ISOLATION AND CHARACTERIZATION POLYHYDROBUTYRATE (PHB) PRODUCING BACTERIA FROM WASTE COOKING OIL USING POMEGRANATE MOLASSES AS CARBON SOURCE

Abstract

In this study, a polyhydroxybutyrate (PHB) producing bacterium was isolated from waste cooking oil and characterized for its morphological and biochemical properties. Staining methods utilizing Sudan Black B and Nile Blue A were used on isolated bacterium to demonstrate good capability for synthesizing PHB. It was shown that the isolated bacterium species was related to Bacillus thuringiensis LMA by using 16S rRNA gene sequences analysis. During the stationary phase, the Bacillus strain was subjected to 10 % (w/v) of pomegranate molasses as a carbon source and 5 g/L of peptone as a nitrogen source. 2 ml of batch fermentation was collected. Samples were collected twice during the incubation period for detection of PHB using Sudan Black B. The PHB production accounted for up to 57.45% of the cell dry weight. The PHB produced was characterized using Fourier Transform Infrared Spectroscopy (FTIR) and Nuclear Magnetic Resonance Spectroscopy (NMR). The drastic absorption band at approximately 1717 cm-1 indicated the stretching vibration of C=O group in PHB polyester, while the functional groups of PHB were identified methyl (-CH3) at 1.28 ppm, methylene (-CH2) 2.0 and 2.5 ppm, and methylene doublet group (CH3) at 5.3 ppm.