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Production of Xylanase by the White-Rot Fungus Ganoderma
Lucidum in Submerged Fermentation Using Wildly Growing Non-
Food Plant Biomass as Substrate
Boddireddy Sridevi
*
Department of Microbiology, Telangana Social Welfare Residential Degree College for Women,
Warangal East, Warangal – 506001, Telangana, India
*
Corresponding Author
DOI:
https://doi.org/10.51583/IJLTEMAS.2026.150300030
Received: 18 March 2026; Accepted: 23 March 2026; Published: 04 April 2026
ABSTRACT
Xylanases are important hydrolytic enzymes responsible for the degradation of xylan, the major hemicellulosic
component of lignocellulosic biomass. These enzymes play an essential role in various industrial processes,
including pulp and paper bleaching, food processing, textile manufacturing, animal feed formulation, and biofuel
production. However, the high cost of enzyme production has limited their widespread industrial application.
One promising approach to reducing production costs is the utilization of inexpensive lignocellulosic biomass
as a fermentation substrate. The present investigation focuses on the production of extracellular xylanase by the
white-rot fungus Ganoderma lucidum using hydrolysates derived from widely growing non-food plant biomass.
Six terrestrial and aquatic weeds—Commelina benghalensis, Cynodon dactylon, Eichhornia crassipes,
Parthenium hysterophorus, Pistia stratiotes, and Setaria viridis—were evaluated as potential substrates for
xylanase production under submerged fermentation conditions. Plant biomass samples were subjected to
autohydrolysis to release hemicellulose-rich soluble compounds that could serve as carbon sources for fungal
growth and enzyme production. Fermentation experiments were conducted at 25 ± 2 °C and pH 5.5 under static
conditions for 7–14 days. Among the substrates tested, hydrolysate derived from Setaria viridis resulted in the
highest xylanase production, followed by Cynodon dactylon and Parthenium hysterophorus. Moderate enzyme
activity was observed with aquatic plant substrates such as Eichhornia crassipes and Pistia stratiotes, whereas
Commelina benghalensis supported minimal enzyme production. The results demonstrate that non-food plant
biomass can serve as an economical and sustainable substrate for xylanase production. Utilization of such
biomass not only reduces production costs but also contributes to effective management of invasive plant species
and lignocellulosic waste. This study highlights the potential of Ganoderma lucidum as a promising organism
for cost-effective industrial enzyme production.
Keywords: Xylanase, Ganoderma lucidum, plant biomass hydrolysate, lignocellulose, submerged fermentation,
white-rot fungi
INTRODUCTION
Lignocellulosic biomass represents the most abundant renewable organic resource available on Earth. It is
primarily composed of cellulose, hemicellulose, and lignin, which together form the structural framework of
plant cell walls. Among these components, hemicellulose accounts for approximately 20–35 % of plant biomass
and consists mainly of complex heteropolymers such as xylan, arabinoxylan, and glucomannan. Efficient
degradation of hemicellulose is essential for the conversion of lignocellulosic materials into valuable products.
Xylanases (EC 3.2.1.x) are glycoside hydrolases that catalyze the hydrolysis of β-1,4-xylosidic linkages in xylan,
leading to the release of xylo-oligosaccharides and xylose. These enzymes play a crucial role in the
bioconversion of plant biomass and have attracted considerable attention due to their broad industrial
applications. In the pulp and paper industry, xylanases are widely used in environmentally friendly biobleaching
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processes that reduce chlorine consumption. In the food industry, they improve dough handling properties and
bread quality. Additionally, xylanases are used in animal feed, textile processing, fruit juice clarification, and
bioethanol production. Microorganisms represent the primary source of industrial enzymes due to their rapid
growth rates and ability to produce large quantities of extracellular enzymes. Various bacteria, actinomycetes,
and filamentous fungi are known to produce xylanases. Among these microorganisms, filamentous fungi are
considered the most efficient producers due to their high secretion capacity and ability to grow on complex
lignocellulosic substrates.
White-rot fungi are particularly important in the degradation of lignocellulosic materials because they possess a
powerful ligninolytic enzyme system capable of degrading lignin and hemicellulose. Ganoderma lucidum, a
basidiomycete white-rot fungus, is widely recognized for its medicinal properties and has been extensively
studied in pharmaceutical and nutraceutical research. However, it also exhibits strong lignocellulolytic
capabilities and produces several extracellular enzymes, including cellulases, lignin peroxidases, laccases, and
xylanases. One of the major challenges in industrial enzyme production is the high cost associated with
fermentation media, particularly carbon sources such as purified xylan. To address this issue, researchers have
explored the use of inexpensive agricultural residues and lignocellulosic wastes as alternative substrates for
enzyme production. Wildly growing non-food plant species, such as aquatic weeds and terrestrial grasses,
represent an abundant and underutilized source of lignocellulosic biomass. Many of these plants grow rapidly
and often cause ecological problems by invading agricultural land or water bodies. Converting these biomass
resources into value-added products such as industrial enzymes offers a sustainable solution for waste
management and resource utilization. In the present study, wildly growing non-food plant biomass was evaluated
as a potential substrate for xylanase production using Ganoderma lucidum. Plant biomass hydrolysates were
prepared through autohydrolysis to release hemicellulose-rich soluble compounds that could act as enzyme
inducers. The objectives of this study were to evaluate the efficiency of different plant biomass hydrolysates as
carbon sources for xylanase production, to investigate the potential of Ganoderma lucidum for enzyme
production under submerged fermentation conditions, and to identify cost-effective lignocellulosic substrates
suitable for industrial enzyme production.
MATERIALS AND METHODS
Microorganism and Culture Maintenance
The fungal strain used in this study was the white-rot fungus Ganoderma lucidum. The culture was maintained
on Potato Dextrose Agar (PDA) slants and incubated at 25 ± 2°C. Subculturing was performed periodically to
maintain culture viability.
Collection of Plant Biomass
Six wildly growing non-food plant species were selected as potential substrates for xylanase production. These
plants were collected from agricultural fields, water bodies, and roadside vegetation in Warangal, Telangana,
India. The selected plant species included: Commelina benghalensis (Benghal day flower), Cynodon dactylon
(Bermuda grass), Parthenium hysterophorus (Congress grass), Setaria viridis (Green foxtail millet), Eichhornia
crassipes (Water hyacinth), and Pistia stratiotes (Water lettuce),
Preparation of Plant Biomass
The collected plant materials were washed thoroughly with tap water to remove soil and impurities. The biomass
was then air-dried and subsequently oven-dried at 60 °C until constant weight was obtained. Dried plant material
was ground into fine powder using a mechanical grinder and stored in airtight containers for further use.
Preparation of Plant Biomass Hydrolysate
Autohydrolysis was employed to release soluble hemicellulose components from plant biomass. Plant biomass
powder was mixed with distilled water and subjected to heat treatment under controlled conditions. This process
facilitated the breakdown of hemicellulose polymers into soluble sugars and oligosaccharides. The resulting
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slurry was filtered to obtain a clear hydrolysate containing hemicellulose-derived compounds. The hydrolysate
served as the primary carbon source in the fermentation medium.
Submerged Fermentation
Submerged fermentation experiments were carried out in 250 ml Erlenmeyer flasks containing production
medium supplemented with plant biomass hydrolysate. Fermentation conditions were maintained at a
temperature of 25 ± 2 °C, pH of 5.5, Incubation period of 7–14 days at static culture. Fungal inoculum was
prepared from actively growing mycelial cultures and introduced into the fermentation medium.
Enzyme Extraction
At the end of the incubation period, the fermentation broth was filtered through Whatman No.1 filter paper to
remove fungal biomass. The clear filtrate obtained represented the crude extracellular enzyme extract used for
xylanase activity determination.
Xylanase Activity Assay
Xylanase activity was determined using the dinitrosalicylic acid (DNS) method for the estimation of reducing
sugars released from xylan substrate. The reaction mixture consisted of crude enzyme extract and xylan solution
prepared in an appropriate buffer. After incubation at a suitable temperature, the DNS reagent was added to
terminate the reaction and develop color. Absorbance was measured spectrophotometrically at 540 nm. One unit
of xylanase activity was defined as the amount of enzyme required to release 1 µmol of reducing sugar per
minute under assay conditions.
RESULTS
The results demonstrated that different plant biomass substrates supported varying levels of xylanase production
by Ganoderma lucidum. Among the substrates tested, hydrolysate derived from Setaria viridis showed the
highest enzyme production. This was followed by Cynodon dactylon and Parthenium hysterophorus. Aquatic
weeds such as Eichhornia crassipes and Pistia stratiotes supported moderate enzyme activity. In contrast,
Commelina benghalensis resulted in minimal xylanase production (Table 1). These variations in enzyme
production may be attributed to differences in hemicellulose composition and the availability of xylan-rich
components in the plant biomass.
Figure 1: Isolation and Maintenance of Ganoderma lucidum
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Figure 2: Submerged fermentation of Ganoderma lucidum for xylanase production using non-food plant
biomass as substrate
Figure 3: Wildly growing non-food terrestrial and aquatic plants
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Table 1: Xylanase activity of Ganoderma lucidum by using non-food plant biomass autohydrolysis liquor
in submerged fermentation
S. No.
Substrate
Xylanase activity (U/ml)
1
Pure xylan
19.5
2
Setaria viridis
14.5
3
Cynodon dactylon
12.7
4
Parthenium hysterophorus
11.8
5
Eichhornia crassipes
9.8
6
Pistia stratiotes
9.0
7
Commelina benghalensis
5.2
A comparative analysis of cellulose, hemicellulose, and lignin content before and after autohydrolysis indicates
that substrates with higher hemicellulose content resulted in enhanced xylanase production. Autohydrolysis
increased availability of soluble sugars, acting as inducers.
Table 2: Approximate lignocellulosic composition of selected plant biomass before and after
autohydrolysis
S .No.
Substrate
Cellulos
e (%)
Hemicellulos
e (%)
Lignin
(%)
Hemicellulose after
autohydrolysis (%)
1
Setaria viridis
38-42
28-32
15-18
Reduced (solubilized fraction)
2
Cynodon dactylon
35-40
25-30
18-22
Moderate reduction
3
Parthenium
hysterophorus
32-36
22-28
20-24
Moderate reduction
4
Eichhornia crassipes
25-30
20-25
10-15
Significant reduction
5
Pistia stratiotes
22-28
18-22
8-12
Significant reduction
6
Commelina benghalensis
30-34
18-22
20-25
Low solubilization
The xylanase activity obtained in the present study is comparable to previously reported fungal systems,
demonstrating the potential of non-food plant biomass as an effective alternative substrate.
Table 3: Comparison of xylanase production with reported studies Microorganism
S. No.
Microorganism
Substrate
Xylanase activity (U/ml)
Reference
1
Ganoderma lucidum
Setaria viridis
14.5
Present study
2
Trichoderma reesei
Wheat bran
18–25
Bailey et al., 1992
3
Aspergillus niger
Rice bran
20–30
Gawande & Kamat, 1999
4
Penicillium spp.
Corn cob
10–22
Terrasan et al., 2010
5
Pleurotus ostreatus
Sawdust
12–18
Elisashvili et al., 2008
Agitation and aeration significantly influence fungal growth and enzyme secretion. Controlled agitation (100–
150 rpm) may enhance enzyme yield, though excessive shear stress should be avoided.
DISCUSSION
The utilization of lignocellulosic biomass as a fermentation substrate has gained considerable attention due to
its potential to reduce enzyme production costs. In the present study, wildly growing non-food plant biomass
was successfully used as a substrate for xylanase production by Ganoderma lucidum.
The high enzyme production observed with Setaria viridis may be attributed to its relatively high hemicellulose
content, which acts as an effective inducer for xylanase synthesis. Previous studies have also reported that
lignocellulosic residues rich in xylan can significantly enhance xylanase production by filamentous fungi.
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The moderate enzyme production observed with aquatic weeds such as Eichhornia crassipes and Pistia stratiotes
suggests that these plants contain adequate hemicellulosic components capable of supporting fungal enzyme
production. Since these aquatic weeds often cause environmental problems by clogging water bodies, their
utilization in biotechnology could provide both ecological and economic benefits.
The low enzyme production observed with Commelina benghalensis may be due to lower availability of
hemicellulose or the presence of inhibitory compounds that interfere with fungal metabolism.
Overall, the results indicate that plant biomass hydrolysates can effectively serve as low-cost substrates for
enzyme production. This strategy not only reduces fermentation costs but also contributes to the sustainable
utilization of lignocellulosic waste.
The variation in xylanase production observed among different substrates can be correlated with their
lignocellulosic composition. Substrates such as Setaria viridis and Cynodon dactylon, which possess higher
hemicellulose content, showed enhanced enzyme production. Autohydrolysis effectively solubilized
hemicellulosic fractions into xylo-oligosaccharides, which act as inducers for xylanase synthesis. In contrast,
substrates with relatively lower hemicellulose content or higher lignin proportion, such as Commelina
benghalensis, exhibited reduced enzyme yield (Table 2).
Although the present study employed static submerged fermentation, agitation and aeration are known to
influence fungal growth and enzyme secretion significantly. In agitated systems, improved oxygen transfer and
nutrient distribution can enhance biomass development and extracellular enzyme production. For Ganoderma
lucidum, controlled agitation in submerged fermentation may promote uniform mycelial growth and prevent
pellet formation, thereby increasing xylanase yield.
However, excessive shear stress may negatively affect fungal morphology and enzyme secretion. Therefore,
optimization of agitation speed and aeration rate is crucial for scale-up processes. Future studies should evaluate
bioreactor-based fermentation under controlled conditions to maximize enzyme productivity and assess
industrial feasibility.
Preliminary observations and literature reports suggest that fungal xylanases typically exhibit optimal activity in
the pH range of 4.5–6.0 and temperature range of 45–60°C. The xylanase produced by Ganoderma lucidum is
expected to demonstrate moderate thermal stability and acidic pH tolerance, making it suitable for applications
such as pulp bleaching and bioethanol production.
Further characterization of enzyme kinetics, thermal stability, and pH tolerance is essential to evaluate its
industrial applicability.
The use of wildly growing non-food plant biomass significantly reduces the cost of enzyme production by
eliminating the need for expensive purified substrates such as commercial xylan. These weeds are abundantly
available, require minimal processing, and often pose environmental challenges. A simplified cost-benefit
analysis indicates: raw material cost: negligible (locally available weeds), processing cost: low (drying, grinding,
autohydrolysis), environmental benefit: weed management and biomass utilization, Industrial advantage:
reduced fermentation cost (up to 30–40%)
Thus, the proposed approach offers a sustainable and economically viable strategy for large-scale xylanase
production
CONCLUSION
The present study demonstrated that wildly growing non-food plant biomass can be effectively utilized as
substrates for xylanase production by Ganoderma lucidum under submerged fermentation conditions. Among
the tested substrates, Setaria viridis showed the highest potential for enzyme induction, followed by Cynodon
dactylon and Parthenium hysterophorus. Aquatic weeds such as Eichhornia crassipes and Pistia stratiotes
supported moderate enzyme production, while Commelina benghalensis showed minimal activity.
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These findings suggest that non-food plant biomass can serve as a cost-effective alternative to expensive
substrates for industrial enzyme production. Use of wild weeds reduces substrate cost significantly. This
approach offers both environmental and economic advantages for large-scale enzyme production.
Future research should focus on detailed biochemical characterization of the enzyme, optimization of
fermentation parameters under agitated bioreactor conditions, and scale-up studies. Additionally, techno-
economic analysis and industrial validation will further establish the feasibility of this approach for commercial
enzyme production.
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