Journal
BIOTECHNOLOGY AND BIOPROCESS ENGINEERING
Volume 14, Issue 4, Pages 482-489Publisher
KOREAN SOC BIOTECHNOLOGY & BIOENGINEERING
DOI: 10.1007/s12257-008-0119-z
Keywords
polyhydroxyalkanoate isolation; Bacillus flexus; Microbispora sp.; protease; aqueous two-phase system; PEG/Phosphate
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Funding
- Council of Scientific and Industrial Research (CSIR), New Delhi, India
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Main objective of present work was to isolate polyhydroxyalkanoate (PHA) from cell lysate of Bacillus flexus by aqueous-aqueous two-phase system (ATPS). Selected ATPS having polyethylene glycol (12%, w/v) and potassium phosphate (9.7%, pH 8.0) containing cell lysate obtained by sonication or hypochlorite treatment of B. flexus biomass (1 g%, dry weight), was held at 28 degrees C for 30 min, which partitioned PHA into top PEG phase and residual cell materials into bottom phase. For enzymatic cell hydrolysis, Microbispora sp. culture filtrate having protease (3 U/mL) was mixed with B. flexus biomass and ATPS, incubated at 37 degrees C for 2 h prior to phase separation. PHA recovered by centrifugation was 19 similar to 51% of cell dry weight, depending on the mode of cell disruption. Protease was recovered along with PHA in the PEG phase and showed 7 fold increase in activity. PHA was characterized by GC, FTIR, and H-1 NMR. Results indicated that ATPS can be used for the isolation of PHA from hydrolyzed bacterial cells and purified protease can be recovered as a by-product, in a single defined experiment. Results have indicated that ATPS can be successfully employed as a non-organic solvent method for the isolation of PHA. (C) KSBB
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