Effect of Solid State Fermented (SSF) Biomass on In Vitro Methanogenesis and Dry Matter Digestibility in Adult Surti Buffaloes

doi:10.21203/rs.3.rs-5139548/v1

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Effect of Solid State Fermented (SSF) Biomass on In Vitro Methanogenesis and Dry Matter Digestibility in Adult Surti Buffaloes

https://doi.org/10.21203/rs.3.rs-5139548/v1

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An in vitro rumen fermentation study was conducted to evaluate the effect of supplementation with solid state fermented biomass (SSF) in total mixed rations (TMRs) on total gas production, methanogenesis and dry matter digestibility by using the rumen liquor of adult Surti buffaloes. SSF biomass was supplemented at 0, 1, 2, 3, 4, 5, 6, 7, and 8% with TMR (65% wheat straw and 35% concentrate) for in vitro gas production trials. The results of the in vitro study revealed significantly (P < 0.05) greater IVDMD (58.43%) and lower CH₄ production (3.58 ml CH₄/100 mg DDM) at a 3% level of SSF biomass addition in TMR. Based on the overall results of in vitro studies, 3% SSF biomass supplementation was found to be most suitable for further in vivo studies in adult Surti buffaloes.

Animal Science

SSF Biomass

In Vitro

Digestibility

Methane

Gas Production

Improvements in feed utilization, animal production, health, and food safety are the goals of rumen microbial research. These goals can be accomplished by encouraging ideal fermentation, minimizing ruminal problems, and preventing infections. It is best to think of supplements as an addition to sound feeding procedures. An animal's behavior can be influenced by a class of feed chemicals known as feed additives, which are only required at trace levels. The use of anti biowaste agents in feed has substantially decreased in the last ten years due to the detrimental effects of these agents on animal health, the residue they left in animal products, and the possibility that microbes could become resistant to them. As a result, the use of microorganisms in animal nutrition has gained popularity.

The development of enzyme supplements that enhance fiber digestion and lower enteric methane emissions from large ruminants has been the main emphasis of research.

The two main techniques for extracting enzymes are solid-state fermentation (SSF) and submerged fermentation (SmF). Due to its decreased energy need, decreased effluent generation, and direct application of fermented products for feeding, the bioconversion of fibrous material by SSF has attracted increasing interest [Yang et al. 2011] and [Surabhiet al. 2024]. Fermentation involving solids in the absence (or almost absence) of free water is referred to as "solid-state fermentation."

For the generation of enzymes by microbial flora, solid-state fermentation has enormous promise. This method is particularly intriguing because it results in a raw, fermented product that may be employed immediately as an enzyme source [C. Senthil kumar & R. Vijay Anand et al. 2023]. Ideally, the SSF system can be used to manufacture practically every known microbial enzyme. The production of enzymes such as proteases, cellulases, xylanases, amylases, and pectinases has been the subject of extensive research [Pandey et al. 1999] and [Partha Sarathi Patra and Ashim Chandra Sinha et al. 2012]. Exogenous fibrinolytic enzymes have significantly increased digestibility and decreased intestinal methane emission and positive effects on increasing the efficiency of feed utilization by ruminants both in vitro [Murad et al. 2009] and in vivo [Arriola et al. 2011] in recent years as a result of being added to animal feeds. Therefore, the present study investigated the effect of SSF biomass supplementation on in vitro rumen fermentation.

The present study was conducted at the Animal Nutrition Research Station, College of Veterinary Science and Animal Husbandry, Kamdhenu University, Anand, Gujarat. Wheat straw, mung gotar, groundnut cake, deoiled rice bran, molasses and mineral mixture were used for preparation of the total mixed rations (TMRs). The TMR was oven dried at 70°C and finely ground in a Wiley mill using 1 mm sieve. The TMR was analyzed for proximate constituents (AOAC, 2005) and fiber fraction [Van Soest et al. 1991]

SSF biomass was procured from the Department of Microbiology, Gujarat Vidhyapeeth, Sadra, Gandhinagar, Gujarat, India. The solid-state fermented (SSF) biomass of jowar hay was carried out with a culture of Aspergillus oryzae and Trichoderma spp. fungi. The experimental TMR without any supplementation of the SSF biomass was the control group and was designated as S0, while the TMR with SSF biomass supplementation at 1, 2, 3, 4, 5, 6, 7 and 8% was designated as S1, S2, S3, S4, S5, S6, S7 and S8, respectively.

Rumen liquor for in vitro rumen fermentation studies was collected from two adult Surti buffaloes using stomach tubes. The buffaloes were fed individually with TMRs prepared to meet their nutrient requirements (ICAR 2013) with free access to water. The collected rumen liquor was strained through four layers of muslin cloth, which was termed the strained rumen liquor (SRL), and was mixed in prepared artificial saliva (McDougall’s) in proper proportions prior to incubation. Substrates (200 mg) with various levels of SSF biomass were incubated with artificial saliva mixed with SRL (40 ml) for 48 h in quadrupling at 39 ± 1°C in a shaker twin water bath [Menke et al. 1979]. After 48 h of incubation, total gas production (TGP) was recorded after subtracting the gas production from the blank. To determine in vitro methane production, gas produced in 100 ml glass syringes after a 24 h of incubation period was used. A gas sample was directly injected into gas chromatograph (GC) from each syringe, and the CH₄ concentration was determined against standard methane gas (22.54%). All the samples were analyzed using a GC instrument, fitted with an SS column (4 ft. long, 3.2 mm inside diameter) packed with Porapack N (80 to 100 mesh), and equipped with a flame ionization detector (FID). The column temperature was maintained at 50°C, and nitrogen was used as the carrier gas, with a flow rate of 30 ml/min. Calibration was completed using standards (22.54%) procured from CHEMIX Specialty Gases & Equipment., Bangalore. After completion of incubation, the content of each syringe was filtered and dried in a preweighed Gooch crucible. The IVDM was calculated by subtracting the residues remaining after incubation from the amount of substrate incubated and was expressed as a percentage.

Statistical analysis

The data generated during the experiment were analyzed by two-way analysis of variance (ANOVA) using the WASP 2.0 method as prescribed by [Snedecor and Cochran 1994].

The proximate composition and fiber fraction data of the prepared TMRs are presented in Table 1. The effects of solid-state fermented (SSF) biomass on IVDMD, total gas production (TGP) and methane (ml/100 g of digestible DM) are presented in Table 2.

Table 1

Chemical Composition and Fiber Fraction of the Total Mixed Ration
Parameters (%, on DM basis)	TMR
Crude protein	11.08
Ether extract	2.33
Crude fiber	27.42
Nitrogen free extract	45.30
Total Ash	13.87
Organic matter	78.84
Neutral detergent fiber	53.13
Acid detergent fiber	35.32
Cellulose	28.12
Hemicellulose	17.81
Lignin	5.62
Calcium	1.50

Table 2

Average *In Vitro* Dry Matter Digestibility (IVDMD, %), Total Gas Production (TGP, Ml) and Methane (Ml/100 G Digestible DM) of Substrates Containing Different Levels of SSF Biomass
Substrates	IVDMD	TGP	Methane
S0	53.12^ab ± 0.37	67.67 ± 2.96	4.01 ± 0.44
S1	48.97^a ± 0.35	35.33 ± 15.07	4.01 ± 0.77
S2	51.90^ab ± 2.78	59.00 ± 11.27	3.85 ± 0.83
S3	58.43^b ± 4.53	52.33 ± 11.79	3.58 ± 0.22
S4	50.80^ab ± 1.53	42.00 ± 14.00	4.02 ± 0.21
S5	52.83^ab ± 0.88	23.37 ± 14.66	4.27 ± 0.22
S6	48.03^a ± 1.27	58.00 ± 4.04	4.35 ± 0.36
S7	56.83^ab ± 2.90	50.33 ± 14.15	4.23 ± 0.22
S8	56.73^ab ± 5.70	64.00 ± 3.06	4.32 ± 0.49
SEM	3.03	11.24	0.48
CD(0.05)	8.99	NA	NA
CV%	9.88	38.76	20.14
* a, b, c superscripts in a column differ significantly (P < 0.05)

The findings indicated that the in vitro dry matter digestibility percentages were 53.12, 48.97, 51.90, 58.43, 50.80, 52.83, 48.03, 56.83, and 56.73 for the S0, S1, S2, S3, S4, S5, S6, S7, and S8 groups, respectively. For the S0, S1, S2, S3, S4, S5, S6, S7, and S8 groups, the corresponding in-vitro total gas production (ml) values were as follows: 67.67, 35.33, 59.00, 52.33, 42.00, 23.37, 58.00, 50.33, and 64.00. The in vitro methane production values for the S0, S1, S2, S3, S4, S5, S6, S7, and S8 groups were 4.01, 4.01, 3.85, 3.58, 4.02, 4.27, 4.35, 4.23, and 4.32, respectively.

A significantly (P < 0.05) greater percentage of patients in the 3% level of SSF biomass-supplemented group (58.43%) IVDMD than did those in the control group (53.12%). The addition of SSF biomass, which may have improved digestibility, resulted in higher fermentation rates in the present study. Additionally, the addition of enzymes enhances the growth of cellulolytic bacteria, accelerates the digestion of fiber, and facilitates the movement of microbial proteins from the rumen [Azzaz et al. 2013]. Similarly, significant (P < 0.05) improvements in IVDMD [Bhasker et al. 2012; Arati, 2013; Reddy et al. 2016; Chaudhari, 2018] were also reported by earlier authors.

A total amount of gas produced during the in-vitro incubation study of 48 h was observed, and the data are presented in Table 2. The values for total gas production were not significantly different. Methane is a substantial contributor to greenhouse gas emissions,; thus, mitigating its effects is also necessary for a better environment. The addition of probiotics, additives, or various nutrients to feed has an established effect on methane (CH₄) generation. [Mamuad et al. 2014, Lamba et al. 2014]. The effect of SSF biomass supplementation on methane production was found to numerically (P > 0.05) lower than that of the control on methane production by 21% in the 3% SSF biomass supplemented group.

It was observed that SSF biomass at the 3% level had a significantly greater IVDMD. At the 3% level of SSF, biomass supplementation numerically decreased in vitro methane production; hence, SSF biomass supplemented at the 3% level in the diet is the most suitable for further in vivo studies.

The permission for animal experiments was granted by the Institutional Animal Ethics Committee (IAEC: 375/ANRS/2022).

Funding

The research grant was received from the project of Kamdhenu University.

Data availability

The authors acknowledge that the data presented in this study will be available upon reasonable request.

Conflict of interest

All the authors affirm that they have no conflicts of interest.

Author contributions

All the contributors contributed to the conception and planning of the study. Material preparation, data collection and analysis were performed by Dr. Gaurang P. Mathukiya, Dr. Paresh R. Pandya, Dr. Kalpesh K. Sorathiya and Dr. Sunil V. Rathod. The first draft of the manuscript was written by Dr. Gaurang P. Mathukiya, and all theauthors commented on previous versions of the manuscript. All the authors read and approved the final manuscript.

Acknowledgment

The authors are grateful to the Animal Nutrition Research Station, College of Veterinary Science and Animal Husbandry, Anand. I am thankful for providing the necessary facilities and financial support for undertaking this study.

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The authors declare no competing interests.

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Effect of Solid State Fermented (SSF) Biomass on In Vitro Methanogenesis and Dry Matter Digestibility in Adult Surti Buffaloes

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Abstract

1. INTRODUCTION

2. MATERIALS AND METHODS

3. RESULTS AND DISCUSSION

4. CONCLUSION

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References

Additional Declarations

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