Microbe Assisted Saccharification of Agriculture Waste Using Bacillus Sp. Enzyme Cocktail
Keywords:
Bacillus, scarification, agriculture waste, lignocellulosic enzyme, Cocktail
Abstract
A lignocellulosic degrading enzyme cocktail producing Bacillus sp. was isolated and assessed for its efficient saccharification efficiency. Maximal production of cellulases (100 U/mL), xylanases (108 U/mL) and laccases (56 U/mL), were produced using moistened 0.3% MD5 medium maintained at 37 °C, pH 9, 200 µM CuSO4 with 4% inoculum volume after 48h of incubation. An ecofriendly and economical mixed agriculture waste comprising of wheat straw, sugarcane bagasse and rice straw, served as good source for bioethanol production due to its carbohydrate contents. Statistical optimization using response surface methodology was done to carry out the enzyme cocktail treated saccharification of alkali-treated mixed agriculture biomass. Enzyme cocktail was used to treat 100 g of alkali-treated mixed agriculture waste maintained at pH 8.5 and 65oC for 18h and gave a net yield of 10.2 and 29 mg/mL of reducing sugars and glucose, respectively. This study highlights the potential of Bacillus to produce a cocktail of lignocellulolytic enzymes that could be used to produce bioethanol, pulp and fruit industry.
References
1. Agrawal S, Yadav RD, Mahajan R (2016) Synergistic effect of xylano-pectinolytic enzymes produced by a bacterial isolate in bleaching of plywood industrial waste. J Clean Prod 118:229-33.
2. Amadi OC, Awodiran IP, Moneke AN, Nwagu TN, Egong JE, Chukwu GC (2022) Concurrent production of cellulase, xylanase, pectinase and immobilization by combined cross-linked enzyme aggregate strategy-advancing tri-enzyme biocatalysis. Bioresour Technol Rep. 18:101019.
3. Bailey MJ, Biely P, Poutanen K (1992) Interlaboratory testing of methods for assay of xylanase activity. J of Biotechnol. 23(3):257-70.
4. Bains J, Capalash N, Sharma P (2003) Laccase from a non-melanogenic, alkalotolerant γ-proteobacterium JB isolated from industrial wastewater drained soil. Biotechnol Lett. 25:1155-9.
5. Bakry MM, Salem SS, Atta HM, El-Gamal MS, Fouda A (2024) Xylanase from thermotolerant Bacillus haynesii strain, synthesis, characterization, optimization using Box-Behnken Design, and biobleaching activity. Biomass Convers Biorefin. 14(8):9779-92.
6. Bala A, Singh B (2019) Development of an environmental-benign process for efficient pretreatment and saccharification of Saccharum biomasses for bioethanol production. Renew Energy. 130:12-24.
7. Benatti AL, Polizeli MD (2023) Lignocellulolytic biocatalysts: the main players involved in multiple biotechnological processes for biomass valorization. Microorganisms. 11(1):162.
8. Buckeridge MS, De Souza AP, editors (2017) Advances of basic science for second generation bioethanol from sugarcane. Cham: Springer International Publishing. 7-9.
9. Cappuccino J (2018) Microbiology a laboratory manual. Pearson.
10. Chadha R and Sivamani G (2021) Critical Minerals for India. Available from: https://csep.org/wp-content/uploads/2021/12/Critical-Minerals-for-Green-Technologies-1.pdf [Assessed on 12 February 2021].
11. Fang H, Li C, Zhao J, Zhao C (2021) Biotechnological advances and trends in engineering Trichoderma reesei towards cellulase hyperproducer. Biotechnol Bioprocess Eng. 26:517-28.
12. Harnentis YM, Rizal Y, Mahata ME (2013) Isolation, characterization and production of mannanase from thermophilic bacteria to increase the feed quality. Pak J Nut. 12(4):360-4.
13. Kannan N (2002) Laboratory manual in general microbiology. Panima publishing corporation.
14. Kasana RC, Salwan R, Dhar H, Dutt S, Gulati A (2008) A rapid and easy method for the detection of microbial cellulases on agar plates using gram's iodine. Curr Microbiol. 57(5):503-7. doi: 10.1007/s00284-008-9276-8
15. Li M, Luo N and Lu Y (2017) Biomass energy technological paradigm (BETP): trends in this sector. Sustainability. 9(4): p.567.
16. Limkar MB, Pawar SV, Rathod VK (2019) Statistical optimization of xylanase and alkaline protease co-production by Bacillus spp using Box-Behnken Design under submerged fermentation using wheat bran as a substrate. Biocatal Agric Biotechnol. 17:455-64.
17. Llimós Turet, J (2022). Lignocellulolytic enzymes production via solid-state fermentation of agroindustrial residues: process optimization and application.
18. Maitan-Alfenas GP, Visser EM, Guimarães VM (2015) Enzymatic hydrolysis of lignocellulosic biomass: converting food waste in valuable products. Curr Opin Food Sci. 1:44-9.
19. Mansora AM, Lima JS, Anib FN, Hashima H and Hoa WS (2019) Characteristics of cellulose, hemicellulose and lignin of MD2 pineapple biomass. Chem Eng 72(1): 79-84
20. Maurya DP, Singla A and Negi S (2015) An overview of key pretreatment processes for biological conversion of lignocellulosic biomass to bioethanol. 3 Biotech. 5(5): 597-609.
21. Miller GL (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugar Analytical Chemistry 31 (3) 426–428.
22. More SS, PS R, K P, Malini S, SM V (2011) Isolation, purification, and characterization of fungal laccase from Pleurotus sp. Enzyme Res. 2011(1):248735.
23. Morin LG, Prox, J (1973) Single Glucose Oxidase- Peroxidase reagent for two-minute determination of serum glucose. Clin Chem. 19(9), 959-962.
24. Nkohla A, Okaiyeto K, Olaniran A, Nwodo U, Mabinya L, Okoh A (2017) Optimization of growth parameters for cellulase and xylanase production by Bacillus species isolated from decaying biomass. J Biotech Res. 8:33.
25. Park SA, Bhatia SK, Park HA, Kim SY, Sudheer PD, Yang YH, Choi KY (2021) Bacillus subtilis as a robust host for biochemical production utilizing biomass. Crit Rev Biotechnol. 41(6):827-48.
26. Phukon LC, Abedin MM, Chourasia R, Singh SP, Tayung K, Rai AK (2024) Valorization of agro-wastes by Bacillus altitudinis XYL17 through simultaneous production of xylanase, xylooligosaccharides, and antioxidant compounds. Ind Crop Prod. 213:118395.
27. Rosado MJ, Rencoret J, Marques G, Gutiérrez A, Del Río JC (2021) Structural characteristics of the guaiacyl-rich lignins from rice (Oryza sativa L.) husks and straw. Front. Plant Sci. 12:640475. doi: 10.3389/fpls.2021.640475.
28. Sharma S, Kundu A, Basu S, Shetti NP, Aminabhavi TM (2020) Sustainable environmental management and related biofuel technologies. J Environ Manag. 1;273:111096.
29. Sreena CP, Vimal KP, Sebastian D (2016) Production of cellulases and xylanase from Bacillus subtilis MU S1 isolated from protected areas of Munnar wildlife division. J Microbiol Biotechnol Food Sci.1;5(6):500.
30. Srivastava N, Srivastava M, Mishra PK, Ramteke PW, Singh RL, editors (2019) New and future developments in microbial biotechnology and bioengineering: From cellulose to cellulase: Strategies to improve biofuel production. Elsevier.
31. Su Y, Liu C, Fang H, Zhang D (2020) Bacillus subtilis: a universal cell factory for industry, agriculture, biomaterials and medicine. Microb Cell Fact. 19:1-2. https://doi.org/10.1186/s12934-020-01436-8.
32. Teather RM, Wood PJ (1982) Use of Congo red-polysaccharide interactions in enumeration and characterization of cellulolytic bacteria from the bovine rumen. Appl Environ Microbiol. 43(4):777-80.
33. Thite VS, Nerurkar AS, Baxi NN (2020) Optimization of concurrent production of xylanolytic and pectinolytic enzymes by Bacillus safensis M35 and Bacillus altitudinis J208 using agro-industrial biomass through Response Surface Methodology. Sci Rep. 10(1):3824.
34. Tien M, Kirk TK (1984) Lignin-degrading enzyme from Phanerochaete chrysosporium: purification, characterization, and catalytic properties of a unique H2O2-requiring oxygenase. Proc Natl Acad Sci. 81(8):2280-4.
35. Tunio AA, Naqvi M, Qureshi AS, Khushk I, Jatt AN, Nizami AS, Naqvi HA, Charan TR, Bhutto MA, Tunio NA (2024) Multi enzyme production from mixed untreated agricultural residues applied in enzymatic saccharification of lignocellulosic biomass for biofuel. Process Saf Environ Prot. 186:540-51.
36. Ugras S, Bicen HE, Emire Z (2024) Determination of Cellulase Enzyme Produced by Bacillus cereus DU-1 Isolated from Soil, and Its Effects on Cotton Fiber. Braz Arch Biol Technol 67:e24230391.
37. Ullah K, Sharma VK, Ahmad M, Lv P, Krahl J, Wang Z (2018) The insight views of advanced technologies and its application in bio-origin fuel synthesis from lignocellulose biomasses waste, a review. Renew Sustain Energy Rev. 1;82: 3992-4008.
38. Vu V, Farkas C, Riyad O, Bujna E, Kilin A, Sipiczki G, Sharma M, Usmani Z, Gupta VK, Nguyen QD (2022) Enhancement of the enzymatic hydrolysis efficiency of wheat bran using the Bacillus strains and their consortium. Bioresour Technol. 343:126092.
39. Wise J (2021) COP26 summit: leaders frustrated at watered down climate deal. Available from:https://en.x-mol.com/paper/article/1460737005525565440 [Assessed on 20 December 2021].
40. Yadav A, Ali AA, Ingawale M, Raychaudhuri S, Gantayet LM, Pandit A (2020) Enhanced co-production of pectinase, cellulase and xylanase enzymes from Bacillus subtilis ABDR01 upon ultrasonic irradiation. Process Biochem. 92:197-201.
41. Zhang X, Al-Dossary A, Hussain M, Setlow P, Li J (2020) Applications of Bacillus subtilis Spores in Biotechnology and Advanced Materials. Appl Environ Microbiol. 18; 86 (17):e01096-20. doi: 10.1128/AEM.01096-20. PMID: 32631858; PMCID: PMC7440806.
2. Amadi OC, Awodiran IP, Moneke AN, Nwagu TN, Egong JE, Chukwu GC (2022) Concurrent production of cellulase, xylanase, pectinase and immobilization by combined cross-linked enzyme aggregate strategy-advancing tri-enzyme biocatalysis. Bioresour Technol Rep. 18:101019.
3. Bailey MJ, Biely P, Poutanen K (1992) Interlaboratory testing of methods for assay of xylanase activity. J of Biotechnol. 23(3):257-70.
4. Bains J, Capalash N, Sharma P (2003) Laccase from a non-melanogenic, alkalotolerant γ-proteobacterium JB isolated from industrial wastewater drained soil. Biotechnol Lett. 25:1155-9.
5. Bakry MM, Salem SS, Atta HM, El-Gamal MS, Fouda A (2024) Xylanase from thermotolerant Bacillus haynesii strain, synthesis, characterization, optimization using Box-Behnken Design, and biobleaching activity. Biomass Convers Biorefin. 14(8):9779-92.
6. Bala A, Singh B (2019) Development of an environmental-benign process for efficient pretreatment and saccharification of Saccharum biomasses for bioethanol production. Renew Energy. 130:12-24.
7. Benatti AL, Polizeli MD (2023) Lignocellulolytic biocatalysts: the main players involved in multiple biotechnological processes for biomass valorization. Microorganisms. 11(1):162.
8. Buckeridge MS, De Souza AP, editors (2017) Advances of basic science for second generation bioethanol from sugarcane. Cham: Springer International Publishing. 7-9.
9. Cappuccino J (2018) Microbiology a laboratory manual. Pearson.
10. Chadha R and Sivamani G (2021) Critical Minerals for India. Available from: https://csep.org/wp-content/uploads/2021/12/Critical-Minerals-for-Green-Technologies-1.pdf [Assessed on 12 February 2021].
11. Fang H, Li C, Zhao J, Zhao C (2021) Biotechnological advances and trends in engineering Trichoderma reesei towards cellulase hyperproducer. Biotechnol Bioprocess Eng. 26:517-28.
12. Harnentis YM, Rizal Y, Mahata ME (2013) Isolation, characterization and production of mannanase from thermophilic bacteria to increase the feed quality. Pak J Nut. 12(4):360-4.
13. Kannan N (2002) Laboratory manual in general microbiology. Panima publishing corporation.
14. Kasana RC, Salwan R, Dhar H, Dutt S, Gulati A (2008) A rapid and easy method for the detection of microbial cellulases on agar plates using gram's iodine. Curr Microbiol. 57(5):503-7. doi: 10.1007/s00284-008-9276-8
15. Li M, Luo N and Lu Y (2017) Biomass energy technological paradigm (BETP): trends in this sector. Sustainability. 9(4): p.567.
16. Limkar MB, Pawar SV, Rathod VK (2019) Statistical optimization of xylanase and alkaline protease co-production by Bacillus spp using Box-Behnken Design under submerged fermentation using wheat bran as a substrate. Biocatal Agric Biotechnol. 17:455-64.
17. Llimós Turet, J (2022). Lignocellulolytic enzymes production via solid-state fermentation of agroindustrial residues: process optimization and application.
18. Maitan-Alfenas GP, Visser EM, Guimarães VM (2015) Enzymatic hydrolysis of lignocellulosic biomass: converting food waste in valuable products. Curr Opin Food Sci. 1:44-9.
19. Mansora AM, Lima JS, Anib FN, Hashima H and Hoa WS (2019) Characteristics of cellulose, hemicellulose and lignin of MD2 pineapple biomass. Chem Eng 72(1): 79-84
20. Maurya DP, Singla A and Negi S (2015) An overview of key pretreatment processes for biological conversion of lignocellulosic biomass to bioethanol. 3 Biotech. 5(5): 597-609.
21. Miller GL (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugar Analytical Chemistry 31 (3) 426–428.
22. More SS, PS R, K P, Malini S, SM V (2011) Isolation, purification, and characterization of fungal laccase from Pleurotus sp. Enzyme Res. 2011(1):248735.
23. Morin LG, Prox, J (1973) Single Glucose Oxidase- Peroxidase reagent for two-minute determination of serum glucose. Clin Chem. 19(9), 959-962.
24. Nkohla A, Okaiyeto K, Olaniran A, Nwodo U, Mabinya L, Okoh A (2017) Optimization of growth parameters for cellulase and xylanase production by Bacillus species isolated from decaying biomass. J Biotech Res. 8:33.
25. Park SA, Bhatia SK, Park HA, Kim SY, Sudheer PD, Yang YH, Choi KY (2021) Bacillus subtilis as a robust host for biochemical production utilizing biomass. Crit Rev Biotechnol. 41(6):827-48.
26. Phukon LC, Abedin MM, Chourasia R, Singh SP, Tayung K, Rai AK (2024) Valorization of agro-wastes by Bacillus altitudinis XYL17 through simultaneous production of xylanase, xylooligosaccharides, and antioxidant compounds. Ind Crop Prod. 213:118395.
27. Rosado MJ, Rencoret J, Marques G, Gutiérrez A, Del Río JC (2021) Structural characteristics of the guaiacyl-rich lignins from rice (Oryza sativa L.) husks and straw. Front. Plant Sci. 12:640475. doi: 10.3389/fpls.2021.640475.
28. Sharma S, Kundu A, Basu S, Shetti NP, Aminabhavi TM (2020) Sustainable environmental management and related biofuel technologies. J Environ Manag. 1;273:111096.
29. Sreena CP, Vimal KP, Sebastian D (2016) Production of cellulases and xylanase from Bacillus subtilis MU S1 isolated from protected areas of Munnar wildlife division. J Microbiol Biotechnol Food Sci.1;5(6):500.
30. Srivastava N, Srivastava M, Mishra PK, Ramteke PW, Singh RL, editors (2019) New and future developments in microbial biotechnology and bioengineering: From cellulose to cellulase: Strategies to improve biofuel production. Elsevier.
31. Su Y, Liu C, Fang H, Zhang D (2020) Bacillus subtilis: a universal cell factory for industry, agriculture, biomaterials and medicine. Microb Cell Fact. 19:1-2. https://doi.org/10.1186/s12934-020-01436-8.
32. Teather RM, Wood PJ (1982) Use of Congo red-polysaccharide interactions in enumeration and characterization of cellulolytic bacteria from the bovine rumen. Appl Environ Microbiol. 43(4):777-80.
33. Thite VS, Nerurkar AS, Baxi NN (2020) Optimization of concurrent production of xylanolytic and pectinolytic enzymes by Bacillus safensis M35 and Bacillus altitudinis J208 using agro-industrial biomass through Response Surface Methodology. Sci Rep. 10(1):3824.
34. Tien M, Kirk TK (1984) Lignin-degrading enzyme from Phanerochaete chrysosporium: purification, characterization, and catalytic properties of a unique H2O2-requiring oxygenase. Proc Natl Acad Sci. 81(8):2280-4.
35. Tunio AA, Naqvi M, Qureshi AS, Khushk I, Jatt AN, Nizami AS, Naqvi HA, Charan TR, Bhutto MA, Tunio NA (2024) Multi enzyme production from mixed untreated agricultural residues applied in enzymatic saccharification of lignocellulosic biomass for biofuel. Process Saf Environ Prot. 186:540-51.
36. Ugras S, Bicen HE, Emire Z (2024) Determination of Cellulase Enzyme Produced by Bacillus cereus DU-1 Isolated from Soil, and Its Effects on Cotton Fiber. Braz Arch Biol Technol 67:e24230391.
37. Ullah K, Sharma VK, Ahmad M, Lv P, Krahl J, Wang Z (2018) The insight views of advanced technologies and its application in bio-origin fuel synthesis from lignocellulose biomasses waste, a review. Renew Sustain Energy Rev. 1;82: 3992-4008.
38. Vu V, Farkas C, Riyad O, Bujna E, Kilin A, Sipiczki G, Sharma M, Usmani Z, Gupta VK, Nguyen QD (2022) Enhancement of the enzymatic hydrolysis efficiency of wheat bran using the Bacillus strains and their consortium. Bioresour Technol. 343:126092.
39. Wise J (2021) COP26 summit: leaders frustrated at watered down climate deal. Available from:https://en.x-mol.com/paper/article/1460737005525565440 [Assessed on 20 December 2021].
40. Yadav A, Ali AA, Ingawale M, Raychaudhuri S, Gantayet LM, Pandit A (2020) Enhanced co-production of pectinase, cellulase and xylanase enzymes from Bacillus subtilis ABDR01 upon ultrasonic irradiation. Process Biochem. 92:197-201.
41. Zhang X, Al-Dossary A, Hussain M, Setlow P, Li J (2020) Applications of Bacillus subtilis Spores in Biotechnology and Advanced Materials. Appl Environ Microbiol. 18; 86 (17):e01096-20. doi: 10.1128/AEM.01096-20. PMID: 32631858; PMCID: PMC7440806.
Published
2024-12-20
How to Cite
Sunny Banyal, Shikha Kumari, & Vijay Kumar. (2024). Microbe Assisted Saccharification of Agriculture Waste Using Bacillus Sp. Enzyme Cocktail. Revista Electronica De Veterinaria, 25(2), 1446 -1457. https://doi.org/10.69980/redvet.v25i2.1832
Section
Articles
