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Optimization of Cellulase Enzyme From Vegetable Waste by Using Trichoderma Atroviride in Solid State Fermentation
Optimization of Cellulase Enzyme From Vegetable Waste by Using Trichoderma Atroviride in Solid State Fermentation
e-ISSN: 2319-2402,p- ISSN: 2319-2399.Volume 10, Issue 5 Ver. II (May. 2016), PP 68-73
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Abstract: The present study deals with production of cellulase enzymes from filamentous soil fungus
Trichoderma atroviride by using vegetable waste as substrate through the process of solid state fermentation.
The enzyme production was further improved by optimizing a number of physical parameters such as incubation
time, temperature, pH, inoculums size and nutritional factors (carbon source, nitrogen source and detergents).
By optimization of different parameters, the maximum activities of cellulase synthesized by Trichoderma
atroviride were observed after 5 days incubation at H 6 and 30C temperature with sucrose, yeast extract and
Tween -80 as carbon, nitrogen and detergents supplements respectively. The high activity of cellulase produced
by the fungus suggests its potential for commercial scale production for various industrial applications.
Keywords: Cellulase, optimization, solid state fermentation, Trichoderma atroviride, vegetable waste.
I.
Introduction
The need for utilizing renewable resources to meet the future demand for fuel has increased the
attention on cellulose. Cellulose is the structural component of the primary cell wall of plant biomass. It is a
polymer of -1, 4 linked glucose units. It is considered as one of the most abundant renewable carbon source on
earth and the dominating waste material from agriculture [1]. It represents about 1.5 x 1012 tons of the annual
biomass production through photosynthesis and is considered to be an almost inexhaustible source of raw
material. Its crystalline structure and insoluble nature represents a big challenge for enzymatic hydrolysis.
Cellulose is generally degraded by multi-complex enzyme called cellulases [2]. Cellulase (E.C 3.2.1.4) refers to
a class of enzyme that catalyzes the hydrolysis of 1, 4 -D glycosidic linkages in cellulose component [3].
Cellulase enzymes play an important role in natural biodegradation processes in which plant materials
are effectively degraded by cellulolytic fungi, bacteria, actinomycetes and protozoa. In industry, these enzymes
have its importance due to major role in the production of fermentable sugars and ethanol, organic acids and
chemicals. The major other applications are biostoning of jeans and biopolishing of cotton and other cellulosic
fabrics, paper recycling and as animal feed additives for improving the nutritional quality and digestibility [4]. It
is also used for deinking of paper industries and to enhance pulp drainage in textile industries. It is used for
bioremediation, waste water treatment and also for single cell protein [5]. It has importance in food sciences like
food processing, drying of beans in coffee. Main function of cellulase enzyme in food industry is extraction,
clarification and stabilization of fruit juices and vegetables.
Bioconversion of cellulosic materials mainly depends on the nature of cellulose component, sources of
cellulase enzyme and optimal conditions for production of enzymes. Cellulase enzymes are synthesized by a
number of microorganisms. Fungi and bacteria are the main natural agents of cellulose degradation [6].
However, fungi are well known agents of decomposition of organic matter due to their elongated hyphea that
creates mechanical pressure on the cellulose structure, causing them to production of large amounts of cellulase
enzymes [7]. Cellulase production from fungi is highly useful for the enzyme production as compared to other
microorganisms. Fungi such as Trichoderma sp., Aspergillus sp. and Penicillium sp. are the most commonly
cellulase producers. However, the most extensively studied cellulases are produced by efficient lignocellulose
degrading fungi, particularly Trichoderma sp. [8].
Improvement of microbial strains for the over-production of industrial products can reduce the process
cost and may also possess some specialized desirable characteristics. Several lignocellulosic materials are
efficient substrates for white-rot fungi, which produce industrially important cellulolytic enzymes. Among
processes used for enzyme production, solid state fermentation (SSF) using agro-wastes are an attractive and
cost effective option because it presents higher productivity involving a simpler operation [9]. Solid state
fermentation (SSF) is gaining interest as a cost effective technology for the production of higher yields of
cellulase as compared to liquid culture. SSF has many advantages over other fermentation processes due to
simple process, energy saving, less water consumption and less production of waste products. Production of
cellulases by the fungal isolates requires optimal conditions for their growth which leads to the release of
extracellular enzymes. The growth conditions as well as extracellular enzyme production conditions is likely to
vary among isolates. Certain fermentation parameters such as temperature, incubation period, carbon source, pH
etc. were found to be critically affecting the cellulase yield [10].
DOI: 10.9790/2402-1005026873
www.iosrjournals.org
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Optimization of Cellulase Enzyme From Vegetable Waste By Using Trichoderma Atroviride in Solid..
Therefore, the present study aims to enhance the production of extracellular cellulases by filamentous fungi
Trichoderma atroviride under solid state fermentation using vegetable waste as substrate and optimizing cultural
parameters favoring the maximal exploration of fungal capacity for overproducing of cellulase.
II.
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Optimization of Cellulase Enzyme From Vegetable Waste By Using Trichoderma Atroviride in Solid..
2.5.7 Statistical Analysis
Data presented on the average of three replicates (SE) obtained from their independent experiments.
III.
CMCase
FPase
100
80
60
40
20
0
3
CMCase
FPase
100
80
60
40
20
0
25
30
35
40
DOI: 10.9790/2402-1005026873
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Optimization of Cellulase Enzyme From Vegetable Waste By Using Trichoderma Atroviride in Solid..
3.3.3 Effect of pH
Among physical parameters, pH of the growth medium plays an important role by inducing
morphological changes in microbes and in enzyme secretion [17]. The pH change observed during the growth of
microbes also affects product stability in the medium. Trichoderma atroviride was allowed to grow in media of
different pH ranging from 3.0 to 8.0. Maximum enzyme production was observed in medium of pH 6 (Fig 3).
Trichoderma atroviride showed maximum CMCase (88.91 U/g) and FPase (50.96 U/g) production at pH 6
during fifth day of incubation at 30C. Reduction in enzymes activities after 6.0 pH may be ascribed to the fact
that change in pH may change the three dimensional structure of the enzymes. These results are in agreement
with the observations of Baig et al. [18] wherein Trichoderma lignorum favoured a pH of 6.0 as optimum for
maximum CMCase production from using banana waste. Our results are also similar with Jaradat et al. [19] who
reported that the cellulase enzyme from the active isolate J2 was active over a pH range of 47 with maximum
activity at pH 6.
CMCase
FPase
100
80
60
40
20
0
3
pH
CMCase
FPase
140
120
100
80
60
40
20
0
Control Sucrose Dextrose Starch
Lactose
Carbon Source (1 %)
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Optimization of Cellulase Enzyme From Vegetable Waste By Using Trichoderma Atroviride in Solid..
are necessary for the metabolism of fungi. The results of present study regarding best nitrogen source were
found to be similar with the results of the previous studies where yeast extract was the best nitrogen source for
the production of cellulase by Trichoderma.
CMCase
FPase
180
160
140
120
100
80
60
40
20
0
Control Yeast Peptone Beef Sodium
Extract
Extract Nitrate
Nitrogen Source (1%)
CMCase
FPase
180
160
140
120
100
80
60
40
20
0
Detergents (1%)
IV.
Conclusions
The cost-effective technologies are needed for economical production of cellulases using vegetable
waste as substrate. Cellulase yields appear to depend on a complex relationship involving a variety of factors
like incubation time, pH value, temperature, presence of carbon, nitrogen and detergent sources. Major
parameters affecting the fermentation process for enzyme production were studied and optimal levels were
identified. Presently our studies investigated that the fungi belonging to Trichoderma atroviride effectively
produced cellulase under laboratory conditions and can be used for various industrial applications.
DOI: 10.9790/2402-1005026873
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Optimization of Cellulase Enzyme From Vegetable Waste By Using Trichoderma Atroviride in Solid..
Acknowledgement
The author gratefully acknowledges support by getting Senior Research Fellowship from the
University Grant Commission (UGC) and lab facilities from Deptt. of Environmental Sciences, M.D.University,
Rohtak.
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DOI: 10.9790/2402-1005026873
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