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University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Microalgae between „Facts and Phantasy“
Rape seed ca. 1300 l/ha
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 1/33
Photosynthesis Growth C-Storage Harvest
Refinement934.025 l/ha
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Algal Biomass to replace fossil carbon ?
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm
What is the aim?
Production of organic carbon for high valuable products (Carotenoids, highly unsaturated fatty acids, proteins ?)Or :Production of feed stock or bulk chemicals ?
C/N
C/P
C/S
2/33
Or :Production of feed stock or bulk chemicals ?Or:Production of storable energy (Hydrogen, Methane, liquid fuels) ?
Growing Biomass: N:C = 1:7Energy requirement N:C = 0,35
Energy requirement per C: 115 kcal/MEnergy requirement per N: 289 kcal/M
C/S
C/O/H
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Energy from Biomass
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm
Two major criteria
1. The Potential What is the percentage contribution to the energy demand? High Efficiency needed
3/33
demand?
2. The EcobalanceWhat is the reduction in greenhouse emission per unit energy?
High Efficiency needed
Life cycle analysis needed
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Efficiency of Photosynthesis 1 red photon: 176 KJ/molC6H12O6: 2816 KJ/mol1 C needs: 468 KJ/mol8 photons needed: 468 *100 = 33%
8*176Real light: 209 KJ/mol, 50% PAROptimum quantum Yield: 10 P/CMaximum efficiency: 0,5* 468*100: 11,9%
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 4/33
Maximum efficiency: 0,5* 468*100: 11,9%10*209
Artificial red: 33% - real light: 11,9%
D. Walker, J. Appl Phycol 2009
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
2. The efficiency is low: what is the real efficiency?
AWD Larkum, Curr Opin Biotechn 2010
P/C35,6
34,7
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 5/33
34,7
33,8
33,9
38,5
1. P/C values do not vary in the light, 2. P/C values are much higher than 103. The overall efficiency is between 3-4 %!!
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Can we improve it? Balancing the energy flux from Photon to Biomass: the approach
light
cell
Qpharphotosynthetic electrons Biomass
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 6/33
3 Steps:• From incident to absorbed photons: Qphar• From „photosynthetic“ to „bio-synthetic“ electrons• From „bio-synthetic electrons“ to biomass
heatMetabolic Costs:- C3/C4- Photorespiration- Mehler-Reaction- ?
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Qphar depends on:
1. Spectral overlap between incident light and absorption
From incident to absorbed photons: Qphar
Qphar = numbers of absorbed photons per area and time
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 7/33
1. Spectral overlap between incident light and absorption spectrum of the algal cells
2. Pathlengths of the light: z3. The In-vivo absorption coefficient: a*ph)
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
In-vivo absorptions coefficient: a*ph
The numbers of pigments is not sufficient to Calculate the number of absorbed quanta perCell and time.
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 8/33
(from Kirk, 1990)
Pigment package is more important than pigmentPattern.
Dense pigment pattern does not only decrease theAbsorption efficiency but increases the Investment cost for the cell
This is the reason why antenna size decreaseincreases growth efficiency
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
5µm
Variation of a*ph as a function of cell size
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 9/33
(from Kirk, 1990)
90 µm
consequence: the biodiversity is limited to small speciesMarch 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
From „photosynthetic“ to „bio-synthetic“ electrons
O2H2O2
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 10/33
Red arrow indicate alternative electron cycling
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
PAM
Doing it in a proper way:
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 11/33
Chlorella
Clark Electrode
Ek
At the light intensity of Ek growth is minimaly light limited at optimum growth
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
How to model biomass production based on quantum efficiency
Model: PIEilers
a 0.00004b 0.02335c 2.66387
0 50 100 150 200 250 300 350
0
5
10
15
20
25
ET
R /
oxyg
en e
volu
tion
Qphar
Photosynthesis-Irradiance-Curve
Oxygen evolution
PAM-Fluorescence
fluorescence
08:00 10:00 12:00 14:00 16:00 18:00 20:00
0
100
200
300
400
500
600
oxygen
O2
[µm
ol m
g C
hl a
-1h-1
]
daytime
O2 = ΦPSII * Qphar * 0.125
12/33
integrated photosynthetic production
(oxygen evolution)
integrated net carbon fixationO2/CO2 gas exchangephotosynthetic/respiratory quotient
modelled biomass productionelemental compositiondry weight
carbon : biomassdry weight : Chl
measured biomass productiongrowth rate dry weight d-1
vs.
Comparison of modelled with weighted biomass allows the validation of the modelMarch 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
A case study with Chlamydomonas (full replete at Ek)
12,7% of absorbed light at EkMaximum growth rate at minimum dissipation
P/C = 24
13/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
„Sites of quantity of Losses“: (given in per cent of Qphar)1. Non-photochemical Quenching at Ek = 30%2. Alternative electron cycling: (Asada, PSII-Cycle) 10- 25%3. Dissipation: (F of RCs 80%, heat of exerg. reactions: 20 % (?)4. Respiration: (Protein biosynthesis, glykoneogenesis, transport): 10% (?)
Photosynthesis is efficient – metabolims for biosynthesis is not !
6,3% of incident light at EkMaximum growth rate at minimum dissipation
4,3% of incident natural daylight Average growth rate
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Beside of metabolic losses the reduction degree of the formed biomass strongly influences the quantum requirement per assimilated carbon.Photon requirement per carbon incorporated into biomass
theor. realwith a Carbohydrate:Lipid:Protein=3:2:5 21 ?with a Carbohydrate:Lipid:Protein= 6:1:3 11 ?
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 14/33
with a Carbohydrate:Lipid:Protein= 6:1:3 11 ?with a Carbohydrate:Lipid:Protein= 1:6:3 19 ?
How can we measure it ?
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
FTIR spectra reflect the macromolecular composition of the biomass
P=O stretch
Amid IIAmid I
Protein
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm 15/33
4000 3500 3000 2500 2000 1500 1000
O-H stretchC-O stretch
Kohlenhydrate
Wellenzahl cm-1
C-O stretchC-H stretch
PE
Nucleotide
rel.
Abs
orpt
ion
FTIR spectroscopy
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
How to extract from FTIR spectra quantitative information ?
16/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
Wagner et al. J. Biophotonics 2010March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Energy balance refined for the carbon pools in the cell
Photon requirement per carbon incorporated into biomass theor. measured
with a Carbohydrate:Lipid:Protein=3:2:5 21 28
17/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
with a Carbohydrate:Lipid:Protein=3:2:5 21 28with a Carbohydrate:Lipid:Protein= 6:1:3 11 12with a Carbohydrate:Lipid:Protein= 1:6:3 19 29
The deviation is the higher the more energy rich compoundsare accumulated. Why ?
The „Biosynthesis of a cell“ is a slow process
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
The deviation is the higher the more energy rich compoundsare accumulated. Why ? Wilhelm and Jakob, Tatup 2012
18/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Can we speed up the slow processes by bio-engineering?
19/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
The answer is: No – because of physical a) Because of the diffusion constants of macromoleculesb) Tripling the carbon content of a cell every hour is physically impossibleTherefore, down-regulation of photosynthesis under full sunlight is essential,because it adapts the time constants of metabolism with the time constants of the physics of light reactions.
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Summary:
1. The light reactions in photosynthesis are far too efficient to convert the captured light energy into biomolecules.
2. Energy dissipation by NPQ and alternative electron cycling are essential mechanismto balance the energy flux between absorption and the biosynthesis of cellular macromolecules.
20/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
macromolecules.
3. The electron partitioning is the rate litmiting step in making new cells
4. Under natural light conditions the photon requirement for fast growing cellscan not decreased below a value of 20 because of the metabolic costs which are inevitably associated with the biochemical conversion of sugars into protein and lipidsand the turn-over of both in light-dark cylces.
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
100
120
140
160
180
200G
HG
-Em
issi
on
s in
g C
O2-
eq. /
MJ
100
120
140
160
180
200
Biofuel
Fossil Fuel
Overall Emissions
Allocated Emissions
Opt PBR : 80 to/ha yrPess PBR: 50 to ha yrOpt OP: 40 to ha yrPess OP: 20 to ha yr
Allocated: Nutrients, Harvest, extraction, refinement
1) Increase in yield little effect !2) Allocated costs are crucial
21/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
0
20
40
60
80
PB
R o
pt
PB
R p
es
OP
opt
OP
pes
Rap
esee
d
Die
sel
PB
R o
pt
PB
R p
es
OP
opt
OP
pes
Whe
at
Gas
olin
e
PB
R o
pt
PB
R p
es
OP
opt
OP
pes
Ene
rgy
crop
s
Nat
ural
Gas
Biodiesel Bioethanol Biogas
GH
G-E
mis
sio
ns
in g
CO
0
20
40
60
80
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
22/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
23/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Photo-MethaneThe Concept
CO2+2 H2O CH4 + 2 O2
No: MixingNo: N, P, S
24/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
No: N, P, SNo: HarvestNo: Refinement
14 (!) conversion stepsfrom CO2 to CH4
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
(from Günther et al. 2013)
Normally glycolate production stops after several hours due to CCM and cell internal metabolization of glycolate (C2-pathway).
Inhibition of glycolate oxidation enhances glycolate excretion
25/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
(from Günther et al. 2013)
The combination of inhibition of glycolate oxidation and CCM leads to highand stable glyocolate excretion.
26/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
(from Günther et al. 2013)
Transgene Chlamydomonas cells without CCM and inactived GDH-Activity show constant glycolate excretion during the whole light period.
0.7
0.8
0.9
1
rela
tive
Gly
kola
texk
reti
on
[ra
tio
]27/33
University of Leipzig - Department of Biology Plant Physiology unit - Prof. Dr. Christian Wilhelm
Wt rates:Measured from freshly harvested cells
0
0.1
0.2
0.3
0.4
0.5
0.6
0 1 2 3 4 5 6 7Zeit [h]
rela
tive
Gly
kola
texk
reti
on
[ra
tio
]
WT
K5-10
K5-15
Transgenic rates:Measured from continuous culture
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
(from Günther et al. 2013)
A selected consortium of archaea converts glycolate to CH4 and CO2 in a Stoichiometric ratio of 5:4. After CO2 elimination the biogas can be used Without any refinement for all purposes (also for cars).
80
100
Bio
gasa
ntei
l [V
ol.%
] 800
1000O2 N2 CH4 CO2 H2S
Glykolatzugabe = 627mg/d, konstant Verweilzeit = 417d
28/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
0
20
40
60
80
610 650 690 730 770 810 850 890 930
Zeit [d]
Bio
gasa
ntei
l [V
ol.%
]
0
200
400
600
800
H2S
[ppm
]
04/2011 12/2011
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
(from Günther et al. 2013)
The technical design for the future reactor
29/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
Source: KIT VIP 2012
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
31/35University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Research needed
1. Instead of fast growing species – biofilm algae2. Instead of accumulation of storage products – Optimized Excretion3. Instead of improved photosynthesis – Metabolic engineering of C-ass.4. Instead of cells with fast nutrient uptake – cells with „zero growth“
32/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
4. Instead of cells with fast nutrient uptake – cells with „zero growth“5. Instead of volume reactors – Biofilm-carrier material
Final RemarksRe-start of research needed
Respecting the principles of Constructive Technology Assessment (CTA)
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
Thanks for your PatienceThanks for your PatienceThanks for your PatienceThanks for your Patience
Reimund Goss
Single cell analysis
Photonenblances
Heiko Wagner
33/33University of Leipzig - Department of Biology
Plant Physiology unit - Prof. Dr. Christian Wilhelm
Norbert Räbiger, BremenSaskia John, Bremen
Clemens Posten, KarlsruheLinda Oeschger, Karlsruhe
Torsten Jakob
FTIR-SpektroskopiePhoto-MethanTheresa Quaas Anja Günther
March 17th 2014 Meeting of the “Deutsche Physikalische Gesellschaft
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