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Journal of Scientific & Industrial Research

Vol. 65, January 2006, pp. 72-76








Bakers yeast production under fed batch culture from apple pomace
Shashi Bhushan
1
and V K Joshi
2,
*

1
Institute of Himalyan Bioresource Technology, CSIR, Palampur 176 061
2
Department of Postharvest Technology, Dr Y S Parmar University of Horticulture and Forestry, Nauni-Solan, 173 230
Received 31 December 2004; revised 30 September 2005; accepted 10 October 2005
Apple pomace extract (APE) in aerobic variable fed batch culture under standardized fermentation parameters was
evaluated for the production of bakers yeast. The substrate was fed at a flow rate of 0.39 dm
3
h
-1
to regulate fermentable
sugar concentration between 1-2 % in bioreactor. An average specific growth rate of 0.24 h
-1
and cellular yield coefficient of
0.48 g/g sugar was obtained during bakers yeast production. Under variable fed batch aerobic bakers yeast fermentation,
yield obtained with APE was 96 % to that of expected theoretical yield and thus, qualified as an alternative to molasses, the
traditional bakers yeast production carbon source. The dough raising capacity of experimentally produced yeast cells
revealed no apparent difference from that of commercial sample.
Keywords: Aerobic fermentation, Apple pomace extract, Bakery products, Bakers yeast, Fed batch culture, Molasses,
Saccharomyces cerevisiae, Specific growth rate
IPC Code: C12N1/18

Introduction
Apple pomace (AP), a leftover residue after apple
juice extraction, poses a lot of problem of
environmental pollution and a loss of nutritive natural
wealth
1
. High biochemical oxygen demand (BOD,
395-37000) of AP makes it easily vulnerable to
fermentation / biodegradation, consequently if not
disposed-off immediately, would generate foul
smell
2,3
. The nutritive value of AP rests mainly on its
total carbohydrates, crude protein, crude fat, fibres
and minerals. AP can be used for the preparation of
jam, sauce, soft drinks, animal feed along with direct
incorporation in bakery products
4-6
, besides
production of citric acid, ethanol, acetone, butanol
and microbial colour
7-9
. Due to high nutritive value
and easy fermentability, it is suitable for the
production of bakers yeast. The fermentable sugar
was found enough in apple pomace extract (APE) to
support the growth of bakers yeast along with crude
protein contents as well as a few trace elements
3,10,11
.
Besides type and concentration of carbon sources,
other factors such as dissolved oxygen (DO) agitation,
pH and temperature, are also known to influence the
fermentation behaviour and overall biomass
production
12-15
. The fermentation parameters such as
temperature, pH and DO, standardized during earlier
investigation
16
, were employed in the present
fermentation.

This paper presents the results of substitution of
molasses (a traditional substrate) by APE and its
effect on the fermentation characteristics such as
specific growth rate (SGR), total biomass yield,
cellular yield coefficient (CYC) and other related
fermentation characteristics.

Materials and Methods
Apple Pomace Extract (APE)
The dried AP, procured from HPMC (Horticultural
Produce Processing and Marketing Corporation),
Fruit processing unit, Parwanoo (HP), was ground
into fine powder, diluted with water in 1:6 ratio and
boiled for h before extraction.The boiled material
was pressed in hydraulic press and the liquid filled
into conical flasks (5 l), was sterilized at 121C for
20 min.

Inoculum
Preserved slant of commercial bakers yeast
(Saccharomyces cerevisiae var. diastatic) was used to
inoculate in yeast malt extract broth and incubated at
30C for 24 h and then, transferred to APE medium
and again incubated at the same temperature for
another 24 h. Inoculum (10
2
-10
3
cells/ml) was added
at the rate of 1 percent to initiate the fermentation.
__________
*Author for correspondence
vkjoshipht@rediffmail.com, winevkj@yahoo.com
BHUSHAN & JOSHI: BAKERS YEAST PRODUCTION UNDER FED BATCH CULTURE FROM APPLE POMACE


73
Fed-batch Cultivation of Bakers Yeast
Replicated fermentations were performed in a
variable fed batch mode in a 5 l bioreactor (Bioflo,
2000, New Brunswick Sci. Co., New Jersy), which
was computer controlled having advanced
fermentation software (Fig. 1). APE (1:6) contained
fermentable sugar (40-45 g l
-1
), supplemented with
yeast extract and beef extract (0.3% each), peptone
(0.5%) and ammonium sulphate (1.8 %). The
fermentation conditions were as per the standardized
parameters used in an earlier experiments
12
. The
initial volume in the vessel of fermenter was kept 1.5 l
and rest of the medium of same concentration was fed
incrementally at a flow rate of 0.39 dm
3
h
-1

to get the
final fermentation volume at 4.5 l. After inoculation,
the fermentation was run in a batch mode for 1 h as a
terminal cell maturity (TCM) step whereas, agitation
and continuous oxygen supply as per the standardized
conditions, was provided. The same was carried out to
mature the cells prior to running the fermentation in
the fed batch mode.
Analytical Methods
The AP medium was initially analyzed for pH and
fermentable sugar. The samples were drawn after an
interval of 2 h from the vessel for the off-line
determination of sugar, pH, cell biomass etc. The
reducing sugar was determined by Nelson Somogyi
method
17
while pH was estimated using digital pH
meter (3030 ELTOP). The weight of fresh biomass
18

after drying at 70+1C was determined.
Fermentation Kinetics
Fermentation kinetics parameters (SGR, CYC and
total biomass) were determined
20,21
, whereas
generation time was calculated
22
. The biomass at any
point of time in a variable fed batch system is:
x
t
= x
0
+Y (S
R
-S) (1)
where x
t
and x
0
are the total amount of biomass in
bioreactor at any time t and zero time, respectively.
S
R
is the initial substrate concentration; S is the
residual substrate concentration and Y is the yield
factor (g biomass/g substrate consumed).
The specific growth is determined by
F.S
R
= u (X/Y) (2)
F.S
R
. Y
u = (3)
X
where X = x v, cell concentration and volume at time
t, respectively. In the equation, F is the flow rate
of medium (dm
3
h
-1
) incrementally fed to the
bioreactor, X the total biomass in a bioreactor, Y
the yield factor and u is the specific growth rate.
The generation time (G) was calculated as:
G = t/n = 1/v (4)
where t is the time, n number of divisions and v
is number of division per hour.
Quantity of CYC was calculated on the basis of (g)
biomass produced at a time t from the total sugar
consumed at that time and volumetric yield obtained
(g biomass/100 ml medium). The total biomass yield
(X) was the cumulative gram biomass produced
during the fermentation from the total medium or
substrate fed to the bioreactor at that particular time
as:
X = x
t
x V
t
(5)
where x
t
and V
t
are the cell biomass and volume of the
medium respectively at a time t. Sugar consumed at
any time was calculated from a material balance
considering sugar fed, sugar present in the bioreactor
and sugar withdrawn during sampling, and expressed
as glucose equivalent.
Dough Raising Capacity (DRC)
A known volume of dough (made with respective
yeasts) was taken in a 250 ml measuring cylinder,

Fig. 1 Schematic representation of bakers yeast production
1) Exhaust port; 2) Head plate; 3) Impeller; 4) Cooling; 5)
Aeration supply; 6) Heating pad; 7) Air flow meter; 8) Water
inlet; 9) Temperature sensor; 10) Incremental feeding; 11) Water
outlet; and 12) DO probe
J SCI IND RES VOL 65 JANUARY 2006


74
smeared with thin layer of oil. It was then kept at
37C and subsequent increases in volume were
recorded at regular intervals. Comparative leavening
activity of experimental yeast produced from APC
along with commercial bakers yeast was made.
Results and Discussion
A general increase in sugar consumption or
decrease in residual sugar concentration was found
with progress in fermentation (Table 1). Initial
average sugar concentration significantly decreased
during first hour of fermentation and thereafter,
remained between 1-2 percent till final hour of
fermentation at terminal cell maturity (TCM), when it
further decreased to a significantly lowest
concentration. The decline was as per expectations in
such type of fermentation, wherein the fermentable
sugar present in the medium is either respired or
fermented by the yeast. A significantly lower quantity
of ethanol was observed during initial few hours of
fermentation indicating that sugar was mainly
respired by the yeast. It increased linearly with the
progress in fermentation time and at the end of
fermentation, a significantly highest concentration of
mean ethanol was recorded. With the increase in
sugar consumption, an increase in volumetric yield of
biomass is obvious in an aerobic fermentation
23,24
. A
continuous increase in volumetric yield with time
took place. The volumetric yield (1.82%) was
recorded at the end of fermentation.
SGR, an important fermentation characteristic of
bakers yeast production, is negatively correlated to
CYC
14,25
. Mean SGR (Table 1) was higher during
initial stages, which gradually declined towards the
end of fermentation. The overall average SGR
throughout the fermentation period remained at
0.24 h
-1
, well in the range (0.18-0.25 h
-1
) optimized
for higher biomass production from glucose and
molasses medium
14,15,26,27
. The higher SGR during
initial hour might be due to the higher sugar
concentration
25
. Similar was the trend with CYC. The
initial rise observed in CYC might be attributed to the
higher yeast efficiency, which was generally more
during early stages and the sugar present during that
phase mainly used for its own growth and
propagation. The overall average CYC (0.48 g/g
sugar consumed) is in conformity with the earlier
findings (0.44-0.54 g/g glucose), based on glucose
and molasses
medium
13,28-32
. A clear relationship was observed
between SGR and CYC, which is the characteristic
feature of fed-batch culture
33
. As per expectation, the
biomass yield increased significantly throughout the
fermentation period. Under aerobic conditions, in
general, a higher biomass yield is obtained
25,34
(85.23
g) at the end of fermentation (overall mean biomass
yield 47.03 g). Total cumulative biomass produced
during this experiment was 85.23 g from 179.26 g of
sugar consumed (CYC, 0.476 g/g sugar) while the
Table 1 Effect of optimized parameters on the growth characteristics** of bakers yeast under fed batch culture
Time (h) Residual sugar
% glucose
Ethanol
%
Volumetric
yield %
Specific growth
rate
h
-1

Cellular yield
coefficient
g/g sugar
Total biomass
yield
g
Theoretical
biomass yield
g
Deviation
0 4.34+0.130 0.00 0.05+0.001 - - - - -
1* 2.15+0.171 0.03+0.018 1.03+0.091 0.32+0.010 0.52+0.019 15.46+1.164 16.42+1.067 0.097
3 1.87+0.044 0.08+0.006 1.15+0.050 0.28+0.021 0.47+0.021 26.20+1.199 28.12+0.725 1.924
5 1.76+0.052 0.24+0.087 1.26+0.035 0.25+0.029 0.46+0.016 38.60+1.143 39.43+0.310 0.833
7 1.56+0.140 0.34+0.054 1.30+0.059 0.24+0.016 0.45+0.022 49.77+2.243 53.44+1.423 3.669
9* 1.30+0.041 0.46+0.032 1.47+0.084 0.21+0.024 0.47+0.017 67.91+1.679 70.22+0.630 2.309
10 0.46+0.160 0.54+0.047 1.82+0.090 0.18+0.033 0.48+0.009 85.23+1.855 89.01+1.227 3.782
Mean 0.94 0.41 1.34 0.24 0.48 47.03 49.11 2.248
CD(p<0.05) 0.217 0.058 0.101 0.024 0.021 7.572 8.745
*Incremental feeding start and stops
**Mean+SE

BHUSHAN & JOSHI: BAKERS YEAST PRODUCTION UNDER FED BATCH CULTURE FROM APPLE POMACE


75
theoretical yield was 89.01 g (CYC, 0.5 g/g sugar)
from the same amount of sugar consumed. Thus,
experimental yield is slightly less than that of
expected yield (Table 1) and found to be around
96 per cent of the expected yield (Fig. 2). It can be
taken as a measure of success for use of AP as an
alternative substrate for the production of bakers
yeast.
In general, the dough raising capacity was higher in
control than the experimental yeast (Fig. 3). The
initial phase seems to be an activation period,
probably due to the transfer of yeast metabolism from
aerobic to anaerobic. After first hour of slow
fermentation, there was a continuous increase in
volume for about 3.5 h, and thereafter a slight but
stable increase took place. So, experimentally
produced yeast possessed similar activity to that of
commercial yeast available in the market.
Comparatively low volume development in all the
treatments might have been affected by the dough
ingredients, retention of CO
2
in the dough,
fermentation temperature, pH, trehalose content of the
yeast etc.
19,23,24,35-39
. While the yeast leavening in
dough is generally affected by the amount of CO
2

production during fermentation and its retention in
dough system.
Conclusions
AP performed well as a carbon source for the
growth of bakers yeast. Also, the optimized
parameters performed quite well and gave maximum
yield under established fermentations conditions. The
repeated fermentations were consistent, with little or
no deviation and had similar pattern for respective
growth characteristics during fermentation of bakers
yeast.
References
1 Joshi V K, Apple pomace utilization- Present status and
future strategies, in Advances in Biotechnology, edited by
Ashok Pandey (Educational Publishers and Distributors,
New Delhi) 1998, 141-155.
2 Hang Y D & Walter R H, Treatment and utilization of apple
processing wastes, in Processed Apple Products, edited by
Donald L Downing (AVI Publ, New York) 1989, 365-377.
3 Joshi V K, Jaswal S & Lal B B, Apple pomace: Effect of
sulphur dioxide and temperature on its preservation and
medium optimization for yeast biomass production, J Sci Ind
Res, 57(1998) 692-697.
4 Smock, R M & Neubert A M, Apples and Apple Products
(Interscience Publ, New York) 1950.
5 Wang H J & Thomas R L, Direct use of apple pomace in
bakery products, J Food Sci, 54 (1989) 618-620.
6 Joshi V K & Kaushal N K, Composition of apple pomace
standardization of pulp making, preparation and evaluation
of apple jam, Res Ind, 40 (1995) 203-207.
7 Sandhu D K & Joshi V K, Development of apple pomace
medium optimization of conditions for pigment production
by Rhodotorula, Adv Food Sci, 19(1/2) (1996) 31-34.
8 Joshi V K & Sandhu D K, Preparation and evaluation of
animal feed using solid state fermentation of apple pomace,
Biores Technol, 56 (1996) 251-255.
9 Attri D & Joshi V K, Optimization of apple pomace based
medium and fermentation conditions for pigment production
by Micrococcus species, J Sci Ind Res, 64 (2005) 598-601.
10 Bhushan S, Joshi V K & Kaushal B B Lal, Process
development and evaluation for the production of bakers
yeast using apple pomace as a carbon source, Indian Fd
Packer (submitted).
11 Joshi V K, Bhushan S, Attri D & Lal B B, Utilization of
apple pomace in comparison to molasses and jaggary, under
fed-batch cultivation for bakers yeast production, Acta
Horticulture (in press).

Fig. 2 Comparison of theoretical and experimental biomass
yield of bakers yeast with optimised parameters

Fig. 3 Comparison of dough raising capacity (DRC) of bakers
yeast from apple pomace extract (experimental yeast) with control
yeast
J SCI IND RES VOL 65 JANUARY 2006


76
12 Oura E, Effect of aeration intensity on the biochemical
composition of bakers yeast I Factors affecting the type of
metabolism, Biotechnol Bioeng, 16 (1974) 1197-1206.
13 Wang H Y, Cooney C L & Wang D C Computer aided
bakers yeast fermentation, Biotechnol Bioeng, 19 (1977)
69-86.
14 Aiba S, Nagai S & Nishizawa Y, Fed-batch culture of
Saccharomyces cerevisiae. A perspective of computer
control to enhance the productivity in bakers yeast
cultivation, Biotechnol Bioeng, 18 (1976) 1001-1016.
15 Ferrari M D, Bianco R, Froche C & Loperena M L, Bakers
yeast production from molasses/cheese whey mixtures,
Biotechnol Lett, 23 (2001) 1-4.
16 Bhushan S & Joshi V K, Effect of temperature, pH,
dissolved oxygen and agitation on the growth of bakers
yeast under variable fed batch culture, Adv Appl Microbiol
(submitted).
17 Sadasivam S & Manickam A, Biochemical Methods, 2nd
edn (New Age International, New Delhi) 1996.
18 Snell F D & Ettre L S, Yeast, in Encylopedia of Industrial
Chemical Analysis (Inter science Publishers, New York)
1974, 514-543.
19 Trevelyan W E & Harrison J S, Studies on yeast
metabolism: The trehalose content of bakers yeast during
anaerobic fermentation, Biochem J, 62 (1956) 177-183
20 Pirt S J, Principles of Microbe and Cell Cultivation (Blacks
Well, Oxford) 1975.
21 Yoshida F, Yamane T & Nakamoto K, Fed-batch
hydrocarbon fermentation with colloidal emulsion feed,
Biotechnol Bioeng, 15 (1973) 257-270.
22 Stainier R Y and Doudoroff M & Adelberg E A (General
Microbiology. McMillan, India) 1970, 298-324.
23 Oura E, The production of energy during aerobic growth of
bakers yeast on glucose and ethanol, Proc 2nd Congress on
Yugoslav Microbiology, Opatija, 1972, 237-246.
24 Oura E, The effect of aeration on the growth energetic and
biochemical composition of bakers yeast, Res Lab State
Alcohol Manopoly, Helsinki, 1972.
25 Reed G, Production of bakers yeast, in Prescott and Dunns
Industrial Microbiology, 4th edn, edited by G Reed (AVI
Publishing Co., Westport, CT) 1982, 583-633.
26 Von Meyenburg H K, Energetic of the budding cell of
Saccharomyces cerevisiae during glucose limited aerobic
growth, Arch Microbiol, 84 (1969) 254-260.
27 Van Hoek P, Van Dijken J P & Pronk J T. Effect of specific
growth rate on fermentation capacity of bakers yeast, Appl
Environ Microbiol, 64 (1998) 4226-4233.
28 Dellweg H, Bronn W K & Hartmeir M, Respiration rate of
growing and fermenting yeast, Ke.-Kemi, 4 (1977) 611-615.
29 Chen S L, Carbohydrate assimilation in actively growing
yeast. S cerevisiae. I. Metabolic pathways for C
14
glucose
utilization by yeast during aerobic fermentation, Biochem
Biphys Acta, 32 (1959) 470-479.
30 Chen S L, Carbohydrate assimilation in activity growing
yeast. S. cerevisiae. II. Synthesis of polysaccharides from
C
14
glucose, Biochem Biophys Acta, 32 (1959) 480-483.
31 Chen S L & Gutmanis F, Carbon dioxide inhibition of yeast
growth in biomass production, Biotechnol Bioeng, 18 (1976)
1455-1462.
32 Vijayalakshmi G & Basappa S C, Enhancement of yield
activity bakers yeast by carbon compounds added during
ripening, Indian J Microbiol, 36 (1996) 189-191.
33 Stanbury P F, Whitaker A, & Hall S J. Principles of
Fermentation Technology (Aditya Books (P) Ltd., New
Delhi) 1997, 13-33.
34 Basappa S C, Bakers yeast: Production, quality and
utilization, in Biotechnology: Food Fermentation-
Microbiology, Biochemistry and Technology (Vol. II), edited
by V K Joshi & Ashok Pandey (Educational Publisher and
Distributors, New Delhi) 1999, 1113-1144.
35 Chen S L, Cooper E J & Gutmanis F, Active dry yeast:
Protection against oxidative deterioration during storage,
Food Technol, 20 (1966) 79-83.
36 Suomalainen H, Changes in the cell constitutions of bakers
yeast in changing growth conditions, Pure Appl Chem, 7
(1963) 634-654.
37 Sato T, Tsumura N, Tanaka T, Onada T & Koyanag Y,
Comparison of practical characters of bakers yeast of the
world, Nippon Jozo Kvokai, 15 (1961) 74-80.
38 Reed G & Peppler H J, Yeast Technology (AVI Publishing
Co., Westport, CT) 1973.
39 Watson T G, Effect of sodium chloride on steady state
growth and metabolism of S cerevisiae, J Gen Microbiol, 64
(1970) 91-99.