Breeding And Efficient Synthesis Of Aerobic Methane Oxidizing Bacteria Research On Polyhydroxyalkanoates

Polyhydroxyalkanoates(PHAs)have potential application prospects for replacing traditional plastic materials.Methane oxidizing bacteria can accumulate PHB in times of unbalanced nutrient supply,and the theoretical synthetic yield of PHB is 67%.In recent years,the demand for PHB products has expanded from initial packaging material applications to biomedical and pharmaceutical sectors,based on their biodegradable and biocompatible properties.Most of the research on ph HB synthesis by methane oxidizing bacteria is fermented using mixed bacteria,while there are fewer studies on PHB production from pure bacteria.Therefore,this paper explores the regulation strategy of isolating pure bacteria from the methane oxide bacteria obtained in the laboratory with methane as the synthesis of PHB by the pure bacteria.This paper performed further isolation based on the enrichment of mixed methane oxidation flora in the laboratory.An aerobic culture based on Hungate method,sequenced after multiple rounds of continuous isolation,identified pure bacteria and named Pseudacidovorax intermedius PHB-1.The growth amount was determined and the corresponding growth curve of the pure bacteria was drawn,the PHB synthesis capacity was determined,and the next PHB synthesis conditions were optimized.To maximize PHB production,optimum methane oxygen ratio is 3:2;optimal copper ion concentration is 35 u mol/L;optimum pH is 7.5;and optimum temperature is 35℃.Nitrate culture effectively improves the synthetic PHB capacity of PHB-1,while ammonia nitrogen culture obviously inhibited PHB synthesis;During the eight nitrate-nitrogen cycle culture,the maximum PHB synthesis of PHB-1 fluctuated around 27.5%,which is slightly higher than PHB-1 cultured with nitrogen as the only nitrogen source(26.37%).It is shown that the addition of a certain amount of nitrate in a short period of time is beneficial to the PHB synthesis of PHB-1;However,during the ammonia nitrogen-nitrogen cycle culture,the peak PHB yield of PHB-1 fluctuated around 23%,which was slightly lower than that of the synthetic PHB of the strains that used nitrogen as a nitrogen source.This indicates that when THE ammonia nitrogen-nitrogen cycle culture of PHB-1 is carried out,the ability of bacteria to synthesize PHB is reduced,and the growth rate of PHB yield is slowed down,which may require a certain buffer adaptation time.In the synthetic PHB stage,the co-substrates most suitable for PHB synthesis are0.2 mmol/L sodium citrate and 0.2 mmol/L acetic acid;After the addition of sodium citrate and sodium lactate,PHB-1 will form 3-HV and 3-HB copolymers;The amount of methanol added as an important intermediate product in the carbon cycle of the strain had a significant effect on the maximum synthesis of PHB and the growth rate of the strain.In the continuous synthesis process of SBR reactor,so the culture conditions with relatively high starvation time ratio are more beneficial to PHB output;the cosubstrate of PHB synthesis capacity is 0.2 mmol/L acetic acid(47.12%);After nitrate culture,the PHB synthesis capacity(28.62%)is relatively stable,effectively increasing PHB production.Therefore,the continuous yield of PHB synthesis under all conditions was higher than the PHB yield of the medium strain in the noncontinuous synthetic state.