Heikki Ojamo
Prof. / bioprocess engineering
M.Sc. (Tech.) Markus Räsänen
Tohtorikoulutettava
Fermentation of the fractions
from silage processing
12.3.2014
Heikki Ojamo
Markus Räsänen
Fractions studied
1. Silage juice
• Applicability as a replacer for high-value, complex components
in growth media for industrially relevant microbes (University of
Oulu / Laboratory of bioprocess engineering)
2. Hydrolyzate from extracted silage
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Feed protein production in Pekilo-fermentation
Applicability of the hydrolyzate as C/energy source for Pekilo
Requirements of supplementary nutrients in fermentation
Techno-economic parameters for the fermentation process
12.3.2014
Heikki Ojamo
Markus Räsänen
Silage juice as a component in
growth media
1. Inhibitory effects of weak organic acids (lactic/formic/acetic acids)
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•
E.coli and Bacillus subtilis : 100 % inhibition of growth with 50 %
juice concentration (replacing water)
L.rhamnosus, S.cerevisiae and Pichia pastoris: growth and glucose
consumption slow down and cellmass yield decreases by 25…50 %
2. Nutrients in the juice
• L.rhamnosus, S.cerevisiae and P.pastoris grow on the juice without
any supplementary nutrients (peptone and yeast extract)
• The addition of peptone and/or yeast extract into the juice have no
effect on growth, rate of glucose consumption or cellmass yield
=> Silage produced with enzyme treatment probably would produce
juice applicable as a nutrient source in media for growing microbes
Ruohosta proteiinia - loppuseminaari
12.3.2014
Heikki Ojamo
Markus Räsänen
Production of feed protein on
silage hydrolyzate
1. Why Pekilo (Paecilomyces variotii)?
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•
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Hydrolytic fungus
Tolerates inhibitors in hydrolyzates better than e.g. yeasts
Is even able to use furfural and HMF as C/energy source
A filamentous organism => easy to filtrate out from the broth to
high dry substance (30 % +)
Long history of use as SCP-product and approved feed
component
Protein content 50 – 60 %
Ruohosta proteiinia - loppuseminaari
12.3.2014
Heikki Ojamo
Markus Räsänen
Production of feed protein on
silage hydrolyzate
1. Inhibitory effect of the process fractions
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•
Hydrolyzate/extract –combination increases remarkably lag-time
The effect can be avoided by adaptation to the fractions (
hydrolyzate 1:5 = appr. 100 g/L glucose)
2. Pullokoesarja - CO2
1400
pO2 %
Glu + PYM
120
1200
H 1:14 + PU 1:10
600
H 1:16 + PU 1:10
400
200
H 1:18 + PU 1:10
pO2 %
CO2 ppm
80
800
60
40
20
0
0
-200
100
H 1:12 + PU 1:10
1000
0
50
100
150
H 1:20 + PU 1:10
0
20
40
60
80
Aika h
Aika h
Ruohosta proteiinia - loppuseminaari
12.3.2014
Heikki Ojamo
Markus Räsänen
100
Production of feed protein on
silage hydrolyzate
2. Consumption of glucose and acids in fermentation
•
In a fed-batch fermentation the final cell dry weight concentration was 37
g/L and cellmass yiedl on glucose was 0,49 g/g (0,49 – 0,67 g/g)
35
30,3
Konsentraatio g/l
30
25
60
20
50
16,8
15
16,3
13,5
12,5
10
0
-5
40
Muurahaishappo
g/l
Maitohappo g/l
30
Etikkahappo
20
Muurahaishappo
Glukoosi
Glukoosi g/l
1,72,1
0,6
0,40,9
1,4
0
-20
Etikkahappo g/l
5,8
5
47,047,6
20
40
Aika h
0,8
0,5
0,3
0,1
60
10
0,4
0,3
0,0
80
100
9,4 9,4
4,8 4,5
1,9 1,8
0
-20
0
20
13,3
10,2
6,4
4,4
3,0
0,9
0,4
0,0
40
Maitohappo
0,9
0,7
0,5
0,4
60
2,6
1,3
0,90,6
0,40,3
0,4
80
100
-10
Ruohosta proteiinia - loppuseminaari
12.3.2014
Heikki Ojamo
Markus Räsänen
Production of feed protein on
silage hydrolyzate
3. Need for supplementary nutrients
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The xtract can be replaced with e.g. CSL (5 g/25 g glucose) and
diammoniumhydrogen phosphate (5 g/25 g glucose)
4. Specific growth rate (µ = dX/(X dt))
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Based on fed-batch experiments µ
0,16 h-1
Continuous chemostat cultivation experiments done µ
0,12 h-1 with
max. glucose concentration 30 g/L limited by the oxygen transfer
capacity of the fermentor and practical arrangements in small-scale
(appr. 2-fold value achievable in large-scale meaning volumetric
cellmass productivity appr. 3,7 g/Lh]
5. Asepticity
•
Continuous fermentation was run > 500 h without any contamination
Ruohosta proteiinia - loppuseminaari
12.3.2014
Heikki Ojamo
Markus Räsänen
Production of feed protein on
silage hydrolyzate
6. Techno-economic parameters
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Continuous fermentation; residence time ≥ 6 h
Cellmass volumetric productivity 3,7 g/Lh
Cellmass yield on glucose 0,5 g/g
Cellmass yield on oxygen 0,85 g/g
Working volume of the fermentor 80 m3
Annual productivity of dry Pekilo 2400 t
CSL and diammoniumhydrogen phosphate consumption appr. 500 t/a
Power consumption in fermentation 5 000 MWh/a
Ruohosta proteiinia - loppuseminaari
12.3.2014
Heikki Ojamo
Markus Räsänen