Surfactant Assisted Extraction of Polyhydroxyalkanoate Sunthesized by Cupriavidus necator

Abstract

Polyhydroxyalkanoates or Bioplastics, are synthesized as reserve food source by microbes under nutrient depleted condition and in the presence of surplus carbon source. Cupriavidus necator, a gram-negative soil microbe, was used to valorize crude glycerol derived from biodiesel plant into PHA in an anaerobic and unsterile mode of batch fermentation. Medium optimization studies were performed by one variable at a time approach and it was found that maximum PHA production of 11.96 g/L, was achieved at optimized medium composition of 9 wt% crude glycerol, 0.75 g/L ammonium sulphate, 2 g/L sodium bicarbonate, 7.5 mM copper sulphate & 4 mM magnesium sulphate, pH adjusted to 6.8 and incubated at 30oC, 150 rpm for 72 hours. NMR & FTIR based structural characterization revealed that the PHA synthesized is a novel long chain length terpolymer and is named as Poly [3-Hydroxybutyrate-co-3- Hydroxyvalerate-co-3-hydroxy 4-methoxyphenyl valerate] (P3HB-co-HV-co- MeOPhHV). Novel PHA synthesized has glass transition temperature of -14.34oC, melting temperature of 104.85oC while the onset of degradation was found to be at 250.64oC. Average molecular weight of the polymer was found to be 994 KDa while the average molecular number is 615 KDa and polydispersity index is 1.616. Different micellar extraction based purification techniques were developed to separate and purify synthesized PHA from biomass present within the crude fermentation broth. The maximum purity of 92.49% was obtained by performing nonionic surfactant based cloud point extraction during batch studies, continuous cloud point extraction in RDC, revealed that variation in operational variables such as flow rate and rotor speed reduced the overall purity to 88%. low frequency sonic waves assisted cloud point extraction increased the PHA purity to 94.28%. Gum arabic – mixed surfactant coacervate complex assisted microfiltration of PHA resulted in a purity of 97.08 % Maximum separation factor of 3.045 was achieved when crude broth was run in reverse phase C18 column with TX100+methanol based hybrid micellar mobile phase