Optimum Condition for Polyhydroxyalkanoate Production from Crude Glycerol by Bacillus sp. Isolated from Lipid-Containing Wastewater
DOI:
https://doi.org/10.48048/tis.2022.2588Keywords:
Crude glycerol, Polyhydroxyalkanoates, Production, Bacteria, Wastewater, Optimum condition, Bacillus sp.Abstract
Polyhydroxyalkanoates (PHAs) are a group of biopolymers used as an alternative to petroleum-based synthetic plastics. Their industrial application is not widely available due to production cost constraints, mainly from raw materials used for carbon sources. This research focuses on selecting a Bacillus strain from lipid-containing wastewater that can produce PHAs from crude glycerol and investigating the optimum conditions for producing PHAs from crude glycerol. The Bacillus strain was isolated from the wastewater storage pond at a fermented pork sausage manufacturing plant in Thailand for PHAs production utilizing Mineral Salt Medium (MSM) with 1 % w/v glucose as a carbon source. PHAs synthesis was preliminarily investigated by staining cells with Sudan Black B. The isolated FMI3 produced the highest PHAs at 43 % of the dry cell weight (DCW). It was identified as Bacillus pumilus FMI3. Crude glycerol was obtained from a by-product of the transesterification of waste cooking oil catalysed by NaOH. It was partially purified by acidification and neutralization and exhibited about 80 % w/w glycerol, 1.90 ± 0.09 % ash content, 2.80 ± 0.14 % water content, light brown colour, and pH 7.11 ± 0.35. Isolated B. pumilus FMI3 grows best and produces the highest PHAs yields under optimum conditions, i.e., MSM containing crude glycerol and ammonium sulphate at 61:1 mole ratio as carbon and nitrogen sources respectively, 15 % inoculum, initial pH of neutral at 7, and shaking at 150 rpm at 40 °C for 72 h. These conditions achieved PHAs at 79 % of DCW, classified as medium-chain-length PHAs comprising 3-hydroxyoctanoate,
3-hydroxydecanoate, 3-hydroxydodecanoate, and 3-hydroxytetradecanoate at 9, 12, 67 and 12 %, respectively. This study reported the feasibility of utilizing crude glycerol as feedstock for PHAs production by the B. pumilus FMI3 strain, which was able to grow on and tolerate salts of crude glycerol.
HIGHLIGHTS
- Bacteria which endure extreme environments (i.e., very limited of nutrients; drought and chemicals) can produce polyhydroxyalkanoates from crude glycerol
- Polyhydroxyalkanoates production from partially purified crude glycerol is presented
- The yield of polyhydroxyalkanoates achieve at 79 % of dry cell weight under the optimum condition
GRAPHICAL ABSTRACT 
Downloads
References
M Koller. Polyhydroxyalkanoate biosynthesis at the edge of water activity-haloarchaea as biopolyester factories. Bioengineering 2019; 6, 34.
C Rigouin, S Lajus, C Ocando, V Borsenberger, JM Nicaud, A Marty, L Averous and F Bordes. Production and characterization of two medium-chain-length polyhydroxyalkanoates by engineered strains of Yarrowia lipolytica. Microb. Cell Factories 2019; 18, 99.
V Mezzolla, OF D’Urso and P Poltronieri. Role of PhaC type I and Type II enzymes during PHA biosynthesis. Polymers 2018; 10, 910.
K Dietrich, MJ Dumont, LFD Rio and V Orsat. Producing PHAs in the bioeconomy-towards a sustainable bioplastic. Sustain. Prod. Consumption 2017; 9, 58-70.
A Choonut, P Prasertsan, S Klomklao and K Sangkharak. Study on mcl-PHA production by novel thermotolerant gram-positive isolate. J. Polymer. Environ. 2020; 28, 2410-21.
ZA Raza, S Abid and IM Banat. Polyhydroxyalkanoates: Characteristics, production recent developments and applications. Int. Biodeterioration Biodegradation 2018; 126, 45-56.
SA Bose, S Raja, K Jeyaram, S Arockiasamy and S Velmurugan. Investigation of fermentation condition for production enhancement of polyhydroxyalkanoate from cheese whey by Pseudomonas sp. J. Microbiol. Biotechnol. Food Sci. 2020; 9, 890-8.
CBCD Paula, FC de Paula-Elias, MN Rodrigues, LF Coelho, NMLD Oliveira, AFD Almeida and J Contiero. Polyhydroxyalkanoate synthesis by Burkholderia glumae into a sustainable sugarcane biorefinery concept. Front. Bioeng Biotechnol. 2021; 8, 631284.
Renewable Energy Market Update Outlook for 2021 and 2022, Available at: http://iea.org, accessed December 2021.
M Hajek, J Kwiecien and F Skopal. Biodiesel: The influence of de-alcoholization on reaction mixture composition after neutralization of catalyst by carbon dioxide. Fuel 2012; 96, 85-98.
FCD Paula, S Kakazu, CBCD Paula, JGC Gomez and J Contiero. Polyhydroxyalkanoate production from crude glycerol by newly isolated Pandoraea sp. J. King Saud Univ. Sci. 2017; 29, 166-73.
C Zhu, CT Nomura, JA Perrotta, AJ Stipanovic and JP Nakas. Production and characterization of poly-3-hydroxybutyrate from biodiesel-glycerol by Burkholderia cepacian ATCC 17759. Biotechnol. Progr. 2010; 26, 424-30.
C Phithakrotchanakoon, V Champreda, S Aiba, K Pootanakit and S Tanapongpipat. Production of polyhydroxyalkanoates from crude glycerol using recombinant Escherichia coli. J. Polymer. Environ. 2015; 23, 38-44.
GYA Tan, CL Chen, L Li, L Ge, L Wang, IMN Razaad, Y Li, L Zhao, Y Mo and JY Wang. Start a research on biopolymer polyhydroxyalkanoate (PHA): A review. Polymers 2014; 6, 706-54.
SS Dryabina, KM Fotina, YV Shulevich, AV Navrotskii and IA Novakov. Treatment of fat-containing wastewater using binary flocculant mixtures based on chitosan and salt of poly (2-dimethylamino) ethyl methacrylate. J. Polymer. Environ. 2019; 27, 1595-601.
A Kumar, BS Bisht, VD Joshi, AK Singh and A Talwar. Physical, chemical and bacteriological study of water from rivers of uttarakhand. J. Hum. Ecol. 2010; 32, 169-73.
O Dangsawat, C Nopsuwan, J Rattanawut and P Permpoonpattana. Isolation of Bacillus species from chicken intestine for use as prebiotic supplement in laying hen: effects on performance and egg quality. Khon Kaen Agr. J. 2019; 47, 397-404.
SS Win and TA Trabold. Chapter 6 - Sustainable waste-to-energy technologies: Transesterification. Sustain. Food Waste-To-energy Syst. 2018; 2018, 89-109.
B Hungund, VS Shyama, P Patwardhan and AM Saleh. Production of polyhydroxybutyrates from Paenibacillus durus BV-1 isolated from oil mill soil. J. Microb. Biochem. Tech. 2013; 5, 13-7.
A Getachew and F Woldesenbet. Production of biodegradable plastic by polyhydroxybutyrate (PHB) accumulating bacteria using low cost agricultural waste material. BMC Res. Notes 2016; 9, 509.
W Ishak and Z Ramlil. Recovery and purification of crude glycerol from vegetable oil transesterification. Separ. Purif. Rev. 2015; 44, 250-67.
MS Ardi and MK Aroura. Progress, prospect and challenges in glycerol purification. Renew. Sustain. Energ. Rev. 2015; 42, 1164-73.
V Sukruansuwan and SC Napathorn. Use of agro-industrial residue from the canned pineapple industry for polyhydroxybutyrate production by Cupriavidus necator strain A-04. Biotechnol. Biofuels 2018; 11, 202.
N Phanse, A Chincholikar, B Patel, P Rathore, P Vyas and M Patel. Screening of PHA (polyhydroxyalkanoate) producing bacteria from diverse sources. Int. J. Biosci. 2011; 6, 27-32.
W Penkhrue, D Jendrossek, C Khanongnuch, W Pathomaree, T Aizawa, R Behrens and S Lumyong. Response surface method for polyhydroxybutyrate (PHB) bioplastic accumulation in Bacillus drentensis BP17 using pineapple peel. Plos One 2020; 15, e0230443.
S Valappil, A Boccaccini, C Bucke and I Roy. Polyhydroxyalkanoates in Gram-positive bacteria: insights from the genera Bacillus and Streptomyces. Antonie Leeuwenhoek 2007; 91, 1-17.
H Dave, C Ramakrishna and J Desai. Production of polyhydroxybutyrate by petrochemical activated sludge and Bacillus sp. IPCB-403. Indian J. Exp. Biol. 1996; 34, 216.
A Rodriguez-Contreras, M Koller, MMDS Dias, M Calafell, G Braunegg and MS Marques-Calvo. Novel poly [(R)-3-hydroxybutyrate]-producing bacterium isolated from a Bolivian hypersaline lake. Food Tech. Biotechnol. 2013; 51, 123-30.
Y Liu, Q Lai, J Du and Z Shao. Bacillus zhangzhouensis sp. nov. and Bacillus australimaris sp. nov. Int. J. Syst. Evol. Microbiol. 2016; 66, 1193-9.
WL Nicholoson, N Munakata, G Horneck, HJ Melosh and P Stelow. Resistance of Bacillus endospores to extreme terrestrial and extraterrestrial environments. Microbiol. Mol. Biol. Rev. 2000; 64, 548-72.
SJ Lee, SB Kim, SW Kang, SO Han, C Park and SW Kim. Effect of crude glycerol-derived inhibitors on ethanol production by enterobacter aerogenes. Bioproc. Biosystems Eng. 2012; 35, 85-92.
YJ Wang, FL Hua, YF Tseng, SY Chan, SN Sin, H Chua, PHF Yu and NQ Ren. Synthesis of PHAs from waste water under various C:N ratios. Bioresource Tech. 2007; 98, 1690-3.
V Saranya and R Shenbagarathai. Effect of nitrogen and calcium sources on growth and production of PHA of Pseudomonas sp. LDC-5 and it’s mutant. Curr. Res. J. Biol. Sci. 2010; 2, 164-7.
EZ Gomaa. Production of polyhydroxyalkanoates (PHAs) by Bacillus subtilis and Escherichia coli grown on cane molasses fortified with ethanol. Braz. Arch. Biol. Tech. 2014; 57, 145-54.
Q Wang and CT Nomura. Monitoring differences in gene expression levels and polyhydroxy-alkanoate (PHA) production in Pseudomonas putida KT2440 grown on different carbon sources. J. Biosci. Bioeng. 2010; 110, 653-9.
A Parvathi, K Krishna, J Jose, N Joseph and S Nair. Biochemical and molecular characterization of Bacillus pumilus isolated from coastal environment in cochin, India. Braz. J. Microbiol. 2009; 40, 269-75.
SP Mohandas, L Balan, N Lekshmi, SS Cubelio, R Philip and ISB Singh. Production and characterization of polyhydroxybutyrate from vibrio harveyi MCCB 284 utilizing glycerol as carbon source. J. Appl. Microbiol. 2016; 122, 698-707.
AU Rehman, A Aslam, R Masood, MN Aftab, R Ajmal and IU Haq. Production and characterization of a thermostable bioplastic (poly--hydroxybutyrate) from Bacillus cereus NRRL-B-3711. Pakistan J. Bot. 2016; 48, 349-56.
G Pagliano, W Gugliucci, E Torrieri, A Piccolo, S Cangemi, FAD Giuseppe, A Robertielb, V Faraco, O Pepe and V Ventorino. Polyhydroxyalkanoates (PHAs) from dairy wastewater effluent: Bacterial accumulation, structural characterization and physical properties. Chem. Biol. Tech. Agr. 2020; 7, 29.
K Sangkharak and P Prasertsan. The production of polyhydroxyalkanoate by Bacillus licheniformis using sequential mutagenesis and optimization. Biotechnol. Bioproc. Eng. 2013; 18, 272-9.
MJ de Smet, G Eggink, B Witholt, J Kingma and H Wynberg. Characterization of intracellular inclusions formed by Pseudomonas oleovorans during growth on octane. J. Bacteriol. 1983; 154, 870-8.
DY Kim, HW Kim, MG Chung and YH Rhee. Biosynthesis, modification, and biodegradation of bacterial medium-chain-length polyhydroxyalkanoates. J. Microbiol. 2007; 45, 87-97.
SP Ouyang, RC Luo, SS Chen, Q Liu, A Chung, Q Wu and GQ Chen. Production of polyhydroxy-alkanoates with high 3-hydroxydodecanoate monomer content by fadB and fadA knockout mutant of Pseudomonas putida KT2442. Biomacromolecules 2007; 8, 2504-11.
AK Singh, R Bhati and N Mallick. Pseudomonas aeruginosa MTCC 7925 as a biofactory for production of the novel scl-lcl-PHA thermoplastic from non-edible oils. Curr. Biotechnol. 2015; 4, 65-74.
RG Saratale, GD Saratale, SK Cho, DS Kim, GS Ghodake, A Kadam, G Kumar, RN Bharabava, R Banu and HS Shin. Pretreatment of kenaf (Hibiscus cannabinus L.) biomass feedstock for polyhydroxybutyrate (PHB) production and characterization. Bioresource Tech. 2019; 282, 75-80.
ADS Moura, R Demori, RM Leão, CLC Frankenberg and RMC Santana. The influence of the coconut fiber treated as reinforcement in PHB (polyhydroxybutyrate) composites. Mater. Today Comm. 2019; 18, 191-8.
S Kumalaningsih, N Hidayat and N Aini. Optimization of polyhydroxybutyrates (PHB) production from liquid bean curd waste by Alcaligenes Latus bacteria. J. Agr. Food. Tech. 2011; 1, 63-7.
AM Gumel, MSM Annuar and T Heidelberg. Biosynthesis and characterization of poly-hydroxyalkanoates copolymers produced by Pseudomonas putida Bet001 isolated from palm oil mill effluent. Plos One 2012; 7, e45214.
AK Bhuwal, G Singh, NK Aggarwal, V Goyal and A Yadav. Isolation and screening of polyhydroxyalkanoates producing bacteria from pulp, paper, and cardboard industry wastes. Int. J. Biomater. 2013; 2013, 752821.
MC Sin, MSM Annuar, IKP Tan and SN Gan. Thermodegradation of medium-chain-length poly(3-hydroxyalkanoates) produced by Pseudomonas putida from oleic acid. Polymer Degrad. Stabil. 2010; 95, 2334-42.
Downloads
Published
Issue
Section
License
Copyright (c) 2022 Walailak University
This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.
