Study On The Polyhydroxyalkanoate (PHA)production By Mixed Culture Using Crude Glycerol As Substrate

Waste oil,besides volatile organic acid(VFA)and methane gas,can be generated during the recycle of kitchen waste,which can be collected for the production of biodiesel through ester exchange reaction.However,large amount of crude glycerol is produced at the same time.Therefore,the utilization of crude glycerol is the critical unit for the proper development of the kitchen waste recycling.Polyhydroxyalkanoate(PHA)is one of the most promising products with great recycle potential,based on the new concept of wastewater treatment.To explore the feasibility of PHA synthesis from crude glycerol,two sequencing batch reactors(SBRs)with different inoculums were used to enrich PHAproducing mixed cultures.Efficiency of the PHA production using crude glycerol was investigated to support the industrial operation of kitchen waste deep recycling.The 1# SBR was inoculated with the sludge from the secondary sedimentation tank of a waste water treatment plant while the 2# SBR was inoculated with the sludge from a long-running PHA enrichment reactor using VFA as substrate.Longer time,about two months,to reach the stable phase of enrichment was found to be needed by studying the changes in the physicochemical characteristics and kinetic parameters of the sludge when compared to VFA as substrate.At the beginning of the enrichment process,the sludge concentration decreased,the zoogloea began to loosen,the Zeta potential increased,the repulsion between the flocs increased,small flocs increased,and sludge settling got worse.However,when the system gradually adapted to new environment,a new structure of zoogloea formed,resulting in quickly aggregation sedimentation of the sludge.Shorter feast period length was observed in 1# SBR,with a maximum PHA yield of 25.93%,compared with 23.06% in 2# SBR.The specific PHA storage rate were 0.27 mg·mg-1·h-1 and 0.21 mg·mg-1·h-1 in 1# and 2# SBR,respectively.These results indicated that the inoculated sludge from the secondary sedimentation tank of the wastewater treatment plant had better performance on the enrichment of PHA-producing cultures when fed with crude glycerol.To explore the effect of organic loading rate(OLR)on the enrichment process,three gradients of OLR(1200,2000 and 2800 mg COD·L-1·d-1)were set gradually in one SBR.The results showed that the substrate uptake rate was low,which ultimately lowered PHA productivity under low OLR.However,under high OLR,the Feast/Famine ratio(F/F)increased,causing the decrease of selective pressure because the non-producing PHA bacteria would use excess carbon source for growth.The stable system performed best under 2000 mg COD·L-1·d-1 both in 1# and 2# SBR,with maximum PHA yields of 36.59% and 36.33% in batch assays,respectively.Meantime,PHA productivity reached the highest,which was 0.35 g·g-1 glycerol in 1# SBR and 0.34 g·g-1 glycerol in 2# SBR.The structure of the microorganisms changed under different OLRs.The OLR affected the species diversity,the relative abundance of bacteria and the succession of dominant bacteria.There were 101 and 130 species survived under all three OLRs in 1# SBR and 2# SBR,respectively,indicating that microorganisms in 2# SBR adapted better to the changes in OLR.To further increase the PHA yield,a mixture of propionic acid and glycerol were used as substrate for PHA accumulation.Two mix ratios were set,which were 20:80 and 50:50 propionic acid versus glycerol(%,based on COD).The results showed that PHAproducing bacteria enriched with crude glycerol could also use propionic.Under the same OLR,the PHA maximum yields decreased in the order of 50:50,20:80 propionic acid versus glycerol,sole crude glycerol and sole propionic acid.The mixed carbon source had mutual promotion effect on the metabolism of glycerol and propionic acid.When the crude glycerol: propionic acid rate was 1:1(based on COD),the PHA maximum yield in 1# and 2# SBR reached 55.69% and 59.56%,respectively.When crude glycerol and propionic acid were mixed as carbon source,the synthetic PHA was composed of two polymers,HB and HV,and the HV content increased with increasing propionic acid content.In summary,the enrichment of PHA-producing bacteria using crude glycerol as substrate requires no fermentation,not only simplifies the PHA production process,significantly reduces the cost of PHA synthesis,but also adapts carbon source more widely and optimizes the PHA structure.More importantly,the resource utilization of crude glycerol is also significant for the healthy development of kitchen waste disposal project.