SCREENING FOR POTENTIAL POLYHYDROXALKANOATES PRODUCING BACTERIA FROM DIFFERENT WASTE SOURCES IN PAKISTAN

Abstract

Polyhydroxyalkanoates (PHAs) are energy and carbon reserve materials stored in the cytoplasm of certain bacteria under stress conditions. PHAs are regarded as biodegradable, biocompatible and environment friendly bioplastics. This study was aimed to isolate bioplastic producers from a variety of environmental samples of Pakistan, grow them on different carbon sources and characterize the bioplastic thus produced. A total of twenty five isolates were obtained from oil/diesel containing soil, cafe waste water, municipal water, garden soil, and waste from paper pulp industry. Out of them only three isolates showed black-blue colouration when stained with Sudan black B, a preliminary screening agent for lipophilic compounds, and positive result with Nile blue A staining, a more specific dye for PHA granules. To confirm the genetic basis for PHA production PhaC gene (540 bp bands) was amplified using specific primers. Moreover, 16S rRNA gene sequence analysis confirmed the isolate with highest productivity as Pseudomonas putida DLL-E4. PHA production over course of time was studied using three different carbon sources i.e. glucose, sucrose and sunflower seed oil. Results indicated highest PHA yield with glucose 10.4 mg/L at 60 hrs, sucrose yielded 9 mg/L at 40-60 hrs and lowest yield 4 mg/L at 40 hrs was obtained with sunflower seed oil. Extracted polymer was characterized by X- ray diffraction and FTIR analysis revealed amorphous nature of PHA. Polymer was tested for pH stability was checked and found to be stable at pH 3 for 3 minutes.