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Enzymatic degradation of polyhydroxyalkanoate using lipase from Bacillus subtilis

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Abstract

Polyhydroxyalkanoates (PHAs) are an important class of biodegradable polymers synthesized by a few bacteria under nutrient-limiting conditions. In this study, the lipase-catalysed degradation of PHA synthesized by Enterobacter sp. was monitored. For this, the lipase-encoding gene from Bacillus subtilis DI2 was PCR-amplified, cloned into a T vector system and sequenced. It was expressed in Escherichia coli DH5α cells, the recombinant enzyme was purified 24.25-fold, and its molecular weight was determined to be around 28 kDa. When PHA biodegradation studies were carried out with this enzyme, gel permeation chromatography showed 21.3 and 28.3 % molecular weight decrease and weight loss, respectively. Further, scanning electron micrographs revealed alterations in polymer surface morphology. Changes in molecular vibrations were noticed in the FTIR spectra. When the chemical shifts in NMR spectra were studied, a steep reduction in area under the peak at 1.57 ppm was observed. In the heating range of 30–930 °C employed during thermogravimetry analysis, the degraded sample showed a total of 45.82 % weight loss, as against 18.89 % for the native sample. The melting temperature (T m) of the polymer was also brought down from 126.22 to 118.18 °C, as inferred from differential scanning calorimetry. Lipase-catalysed chain scission reactions could thus be used to generate low molecular weight functional biopolymers with wide-ranging pharmaceutical applications, such as in sustained drug release.

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References

  • Bhuwal AK, Singh G, Aggarwal NK, Goyal V, Yadav A (2013) Isolation and screening of polyhydroxyalkanoates producing bacteria from pulp, paper, and cardboard industry wastes. Int J Biomater 2013, 1–10, Article ID 75282

  • Chen C, Yu CH, Cheng YC, Yu PHF, Cheung MK (2006) Preparation and characterization of biodegradable nanoparticles based on amphiphilic poly(3-hydroxybutyrate)epoly(ethylene glycol)epoly(3-hydroxybutyrate) triblock copolymer. Eur Polym J 42:2211–2220

    Article  CAS  Google Scholar 

  • Colak A, Guner S (2004) Polyhydroxyalkanoate degrading hydrolase-like activities by Pseudomonas sp. isolated from soil. Int Biodeterior Biodegr 53:103–109

    Article  CAS  Google Scholar 

  • Fiorese M, Freita F, Pais J, Ramos AM, Aragao C, Reis M (2009) Recovery of polyhydroxybutyrate (PHB) from Cupriavidus necator biomass by solvent extraction with 1,2-propylene carbonate. Eng Life Sci 9:454–461

    Article  CAS  Google Scholar 

  • Gumel AM, Annuar MSM, Chisti Y, Heidelberg T (2012) Ultrasound assisted lipase catalyzed synthesis of poly-6-hydroxyhexanoate. Ultrason Sonochem 19:659–667

    Article  CAS  Google Scholar 

  • Hauttecoeur B, Jolivet M, Gavard R (1994) Depolymerization, by a controlled chemical process, of beta hydroxybutyric lipid (or PHB) of Bacillus megaterium. C R Acad Sci Hebd Seances Acad Sci D 274(19):2729–2732

    Google Scholar 

  • Horowitz DM, Sanders JKM (1994) Amorphous, biomimetic granules of polyhydroxybutyrate: preparation, characterization, and biological implications. J Am Chem Soc 116:2695–2702

    Article  CAS  Google Scholar 

  • Khanna S, Srivastava AK (2005) Recent advances in microbial polyhydroxyalkanoates. Process Biochem 40:607–619

    Article  CAS  Google Scholar 

  • Kimura M (1980) A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16(2):111–120

    Article  CAS  Google Scholar 

  • Kuntanoo K, Promkotra S, Kaewkannetra P (2013) Biodegradation of polyhydroxybutyrate-co-hydroxyvalerate (PHBV) blended with natural rubber in soil environment. Int Sci Index 7(12): waset.org/Publication/16662 (2013)

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  CAS  Google Scholar 

  • Lauzier C, Revol JF, Debzi EM, Marchessault RH (1994) Hydrolytic degradation of isolated poly(b-hydroxybutyrate) granules. Polymer 35:4156–4162

    Article  CAS  Google Scholar 

  • Ma J, Zhang Z, Wang B, Kong X, Wang Y, Cao S, Feng Y (2006) Overexpression and characterization of a lipase from Bacillus subtilis. Protein Expres Purif 45:22–29

    Article  CAS  Google Scholar 

  • Motiwalla M, Punyarthi P, Mehta M, D’Souza J, Kelkar-Mane V (2013) Studies on degradation efficiency of polycaprolactone by a naturally occurring bacterium. J Environ Biol 34:43–49

    Google Scholar 

  • Nguyen S, Yu G-E, Marchessault RH (2002) Thermal degradation of poly(3-hydroxyalkanoates): preparation of well-defined oligomers. Biomacromol 3(1):219–224

    Article  CAS  Google Scholar 

  • Obruca S (2010) Controlled production and degradation of selected biomaterials. Dissertation, Brno University, Czech Republic

  • Philip S, Keshavarz T, Roy I (2007) Polyhydroxyalkanoates: biodegradable polymers with a range of applications. J Chem Technol Biotechnol 82(3):233–247

    Article  CAS  Google Scholar 

  • Rahayu A, Zaleha Z, Yahya ARM, Majid MIA, Amirul AA (2008) Production of copolymer poly(3-hydroxybutyrate-co-4-hydroxybutyrate) through a one-step cultivation process. World J Microbiol Biotechnol 24:2403–2409

    Article  CAS  Google Scholar 

  • Rodríguez-Contreras A, Calafell-Monfort M, Marqués-Calvo MS (2012a) Enzymatic degradation of poly(3-hydroxybutyrate) by a commercial lipase. Polym Degrad Stabil 97:2473–2476

    Article  Google Scholar 

  • Rodríguez-Contreras A, Calafell-Monfort M, Marqués-Calvo MS (2012b) Enzymatic degradation of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) by commercial lipases. Polym Degrad Stabil 97:597–604

    Article  Google Scholar 

  • Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Springs Harbor Laboratory, New York

    Google Scholar 

  • Sanchez J, Tsuchii A, Tokiwa Y (2000) Degradation of polycaprolactone at 50°C by a thermotolerant Aspergillus sp. Biotechnol Lett 22:849–853

    Article  CAS  Google Scholar 

  • Sandhya M, Aravind J, Kanmani P (2013) Production of polyhydroxyalkanoates from Ralstonia eutropha using paddy straw as cheap substrate. Int J Environ Sci Technol 10(1):47–54

    Article  CAS  Google Scholar 

  • Sharma L, Mallick N (2005) Accumulation of poly—hydroxybutyrate in Nostoc musocrum: regulation by pH, light-dark cycles, N and P status and carbon sources. Bioresour Technol 96:1304–1310

    Article  CAS  Google Scholar 

  • Sudesh K, Abe H, Doi Y (2000) Synthesis, structure and properties of polyhydroxyalkanoates: biological polyesters. Prog Polym Sci 25:1503–1555

    Article  CAS  Google Scholar 

  • Tanamool V, Imai T, Danvirutai P, Kaewkannetra P (2011) Biosynthesis of polyhydroxyalkanoate (PHA) by Hydrogenophaga sp. isolated from soil environment during batch fermentation. J Life Sci 5(12):1003–1012

    CAS  Google Scholar 

  • Vinish V, Sangeetha SH, Aravind J, Kanmani P, Sathishkumar T (2015) Optimizing the nutrient feeding strategy for PHA production by a novel strain of Enterobacter sp. Int J Environ Sci Technol. doi:10.1007/s13762-015-0784-3

    Google Scholar 

  • Winkler UK, Stuckmann M (1979) Glycogen, hyaluronate and some other polysaccharides greatly enhance the formation of exolipase by Serratia marcescens. J Bacteriol 138:663–670

    CAS  Google Scholar 

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Acknowledgments

The authors would like to thank the BU-DRDO Centre for Life Sciences, Coimbatore, for the SEM analysis; PSG College of Technology, Coimbatore, for the FTIR analysis; Department of Chemistry, IITM, Chennai, for TGA and DSC analyses; and SAIF-IITM, Chennai, for the NMR analyses.

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Authors and Affiliations

  1. Department of Biotechnology, Kumaraguru College of Technology, Coimbatore, 641049, India

    P. Kanmani, K. Kumaresan, J. Aravind & R. Balan

  2. School of Biosciences and Technology, VIT University, Vellore, 632014, India

    S. Karthikeyan

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  1. P. Kanmani
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  2. K. Kumaresan
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  3. J. Aravind
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  4. S. Karthikeyan
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  5. R. Balan
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Correspondence to J. Aravind.

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Kanmani, P., Kumaresan, K., Aravind, J. et al. Enzymatic degradation of polyhydroxyalkanoate using lipase from Bacillus subtilis . Int. J. Environ. Sci. Technol. 13, 1541–1552 (2016). https://doi.org/10.1007/s13762-016-0992-5

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  • DOI: https://doi.org/10.1007/s13762-016-0992-5

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