International Journal of Applied Science and Engineering
Published by Chaoyang University of Technology

Sathya Priya B 1*, Soundrya S. L 1, Stalin T 2, Charles J. Banks 3, Huang-Mu Lo 4*

1 Department of Environmental Sciences, Bharathiar University, Coimbatore,
Tamil Nadu, India
2 Clonal Propagation and Research Centre, Forestry Research & Development, Karur, Tamil Nadu, India
3 University of Southampton, University Road, Southampton, United Kingdom
4 Department of Environmental Engineering and Management, Chaoyang University of Technology, Taiwan

Download Citation: |
Download PDF


ABSTRACT

The biopolymer polyhydroxybutyrate (PHB) and biopigment carotenoid produced by microbes holds significant industrial and environmental potential. This research aimed at isolating pigmented bacteria from the Western Ghats of Tamil Nadu, India, which effectively co-produced PHB, adding novelty to the study. The orange pigmented bacterium was further screened for PHB synthesis by Sudan Black B staining. Subsequent 16S rRNA gene sequencing identified the positive isolate as Bacillus chungangensis. The isolate exhibited optimal performance at a pH of 10, a temperature of 45°C, and a salinity of 6%, confirming its extremophilic nature. The isolate achieved cost-effective biopolymer and biopigment production by leveraging agro-waste substrates like sugarcane bagasse, groundnut shell, and coconut oil cake. Notably, sugarcane bagasse yielded a remarkable 63% PHB yield and groundnut shell yielded more biopigment, 4.89 g/L surpassing other waste substrates and control. The biopigment exhibited λmax at 450 nm and the FTIR studies confirmed the carotenoid nature of biopigment. Fourier transform infrared (FTIR) analysis of extracted biopolymer from fermented samples of these substrates exhibited a C=O stretch at 1635.64/cm and 1627.92/cm, confirming the presence of polyhydroxybutyrates. This study unveils the potential of Bacillus chungangensis for sustainable PHB synthesis with biopigment carotenoid as a co-product by utilizing agro-waste substrates and contributing to ecofriendly biopigment and biopolymer.


Keywords: Polyhydroxybutyrate (PHB), Agro-waste, Biopigments, Biopolymer, Bacillus, Ecofriendly, Co-production, Carotenoid


Share this article with your colleagues

 


REFERENCES

  1. Aluru, R.R., 2020. Screening and biochemical characterization of PHB producing bacterium isolated from costal region of Andhra Pradesh. Environmental and Earth Sciences Research Journal, 7(3), 116–120.

  2. Anjali, M., Sukumar, C., Kanakalakshmi, A., Shanthi, K. 2014. Enhancement of growth and production of polyhydroxyalkanoates by Bacillus subtilis from agro-industrial waste as carbon substrates. Composite Interfaces, 21(2), 111–119.

  3. Aryaraj, D., Pramitha, V. S. 2021. Extraction and characterization of Polyhydroxybutyrate (PHB) from Bacillus flexus MHO57386.1 isolated from marine sponge Oceanopia arenosa (Rao, 1941). Marine Science and Technology Bulletin, 10(2), 170–185.

  4. Ashby, R.D., Solaiman, D.K., Strahan, G.D. 2011. Efficient utilization of crude glycerol as fermentation substrate in the synthesis of poly (3-hydroxybutyrate) biopolymers. Journal of the American Oil Chemists' Society, 88, 949–959.

  5. Bhatia, S.K., Yoon, J.J., Kim, H.J., Hong, J.W., Hong, Y.G., Song, H.S., Moon, Y.M., Jeon, J.M., Kim, Y.G., Yang, Y.H. 2018. Engineering of artificial microbial consortia of Ralstonia eutropha and Bacillus subtilis for poly (3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer production from sugarcane sugar without precursor feeding. Bioresource technology, 257, 92–101.

  6. Bhuwal, A.K., Singh, G., Aggarwal, N.K., Goyal, V., Yadav, A. 2013. Isolation and screening of polyhydroxyalkanoates producing bacteria from pulp, paper, and cardboard industry wastes. International Journal of Biomaterials, 2013, 752821.

  7. Cho, S. L., Jung, M. Y., Park, M. H., Kim, W. 2010. Bacillus chungangensis sp. nov., a halophilic species isolated from sea sand. International Journal of Systematic and Evolutionary Microbiology, 60(6), 1349–1352.

  8. Choi, T.R., Jeon, J.M., Bhatia, S.K., Gurav, R., Han, Y.H., Park, Y.L., Park, J.Y., Song, H.S., Park, H.Y., Yoon, J.J., Seo, S.O. 2020. Production of low molecular weight P(3HB-co-3HV) by butyrateacetoacetate CoA-transferase (cftAB) in Escherichia coli. Biotechnology and bioprocess engineering, 25, 279–286.

  9. Dalsasso, R.R., Pavan, F.A., Bordignon, S.E., de Aragão, G.M.F., Poletto, P. 2019. Polyhydroxybutyrate (PHB) production by Cupriavidus necator from sugarcane vinasse and molasses as mixed substrate. Process Biochemistry, 85, 12–18.

  10. Danial, A.W., Hamdy, S.M., Alrumman, S.A., Gad El-Rab, S.M., Shoreit, A.A., Hesham, A.E.L. 2021. Bioplastic production by Bacillus wiedmannii AS-02 OK576278 using different agricultural wastes. Microorganisms, 9(11), 2395.

  11. Dawoud, T.M., Alharbi, N.S., Theruvinthalakal, A.M., Thekkangil, A., Kadaikunnan, S., Khaled, J.M., Rajaram, S.K. 2020. Characterization and antifungal activity of the yellow pigment produced by a Bacillus sp. DBS4 isolated from the lichen Dirinaria agealita. Saudi Journal of Biological Sciences, 27(5), 1403–1411.

  12. Desouky, S.E.S., Abdel-Rahman, M.A., Azab, M.S. and Esmael, M.E. 2017. Batch and fed-batch production of polyhydroxyalkanoates from sugarcane molasses by Bacillus flexus Azu-A2. Journal of Innovations in Pharmaceutical and Biological Sciences, 4, 55–66.

  13. Dhangdhariya, J.H., Dubey, S., Trivedi, H.B., Pancha, I., Bhatt, J.K., Dave, B.P., Mishra, S. 2015. Polyhydroxyalkanoate from marine Bacillus megaterium using CSMCRI's dry sea mix as a novel growth medium. International Journal of Biological Macromolecules, 76, 254–261.

  14. El-Sheekh, M.M., El-Abd, M.A., El-Diwany, A.I. 2015. Poly-3 hydroxybutyrate production by Bacillus flexusME-77 using some industrial wastes. Rendiconti Lincei, 26(2), 109–119.

  15. Endres, H.J. 2017. Bio-based thermoplastic and thermosets polymer. Lightweight and Sustainable Materials for Automotive Applications, CRC Press, 139–166.

  16. Getachew, A., Woldesenbet, F. 2016. Production of biodegradable plastic by polyhydroxybutyrate (PHB) accumulating bacteria using low-cost agricultural waste material. BioMed Central Research Notes, 9, 1–9.

  17. Gowda, V., Shivakumar, S. 2014. Agrowaste-based Polyhydroxyalkanoate (PHA) production using hydrolytic potential of Bacillus thuringiensis IAM 12077. Brazilian Archives of Biology and Technology, 57, 55–61.

  18. Han, J., Wu, L.P., Hou, J., Zhao, D. and Xiang, H. 2015. Biosynthesis, characterization,and hemostasis potential of tailor-madepoly (3-hydroxybutyrate-co-3-hydroxyvalerate) produced by Haloferax mediterranei. Biomacromolecules, 16(2), 578–588.

  19. Hong, K., Sun, Y., Tian, X., Chen, D., Zhou, J., Huang, W. 2019. Production of poly-hydroxybutyrate (PHB) by Halomonas sp. LY-01 using mixed carbon sources and its characterization. International Journal of Biological Macromolecules, 136, 706–711.

  20. Joseph, J. and Chithira, O.S., 2021. Characterisation of polyhydroxy Butyrate (PHB) from Bacillus subtilis strain P isolated from rhizosphere soil. InternationalJournal of Current Microbiology and Applied Sciences, 10(01), 3004–3014.

  21. Juengert, J.R., Bresan, S., Jendrossek, D. 2018. Determination of polyhydroxybutyrate (PHB) content in Ralstonia eutropha using gas chromatography and Nile red staining. Bio-protocol, 8(5), e2748.

  22. Kalaivani, R., Sukumaran, V. 2015. Enhancement of technique for optimized production of PHA frommarine bacteria, utilizing cheaply available carbonsources at Thanjavur district, India. International Journal of Current Microbiology and Applied Sciences, 4(4), 408–417.

  23. Kazi, Z., Hungund, B.S., Yaradoddi, J.S., Banapurmath, N.R., Yusuf, A.A., Kishore, K.L., Soudagar, M.E.M., Khan, T.M., Elfasakhany, A. and Buyondo, K.A. 2022. Production, characterization, and antimicrobial activity of pigment from streptomyces species. Journal of Nanomaterials, 2022, 3962301.

  24. Koch, M., Doello, S., Gutekunst, K., Forchhammer, K. 2019. PHB is produced from glycogen turn-over during nitrogen starvation in Synechocystis sp. PCC 6803. International Journal of Molecular Sciences, 20(8), 1942.

  25. Korumilli, T., Mishra, S. 2014. Carotenoid production by Bacillus clausii using rice powder as the sole substrate: Pigment analyses and optimization of key production parameters. Journal of Biochemical Technology, 5(4), 788–794.

  26. Kourmentza, C., Plácido, J., Venetsaneas, N., Burniol-Figols, A., Varrone, C., Gavala, H.N., Reis, M.A. 2017. Recent advances and challenges towards sustainable polyhydroxyalkanoate (PHA) production. Bioengineering, 4(2), 55.

  27. Krueger, C.L., Radetski, C.M., Bendia, A.G., Oliveira, I.M., Castro-Silva, M.A., Rambo, C.R., Lima, A.O. 2012. Bioconversion of cassava starch by-product into Bacillus and related bacteria polyhydroxyalkanoates. Electronic Journal of Biotechnology, 15(3), 8–8.

  28. Kumalaningsih, S., Hidayat, N., Aini, N. 2011. Optimization of polyhydroxyalkanoates (PHA) production from liquid bean curd waste by Alcaligenes latus bacteria. Journal of Agricultural Food Technology, 1, 63–67.

  29. Kumar, V., Darnal, S., Kumar, S., Kumar, S., Singh, D. 2021. Bioprocess for co-production of polyhydroxybutyrate and violacein using Himalayan bacterium Iodobacter sp. PCH194. Bioresource Technology, 319, 124235.

  30. Lamberti, F.M., Román-Ramírez, L.A., Wood, J. 2020. Recycling of bioplastics: routes and benefits. Journal of Polymers and the Environment, 28, 2551–2571.

  31. Law, J.H., Slepecky, R.A. 1961. Assay of poly-β-hydroxybutyric acid. Journal of Bacteriology, 82(1), 33–36.

  32. Liu, Y., Huang, S., Zhang, Y., Xu, F. 2014. Isolation and characterization of athermophilic Bacillus shackletonii K5 from a biotrickling filter for the production of polyhydroxybutyrate. Journal of Environmental Sciences, 26(7), 1453–1462.

  33. Lu, W.K., Chiu, T.Y., Hung, S.H., Shih, L., Chang, Y.N., 2004. Use of response surface methodology to optimize culture medium for production of poly-γ-glutamic acid by Bacillus licheniformis. International Journal of Applied Science and Engineering, 2(1), 49–58.

  34. Mahitha, G., Madhuri, R. 2015. Purification and characterization of polyhydroxybutyrate produced from marine bacteria. International Journal of Scientific & Engineering Research, 6, 71–75.

  35. Mayeli, N., Motamedi, H., Heidarizadeh, F. 2015. Production of polyhydroxybutyrate by Bacillus axaraqunsis BIPC01 using petrochemical wastewater as carbon source. Brazilian Archives of Biology and Technology, 58, 643–650.

  36. McAdam, B., Brennan Fournet, M., McDonald, P., Mojicevic, M. 2020. Production of polyhydroxybutyrate (PHB) and factors impacting its chemical and mechanical characteristics. Polymers, 12(12), 2908.

  37. Moorkoth, D., Nampoothiri, K.M. 2016. Production and characterization of poly (3-hydroxy butyrate-co-3 hydroxyvalerate) (PHBV) by a novel halotolerant mangrove isolate. Bioresource Technology, 201, 253–260.

  38. Mostafa, Y.S., Alrumman, S.A., Otaif, K.A., Alamri, S.A., Mostafa, M.S., Sahlabji, T. 2020. Production and characterization of bioplastic by polyhydroxybutyrate accumulating Erythrobacteraquimaris isolated from mangrove rhizosphere. Molecules, 25(1), 179.

  39. Nair, A.M., Annamalai, K., Kannan, S.K., Kuppusamy, S. 2014. Utilization of sugarcane molasses for the production of polyhydroxyalkanoates using Bacillus subtilis. Malaya Journal of Bioscience, 1, 24–36.

  40. Numan, M., Bashir, S., Mumtaz, R., Tayyab, S., Rehman, N.U., Khan, A.L., Al-Harrasi, A. 2018. Therapeutic applications of bacterial pigments: a review of current status and future opportunities. 3 Biotech, 8, 1–15.

  41. Pallath, N., Francis, B., Devanesan, S., Farhat, K., Balakrishnan, M. 2023. Isolation and characterization of novel carotenoid pigment from marine Planococcus maritimus MBP-2 and their biological applications. Journal of King Saud University-Science, 35(8), 102872.

  42. Poomipuk, N., Reungsang, A., Plangklang, P. 2014. Poly-β-hydroxyalkanoates production from cassava starch hydrolysate by Cupriavidus sp. KKU38. International Journal of Biological Macromolecules, 65, 51–64.

  43. Pradhan, S., Dikshit, P.K., Moholkar, V.S. 2018. Production, ultrasonic extraction, and characterization of poly (3 hydroxybutyrate) (PHB) using Bacillus megaterium and Cupriavidus necator. Polymers for Advanced Technologies, 29(8), 2392–2400.

  44. Priya, M., Selvi, C.S., Kousalya, M. 2017. Screening for potential carotogenic producing bacteria from rhizospheric soil and it act as an antibacterial activity. World Journal of Pharmacy and Pharmaceutical Sciences, 6(12), 1033–1043.

  45. Ram, S., Mitra, M., Shah, F., Tirkey, S.R., Mishra, S. 2020. Bacteria as an alternate biofactory for carotenoid production: A review of its applications, opportunities and challenges. Journal of Functional Foods, 67, 103867.

  46. Ramezani, M., Amoozegar, M.A., Ventosa, A. 2015. Screening and comparative assay of poly-hydroxyalkanoates produced by bacteria isolated from the Gavkhooni Wetland in Iran and evaluation of poly-β-hydroxybutyrate production by halotolerant bacterium Oceanimonas sp. GK1. Annals of Microbiology, 65, 517–526.

  47. Ratnakaran, P., Bhoir, M., Durve-Gupta, A. 2020. Isolation and characterization of pigment producing bacteria isolated from waste. International Journal of Applied Research, 6(4), 252–260.

  48. Ratnawati, R., Hamidah, N.Y., Pradana, M.A. and Prasetyaningrum, A. 2024. Biodegradable plastic from cross-linked rice flour: Effect of cross-linking agent and plasticizer. International Journal of Applied Science and Engineering, 21(1), 2023061.

  49. Sabapathy, P.C., Devaraj, S., Parthiban, A., Pugazhendhi, A., Kathirvel, P. 2019. Aegle marmelos: A novel low-cost substrate for the synthesis of polyhydroxyalkanoate by Bacillus aerophilus RSL-7. Biocatalysis and Agricultural Biotechnology, 18, 101021.

  50. Sakthiselvan, P., Madhumathi, R. 2018. Kinetic evaluation on cell growth and biosynthesis of polyhydroxybutyrate (PHB) by Bacillus safensis EBT1 from sugarcane bagasse. Engineering in Agriculture, Environment and Food, 11(3), 145–152.

  51. Saratale, R.G., Saratale, G.D., Cho, S.K., Kim, D.S., Ghodake, G.S., Kadam, A., Shin, H.S. 2019. Pretreatment of kenaf (Hibiscus cannabinus L.) biomass feedstock for polyhydroxybutyrate (PHB) production and characterization. Bioresource Technology, 282, 75–80.

  52. Sathiyanarayanan, G., Bhatia, S.K., Song, H.S., Jeon, J.M., Kim, J., Lee, Y.K., Kim, Y.G., Yang, Y.H. 2017. Production and characterization of medium-chain-length polyhydroxyalkanoate copolymer from Arctic psychrotrophic bacterium Pseudomonas sp. PAMC 28620. International Journal of Biological Macromolecules, 97, 710–720.

  53. Sawant, S.S., Salunke, B.K., Kim, B.S., 2014. A laboratory case study of efficient polyhydoxyalkonates production by Bacillus cereus, a contaminant in Saccharophagus degradans ATCC 43961 in minimal sea salt media. Current Microbiology, 69, 832–838.

  54. Shah, K.R. 2014. Optimization and production of polyhydroxybutarate (PHB) by Bacillus subtilis G1S1 from soil. International Journal of Current Microbiology and Applied Sciences, 3(5), 377–387.

  55. Shah, K.R. 2012. FT-IR analysis of polyhydroxyalkanoates bynovel Bacillus sp. AS 3-2 from soil of Kadi region, North Gujarat, India. Journal of Biochemical Technology, 3(4), 380–383.

  56. Sharma, R., Ghoshal, G. 2021. Characterization and cytotoxic activity of pigment extracted from Rhodotorula mucilaginosa to assess its potential as bio-functional additive in confectionary products. Journal of Food Science and Technology, 58, 2688–2698.

  57. Sukan, A., Roy, I., Keshavarz, T. 2014. Agro-industrial waste materials as substrates for the production of poly (3-hydroxybutyric acid). Journal of Biomaterials and Nanobiotechnology, 5(4), 229–240.

  58. Suwannasing, W., Imai, T., Kaewkannetra, P. 2015. Cost-effective defined medium for the production of polyhydroxyalkanoates using agricultural raw materials. Bioresource Technology, 194, 67–74.

  59. Vila, E., Hornero-Méndez, D., Azziz, G., Lareo, C., Saravia, V. 2019. Carotenoids from heterotrophic bacteria isolated from fildes peninsula, king george island, antarctica. Biotechnology Reports, 21, e00306.

  60. Williams S. T., Goodfellow M., Alderson G. 1989. Bergey's Manual of Systematic Bacteriology, in Genus Streptomyces Waksman and Henrici, 1943, 339AL, Vol. 4.


ARTICLE INFORMATION

Received: 2023-09-18
Revised: 2023-11-01
Accepted: 2024-02-08


Cite this article:

Priya B, S., Soundrya, S.L., Stalin, T., Banks, C.J., Lo, H.M. 2024. Enhanced co-production of biopolymer polyhydroxybutyrate (PHB) and biopigment (carotenoid) through wild Bacillus chungangensis by utilizing agro-waste substrates. International Journal of Applied Science and Engineering, 21, 2023332. https://doi.org/10.6703/IJASE.202406_21(2).009

  Copyright The Author(s). This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are cited.

Other people also read ...

doaj logo

 

2.3
2022CiteScore
 
 
48th percentile
Powered by  Scopus
 
 

SCImago Journal & Country Rank

IJASE - Most Read Articles

IJASE - Most popular articles