Evaluation of short-chain-length polyhydroxyalkanoate accumulation in Bacillus aryabhattai

This study was focused on the polyhydroxybutyrate (PHB) accumulation property of Bacillus aryabhattai isolated from environment. Twenty-four polyhydroxyalkanoate (PHA) producers were screened out from sixty-two environmental bacterial isolates based on Sudan Black B colony staining. Based on their PHA accumulation property, six promising isolates were further screened out. The most productive isolate PHB10 was identified as B. aryabhattai PHB10. The polymer production maxima were 3.264 g/L, 2.181 g/L, 1.47 g/L, 1.742 g/L and 1.786 g/L in glucose, fructose, maltose, starch and glycerol respectively. The bacterial culture reached its stationary and declining phases at 18 h and 21 h respectively and indicated growth-associated PHB production. Nuclear Magnetic Resonance (NMR) spectra confirmed the material as PHB. The material has thermal stability between 30 and 140 °C, melting point at 170 °C and maximum thermal degradation at 287 °C. The molecular weight and poly dispersion index of the polymer were found as 199.7 kDa and 2.67 respectively. The bacterium B. aryabhattai accumulating PHB up to 75% of cell dry mass utilizing various carbon sources is a potential candidate for large scale production of bacterial polyhydroxybutyrate.(AU)

Variation analysis of bacterial polyhydroxyalkanoates production using saturated and unsaturated hydrocarbons

ABSTRACT Polyhydroxyalkanoates (PHA) are efficient, renewable and environment friendly polymeric esters. These polymers are synthesized by a variety of microbes under stress conditions. This study was carried out to check the suitability of waste frying oil in comparison to other oils for economical bioplastic production. Six bacterial strains were isolated and identified as Bacillus cereus (KF270349), Klebsiella pneumoniae (KF270350), Bacillus subtilis (KF270351), Brevibacterium halotolerance (KF270352), Pseudomonas aeruginosa (KF270353), and Stenotrophomonas rhizoposid (KF270354) by ribotyping. All strains were PHA producers so were selected for PHA synthesis using four different carbon sources, i.e., waste frying oil, canola oil, diesel and glucose. Extraction of PHA was carried out using sodium hypochlorite method and maximum amount was detected after 72 h in all cases. P. aeruginosa led to maximum PHA production after 72 h at 37 °C and 100 rpm using waste frying oil that was 53.2% PHA in comparison with glucose 37.8% and cooking oil 34.4%. B. cereus produced 40% PHA using glucose as carbon source which was high when compared against other strains. A significantly lesser amount of PHA was recorded with diesel as a carbon source for all strains. Sharp Infrared peaks around 1740-1750 cm-1 were present in Fourier Transform Infrared spectra that correspond to exact position for PHA. The use of waste oils and production of poly-3hydroxybutyrate-co-3hydroxyvalerate (3HB-co-3HV) by strains used in this study is a good aspect to consider for future prospects as this type of polymer has better properties as compared to PHBs.(AU)

Optimization of biodegradable plastic production on sugar cane molasses in Enterobacter sp. SEL2

Contaminated environments have a large number of bacteria which can accumulate PHA as their energy reserves. Out of 54 isolated bacterial strains from three groups of contaminated sites 48 were found PHA positive. The sites were grouped on the basis of the type of carbon sources i.e. sugars, fatty acids and much diverse type. Strains MFD5, MFD11, UML3, USL2, SEL2, SEL3, SEL10 and PFW1 produced 69.9 ± 0.29, 75.27 ± 0.45, 65.43 ± 0.1, 72.54 ± 0.27, 76.61 ± 0.28, 61.81 ± 0.05, 71.16 ± 0.09 and 74.92 ± 0.5 percent of PHA to their constant cell weight (CCW) respectively in PHA detection media supplemented with 2% glucose. Molasses, whey, crumbs hydrolysate and palm oil were checked as inexpensive carbon sources. Molasses alone could supply the required nutrients for growth and PHA production. Strain SEL2 produced 47.36 ± 0.45% PHA using 2% molasses at 37 °C and pH 7.0. Upon production optimization the best accumulation (80.95 ± 0.01%) was observed in PHA detection media with 0.2% nitrogen source, 3% molasses, pH 5.0 and 37 °C by the strain SEL2. The overall effect of the presence of increased molasses concentration in the media was positive it increased the accumulation period till 72 h. Enterobacter sp. SEL2 (JF901810) is first time being reported for PHA production.

Growth kinetics, effect of carbon substrate in biosynthesis of mcl-PHA by Pseudomonas putida Bet001

Growth associated biosynthesis of medium chain length poly-3-hydroxyalkanoates (mcl-PHA) in Pseudomonas putida Bet001 isolated from palm oil mill effluent was studied. Models with substrate inhibition terms described well the kinetics of its growth. Selected fatty acids (C8:0 to C18:1) and ammonium were used as carbon and nitrogen sources during growth and PHA biosynthesis, resulting in PHA accumulation of about 50 to 69% (w/w) and PHA yields ranging from 10.12 g L-1 to 15.45 g L-1, respectively. The monomer composition of the PHA ranges from C4 to C14, and was strongly influenced by the type of carbon substrate fed. Interestingly, an odd carbon chain length (C7) monomer was also detected when C18:1 was fed. Polymer showed melting temperature (Tm) of 42.0 (± 0.2) °C, glass transition temperature (Tg) of -1.0 (± 0.2) °C and endothermic melting enthalpy of fusion (ΔHf) of 110.3 (± 0.1) J g-1. The molecular weight (Mw) range of the polymer was relatively narrow between 55 to 77 kDa.

Volatile fatty acids influence on the structure of microbial communities producing PHAs

Polyhydroxyalkanoates (PHAs) can be produced by microorganisms and are a biodegradable alternative to fossil-fuel based plastics. Currently, the focus is on reducing production costs by exploring alternative substrates for PHAs production, and on producing copolymers which are less brittle than monomers. Accordingly, this study used a substrate consisting of wastewater from waste-glycerol fermentation, supplemented with different amounts of acetic and propionic acids. These substrates were used to feed mixed microbial communities enriched from activated sludge in a sequencing batch reactor. A reactor supplemented with 2 mL of acetic acid produced 227.8 mg/L of a homopolymer of hydroxybutyrate (3HB); 4 mL of acetic acid produced 279.8 mg/L 3HB; whereas 4 mL of propionic acid produced 673.0 mg/L of a copolymer of 3HB and 3HV (hydroxyvalerate). Ribosomal Intergenic Spacer Analysis (RISA) was used to show the differences between the communities created in the reactors. Thauera species predominated in biomass that produced 3HB; Paracoccus denitrificans in the biomass that produced 3HB-co-3HV. Because P. denitrificans produced the more desirable copolymer, it may be advantageous to promote its growth in PHAs-producing reactors by adding propionate.

Semi-scale production of PHAs from waste frying oil by Pseudomonas fluorescens S48

The present study aimed at developing a strategy to improve the volumetric production of PHAs by Pseudomonas fluorescens S48 using waste frying oil (WFO) as the sole carbon source. For this purpose, several cultivations were set up to steadily improve nutrients supply to attain high cell density and high biopolymer productivity. The production of PHAs was examined in a 14 L bioreactor as one-stage batch, two-stage batch, and high-cell-density fed-batch cultures. The highest value of polymer content in one-stage bioreactor was obtained after 60 h (33.7%). Whereas, the two-stage batch culture increased the polymer content to 50.1% after 54 h. High-cell-density (0.64 g/L) at continuous feeding rate 0.55 mL/l/h of WFO recorded the highest polymer content after 54 h (55.34%). Semi-scale application (10 L working volume) increased the polymer content in one-stage batch, two-stage batch and high cell density fed-batch cultures by about 12.3%, 5.8% and 11.3%, respectively, as compared with that obtained in 2 L fermentation culture. Six different methods for biopolymer extraction were done to investigate their efficiency for optimum polymer recovery. The maximum efficiency of solvent recovery of PHA was attained by chloroform-hypochlorite dispersion extraction. Gas chromatography (GC) analysis of biopolymer produced by Pseudomonas fluorescens S48 indicated that it solely composed of 3-hydrobutyric acid (98.7%). A bioplastic film was prepared from the obtained PHB. The isolate studied shares the same identical sequence, which is nearly the complete 16S rRNA gene. The identity of this sequence to the closest pseudomonads strains is about 98-99%. It was probably closely related to support another meaningful parsiomony analysis and construction of a phylogenetic tree. The isolate is so close to Egyptian strain named EG 639838.(AU)