Analysis of polyhydroxyalkanoates granuales in bacillus Sp. MFD11 and enterobacter Sp. SEL2

Bacillus sp. MFD11 (JF901809) and Enterobacter sp. SEL2 (JF901810) were isolated from agriculture waste contaminated sites. When fed with 2% glucose as a carbon source, these bacteria produced 75.26±0.45% and 76.61±0.28% PHA of their wet weight respectively. The accumulated PHA was extracted by direct addition of sodium dodysyl sulphate in the culture medium, which yielded 52.3±0.56 micro g/l and 136.21±0.45 micro g/l PHA respectively when assayed with Crotonic acid. The PHA detection medium (PDM) provided nutrient limitation condition which favored accumulation of PHA granules. A tremendous increase in cell size was observed when strain MFD11 was grown in PDM. The size of the granules as revealed by TEM micrographs spanned from 0.1 to 1.5 micro m which is quite large as compared to the size reported in the literature 0.2 to 0.5 micro m 18). (PHA polymer was analyzed by FTIR, GC/MS and proton Nuclear magnetic resonance. The intense absorption band in the spectrum at 1724-1740 cm -1 and 1215 cm -1 to 1280 corresponding to C=O and C-O stretching group, respectively, indicated that the both strains were PHA producers. GC/MS analysis indicated that the polymer produced were copolymers of PHB-co-PHV. NMR also suggested that the extracted PHA was not a homopolymer but was the blend of copolymers with 3HV in lower abundance. Differential calorimetric thermal analysis showed melting temperature of 163 and 169 degree C for PHA produced by both strains, respectively. However, the observed melting temperature was found to be lower than the standard PHB (Signa-aldrich). (author)