title: Validation of thermally assisted hydrolysis and methylation-gas chromatography for rapid and direct compositional analysis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) in whole bacterial cells.
abstract: Thermally assisted hydrolysis and methylation-gas chromatography (THM-GC) in the presence of an organic alkali was validated for the compositional analysis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] accumulated in whole bacterial cells. Recombinant Cupriavidus necator Re2058/pCB113 was grown in a batch fermentation with different concentration of palm oil and fructose in order to control the molar fraction of 3HHx in P(3HB-co-3HHx) produced in the cells. Trace amounts (30μg) of freeze-dried cells were directly subjected to THM-GC in the presence of tetramethylammonium hydroxide (TMAH) at 400°C. The obtained chromatograms clearly showed nine characteristic peaks, attributed to the THM products from 3HB and 3HHx units in the polymer chains, without any appreciable interference by the bacterial matrix components. Based on these peak intensities, the copolymer compositions were determined rapidly without using any cumbersome and lengthy sample pretreatment as in conventional GC method. Moreover, the compositions thus obtained were strongly correlated with those by NMR and conventional GC involving solvent extraction.
authors: School of Industrial Technology, Universiti Sains Malaysia, 11800, Minden, Penang, Malaysia. Electronic address: sitibaidurah@usm.my.
authors: Siti
authors: S
authors: Baidurah
authors: School of Biological Sciences, Universiti Sains Malaysia, 11800, Minden, Penang, Malaysia.
authors: Paramasivam
authors: P
authors: Murugan
authors: School of Biological Sciences, Universiti Sains Malaysia, 11800, Minden, Penang, Malaysia.
authors: Lee
authors: L
authors: Joyyi
authors: Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan.
authors: Junya
authors: J
authors: Fukuda
authors: Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan.
authors: Masaya
authors: M
authors: Yamada
authors: School of Biological Sciences, Universiti Sains Malaysia, 11800, Minden, Penang, Malaysia.
authors: Kumar
authors: K
authors: Sudesh
authors: Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan.
authors: Yasuyuki
authors: Y
authors: Ishida
mesh_terms: D020155:3-Hydroxybutyric Acid / Q000737:chemistry; D002208:Caproates / Q000032:analysis* / Q000737:chemistry; D002623:Chemistry Techniques, Analytical / Q000295:instrumentation / Q000379:methods*; D002849:Chromatography, Gas; D020118:Cupriavidus necator / Q000737:chemistry*; D005285:Fermentation; D006358:Hot Temperature; D006868:Hydrolysis; D009682:Magnetic Resonance Spectroscopy; D008745:Methylation; D000073878:Palm Oil; D010938:Plant Oils / Q000378:metabolism; D011108:Polymers; D000094122:Polyhydroxybutyrates
chemical_list: D020155:3-Hydroxybutyric Acid; D002208:Caproates; D000073878:Palm Oil; D010938:Plant Oils; D011108:Polymers; C115940:poly(3-hydroxybutyrate-co-3-hydroxyhexanoate); D000094122:Polyhydroxybutyrates; C045051:3-hydroxyhexanoic acid
keywords: Biodegradable polymer; Copolymer composition; Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate); Tetramethylammonium hydroxide; Thermally assisted hydrolysis and methylation-GC; Validation