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Journal of Microbiology Research and Reviews

Vol. 3(3): 24-42, July, 2015

ISSN: 2350-1510


Bioprospecting Starchy Feedstocks for Bioethanol

Production: A Future Perspective
Suman Jagatee1, Shuvashish Behera2, Preeti Krishna Dash1, Santilata Sahoo1* and Rama
Chandra Mohanty1

Department of Botany, Utkal University, Bhubaneswar 751004, Orissa, India

Biochemical Conversion Division, Sardar Swaran Singh National Institute of Renewable Energy, Kapurthala-144601,
Punjab, India

Equal contributors
Email for correspondence: santi.bot.uu@gmail.com
Ethanol is one of the main bio-based molecule produced worldwide, mainly from corn and other starchy
crops. Starchy substrates employed as raw materials for ethanol production which cannot be directly
fermented to ethanol by certain microorganisms. This is because yeasts, which are employed for
fermentation, cannot utilize the starch molecules made up of long chains of glucose molecules hence
prior to fermentation need to hydrolyze to simpler fermentable sugars into simple glucose molecules.
Although corn starch to ethanol is a mature process, corn production is not feasible in different region.
Except corn starch, there are some low impact tuber crops including cassava, sweet potato, yam,
aroids, sugar beet, etc. which offer a viable alternative starchy raw material that can be converted to
useful sugar feed stocks needed for the production of ethanol and other value added products. It
requires simultaneous saccharification and fermentation for a better ethanol yield. Production of this
renewable fuel, especially from starchy materials such as tuber crops, holds a remarkable potential to
meet the future energy demand because of its high production and comparatively less demand for use
as food and fodder. Thus, the present review focuses on various starchy crops for bioethanol
production, fermentation techniques and microorganisms used in fermentation process along with its
future prospective.
Keywords: Starchy feedstocks, pretreatment, saccharification, fermentation, bioethanol

The global rise in energy consumption, the depletion of low extraction cost fossil fuel reserves, and climate change are
forcing the search for new energy sources and alternative ways to power the worlds motor vehicles (Bhatia et al., 2012;
Glithero et al., 2013). Concerns about energy security and the environmental impacts of energy production have led to
the implementation of policies designed to encourage the production and use of renewable bioenergy (Dias et al., 2013;
Glithero et al., 2013). The increasing need for ethanol as an energy source has stimulated worldwide investigations in
search of cheaper substrates for bulk ethanol production. Therefore, a considerable emphasis was thrusted on the
development of biofuel production technologies from plant sources and bioethanol production from plant biomass. The
industrial processes for bioethanol production mainly use grain, tuber and root starches (Tasic et al., 2009). A number of
less expensive substrates such as corn stover (Bondesson et al., 2013), cassava bagasse (Amenaghawon et al., 2013),
pine apple (Zain et al., 2012), jerusalem artichoke juice (Onsoy et al., 2007), mahula flowers (Behera et al., 2011),
sugarcane molasses (Nofemale et al., 2012; Behera et al., 2012) and sweet potato (Zhang et al., 2011) have been used
for such purposes.
The consumption of foods from root and tuber crops is considerably smaller in developing countries. Thus surplus


tuber crops that are not used for food can be utilized for production of fuels without affecting the food supply for the
human beings. In India, major tuber crop productions are cassava and sweet potato as compared to yam and other
tuber crops. Therefore, among various starchy materials available throughout the world; corn, wheat, potato, sweet
sorghum, sweet potato and cassava have been successfully utilized for the commercial production of bioethanol (Zhang
and Oates, 1999; Duvernay, 2008; Zhang et al., 2011). In general, root and tuber crops are preferred for the bioethnaol
production due to high starch content (80%) (Hoover, 2001). The study also indicates that the root crops have greater
potential than corn grains as ethanol sources, if economical harvesting and processing techniques are developed
(Thatoi et al., 2014). Starch is a complex carbohydrate which needs conversion into simpler sugars before being
converted into ethanol. The ethanol fermentation processes from these starchy materials commonly involves two stages:
(i) liquefaction of starch by -amylase and enzymatic saccharification of the low molecular weight liquefaction products
such as dextrin to produce glucose;
(ii) fermentation of glucose to ethanol. In this process, the energy input of the first stage is about 30-40% of the total
energy during bioethanol production from starch (Lin and Tanaka, 2006; Lee et al., 2012).
Yeasts, particularly Saccharomyces cerevisiae is the first choice for industrial ethanol production, because of their
good fermentative capacity, high tolerance to ethanol and other inhibitors (either formed during raw materials pretreatments or produced during fermentation) and the capacity to grow rapidly under the anaerobic conditions that are
characteristically established in large scale fermentation vessels. Hexoses are readily converted to ethanol by the
conventional yeast S. cerevisiae (Agbogbo and Kelly, 2008; Watanabe et al., 2007). In recent years, research has been
focused on the processes involving gram negative anaerobic bacterium, Zymomonas mobilis,because of several better
fermentation attributes such as conversion of glucose to ethanol and CO2, grows more rapidly and demonstrates highest
ethanol productivity at industrial-scale during fermentation when compared to other fermentation organisms. Further, the
ethanol tolerance of Z. mobilis is comparable with the strains of S. cerevisiae and produce less by-products (Busche et
al., 1992). Further, the implementation and development of the SSF, partial-SSF (PSSF) and separate hydrolysis and
fermentation (SHF) are some of the important advances related to overall process from different starchy feedstocks in
the industrial scale of ethanol production (Montesinos and Navarro, 2000).
Based on these observations, the present review summarizes the progress made on agro-industrial technology for
production of bioethanol from different starchy feedstocks with respect to its overall conversion technology. This review
also provides a brief summary of the current challenges and emphasizes on its future perspectives.
An overview of starchy feedstocks
The economics of ethanol production by fermentation are significantly influenced by the cost of the raw materials, which
accounts for more than half of the production costs. The type of feedstock chosen for ethanol production has a
significant impact on the design of fermentation process. In recent years, efforts have been directed towards the
utilization of cheap and renewable agricultural resources, such as starch as the alternative substrate for commercial
ethanol production. There are different starchy crops like cassava (Manihot esculenta), potato (Solanum tuberosum),
sorghum (Sorghum sp) grains and sweet potato being utilized for the bioethanol production (Maarel et al., 2002; Mojovic
et al., 2009). The development process for the bioethanol production from starchy feedstocks has been depicted in
Figure 1.
Starch is a well-known polymer of glucose linked bonds. The two major types of molecules in starch are amylase and
amylopectin (Maarel et al., 2002). Amylase, the minor linear polymer in sweet potatoes, consists mainly of -1,4 linked
D-glucopyranosol residues (Hoover, 2001). It contains up to 6000 glucose units. Amylopectin, the major polymer in
sweet potatoes is composed of short -1,4 linked linear chains of 10-60 glucose units and -1,6 linked side chains
containing 15-45 glucose units. Amylopectin contributes to the side branching of starch. One molecule of amylopectin
usually contains around 2,000,00 glucose units, making it one of the largest molecules existing in nature. The different
type of saccharification and further fermentation for various starchy feedstocks are summarized in the Table 1.
Starchy feedstocks for bioethanol production
Cassava (Manihot esculenta), the tropical root crop, is the third most important source of calories in the tropics, after rice
and corn. According to FAO, more than 600 million people depend on cassava in Africa, Asia and Latin America (Onu
and Edon, 2009). Africa is the largest centre of cassava production which is grown on 7.5 million hectare and produces
about 60 million tonnes per year. Other important cassava producing countries are Brazil (upcoming), Indonesia,


Figure 1. Bioethanol production process from starch based feedstocks

Thailand, Congo and Mozambique (upcoming) (Kuiper et al., 2007; Qiu et al., 2010). It gives the highest yield of starch
per hectare of any crop ranking it as 4th crop in the worlds production after rice, wheat and maize. It can be considered
as a suitable substrate for the production of bioethanol because it is a starch-rich (sugar content is more than 35%),
cold-resistant, fast-growing crop and could grow on a poor soil (Akpan et al., 2004). Cassava is cultivated as an annual
crop in tropical and subtropical regions for its edible starchy tuberous root, a major source of carbohydrate. The plant is
tolerant to extreme stress condition like drought which fits nicely within traditional farming systems (El-Sharkawyand
Mabrouk, 2007). On infertile land where the cultivation of other crops is difficult, unless considerable inputs are applied,
cassava still has a reasonable yield. The major industrial uses of fresh cassava roots are production of chips, pellets and
starch. However, the high starch content (70-85%) makes this crop a suitable feedstock for bioethanol production (Balat
et al., 2008; Patrascu et al., 2009).
Zamora et al. (2010) carried out the study of the optimization process for producing ethanol from cassava starch
through saccharification and further fermentation using Saccharomyces cerevisiae. The development of a semicontinuous process showed an 89.84% conversion of starch yielding an ethanol concentration of 49.76%. Shanavas et
al. (2011) used new enzymes like SpezymeXtra and Stargen 001 for bioethanol production from cassava starch
which yield around 558 g ethanol/kg starch with 98.4% fermentation efficiency. The study also showed that Spezyme
level beyond 20.0 mg for 10% (w/v) starch slurry was not critical for optimizing bioethanol yield from cassava starch.
However, Nuwamanya et al. (2011) investigated the feasibility of using non-food parts of cassava for energy production.
The promising results revealed that at least 28% of peels and stems comprise dry matter, and 10 g feedstock yields >8.5
g sugar, which in turn produced >60% ethanol, with pH 2.85, indicating a potential use of cassava feedstock for
ethanol production. Further, Ademiluyi and Mepba (2012) used cassava flour for the production of ethanol from 5
different types of cassava types and concluded highest ethanol production (0.61 ml/g of cassava flour) from TMS
92B/00068 cassava variety. While another variety of cassava TMS 98/0505 produced 0.42 ml ethanol per g of cassava
flour. The different type of saccharification and further fermentation from cassava is summarized in the Table 1.
Recently, industrial experiments have been developed to obtain starch and ethanol from potato based on the utilization
of wastes of potatoes destined to human consumption, thereby achieving a very low net energy balance (Vazirzadeh
and Robati, 2013). Although there are examples of genetic engineering approaches to improve production of ethanol
from crops (Torney et al., 2007), there has been no industrial development based on the specific potato cultivars
optimized for the ethanol industry due to its high cost for fuel ethanol production. Liimatainen et al. (2004) analyzed the
production of bioethanol from waste potatoes that correspond to approximately 5 to 20% of crops and obtained certain
by-products. Rani et al. (2010) used potato tuber flour for ethanol production through liquefaction with -amylase (2.05
DUN U/g starch) at 80C for 30 min followed by saccharification with glucoamylase (20.5 GA U/g starch) at 60 C for 2h
which generated 15.2% of total reducing sugars in the hydrolysate. In this study, they obtained 56.8 g l of ethanol by
fermentation of hydrolysate using Saccharomyces cerevisiae at 30 C for 48h. Arapoglou et al. (2010) studied a number
of batches of potato peel waste (PPW) which were hydrolyzed with three types of enzymes releasing 18.5 g l reducing


Table 1. The different type of saccharification and further fermentation processes for different feed stocks.
Sl. No


Liquifaction and


Sweet sorghum
Sweet potato
Raw Sweet Potato


Fermentation condition


Amylase, cellulase
Amylase, cellulase
-amylase and



time (h)

Corn cob

Amylase, cellulase
A. niger






-amylase and



Sweet potato flour






S. cerevisiae
Saccharomyces sp.
Saccharomyces sp.
S. cerevisiae
Sacharomyces sp.
Cocultureof A.niger
and S. cerevisiae

7.95% (v/v)

Chen et al. 2014

Prasad, 2014
Prasad, 2014

10.08% (v/v)

Prasad, 2014
Itelima et al. 2013


Sweet potato

-amylase and


S. cerevisiae
Co-culture of
Trichoderma sp. and S.
S. cerevisiae


Waste Sweet
Waste potatoes

-amylase and
-amylase and Termamyl



Z.mobilis AX101





95% (v/v)

Vazirzadeh and
Robati, 2013

Sulphuric acid, cellulose


S. cerevisiae


Erdei et al. 2013


Wheat meal
Wheat straw
Sweet sorghum

cellulose Cellic CTec3




91.9 kg

Li et al. 2013


Sweet potato
Wheat straw

Sulphuric acid and -amylase

Perchloric acid





Waste potatoes





Sweet potato

-amylase and



S. cerevisiae TSH1
Z. mobilis TSH-01
S. cerevisiae
Co culture of S.
cerevisiae and P. stipitis
S. cerevisiae (ATCC
BCRC 21494 + A.
oryzae BCRC 30289
and M. purpureus BCRC
31615 (1:2)



Sweet potato



91.39% (v/v)

Kumar et al. 2014

Duhan et al. 2013

Swain et al. 2013

Lareo et al. 2013

Dewan et al. 2013

11.42 g/l

Ferrari et al. 2013


Izmirlioglu and
Demirci, 2012

Ismail et al. 2012

Lee et al. 2012

Chu et al. 2012


Table 1 Contd.

Sweet potato

Thermostable -amylase and




S. cerevisiae

96.7% (v/v)

Purohit and Mishra,



Sweet potato

Celluclast , Pectinex,
Viscozyme and Liquozyme






Srichuwong et al.


Corn cob


S. cerevisiae (3090)

24.79% (v/v)



2.5 N sulphuric acid and

Spezymeand Stargen



558 g

Shanavaset al. 2011

60% (v/v)

Nuwamanya et al.


Zhang et al. 2011

Issatchenkia orientalis
IPE 100

0.25g/g dry

Kwon et al. 2011

S. cerevisiae
Z. mobilis

87.8% (v/v)






Nonfood parts of
Sweet potato

Sweet sorghum

Sweet potato
Sweet potato

Potato peel waste
sweet potato

1M HCL and 1M NaOH,

Amyloglucosidases, amylase,
Glucoamylase (1.6AGU/g)
and glucoamylase (1.6




Cellulase, thermostable amylase and glucoamylase


mutant strain of Aspergillus

niger isolated from mildewed
sweet potato used for
-amylase and glucoamylase

Sulphuric acid
Thermostable -amylase,
xylanase, glucoamylase

Wheat meal
Wheat straw

-amylase and
impregnation sulphuric acid


Sweet potato


Industrial purplefleshed sweet


1000 u/g
Liquozyme SC and Spirizyme











S. cerevisiae

S. cerevisiae CCTCC


S. cerevisiae HAU-1
S. cerevisae var.

Salve et al. 2012

Cao et al. 2011


Zhang et al.2011


Rani et al. 2010


S. cerevisiae CCTCC


Arapoglou et al. 2010

Zamora et al. 2010
Zhang et al. 2010

Erdei et al. 2010



S. cerevisiae








Z. mobilis
ATCC 29191
Ethanol Red Yeast

Zhang and Feng,


Bridgers et al. 2010


Table 1 Contd.

Potato mash




Wheat bran flour


Sweet potato


Hybrid sorghum


Waste potato

Thermostable -amylase,
A. niger

A. niger



S. cerevisiae NBR 0224

16.61% (v/v)



Co culture of A.niger
GS4 and S. cerevisiae
Co culture of A.niger and
Saccharomyces sp.







Sweet potato

-amylase and glucoamylase



Sweet sorghum




Thermostable amylase and

-amylase and glucoamylase



Srichuwong et al.
Ado et al. 2009


Manikandan and
Viruthagiri, 2009


Berehoiu et al. 2009



Duvernay, 2008

Immobilized S.
S. cerivisiae and S.

93.24% (v/v)
9.5 g/100g

Liu and Shen, 2008b

Aithal and Kulkarni,
Liimatainen et al.

sugar and produced 7.6 g l-1 of ethanol using Saccharomyces cerevisiae. In order to obtain maximum glucose conversions, the relationship among parameters of
the liquefaction and saccharification process was investigated by a response surface method. The optimum combination of temperature, dose of enzyme (amylase) and amount of waste potato mash were 95C, 1 ml of enzyme (18.8 mg protein/ml) and 4.04 g dry-weight/100 ml water with 68.86% loss in dry weight
for liquefaction. For saccharification, temperature, dose of enzyme and saccharification time were optimized and optimum conditions were determined as 60C72 h-0.8 ml (300 Unit/ml) of amyloglucosidase combination which yielded 34.9 g l -1of glucose. After optimization of hydrolysis of the waste potato mash, ethanol
fermentation was studied. Effects of pH and inoculums size were evaluated to obtain maximum ethanol. Results concluded with 5.5 optimum pH and 3%
inolculum size, respectively for maximum ethanol concentration (30.99 g l-1) (Izmirlioglu and Demirci, 2012). The different type of saccharification and further
fermentation from potato has been summarized in Table 1.
Sweet sorghum
Sweet sorghum (also called Sorgo) is an indigenous African plant which has more tolerance to drought and water logging than corn (Qiu et al., 2010). Besides
that, less chemicals and fertilizer are used in the production of sorghum. Mostly these are planted in saline-alkali soil within temperature 10-40oC. It is also
termed as a crop with high energy content, photosynthesis efficiency, biomass production capacity and high sugar content. Sweet sorghum produces grain,
which is harvested for human consumption and contains sweet juice in the stalk. After harvest, the stalks are squeezed for the sweet juice, which can be turned
into sugar or fermented to ethanol. The stalk material remaining after the sugar juice extraction is called as bagasse and can be used as animal feed (Liu and
Shen, 2008a).
Sweet sorghum has been considered as one of the most promising crops by various researchers for the production of ethanol at very low cost (Barbanti et al.,
2006; Yun-long etal., 2006; Wang and Liu, 2009). Fermentation of sweet sorghum using yeast has an advantage of rapid fermentation (Liu and Shen, 2008b).
Sufficient nutrients like carbon and nitrogen are required for the yeast to grow and reproduce but inorganic salts present in sweet sorghum juice is not enough to


meet the need of fermentation (Mei et al., 2009). Sipos et al. (2010) have shown that yeast extract, ammonium, urea,
calcium and magnesium have effects on both growth and fermentation, thus stimulating fermentation rate and ethanol
production. Asli (2010) studied the fermentation of sweet sorghum juice at different pH levels using different nitrogen
sources and dilution rates and found highest ethanol yield at pH 4.5 using ammonium sulphate as a nitrogen source and
an initial 100 g/l sugar concentration. Kundiyana et al. (2010) studied the fermentation of sweet sorghum juice at
different pH levels using urea as the nitrogen source. The authors concluded that the highest ethanol yield could be
obtained at a pH of 4.3 without the addition of urea and without prior sterilization of the juice. The different type of
saccharification and further fermentation from sorghum has been summarized in Table 1.
Sweet potato
Sweet potato (Ipomoea batatas L.) rank as the fifth most important food crops on a fresh-weight basis in developing
countries, after rice, wheat, corn and cassava. It is one of the most important starch producing crops grown worldwide
which is resistant to drought, pests and diseases, and can grow in poor soils (Zhang and Feng, 2010, Zhang et al.,
2011). The global annual production of sweet potato per year is more than 140 million tonnes. The total annual
production of sweet potato in India was 1046 tonnes in the year 2009-2010 (Indian Horticulture Database 2011). It is a
native crop of the state of Odisha (India) and available in plenty with a total production of 438.80 tonnes (Jata et al.,
2012). On emphasizing the nutritional value of sweet potato, the Center for Science in the Public Interest has compared
it with other vegetables in respect of fiber content, complex carbohydrates, protien, vitamin A, vitamin C, iron, and
calcium, which ranked highest.
The dry matter content in sweet potato ranges from 21 to 30% which consists of 80% of starch. It is a low impact crop
which offers a viable alternative starchy raw material that can be converted to useful sugar feed stocks needed for the
production of ethanol and other value added products. It has been considered a good substrate for alcohol fermentation
since they have a higher starch yield per unit land cultivated than grain, and continue to increase in weight during its
long growing season until harvested. It requires simultaneous saccharification and fermentation for a better ethanol
(Huntrods, 2008; Qiu et al., 2010; Srichuwong et al., 2009; Ziska et al., 2009; Lee et al., 2012; Duvernay et al., 2013).
The different type of saccharification and further fermentation from sweet potato has been summarized in Table 1.
Industrial sweet potatoes are not intended for use as a food crop. They are bred to increase its starchy content,
significantly reducing its attractiveness as a food crop when compared to other conventional food cultivars (visual
aspect, color, taste). Therefore, the fermentable sugar yields from a sweet potato crop increases. Some industrial sweet
potatoes breeding lines developed could produce ethanol yields of 4500-6500 l/ha compared to 2800-3800 l/ha for corn
(Ziska et al. 2009; Duvernay et al., 2013). However, a comparative analysis of bioethanol production from different
starchy crops with other substrates has been depicted in Table 2.
Generally, native starch granules are semi-crystalline, round to oval in shape and have smooth surface. Native starch
generally resists amylase activity. However, they are readily hydrolyzed when get gelatinized (Tester and Karkalas,
2006). Enzymatic hydrolysis of starch have better hydrolysis rate as compared to acid hydrolysis (Purohit and Mishra,
2012). Fresh sweet potato contains high water content. The drying process of this material is an important aspect for
bioethanol processing. Therefore, handling of high viscous material and extra cost of drying effects on the performance
of the process (conversion of starch to fermentable sugars) was carried out by, Moorthy 2002. Some of the studies on
sweet potato revealed that, simultaneous saccharification and fermentation yields 91.4% ethyl alcohol on its starch
content (Zhang et al., 2011), where simultaneous saccharification and fermentation of high gravity slurry of 304 g l1
showed fermentation efficiency of 89.7% (Srichuwong et al., 2009). Similarly, Lareo et al. (2013) followed simultaneous
saccharification and ethanol fermentation using fresh and dry flour of sweet potato and got 38-45 g/l ethanol
concentration in 16h of fermentation time with 100% sugar conversion. A comparative analysis of bioethanol production
from different starchy crops with other substrates has been done in Table 2.
Corn is the most widely grown grain crop throughout the United States. It is a lignocellulosic biomass which can be
classified as the waste produced as a low value byproduct of industrial sectors such as agriculture, and can be utilized
to produce ethanol due to its high sugar content. This plant contains leafy stalk producing ears which enclose the grain,
in the form of seeds and emphasizing on the nutritional value of sweet, yellow and raw corn grown in US. It has
carbohydrate, fat and protein factor of 3.57, 8.37 and 2.44 respectively. Currently rising energy demand and global
warming have encouraged worldwide research and development of biofuels, especially from lignocellulosic materials
which includes sugarcane bagasse, straw, corn etc. No doubt bio-ethanol production from corn is a mature technology
that is not likely to see considerable reduction in the production costs. A comparative analysis of bioethanol production


Table 2. Comparative analysis of different starchy crops with other substrates

involved in the current and future ethanol production
Starchy and
sugary substrates
Sweet potato
Sugar beet
Sweet sorghum
Sugar cane

production (L/ha)



Drapoch, 2008
Lareo et al. 2013
Kymalainen, 2007
Osuji et al. 2010
Cheng, 2010
Osuji et al. 2010
Kraatz, 2008
Rutto et al. 2013
MINO, 2010
Cheng et al. 2011

from different starchy crops with other substrates has been done in Table 2.
Starch can be extracted from corn through the milling process, which can be enzymatically treated for obtaining glucose
syrup, which further fermented to ethanol. There are two types of corn milling process in the industry: wet and dry.
During wet-milling process, corn grain is separated into its components through which starch is converted into ethanol
and the remaining components are sold as co-products. During dry-milling, grains are not fractionated and all their
nutrients enter the process and are concentrated into a distillation co-product utilized for animal feed called Dried
Distillers Grains with solubles (DDGS) (Sanchez and Cardona, 2008). Itelima et al. (2013) stated that significant cost
reductions may be possible if cellulose based agricultural wastes such as corn cobs are used instead of corn, and their
study resulted optimal ethanol yield of 10.08 v/v after 7 days of fermentation at varied pH range i.e. between 3.05 and
7.58 using co culture of Aspergilus niger and Saccharomyces cerevisiae. According to Perlack et al. (2005), corn stover
has the greatest potential to be used for cellulosic ethanol production with a projected availability of 170~256 million tons
annually in the United States.
Wheat is a cereal grain worldwide cultivated plant belonging to family poaceae. It is the first known domesticated cereal
which has the ability of self pollination and are generally cultivated on land area. In the present era, it secures the third
position as a cereal crop product after maize and rice. Whereas, it is the second main food crop for human feed and also
used as animal feed. Along with that it was one of the first food crops which could be easily cultivated on large scale and
had the additional advantage of yielding a harvest that could provide long term storage of food. Instead of being the
leading source of vegetable protein globally and having higher protein content than other cereal crops such as maize
and rice, it is preferred as a suitable substrate for production of ethanol because the whole grain contains concentrated
amount of vitamins, minerals and proteins while the refined grain contains mostly starch.
According to Manikandan and Viruthagiri (2009), wheat bran flour yields 23.1 g l-1ethanol by SSF process using co
culture of A. niger and K. marxianus when it is fermented for 48h at pH 5.5 and 30C. Besides that, the study by Ismail
et al. (2012) reveals that when wheat straw is pretreated with perchloric acid and fermented at temperature 30C, pH 5.5
for 42h with co culture of S. cerevisiae and P. stipitis, it resulted in 11.42 g l-1 of ethanol. Erdei et al. (2010) took both
wheat meal and wheat straw for ethanol production. They liquefied and saccharified wheat meal with the help of amylase, amyloglucosidase and impregnated wheat straw with sulphuric acid. They fermented by using S. cerevisiae
and resulted 56.5 g l-1 of ethanol when incubated for 72 and 96h at 36.5C and pH 4.8, respectively. Further,Erdei et al.
(2013) resulted 60 g l-1 of ethanol with wheat straw and wheat meal using S. cerevisiae. In this experiment, the wheat
meal was pretreated with -amylase, amyloglucosidase and cellulases and then incubated for 48h at 32C having pH 5
along with that wheat straw was pretreated with sulphuric acid and cellulose and incubated for 120h at 35C and pH 5.
Yam (Dioscorea rotundata) is a starchy staple foodstuff, belongs to Dioscoreaceae family, normally eaten as a
vegetable, boiled, baked or fried which is found in different countries like Africa, America, Caribbean, South Africa and


Asia. Several edible yams such as bitter yam (D. dumetorum), water yam (D. alata), white yam (D. rotundata) and yellow
yam (D. cayenensis) are widely grown throughout the tropics. In West Africa they are consumed mainly as "fufu", stiff
glutinous dough. The starch present in the starchy tuber can be hydrolyzed, fermented and then distilled to produce
ethanol (Thatoi et al., 2014). Maldakar and Maldakar (2000) used pulp of Diascorea sativa through gelatinization at 90
C with alpha amylase, saccharification with a-amyloglucosidase, and then fermentation using yeast for the production of
ethanol. Similarly Yan et al. (2007) produce ethanol from Diascorea after gelatinization and liquefaction at 90-93 oC and
further saccharification using -amylase.
Sugar beet
Ethanol production from sugar crops such as sugarcane and sugar beet account for about 40% of the total bioethanol
produced and nearly 60% corresponding to starch crops. The high yield of sugar per acre, lower cycle of crop
production, high tolerance of a wide range of climatic variations, low fertilizer requirement and low conversion costs are
the major advantages of these crops. The main disadvantage of using these crops is their natural seasonal availability.
Sugar beets can grow in temperate climate with a lower rain fall than is needed for sugar cane and generate good yields
(25-50 tons per acre). The major sources of sugars containing sugar beet are found mainly in Europe, France and North
which has been exploited for biofuel production. On average, the ethanol yield is 25 gallons per ton of sugar beets;
however, ethanol production from sugar beet requires a greater chemical and energy input and consequently is more
expensive than the process using sugar cane (Wagner, 2005; Mussatto et al., 2010; Vohra et al., 2013).
Sugar beets are the underground root crops which belongs to the Amaranthaceae family whose root contains a
significant quantity of sucrose. A sugar beet weighs two to five pounds and produces about three teaspoons of sugar
after fully grown. Theoretically a ton of sugar beet will give 110 L ethanol which is comparable to a ton of barley giving
340 l of ethanol. A comparative analysis of bioethanol production from different starchy crops with other substrates has
been done in Table 2. Beet sugar can boost ethanol production by adding crystallized sugar to corn slurry to speed up
the fermentation process (Thatoi et al., 2014). Ethanol production was studied by Grahovac et al. (2012) in batch culture
using free cells of S. cerevisiae from intermediates of sugar beet processing. The ethanol production based on the
intermediates of sugar beet was amounted to 490 g/kg. Bioethanol has also been produced from sugar beet molasses
by calcium (Ca)-alginate immobilized Saccharomyces cerevisiae yeast in the presence of castor oil without heat or filter
sterilization. The bioethanol production process can also act on the debris from sugar beet increasing the conversion by
an additional 30% by using Bacillus strearothermophilus bacterium that digests hemicelluloses, a sugar that yeast is
unable to break down.
Aroids comprise several plant species under the family Araceae that are cultivated for food in most of the tropical and
subtropical parts of the world. These are mostly herbaceous plants often within large root stocks that act as storage
organs, which are mostly consumed (Lin and Tanaka, 2006). Among various edible aroids, commercially cultivated
promising tuber crops are Colocasia, Xanthosoma, Alocasia and Amorphophallus which can serve as good substrates
for fermentation (Ray and Ward, 2006; Thatoi et al., 2014). Edible aroids that are grown commercially in the U.S. are
taro (Colocasia esculenta), tannia (Xanthosoma sagittifolium) and giant taro (Alocasia spp.). Popular aroids cultivated in
India are taro (Colocasia esculenta (L.) Schott.), tannia (Xanthosoma sagittifolium (L.) Schott.) and elephant foot yam
(Amorphophallus paeoniifolius (Dennst.) Nicolson) (Sunitha et al., 2013). The yield of taro, elephant foot yam and tannia
is 5-10, 20 and 25 t/ha with 10-20% varies in starch contents, respectively.
Pretreatment of starchy feedstocks
Pretreatment basically refers to the physical or thermal actions which can be applied to several starchy feedstocks in
order to improve hydrolysis efficiency by exposing the granules to the enzymes. The pre-treatment step changes the
native properties of the substrate through the destruction of the cell structure in order to prepare the materials for
enzymatic degradation. The mechanical milling grounds the dried tubers to a certain particle size that can pass through
a mesh screen and then the flour can be collected and used for further processing i.e. hydrolysis of the substrate (Thatoi
et al., 2014; Cinelli et al., 2015). When starch is heated in water, it forms a very thick gel, which is known as
gelatinisation. Starch gelatinisation is essential for efficient enzymatic hydrolysis (Takashi et al., 2010). The
gelatinisation is followed by hydrolysis of starch.
Lamsal et al. (2011) applied two physical pretreatment methods like roller mill flaking and hammer mill grinding before
starch hydrolysis for the production of ethanol from corn using a commercial S. cerevisiae strain. In this study, ethanol
productivity was higher in case of hammer mill pretreatment rather than roller mill flaking with the production of similar


ethanol concentrations (18-19% v/v). Shariffa et al. (2009) showed effect of preheating of cassava and sweet potato
starch granules at 60 oC for 30 min which resulted in a 14% increase in the degree of hydrolysis. Similarly Li et al. (2012)
reported the importance of preheating stage in corn below the gelatinization temperature which increases about 80% of
hydrolysis. However, Uthumporn et al. (2013) followed another defatting step before hydrolysis which increases 3-fold of
corn starch hydrolysis. Sapinska et al. (2014) applied several enzymes like xylanases, cellulases, cellobiases and
pullulanases for the hydrolysis of rye and corn mashes after the pretreatment of the raw materials in a process defined
as pressure-less starch liberation method, aiming at beverage applications.
Necessity for saccharification
The development process for simultaneous liquefaction, saccharification and fermentation of starch would reduce the
energy input and increase the efficiency of substrate utilization (Koutinas et al., 2007; Chu et al., 2012). Starchy
substrates employed as raw materials for ethanol production which cannot be directly fermented to ethanol. This is
because yeast like S. cerevisiae cannot utilize the starch molecules which are made up of long chains of glucose
molecules hence prior to fermentation, are needed to hydrolyze it to simpler fermentable sugars by breaking starch
molecules into simple glucose molecules (Altintas et al., 2003). Enzymes and acids can be employed for hydrolysis, by
following a reaction of starch with water (hydrolysis) to break down the starch into fermentable sugars (saccharification)
which include enzymes such as amylases, amyloglucosidases, glucoamylases, pullulanase etc. from either fungal or
bacterial origin and hydrochloric acid or sulphuric acid as an acid source for hydrolysis. Typically, hydrolysis is
performed by mixing the starch with water to form slurry which is then stirred and heated to rupture the cell walls.
Specific enzymes and acid that will break the chemical bonds are added at various times during the heating cycle to
dissociate glucose molecules. Mostly, -amylase and glucoamylase are preferred for the process of hydrolysis in which
-amylase is an endo-amylase attacking -1, 4 bonds in random fashion which rapidly reduce molecular size of starch
and consequently its viscosity. Glucoamylase is an exo-amylase which can hydrolyze the -1, 4 glycosidic linkage of the
nonreducing terminal glucose unit. It can also hydrolyze the -1-6 linkage from the amylopectin (Ray and Swain, 2011).
Microorganisms generally prefer gelatinized starch. But large quantity of energy is required for gelatinization. So it will
be better to use organisms growing well on raw (ungelatinised) starch which on hydrolysis would further act as a suitable
media for microorganisms to carry out the fermentation process (Prasad et al., 2007). Rattanachomsri et al. (2009)
showed efficient saccharification of non-gelatinized cassava pulp by the combined action of several accessory enzymes,
using commercial cellulases (Celluclast 1.5l), b-glucosidase (Novozym 188), pectinases (Pec-tinex Ultra SP-l) and bglucanases (Optimash BG), in association with glucoamylase (AMG 300 l) and a-amylase (Termamyl 120 l). Several
recent studies have reported the effectiveness of simultaneous starch saccharification and fermentation (SSSF)
processes for increasing ethanol productivity from starch (Takashi et al., 2010; Balcerek and Pielech-Przybylska, 2013;
Szymanowska-Powaowska et al., 2014).Zhang et al. (2011) applied simultaneous saccharification and fermentation
(SSF) from viscosity reducing of raw sweet potato for bioethanol production using Saccharomyces cerevisiae at
laboratory, pilot and industrial scales.In this study, The maximum ethanol concentration, average ethanol productivity
rate and yield of ethanol after fermentation in laboratory scale (128.51 g/l, 4.76 g/l/h and 91.4%) were satisfactory with
small decrease at pilot scale (109.06 g/l, 4.89 g/l/h and 91.24%) and industrial scale (97.94 g/l, 4.19 g/l/h and 91.27%)
In recent years, immobilization of enzymes or microbial cells or co-immobilization of both for the simultaneous starch
saccharification and fermentation (SSSF) process has attained scientific and technical interest for alcoholic
fermentation. Immobilization can be carried out in different ways; adsorption and entrapment in matrices are the
methods most commonly used (Yu et al. 1996). Among the various immobilized methods tested, a co-immobilized
system using microorganisms co-immobilized in different matrix appeared most promising with respect to ethanol
productivity from various starchy feed stocks (Nellaiah and Gunasekaran, 1992; Lee et al., 2012). Also, most of these
studies utilize commercial enzymes which may lead to the higher cost of production. However, the starch content can be
saccharified by acid-enzyme or enzyme-enzyme hydrolysis to obtain lower molecular weight carbohydrates and finally to
monomeric sugars (Raman and Pothiraj, 2008; Behera et al.,2015). The resulting hydrolysate can be used as substrate
in fermentation industries. The different type of saccharification for different feed stocks has been summarized in the
Table 1.
Exploration of fermentation
Fermentation is a metabolic process of microorganisms to obtain energy by breaking down organic compounds. It is the


third step which leads to the production of ethanol from saccharified biomasses. In this process, the large organic
molecules is slowly decomposed (such as starch) into smaller molecules such as ethanol. Ethanol fermentation can be
described as the biochemical process by which sugar such as glucose; fructose and sucrose are converted into cellular
energy there by producing ethanol and carbon dioxide as metabolic waste products (Shanavas et al., 2011).
Fermentation typically requires 12 to 72h of incubation period depending on the amount and type of microorganism used
to start fermentation. In general, root and tuber crops are preferred for the bioethanol production due to its high starch
content (16-24%) (Hashem and Darwish, 2011).
Fermentation of starchy materials is a process which generally involves several steps. After saccharification the slurry
containing fermentable sugars needs to be fermented for production of ethanol and further separation and purification of
the ethanol, usually by distillation. Fermentation can be followed by 2 methods such as (1) Solid state fermentation
(SSF) and (2) Submerged fermentation (SmF).
Solid state fermentation
It refers to the process where microbial growth and fermentation occurs on the surface of the solid materials. This
process occurs in the absence of free water, where the moisture is absorbed to the solid matrix (Pandey et al., 2000).
The moist solid materials which are polymeric in nature and insoluble in water act as a carbon source as well as
providing anchorage for the microorganisms. Solid state fermentation has a series of advantages over submerged
fermentation including lower cost, improved product characteristics, higher product yield, easiest product recovery and
reduced energy requirement (Ray et al., 2008). The main process for the solid support in SSF is that use natural solid
substrate like starch, flour or (ligno) cellulose residue or agroindustrial sources such as cassava, potato, sweet potato
and pulp/ residues, etc. In these cases substrates are also used as the source of carbon and nutrients for the microbial
growth (Tengerdy and Szakaca, 2003; Ray et al., 2008). A consolidate solid state fermentation system was applied to
corn and cassava starch by Moukamnerd et al. (2010) for the continuous recovery of alcohol. However, the processing
cost of starch might reflect on the overall production cost of ethanol from cassava.
Approximately 90% of all industrial enzymes are produced in submerged fermentation, frequently using specifically
optimized and genetically manipulated microorganisms. However, SF holds tremendous potential for the production of
enzymes. Solid state fermentation (SSF) constitutes an interesting alternative since the metabolites so produced are
concentrated and purification process costs less over submerged fermentation (Nigam and Singh, 1995, Pandey et al.,
2000). The aim of SSF is to bring the cultivated microorganisms into tight contact with the insoluble substrate and thus
to achieve the highest substrate concentration during fermentation.Much published information is available on the
production of enzymes of industrial importance, such as protease, cellulase, ligninases, xylanase, pectinase, amylase,
glucoamylase, etc. Several enzymes such as -amylase, glucoamylase and pullulanase used for starch processing have
been produced through SSF process using various microbial systems (Murthy et al., 2009; Parbat and Singhal, 2011;
Saxena and Singh, 2011; Sugumaran et al., 2014). The enzyme -amylase can be obtained from Aspergillus oryzae,
Bacillus amyloliquefaciens and Bacillus licheniformis. The major end products from the action of a-amylase on starch
are glucose, maltose, maltotriose, maltotetrose, maltopentaose and maltohexose. Glucoamylases are used in glucose
syrup production from liquefied starch, and are usually obtained from Aspergillus niger and Rhizopus species.
Pullulanase are used in debranching starch in sugar syrup manufacture, and are prepared from Klebsialla pneumonia
and Bacillus acidopullulyticus (Nigam and Singh, 1995). Swain and Ray (2007) carried out to investigate the -amylase
production by B. subtilis strain CM3 isolated from cowdung microflora in SSF using CFR as the substrate and
optimization of the fermentation parameters (incubation period, medium initial pH, moisture holding capacity (MHC) and
temperature) by applying RSM.The experimental results showed that the optimum incubation period, initial medium pH,
moisture holding capacity and temperature were 6 days, 8.0, 70% and 50 C, respectively.

Submerged fermentation
It is the process in which the growth and anaerobic or partially anaerobic decomposition of the carbohydrates by microorganisms in liquid medium occur with plenty availability of free water (Ray and Ward, 2006). Fermented industrial
products like acetone-butanol, ethanol, lactic acid, etc. are the products of SmF. It is the process of choice for industrial
operation due to the very well-known engineering aspects, such as fermentation modeling, bioreactor design and
process control (Gutierrez-Correa and Villena, 2003), etc. The submerged fermentation processes for amylase
production have become economically feasible, and A. niger growing in starch-salt medium has been used for largescale production of enzyme (Nigam and Singh, 1995).vRattanachomsri et al. (2009) investigated the action of their own
enzyme preparation produced by submerged fermentation of Aspergillus niger on 40 g/l of cassava pulp, obtaining up to


91% of hydrolysis efficiency. The different type of saccharification and further fermentation process though submerged
method for different feed stocks has been summarized in the Table 1.
Product Recovery
As biomass hydrolysis and fermentation technologies approach commercial viability, advancements in product recovery
technologies will be required. Bioethanol obtained from a fermentation conversion requires further separation and
purification of ethanol from water through a distillation process. Distillation technologies that will allow the economic
recovery of dilute volatile products from streams containing a variety of impurities, have been developed and
commercially demonstrated (Balat, 2011). Fractional distillation is a process implemented to separate ethanol from
water, based on their different volatilities. The first step is to recover the bioethanol in a distillation column, where most
of the water remains with the solid part. For purification of ethanol distillation is the most dominant and recognized
industrial technique. It utilizes the differences of volatilities of components in a mixture. The basic principle of distillation
is heating a mixture, where low boiling point components are concentrated in the vapor phase and by condensing this
vapor, more concentrated less volatile compounds such as ethanol is obtained in liquid phase (Karuppiah et al., 2008;
Bhalla et al., 2013).

Ethanologenic microorganisms
Ethanol is produced by fermentation of sugar by microorganisms. For efficient ethanol production there is requirement of
efficient microorganisms that are able to ferment a variety of sugars (pentoses and hexoses) while at the same time be
able to tolerate the alcohol and sugar stress conditions that accompany fermentations. Generally microorganisms
participate in two major applications during the process of fermentation, that is, microorganisms convert fermentable
substrates into ethanol and in the other way, microorganisms produce enzymes to catalyse conversion of complex
carbohydrates into simpler sugars (Thatoi et al., 2014). Several microorganisms, Zymomonas mobilis, Saccharomyces
cerevisiae, Saccharomyces uvarum, Candida tropicalis, Candida shehatae, Pichia stipitis, Clostridium sp., have been
considered as ethanologenic microbes. The yeast S. cerevisiae and facultative bacterium Z. mobilis are very promising
candidates for industrial alcohol production. The best xylulose fermenting yeasts so far identified are species of
Brettanomyces sp., Candida, Hansenula and Torulospora. Pentose utilising yeasts also convert xylose into xylitol in
addition to ethanol. However, important yeast, P. stipites apparently produces no xylitol during sugar fermentation
(Behera et al., 2014).
Fermentation is a biological process, in which ethanologenic microorganisms convert organic material in to simpler
compounds, such as sugars. The microorganisms then ferment the fermentable compounds to produce ethanol and
CO2. During the whole process of ethanol fermentation, there are requirement of mainly two types of microorganisms.
One is to produce the enzyme by microorganisms or supplied manually to catalyze chemical reactions that hydrolyze the
complicated substrates into simpler compounds and the other is to convert fermentable substrates into ethanol by the
microorganisms. Among many microorganisms being exploited for ethanol production, the genus of
Saccharomyces cerevisiae still remains as the prime species. Zymomonas mobilis is another one of the most intensively
investigated species within the past three decades because it possesses some characteristics compared to its
counterpart S. cerevisiae.

Saccharomyces cerevisiae
Traditionally, the yeast, Saccharomyces cerevisiae has been used all over the world as the major ethanol producing
microorganism. The sugars that are metabolizable by this organism include glucose, fructose, mannose, galactose,
sucrose, maltose, and maltotriose. Ethanol production by S. cerevisiae is carried out via the glycolytic pathway (also
known as the Embden-Myerhof-Parnas or EMP pathway) (Thomas et al., 1996). However, under anaerobic ethanol
fermentations, yeast cells suffer from various stresses. Some of them are environmental, while the others are generated
by the yeast cells themselves, for example ethanol accumulation, and correspondingly strong inhibition on yeast cell
growth and ethanol fermentation. The yeast cells in ethanol fermentation are under harsh condition when compared with
their aerobic cultures in which the process parameters are optimized with optimum pH, sufficient nutrients and very low
ethanol that does not exert inhibition. Their metabolic activities are seriously affected, especially under industrial


conditions in which the optimum conditions for growth and fermentation are overwhelmed by pursuing economic
Zymomonas mobilis
It is an anaerobic and gram-negative bacterium producing ethanol from glucose via the Entner-Doudoriff (ED) pathway
(Conway 1992) in conjunction with the enzymes PDC and ADH. Z. mobilis catabolizes only three sugars, D-glucose, Dfructose and sucrose, as its sole carbon and energy sources. Its growth on sucrose is accompanied by the extracellular
formation of the fructose oligomers (levan) and sorbitol with a significant reduction in its yield for ethanol, which makes it
unsuitable for production of the alcohol using molasses. Meanwhile, it can only ferment glucose in the hydrolysate of
starch materials but cannot utilize other sugars like the species of S. cerevisiae, making it unsuitable for ethanol
production using starch materials.
Zymomonas is the only microorganism that metabolizes glucose anaerobically using the ED pathway as opposed to
the EM or glycolytic pathway. The ED pathway yields only half as much ATP per mole of glucose as the EM pathway. As
a consequence, Zymomonas produces less biomass than yeast, and more carbon is funneled to fermentation products.
Also, as a consequence of the low ATP yield, Zymomonas maintains a high glucose flux through the ED pathway. All the
enzymes involved in fermentation are expressed constitutively, and fermentation enzymes comprise as much as 50% of
the cells total protein. Despite its advantages as an ethanologen, Z. mobilis is not well suited for all of the biomass
resources conversion because it ferments only glucose, fructose, and sucrose. Moreover, Z. mobilis on synthetic media
containing glucose, fructose or sucrose, the specific rates of sugar uptake and ethanol production are at a maximum
when utilizing the glucose medium.
Escherichia coli
Escherichia coli is another valuable bacterial resource for ethanol production. One of the first successful applications
of metabolic engineering was the construction of E. coli strains to selectively produce ethanol. It has several
advantages as a biocatalyst for ethanol production, including the ability to ferment a wide spectrum of sugars, in addition
with no requirements for complex growth factors, and prior industrial use (e.g., for production of recombinant protein).
The major disadvantages associated with E. coli cultures, are a narrow and neutral pH growth range (6.0-8.0), less
hardy cultures compared to yeast, and public perceptions.
Besides having several benefits, the production and utilization of bioethanol using starchy feedstocks also have several
challenges. Supplying continuously the demand of starchy feedstocks for subsistence and commercialization oriented to
bioethanol and food related industries depend on the sustainable production system. To sustain crop and tuber
production system, severe biotic and abiotic stresses have to be alleviated. The challenges in using sweet sorghum as a
feedstock includes the breeding of new sorghum varieties, planting techniques, harvesting time (especially during the
rainy season), crushing and fermentation, as well as reduction of compounds that affect industrial processes such as
aconitic acid, phenolic compounds, and starch (Amorim et al., 2011). The use of biotechnology can potentially address
many of these technical challenges and environmental concerns. In recent years, there is a significant progress in the
use of biotechnological techniques such as molecular biology, protein engineering, genetic engineering to increase the
activity of enzymes and the microbes for enhanced biofuel production (Chandel et al., 2011). This progress will certainly
help us to make bioethanol a replacement for our present transportation fuel.
Several biotechnological tools such as recombinant DNA, metabolic engineering and mutagenesis are available for
the improvement of yeast strains capable of withstanding the harsh environments typically of certain industrial
fermentation process. The advances in metabolic pathway engineering and genetic engineering techniques have led to
the development of micro-organisms capable of efficiently converting biomass sugars into ethanol (Neves et al., 2006).
Strict anaerobic hemophilic bacteria such as Clostridium sp. and Thermoanaerobacter sp. have been proposed to
explore the benefits of fermentation at elevated temperatures (Sanchez and Cardona, 2008). In a recent study, E. coli
cells for efficient ethanol production from hexoses and pentoses were developed using elementary mode analysis to
dissect the metabolic network into its basic building blocks (Trinh et al., 2008). More recently an attempt has been made
to engineer E. coli for the production of ethanol from fatty acid feed stocks, resulting in ethanol yield higher than the
theoretical maximum obtained from sugars (Clomburg and Gonzalez, 2010).The technological improvement could help


to improve the system efficiency and provide value added co-products, which will reduce the production cost. Further,
some of the technological challenges need further research work.
Conclusions and future prospective
The utilization of bioethanol for transportation has the potential to contribute to a cleaner environment. With the
increasing demands for energy and shrinking energy resources, the utilization of starchy biomass for the production of
biofuel offers a renewable alternative. Recently, bioethanol production from tuber crops also seems to be a promising
aspect because of their abundant availability, high starch content and cost effective processing, making it one of the
desirable substrate among all the other substrates such as lignocelluloses and algal biomass, which have high
processing and maintenance cost. The development of bioethanol production from different starchy feedstocks has been
important not only to reduce the fossil fuels dependency but also to lower the contribution of petroleum derivatives to
climate changes and air pollution. Apart from biofuels, other value-added products such as fermentable sugars, organic
acids, solvents and drink softeners etc. may also be produced from starchy biomass using appropriate technologies.
Theoretically this is quite possible; however, technologically it is not an easy task because of various technological gaps.
Genetic engineering approaches should be more focused on developing new improved strains with higher substrate
tolerance and improved production kinetics. One of the major bottlenecks in the industrial application of enzymes and
metabolites from thermophiles is the low productivity that is typical of the fermentation processes for thermophiles. To
enhance productivity, physiology of these unique microorganisms must be searched. Key approaches can be
bioprocesses and media design, implementation of innovative bioreactors and overproduction in mesophilic host at
molecular level.
We are thankful to University Grant Commission, New Delhi [(Letter F. No. 41-478/2012 (SR), dated 16.07.2012], for
financial support to carry out this work.
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