Microbial Synthesis of Poly (3-Hydroxybutyrate- co-4-Hydroxybutyrate) by Cupriavidus sp. USMAA1020 Isolated from Malaysian Environment (original) (raw)
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Bioresource Technology, 2008
Cupriavidus sp. USMAA1020 was isolated from Malaysian environment and able to synthesize poly(3-hydroxybutyrate-co-4-hydroxybutyrate), [P(3HB-co-4HB)] when grown on c-butyrolactone as the sole carbon source. The polyester was purified from freeze-dried cells and analyzed by nuclear magnetic resonance (NMR) spectroscopy. 1 H and 13 C NMR results confirmed the presence of 3HB and 4HB monomers. In a one-step cultivation process, P(3HB-co-4HB) accumulation by Cupriavidus sp. USMAA1020 was affected by carbon to nitrogen ratio (C/N). A two-step cultivation process accumulated P(3HB-co-4HB) copolyester with a higher 4HB fraction (53 mol%) in nitrogen-free mineral medium containing c-butyrolactone. The biosynthesis of P(3HB-co-4HB) was also achieved by using 4-hydroxybutyric acid and alkanediol as 1,4-butanediol. The composition of copolyesters varied from 32 to 51 mol% 4HB, depending on the carbon sources supplied. The copolyester produced by Cupriavidus sp. USMAA1020 has a random sequence distribution of 3-hydroxybutyrate (3HB) and 4-hydroxybutyrate (4HB) units when analyzed by nuclear magnetic resonance (NMR) spectroscopy. When c-butyrolactone was used as the sole carbon source, the 4HB fraction in copolyester increased from 25 to 60 mol% as the concentration of c-butyrolactone in the culture medium increased from 2.5 g/L to 20.0 g/L.
Applied Biochemistry and Biotechnology, 2011
Poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-4-hydroxybutyrate) [P(3HB-co-3HV-co-4HB)] terpolymer was produced using Cupriavidus sp. USMAA2-4 via one-step cultivation process through combination of various carbon sources such as 1,4-butanediol or γ-butyrolactone with either 1-pentanol, valeric acid, or 1-propanol. Oleic acid was added to increase the biomass production. The composition of 3HV and 4HB monomers were greatly affected by the concentration of 1,4-butanediol and 1-pentanol. Terpolymers with 3HV and 4HB molar fractions ranging from 2 to 41 mol.% and 5 to 31 mol.%, respectively, were produced by varying the concentration of carbon precursors. The thermal and mechanical properties of the terpolymers containing different proportions of the constituent monomers were characterized using gel permeation chromatography (GPC), DSC, and tensile machine. GPC analysis showed that the molecular weights (M w ) of the terpolymer produced were within the range of 346 to 1,710 kDa. The monomer compositions of 3HV and 4HB were also found to have great influences on the thermal and mechanical properties of the terpolymer P(3HB-co-3HV-co-4HB) produced.
Polymer Degradation and Stability, 2010
Poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-4-hydroxybutyrate) [P(3HB-co-3HV-co-4HB)] terpolymer was produced using Cupriavidus sp. USMAA2-4 via one-step cultivation process through combination of various carbon sources such as 1,4-butanediol or γ-butyrolactone with either 1-pentanol, valeric acid, or 1-propanol. Oleic acid was added to increase the biomass production. The composition of 3HV and 4HB monomers were greatly affected by the concentration of 1,4-butanediol and 1-pentanol. Terpolymers with 3HV and 4HB molar fractions ranging from 2 to 41 mol.% and 5 to 31 mol.%, respectively, were produced by varying the concentration of carbon precursors. The thermal and mechanical properties of the terpolymers containing different proportions of the constituent monomers were characterized using gel permeation chromatography (GPC), DSC, and tensile machine. GPC analysis showed that the molecular weights (M w ) of the terpolymer produced were within the range of 346 to 1,710 kDa. The monomer compositions of 3HV and 4HB were also found to have great influences on the thermal and mechanical properties of the terpolymer P(3HB-co-3HV-co-4HB) produced.
Biosynthesis and characterization of poly(?-hydroxybutyrate) produced byBacillus circulans
Polymer Bulletin, 1996
The biosynthesis and characterization of medium chain length poly-3-hydroxyalkanoates (mcl-PHA) produced by Pseudomonas putida Bet001 isolated from palm oil mill effluent was studied. The biosynthesis of mcl-PHA in this newly isolated microorganism follows a growth-associated trend. Mcl-PHA accumulation ranging from 49.7 to 68.9% on cell dry weight (CDW) basis were observed when fatty acids ranging from octanoic acid (C 8:0 ) to oleic acid (C 18:1 ) were used as sole carbon and energy source. Molecular weight of the polymer was found to be ranging from 55.7 to 77.7 kDa. Depending on the type of fatty acid used, the 1 H NMR and GCMSMS analyses of the chiral polymer showed a composition of even and odd carbon atom chain with monomer length of C4 to C14 with C8 and C10 as the principal monomers. No unsaturated monomer was detected. Thermo-chemical analyses showed the accumulated PHA to be semi-crystalline polymer with good thermal stability, having a thermal degradation temperature (T d ) of 264.6 to 318.8 (60.2) o C, melting temperature (T m ) of 43. (60.2) o C, glass transition temperature (T g ) of 21.0 (60.2) o C and apparent melting enthalpy of fusion (DH f ) of 100.9 (60.1) J g 21 .
Applied Microbiology and Biotechnology, 2011
Burkholderia sp. synthase has been shown to polymerize 3-hydroxybutyrate (3HB), 3-hydroxyvalerate, and 3-hydroxy-4-pentenoic acid monomers. This study was carried out to evaluate the ability of Burkholderia sp. USM (JCM 15050) and its transformant harboring the polyhydroxyalkanoate (PHA) synthase gene of Aeromonas caviae to incorporate the newly reported 3-hydroxy-4-methylvalerate (3H4MV) monomer. Various culture parameters such as concentrations of nutrient rich medium, fructose and 4-methylvaleric acid as well as harvesting time were manipulated to produce P(3HB-co-3H4MV) with different 3H4MV compositions. The structural properties of PHA containing 3H4MV monomer were investigated by using nuclear magnetic resonance and Fourier transform infrared spectroscopy (FTIR). The relative intensities of the bands at 1,183 and 1,228 cm −1 in the FTIR spectra enabled the rapid detection and differentiation of P(3HB-co-3H4MV) from other types of PHA. In addition, the presence of 3H4MV units in the copolymer was found to considerably lower the melting temperature and enthalpy of fusion values compared with poly(3-hydroxybutyrate) (P(3HB)). The copolymer exhibited higher thermo-degradation temperature but similar molecular weight and polydispersity compared with P(3HB).
Prospect of Synechocystis sp. PCC 6803 for synthesis of poly(3-hydroxybutyrate-co-4-hydroxybutyrate)
Algal Research, 2020
The rapidly growing use of petroleum-based plastics is contributing to severe environmental pollution, thereby putting the environment to a hard test. Research for alternative plastics is essential to substitute conventional plastics with a certain grade of biodegradability. One such potential material to replace petrochemical-based plastics is microbially originated polyhydroxyalkanoates. Among them, poly-β-hydroxybutyrate (PHB) is the most common and well-characterized member. However, studies demonstrate that the properties of PHB such as brittleness, low extension-to-break, and lack of flexibility limit its possible application whereas; copolymers have the properties which can overcome the limitations of PHB. In this study, Synechocystis sp. PCC 6803, a unicellular, non-diazotrophic cyanobacterium has been found to accumulate poly(3-hydroxybutyrate-co-4-hydroxybutyrate) copolymer under nitrogen and phosphorus-deficient conditions either using γ-butyrolactone alone or along with acetate as carbon source. The maximum copolymer accumulation i.e. 37.64% dry cell weight was recorded when the cultures were supplemented with 0.4% acetate and 0.01% γ-butyrolactone. The purity and mole fraction of the polymer was confirmed by proton nuclear magnetic resonance and fourier transform infrared spectroscopy. The material properties were assessed and found comparable with other polymers from bacteria and cyanobacteria. Thus, the test organism has the potential to be utilized for large scale biopolymer production and applications in various fields.
Biomacromolecules, 2001
A series of poly(3-hydroxybutyrate-co-4-hydroxybutyrate)s [P(3HB-co-4HB)s] with different 4HB content, biosynthesized by Ralstonia eutropha H16 with mixed carbon sources of 4-hydroxybutyric acid (4HBA) and butyric acid, were fractionated by solvent/nonsolvent fractionation into copolyester fractions with different 4HB content and narrower compositional distribution. The fractions obtained were classified into two groups, 3HB-and 4HB-rich P(3HB-co-4HB)s. The thermal properties were investigated for these fractionated copolyesters. With increasing 4HB content, the melting temperature at first decreased while 3HB content was rich, and then increased while 4HB content was rich. The glass transition temperature decreased linearly with increasing 4HB content. The 4HB-rich P(3HB-co-4HB) was found to be immiscible with the 3HBrich P(3HB-co-4HB), as two glass transitions corresponding to those of respective P(3HB-co-4HB)s were observed by DSC. It was concluded that as-produced bacterial P(3HB-co-4HB) samples used in this study should be considered as immiscible polymer blends.
Thermal degradation of microbial poly(4-hydroxybutyrate)
Macromolecules, 1994
We obtained by microbial synthesis an almost pure poly(4-hydroxybutyrate) (P(4HB)) sample (i.e., a P(3HB-co-4HB) copolymer containing only 3% of 3HB units). The thermal degradation of this material was investigated using thermogravimetry, direct pyrolysis mass spectrometry and also by preparative pyrolysis and subsequent NMR analysis of the pyrolyzate. Previous studies' on P(3HB-co-4HB) copolyesters have reported that the thermal degradation process occurs by a P-CH hydrogen transfer, similar to the well-known case of P(3HB). However, the latter work did not distinguish between the thermal degradation fate of 3HB and 4HB units. Our findings indicate that the thermal decomposition of P(4HB) yields a series of cyclic oligomers (i.e., y-butyrolactone and its higher homologs) by an intramolecular exchange mechanism. Detailed mass spectrometric and 'H-NMR evidence is presented to prove this point.
Synthesis of biodegradable polyesters by Gram negative bacterium isolated from Malaysian environment
World Journal of Microbiology & Biotechnology, 2008
A locally isolated Gram negative bacterium, Cupriavidus sp. USMAA9-39 was able to produce various types of biodegradable polyesters through a two-step cultivation process. These are copolymer poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)], copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] and terpolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-4-hydroxybutyrate) [P(3HB-co-3HV-co-4HB)]. These polymers were synthesized by this bacterium when grown with a combination of some carbon sources. The biosynthesis of P(3HB-co-4HB) was achieved by using carbon sources such as γ-butyrolactone or 1,4-butanediol or by a combination of oleic acid with either γ-butyrolactone or 1,4-butanediol. Meanwhile, poly(3-hydroxybutyrate-co-3-hydroxyvalerate) was produced using 1-pentanol or valeric acid or by a combination of oleic acid with either 1-pentanol or valeric acid. When γ-butyrolactone or 1,4-butanediol with either valeric acid or 1-pentanol were used as mixed carbon sources, P(3HB-co-3HV-co-4HB) terpolymer were produced. The presence of 3HB, 3HV or/and 4HB monomers were confirmed by gas chromatography and nuclear magnetic resonance (NMR) spectroscopy.