloses.

AP: Atmospheric Pressure, CE: Capillary Electrophoresis, ESI: Electron Spray Ionisation, HPLC: High Performance Liquid Chromatography, MALDI: Matrix Assisted Laser Desorption Ionisation, MS: Mass Spectrometry, MSQ: Quadrupole Mass Spectrometry, TOF: Time of Flight, other abbreviations as in Table 8. * For method comparison, see text above and ref. Xu et al. 2013. ** For application example and details, see text above and ref. Tolonen et al. 2014.

Isolated lignin Obtained information

Composition

- Ash content (metals) by incineration Organic content

- Carbohydrate content: hydrolysis and HPAEC/PAD Lignin-Carbohydrate Complexes (LCC)

Structure

- Derivatisation-31P NMR Frequency of hydroxyl functionalities

- 1D and 2D NMR techniques Substructures and inter-unit linkages

- Pyrolysis-GC/MS Fingerprint, mainly for sample comparison

Molar mass

- HPSEC-UV in NaOH Molar mass distribution

Elemental analysis (CHONS) Elemental composition

Thermal properties

- Glass transition temperature by DSC, degradation

by TGA

Transitions and degradation as a function of tempe-

rature

Table 10. Analysis methods for isolated lignin.

CHONS: Carbon, Hydrogen, Oxygen, Nitrogen and Sulfur, DSC: Differential Scanning Calorimetry, NMR: Nuclear Magnetic Resonance, TGA: Thermogravimetric Analysis, other abbreviations as in Tables 8 and 9.


6. Networking

The research was carried out jointly by Aalto University, the Finnish Forest Research Institute, Lappeenranta University of Technology, University of Helsinki, University of Jyväskylä, VTT Technical ResearchCentreofFinland,ÅboAkademiUniversityandFinnishBioeconomyClustercompanies.Table 11 presents the research partners and their roles. Table 11. Partner organizations and their research roles.


Aalto University Pressurized hot water extraction, near- and

supercritical water treatment, delignification

Andritz Industrial tutor

Finnish Forest Research Institute Pressurized hot water extraction, purification,

delignification

Kemira Work Package Coordinator

Lappeenranta University of Technology Membrane filtration, separation, purification



University of Helsinki Enzymatic hydrolysis, analytical methods

University of Jyväskylä Delignification, characterization of black liquor


VTT Pressurized hot water extraction, analytical

methods, enzymatic hydrolysis, modelling,

delignification, green liquor extraction

Åbo Akademi University WP leader. Pressurized hot water extraction,

analytical methods, purification, reaction

kinetics, hydrolysis by heterogeneous

catalysis, chemical characterization,

delignification



Bardot, F. Cellulose stabilization during alkaline pulping for the production of high‐purity dis-solving‐grade pulp, Master’s thesis, Aalto Uni-versity, 2012.

Borrega, M., Tolonen, L.K., Bardot, F., Testova, L. and Sixta, H. Potential of hot water extrac-tion of birch wood to produce high‐purity dis-solving pulp after alkaline pulping, Bioresource Technology, 135, 2012, 665–671, DOI: 10.1016/j.biortech.2012.11.107.

Borrega, M., Niemelä, K. and Sixta, H. Effect of hydrothermal treatment intensity on the for-mation of degradation products from birch-wood, Holzforschung, 67, 2013, 8, 871–879, DOI: 10.1515/hf-2013-0019.

Borrega, M. and Sixta, H. Purification of cel-lulosic pulp by hot water extraction, Cellulose, 20, 2013, 2803–2812, DOI 10.1007/s10570-013-0086-1.

Garcia de Castro Insua, R. Acid hydrolysis of birch and spruce hemicelluloses by heteroge-neouscatalysis,Master’sthesis,ÅboAkademiUniversity, 2013.

Grénman, H., Eränen, K., Krogell, J., Willför, S., Salmi, T. and Murzin, D. Kinetics of Aqueous Extraction of Hemicelluloses from Spruce in an Intensified Reactor System, Ind. Eng. Chem. Res., 50, 2011, 7, 3818–3828.

Kilpeläinen, P., Leppänen, K., Spetz, P., Ki-tunen, V., Ilvesniemi, H., Pranovich, A. and Willför, S. Pressurised hot water extraction of acetylated xylan from birch sawdust, Nord. Pulp Pap. Res. J., 27, 2012, 4, 680–688.

Kilpeläinen, P., Kitunen, V., Pranovich, A., Il-vesniemi, H. and Willför, S. Pressurized hot water flow-through extraction of birch saw-dust with acetate pH buffer, BioRes., 8, 2013, 4, 5202–5218.

Kilpeläinen, P., Hautala, S. Byman, O., Tan-ner, J., Korpinen, R., Lillandt, M., Pranovich, A., Kitunen, V., Willför, S. and Ilvesniemi, H. Pressurized hot water flow-through extraction system scale up from laboratory to pilot scale, Green Chem., 2014, DOI: 10.1039/C4GC00274A.

Koivula, E., Kallioinen, M., Sainio, T., Antón, F.E., Luque, S. and Mänttäri, M. Enhanced membrane filtration of wood hydrolysates for hemicelluloses recovery by pretreatment with polymeric adsorbents, Biores. Tech., 143, 2013, 275-281.

Korpinen, R., Kallioinen, M., Hemming, J., Pranovich, A., Mänttäri, M. and Willför, S. Comparative evaluation of various lignin deter-mination methods on hemicellulose-rich frac-tions of spruce and birch obtained by pres-surized hot-water extraction (PHWE) and subsequent ultrafiltration (UF), Holzforschung, 2014, DOI: 10.1515/hf-2013-0233.

Krogell, J., Eränen, K., Granholm, K., Pranovich, A. and Willför, S. High-temperature pH measuring during hot-water extraction of hemicelluloses from wood, Industrial Crops and Products, 61, 2014, 9–15.

Krogell, J., Korotkova, E., Eränen, K., Pranovich, A., Salmi, T., Murzin, D. and Willför, S. Intensification of hemicellulose hot-water extraction from spruce wood in a batch extrac-tor - Effects of wood particle size, Bioresource Technology, 143, 2013, 212–220.


Kuivanen, J. Kuusen ja koivun jatkuvavirtauk-sellinen paineistettu kuumavesiuutto, Master’s thesis, University of Jyväskylä, 2012.

Lehto, J. and Alén, R. Purification of hardwood-derived autohydrolysates, BioResources, 7, 2012, 1813–1823.

Lehto, J., Alén, R. and Malkavaara, P. Multivar-iate correlation between analysis data on dis-solved organic material from Scots pine (Pinus sylvestris) chips and their autohydrolysis pre-treatment conditions, BioResources, 9, 2014, 93–104.

Lehto, J., Alén, R., and Malkavaara, P. Multi-variate correlation between analytical data for various organics dissolved during autohydrol-ysis of silver birch (Betula pendula) chips and treatment conditions, BioResources, 9, 2014, 4958–4970.

Leppäaho, M. Kuumavesiuutetun kuusisahan-purun sooda-antrakinonikeitto, Master’s thesis, University of Jyväskylä, 2013.

Li, Z. Optimisation of alkaline flow-through ex-traction of spruce wood for lignin recovery, Master’sthesis,ÅboAkademiUniversity,2013.

Nevalainen, M. Ultrasuodatusmembraanin mo-difiointi entsyymeillä suodatusominaisuuksien parantamiseksi, Master’s thesis, Lappeenranta University of Technology, 2013.

Penttilä, P.A., Kilpeläinen, P., Tolonen, L., Su-uronen, J.-P., Sixta, H., Willför, S. and Serimaa, R. Effects of pressurized hot water extraction on the nanoscale structure of birch sawdust, Cellulose, 20, 2013, 5, 2335–2347.

Retsja, L. Puuperäisten biomassahydrolysaat-tien likaamien membraanien pesu, Master’s the-sis, Lappeenranta University of Technology, 2013.

Rissanen, J., Grénman, H., Willför, S., Mur-zin, D. and Salmi, T. Spruce Hemicellulose for Chemicals Using Aqueous Extraction: Kinet-ics, Mass Transfer, and Modeling, Industrial & Engineering Chemistry Research, 53, 2014, 15, 6341–6350.

Song, T. Extraction of polymeric galacto-glu-comannans from spruce wood by pressurised hotwater,Doctoralthesis,ÅboAkademiUni-versity, 2013.

Testova, L., Borrega, M., Tolonen, L.K., Pent-tilä, P.A., Serimaa, R., Larsson, P.T. and Sixta, H. Dissolving-grade birch pulps produced un-der various prehydrolysis intensities: quality, structure and applications, Cellulose, 2014, DOI 10.1007/s10570-014-0182-x.

Tikkanen, T. Paineistetulla kuumavesiuutolla tuotetun hemiselluloosarikkaan jakeen entsy-maattinen puhdistaminen, Master’s thesis, Uni-versity of Oulu, 2013.

Tolonen, L.K., Penttilä, P.A., Serimaa, R., Kruse, A. and Sixta, H. The swelling and disso-lution of cellulose crystallites in subcritical and supercritical water, Cellulose, 20, 2013, 6, 2731-2744. doi:10.1007/s10570-013-0072-7.

Tolonen, L.K., Juvonen, M., Mikkelson, A., Nie-melä, K., Tenkanen, M. and Sixta, H. Super-critical water treatment for cello-oligosaccha-ride production from microcrystalline cellulose, Manuscript submitted to ChemSusChem 2014.

Xu, C., Korpinen, R., Hirsilä, P., Liitiä, T., Tuo-mainen, P., Tamminen, T., Willför, S. and Ten-kanen, M. Effect of lignin on the molar mass analysis of hot water extracted hemicelluloses, FuBio Seminar, 27.8.2013, Helsinki, Finland.


IONIC LIQUIDS FOR WOOD

FRACTIONATION

CONTACT PERSON – Work Package 2 leader Alistair W. T. King, [email protected]

Aalto University: Ville Alopaeus, Lauri Hauru, Michael Hummel, Yibo Ma, Alexandr Ostonen, Annariikka Roselli, Erlin Sapei, Herbert Sixta, Petri Uusi-Kyyny Andritz: Christian Järnefelt, Tiina Rauhala Lappeenranta University of Technology: Mika Mänttäri, Olli Nakari, Arto PihlajamäkiMetsä Fibre: Pirkko Liias Stora Enso: Paula Rantamäki, Heidi Saxell University of Helsinki: Jussi Helminen, Ashley Holding, Uula Hyväkkö, Tia Kakko, Ilkka Kilpeläinen, Arno Parviainen UPM-Kymmene: Ulf Hotanen, Mika Hyrylä VTT Technical Research Centre of Finland: Anna Suurnäkki, Ronny Wahlström Åbo Akademi University: Ikenna Anugwom, Valeri Eta, Jyri-Pekka Mikkola, Päivi Mäki-Arvela,Pasi Virtanen


ABSTRACT

During FuBio 1, new concepts in wood processing with novel ionic liquids as potentially environmentally benign reaction media were presented. These include a refinement in our understanding about the effects that these unique solvators have on woody material and examples of the new structures under development, which offer increased process sustainability over the initial generations of ionic liquids.

Herein we present the updated results from these fractionation concepts, including switchable ionic liquid (SIL, selective extraction of components) fractionation and ionic liquid-aided fractionation (ILAF, homogeneous dissolution and selective precipitation of components). One new concept is presented, called the IONCELL-P process, which is a process for fractionation of kraft pulp into its higher-value pure components (cellulose and hemicellulose). As ionic liquids must be recovered in all processes, we also present the most up-to-date recyclable ionic liquid structures that have been developed during the FuBio project (switchable: SIL; distillable: DIL; phase-separable: PSIL) and the methods that are being used to assess ionic liquid purification (thermodynamics of distillation in ionic liquids and membrane purification methods).

Overall, we have developed understanding that makes us highly competitive on the international scale in the area of bioprocessing with ionic liquids.

Keywords:fractionation, dissolution, biomass, distillable, switchable, sustainable, IONCELL


1. Background

1.1 Ionic Liquid Bioprocess Sustainability

Wood is regarded as the number one sustainable resource to replace fossil fuels. However, challenges exist in the incorporation of wood-based feedstocks and processes into traditionally fossil-based value chains. Therefore, new ‘tuneable’ methods offering more varied and improved selectivities for the fractionation and processing of wood biopolymers into materials, chemicals and energy are necessitated. The reported high efficiency in the solvation of cellulose, as a major wood component, by ionic liquids (ILs) has thus afforded new processing opportunities whereby wood itself can be effectively solvated and processed accordingly. In addition, room temperature ionic liquids (RTILs), such as 1-ethyl-3-methylimidazolium acetate ([emim][OAc], m.p. -45°C), offer so effective cellulose solvation capabilities that they are now considered to be industrially viable media for existing and novel cellulose processing applications. An important example is in the replacement of N-methylmorpholine-N-oxide hydrate (NMMO∙H2O) in a ‘Lyocell’ process, circumventing hazardous thermal stability issues. [emim][OAc] has been so successful in cellulose solvation that BASF is now producing it on a ton-scale. Publications are appearing, however, that highlight the instability of [emim][OAc] in the presence of lignocellulosic solutes, although this may be beneficial for homogeneous dissolution of wood as certain [emim][OAc] decomposition products are known to catalyse certain chemistries. In regard to process sustainability, basic ILs such as [emim][OAc] are also known to have reduced thermal stabilities. This effectively prevents the recovery of the IL on an industrial-scale by distillation. As such, other methods of recycling or more recyclable structures/systems were sought during FuBio 1 & 2, e.g., switchable

ionic liquids (SILs), distillable ionic liquids (DILs) and phase-separable ionic liquids (PSILs). The results from fractionation of wood and subsequent recycling, using all classes of ILs described above, are presented in this chapter.

2. Objectives

The main application objectives of this work package were focussed on the development of processes that utilize ionic liquids, namely:

• Developrecyclable,cheapandlowtoxicityionic liquids that are capable of dissolving cellulose, wood or extracting wood.

• Assesstheirpotentialforfractionationof wood into its components of specified quality for novel processes or to fit into existing value chains.

• Assesstherecyclabilityandpurificationof these ionic liquids before and after fractionation steps.

On the whole, the objectives called for the assessment of existing structures for their efficiency in the fractionation of woody material and for the development of new ionic liquids. This was necessary due to the poor stability and recyclability of existing structures. A general pre-requisite for new ionic liquids for the above application was that they be effective at dissolving both lignin and wood polysaccharides, such as cellulose or hemicelluloses. Determining which biopolymers these ionic liquids dissolve or extract from the substrate will dictate which fractionation schemes can be applied.



Due to the infancy of ionic liquids research and the structural complexity of ionic liquids in comparison to molecular solvents, this work package demanded a more academic approach. This was conducted alongside assessment of ionic liquids for their efficacy for the processing of lignocellulosics. Three main areas were focused on (Scheme 1).

The research strategy outlined in Scheme 1 can be summarized as follows:

• Thestartingpointwasthesynthesisofexisting imidazolium-based ionic liquids

• Nextwastofurtherourunderstandingofdifferent effects and phenomena that occur when lignocellulosics are contacted with ionic liquids (e.g. dissolution capabilities, fractionation efficiency, fibrillation, chemical reaction).

• Developstructurepropertyrelationshipsin regard to the physical properties of the ionic liquids (e.g. thermal stabilities) and the observed effects and phenomena (mainly biopolymer solubilities). This was achieved through a process of parameterization of effective and non-effective ionic liquids. Computational methods were also used to predict physiochemical properties of structures.

• Throughthisunderstandingofeffects,parameters and the generation of new structures, it was possible to develop hypotheses about which structural features would enable more advantageous effects (e.g. lower viscosities or more recyclable structures)

• Newseriesofstructuresweresynthesizedbringing an improved understanding of physical properties and recycling issues, in particular

• Thiswasfinalizedbyassessmentoftheirpotential in fractionation processes

Scheme 1. Strategy for academic development of novel and effective ionic liquids.


4. Results

4.1 Switchable Ionic Liquid Fractionation

The main objective of the work was to devel-op a process to fractionate the main biomass components to enable their utilization in the production of materials, chemicals and fuels for the future bioeconomy.

Several Switchable Ionic Liquids (SIL) were synthesized and characterized. The prepared SILs were used in a novel process involving the fractionation of Nordic woody biomass by se-lective dissolution. The process was discov-ered and extensively developed during the FuBio programme, with the duration of frac-tionation successfully reduced from five days to just two hours. In addition, relatively high purity of the resulting fractions was achieved; for example, cellulose-rich material contain-ing 79 wt-% cellulose, 11 wt-% hemicelluloses and 5 wt-% lignin was obtained from spruce chips. All of the produced fractions were char-acterized in detail using several methods. The SIL treatment enables the fractionation of biomass into relatively pure fractions un-der milder conditions than in current industri-al processes, thus consuming less energy. The process could be utilized where the production of hemicelluloses and lignin together with cel-lulose fibres is targeted. Hemicelluloses and lignin fractions could be processed further as bio-based chemical and materials, thus adding value to these raw materials compared to the

current kraft process, in which they are burnt for energy recovery.

The SILs were synthesized by bubbling CO2 or SO2 through the liquid mixture with the ratio of amidine/guanidine to hydroxyl-containing compound calculated based on the number of hydroxyl groups in the alcohol or alkanol. Here a sample case is presented as an example procedure. DBU-MEA-SO2-SIL was prepared by passing SO2 through the mixture containing 1:1 molar amounts of 1,8-diazabicyclo-[5.4.0]-undec-7-ene (DBU) and monoethanolamine (MEA). The weight increase with the SIL corresponds to a molar ratio of 1:1:1 of all components. These results indicate that all OH groups of the MEA react upon formation of the ionic liquid according to Scheme 2. This observation is supported by NMR and FTIR studies.

In previous studies it has been demonstrated that SILs are potential novel solvents for the fractionation of lignocellulosic material into suitable fractions. However, the fractionation efficiency has not been comparable to those obtained with conventional ILs mainly due to the low treatment temperature. Therefore, the fractionation of lignocellulosic material was investigated based on new types of SILs. These were based on glycerol or alkanol amine, CO2 or SO2 and an amidine (DBU). The new SILs have one major advantage

Scheme 2. Proposed reaction scheme for the formation of DBU-MEA-SO2-SIL (adapted from Anugwom et.al. 2014. ChemSusChem 7, 1170).


compared to previously presented SILs: their decomposition temperatures are significantly higher, thus allowing higher treatment temperatures, which leads to more efficient fractionation. Wood was fractionated without any mechanical agitation under normal pressure and at 100-120°C. However, the treatment time was found to be too long and too cost-intensive to implement at an industrial scale. Furthermore, the use of large amounts of SILs and the need for drying of the wood raw material would also add to the process cost. However, addition of water to the wood/SIL mixture or/and use of fresh non-dried wood was found to be effective in reducing SIL consumption. The optimization of conditions for the selective fractionation of woody biomass was investigated via a novel and economically feasible fractionation method using an alkanol amine (MEA) and an organic superbase (DBU) derived SIL with water. The Short Time High Temperature (STHT) approach was used, where the wood was immersed in the SILs and water added to achieve a 1:3:5 weight ratio. The mixture was kept at 160°C under normal atmospheric pressure for 2 hours without stirring. While the mixture was still hot, the undissolved wood fraction was separated using vacuum

filtration. The undissolved wood material was washed several times with isopropanol at about 40°C until all the SIL was visually removed. After the dissolution/extraction of wood, the undissolved residues as well as precipitated materials from the SIL were analysed by various methods. Weight losses were recorded and the materials were analysed in order to determine their composition. The latest results showed that the STHT method can be used to remove almost all lignin and the majority of hemicelluloses from wood in just two hours. For example, DBU/MEA/CO2-water treated spruce contained 79 wt-% cellulose, and only 11 wt-% hemicelluloses and 5 wt-% lignin. In addition, the resulting fibres are very light in colour (Figure 1).

Significant further improvements to the process can be made. Firstly, the recovery of dissolved hemicelluloses and lignin from the spent SIL should be improved. In addition, the process is still far from optimized, thus, substantially improved results could be achieved through optimization. Furthermore, the number of new SIL designs is considerable and remains so far unexplored. Most importantly, SILs ‘triggered’ with acid gases other than those studied here could give rise to a rich family of yet unknown potential.

Figure 1. Undissolved fluffy material recovered from A) birch and B) spruce after SIL treatment applying the STHT method. The SIL applied was DBU-MEA-SO2 with water at 160°C for 2 h.


4.2 Recyclable Ionic Liquid Design

Throughout FuBio 1 & JR2, emphasis was placed on the development of recyclable systems for biomass processing at the University of Helsinki. The motivation for this was that no workable strategies were available for recycling ionic liquids after a fractionation step. Typically, oligomeric materials and inorganics are present that are very difficult to remove due to the non-volatility of traditional ionic liquids. Therefore, we sought to introduce recyclability to the ionic liquid in the form of the classical purification methods of distillation and phase separation (Figure 2). In this case the distillation of [TMGH][CO2Et] required 130°C and 5 mbar in a Kugelrohr. In the case of recycling by phase-separation, dissolved cellulose and [P8881][OAc] could be recovered by addition of pure water or kosmotropic salts. [P8881][OAc] could then be reused in the dissolution of cellulose.

After continued optimization of different distillable acid-base conjugate ionic liquids for biomass processing, it was found that [DBNH][OAc] was a highly effective ionic liquid for cellulose dissolution. This was a result of the very low viscosity achieved by the combination of 1,5-diazabicyclo(4.3.0)non-5-ene (DBN, superbase) and acetic acid (organoacid), which was similar to that of the benchmark ionic liquid [emim][OAc] (Figure 3). When combining different bases with acetic and propionic acid, a clear trend developed concerning cellulose solubility. Combinations of these acids with the superbase range proved effective at dissolving cellulose, whereas their combination with normal organic bases was ineffective. This was rationalized by the fact that the basicity of the unconjugated base is also a measure of the ionic liquids’ cation acidity. It was concluded that if the cations are too acidic they stabilize the anion to such an extent that solvation of cellulose, with significant enthalpy of dissolution gain through cellulose hydrogen bond breakage, is inhibited.

[P8881][OAc], which was demonstrated to be phase-separable upon addition of water or kosmotropic electrolyte solutions (e.g. sodium acetate solution), was also found to be an excellent solvent for cellulose as the DMSO electrolyte. Its ability to form isotropic solutions was so strong that it was possible to obtain 1H-13C heteronuclear single-quantum correlation (HSQC) NMR spectra where the cellulose backbone was free from any ionic liquid resonances (Figure 4). To the best of our knowledge, this was the first time that this has been achieved with high DP cellulose (MCC). Moreover, we were able to assign the terminal glycosidic C1 and anomeric C1 signals from the chain ends in the 1H NMR spectra (Figure 5). Cellulose regenerated from this solution mainly as the more thermodynamically stable cellulose II crystalline polymorph, together with some residual cellulose I and amorphous cellulose (Figure 4). Therefore, these are highly suitable media for further biomass pre-treatments, fractionations or chemical modification. This will be thoroughly investigated in the future.


Figure 2. Distillation of the cellulose-dissolving [TMGH][CO2Et] (a) and the recycling strategy for phase-separable ionic liquids, such as [P8881][OAc] (b), which dissolve cellulose as their DMSO electrolytes (Adapted from King et al. 2011, Angew. Chem. Int. Ed. 50, 6301 and Holding et al. 2014. ChemSusChem 9, 1565).

a)

b)


Figure 4. (a) Solution-state HSQC NMR of MCC dissolved in [P8881][OAc]:d6-DMSO. (b) XRD of untreated and regenerated MCC (adapted from Holding et al. 2014. ChemSusChem 9, 1565).

Figure 5. Assigned 1H NMR of MCC dissolved in [P8881][OAc]:d6-DMSO showing polymeric and terminal C1 peaks.

Figure 3. Viscosities vs. temp. of a range of acid-base conjugate ionic liquids (a), with [DBNH][OAc] having the same viscosities as the benchmark [emim][OAc]. X-ray crystal structure of [DBNH][OAc] (b) indicating strong hydrogen bonding between anion and cation. Proposed model for how cation acidity influences the enthalpy gain for Gibbs free energy of dissolution (c). (Adapted from Parviainen et al. 2013 ChemSusChem 6, 2161).

a) b) c)

a) b)


4.3 Homogeneous Ionic Liquid-Aided Fractionation

The objectives of this task were to define ad-equate pretreatments to enhance IL frac-tionation of wood components and to define quantitative measures of IL’s wood polymer solvation capability. We employed a standard process, explained in Figure 6, to investigate the effects of pretreatments.

Comparing native and autohydrolyzed birch, the latter produced better results (Figure 7).

Without autohydrolysis, 7% of birch was in-soluble. Thus, after autohydrolysis (P-factor 500) the purity of precipitate 1 increased sig-nificantly (LIG/CH = 0.27 to 0.16) without loss of cellulose yield or molar mass.

The process was repeated with different particle sizes and P-factors. Compared to <125 µm Wiley meal, more insolubles were produced with commercial sawdust, but this could be compensated by increasing the autohydrolysis P-factor from 500 to 1500. In order to fragment the remaining lignin, ozonation was applied on

Figure 6. Standard fractionation scheme for optimization of processing conditions.

Figure 7. (a) ILAF of wood; (b) ILAF of autohydrolyzed wood.

a) b)


autohydrolyzed wood. However, no increased selectivity in terms of molar mass or solubility was gained.

A final cellulose II pulp with a lignin content of 13% was thus produced. On centrifugation, the pulp compacted into a form containing 96% water, which oven-dried to a black, hard sol-id. As is, the pulp could be blended with card-board pulp to potentially reduce porosity. With further conventional delignification, dissolving pulp could be produced.

4.4 IONCELL-P Process

The objective of this research task was to define the optimal conditions for the quantitative fractionation of paper grade pulps using the IONCELL-P(ulp) process in order to upgrade them to acetate grade or dissolving pulps. The study was based on the differences in solubility behaviour of cellulose and xylan in ionic liquid systems. Different co-solvents mixed with the ionic liquid alter the ionic liquid’s dissolution abilities to different levels. Water addition deteriorates the ionic liquid’s dissolution abilities. When a sufficient amount of water is added, the solvent system enters a so-called

hemicellulose selective dissolution window, as illustrated in Figure 8. Maximum removal of xylan is at the border line between the selective and unselective areas of the chart. When the molar fraction of water exceeds a certain threshold value, hemicelluloses are no longer soluble in the system. This means that the water can also be used to regenerate the dissolved material.

As a result of testing several ionic liquid systems and different sources of pulps for optimizing IONCELL-P fractionation, it can be reported that fractions separated in optimal conditions (3h, at 60˚C, water content dependent on used ionic liquid system) have no molecular weight degradation, have high recovery yields of fractionated polysaccharides, and maintain the cellulose I crystallinity of the cellulose fraction.

The pulp extraction processes can be concluded to be successful, since it has been shown that high yields and high purity polymeric components can be retrieved from the IONCELL-P fractionation processes, with the cellulose fractions having promising properties to enter the market for value-added acetate grade pulps.

Figure 8. Selective hemicellulose dissolution window of [emim][OAc] as a function of ionic liquid, water and pulp content as weight %. The selective dissolution window is highlighted dark blue (a). Molar mass distributions (b): of the initial birch kraft pulp, the separated cellulose fraction (blue), the precipitated hemicellulose fraction (red), and the calculated sum of the fractions (treatment, 15 wt % water, 60°C, 3 h; consistency, 10.5 wt %).

a) b)


4.5 Cellulase Modification of Cellulose in Ionic Liquid

Ionic liquids (ILs) dissolve cellulose. This opens up new interesting opportunities for modifying cellulose physically, chemically and enzymatically. Enzymatic cellulose modifications in IL solutions have so far not been studied significantly. The benefits of using enzyme catalysis include, e.g., mild reaction conditions leading to energy savings and less product degradation, and highly specific reactions meaning fewer by-products as compared to chemical reactions. With different enzymes it could be possible to catalyse oxidation, hydrolysis, acylation and other cellulose modifications in ILs. In FuBio JR2 WP2, enzyme work was concentrated on studying the action of glycosyl hydrolases, mainly endoglucanases, in cellulose hydrolysis in aqueous ILs. By designing functional combinations of glycosyl hydrolases and ILs, new concepts can be considered for tailoring pulp properties (e.g. controlling DP and removing hemicelluloses) and for the total hydrolysis of plant cell wall polysaccharides to monosaccharides in the production of biofuels and biochemicals.

Cellulases are severely inactivated by cellulose-dissolving ILs. In this study, the reasons for cellulase inactivation were studied in detail. At the beginning of the study, it was noticed that [DMIM]

DMP and [EMIM]AcO, two cellulose-dissolving ILs, were very basic in aqueous solution and, thus, the medium basicity appeared a plausible reason for the observed enzyme inactivity. Studies on enzymatic cellulose hydrolysis in different basic solutions without IL, as well as studies with alkaline cellulases, did not, however, support the theory of IL basicity being a major contributor to enzyme inactivation. On the other hand, cellulase thermostability appeared to correlate with higher hydrolytic performance in IL solutions. In the programme, novel cellulose-dissolving and distillable ILs based on the TMG and DBN superbases were developed. The cellulase-compatibility of these ILs was tested, but thee ILs were found not to offer any increased cellulase activity compared to other IL types. Analysis of saccharide content in IL solutions was found to be difficult when developing a capillary electrophoresis method for the analysis of oligosaccharides tolerating the presence of IL.

Radiolabeled cellulases were used to study their cellulose-binding capability in IL solutions (Figure 9). The binding studies were done pairwise with intact cellulases and their core domains, i.e. cellulases lacking a carbohydrate-binding module (CBM). The results clearly show the cellulose binding to be sensitive to IL presence. CBM binding was found to be highly IL-sensitive, whereas cellulose binding through the active site tunnel in cellobiohydrolases was much less affected

Figure 9. Binding isotherms for Trichoderma reesei endoglucanase Cel5A (A), cellobiohydrolase Cel7A (B) and the core domain of Cel7A (C) in five different ionic liquid media.


by ILs. Endoglucanases were found to be very dependent on their CBM for cellulose-binding, which translated into a high IL sensitivity.

Although cellulases were generally inactivated in IL solutions, the IL [DMIM]DMP was found to be stabilizing for cellulases during prolonged incubation periods. Endoglucanases were found to reduce the DP of partially dissolved microcrystalline cellulose in 90% [DMIM]DMP, representing a new cellulase activity on dissolved, non-derivatized cellulose.

4.6 Thermodynamics and Modelling of Ionic Liquid Distillation

The work done consisted of Knudsen effusion apparatus construction, vapour pressure measurements of [TMGH][CO2Et], [DBNH][CO2Et] and [DBNH][OAc] distillable ionic liquids (DILs), distillation experiments of [DBNH][OAc] + water mixture and phase equilibrium measurements and modelling of CO2 + DBU +

Figure 10. Schematic of the Knudsen effusion apparatus: a) wide range gauge; b) turbo pump Edwards STPA 1303C; c) scroll pump XD510; d) gate valve; e) cold finger for liquid nitrogen; f) ball valve; g) air admittance valve; h) gas admittance valve; i) vacuum chamber; j) aluminium blocks (ovens); k) temperature panel Nokeval; l) PID temperature controllers; m) computer.

glycerol switchable ionic liquid (SIL) systems. In addition to the experimental work, thermal separation processes for ionic liquid (IL) recycling in biomass fractionation processes using ionic liquids as fractionating agents were modelled.

The Knudsen effusion apparatus is depicted in Figure 10. Initial tests were carried out. The apparatus is effective for measuring compounds with very low vapour pressures (e.g. aprotic ionic liquids).

The measurement results indicate that DILs can be distilled at pressures below 45 mbar without significant decomposition. Distillation at lower pressures and temperatures decreased the decomposition of DILs. In addition, pure DBN was regenerated, possibly from hydrolysis products of DBN, which may have an important role in improving process feasibility. In the distillation experiments, the DIL was purified to 90 m-% purity, without further optimization.

The structure of CO2 + DBU + glycerol SILs was highly dependent on the DBU:glycerol ratio. SIL systems were stable in the 30-80°C temperature range. Addition of water into the system caused formation of competing CO2 + DBU + H2O SIL.


For modelling the recovery processes of ILs, three promising ILs having water as the antisolvent were chosen: 1) [emim][OAc]; 2) a distillable ionic liquid formed from 1,1,3,3-tetramethylguanidine and propanoic acid; 3) a switchable ionic liquid formed from 1,8-diazabicyclo[5.4.0]undec-7-ene and butanol. The modelling of the chemical systems relied on various estimation methods for modelling physical property, VLE and energy variables. The simulated pressure levels and the presence of reactions in the chemical systems indicate short-path distillation in the technical implementation of the recovery processes of ILs.

4.7 Membrane Purification of Ionic Liquids

The focus of this study was on the recovery and recycling of Ionic Liquids (ILs) by pressure-driven membrane separation processes. ILs were used for processing whole wood. First, the filterability and tolerance of selected membranes towards [emim][DMP] (IL) and DBU-MEA-SO2 (Switchable Ionic Liquid, SIL)

were examined. Membrane filtration of these ILs is challenging, mainly due to their high viscosity. This can be overcome either by using low-viscosity co-solvents, such as molecular solvents (methanol, ethanol, isopropanol, etc.), or by performing filtrations at elevated temperatures. As very high temperatures cannot be used with polymeric membranes, ceramic membranes were also tested. However, ceramic membranes offer restricted cut-off options at the nanofiltration scale. After identifying suitable membranes and proving their usability with the ILs, the final step involved trials with real residue solutions of the ILs. The NF270 nanofiltration membrane and TiO2 ceramic nanofiltration membranes were tested with real residues and the results were very positive: good retentions for sugar-like organics were gained with these membranes for both ILs in the studied conditions (see Figure 11 below for [emim][DMP] residue). The separation processes can also be optimized for better performance through optimal combinations of parameters such as temperature, cross-flow velocity and pressure.

Figure 11. a) Flux of [emim][DMP]-Isopropanol (1:4) solution through the ceramic Inopor®ultra TiO2 membrane (mean pore size 5 nm, cut-off 8500 g/mol) showing the retention of saccharides at different times during the test. Batch 2 L, Δp 3 bar, temperature 30°C, cross-flow velocity 0.8 m/s, filtration area 1040 cm2 and VRF = 2.0. b) The filter unit for high-pressure cross-flow filtrations showing the module for polymeric flat sheet membranes. Inset: module with ceramic membrane.

a)

b)



Our academic understanding of the area has developed to a very high level, and is reflected in the quality of the publications produced. Several new processes have been presented, with both SIL fractionation and the IONCELL-P process representing highly promising methods of wood and pulp fractionation (respectively). Three new classes of recyclable ionic liquids, switchable (SILs), distillable (DILs) and phase-separable (PSILs), are being developed, which should be placed in their appropriate applications in future work. All three are known to dissolve cellulose. SILs are best understood in regard to wood fractionation to produce high-purity cellulose through extraction of lignin and hemicellulose. DILs and PSILs are known superb solvents for cellulose, although their ability to fractionate wood is less studied. Cellulase enzyme activity is reduced in ionic liquids but not destroyed. Mechanistic understanding of this process is being developed and further studies are planned. Methodologies and equipment for studying and modelling the thermodynamics of ionic liquid recycling are also being developed, as are those for membrane purification of the used ionic liquid. For those membranes that are stable in ionic liquids, unoptimized cross-flow nanofiltration of the used ionic liquid demonstrated reasonable fluxes and good saccharide retention. Important progress by international standards has been made in the area of the processing of lignocellulosics using ionic liquids, and the potential of this area of research is clear. Further academic understanding and process optimization are clearly needed to prove the use of ionic liquids for wood fractionation.

In contrast to our understanding of molecular solvents, the area of ionic liquids is a highly demanding area with more complex chemistry

due to their relative novelty. Therefore, much effort has been targeted at understanding these enigmatic structures and the phenomena that they present. To achieve future success in this area, work should be focused on selected target processes and sufficient resources for continued research should be continued as new unique effects and phenomena are constantly being discovered with these media that are not present or achievable with molecular solvents.

Switchable ionic liquid (SIL) fractionation and the IONCELL-P process offer interesting avenues for future development. SIL fractionation clearly needs further work to demonstrate full recyclability of the system, as this was only partly touched upon during the project. Improvements in recovered pulp quality will also likely emerge as a result of this development. The IONCELL-P process, as the simplest process studied, is already quite mature. More detailed techno-economic studies would be useful to refine these processes further. To reduce research costs and enable up-scaling, future programmes should establish protocols for the recycling and reuse of ionic liquids under study. Investment in equipment judiciously located in the appropriate departments would facilitate this and the development of competence in this area.


6. Networking

Close collaboration between the different partners and international research groups was encouraged during the project. Two international symposia were organized on the topic of the work package. One was held in May 2012 at Haikko Manor, Porvoo, and one in August 2013 at the University of Helsinki. Respected international researchers in the area of ionic liquids and biomass processing presented their

work. These included Tom Welton and Jason Hallet from Imperial College, Seema Singh from the Joint Bioenergy Institute, Roberto Rinaldi from the Max Planck Institute for Coal Research, Joern Viell from AVT-Aachen and Martin Lawoko from the Wallenberg Wood Science Centre.

Table 1 shows both the partners and their role in the work package.

Work package partners Role of the participating organisation

Aalto University

•ForestProductsTechnology(FPT)

•BiotechnologyandChemical

Technology (BCT)

(FPT) IONCELL-P Process, Homogeneous IL-aided

fractionation,

(BCT) Thermodynamics and Modelling of Ionic Liquid

Distillation

Andritz Industrial tutor

Lappeenranta University of

Technology

Membrane Purification of Ionic Liquids

Metsä Fibre Work Package Coordinator


University of Helsinki WP leader. Recyclable Ionic Liquid Design


VTT Cellulase Modification of Cellulose in Ionic Liquids

Åbo Akademi University Switchable Ionic Liquid Fractionation

Table 1: Work package partners and their roles


7. Publications

Anugwom, I., Eta V., Mäki-Arvela, P., Virtanen, P., Hedenström, M., Hummel, M., Sixta, H. and Mikkola, J.-P. 2014. Switchable Ionic Liquids as Delignification Solvents for Lignocellulosic Ma-terials. ChemSusChem 7, 1170-1176.

Anugwom, I., Eta, V., Mäki-Arvela, P., Vir-tanen, P., Hedenström, M., Ma, Y., Hummel, M., Sixta, H. and Mikkola, J.-P. 2014. Towards optimal selective fractionation for Nordic woody biomass using Novel Amine–Organ-ic Superbase derived Switchable Ionic Liquid (SIL). Manuscript Submitted.

Anugwom, I., Eta, V., Mäki-Arvela, P., Vir-tanen, P., Lahtinen, M. and Mikkola, J.-P. 2014. The composition and crystallinity of birch chips (Betula pendula) upon 'DBU-DEA-CO2-SIL' alkanol amine - organic super base - de-rived Switchable Ionic Liquid (SIL) treatment. Green Processing & Synthesis 3, 147-154.

Anugwom, I., Mäki-Arvela, P., Virtanen, P., Damlin, P., Sjöholm, R. and Mikkola, J.-P. 2011. Switchable Ionic liquids (SILs) based on glyc-erol and acid gases. RSC Advances 1, 452-457.

Anugwom, I., Mäki-Arvela, P., Virtanen, P., Willför, S., Sjöholm, R. and Mikkola, J.-P. 2011. Treating birch wood with a switchable 1,8-diazabicyclo[5.4.0]-undec-7-ene-glycerol car-bonate ionic liquid. Holzforschung 66, 809-815. Anugwom, I., Mäki-Arvela, P., Virtanen, P., Willför, S., Sjöholm, R. and Mikkola, J.-P. 2012. Selective extraction of hemicellulose from spruce with Switchable Ionic liquids. Carbohy-drate Polymers 87, 2005-2011.

Ebner, G., Vejdovszky, P., Wahlström, R., Su-urnäkki, A., Schrems, M., Kosma, P., Rosenau, T. and Potthast, A. 2014. The effect of 1-ethyl-3-methylimidazolium acetate on the enzymat-ic degradation of cellulose. Journal of Molecu-lar Catalysis B: Enzymatic 99, 121-129.

Eta, V., Anugwom, I., Virtanen, P., Eränen, K., Mäki-Arvela, P. and Mikkola, J.-P. 2014. Loop vs. batch reactor setups in the fractionation of birch chips using switchable ionic liquids. Chemical Engineering Science 238, 242-248.

Eta, V., Anugwom, I., Virtanen, P., Mäki-Arvela, P. and Mikkola, J.-P. 2014. Enhanced mass trans-fer upon switchable ionic liquid mediated wood fractionation. Industrial Crops & Products 55, 109-115.

Froschauer, C., Hummel, M., Iakovlev, M., Ro-selli, A., Schottenberger, H. and Sixta, H. 2013. Separation of Hemicellulose and Cellulose from Wood Pulp by Means of Ionic Liquid/Cosolvent Systems. Biomacromolecules 14, 1741-1750.

García, V., Valkama, H., Sliz, R., King, A. W. T., Myllylä, R., Kilpeläinen, I. and Keiski, R. L. 2013. Pervaporation recovery of [amim]Cl dur-ing wood dissolution; effect of [amim]Cl prop-erties on the membrane performance. Journal of Membrane Science 444, 9-15.

Hauru, L. K. J., Hummel, M., King, A. W. T., Kilpeläinen, I. and Sixta, H. 2012. New insights into the requirements for dissolution of cellu-lose into ionic liquids. Biomacromolecules 13, 2896−2905.


Hauru, L. K. J., Ma, Y., Hummel, M., Alekhi-na, M., King, A. W. T., Kilpeläinen, I. A., Pent-tilä, P. A., Serimaa, R. and Sixta, H. 2013. En-hancement of ionic liquid-aided fractionation of birchwood. Part 1: Autohydrolysis pretreat-ment. RSC Advances 16365-16373.

Holding, A. J., Heikkilä, M., Kilpeläinen, I. and King, A. W. T. 2014. Phase-separable ionic liq-uids and electrolytes for biomass processing. ChemSusChem 9, 1565-1577.

Hummel, M., Froschauer, C., Laus, G., Röder, T., Kopacka, H., Hauru, L. K. J., Weber, H. K., Sixta H. and Schottenberger H. 2011. Dimethyl phosphorothioate and phosphoroselenoate ionic liquids as solvent media for cellulosic ma-terials. Green Chemistry 13, 2507-2517.

Hyväkkö, U., King, A. W. T. and Kilpeläinen, I. 2014. Extraction of wheat straw with aqueous tetra-n-butylphosphonium hydroxide. BioRe-sources 9, 1565-1577.

Hyvärinen, S., Mikkola, J.-P., Murzin, D. Yu., Vaher, M., Kaljurand, M. and Koel, M. 2014. Sugars and sugar derivatives in ionic liquid media obtained from lignocellulosic biomass: Comparison of capillary electrophoresis and chromatographic analysis. Catalysis Today 223, 18-24.

King, A. W. T., Asikkala, J., Mutikainen, I. and Kilpeläinen, I. 2011. Distillable Acid-Base Con-jugate Ionic Liquids for Cellulose Dissolution and Processing. Angewandte Chemie Interna-tional Edition 50, 6301-6305.

King, A. W. T., Parviainen, A., Karhunen, P., Matikainen, J., Hauru, L. K. J., Sixta, H. and Kilpeläinen, I. 2012. Relative and inherent nu-cleophilicity/basicity/diffusivity of imidazoli-um-based ionic liquids – the implications for lignocellulose processing applications. RSC Advances 2, 8020-8026.

King, A. W. T., Xie, H., Fiskari, J. and Kilpeläin-en, I. Reduction of biomass recalcitrance via bi-omass pre-treatments, Chapter 7; Materials for Biofuels, World Scientific Publishing, Ed. Arthur Ragauskas, 2013. (ISBN: 978-981-4513-27-2).

Kyllönen, L., Parviainen, A., Deb, S., Lawoko, M., Gorlov, M., Kilpeläinen, I. and King, A. W. T. 2013. On the solubility of wood in ionic liquids. Green Chemistry 15, 2374-2378.

Leskinen, T., King, A. W. T. and Argyropou-los, D. S. Fractionation of lignocellulosic ma-terials with ionic liquids; Production of biofuels and chemicals with ionic liquids, Springer Book Series - Biofuels and Biorefineries, Ed. Zhen Fang, 2013. (ISBN 978-94-007-7710-1).

Leskinen, T., King, A. W. T., Kilpeläinen, I. and Argyropoulos, D. S. 2013. Fractionation of lig-nocellulosic materials using ionic liquids: Part 2. Effect of particle size on mechanisms of fractionation. Industrial & Engineering Chem-istry Research 52, 3958-3966.

Mikkola, S-K, Robciuc, A., Lokajova, J., Hold-ing, A. J., Lämmerhofer, M., Kilpeläinen, I., Holopainen, J. M., King, A. W. T. and Wied-mer, S. K. 2014. Impact of amidinium, imidazo-lium and phosphonium based ionic liquids on biological cells and liposomes. Submitted to Green Chemistry.


Ostonen, A., Sapei, E., Uusi-Kyyny, P., Kle-melä, A. and Alopaeus, V. 2014. Measure-ments and modeling of CO2 solubility in 1,8 diazabicyclo-[5.4.0]-undec-7-ene - glycerol solutions. Fluid Phase Equilibria 347, 25-36.

Parviainen, A., King, A. W. T., Mutikainen, I., Hummel, M., Selg, C., Hauru, L. K. J., Sixta, H. and Kilpeläinen, I. 2013. Predicting cellulose solvating capabilities of acid-base conjugate ionic liquids. ChemSusChem 6, 2161-2169.

Rauhala, T., King, A. W. T., Zuckerstatter, G., Suuronen, S. and Sixta, H. 2011. Effect of au-tohydrolysis on the lignin structure and the ki-netics of delignification of birch wood, Nordic Pulp & Paper Research Journal 26, 386-391.

Roselli, A., Hummel, M., Monshizadeh, A., Maloney, T. and Sixta, H. 2014. Ionic liquid extraction method for upgrading eucalyptus kraft pulp to high purity dissolving pulp. Sub-mitted to Cellulose.

Stépán, A. M., King, A. W. T., Kakko, T., Toriz, G., Kilpeläinen, I. and Gatenholm, P. 2013. Fast and highly efficient acetylation of xylans in ionic liquid systems. Cellulose 20, 2813-2824.

Wahlström, R., King, A. W. T., Parviainen, A., Kruus, K. and Suurnäkki, A. 2013. Cellulose hydrolysis with thermo- and alkali-tolerant cellulases in cellulose-dissolving superbase ionic liquids. RSC Advances 3, 20001-20009.

Wahlström, R., Rahikainen, J., Kruus, K. and Suurnäkki, A. 2014. Cellulose hydrolysis and binding with Trichoderma reesei Cel5A and Cel7A and their core domains in ionic liquid so-lutions, Biotechnology & Bioengineering 111, 726-733.

Wahlström, R., Rovio, S. and Suurnäkki, A. 2012. Partial Enzymatic Hydrolysis of Micro-crystalline Cellulose in Ionic Liquids by Tricho-derma reesei Endoglucanases RSC Advances 2, 4472-4480.

Wahlström, R., Rovio, S. and Suurnäkki, A. 2013. Analysis of Mono- and Oligo-saccharides in Ionic Liquid Containing Matrices. Carbohy-drate Research 373, 42-51.


HYDROXY ACIDS(AND OTHER ACIDS)

FROM BLACK LIQUOR ADVANCED SEPARATION TECHNOLOGIES

POLYMERIZATION OF HYDROXY ACIDS

BIOTECHNICAL PRODUCTION OF GLYCOLIC ACID

CONTACT PERSON – Work Package 6 leader Jarmo Ropponen, [email protected]

Hycail: Svante Wahlbeck Kemira: Jukka HietalaMetsä Fibre: Ismo Reilama Lappeenranta University of Technology: Sanna Hellstén, Mari Kallioinen, Jussi Lahti,Mika Mänttäri, Tuomo Sainio University of Helsinki: Sari Rautiainen, Timo Repo University of Jyväskylä: Raimo Álen, Pauli MoilanenUPM-Kymmene: Päivi VarvemaaVTT Technical Research Centre of Finland: Thomas Gädda, Sakari Kaijaluoto,Katariina Kemppainen, Outi Koivistoinen, Klaus Niemelä, Minni Pirttimaa, Peter Richard


ABSTRACT

The purpose of this study was to develop technologies and competences to recover hydroxy acids from black liquor and produce glycolic acid biotechnically from sugars. A novel multistep separation process for recovering hydroxy acids from black liquor, combining membrane filtration, chromatographic separation and electrodialysis, was developed. Further polymerization of the hydroxy acids was also conducted to evaluate their properties as value-added products. The results suggest that in the demonstrated process the neutralization of black liquor is not required, and that the purity of the end product is suitable for polymerization. In addition, the study demonstrates for the first time the conversion of purified hydroxy acid mixtures obtained from black liquor to polymeric material. Furthermore, production of 17 g/l of glycolic acid was achieved by fermentation of the engineered Kluyveromyces lactis strain. Development of bioreactor cultures and the selection of a suitable yeast host led to improvements in production levels that demonstrate technical feasibility. The results identify black liquor as an untapped source of hydroxy acids which can be further developed into value-added products.

Keywords:hydroxy acids, separation, polymerization, biotechnical production


1. Background

Globally, more than 140 million tonnes of wood kraft pulp and 20 million tonnes of non-wood kraft and soda pulps are produced each year. Only a fraction of the dissolved organic matter originating from this production – including tall oil (1.4 million tonnes), turpentine (200 000 tonnes) and kraft and soda lignin (100 000 tonnes) – is currently isolated as pulping by-products for chemical or material applications. The spent cooking liquor from kraft pulping, or ‘black liquor’, consists mainly of lignin and wood extractives, but also contains salts of hydroxy carboxylic acids, which are formed from wood carbohydrates in alkaline conditions during pulping. All black liquors contain varying amounts of glycolic, lactic, 2-hydroxybutanoic, 2,5-di-hydroxypentanoic, xyloisosaccharinic, and glucoisosaccharinic acids as the main hydroxy acids, in addition to numerous minor ones. The formation of hydroxy acids may account for up to 10–15% of wood raw material consumption in pulping. Their isolation as new pulping by-products has not been realized at the mill scale to date due to a number of technical challenges as well as limited markets for many compounds. The annual production of these hydroxy acids in kraft pulping is approximately 6 million tonnes in Europe alone, representing a vast underutilized raw material source. Hydroxy acids are building block chemicals which could be widely utilized in the synthesis of a range of chemicals and polymeric materials as replacements for fossil-derived materials. Potential spearhead applications include biodegradable plastics, packaging films, adhesives and cosmetics.

Most previous studies addressing the isolation of hydroxy acids from black liquors deal with isolation in small volumes for analytical purposes. In the FuBio programme, three approaches for larger-scale isolation of hydroxy acids were investigated: electrodialysis,

chromatography, and a combination of membrane processes with precipitation-crystallization steps. Significant progress was made in each of these areas, resulting in a number of potential process concepts for further optimization and development. In order to achieve reasonable amounts of hydroxy acids to enable practical application evaluations, efforts were focused on developing a multistep separation and purification process encompassing all of the abovementioned purification methods.

2. Objectives

The main objective was to design a novel value chain based on hydroxy acids separated from black liquor or biotechnically produced from sugars. These hydroxy acids would be converted into novel biopolymers and developed and tested for the technologies needed to enable such a value chain in reality. The specific aims were to develop:

• Conceptfortheseparationoforganicacidsfrom black liquor

• Fermentation-basedtechnologyfortheproduction of glycolic acid

• Potentialapplicationsfortheseorganicacids

3. Research approaches

As there were no existing feasible separation processes for the recovery and purification of hydroxy carboxylic acids from black liquor to serve as a starting point, the research approach in process development was strongly experimental in nature. It was concluded early on that a multistage separation process will be needed due to the complexity of the feedstock and the high purity requirements of the final product. The majority of process development was conducted using authentic soda black


liquor instead of kraft black liquor mainly for safety reasons. The selection of ultrafiltration modules and membranes, however, was carried out with kraft black liquor. Experiments with synthetic solutions were also carried out to investigate the most relevant separation mechanisms in more detail.

Both hardwood (HW) and softwood (SW) soda black liquors were used. The main differences in the acid compositions of HW and SW soda black liquors are the larger amount of glucoisosaccharinic acid (GISA) in SW black liquor and the higher concentrations of 2-hydroxy butanoic acid and acetic acid in HW black liquor.

These hydroxy acids offer many potential applications, e.g., as polymer precursors. As the separated hydroxy acids were obtained in acid form, polymerisations were carried out using condensation polymerisation to obtain polymeric materials. In addition, ring-opening polymerisations were used to polymerize glycolide produced from glycolic acid and benchmarked against commercial glycolide-based polyglycolic acid. Moreover, the development of glycolic acid production based on yeast fermentation was continued from the first phase of the programme.

4. Results

4.1 Separation process for recovery and purification of organic acids from black liquor

A process sequence consisting of ultrafiltration, size-exclusion chromatography, ion exchange, adsorption, and evaporation was successfully implemented at laboratory scale (Figure 1) to recover and purify a mixture of hydroxy acids from black liquor. The purpose of the ultrafiltration stage was to remove the majority of lignin and thus facilitate the subsequent chromatographic step, in which the carboxylic acids and NaOH were separated from each other. The acids were liberated using an ion exchange resin and concentrated by evaporation. Evaporation also effectively eliminated volatile acids from the hydroxy acid fraction. Finally, residual lignin was removed by adsorption. Mixtures of hydroxy acids were obtained in high purity both from softwood and hardwood soda black liquors. An example of concentrations of individual acids and lignin after each process step is displayed in Figure 2.

Figure 1. Separation process for recovering hydroxy carboxylic acids, volatile acids, and NaOH from black liquor.


Removal of lignin and recovery of hydroxy acid salts from soda black liquorBased on preliminary experiments, UP010 and NP010 membranes with cut-off values of 10 kDa and 1 kDa, respectively, were chosen for more detailed studies. Filtration experiments were carried out with a rectangular cross-flow module at a constant permeate flux of 70 kg/(m2h) and 40 kg/(m2h) until the pressure needed for constant flux increased to a predefined upper limiting value (10 bar with UP010, 20 bar with NP010). With both membranes, lower pressures were needed for constant flux with softwood soda cooking liquor than with hardwood cooking liquor.

In the filtration of hardwood soda black liquor, lignin concentration decreased from 79.5 g/L to 16.2 g/L when NP010 membrane was used and to 32.6 g/L when using UP010 membrane. The dry solids content decreased from 14.1 wt-% to 9.0 wt-% using NP010 and to 10.4 wt-% using UP010. For the softwood soda black liquor, the amount of lignin decreased from 72 g/L to 14.6 g/L using NP010 membrane and to 22.5 g/L using UP010. The amount of dry solids decreased from 14.0 wt-% to 10.8 wt-% with NP010 and 10.3 wt-% with UP010. Subsequent chromatographic tests confirmed

that the permeate of UP010 requires excessive washing of the chromatographic column, and therefore the 1 kDa NP010 is recommended for ultrafiltration.

The concentrations of acids in the ultrafiltration permeates are shown in Table 1. It was found that the hydroxy acids were well recovered in the permeate. For instance, more than 90% of the isosaccharinic acids (softwood soda cooking liquor) were in the permeate stream (1 kDa, NP010 membrane) at a volume reduction of about 80%. Generally, acids retention was very low or even negative. Due to the analytical uncertainties, small differences in acid concentrations are not meaningful.

A significant purification of acids was achieved. For instance, the purity (based on total organic matter) of isosaccharinic acids increased from 20% to 40% when the softwood soda cooking liquor was filtered (1 kDa membrane). Purity increase was caused by removal of lignin and probably also hemicelluloses. Only about 10% of inorganic compounds (NaOH) were retained by the membrane.

Figure 2. Typical average compositions of the hydroxy acid containing streams after each process step.


Chromatographic separation of hydroxy acids and sodium hydroxideThe key step in the multistep separation process developed here is chromatographic separation of hydroxy acids from the inorganic compounds in black liquor. As no method was available to conduct such a separation in alkaline conditions, considerable efforts were invested in finding a suitable chromatographic separation material for this purpose.

After testing six different types of separation media and several ionic forms of various ion exchange resins, Sephadex G–10 size-exclusion gel was found to be suitable for separating hydroxy acids in the form of sodium salts from sodium hydroxide. A typical chromatogram, i.e. concentration history at the column outlet, from the separation of hardwood soda black liquor is

overlaid on a photograph of the sample bottles in Figure 3. NaOH elutes between approximately 0.8 and 1.0 bed volume (fractions 7 and 8), whereas the hydroxy acids elute between 0.5 and 0.8 bed volume (fractions 3 to 6).

Some fractionation of the acids by size-exclusion occurred in the Sephadex column, although this was not the main objective of this separation step. Isosaccharinic acids elute rapidly due to their large molecular size, and therefore the largest concentrations were observed in fractions 3 and 4 obtained from hardwood soda black liquor. Interestingly, dicarboxylic acids were also enriched in fractions 3 and 4. Smaller 2-hydroxy butanoic acid was accumulated in fractions 5 and 6. GC/MS analysis also revealed the presence of polar aromatic acids in fraction 6. Highest acid

XISA,g/L

GISA,g/L

2,5-DHPAg/L

2-OH-butanoic acid,g/L

Lactic acid,g/L

Glycolic acid,g/L

Oxalic acid,g/L

Acetic acid,g/L

Formic acid,g/L

Total,g/L

HW Feed

NP010

UP010

SW Feed

NP010

UP010

3.2

2.8

4.5

3.7

4.4

4.9

7.3

6.8

6.0

15.7

18.3

18.7

1.1

0.9

0.8

1.3

1.1

1.3

4.0

4.4

4.0

1.3

1.3

1.4

2.8

1.8

2.5

4.2

0.0

4.8

1.2

1.2

1.4

1.9

1.7

1.9

0.5

0.3

0.5

0.5

0.7

0.7

13.7

13.6

16.0

4.4

1.6

5.2

5.4

4.7

6.0

5.4

5.9

5.9

39.2

36.6

41.7

38.4

35.1

44.6

Table 1. Concentrations of carboxylic acids based on CE analysis in the feed and in the ultrafiltration permeates of NP010 and UP010 membranes in ultrafiltration of hardwood (HW) and softwood (SW) soda black liquors.

Figure 3. Chromatographic separation of hydroxy acids and sodium hydroxide from ultrafiltration permeate (NP010) of hardwood soda black liquor using Sephadex G–10.


concentrations were found in fractions 4 and 5, while the concentration of acids in fraction 6 was very low. The total recovery yield of hydroxy acids was 95% with the fractionation scheme chosen. Similar results were also obtained for softwood soda black liquors.

It was further found that the recovery of hydroxy acids from soda black liquor is possible also when the feed is concentrated to 25 wt-% dry solids. This is attractive, as a higher feed concentration markedly increases the productivity of the process. Moreover, good separation was achieved with injection volumes significantly larger than 8% of bed volume (Figure 4). This is because the elution of sodium hydroxide slows down with increasing amount

of hydroxy acids present. Injection volumes up to 24% were found to provide satisfactory purity of the hydroxy acids with respect to sodium hydroxide. However, the concentration of lignin in the hydroxy acid fraction increases with increasing injection volume.

A series of 44 separation runs with hardwood soda black liquor were conducted in the Sephadex G–10 column to determine whether the material is fouled by lignin. No shifts in the retention times or concentration profiles of the hydroxy acids and sodium hydroxide were observed in the conductivity and RI-signals, indicating that fouling does not affect the separation of hydroxy acids and NaOH. On the other hand, the amount of lignin in the

Figure 4. Effect of injection volume on the separation of hydroxy acids from hardwood soda black liquor using Sephadex G–10.


hydroxy acid fraction doubled during the 44 consecutive cycles. This does not necessarily pose major problems, as the residual lignin can be effectively removed in the subsequent process steps.

The pH range recommended by the manufacturer for Sephadex G–10 is 2 to 13. Therefore, its long-term stability in highly alkaline conditions and elevated temperature was investigated. Moisture retention capacity is often used as a measure of effective cross-link density in polymer gels. The changes in the moisture retention capacity of Sephadex G–10 during 4 weeks exposure to 1 M NaOH at 60 oC were below the experimental accuracy, which suggests that its cross-link density was not significantly reduced. Nevertheless, two unidentified compounds were observed in HPLC analyses of liquid phase samples after aging of Sephadex G–10 in 1 M NaOH.

Liberation of acids by ion exchangeIon exchange with strong acid cation exchange resins was found to be suitable for liberation of the acids (i.e. replacement of Na+ with H+). The fractions obtained in the chromatographic separation step were fed through an ion exchange column packed with CS11GC resin in H+ form. The pH of the solutions collected at the column outlet after ion exchange ranged from 1.6 to 2.9. The conversion of sodium salts to acid form was 95–100% for all fractions. No loss of acids due to adsorption to the stationary phase was observed. The acid composition was unchanged, with the exception that GISA is converted to its lactone form due to the decrease in pH.

An aspect that deserves attention when applying ion exchange for liberation of the acids is the formation of sodium salts during regeneration of the resin. For example, if sulphuric acid is used for regeneration, a sodium sulphate stream is produced. To avoid disturbing the sodium–sulphur balance of

the pulp mill, this stream must be treated separately, e.g. by electrodialysis.

Removal of residual lignin by adsorptionMost of the lignin was precipitated from the hydroxy acid fraction in the ion-exchange stage. For example, from the hardwood soda black liquor the average lignin content decreased from approximately 3 g/L to less than 1 g/L. Removal of residual lignin was done by adsorption on a neutral hydrophobic polymer resin (XAD–16). After adsorption treatment, the lignin contents of the hardwood and softwood soda black liquors were approximately 0.2 g/L and 0.1 g/L.

No significant losses of hydroxy acids were observed during adsorptive lignin removal. The yield of acids in the lignin removal step was 96%. XAD–16 was thus concluded to be quite selective towards lignin over hydroxy acids.

Concentration by evaporationThe hydroxy acid fractions were concentrated by evaporation at reduced pressure after the adsorption treatment step. The relative amount of volatile acids was reduced by 40–70%. The final purity of hydroxy acids was 38–91%. If higher purity is required the product can be purified, for example by crystallization or chromatographic separation. The concentrations of the main components in the final products are presented in Table 2. In addition, GC/MS analysis revealed the presence of some glycerol and sugars (not shown).

The final concentration could also be performed using membrane filtration, provided that the membrane can withstand acidic conditions (initial pH 2, final pH approximately 1). Using a suitable membrane, small volatile acids could be separated in the concentration step. Concentration by membrane filtration is most probably not feasible before liberation of the acids due to high lignin solubility at alkaline pH.


Further results on recovery and purification of hydroxy acids Further findings regarding the recovery and purification of hydroxy acids from black liquor are described briefly below.

Recommended ultrafiltration module. In order to select an appropriate type of membrane module for further studies, low shear rate (DSS) and high shear rate (CR, VSEP) membrane filtration modules were tested. 10 wt-% kraft black liquor (UPM Kaukas mill) was successfully concentrated to about 90% volume reduction with both low and high shear rate modules using NP010 (1 kDa) membrane. The flux of the CR-filter was significantly higher than that of the VSEP-filter or the DSS-filter. In the CR-filter, the average flux was over 130 kg/(m2h) up to a volume reduction of 80%. When the volume reduction exceeded 75% the flux started to decrease very rapidly from 100 kg/(m2h) to 5 kg/(m2h) at a volume reduction of 90%. It was thus concluded that the CR-filters with a rotating blade near the membrane surface were the best choice for ultrafiltration of black liquor.

Recovery and liberation of carboxylic acids by electrodialysis. Electrodialysis (ED) was investigated as an alternative method for recovery and purification of hydroxy acids from black liquor. It was noted that electrodialysis membranes were chemically not very durable against black liquor at any concentration, even when the solution pH was decreased by carbonation. This was mainly due to the precipitation of lignin (and partly extractives) on the membrane surface, which rapidly led to permanent losses in ion exchange efficiency of the membrane. In a typical case, the efficiency for the first 4 hours was 0.25 kWh/mol and during 4-9 hours 0.42 kWh/mol.

Electrodialysis was also tested as an alternative for ion exchange in liberation of the acids. A fraction of hydroxy acids obtained by ultrafiltration and chromatography was used as the feed. The fraction used as feed contained approximately 3.3 g/L of lignin.

In the first phase, the fraction was processed by ED. After this experiment the membranes

Fraction Mass

reducti-

on,

wt-%

Lignin,

g/L

XISA,

g/L

GISA,

g/L

2,5-

DHPA,

g/L

2-OH-

butanoic

acid,

g/L

Lactic

acid,

g/L

Glycolic

acid,

g/L

Oxalic

acid,

g/L

Acetic

acid,

g/L

Formic

acid,

g/L

HW black

liquor

3 85 1.21 6.2 7.4 0.00 0.00 0.4 1.0 3.0 2.5 0.0

4 89 1.41 46.2 22.9 0.00 10.2 7.9 9.7 1.5 23.3 3.9

5 85 2.13 5.0 2.2 0.00 16.3 4.9 3.5 0.0 20.9 10.6

6 92 1.92 0.00 0.00 0.00 4.1 0.6 0.3 0.5 2.8 2.8

SW black

liquor

1 92 1.03 5.2 45.8 0.00 0.00 1.7 2.9 3.8 3.6 0.7

2 96 4 53.4 148 4.38 7.9 34.1 24.0 0.0 13.0 9.2

3 89 2.49 8.4 2.0 2.20 5.6 8.5 5.0 0.0 7.0 15.2

Table 2. Concentrations of lignin and carboxylic acids in the concentrated product fractions recovered and purified from hardwood (HW) and softwood (SW) soda black liquors.


were kept in the initial fraction and, after two weeks, the same membranes were used again for ED. The same procedure was repeated after six weeks. As can be seen from Table 3, efficiency clearly dropped already after 2 weeks from 0.23 kWh/mol to 0.29 kWh/mol. After six weeks, the efficiency was 0.28 kWh/mol. This efficiency deterioration was probably due to the conditions used in the durability tests, i.e., when bipolar membranes were soaked in the hydroxy acid fraction also its anionic side was influenced, which would not be the case under normal operating conditions.

Membranes were visually estimated after 2 and 6 weeks of soaking and no visual changes were observed. After ED runs a thin layer of lignin was precipitated on the surface of the bipolar membrane’s cationic side. This layer could be rinsed off with water. The results indicate that at least 90% of initial lignin is removable from the black liquor by ED treatment.

Fractionation of hydroxy acids by nanofiltration. The use of nanofiltration to separate carboxylic acids from each other was demonstrated using Desal-5 DK membranes. A kraft black liquor that was pre-treated by cooling crystallization and acid precipitation to remove lignin was used as the feed. The effect

of pH on the retention of acid compounds was investigated by increasing the pH of the feed liquor stepwise from pH 2.3 to 12 with NaOH.

It was found that pH had a marked effect on permeate flux and retention, and thus on hydroxy acid separation. Higher pH can either increase retention, as with sulphate and xyloisosaccharinic acid XISA, or decrease it, as with formic acid (Figure 5). Nanofiltration can thus be used to fractionate organic acids into two fractions that are rich in different hydroxy (or volatile) acids. Acetic and formic acids permeated the NF membrane especially at high pH, and isosaccharinic acids and sulphate ions were concentrated in the retentate stream. Nanofiltration at pH 2.3 led to a concentrated stream with an acids content of about 80% of total organic matter. Furthermore, GISA purity in the concentrate was almost 40% based on total organic matter.

4.2 Polymeric materials from fractionated black liquor

Fractionated black liquor samples for the polymerization were obtained from the partners in the project. One fraction (HW1) from hardwood black liquor and three fractions (SW1, SW2, and SW3) from softwood black liquor

Parameter Fresh 2 weeks 6 weeks

Hydroxy acids (g/L) 19.6 20.1 20.1

Efficiency after 1 h

(kWh/mol)

0.23 0.29 0.28

pH after ED 2.37 2.23 2.07

Lignin (at 280 nm; alkali so-

luble) (g/L)

0.36 0.26 0.24

Lignin (at 205 nm; acid so-

luble) (g/L)

1.36 1.09 1.11

Residual sodium (g/L) 0.30 0.19 0.01

Table 3. Results obtained from the Lappeenranta fraction (as such and after the ED experiments with varying treatment time).


were used. The fractions were purified by the abovementioned combination of ultrafiltration and size-exclusion chromatography. Two samples of black liquor were purified by electrodialysis (ED). The black liquors originated from soda-AQ pulping. The compositions of the various fractions were calculated from results obtained by capillary electrophoresis (Table 4).

The dilute aqueous fractions of black liquor were polymerized by condensation polymerization. A predetermined reaction volume was placed into a suitable round-bottomed flask equipped with a Teflon® covered magnetic stirring bar and a distillation head. Sn(Oct)2 (0.1 wt-% based on analysed acid content) was used as

a catalyst. Temperature of the reaction was increased gradually from 130 °C to 160 °C while maintaining a pressure of 500 mbar. Pressure was then gradually decreased to 30 mbar. Once the target pressure was achieved, the reaction was allowed to continue for 24 h. Polymeric materials were successfully obtained from all of the fractions by condensation polymerization. The distillate obtained from the dilute hydroxy acid mixtures was analysed to determine how the polymerization proceeds. The distillates analysed were shown to contain in all cases formic and acetic acid. Glycolic, lactic and 2-hydroxybutyric acids were also detected in some cases.

Figure 5. Effect of pH on the retention of various organic acids during nanofiltration with Desal-5 DK membrane (70 °C, 25 bar).

OA

g/L

(χOA)

FA

g/L

(χFA)

AA

g/L

(χAA)

GA

g/L

(χGA)

LA

g/L

(χLA)

2HBA

g/L

(χ2HBA)

2,5-DHPA

g/L

(χDHPA)

XISA

g/L

(χXISA)

GISA

g/L

(χGISA)

HW1 1.5 (1.0) 3.9 (5.2) 23.3 (23.9) 9.7 (7.8) 7.9 (5.4) 10.2 (6.1) 46.2 (18.9) 22.9 (8.5)

SW1 3.8 (2.8) 0.7 (1.0) 3.6 (4.0) 2.9 (2.5) 1.7 (1.2) 5.2 (2.3) 45.8 (18.4)

SW2 9.2 (6.2) 13.0 (6.8) 24.0 (9.8) 34.1 (11.8) 7.9 (2.4) 4.4 (1.0) 53.4 (11.1) 148.1 (27.8)

SW3 15.2 (27.7) 7.0 (9.8) 5.0 (5.5) 8.5 (7.9) 5.6 (4.5) 2.20 (1.4) 8.4 (4.7) 2.0 (1.0)

SODAAQ1 0.4 (3.2) 0.9 (6.1) 0.3 (1.8) 0.5 (2.2) 0.2 (1.0) 1.2 (3.5) 3.0 (6.7)

SODAAQ2 2.0 (14.8) 1.9 (10.5) 0.9 (4.1) 1.1 (4.3) 0.6 (1.8) 0.4 (1.0) 1.6 (1.0) 5.0 (10.1)

Table 4. Compositions [g/L] and molar ratios (χ) of black liquor fractions analysed by capillary electrophoresis.


All products had the physical appearance of solid materials or highly viscous fluids. This is taken as primary evidence for the formation of polymers. Similarly, the thermal properties of the samples suggest that polymers were obtained. The Tgs varied between -1 and 49 °C, depending on the sample. The reason for this variance is not understood and further experiments should be done with larger amounts of polymer formed. The properties of the characterized products, reaction volumes, yields, theoretical yields calculated, thermal properties and molecular weights are presented in Table 5.

The obtained polymer yields were generally higher than the calculated yield based on the amount of acids. This indicates that the condensation is incomplete and there is still unreacted material remaining. This result was expected as the OH/COOH ratios are not even. The polymers obtained were found to be relatively poorly soluble in most common organic solvents. The polymers were insoluble in chlorinated solvents, hydrocarbons and alcohols. The solubilities varied from poor to good when dissolved into polar aprotic solvents such as DMSO and THF. The solubility of the materials depended on the material composition, although thorough analysis of this was not possible due to the limited amounts of material available.

Glycolide production and ring-opening polymerization of glycolide In addition, a method for glycolide production by polycondensation of glycolic acid using a catalyst under reduced pressure suitable for large-scale production was successfully developed. The production of glycolide using the chosen distillation method produced a yield of only 20% glycolide from the polycondensate of glycolic acid. Using a catalyst, glycolide formation reached 100 g/kg polycondensate/hour. However, the simultaneous formation of insoluble residue was a main concern for the development of an industrial continuous production process based on the studied method. The purity of the produced glycolide was comparable to commercial polymerization grade glycolide. Collecting the distillate of glycolide in a nonpolar solvent and re-crystallization yields a purity of 99%, which is suitable for production of high molecular weight polymers. Polymerization of high molecular weight polyglycolic acid (PGA) was demonstrated using ring-opening polymerization in the melt (0.1 wt-% SnOct, 250°C, 4h). The glycolide samples along with a commercial reference were polymerized successfully (Table 6). The polymers obtained showed some differences in structural analysis (Figure 6). The additional peaks seen with the polymers prepared from glycolide samples 53 and 102 are due to the polymer end groups. The minorpeaksobservedat~69ppmand~66ppmare due to residual glycolide.

Volume

[mL]

Acid

content

[g/L]

Yield

[g]

Yield

[%]

Tg

[°C]

Tg,calc.

[°C]

Mn,theor

[g/mol]

Mn

[g/mol]

Mw

[g/mol]

HW1 10 125.5 1 96 -1 11 770 734 1010

SW1 20 63.6 1.38 128 27 6 1740 1030 1420

SW2 2 294.0 0.56 112 6 8 620 540 750

SW3 20 53.8 0.82 88 49 57 3560 1600 2400

SODAAQ1 200 6.5 1.27 117 42 8 670 1330 1900

SODAAQ2 200 13.6 1.72 78 32 9 1010 1030 1410

Table 5. Reaction volumes, obtained and theoretical yields, obtained and theoretical glass transition temperatures and obtained molecular weights of the polymers obtained from black liquor fractions.


4.3 Glycolic acid by fermentation

Biotechnical production of glycolic acid was demonstrated using host organisms Saccharomyces cerevisiae, Aspergillus niger, and Kluyveromyces lactis. Glycolic acid was produced in the glyoxylate cycle of the cell. Expression of the glyoxylate reductase gene and deletion of the malate synthase gene are the minimum requirements for glycolic acid production.

Yeast codon optimized glyoxylate reductase gene, GLYR1, from the plant Arabidopsis thaliana was introduced to the production host. Overexpression of this gene combined with other modifications boosts the glyoxylate

cycle and directs the metabolic flux towards the cycle, thus allowing the yeast to produce glycolic acid (Figure 7). Development of bioreactor cultures and the selection of a suitable yeast host led to improved production levels that demonstrate technical feasibility. Production of glycolic acid was shown to be possible also at low pH, which is an advantage for the purification process and a clear benefit when compared to Escherichia coli, which has also been reported to be used as a host for glycolic acid production. At low pH there is less need for neutralization and the separation process is thus easier because less gypsum is formed and the glycolic acid is in acid form. Low pH also decreases the risk of bioreactor contamination.

Glycolide Yield GPC

Mw/Mn

Tg

(°C)

Tm

(°C)

PURAC 99% 5,890 / 3,400 38 217

Hycail 1 98% 25,700 / 13,400 43 210, 221

Hycail 53 99% 22,100 / 11,500 44 209, 222

Hycail 102 99% 36,600 / 17,400 44 221

Table 6. Results from glycolide polymerizations.

Figure 6. 13C NMR comparison showing only minute differences in polymer structures.


The best result (17.2 g/l) in the production of glycolic acid was achieved by fermentation of the engineered Kluyveromyces lactis strain. The benefit of K. lactis compared to S. cerevisiae is that it is a Crabtree negative yeast which favours respiratory metabolism, meaning that the organism does not catabolize sugars to ethanol as efficiently as S. cerevisiae.

The first step in constructing the glycolic acid production strain in K. lactis was to delete malate synthase in order to prevent the conversion of glyoxylate to malate. In addition, the cytosolic isocitrate dehydrogenase was deleted and, finally, the S. cerevisiae codon-optimized glyoxylate reductase, GLYR1, was overexpressed from a multicopy plasmid.

After five days, about 3 g l-1 of glycolic acid was produced in flask cultivations. The modified strain was grown on unbuffered media containing 20 g l-1 xylose and 20 g l-1 ethanol and fed daily with additional 10 g l-1 ethanol. Further work is needed to increase the production rate and also to bypass the ethanol step, as it has been shown that only ethanol is converted to glycolic acid, whereas xylose and other sugar substrates are either used solely for cell growth or are first converted to ethanol before converting to glycolic acid. The result shows that K. lactis is a promising host for glycolic acid production as it is producing more glycolic acid with fewer modifications than S. cerevisiae strains.

Figure 7. Modified glyoxylate cycle in S. cerevisiae. Overexpressed genes are presented in green, deleted genes in red.


Kluyveromyces lactis bioreactor cultivationsK. lactis strain H3986 was grown in a Biostat CT-DCU bioreactor at pH 5.0, at 30°C, 1 volume air [volume culture]-1 min-1 (vvm) and 500 rpm agitation. Xylose (20 g l-1) and ethanol (20 g l-1) were provided as carbon source at the beginning of the fermentation. Xylose and ethanol were fed to the bioreactor as separate feeds based on their consumption. The fermentation was monitored for 12 days (Figure 8).

Extracellular metabolites in cell-free spent culture medium were analysed by capillary electrophoresis. H3986 produced extracellularly 17.2 g glycolic acid l-1 after 9 days at pH 5.0 (Table 7). The yield of glycolic acid on xylose-ethanol media was approximately 0.13 g glycolic acid [g ethanol + xylose]-1. Acetate was also produced towards the end of fermentation, but as the glycolic acid level was already close to maximum at this stage, it would have been possible to stop the fermentation before acetate accumulation, thus facilitating the separation process.


The work focuses on separation of hydroxy acids from black liquor and their further upgrading, e.g. as hot melts or reactive monomers. From the industry point of view, if the separation concept proves economically feasible, hydroxy acid based products recovered from black liquor would offer opportunities for pulp and papermaking companies to utilize this side-stream as a valuable raw material for environmentally friendly products.

Figure 8. Xylose, ethanol and oxygen consumption and glycolic acid, acetate and CO2 production.

Glycolic acid production (g l-1)

Day 0 Day 2 Day 4 Day 6 Day 8 Day 9

0 9.9 13.8 14.8 16.1 17.2

Table 7. Glycolic acid production by strain H3986 at pH 5.0 in a bioreactor.


6. Networking

The research was carried out jointly by research organisations and Finnish bioeconomy clustercompanies. Table 8 below presents the research partners and their roles.


Hellstén, S. Recovery of biomass-derived val-uable compounds using chromatographic and membrane separations, Doctoral dissertation, Lappeenranta University of Technology, 2013.

Hellstén, S., Heinonen, J. and Sainio, T. Size-ex-clusion chromatographic separation of hydroxy acids and sodium hydroxide in spent pulping liquor, Sep. Purif. Technol., 118, 2013, 234–241.

Hellstén, S., Lahti, J., Heinonen, J., Kallioinen, M., Mänttäri, M. and Sainio, T. Purification pro-cess for recovering hydroxy acids from soda black liquor, Chem. Eng. Res. Des., 91, 2013, 2765-2774.

Koivistoinen, O. Catabolism of biomass-de-rived sugars in fungi and metabolic engineering as a tool for organic acid production, Doctoral dissertation, University of Helsinki, 2013.

Koivistoinen, O.M., Kuivanen, J., Barth D., Turk-ia H., Pitkänen, J-P., Penttilä, M. and Richard, P. Glycolic acid production in the engineered yeasts Saccharomyces cerevisiae and Kluyvero-myces lactis, Microb Cell Fact 12, 2013, 16 pp.

Koivistoinen, O., Kuivanen, J., Penttilä, M. and Richard, P. Eukaryotic cell and method for pro-ducing glycolic acid, WO13050659A1.

Pirttimaa, M., Rovio, S., Harlin, A., Ropponen, J. and Gädda, T. Polymers derived from isolat-ed hydroxy acids obtained from hardwood and softwood black liquor, submitted.



Hycail Preparation of glycolide from glycolic acid

Kemira Industrial tutor

Lappeenranta University of

Technology

Fractionation and purification of hydroxyl acids from black liquor by

membrane filtration and chromatographic separation


University of Helsinki Expertise in oxidation of sugars to hydroxy acids

University of Jyväskylä Developing the separation of hydroxy acids from black liquor by

electrodialysis

UPM-Kymmene Work Package Coordinator

VTT WP leader. Development of a chromatographic separation method

for hydroxy acids from black liquor and analysis. Polymerization and

copolymerization of hydroxy acids. Development of an enzymatic route to

glycolic acid from sugars.


BIOLOGICAL EFFECTS OF WOOD-BASED EXTRACTS

AND COMPOUNDSIN MODELS OF HUMAN

DISEASE

CONTACT PERSON - Work Package 5 leader Niina Saarinen-Aaltonen, [email protected]

Stora Enso: Jukka KahelinUniversity of Helsinki: Raisa Haavikko, Vânia Moreira, Jari Yli-KauhaluomaUniversity of Tampere: Heikki Eräsalo, Mirka Laavola, Tiina Leppänen, Eeva Moilanen University of Turku: Sari Mäkelä,Lauri Polari, Niina Saarinen-Aaltonen, Emrah Yatkin UPM-Kymmene: Mika Hyrylä, Suvi PietarinenVTT Technical Research Centre of Finland: Kirsi Bromann, Tiina Nakari-Setälä, Mervi ToivariÅbo Akademi University: Annika Smeds, Stefan Willför


ABSTRACT

The project aimed at building a Finnish knowledge platform on the health-promoting potential of wood-derived compounds. The focus was on obesity-associated disorders, inflammatory diseases – especially (osteo)arthritis and allergy, and cancer, i.e. diseases that are common and increasingly prevalent in Finland and other Western countries.

The work demonstrated through various cell-based assays in vitro and in animal models in vivo that pine knot extract and its components have anticarcinogenic and immunomodulatory / anti-inflammatory properties and alleviate obesity-associated disorders. Mechanisms of action relevant to several diseases in the focus areas were identified, along with signalling mechanisms relevant to specific diseases.

The results contribute greatly to our general understanding of the effects of wood-derived compounds on human health. A unique, interdisciplinary research environment was created, providing an innovative knowledge platform enabling identification of novel applications for wood-derived compounds. The focus of the research on bioactive compounds derived from raw material sources available for large-scale production is also unique and increases Finnish competence for further exploitation of innovations in this field. The work paves the way for the development of high-value products from unexploited raw material sources as an integral part of the Finnish forest industry renewal process.

Keywords:pine knot extract, stilbenoids, obesity, adipose tissue, aromatase, breast cancer, prostate cancer, inflammation, arthritis, cartilage, allergy


1. Background

Nature is a rich source of bioactive compounds with therapeutic potential. It is estimated that about half of all drugs used in current medical treatment originate from plants and other natural sources. Nordic wood, especially conifer knotwood, is a rich source of compounds with health-promoting potential, such as polyphenols. Some of these compounds are similar to those found in other sources, such as edible plants. However, unlike in many other plants, the majority of polyphenolic compounds in wood are present in unconjugated form. In addition, polyphenol-rich knot material is available in large quantities as a side product of current industrial processes. Therefore, isolation of polyphenols from knot-enriched wood material in high quantities for further exploitation is feasible.

In this project, wood materials that are not important for pulp production were investigated. The research focused on health problems that are common and continuously increasing in Finnish and other Western populations: obesity, inflammatory diseases, especially osteoarthritis and allergy, and cancer. Obesity is a risk factor for development of metabolic syndrome, which is associated with type 2 diabetes and cardiovascular disease. In Finland, 60% of men and 50% of women are overweight or obese and 15% of men and 5% of women have metabolic syndrome. It has been estimated that half a million Finns have symptomatic osteoarthritis and about 30% of Finns suffer from various forms of allergic diseases. In addition, the incidence of certain cancers, in particular prostate cancer and breast cancer, is increasing in Western countries. By 2020, prostate cancer will constitute almost 50% of diagnosed cancer cases in men. Breast cancer is now, and will continue to be, responsible for more than 40% of cancer cases in women. Clearly, there is an urgent need for novel preventive means and treatment options against these diseases.

2. Objectives

The project was designed to contribute to building a globally competitive and unique Finnish knowledge platform on wood-derived compounds and their potential use as health-promoting agents.

The main objective was to identify and biologically characterize health-promoting wood knot and bark components with the aim of providing preclinical proof-of-concept level evidence in at least one of the focus areas: obesity-associated health problems, cancer, and inflammation, especially in relation to (osteo)arthritis and allergy. Each line of research began by recognizing relevant target(s) and continued by identifying effective extract(s) and/or compound(s) and further documenting the efficacy and preliminary safety of the selected materials in cell-based assays and/or animal experiments (in vivo disease models) in comparison with previously known control compounds.


Initially, the project involved three main raw materials: pine knot extract, spruce knot extract, and birch bark extract, and purified compounds of these, respectively. Based on the findings during the project, the work was further focused on pine knot extract and its compounds.

3.1 Obesity-associated health problems

The work was focused on obesity-induced endocrine disturbances caused by dysregulation of aromatase (CYP19A1) gene expression in white adipose tissue. Aromatase enzyme catalyzes the final step in oestrogen biosynthesis, i.e. conversion of androgens (androstenedione and testosterone) to estrogens (oestrone and oestradiol). White adipose tissue is an


important site for extragonadal aromatization and oestrogen production in man. To investigate pine knot extract effects in vivo, a humanized transgenic hAro-Luc reporter mouse was utilized. These mice express the full-length promoter region of the human aromatase gene, and can be used to study, for example, obesity-related factors that regulate aromatase expression in vivo. Obesity was induced by high-fat diet (HFD). Effect of pine knot extract on weight gain and body composition (adiposity) was determined and serum biomarkers of metabolic disturbance were measured. Obesity-induced adipose tissue inflammation was assessed using histological parameters in the tissue and by measuring adipose tissue proinflammatory cytokine levels. Adipose tissue cultures and bone marrow stromal cell cultures were established as novel tools to investigate factors that regulate aromatase expression in fat tissue.

3.2 Effects on prostate cancer

Anticarcinogenic potential, mechanism of action, and effective concentration range of pine knot extract and its main polyphenols and their metabolites were evaluated in in vitro assays by using human prostate and breast cancer cells and in vivo by using human cancer cell derived tumours in mice. in vitro, the effects of pine knot extract and compounds derived therefrom were tested for proliferation and apoptosis in androgen-independent human prostate cancer cells.

The effective dose of pine knot extract in vivo was assessed in prostate cancer tumours grown in mice. Growth of the tumours was followed and several biomarkers of tumour development and mechanisms of chemoprevention were assessed using morphological and biochemical analyses. Statistical multivariate analyses were applied to combine essential tumour growth markers and to identify effective dosing of pine knot extract.

3.3 Immunomodulatory / anti-inflammatory properties of pine knot extract and its compounds

To screen the anti-inflammatory potential of the pine knot extract and its known constituents, a widely used animal model in pharmacology research, i.e. carrageenan-induced paw inflammation, was used. The advantage of this model is that the effects can be compared to large background data on the activity of various anti-inflammatory compounds and series of other natural products. Also, the early inclusion of in vivo disease models in the evaluation of the biological activity of the extract and compounds of interest also gave a preliminary understanding of the bioavailability of the investigated materials. Furthermore, additional in vivo disease models were developed.

Cell-based assays were established to investigate the effects of the extract and its constituents on inflammatory gene expression in various inflammatory cells and in chondrocytes. The cell culture conditions were adjusted to mimic the inflammatory milieu in vivo and the cells were stimulated accordingly. In addition to appropriate cell lines, also primary human cell cultures were used when possible. Inflammatory gene expression was measured using standard procedures, e.g. quantitative real-time reverse transcriptase polymerase chain reaction (RT-qPCR) to detect mRNA levels, and Western blotting and enzyme-linked immunosorbent assay (ELISA) to measure protein levels. Also, an array-based approach was used where appropriate. Furthermore, the effects on intracellular signalling mechanisms were investigated using, inter alia, phosphoprotein and reporter gene assays.


4. Results

4.1 Pine knot extract attenuates obesity-induced metabolic disturbances and aromatase expression in adipose tissue

A mouse model of diet-induced obesity allowed us to study simultaneously multiple aspects of obesity-associated disorders in vivo. The effects of two dietary doses of a pine knot extract were investigated: “low dose” and “high dose”, which resulted in 32.5 mg/kg and 162.5 mg/kg body weight exposures, respectively. The “low dose” corresponds roughly to 1g daily intake of phenolics in a 70 kg human, i.e. an intake achievable through diet, while the “high dose” corresponds roughly to a 2g daily intake of pinosylvins in a 70 kg human. As expected, high-fat diet (HFD) increased body weight and body fat content in male mice compared to low-fat diet (LFD). Administration of pine knot extract for 8 weeks in HFD diet was well tolerated. Neither dose of pine knot extract significantly affected body weight gain or fat content. However, a higher dose of pine knot extract administered in the HFD diet decreased serum leptin, resistin, insulin, and fasting blood glucose levels, which are hallmarks of metabolic disturbances in humans.

Obesity induces chronic low-grade inflammation in adipose tissue, which causes numerous functional changes that promote the development of obesity-associated diseases. Formation of crown-like structures (CLS), i.e. necrotic adipocytes surrounded by macrophages, and increased levels of tissue proinflammatory cytokines are typical for obesity-related adipose tissue inflammation and dysfunction. The higher dose of dietary pine knot extract decreased the number of CLS in visceral and subcutaneous white adipose tissues (Figure 1) and reduced the tissue level of proinflammatory cytokines. In summary, these data indicate that dietary pine knot extract can attenuate obesity-induced adipose tissue inflammation in vivo.

Functional changes induced by obesity include induction of aromatase expression in adipose tissue, which, in turn is one of the key events leading to obesity-associated endocrine disturbances. Aromatase is an enzyme necessary for the production of oestrogen from androgens. Downregulation of oestrogen biosynthesis in adipose tissue has an impact, for example, on the prevention of obesity-related hypoandrogenemia in obese men and on the development of oestrogen-responsive breast cancer in postmenopausal obese women.

Figure 1. Density of crown-like structures (CLS) in visceral (A) and subcutaneous fat (B). LFD, low-fat diet; HFD, high-fat diet, HFD-PKE pine knot extract containing high-fat diet.


In humans, expression of aromatase is highly regulated through untranslated first exons that are differently utilized in different tissues. In rodents, the regulatory region of the aromatase gene is much less complex and, unlike in humans, the expression level in adipose tissue is very low. Therefore, we used a humanized transgenic hARO-Luc reporter mouse. These mice express the full-length regulatory region of the human aromatase gene attached to a luciferase (Luc) reporter gene.

In hAro-Luc mice, we showed that HFD-induced adiposity was associated with upregulation of Luc expression in adipose tissue. In line with this, adipose tissue Luc activity correlated positively with adipose tissue pro-inflammatory cytokine levels. Importantly, we demonstrated that dietary pine extract down-regulates the expression of human aromatase gene in subcutaneous fat of obese HFD-fed male mice (Figure 2).

In human adipose tissue, aromatase is mainly expressed in mesenchymal stromal cells (MSCs). We isolated and cultured MSC cells from hARO-Luc mouse bone marrow. As expected, Luc expression in MSCs was stimulated by glucocorticoids and proinflammatory cytokines through the aromatase gene promoter I.4. Interestingly,

aromatase gene reporter expression was modulated by pine knot extract in MSCs. In addition, other more indirect effects of pine knot extract seen in the in vivo setting may, at least in part, account for the downregulation of Luc expression in subcutaneous fat. In summary, these data suggest that pine knot extract affects oestrogen biosynthesis in male adipose tissue by modulating aromatase gene expression and may thus have potential to attenuate excessive oestrogen production in white adipose tissue of obese men.

4.2 Pine knot extract shows anticarcinogenic efficacy in prostate cancer

In this project, we showed that pine knot extract and its stilbenoids (monomethyl pinosylvin and pinosylvin), and lignans (matairesinol and nortrachelogenin) have antiproliferative and proapoptotic efficacy at ≥40 µM concentration in vitro (Figures 3 and 4). We also showed that pine extract (≥ 10 mg/L) and its stilbenoids enhance TRAIL-induced apoptosis already at ≥10 µM concentration in vitro (Figure 4B).

Four main compounds of pine extract B1, pinosylvin, monomethyl pinosylvin, matairesinol, and nortrachelogenin, were identified to account for the observed pine knot extract effects. These compounds, when combined in the same ratio as present in the extract, resulted in similar responses in the cell proliferation and apoptosis assays as the extract and, when tested as single compounds, were effective at ≥ 10µM concentrations (Figures 3C and 4). These data indicate that concentrations of ≥ 10 mg/L of pine extract or ≥ 40µM single pine extract compounds are required for inhibition of proliferation and induction of apoptosis in human prostate cancer cells in vitro. Figure 2. Aromatase reporter expression in male

subcutaneous white adipose tissue. LFD, low-fat diet; HFD, high-fat diet; HFD-PKE pine knot extract containing high-fat diet.


Two in vivo studies investigating the effects of pine knot extract on advanced androgen-independent prostate cancer in vivo were completed. Two doses of the extract were investigated: “low dose” resulting in 32 mg/kg/body weight exposure and “high dose” resulting in 162.5 mg/kg/body weight exposure. Three-week peroral exposure to high-dose pine knot extract B1 (52 mg of lignans and stilbenoids per kg body weight) showed anti-tumorigenic efficacy, demonstrated by multivariate analysis combining essential markers of tumour growth

(i.e. tumour volume, vascularization, and cell proliferation) (Figure 5).

Urine and serum analyses of “high dose” pine knot extract fed mice revealed significant absorption and metabolism of the extract compounds. In serum, high micromolar concentrations of nortrachelogenin, and monomethyl pinosylvin along with resveratrol were measured. Resin acids that are present in the pine knot extract were not detected in urine or serum samples.

Figure 3. Pine knot extract and its polyphenols inhibit proliferation of PC-3M-luc2 human prostate cancer cells. PKE, pine knot extract; LS, lignan-stilbenoid mixture containing pinosylvin (PS), monomethyl pinosylvin (MEPS), matairesinol (MR), and nortrachelogenin (NTG). Ab, abietic acid; RV, resveratrol. Figure published in PlosOne 2014, 9(4):e93764.

Figure 4. Pine knot extract and its compounds induce apoptosis in PC-3M-luc2 human prostate cancer cells (A) and sensitize these cells to TRAIL-induced apoptosis (B). PKE, pine knot extract; LS, lignan-stilbenoid mixture containing pinosylvin (PS), monomethyl pinosylvin (MEPS), matairesinol (MR), and nortrachelogenin (NTG). Ab, abietic acid; RV, resveratrol. Figure published in PlosOne 2014, 9(4):e93764.


In summary, these data demonstrate that orally-administered pine knot extract is well tolerated and inhibits the growth of prostate cancer tumours. The in vitro results indicate that pine knot extract derived lignans and stilbenoids and their metabolites account for the anticarcinogenic effect. The main polyphenol in pine knot extract, monomethyl pinosylvin, was identified as a novel bioavailable stilbenoid with anticarcinogenic potential against prostate cancer. In addition, ingestion of pine knot extract increased serum resveratrol concentrations markedly, indicating that the extract is a novel and cost-effective source of resveratrol, monomethyl pinosylvin and other potentially beneficial lignans and stilbenoids.

4.3 Pine knot extract and its stilbenoid constituents have anti-inflammatory properties in vivo

Inflammation is a protective response in the human body that aims to eliminate microbes and other offending agents as well as cancer cells and injured or necrotic tissue. However,

if aberrantly activated or regulated, prolonged or inappropriately focused (e.g. against host tissues, as in autoimmune diseases, or against normally harmless allergens, as in allergic diseases) the inflammatory response has considerable potential to cause harm and may lead to the development of inflammatory diseases, such as allergy, asthma or arthritis. Despite recent successes in the development of especially biological anti-inflammatory drugs, there are still considerable unmet needs in the treatment of inflammatory diseases. In fact, for most inflammatory diseases, current drugs can only offer symptoms relief and no curative or disease-modifying treatments are available. In the present research programme, we investigated the anti-inflammatory / immunomodulatory properties of pine knot extract and its constituents, especially in relation to disease mechanisms of (osteo)arthritis and allergic inflammation. Examples of the results are presented below.

To determine whether pine knot extract has anti-inflammatory potential in vivo, we utilized a widely-used animal model in

Figure 5. Relation between tumour vascularization, tumour cell proliferation and tumour volume. The hyperplane indicates that two of the groups (vehicle in black and high dose in green) were linearly separable when the three factors were considered together. Figure published in PlosOne 9(4): e93764.


pharmacology research, i.e. carrageenan-induced paw inflammation, in the mouse. Interestingly, the standard pine knot extract when given as a single oral dose of 100 mg/kg significantly attenuated carrageenan-induced inflammation, as shown in Figure 6A.

Following this finding, we investigated whether the stilbenoid constituents of the extract might be responsible for the anti-inflammatory bioactivity discovered in the pine knot extract. The extract is known to contain two principal stilbenoids, i.e. pinosylvin and monomethyl pinosylvin. These compounds had a clear anti-inflammatory effect in the carrageenan-induced paw inflammation model in the mouse, as can be seen in Figure 6B. Furthermore, they were more potent than resveratrol, an earlier known natural stilbenoid, which was used as a control compound. The efficacy of the two pine stilbenoids was comparable to that of the standard anti-inflammatory drug dexamethasone, although the pine stilbenoids were needed in higher doses to obtain the efficacy of the anti-inflammatory steroid dexamethasone.

4.4. Pine knot extract and its stilbenoid constituents down-regulate inflammatory gene expression in inflammatory cells and in chondrocytes

In order to understand the anti-inflammatory properties of the pine knot extract and its stilbenoid constituents at the cellular level and their effects on the mediators of inflammation, we investigated their effects on various cells participating in different types of inflammation. Selected effects of the pine knot extract and its stilbenoids pinosylvin and monomethyl pinosylvin on macrophages and chondrocytes in cell culture conditions mimicking inflammatory milieu are presented below.

MacrophagesMacrophages are key inflammatory cells that are differentiated in tissues from blood monocytes. Macrophages have various functions in the first line of defence against microbes and in the development of specific lymphocyte-mediated immune response as well as in the regulation of inflammatory and repair mechanisms through the production of a variety of inflammatory mediators.

Figure 6. Anti-inflammatory properties of pine knot extract and its stilbenoid constituents pinosylvin and monomethyl pinosylvin in carrageenan-induced paw inflammation in the mouse. In (A) the pine knot extract (100mg/kg) was dosed orally and in (B) pinosylvin (100 mg/kg) and monomethyl pinosylvin (100 mg/kg) as well as control compounds resveratrol (100 mg/kg) and dexamethasone (2 mg/kg) were given intraperitoneally 2h prior to the inflammation was induced by injecting carrageenan into the paw. Results are expressed as mean ± SEM, n=6.


In the present study, we investigated the effects of pine knot extract and its stilbenoids on inflammatory gene expression in macrophages activated through the TLR4 pathway by adding bacterial endotoxin lipopolysaccharide (LPS) into the culture. As seen in Table 1 below, pine knot extract inhibited the production of inflammatory factors interleukin 6 (IL-6), monocyte chemotactic protein 1 (MCP-1) and inducible nitric oxide synthase (iNOS) and its product nitric oxide (NO) in a dose-dependent

manner. Interestingly, expression of these inflammatory genes was also inhibited by pinosylvin and monomethyl pinosylvin (stilbenoids present in the extract) as well as by resveratrol, which was used as a control stilbenoid.

ChondrocytesOsteoarthritis (OA) is the most common joint disease worldwide. The disease affects the entire joint causing disability and pain.

Table 1. The effects of pine knot extract and its components pinosylvin and monomethyl pinosylvin as well as the control stilbenoid resveratrol on nitric oxide (NO), interleukin 6 (IL-6) and monocyte chemotactic protein 1 (MCP-1) production and inducible nitric oxide synthase (iNOS) expression in activated macrophages. The results are expressed as mean ± SEM; * indicates p<0.05 and ** = p<0.01.


The primary cause of OA remains unknown. Degradation of the cartilage matrix is the most important feature of OA pathology. The degradation is caused by an imbalance between catabolic and anabolic mediators in the joint, and by the inflammatory mediators produced by the cells in the cartilage and synovial membrane. Matrix metalloprotease enzymes, especially MMP-1, MMP-3 and MMP-13, are produced by chondrocytes in the OA cartilage and play a significant role in cartilage matrix degradation. (Figure 7) Compounds capable of inhibiting the synthesis of MMP enzymes are being intensively sought and investigated as potential future drugs to alleviate cartilage degradation associated with OA.

In the present study, we investigated the effects of pine knot extract and its stilbenoid constituents on the production of various catabolic, anabolic and inflammatory factors in human chondrocytes by using a chondrocyte cell line and primary human chondrocytes isolated from human cartilage tissue from OA patients removed during joint replacement operations. As can be seen in Table 2, the

pine knot extract inhibited the production of the three MMP enzymes typical of OA pathogenesis (i.e. MMP-1, MMP-3 and MMP-13) in a dose-dependent manner. The effect could be due to the stilbenoids present in the extract, because pinosylvin and monomethyl pinosylvin, as well as the control compound resveratrol, also had an inhibitory effect on the synthesis of MMP enzymes.

4.5 Pine knot extract and its stilbenoid constituents attenuate the activity of two major inflammatory signalling pathways

The cell culture studies were extended in order to further understand the mechanisms of action of the pine knot extract and its stilbenoid constituents in inflammatory cells. Nuclear factor-kB (NF-kB) is an important inflammatory transcription factor that is known to be activated by various inflammatory stimuli. Once activated and translocated to the nucleus, NF-kB is able to augment the transcription of various inflammatory genes, including many of those found to be regulated by the pine knot extract and its constituents. Inhibitors of NF-kB are under development for the treatment of arthritis and asthma, and other inflammatory diseases.

To investigate the effects of the pine knot extract and its stilbenoid compounds on NF-kB activation and NF-kB dependent transcription, we generated two different types of reporter gene containing cell lines. The cells were transfected with a reporter gene construct, in which the luciferase gene (which is not normally expressed in human cells) was linked to an NF-kB dependent promoter, as shown in Figure 8A. In those cells, the promoter activity could be enhanced by inflammatory factors which are known to activate NF-kB (e.g. LPS, IL-1 or TNF), and this in turn could be measured as increased luciferase mRNA levels or as

Figure 7. Pathological changes and mechanisms in osteoarthritis.


Table 2. Effects of pine knot extract and its components pinosylvin and monomethyl pinosylvin as well as the control stilbenoid resveratrol on the production of matrix metalloprotease enzymes MMP-1, MMP-3 and MMP-13 in a human chondrocyte cell line activated with the inflammatory cytokine interleukin 1. The results are expressed as mean ± SEM; ** indicates p<0.01.

Figure 8. Effects of pine knot extract and its components pinosylvin and monomethyl pinosylvin on NF-kB mediated transcription. Figure A is a schematic presentation of the cell model generated and used in the studies. Figure B shows the effects of the pine knot extract and its stilbenoid constituents pinosylvin and monomethyl pinosylvin as well as the effects of the control stilbenoid resveratrol and an earlier known NF-kB inhibitor MG132 on NF-kB mediated transcription, which was measured as luciferase activity in the model described in A. The results are expressed as mean + SEM; ** indicates p<0.01.


increased luciferase enzyme activity in the cells.

Figure 8B shows an example of the results of the studies in which the effects of the pine knot extract and its stilbenoid constituents on NF-kB mediated transcription were investigated. The pine knot extract significantly inhibited NF-kB mediated transcription, and the effect was shared by pinosylvin and mono-methyl pinosylvin as well as by resveratrol, which was used as a control stilbenoid.

Heme oxygenase 1 (HO-1) is a factor known to play a role in cellular defence against oxidative stress and to have anti-inflammatory and cytoprotective properties. Those effects are, at least partly, mediated through inhibition of NF-kB activation. Therefore, we aimed to investigate whether the pine knot extract and its stilbenoid compounds, which were able to inhibit NF-kB mediated transcription, could also augment the expression of HO-1.

As shown in Figure 9A, the pine knot extract enhanced the expression of the cytoprotective

and anti-inflammatory factor HO-1 in a dose-dependent manner. Interestingly, pinosylvin and monomethyl pinosylvin also enhanced cellular HO-1 levels, at least when used at 100 µM concentrations, while the effects of resveratrol were minor. The increased HO-1 expression may contribute to or mediate the inhibitory effect on NF-kB mediated transcription caused by the pine knot extract and its stilbenoid constituents.

PI3K/Akt is another major intracellular signalling pathway involved in inflammation and cancer. PI3K (phosphoinositide 3 kinase) activation leads to phosphorylation of Akt, which regulates the activation of many inflammatory genes. Therefore, we were curious to investigate whether the pine knot extract and its bioactive stilbenoid constituents have an effect on the PI3K/Akt pathway in activated inflammatory cells.

As expected, inflammatory stimuli, such as LPS, activated the PI3K/Akt pathway in macrophages, measured as Akt phosphorylation. The pine knot extract inhibited Akt phosphorylation in those conditions, and the effect was shared by

Figure 9. Effects of (A) pine knot extract and (B) its components pinosylvin and monomethyl pinosylvin as well as the control stilbenoid resveratrol on heme oxygenase 1 (HO-1) expression in macrophages activated with the inflammatory stimulus LPS. Hemin was used as an earlier recognized positive control compound to enhance HO-1 expression. The results are expressed as mean + SEM; *indicates p<0.05, ** = p<0.01 and *** = p<0.001.


Figure 10. Effects of pine knot extract and its components pinosylvin and monomethyl pinosylvin as well as the control stilbenoid resveratrol on the PI3K/Akt pathway, measured as Akt phosphorylation. LY294002 was used as a control compound known to inhibit PI3K. The results are expressed as mean + SEM; **indicates p<0.01.

the pine stilbenoids pinosylvin and monomethyl pinosylvin as well as by resveratrol, which was used as a control stilbenoid. An example of the results is shown in Figure 10. These results suggest that stilbenoids are responsible for the inhibitory effect of the pine knot extract on the PI3K/Akt pathway. Furthermore, the studies of NF-kB and the PI3K/Akt pathway strongly suggest that those mechanisms may explain, at least partly, the anti-inflammatory properties of the pine knot extract and its bioactive stilbenoids on inflammatory gene expression and inflammatory responses in vivo.


This project contributes to building a globally competitive and unique Finnish knowledge platform on wood components and their potential use as health-promoting agents. The

diseases targeted by the research (obesity, osteoarthritis, allergy, and cancer) are major public health problems of increasing incidence, the impact of which at the individual and societal level is growing substantially along with their cost to the health care system. Thus, there is a clear need for new preventive means and treatment options for these diseases. In the present project, several health targets and product applications were identified for further development.

Finnish wood resources hold vast potential for the development of novel and innovative high-value products. This project identified wood components that are not yet exploited but have potential to be utilized for the development of health-promoting high-value products, such as functional foods, nutraceuticals, natural drugs or pharmaceuticals. The focus was on wood extracts rich in bioactive components and with broad exploitation potential. This opens opportunities for the development of new value chains and business concepts from raw material supply to further product development and commercialization, thus helping drive the renewal of the forest industry.


6. Networking

The research was carried out in cooperation by the partners, see Table 3. the work package partners and their roles.

Table 3. Research partner organizations and their research roles.

Work package partner Role of the participating organization


University of Helsinki Chemical modification of wood-derived

compounds.

University of Tampere Investigation of bioactivity of wood-derived

extracts and compounds in inflammation with

special reference to (osteo)arthritis and allergy

University of Turku WP leader. Investigation of bioactivity of wood-

derived extracts and compounds in obesity and

cancer


VTT Biotechnological modification of wood-derived

compounds.

ÅboAkademiUniversity Composition analysis of wood extracts and their

metabolites.



Bromann, K., Toivari, M., Viljanen, K., Vuoristo, A., Ruohonen, L. and Nakari-Setälä, T. Identification and characterization of a novel diterpene gene cluster in Aspergillus nidulans. Plos One, 2012, 7(4):e35450.

Eräsalo, H., Laavola, M., Hämälainen, M., Leppänen, T., Nieminen, R. and Moilanen, E. PI3K inhibitors LY294002 and IC87114 reduce inflammation in carrageenan-induced paw edema and down-regulate inflammatory gene expression in activated macrophages. Basic and Clinical Pharmacology and Toxicology, 2014 in press.

Laavola, M., Nieminen, R., Leppänen, T., Eckerman, C., Holmbom, B. and Moilanen, E. Pinosylvin and monomethyl pinosylvin, constituents of extract from knots of Pinus sylvestris, reduce inflammatory gene expression and inflammatory response in vivo. Submitted for publication.

Polari, L., Ojansivu, P., Mäkelä, S., Eckerman, C., Holmbom, B. and Salminen, S. Galactoglucomannan extracted from spruce (Picea abies) as a carbohydrate source for probiotic bacteria. Journal of Agricultural and Food Chemistry, 2012, 60(44):11037-43.

Qiu, Yaqi. Dietary factors affecting human aromatase gene reporter expression in normal tissues and cancer; Focus on mammary gland and breast cancer. Master’s thesis. ÅboAkademi University, Dept. of Biosciences. 2013.

Yatkin, E., Polari, L., Laajala, TD., Smeds, A., Eckerman, C., Holmbom, B., Saarinen, NM., Aittokallio, T. and Mäkelä, S. Novel lignan and stilbenoid mixture shows anticarcinogenic efficacy in preclinical PC-3M-luc2 prostate cancer model. PlosOne 2014, 9(4):e93764.


THERMOPLASTIC LIGNIN AND REINFORCING CELLULOSE FIBER

COMPOSITESFOR ADVANCED BIOCOMPOSITE APPLICATIONS

CONTACT PERSON – Work Package 3 leader Antti Ojala, [email protected]

Metsä Fibre: Pirkko LiiasNovoplastik: Johannes Heiskanen, Pietari Heiskanen, Merja Lehto Stora Enso: Kalle Ekman, Janne Pynnönen University of Helsinki: Juha Fiskari, Ilkka Kilpeläinen, Maarit LahtinenUniversity of Jyväskylä: Markku Kataja, Arttu Miettinen UPM-Kymmene: Mika Hyrylä, Eeva Jernström VTT Technical Research Centre of Finland: Stina Grönqvist, Karita Kinnunen-Raudaskoski,Jani Lehmonen, Kalle Nättinen, Lisa Wikström


ABSTRACT

The processability and thermomechanical performance of thermoplastic lignin composites were studied. The aim was to develop processes and materials to enable the production of composites with high lignin (>50% of the matrix) and wood-based material content. Processing of the composites was carried out using twin-screw extrusion and injection moulding techniques. The focus was on softwood kraft lignin, but also lignins from alkali and hot water extractions were evaluated. To reduce the high glass transition temperature of kraft lignin and to enhance the ductility of the composites, both external as well as internal plasticization routes were employed. Thermomechanical (TMP) and bleached (Fusellu) softwood pulps were used to improve the processability and mechanical performance of the composites. Materials reinforced with Fusellu demonstrated slightly higher mechanical performance than the TMP-reinforced composites. The best performance was obtained using a mixture of acetylated starch (Ac-starch) and triethyl citrate (TEC) as the external plasticizer and Fusellu reinforcements. Acetylation, hydroxypropylation and palmitic acid grafting methods were used to internally plasticize kraft lignin. Of the internally plasticized materials, composites with the Ac-starch-TEC plasticized acetylated lignin (Acet-L) matrix and Fusellu reinforcement gave the best mechanical properties. The fabricated composites had a tensile strength of 35 MPa and modulus of 6.6 GPa. The overall content of the wood-based materials was 70%. The structure of the composites was characterized using X-ray computed microtomography (X-µCT). The focus of the study was on fibre dispersion and breakage. The fibre dispersion in the thermoplastic lignin matrices was observed to be good. However, the processing conditions had a significant detrimental impact on fibre length.

Keywords:thermoplastic lignin, injection moulding, thermomechanical pulp, bleached pulp,X-ray microtomography


1. Background

Modern biorefineries (e.g. pulping mills and bioethanol plants) using lignocellulosic feedstocks produce as a by-product large amounts of lignin, which is currently almost exclusively burned for energy. For example, it is estimated that the annual global production of lignin in kraft pulp mills exceeds 50 million tonnes. Lignin is therefore a vastly underutilized resource.

Global annual plastics production currently stands at around 250 million tonnes, of which thermoplastic materials account for 65%. By far the largest thermoplastic segments are petroleum-based PE, PP and PVC at 33%, 21% and 17%, respectively. The share of non-petroleum based materials remains highly marginal (<1% of all produced thermoplastics). Furthermore, current renewable materials are almost exclusively based on starch, and thus compete directly with global food production. There is therefore a clear demand for low-cost non-food based plastics from renewable feedstock. One of the most promising candidates is thermoplastic lignin – the world’s second most abundant polymer after cellulose.

Initial trials on kraft lignin were carried out in the FuBio programme (2009-2011), in which the thermoplastic properties and processability of lignin were briefly evaluated. Based on these promising results, it was decided to dedicate a separate work package to thermoplastic lignin composites under the FuBio Joint Research 2 programme launched in 2011.

2. Objectives

The aim of the research was to develop technologies and competences enabling the production of novel biocomposite materials in which wood-derived lignin and cellulose fibre are the main constituents. For best performance, both the fibre and lignin fractions were modified. Modification of the lignin fraction included internal and external plasticization for improved ductility and processability. Chemical modifications of the fibres were targeted at enhancing bonding with the matrix lignin and the subsequent reinforcement effect.

The target application sectors were construction and furniture.


The main task was to evaluate the potential of lignin as a thermoplastic matrix material in wood pulp reinforced composites. The focus was on the processability of lignin and the mechanical performance of the composites. Softwood kraft lignin (LignoBoost™) was used as the main component, in addition to which hot water extracted and alkali lignins and commercial lignins were also tested. The reinforcing wood fibres were chemically bleached softwood pulp (Fusellu) and thermomechanical softwood pulp (TMP) supplied by Metsä Fibre and UPM, respectively. Additionally, PHWE extracted and ionic liquid extracted fibres supplied by Metla were evaluated in the composites.

The research was divided into the following four tasks:

• Developmentofligninmodifications(“internal plasticization”) with the goal of reducing glass transition temperature (Tg) and brittleness.


• Chemicalandenzymaticmodificationoffibres with the goal of enhanced fibre and matrix lignin interaction.

• Manufacturingofthecomposites.Twosimultaneous routes were employed: i) selection and development of the actual processes and equipment needed for manufacturing lignin-fibre composite compounds and ii) iterative development of the lignin-fibre composite compounds at laboratory and pilot scale.

• Structuralcharacterizationofthecomposites using X-ray computed microtomography (X-µCT)

Figure 1 shows the key processing steps involved in producing the thermoplastic lignin composites. In the first step, the extracted lignin was plasticized to improve its processability (i.e. to reduce the glass transition temperature, Tg). Two different approaches, internal and external plasticization, were used. In the second step, plasticized lignin was processed with wood pulp to produce wood fibre reinforced thermoplastic lignin composites. Finally, the composite parts were produced by injection moulding.

4. Results

4.1 Structural and thermal characterization of lignins

The main lignin source was softwood kraft lignin LignoBoost (LB, purity 96.7%) supplied by Metso. Table 1 shows the chemical composition of LB determined by the 31P NMR method. The total amount of hydroxyl groups (mmol/g) was 5.28, of which 3.39 were phenolic and the rest were aliphatic, condensed or carboxylic acid. Additionally, the molecular weight (Mw) distribution for LB as well as for alkali lignin (AL) and pressurized hot water extracted lignin (PHWE) were determined using GPC methods. The results of the GPC analyses are presented in Table 2. LB has the highest Mw at 4450 Da. AL and PHWE have Mws of 2800 and 1700 Da, respectively, clearly lower than that of LB.

Thermal properties of the lignin samples were analysed using differential scanning calorimetry (DSC). Figure 2 and Table 3 present the results of the DSC scans. All lignin samples show two glass transition points (Tg) at 60-70 °C (Tg1) and at 100-180 °C (Tg2). LB has the highest Tg2 at 152.5

Figure 1. Schematic presentation of the processing of lignin composites.

Lignin AliphaticOH

Carboxylic acid

Condensed S G Ca PhenolicOH

TotalOH

LB 1.60 0.29 1.46 0.00 1.89 0.05 3.39 5.28

Table 1. Amount of different hydroxyl group species (mmol/g) in lignin samples as detected by NMR .


Sample Mn (Daltons) Mw (Daltons) Polydispersity

LB 2289 4450 1.94

AL 1800 2800 1.51

PHWE 1200 1700 1.36

Table 2. Molar mass distributions of the LB, AL and PHWE samples.

Figure 2. Second heating cycle scans for the AL (red), PHWE (green) and LB (blue) samples. The heating ramp was 10 °C/min from -50 °C to 200 °C.

Lignin Description Tg1 / °C Tg2 / °C Tdeg10% / °C

LB Softwood kraft lignin 68.5 152.5 328.5

PHWE Hardwood PHWE lignin 71.8 120.8 282.0

AL Softwood alkali lignin 61.0 102.4 282.5

PL Palmitic acid modifiedLB lignin

n/d n/d 245.1

HPL HydroxypropylatedLB lignin

63.0 112.4 300.9

Acet-L Acetylated LB lignin 46.0 101.4 272.6

Table 3. Results of the thermal analyses for the LB, AL and PHWE lignins as well as for the internally plasticized PL, HPL and Ace-L lignins.


Figure 3. Second heating cycle scans for the HPL (green), PL (red) and Acet-L (blue) samples. The heating ramp was 10 °C/min.

°C, whereas AL and PHWE have Tg2 at 102.4 and 120.8 °C, respectively. Note that the Tg2 of the PHWE sample is substantially higher than that for AL, although the Mw of PHWE is clearly lower.

The thermal stabilities of the lignin samples were determined using thermogravimetric analysis (TGA). The results are presented in Table 3. LB has the highest thermal stability of the analysed samples. The point at which 10% of the sample weight is lost (Tdeg10) is observed at 329 °C. AL and PHWE are, in turn, less sta-ble with Tdeg10 at 283 and 282 °C, respectively. Compared to the DSC analyses, it is clear that the degradation temperatures for the lignin samples are significantly higher than the sof-tening temperatures. The degradation tem-peratures are also significantly higher than the corresponding temperatures for the employed wood pulps (approx. 220 °C). Hence, we can conclude that the lignins are suitable for melt processing with the wood pulps.

Together with external plasticization, inter-nally plasticized lignins were evaluated as the matrix materials in the composites. The aim of the internal plasticization was to reduce the high Tg of the native kraft lignin (LB) and, successively, to improve the ductility of the composites. Palmitic acid grafted lignin (PL), hydroxypropylated lignin (HPL) and acetylat-ed lignin (Acet-L) were selected as the main target modifications. The samples were syn-thesized using LB as the starting material. Based on the 31P NMR analyses the degree of substitution (DS) of the aromatic and alkyl hydroxyl groups for HPL, PL and Acet-L were 100, 88, and 97%, respectively.

The results of the TGA analyses for the modified lignins are presented in Table 3. Compared to native LB, the modified lignins have somewhat reduced thermal stabilities. The Tdeg10 was observed for the PL, HPL and Acet-L samples at 245, 301 and 273 °C, respectively.


The main purpose of the internal plasticization was to reduce the softening temperatures of lignin and hence improve the ductility of the composites. Figure 3 presents the results of the DSC scans for the modified lignins. The PL sample gives a complex graph with endo-thermic peaks at 0.4 °C and 47.3 °C and a very weak endothermic decline at 78.7 °C. It is un-clear whether some of these peaks are related to Tg. In the HPL and Acet-L scans, the clear endothermic declines at 112 and 101 °C, respec-tively, are attributed to Tg2. Furthermore, the two weak endothermic declines at 63 (HPL) and 46 (Acet-L) °C are attributed to Tg1. 4.2 Processing of lignin composites

Thermoplastic lignin composites were processed at two different size scales. Preliminary testing of the new materials and the material combination was carried out at the laboratory scale. Lignin, additives and fibres were thermally mixed in a Brabender Plasticorder batch mixer with a sample size of 44 g. The test specimens (see Figure 4) were then injection moulded using a MiniJet (ThermoHaake) and mechanically tested using a tensile tester (Instron).

Based on the preliminary laboratory scale results, the most interesting materials and their combinations were further evaluated using a continuous pilot scale process. Figure 5 shows the main processing steps in the manufacture of thermoplastic lignin composites at the pilot scale. Compacting was used to reduce the large volume of the pulps prior to compounding. The fibres were compacted into pellets approximately 15mm x 4mm in size, after which the fibre pellets, lignin, plasticizers and possible additives were thermally mixed to form uniform compounds. The compounds were prepared by using a twin-screw extruder (ZE25 x 48D, Berstorff GmbH). Finally, the compounds were injection moulded as test specimens using an Engel (ES 200/50 HL) and mechanically tested using an Instron tensile tester.

The charpy method was used to measure the impact strengths for the un-notched samples.

4.3 Thermoplastic lignin composites

Various external plasticizers for LB were tested in laboratory scale experiments. Based on the preliminary trials, polyethylene

Figure 4. Lignin composite in a Brabender batch mixer (a). Two tensile test specimens prepared at laboratory and pilot scale (b).

b)a)

Figure 5. Processing steps in the pilot scale manufacture of lignin composites.


glycol (PEG) was observed to give the best processability and tensile properties. Samples with TMP and Fusellu reinforcements were produced. The TMP-reinforced composites had somewhat higher mechanical performance than that of Fusellu. To further evaluate the processability of kraft lignin, externally plasticized composites based on LB, PEG and TMP reinforcements were fabricated at the pilot scale. To determine optimal LB lignin/TMP/PEG ratios, a series of composites with different compositions were fabricated. The best performing composites had a composition (by weight) of 49% LB lignin, 30% TMP and 21% PEG. These composites had a tensile strength of 28.8 MPa, Young’s modulus of 5.1 GPa and impact strength of 4.6 kJ /m2. Further increases in TMP content did not improve the tensile properties. Additionally, higher PEG contents (relative to lignin) resulted in improved impact strength but also significant reductions in tensile strength and stiffness.

PEG has two major drawbacks as a plasticizer: 1) it is of non-renewable origin and 2) it is highly

water soluble, which significantly reduces the weather resistance of the composites. Therefore, a new plasticizer for lignin based on a mixture of starch acetate (Ac-starch) and triethyl citrate (TEC) was developed. Previous studies with flax fibre reinforced Ac-starch-TEC composites have shown that the mixture has excellent compatibility with cellulosic fibres. In addition, Ac-starch and TEC can be obtained from renewable sources, and they are scarcely soluble in water. Preliminary evaluation of the optimal Ac-starch/TEC ratio was performed at the laboratory scale followed by more comprehensive pilot scale trials. The optimal Ac-starch/TEC ratio was found to be approximately 60/40 (by weight).

After the preliminary evaluations, the effect of lignin content was studied at the pilot scale (Figure 6). In all prepared samples, the Ac-starch/TEC ratio was 60/40 and the TMP fibre content was 20%. Tensile strength of the composites was found to decrease with increasing lignin content. The composites with 32% lignin demonstrated a tensile strength

Figure 6. Effect of lignin content on the mechanical properties of Ac-starch-TEC plasticized composites. The Ac-starch/TEC ratio was 60/40 and the TMP content was 20% in all cases. The best PEG-plasticized composites (49% LB lignin/30% TMP/21% PEG) are shown as a reference.


of 26.6 MPa, whereas increasing the lignin content to 49% decreased the strength to 19.6 MPa. In turn, stiffness of the composites improved with increasing lignin content. Compared to the PEG-plasticized composites, the tensile strength of the Ac-starch-TEC plasticized composites were comparable.

Water absorptionTo study weather stability, a series of water absorption trials was performed for the injection moulded PEG (49% LB/21% PEG/30% TMP) and Ac-starch-TEC (45% LB/27.5% Ac-starch-TEC/27.5% TMP) plasticized composites. The composites were immersed in water and the weight changes were monitored over a period of two weeks. Figure 7 presents the results of the trials. During the first week, the weight of the PEG-plasticized composites increased rapidly until levelling off at 41% after 1.5 weeks. In turn, Ac-starch-TEC plasticized composites demonstrated very slow water uptake with a weight increase of only 6% after two weeks. To evaluate the dissolved material, the used water bath solutions were collected and evaporated. The residual solids were weighted and analysed with FTIR. The amount of dissolved material from the PEG-plasticized samples was 7.5%

of the initial dry weight. Furthermore, the FTIR analysis revealed that the dissolved solid was almost exclusively PEG. Thus, the large water uptake (weight increase) of the PEG-plasticized composites can be explained by the dissolution of the highly water soluble PEG component and the successive formation of a hollow structure that readily absorbs water. In turn, the dissolved solid from the Ac-starch-TEC plasticized samples was only 0.3% of the initial dry weight.

Effect of lignin type on performance The properties of AL and PHWE lignins supplied by Metla and a commercial alkali lignin (GV) from GreenValue in composites were evaluated. GV is extracted from plant feedstock by an aqueous sodium hydroxide method. Identical samples with TMP reinforcement and PEG plasticization of AL and GV were fabricated at laboratory scale. Figure 8 presents the results of the tensile tests. Compared to the composites with the LB matrix, the samples with GV and AL show reduced tensile strengths and modulus, whereas the strains at break are significantly increased. The notably high ductility of the AL composites is attributed to the low Tg2 of the material.

Figure 7. Results of the water absorption tests for PEG (diamonds) and Ac-starch-TEC (circles) plasticized composites.


Moreover, composites with Ac-starch-TEC plasticization were fabricated at laboratory scale using AL and PHWE lignins as the matrix materials. Compared to PEG plasticization, Ac-starch-TEC plasticized AL composites show higher tensile strength and modulus, whereas the strain at break is lower. In turn, the Ac-starch-TEC plasticized PHWE composites have notably higher tensile strength and modulus with respect to the

corresponding AL composites. The viscosity of the LB composites was so high that the composites could not be injection moulded.

Following the promising laboratory scale tests, AL and PHWE lignins were also tested at the pilot scale. Figure 9 shows the results for the Ac-starch-TEC plasticized composites. The properties are highly similar between the samples. The only exception is observed in

Figure 8. Tensile properties of the PEG and Ac-starch-TEC plasticized composites with different lignin matrices.

Figure 9. Mechanical properties of composites based on the LB, AL and PHWE matrices. In all cases, the composition was 41% lignin/39% Ac-starch-TEC/20% TMP. The composites were fabricated at the pilot scale.


impact strength, which was 2.1 and 2.3 kJ/m2 for the AL, PHWE composites, respectively, and 1.6 kJ/m2 for the composites with the LB matrix.

Composites based on acetylated lignin In addition to external plasticization, internally modified (i.e. internally plasticized) lignins were also tested in composites. The selected modification methods were acetylation, hydroxypropylation and palmitic acid grafting. The aim of internal plasticization was to replace the external plasticizers or, at least, to significantly reduce the need for external plasticizers. The composites based on palmitic acid grafted lignin (PL) were processable without external plasticizers. However, processing of the composites based on the acetylated (Acet-L) and the hydroxypropylated (HPL) lignin matrices was not possible without external plasticizers. Moreover, both modifications required the same amount of Ac-starch-TEC plasticizer as the composites based on unmodified LB.

In the preliminary laboratory scale tests, the best performance was obtained with Acet-L composites. This material was therefore selected for closer evaluation in pilot scale trials using Ac-starch-TEC as the external plasticizer. Fusellu was employed as the reinforcing fibre as it gave generally better results in Ac-starch-TEC plasticized systems compared to TMP. The composites with the Acet-L matrix had higher improved properties compared to the unmodified LB composites. Composites with 36% Acet-L, 34% Ac-starch-TEC and 30% Fusellu had the best mechanical properties, and their recorded tensile strength (35 MPa) was the highest of all the injection moulded composites. In addition, their impact strength (3.8 kJ/m2) was the highest of all the Ac-starch-TEC plasticized composites.

Composites based on chemically modified TMPThe low water stability of typical lignin

plasticizers such as PEG is a significant limitation on the wider utilization of the material. One possibility for improving the stability is to reduce the content of free PEG. However, a simple reduction of PEG content in the composites leads to significantly weaker tensile properties, such as lower ductility and increased brittleness. To circumvent this problem and to optimize the compatibility between the fibres and the matrix, TMP was chemically cross-linked with PEG diepoxide [PEGDE, polyethylene glycol (200) diglycidyl ether, Polysciences, Inc.] to produce PEG-grafted TMP.

For the composite trials, several samples with different degrees of cross-linking were prepared. The results of tensile tests on two samples with a lower (6%) and a higher (20%) degree of cross-linking are shown in Figure 10. The details of the compounded samples are presented in Table 4. Based on these results, the tensile strengths were significantly increased when the sample had a higher degree of cross-linking (20%). Furthermore, a lower degree of cross-linking (6%) had a positive effect. In comparison with the reference sample, after PEG grafting the tensile strength improved by 45%. Simultaneously, the amount of free PEG could be reduced by 4 wt% without compromising ductility or stiffness, which improved by 23%.

4.4 Structural characterization of the composites using X-ray microtomography

The utilized processing techniques (twin-screw extrusion and injection moulding) expose the fibres to harsh conditions such as high temperatures and shear forces. The process is particularly harmful to cellulosic fibres, which have been observed to undergo substantial decomposition. Reduced fibre length is especially problematic as this is one of the key


factors defining the mechanical performance of the reinforced composites. Particularly challenging is the low impact strength, which limits the utilization of lignin composites in more demanding applications, such as in vehicles. Therefore, the effect of processing conditions on the reinforcing fibres in the thermoplastic lignin composites was analysed using X-ray computed microtomography (X-µCT). Particularly, the focus was on fibre length and distribution in the matrix.

The characterization study began with an initial search of imaging parameters. This process, done using thermoplastic lignin composite materials, indicated that the image contrast between fibre and matrix is sufficient to enable the phases to be differentiated from each other. Figure 11 shows a composite material sample consisting of a PEG-plasticized LB matrix and 25% TMP. The fibres are clearly

visible and some fibre agglomerates (white lumps) can be seen indicating poor dispersion.

Before quantitative assessments of the geometrical structure of the samples can be made, the images must be pre-processed to remove spurious noise. Different methods for noise removal filtering were tested. The best of these appear to be simple Gaussian filtering and variance weighted mean filtering. In the first method, the final value of a pixel is the weighted mean of the surrounding pixel values where the weight function is a Gaussian. In the second method, the mean around a pixel is weighted using the inverse of local variance around the pixel, thus reducing blurring of sharp edges. The method of choice depends on the quality of the original image and the type of analysis carried out on the sample.

The methods of analysis found to be most

Figure 10. The effect of cross-linking with PEG diepoxide on strength properties of the compounded material at laboratory scale. The amounts of different components and the degree of cross-linking are shown in Table 4.

Sample Lignins wt-% Plasticiser wt-% Fibre wt-%

FU88, Ref LB 56 PEG 24 TMP 20

FU112 LB 56 PEG 22,8 TMP-g-PEG (6%) 21,2

FU113 LB 56 PEG 20 TMP-g-PEG (20%) 24

Table 4. Composition of the compounded samples cross-linked with PEG diepoxide (strength test results are presented in Fig. 10).


useful for the materials include measurement of fibre length using a granulometric technique, measurement of fibre orientation using the structure tensor method, and measurement of fibre cross-sectional shape parameters using a novel technique where local fibre orientation information is used to construct 2-dimensional slices of fibres in the direction of their orientation. The slices can then be used to measure e.g. the cross-sectional area of fibres or shape descriptors of the fibre cross-sections (e.g. circularity or

approximate diameter). All of the methods yield the distribution of the measured value (e.g. fibre length distribution).

The homogeneity of the fibre volume fraction in the PEG-plasticized LB matrix with 20% TMP tensile test specimens was analysed (See Figure 12). Depending on the sampling position, the measured fibre volume fraction was found to increase such that the volume fraction is lowest close to the injection cavity and highest at the other end of the specimen.

Figure 12. Volume fraction of fibres in different locations (a) in the thermoplastic lignin test specimen (b). The injection direction was from “broken end” to “non-broken end”.

b)

a)

Figure 11. X-µCT images of a PEG-plasticized lignin/20% TMP sample processed at laboratory scale. Cross-section of the sample (a) shows fibres visible as brighter areas. Two 3D visualizations of the same sample with different, b) low and c) high resolution. Large white areas in the middle image are fibre agglomerates.

a) b) c)


Following these results, further tomographic analyses were conducted on samples taken from the middle part of the specimen.

It was observed that the aspect ratio of the fibres decreases significantly when the intact fibres are processed as injection moulded specimens. A good example of the results is given in Figure 13, where the aspect ratio distributions and the corresponding means for pulp fibres, pelletized fibres and the final injection moulded tensile test specimen are shown. The material in FUMO68 consisted of 20% Fusellu, 39% Ac-starch-TEC and 41% lignin. The reduction in mean aspect ratio is clearly visible both in the distributions and in the means.

The mean fibre length of the injection moulded specimens was in the range 150 µm–300 µm, and the mean aspect ratio in the range 10 – 20 µm (see Figures 14 and 15). The composition of the composites is presented in Table 5. The mean fibre length of the raw chemical (Fusellu) and mechanical (TMP) pulp, measured with the X-µCT technique, was 1100 µm and 600 µm, respectively. The decrease in fibre length due to processing is thus very large.

In some manual methods fibres less than 500 µm in length (typically classified as “fines”) are not included in the mean fibre length determination.

To compare the present results with such measurements, the mean fibre lengths were also determined for the L > 500 µm fractions. The results indicate mean fibre lengths in the range 650 µm – 900 µm for the tensile test specimens and 1500 µm and 960 µm for intact chemical and mechanical pulps, respectively (see Figure 14). It is thus evident that the pelletization, compounding and injection moulding processes significantly increase the fraction of fines, possibly decreasing the mechanical properties of the composite materials.

A key question concerning the fracture mechanisms of composite materials is whether the reinforcement fibres are mainly broken at the fracture surface, or whether some of them are pulled out of the matrix at fracture. The latter case would be indicative of inadequate bonding between the matrix and the fibres. The fracture surfaces of two material samples manufactured in this project, FUMO18 (20% TMP in PEG-plasticized LB matrix) and FUMO39 (20% TMP in Ac-starch-TEC plasticized LB matrix), were studied using X-µCT imaging. Based on visual inspection of the images, it appears that in both of the studied samples only a minor proportion of fibres were pulled out, with most remaining firmly attached to the matrix, see Figure 16. Hence, we can conclude that the fibres have a strong positive reinforcing effect on the composites.

Figure 13. a) Length/diameter distributions estimated for FUMO68 b) Mean length/diameter distributions for FUMO68. Mean values are calculated based on both major (d1) and minor (d2) diameters of fibres, but the distributions are shown only for L/ d1.

a) b)


Figure 15. Mean fibre aspect ratio of the selected composites, raw fibres and fibre pellets. The cross-section of the fibre has been approximated with an ellipse, where d1 refers to the length of the major axis and d2 to the length of the minor axis of the ellipse.

Figure 14. Mean fibre length of the selected composites, raw fibres and pellets. Means are calculated from full data (blue bars) and from the >500 µm length fraction (red bars).

Sample Fibre type Lignin wt-% Acetylated

starch wt-%

TEC wt-%

FUMO39 TMP 40.8 23.2 16.0

FUMO68 Fusellu 40.8 23.2 16.0

Table 5. Composition of the composites. Fibre weight fraction was 20% in all cases.


4.5 Demonstrators

The processability of the thermoplastic lignin composites was demonstrated at VTT by injection moulding parts using commercial moulds provided by Rinotop (Figure 17). The demonstrator parts were prepared using the best material combinations (PEG as well as Ac-starch-TEC plasticized LB matrix and

Figure 16. Cross-sectional X-µCT slices of FUMO18 (a) and FUMO39 (b). Dark, medium grey and light grey colours indicate air, matrix and fibres, respectively. Only a few pulled-out fibres and micro-cracks extending in the bulk material are visible.

a) b)

a) b)Figure 17. Injection moulded demonstrator parts with wood-derived material contents up to 77%.

Fusellu reinforcements). Two different moulds were employed to produce the parts. The “spacer mould” is a commercial mould used to manufacture complex, thin-walled parts employed in concrete casting (a). The other mould was a “ball mould” used for producing knobs for skipping ropes (b). The ball mould has thick walls setting special requirements for the heat conductivity of the material.



In this project, novel thermoplastic lignin composites reinforced with wood pulp were developed. The processability of thermoplastic lignin was demonstrated using typical processing equipment, such as twin-screw extruders and injection moulders. The tensile properties of the lignin composites are close to its main competitors: wood fibre composites with petroleum-based matrix polymers. Therefore, the potential application areas for the lignin composites are the same as for the aforementioned polymers and reinforced composites. These areas include the construction (e.g. decking, fencing, window frames), furniture (e.g. chairs, tables, cupboards) and automotive (e.g. interior parts) industries.

The most promising target applications with the current material performance profile are foreseen to be found in construction. At present, the inadequate impact strength of the injection moulded composites limits the utilization of the material in more demanding applications, such as in vehicles or furniture. However, the

potential is substantial, and furthermore, large quantities of lignin are commercially available after the recent introduction of the LignoBoost technology. The results of this work serve as a platform for further development of high-performance composite materials based on thermoplastic lignin.

6. Networking

The research was carried out jointly by research organizations and Finnish forest cluster companies. Table 6 presents the partners and their role in the research.

Lignin samples and related mechanical performance data were exchanged between the project partners and FPinnovations (Canada) during the project.


Metsä Fibre Industrial tutor. Supplier of bleached softwood pulp.

Novoplastik Testing and evaluation of composite samples.


University of Helsinki Chemical activation and modification of fibres enabling

binding with matrix lignin in composite materials.

University of Jyväskylä Characterization and analysis of composites by X-ray

computed microtomography.

UPM-Kymmene Work Package Coordinator. Supplier of thermomechanical

softwood pulp.

VTT WP leader. Processing and production of thermoplastic

lignin composites. Chemical modification of lignin and

enzymatic modification of fibres.




Lahtinen, M., Ojala, A., Wikström, L., Hietala, S., Nättinen, K., Fiskari, J. and Kilpeläinen, I. Injection-molded fiber-reinforced lignin com-posites from chemically modified thermome-chanical pulp (TMP) fibers, in preparation.

Miettinen, A., Joffe, R., Madsen, B., Nättin-en, K. and Kataja, M. Quantitative analysis of length-diameter distribution and cross-sec-tional properties of fibers from three-dimen-sional tomographic images, Proceedings of the 34th Risø International Symposium on Ma-terials Science, 2013.

Miettinen, A., Ojala, A., Wikström, L., Joffe, R., Madsen, B., Nättinen, K. and Kataja, M. Non-destructive automatic determination of aspect ratio and cross-sectional properties of fibres, in preparation.

Myllymäki, T. Ligniinin rakenne ja kemiallinen muokkaus, Master’s thesis, University of Hel-sinki, 2014.

Nättinen, K., Kouisni, L., Paleologou, M., Wik-ström, L. von Weymarn, N. and Browne, T. Composites of kraft lignin, Poster presentation together with FPInnovations in NWBC, Helsin-ki, 2012.

Ojala, A., Wikström, L., Lehmonen, J. and Nät-tinen, K. Foam-laid formed thermo-mouldable nonwovens, Proceedings of Composites Aus-tralia and the CRC for Advanced Composite Structures, Newcastle, Australia, 2014.


BIOREFINERYPRODUCTS

INITIAL PROCESS PILOTING ANDMATERIAL PROTOTYPE PRODUCTION

IN THE CASE OF BARRIERS,3D-MOULDABLE PACKAGING

AND FILTERS

CONTACT PERSON – Work Package 4 leader Jonas Hartman, [email protected]

Aalto University: Ramarao Abburi, Ilari Filpponen, Jiagi Guo, Karoliina Junka, Janne Laine,Alexey Khakalo, Eve Saarikoski, Jukka Seppälä, Ola Sundman, Monika ÖsterbergKemira: Mari Ojanen, Reetta Strengell, Tarja TurkkiLappeenranta University of Technology: Kaj Backfolk, Henry Lindell, Daria Nevstrueva,Arto Pihlajamäki, Sami-Seppo Ovaska, Esa Saukkonen, Panu TanninenMetsä Board: Pekka SuokasMetsä Fibre: Pirkko Liias, Raili KoponenStora Enso: Mari Hiltunen, Outi Kylliäinen, Titta Lammi, Jari RäsänenTampere University of Technology: Hanna Christophliemk, Johanna LahtiUniversity of Helsinki: Juha Fiskari, Susanna Heikkinen, Pirkko Karhunen, Ilkka Kilpeläinen,Sirkka-Liisa Maunu, Kirsi Mikkonen, Pirita Rämänen, Joonas Siirilä, Maija TenkanenUniversity of Oulu: Aleksi Kolehmainen, Henrikki Liimatainen, Juho Sirviö, Miikka Visanko UPM-Kymmene: Harri Kosonen, Leena Kunnas, Jutta Nuortila-Jokinen, Irmeli Sinisalo, Heli ViikValmet: Ismo Mäkinen, Pasi Nurminen, Ari Puurtinen, Peter Rosenberg, Timo SutelaVTT Technical Research Centre of Finland: Martta Asikainen, Marie Gestranius, Antti Grönroos, Minna Hakalahti, Ali Harlin, Pirjo Heikkilä, Juha Heikkinen, Jaakko Hiltunen, Juha Houni, Petri Jetsu, Saila Jämsä, Timo Kaljunen, Kari Kammiovirta, Annaleena Kokko, Hanna Koskela, Vesa Kunnari, Hanna Kyllönen, Christiane Laine, Marjo Määttänen, Heikki Pajari, Jaakko Pere, Pentti Pirkonen, Pauliina Pitkänen, Hille Rautkoski, Elias Retulainen, Erkki Salo, Harri Setälä, Asko Sneck, Riku Talja, Tekla Tammelin, Alexey VishtalÅbo Akademi University: Pedro Fardim, Jasmina Obradovic, Joanna Narewska, Victor Kisonen, Stefan Willför


ABSTRACT

Demonstration and piloting under the title “Improving traditional fibre products” focused on multilayer barrier packaging, 3D-mouldable packaging and new filter materials.

Different types of biobarriers were successfully prepared and coated as water dispersions on precoated base board. The biopolymer-coated webs were also extrusion coated and finally converted into tray form. Six different types of bio-based derivatives were developed and tested during the project. The tested materials were modified GGM, modified xylan, TOFA esters and TOFA alkyd acrylates, dialdehyde cellulose and DAC alginates and cellulose-copolymer blends. Good grease barrier properties were obtained in several barrier material groups. Additional studies on the effect of the base substrate fibre composition revealed that the hemicellulose content of the base substrate affects the convertability of the paperboard and indicated changes in the dispersion coating hold-out and grease resistance of dispersion-coated paper.

In the case of 3D-mouldable packaging, the maximum extensibility obtained for the paper was around 21% when the polymer dosage was 8%. In-plane compaction increased extensibility up to 35% in the machine direction. Gelatin effectively improved the extensibility and tensile strength of the paper and readily adsorbed on the fibres. In order to enhance the effects, a combination of several chemicals is possible. The treatments are compatible with current papermaking environments and can be used for the production of food packaging materials. The convertability of the produced paper materials was demonstrated in all main forming process types.

Several different new filter materials and manufacturing methods were tested. The results obtained with some of the demo materials were very promising. Cellulose membrane cast from ionic liquid was produced for industrial trials and for comparison with commercial references. Retentions of the studied sample were similar or slightly lower than those of the commercial references and in general the demo sample showed very stable but somewhat lower flux in all tested effluents compared to the commercial membranes. Dicarboxylic acid cellulose (DCC) nanofibre-coated membranes gave promising results. The vacuum filtration method of producing the samples proved effective at the laboratory scale, indicating that efficient production of larger membranes, for example by roll-to-roll coating, could be achieved.

Initial process piloting and material prototype production gained wide interest from the partner companies. Each of the three product groups had an engaged group of industrial tutors whose input focused the research and contributed considerably to successfully improving on conventional fibre products.

Keywords:3D-forming, biobarrier, filter, forging, formability, membrane, moldability, multilayer barrier, packaging


1. Background

There is a fast-growing need to find replacements for oil-based products. Increasing demand for packaging creates opportunities for bio-based barrier materials that can be produced from industrial sidestreams, such as from paper and board production. The replacement of oil-based products with products derived from natural sources has been the focus of intensive research in recent years. Due to their renewability, biocompatibility, biodegradability, wide availability and non-toxicity, natural bio-based products offer a sustainable means of obtaining chemicals and materials and replacing non-renewable resources. One such important group of biorenewable products is biopolymers, including polysaccharides, among which cellulose, hemicellulose and starch have been extensively studied. Due to their positive properties compared to oil-based materials, one of the most promising target applications of biopolymers is in environmentally friendly food packaging materials.

The focus of the FuBio Joint Research 2 programme was on biomass fractionation and the development of current and new biorefinery products. The work was based on the research results of the first FuBio programme targeting the main fractions of wood (e.g. cellulose, hemicelluloses), in which several bio-based polymers and polymer blends were developed for a range of interesting applications. These include dialdehyde cellulose, which is a cellulose derivate produced by a regioselective oxidation of cellulose using periodate as an oxidation agent, modified hemicelluloses (galactoglucomannan and xylan), tall oil including alkyd-acrylate hybrid polymers and tall-oil modified cellulose as well cellulose-polyethylene copolymer blends (CePE).

From the point of view of 3D-packaging, cellulose is a recyclable, biodegradable, and

renewable material that has certain advantages over plastic-based materials. However, fibre-based packaging materials cannot yet compete with plastic packaging in terms of convertability or package design – fibre-based packaging exists in rather simple geometrical forms, whereas plastics can be formed into a vast variety of shapes. The main limiting factor for the production of advanced 3D shapes from paperboard is its poor formability, i.e. its limited ability to withstand certain types of plastic deformation without damage.

Formability is the ability of a material to be formed into complex geometries through forming processes such as hot pressing, vacuum forming, stamping or deep-drawing. The formability of fibres and paper is primarily limited by poor elongation, which is typically below 5%. Formability determines the maximum depth of shapes and, to certain extent, the material appearance. The forming process and process variables (including moisture and temperature) also play a central role in formability.

Membranes are semipermeable barrier materials allowing certain compounds to pass through whilst rejecting others. Natural membranes perform an important role in many biological membrane processes, although the use of artificial membranes has become established in recent decades in a wide range of separation processes. The first artificial membranes were made using the cellulose-based material cellulose nitrate. Today, cellulose-based materials, mainly cellulose acetate and nitrate, are widely used in microporous membranes for micro- (MF) and ultra-filtration (UF) and haemodialysis. In addition cellulose-based membranes can be used in gas separation as dense, non-porous films. Cellulose membranes can be either symmetric or asymmetric, and their physical strength can be improved with the use of support layers. Cellulosic materials are


intrinsically highly hydrophilic and therefore ideal for water-based membrane processes. Although synthetic polymers have taken over some membrane applications, it is believed that novel cellulose materials and advanced processing technologies will open up new possibilities for cellulose, thus reclaiming some markets from synthetic polymers.

The pulp and paper industry is a large consumer of water, the quality of which varies widely, for example in terms of amount and type of solutes, pH and temperature. As an example, selected properties of three waters from pulp mill UF applications are presented in Table 1. Membrane life cycles are typically expected to be several years, and the typical water processing cost is less than 1 €/m3, a third of which is attributed to membranes. Cost reduction is needed, as increasing requirements for process water recycling are expected in the future. Typical water consumption of a pulp mill is currently around 20-30 m3 per tonne of pulp. With more advanced membrane technology this could be reduced to less than 10 m3 / tonne. Filtration performance, stability in harsh conditions, long life span, and low price are key factors for membranes used in the pulp and paper industry.

Water pH Temperature COD

White water from the pulp drying machine 4 – 5 50 °C 500-650 mg/l

EOP filtrate from the bleaching plant 9 – 10 60 °C 2000-5000 mg/l

Press washer filtrate after chemical cooking 13 70 °C > 100000 mg/l

Table 1. Typical examples of pulp mill ultrafiltration waters (pH, temperature and chemical oxygen demand (COD) – commonly used as an indirect measure of organic compounds).

2. Objectives

New types of bio-based materials were tested in different packaging applications with the aim of improving traditional fibre products. The main objectives were:

• Todevelopmulti-layeredboardpackagingstructures, including layers made of possible sidestreams such as xylan, GGM, TOFA or dissolved cellulose, with good grease barrier properties at high relative humidity to create holistic usage of all materials in a mill/production site

In 3D-mouldable packaging structures, the ob-jectives were:

• Toimproveoftheformabilityofpaperwebsto a qualitatively new level and develop fibre web based materials for producing deep 3D shapes (2-3 cm) using industrially feasible technology

• Todemonstratetheconvertabilityofthedeveloped paper-based materials in existing and emerging forming processes

• Toincreaseknowledgeoftheroleofmechanical properties and process parameters in the formability of paper webs

In filter development, the objective was:

• Toimprovetheflux,retention,and/or prolong the lifespan of membranes, especially those used in pulp and paper industry water systems.



Three product applications were selected for initial piloting and demonstration: 1) grease barrier in multilayer board packaging structures, and 2) 3D-mouldable webs and 3) filters. When the results for the different materials developed in the first FuBio programme for similar applications were compared, the importance of centralized testing environments and measurements became clear. Therefore, in the present programme centralized testing platforms were designed.

In packaging barrier research, material development was aimed at achieving a grease barrier by means of dispersion coating, improving the mouldability of conventional fibre webs, and demonstrating and piloting the developed materials.

Figure 1 shows a schematic of the barrier development approach. In the barrier research, each of the six material groups developed their products individually (Figure 1: Synthesis). Studies on self-standing films and spin coating assisted in the evaluation of the film formation properties and application recipe of the materials (Figure 1: Self-standing films, Spin coating). All materials were applied on a selected base board and the resulting barrier properties were tested (Figure 1: Laboratory coating, Pilot coating). After each coating application round (several laboratory rounds and a pilot trial) feedback was given to the material development groups and improvement targets were agreed (e.g. moving from solvent-based system towards water dispersion, aiming for a less tacky product, etc.). After two years, the industrial tutors selected the three most promising derivatives for an additional pilot

Figure 1. Barrier research approach.

• Comparison between samples that spread well

• Clear effect of substrate and barrier sample chemistry

Synthesis

Pilot coating Extrusion Converting

Self-standing films Spin coating Laboratory coating


scale trial. After pilot coating the biobarrier-coated webs were extrusion coated and further converted into tray form, representing the final product (Figure 1: Extrusion, Converting).

As part of the dispersion barrier development, additional studies with dual-polymer systems with and without high aspect ratio plate-like pigment addition were also conducted in order to increase the grease resistance and convertability of paperboard. The effect of the base substrate on flexible packaging was evaluated with special attention to the fibre composition. The effect of fibre composition on convertability was studied by preparing a novel papermaking fibre with cellulose-rich fibre surfaces by enzymatic hemicellulose removal.

In the 3D-mouldable web research task, various approaches were taken to improve the formability of paper-based materials. The

formability was improved via two approaches: 1) increasing the extensibility potential of fibres and paper, and 2) subjecting swelled pulp fibres to extremely high pressure loads. Table 2 shows the research approach.

In filter development, the used test waters were kept as constant as possible throughout the project and fibres and new technologies were studied with the aim of producing materials applicable in reactive and active filter and membrane applications. The work included membrane and membrane component development and functionalization as well as study of membrane properties. The main approaches to membrane development included casting of membranes using ionic liquid (IL) and preparation of nano- and micro-fibrous materials for layered structures using coatings of dicarboxylic acid cellulose (DCC) nanofibres and electrospun micron and sub-micron fibres.

Table 2. Mouldable web research approach.


4. Results

4.1 Improving grease resistance in multilayered board packaging structures

In the barrier research, new coating materials for achieving grease resistance of paperboard in packaging applications were developed. One aim was to obtain the coating materials as water-based dispersions. Six different types of bio-based derivatives were developed and tested during the project. These were modified GGM(ÅboAkademiUniversity),modifiedxylan(VTT), TOFA esters and TOFA alkyd acrylates (University of Helsinki), dialdehyde cellulose alginates (University of Oulu) and cellulose-copolymer blends (Aalto University). Additionally, the barrier coatings were up-scaled from lab to pilot scale and the multi-layer paperboard packaging structures with good grease barrier properties were demonstrated. More than 60 derivatives were tested during the project. Centralized coating tests were performed at VTT facilities (Figure 2) and pilot coated samples were further sent to TUT for extrusion coating and LUT for conversion into tray form.

Sample characterisation was also conducted in a centralized testing environment, including grease, water vapour transmission (WVTR) and oxygen barrier (OTR) measurements.

When considering the grease barrier, the most promising coatings are cellulose-copolymer blends (CePE) and hydroxypropylated xylan coatings. Figure 3 shows the results for the best candidate of these two (CePE and xylan) as well as TOFA hybrid latex (HL) derivative families as normalized to 20 µm thickness. In grease resistance measurements our target was to attain extended resistance, i.e. high values when measured in days (y-axis) as well as low water vapour and oxygen transfer rates (WVTR and OTR, respectively). The blue circle represents the combined target value

area with respect to 3a) grease resistance and water vapour barrier or 3b) oxygen and water vapour barrier.

Regarding the cellulose-copolymer blends (CePE), grease resistance of over 65 days was obtained with a dissolved cellulose/poly(ethylene-co-acrylic acid) blend with a 95/5 polymer/polymer ratio. After dispersion coating, the best OTR achieved was 130 cc/(m2d bar); however after PE extrusion OTR the value was as low as 2 cc/(m2d bar). The WVTR target was also achieved. In this group, the blending capability of commercial co-polymers with alkaline aqueous cellulose solution was studied and the mixing procedure and blend compositions developed. This was done in order to obtain the best morphology of the solutions/suspensions for the coating trials.

TOFA-based hybrid latexes (HL) were synthesized and applied on paperboard and water and grease barrier properties were analysed. Good water and water vapour barrier properties were achieved, whereas the grease barrier was moderate. Oxygen barrier property was improved when talc or cellulose was used as filler. The materials were also heat-sealable. However, some process scale-up issues (e.g. adhesion problems) remained unresolved.

The xylan derivatives – in particular hydroxypropylated xylan (HPX) dispersions – could be coated as such, cross-linked, plasticized and filled with talc or nanoclay and showed good grease barrier properties in all cases. The barrier-enhancing fillers concept was demonstrated and WVTR of 46 g/(m2d) and OTR of 72 cc/(m2d bar) were achieved. In addition, the concept of low DS combined with external plasticization, enabling a bio content of up to 90%, was tested successfully. The colour of the coating material colour was white and remained stable during drying and converting. Heat sealability was achieved, and


Figure 2. Pilot trial equipment at VTT.

PE extrusion coating was possible leading to OTR values below 10 cc/(m2d bar). Die-cutting and press-forming into trays was successful.

In conclusion, several potential barrier dispersion coatings were developed and demonstrated

and multi-layer paperboard structures with several beneficial characteristics, especially grease barrier properties, were produced. All of the derivatives require further optimization according to the target end use, but can be readily utilized at the industrial scale.

Figure 3. Best barrier properties of the three most promising derivatives normalized at 20 µm thicknesses, measured at 23 oC, 50%RH. The targeted area in Figure a) is in the top left corner and in Figure b) in the bottom left corner.

a) b)


Improving grease resistance of starch-based coatings When synthetic polymer was added to modified starch-based dispersion (dual-polymer system) at optimized amounts, efficient grease resistance was achieved. The addition of synthetic polymer to starch-based coatings improved the elastic properties and plasticity and, subsequently, the mechanical convertability of the product. Addition of plate-like like pigment to the renewable dispersion coating increased the grease resistance of the non-converted blanks, whereas micro-scale cracks in the barrier were frequently observed after tray conversion, resulting in poor grease resistance.

Effect of hemicellulose removal on convertability The effect of fibre composition on convertability was studied by preparing a novel papermaking fibre with cellulose-rich fibre surfaces by enzymatic hemicellulose removal. Xylanase treatment of refined never-dried bleached birch kraft pulp demonstrated selective hydrolysis of xylan from the outermost fibre layers, reducing 14.2 wt% of the total amount of xylan in the studied pulp. The mechanical properties of paperboard at 180 g/m2 grammage were mainly preserved despite a 14.2 wt% reduction in xylan content. Press-forming of the produced materials revealed differences related to the end moisture content of the produced paperboard (i.e. drying severity), fibre composition (i.e. hemicellulose content), and press-forming parameters. Consequently, the suitability of the paperboard could be optimized for flexible packaging by tailoring the carbohydrate composition of the base substrate.

4.2 Improving formability of paper-based materials

The approach used for improved formability of the fibre web included three major stages (Figure

4): a) mechanical treatment of fibres to reduce their axial stiffness, create microcompressions and dislocations (steps 2 & 3); b) improvement of inter-fibre bonding by spray application of polymers (step 4) c) structural modification of the fibre network by in-plane compaction treatment and unrestrained drying of the paper to allow free shrinkage (steps 5 & 6). Used in combination, these stages could achieve paper elongation of 15…30%. In addition to these treatments, the forming conditions (mainly temperature and moisture content of paper) that affect the softening of the polymers (cellulose, hemicelluloses, added polymers) could improve the extensibility by 2–4 percentage points. It was found that paper at a moisture content of 8-11% can soften at 80 °C, but further increases in temperature or moisture decrease formability. The combined approach for the improvement of formability is shown in Figure 4.

The application of several natural polymers was tested (starch, xyloglucan, hydroxypropylcellulose, CMC, gelatin, agar). Spray addition of these biochemicals was found to increase both strength and elongation. The most interesting chemicals were agar and gelatin. For example, gelatin improved the extensibility and tensile strength of unrestrained and restrained dried paper. A 4% gelatin addition improved the tensile strength from ca. 50 Nm/g to ca. 80 Nm/g, and extensibility from ca. 9% to 13% for unrestrained dried paper. However, besides strengthening fibre bonding, chemical addition also increased drying shrinkage.

Papers prepared using the combined approach (as shown in Figure 4, excluding the compaction stage) were used to produce three sets of demo materials. The test results clearly demonstrated that different forming processes have different formability requirements. One key difference between the processes is whether the paper blank is allowed to slide into the mould (sliding blank) or whether it is strained into the mould (fixed blank). In the deep-drawing process


the paper is allowed to slide and the depth of the shape is almost unlimited, although the appearance worsens with increased depth. The basic process types tested in the project are classified in Figure 5. Detailed photos of samples made in fixed blank processes are shown in Figures 6-8.

Maximum paper extensibility of around 21%

was obtained at a polymer dosage of 8%. In-plane compaction further increased the extensibility up to 35% in the machine direction. Gelatin effectively improved the extensibility and tensile strength of the paper and readily adsorbed on the fibres. For enhanced effects, a combination of several chemicals can be used. These treatments are compatible with current papermaking environments and can be used for

Figure 4. Combined approach for improvement of formability.

Figure 5. Classification of the basic forming methods tested and examples of shapes obtained. The hot pressing mould is a VTT Design.


Figure 6. Tray-like shapes (underside and top side) obtained by a hot pressing process developed by the Moldable Web project at VTT. The shape depth is around 16 mm.

Figure 7. Double-curved structure produced by hydroforming. Around 20% extensibility is required for this formation. (Hydroforming performed by the Royal Institute of Technology (KTH), Sweden.)

Figure 8. Tray-like shape air-formed by PACCOR (Coveris).Maximumshapedepth~25mm.

Figure 9.Schematicapproachfortheimprovementofmouldabilityofcellulose(ÅA):1)Startingmaterial:softwood dissolving pulp; 2) Swelling of cellulose in DMAc-LiCl; 3) Forging the swelled cellulose at 70MPa pressure; 4) The resulting cup-like form, height ca. 50 mm.

1) 2) 3) 4)


the production of food packaging material. The convertability of the produced paper materials was demonstrated in all main forming process types.

Preparation of three-dimensional cellulose objects from fibres previously swelled in DMAc/LiCl solvent system In this approach, the aim was to take advan-tage of the modifications in intermolecular in-teractions that swelling treatment induces in cellulose in order to be able to subject the cel-lulose to moulding and high pressure. Cellulose pulp was swelled with DMAc/LiCl solvent sys-tem, washed and dried before further process-ing. In addition, acrylated epoxidized soybean oil was incorporated into the swollen fibres to improve mouldability. The formability of these treated cellulose samples was then evaluated.

This approach is unique in that the core treat-ment involves the swelling of the dissolving pulp in non-aqueous salt solution to disrupt the inter- and intramolecular hydrogen bonding and decrease the crystallinity of the cellulose, with subsequent forging under high pressure to create a dimensionally stable shape. Swell-ing of cellulose in non-aqueous salt solution with consequent forging may be used for the production of three-dimensional translucent shapes from cellulose.

4.3 Main results for filter materials

IL castingCasting based on phase inversion by immersion precipitation typically produces an asymmetric membrane structure with a thin, tighter skin lay-er for good separation on one side and an open pore structure bulk layer providing good flux. Such materials are typical in UF filtration appli-cations. LUT’s casted membranes were based on commercial cellulose dissolved into IL liquid. The solution was cast and the resulting wet film was soaked in a non-solvent bath. The struc-

ture of the obtained material was unique and the properties of the produced membranes var-ied according to the process parameters, such as coagulation bath temperature and used ad-ditives. In lab-scale testing, the prepared mem-branes were compared with one of the best commercial RC70PP membranes.

One membrane sample was produced for in-dustrial trials for comparison with commer-cial references. The sample was pure cellulose (8 wt-%) without any additives or any surface functionalization supported by PET nonwo-vens. Industrial trials were carried out by Valmet Technologies at the Metsä Fibre Rauma mills using a CR250/2 unit (effluents listed in Table 3). Retentions of the studied sample were simi-lar or slightly lower than the commercial refer-ences (see white water example in Table 3), and in general the demo sample showed very sta-ble, but somewhat lower flux in all tested efflu-ents compared to the commercial membranes.

The results obtained are very promising, as there are currently no commercial cellulose membranes fabricated directly from cellulose. The membrane preparation process however requires further development in order to opti-mise the membrane properties for specific end uses, to enable the production of larger mem-brane pieces for flat membranes, and to trans-fer the technology to hollow fibre (HF) mem-brane production.

Functional IL-cast membraneIL casting was also used to create temperature-responsive membrane in cooperation with LUT and VTT. VTT’s allylated cellulose was cast at LUT and subsequently grafted with poly(N-iso-propylacrylamide) (PNIPAAm) at VTT. A non-grafted reference and the PNIPAAm grafted demo material were tested at the lab scale. Stimuli-responsive behaviour of the grafted membrane was observed in pure water flux testing as well as anti-fouling and easy-clean-


ing properties during wood hydrolyzate filtra-tion, see Figure 10. The cycle was repeated twice and the properties of the grafted mem-brane restored fully after each cycle.

DCC nanofibre coatingLayered structures are characteristic of fibrous materials. Whereas an asymmetric pore size distribution is obtained inherently in casting, a similar effect can be obtained by combining layers of different pore size. Support materi-al layers provide mechanical strength without significantly affecting flux, while layer(s) with smaller fibres and pore sizes provide effective depth filtration. University of Oulu prepared a new type of membrane barrier layer structure

on top of a commercial support material using DCC-nanofibrils (see Figure 11). The properties of the produced membranes depended on the various process parameters, such as the layer thickness and used cross-linking agent.

These cellulose-based membranes are best suited to neutral conditions with a pH range of 1-11, preferably 5-9. The membranes can puri-fy clear water from feed solutions where the largest contaminant particles have been re-moved and the temperature is below 70˚C. The functionality of the fabricated membranes was demonstrated with real industrial process wa-ter with a high level of purification achieved with paper machine disc filter fractions (cloudy

Sample COD to-tal mg/l

COD soluble

mg/l

pH Conductivity mS/m

Charge µekv/l

TSS GF/A mg/l

Turbidity NTU

White water feed

610 520 4.5 230 -40 167 128

LUT demo sample

470 460 4.8 220 -32 0 0.4

Reduction 23% 12% 4% 20% 100% 100%

RC 30 kDa 500 490 4.8 220 -15 0 0.3

Reduction 18% 6% 4% 63% 100% 100%

Table 3. Analyzed compounds in the feed and the permeate samples of white water using the LUT demo sample and commercial RC 30 kDa membrane.

Figure 10. a) Pure water flux at different temperatures; and b) at different stages of wood hydrolyzate filtration; and c) appearance of non-grafted and PNIPAM grafted allylated cellulose membrane. In (c) non-grafted sample on left, grafted on right.

a) b) c)


and clear filtrate). The filtration efficiency in terms of fouling, flux and purification degree was as good as the commercial reference UH030 when CaCl2 was used as a cross-linking agent in membrane preparation.

The results obtained with this demo materi-al were very promising. The vacuum filtration method of producing samples proved effective at the laboratory scale, indicating that efficient production of larger membranes by roll-to-roll coating can be achieved.


Initial process piloting and material prototype production tasks focused on improving con-ventional fibre products. Whilst different types of biobarriers were successfully prepared and coated as water dispersions on precoated base board many process and product issues still need to be solved before any of the materials can be taken to products. In the case of 3D-mouldable packaging, on the other hand, the studied treatments are compatible with current papermaking environments and can be used for the production of food packaging materials. Also the convertability of the produced paper

Figure 11. DCC barrier layer (a) surface structure and (b) cross-sectional structure.

materials was demonstrated in all main forming process types. Here the exploitation potential of the results depends on the raw material pric-es of addition chemicals and costs for new unit operations. The development of new filter ma-terials shows promising results, however this area needs a lot of research before industrial products can be talked about.

This work will have it’s largest impact on im-proving conventional fibre products through increased knowledge among researchers and industry in the areas of biobarriers, mouldable packaging and filter.

6. Networking

Several research partners were involved this study. In addition, industry partners actively participated in steering and guiding the work and in conducting measurements and trials, see Table 4. Partner organizations and their re-search roles.

a) b)




Aalto University•PolymerTechnology(P)•ForestProductsTechnology(FP)

(P) Blending capability of commercial co-polymers with alkaline aqueous cellulose solution, mixing procedure to obtain best morphology in the solutions and to function well as barrier coatings.(FP) Preparation of model surfaces, analysis of model surfaces and spin coating. Investigation of the effect of different polysaccharides and their properties on web mouldability. Membrane functionalization by click chemistry.

Kemira Assisted partners with dispersion knowledge, Industrial tutor.

Lappeenranta University of Technology•PackagingTechnology(PT)•FiberandPaper(FP)•SeparationTechnology(ST)

(PT) Production and collection of commercial samples for the coating testing platform.(FP) Studies on coating hold-out, infiltration with an aim to identify the effect of substrate properties on paper-coating interaction.(ST) Preparation of Cellulose Membranes.

Metsä Board Industrial tutor


Stora Enso Task 2 Coordinator, study on microwavable packaging materials, barrier measurements, converting trials.

Tampere University of Technology•PaperConvertingandPackagingTechnology

Provided the dissolved cellulose for Aalto Polymer TechnologyExtrusion coatings, measurement of barrier properties

University of Helsinki•OrganicChemistry(OC)•PolymerChemistry(PC)•FoodandEnvironmentalSciences(FES)

(OC) prepared materials with good grease barrier properties for coating experiments using dissolving pulp as starting material.(PC) Preparation of alkyd-acrylate copolymers. Structural characterization of products.(FES) studies of free films, mechanical and barrier properties.

University of Oulu•FibreandParticleEngineering(FPE)•Organicchemistry(OC)

(FPE) Mechanical activation, production of micronized cellulose and application testing. Membrane structures based on DAC/DCC microfibers.(OC) Chemical derivatization and functionalization of fibre material, production of new modified carbohydrates.

UPM-Kymmene Task 1 Coordinator

Valmet Task 3 Coordinator, industrial testing of developed membrane material.

VTT WP leader. Chemical modification of xylan, development of TOFA-based alkyds, model surface studies, barrier testing, laboratory and pilot coating. Chemical modification of pulp, grafting of thermoplastic polymers and impregnation of webs with polymers. Verification of the testing platform for formability. Functionalization of fibres for membranes and commercial cellulose-based membranes, especially stimuli-responsive materials. Electrospun membranes. Membrane testing, lab-scale preparation of (demo) membrane materials.

Åbo Akademi University•Woodandpaperchemistry(WPC)•Fibreandcellulosetechnology(FCT)

(WPC) Chemical modification of GGM, optical method for grease barrier determination.(FCT) Preparation of cellulose 3D objects using physicochemical pretreatments of pulp fibres in combination with mechanical treatment or forging.



Heikkinen, S.L., Mikkonen, K.S., Koivisto, P., Heikkilä, M., Pirkkalainen, K., Liljeström, V., Serimaa, R. and Tenkanen, M. Long-term phys-ical stability of plasticized hemicellulose films, BioResources, 2014, 9, 906-921.

Laine, C., Harlin, A., Hartman, J., Hyvärinen, S., Kammiovirta, K., Krogerus, B., Pajari, H., Rautkoski, H., Setälä, H., Sievänen, J., Uoti-la, J. and Vähä-Nissi, M. Hydroxyalkylated xy-lans - their synthesis and application in coat-ings of packaging and paper, Industrial Crops and Products, 2013, 44, 692–704. http://dx.doi.org/10.1016/j.indcrop.2012.08.033

Mikkonen, K.S., Schmidt, J., Vesterinen, A.-H. and Tenkanen, M. Crosslinking with am-monium zirconium carbonate improves the formation and properties of spruce galactoglu-comannan films, Journal of Materials Science, 2013, 48, 4205-4213.

Matikainen, J., Karhunen, P., Kulomaa, T., Fis-kari, J., Mikkonen, K.S., Tenkanen, M. and Kil-peläinen, I. Films from dissolving pulp modified with fatty acids, EWLP Helsinki August 27–30, 2012, Proceedings, 338–341.

Mikkonen, K.S. and Tenkanen, M. Sustaina-ble food-packaging materials based on future biorefinery products: xylans and mannans, Trends in Food Science & Technology, 2012, 28, 90-102.

Narewska J., Lassila L. and Fardim P. Prepa-ration and characterization of new mouldable cellulose-AESO biocomposites, Cellulose Jour-nal, DOI 10.1007/s10570-013-0157-3

Rämänen, P. and Maunu, S.L. Structure of tall oil fatty acid-based alkyd resins and alkyd-acrylic copolymers studied by NMR spectros-copy, Prog. Org. Coat., 2014, 77, 361–368.

Rämänen, P., Penttilä, P.A., Svedström, K., Maunu, S.L. and Serimaa, R. The effect of dry-ing method on the properties and nanoscale structure of cellulose whiskers, Cellulose, 2012, 19, 901-912.

Rämänen, P. Pitkänen, P., Jämsä, S. and Maunu, S.L. Natural oil-based alkyd-acrylic co-polymers: New candidates for barrier materials, J. Polym. Env., 2012, 20, 950-958.

Saarikoski, E., Rautkoski, H., Rissanen, M., Hartman, J. and Seppälä, J. Cellulose/acrylic acid copolymer blends for films and coating ap-plications, Journal of Applied Polymer Science, 2014, 131, 402864. DOI 10.1002/app.40286

Schmidt, J. Ristisidotut kuusen galaktoglu-komannaanikalvot, Master’s thesis, EKT Series 1581, 2012, University of Helsinki.

Sirviö, J. A., Kolehmainen, A., Liimatainen, H., Niinimäki, J. and Hormi, O. E. O. Biocomposite cellulose-alginate films: Promising packaging materials, Food Chemistry, 2014, 151, 343.

Sirviö, J. A., Liimatainen, H., Niinimäki, J. and Hormi, O. Sustainable packaging materials based on wood cellulose, RSC Advances, 2013, 3, 16590 – 16596.

Tanninen, P., Lindell, H., Saukkonen, E. and Backfolk, K. Thermal and mechanical stability of biopolymer barrier coatings in press forming of paperboard, Packaging Technology and Sci-ence, 2014, 27, 5, 353-363.

Vishtal, A., Hauptmann, M., Zelm, R., Ma-jschak, J.-P. and Retulainen, E. 3D Forming of Paperboard: The Influence of Paperboard Prop-erties on Formability, Packag. Technol. Sci., 2013, doi: 10.1002/pts.2056.


Vishtal A. and Retulainen E. Deep-drawing of paper and paperboard: the role of material properties, BioResources, 2013, 7, 3, 424-450.

Vishtal, A. and Retulainen, E. Improving the extensibility, wet web and dry strength of pa-per by addition of agar, Nordic Pulp and Paper Research Journal. 29 (3): in press (2014).

Zeng X., Retulainen, E., Heinemann, S. and Fu, S. Fibre Deformations Induced by Different Me-chanical Treatments and Their Effect on Zero-Span Strength, Nord. Pulp Pap. Res. J., 2012, 27, 2, 335-342.

Zeng, X., Vishtal, A., Retulainen, E, Sivonen, E. and Fu S. The Elongation of Paper – How fi-bres should be deformed to make paper exten-sible?, BioRes, 2013, 8, 1, 472-486.

The main objective of FuBio is to establish, in Finland, globally competitive

knowledge platforms within the field of wood biorefinery R&D for the

renewal of the forest industry and creation of new business.

FuBio is focused on development of novel value chains, in which wood is

refined into especially materials and chemicals.

www.fibic.fi

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