The amazing ability of continuous chromatography to adapt to a

moving environment. Roger-Marc Nicoud, Founder of Novasep

Barcelona, October 2013

Introduction

My thesis:

the environment pressure pushes Continuous Chromatography to evolve like animals evolved

according to Darwin’s law ….

and for that, I will use shortcuts , approximations and manipulations …

Improving what ?

System size Eluent consumption

Yield Operations

Comment Productivity kg/kg/day normally presented as the main objective function

Minimizing solvent consumption (prior to recycling) is a must in all large scale applications.

Can be a major cost contributor

Control / Process Stability / Cleaning can be critical costs contributors.

Mitigation large LP systems can be cheaper than smaller HP systems

Biopharma industry still a bit less concerned

Ancillary equipment is even more important than chromatography itself.

Life is often a matter of compromize

Continuous can helpCounter-current can help

Continuous is NOT synonymous with counter-current

A continuous but NOT counter-current animal

Be careful with comparisons

Batch My clever process

0

2

4

6

8

10

12

Productivity kg/kg/d

Optimum particle size, eluent composition … are different for Batch and continuous/CC processes.

S.M.B.: the key evolution …..

solid liquid

I

II

III

IVMinimizin

g E.C.

Evolution lead to the first continuous AND counter-current animal

Maximize productivity

Feed

Eluent

The first evidence of continuous chromatography

Environment pressure : Paraxylene• Scale (500,000 Mt/Y/Unit in 2010)

• Chromatographic conditions:– Selectivity about 2– Particle size about 1000 microns (inducing “poor” HETP ….)– Temperature about 180 °C (inducing low viscosity)

• Technical constraints– Target purity: > 99.8 %– “individual bed length” : limited to 1 m due to mechanical constraints

24 columns to reach total length

significant bed length of 24 m(no pb because of low viscosity)

Environment pressure : Fructose• Scale (50,000 Mt/Y/Unit in 2010): from Glucose-Fructose

mixtures• Chromatographic conditions:

– Selectivity about 1.5– Particle size about 300-500 microns (inducing “poor” HETP)– Temperature: 60 °C (thus viscosity about 10 cp )

• Technical constraints– Target purity: about 90 %– Minimize energy consumption for evaporating Water

total bed length of about 8-12 meters

Improving the classical SMB

• Add some degree of freedom by: – Connecting columns in different ways– Selecting different switching modes– Varying I/O lines composition and flows

• New processes are normally: – quasi-continuous instead of being truly continuous – taking advantage of «counter-current» contact

Fructose: further improvement

Commercial solutions implemented: • 4 or 6 columns• Eluent consumption decreased by about 10-15 % compared to SMB

Two sub periods

«Cousin animals»: • SSMB Novasep• ISMB Mitsubishi

A new animal

Environment pressure : optical isomers• Scale (1-100 Mt/Y/Unit in 2010)• Chromatographic conditions:

– Selectivity: typically between 1.2 and 2 – Little influence of particle size on packing cost– HETP depending on particle size …

• Technical constraints– Target purity: 95-99 % – Minimize system size / CSP inventory

Use small particle size

Total bed length about 0.5 m ….To be distributed between Ncol …At large scale L/D < 0.1 …..

The first SMB animal for separating optical isomers (1992):

Influence of petrochemical design is very visible ….

and the modern version ….

Optical isomers: further improvement

VariCol:• Truly continuous• Same hardware as SMB• Typically saves one

column out of 5-6.

VariCol

Key successes

For main successes, continuous chromatography is coupled with crystallization, isomerization and multiple effect evaporators

Very large scale does not mean « second-class » performance

In Line Dilution is key

Environment pressure : need for multicomponent (bio)separations.

Technically one can design a continuous three pure fractions TMB.

III

IIIIVV

VI

VIIVIII

Imposes to work with two suboptimal systems ?

The generic purification problem :

reduces to a binary separation if one wants the green or the red product.SMB is perfectly adapted to these situations(ex.: desalting, xylenes, capture …),

timeW P S

or change concept …

An animal with no progeny ?

Prefer two SMB in series :Paclitaxel, Cyclosporine, EPA …

Improving multicomponent separations

SW

P

S

W

P

I

II

III

IV

V

VI

P+S

P+W

A truly continuous gradient chromatographic process with 6-columns

MCSGP (Multicolumn Countercurrent Solvent Gradient Purification) from ETH.

Important features of the animal:• Opening liquid loop• Short-circuit• Gradient

Feed

S

W

P

I

II

III

IV

V

VI

I

II

III

IV

V

VI

P+S

P+W

I

II

III

IV

V

VI

P+S

P+W

Can be done with three columns

Not strictly continuous but counter current

Two sub periods

3-column MCSGP

“Cousin animal” from Novasep : GSSR Gradient (with) Steady-State Recycling

Environment pressure : need for capturing (biomolecules)

S Very strongly

retained

WNon retained

P

Selectivity is such that a solvent change is mandatory

Loading

Washing

Elution

t

RegenerationEquilibrationtcycle

Bioseparations: the BioSc concept

Use sequential feeding to reduce the number columns (as low as two).

Regenerate

Equilibrate

Target

Feed

Elute

Waste

S

W

PWash

WWash *

Two sub periods

Regenerate

Equilibrate

Target

Elute

Waste

S

W

PWash

W“Cousin animals” from : • Chromacon (Capture SMB)• GE (3C PCC)

Lysine production main steps

About 50,000 MT/year/unit

FERMENTATION

MEMBRANE FILTRATION

BIOMASS

ION EXCHANGEPURIFICATION

EVAPORATION

CRISTALLISATION

CRUDE L LYSINE HCl

ACIDIFICATION

Lysine purification: switch from batch to continous (a decade ago)

• First step: Strong Cation Exchange• Feed pH 3-4 so that Lys+ (maximizes loading and separation)• Then pH lowered to 1-2 so that Lys++ (maximizes capture of small

concentrations)• Wash • Elution by ammonia at pH 9 (Lys0)• Ammonia recycled through stripping

97% Lys with microbial byproducts and mineral cations

• Second step: Weak Cation Exchange• Feed at pH 9 (Lys0 goes through)• Regeneration at pH with diluted sulphuric acid• Wash

Gain continuous (about 10-column systems) vs Batch: resin inventory and effluent divided by about 3

Biopharma

Still looking for a first Biopharma industrial application of continuous chromatography ?

Many proof of concept published:• mAbs• pegylated proteins• peptides• fatty acids …..

Evolution ….

1960 1980 2000 2010 2020 ?

Num

ber o

f col

umns

Xylene

Sugars

I-SMBS-SMB

VariColMCSGP

MCSGPGSSRBioSc 3C PCC

12

6

4

24

2

Continuous character

100 %

50 %

Thank you !

Process integration may lead to non intuitive processes …

HFS 55 %

Glu 95 % Isomerization Glu-Fru 58-42SMB

Glu 90 %

HFS 55 %

Intuition

SMB

Glu 95 % Isomerization Glu-Fru 58-42

Glu 90 %

Fru 90 %

Optimized