Characterization of Pseudomonas fluorescens polyhydroxyalkanoate produced from molasses as a carbon source

Yusuf Hendrawan, Dikianur Alvianto, Sumardi Hadi Sumarlan, Yusuf Wibisono


Polyhydroxyalkanoate (PHA) is a biopolymer produced by various types of bacteria under conditions of unbalanced growth. One of the bacteria generating PHA is Pseudomonas fluorescens with a carbon source in the form of molasses. This study aimed to determine the effect of molasses concentration and incubation period on biomass and PHA characteristics. This study utilised two factors, such as molasses concentration of 40, 60, 80 g/L, and incubation period for 3, 5, 7 days. The research result indicated the highest biomass yield was obtained from the treatment of 40 g/L molasses concentration and in 7-day incubation period, with dry cell weight values of 1,955 g/L, PHA weight of 0.756 g/L, dry cell yield of 2.036%, and PHA yield of 39.06%, respectively. The visual characteristics of PHA samples were indicated with brownish and granular state, while the melting point and functional groups were identical to pure polyhydroxybutyrate (PHB). The characteristics of the PHB structure obtained from Fourier Transform Infrared Spectroscopy (FTIR) analysis, depicting: C = O esters, CH3, CH2, -C-O-, -C-O-C Polymers, and C-C. The melting point of PHA samples,using Differential Scanning Calorimetry (DSC) analysis, was 166.4 oC and an enthalpy of -13.885 J/g with 9.5% degree of crystallisation.  


Incubation period; Molasses; polyhydroxyalkanoate; Pseudomonas fluorescens

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Albuquerque, P.B.S., and Malafaia, C.B. (2018) ‘Perspectives on the production, structural characteristics and potential applications of bioplastics derived from polyhydroxyalkanoates’, International Journal of Biological Macromolecules, 107(A), pp 615-625

Anjum, A., Zuber, M., Zia, K.M., Noreen, A., Anjum, M.N., and Tabasum, S. (2016) ‘Microbial production of polyhydroxyalkanoates (PHAs) and its copolymers: a review of recent advancements’, International Journal of Biological Macromolecules, 89, pp 161-174

Ashton, E.G., Wilson, K.J., Demori, R., Candido, L.H.A., and Mauler, R. (2016) ‘Recycling polymeric multi-material products through micronization’, Journal of Cleaner Production, 116, pp 268-278

Atifah, N., Khaswar S., and Ani S. (2007) ‘Study on bioplastic fermentation Poly-(3-hydroxyalkanoate) by Ralstonia eutropha using sago starch as carbon source’, Journal of Agricultural Technology, 8(3), pp 160-171

Aversa, C., Puopolo, M., and Barletta, M. (2019) ‘Tailor-made bioplastics for environmentally friendly food packaging: a methodological approach to a challenging problem’, Reference Module in Materials, Science and Materials Engineering,

Blanco, P.G., Cortes, O., Poznyak, T., Chairez, I., and Pena, E.I.G. (2018) ‘Polyhydroxyalkanoates (PHA) production by photoheterotrophic microbial consortia: effect of culture conditions over microbial population and biopolymer yield and composition’, European Polymer Journal, 98, pp 94-104

Bugnicourt, E., Cinelli, P., Lazzeri, A., and Alvarez, V. (2014) ‘Polyhydroxyalkanoate (PHA): review of synthesis, characteristics, processing and potential applications in packaging’, eXPRESS Polymer Letters, 8 (11), pp 791-808

Chan, C.M., Pratt, S., Halley, P., Richardson, D., Werker, A., Laycock, B., and Vandi, L.J. (2019) ‘Mechanical and physical stability of polyhydroxyalkanoate (PHA)-based wood plastic composites (WPCs) under natural weathering’, Polymer Testing, 73, pp 214-221

Chanprateep, S. (2010) ‘Current trends in biodegradable polyhydroxyalkanoates’, Journal of Bioscience and Bioengineering, 110(6), pp 621-632

Chen, G.Q., and Jiang, X.R. (2017) ‘Engineering bacteria for enhanced polyhydroxyalkanoates (PHA) biosynthesis’, Synthetic and Systems Biotechnology, 2(3), pp 192-197

Dietrich, K., Dumont, M.J., Rio, L.F.D., and Orsat, V. (2017) ‘Producing PHAs in the bioeconomy – towards a sustainable bioplastic’, Sustainable Production and Consumption, 9, pp 58-70

Din, M.F.M., Ujang, Z., Loodsrecht, M.V., Yunus, M., and Ahmad, M.A. (2006) ‘Polyhydroxyalkanoates (PHAs) production from aerobic-mixed cultures’, Malaysian Journal of Civil Engineering, 18(2), pp 109-128

Emadian, S.M., Onay, T.T., and Demirel, B. (2017) ‘Biodegradation of bioplastics in natural environments’, Waste Management, 59, pp 526-536

Fabricio, C.P., Kakazu, S., Carolina, B.C.P., Gomez, J.G.C., and Contiero, J. (2017) ‘Polyhydroxyalkanoate production from crude glycerol by newly isolated Pandoraea sp’, Journal of King Saud University-Science, 29(2), pp 166-173

Figols, A.B., Varrone, C., Simone, B.L., Dugaard, A.E., Skiadas, I.V., and Gavala, H.N. (2018) ‘Combined polyhydroxyalkanoates (PHA) and 1,3-propanediol production from crude glycerol: selective conversion of volatile fatty acids into PHA by mixed microbial consortia’, Water Research, 136, pp 180-191

Gumel, A.M., Annuar M.S.M., and Heidelberg, T. (2012) ‘Biosynthesis and characterization of polyhydroxyalkanoates copolymer produced by Pseudomonas putida Bet001 isolated from palm oil mill effluent’, Plos One, 7(9): e45214.

Hahn, S.K., Chang, Y.K., and Lee, S.Y. (1995) ‘Recovery and characterization of poly(3-hydroxybutyric acid) synthesized in Alcaligenes eutrophus and recombinant Escherica coli’, Applied and Environmental Microbiology, 61(1), pp 34-39

Jiang,Y., Xin, S., Lei G., Ping, L., Chuancao, D., and Weilan, S. (2008) ‘High poly(β-hydroxybutyrate) production by Pseudomonas fluorescens A2a5 from inexpensive substrates’, Enzyme and Microbial Technology, 42, pp 167-172

Jost, V., Schwarz, M., and Langowski, H.C. (2017) ‘Investigation of the 3-hydroxyvalerate content and degree of crystallinity of P3HB-co-3HV cast films using Raman spectroscopy’, Polymer, 133, pp 160-170

Justyna, M.C., and Kiewisz, R. (2016) ‘Bacterial polyhydroxyalkanoate: still fabulous?’, Microbiological Research, 192, pp 271-282

Kamnev, A.A., Tugarova, A.V., Dyatlova, Y.A., Tarantilis, P.A., Grigoryeva, O.P., Fainleib, A.M., and Luca, S.D. (2018) ‘Methodological effects in fourier transform infrared (FTIR) spectroscopy: implications for structural analyses of biomacromolecular samples’, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 193, pp 558-564

Kansiz, M., Billman-Jacobe, H., and McNaughton, D. (2000) ‘Quantitative determination of the biodegradable polymer poly(ß-hydroxybutyrate) in a recombinant Escherichia coli strain by use of mid-infrared spectroscopy and multivariative statistics’, Applied and Environmental Microbiology, 66(8), pp 3415–3420

Keshavarz, T., and Roy, I. (2010) ‘Polyhydroxy-alkanoates: bioplastics with a green agenda’, Current Opinian in Microbiology, 13(3), pp 321-326

Koller, M., Marsalek, L., Dias, M.M.S., and Braunegg, G. (2017) ‘Producing microbial polyhydroxy-alkanoates (PHA) biopolyesters in a sustainable manner’, New Biotechnology, 37(A), pp 24-38

Lee, S.Y, Choi, J., Han, K., and Song, J.Y. (1999) ‘Removal of endotoxin during purification of poly(3-hydroxybutyrate) from gram-negative bacteria’, Applied and Environmental Microbiology, 65(6), pp 2762-2764

Liu, F., Li., W., Ridgway, D., Gu, T., and Shen, Z. (1998) ‘Production of poly-beta-hydroxybutyrate on molasses by recombinant Escherichia coli’, Biotechnology Letters, 20(4), pp 345-348

Liu, R., Liang, L.L., Cao, W., Wu, M., Chen, K., Ma, J., Jiang, M., Wei, P., and Ouyang, P. (2013) ‘Succinate production by metabolically engineered Escherichia coli using sugarcane bagasse hydrolysate as the carbon source’, Bioresource Technology, 135, pp 574-577

Lopez, C.O., Santiago, C.A., and Margarita, R.C. (2015) ‘Modelling of microbial growth and ammonia consumption at different temperatures in the production of a polyhydroxyalkanoate (PHA) biopolymer’, Journal of Applied and Technology, 13, pp 498-503

Michailides, M.K., Tekerlekopoulou, A.G., Akratos, C.S., Coles, S., Pavlou, S., and Vayenas, D.V. (2015) ‘Molasses as an effective low-cost carbon source for biological Cr(VI) removal’, Journal of Hazardous Materials, 281, pp 95-105

Moura, A.S., Demori, R., Leao, R.M., Frankenberg, C.L.C., and Santana, R.M.C. (2019) ‘The influence of the coconut fiber treated as reinforcement in PHB (polyhydroxybutyrate) composites’, Materials Today Communications, 18, pp 191-198

Nair, A.M., Kanakalakshmi A., Seralathan K.K., and Shanty K. (2014) ‘Utilization of sugarcane molasses for the production of polyhydroxyalkanoates using Bacillus subtilis’, Malaya Journal of Bioscience, 1(1), pp 24-30

Perez, S.R., Serrano, A., Pantion, A.A., and Farinas, B.A. (2018) ‘Challenges of scaling-up PHA production from waste streams: a review’, Journal of Environmental Management, 205, pp 215-230

Purama, R.K., Al-Sabahi, J.N., and Sudesh, K. (2018) ‘Evaluation of date seed oil and date molasses as novel carbon sources for the production of poly(3Hydroxybutyrate-co-3Hydroxyhexanoate) by Cupriaavidus necator H16 Re 2058/pCB113’, Industrial Crops and Products, 119, pp 83-92

Rashid, K., Tariq, S., and Shaukat, W. (2019) ‘Attribution of molasses dosage on fresh and hardened performance of recycled aggregate concrete’, Construction and Building Materials, 197, pp 497-505

Raza, Z.A., Abid, S., and Banat, I.M. (2018) ‘Polyhydroxyalkanoates: characteristics, production, recent developments and applications’, International Biodeterioration & Biodegradation, 126, pp 45-56

Sabapathy, P.C., Devaraj, S., Parthiban, A., Pugazhendi, A., and 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, pp 101021

Sanhueza, C., Acevedo, F., Rocha, S., Villegas, P., Seeger, M., and Navia, R. (2019) ‘Polyhydroxyalkanoates as biomaterial for electrospun scaffolds’, International Journal of Biological Macromolecules, 124, pp 102-110

Saratale, R.G., Saratale, G.D., Cho, S.K., Kim, D.S., Ghodake, G.S., Kadam, A., Kumar, G., Bharabava, R.N., Banu, R., and Shin, H.S. (2019) ‘Pretreatment of kenaf (Hibiscus cannabinus L.) biomass feedstock for polyhydroxybutyrate (PHB) production and characterization’, Bioresource Technology,

Sarka, E., Bubnik, Z., Hinkova, A., Gebler, J., and Kadlec, P. (2012) ‘Molasses as a by-product of sugar crystallization and a perspective raw material’, Procedia Engineering, 42, pp 1219-1228

Sen, K.Y., Hussin, M.H., and Baidurah, S. (2019) ‘Biosynthesis of poly(3-hydroxybutyrate) (PHB) by Cupriavidus necator from various pretreated molasses as carbon source’, Biocatalysis and Agricultural Biotechnology, 17, pp 51-59

Snoei, J., Luong, D.N., and Jukka, S. (2015) ‘Fatty acids an alternative carbon source for Mcl polyhydroxyalkanoate production by Pseudomonas fluorescens SC4’, International International Journal of Latest Research in Science and Technology, 4(1), pp 7-12

Soroudi, A., and Jakubowicz, I. (2013) ‘Recycling of bioplastics, their blends and biocomposites: a review’, European Polymer Journal, 49(10), pp 2839-2858

Thellen, C., Megan, C., Danielle, F., Margaret, A., Wirsen, C., and Ratto, J.A. (2008) ‘A processing, characterization and marine biodegradation study of melt-extruded polyhydroxyalkanoate (PHA) films’, Journal Polymer Environmental, 16, pp 1-11

Xu, L., Lu, Y., Zhang, P., Han, J., Song, G., Wang, G., and Chen, K. (2019) ‘Polysaccaride produced by Bacillus subtilis using burdock oligofructose as carbon source’, Carbohydrate Polymers, 206, pp 811-819

Yan, Q., Zhao, M., Miao, H., Ruan, W., and Song, R. (2010) ‘Coupling of the hydrogen and polyhydroxyalkanoates (PHA) production through anaerobic digestion from Taihu blue algae’, Bioresource Technology, 101(12), pp 4508-4512

Zhila, N., and Shishatskaya, E. (2018) ‘Properties of PHA bi-, ter-, and quarter-polymers containing 4-hydroxybutyrate monomer units’, International Journal of Biological Macromolecules, 111, pp 1019-1026


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