Study On Synthesis Of PHA By Using Hydrolysis Acidification Solution Of Food Waste As Carbon Source

White pollution was caused by undegradable petroleum-based plastics,so it is urgent to develop biodegradable plastics that can replace traditional plastics.Polyhydroxyalkanoate(PHA)can be synthesized by microbial fermentation and is a biodegradable polymer.Currently,PHA productions are very expensive due to the expensive carbon sources.So find an inexpensive carbon source is one efficiently method to reduce expense.In this study,food waste hydrolysis acidification liquid was selected as an inexpensive carbon source for microbial synthesis of PHA.And,a high-yield salt-tolerant strain synthesizing PHA using food waste hydrolysis acidification solution as a carbon source was screened from high salinity kitchen waste leachate treatment system.Different volatile fatty acids(VFA)were used as carbon sources,and the experimental conditions were optimized to establish a fermentation kinetic model.Finally,the mechanism of synthesis of PHA based on metabolomic bacterial cells was investigated and potential metabolic markers were identified.This study can provide a basis for optimizing the PHA synthesis medium and further increasing the PHA production.The research contents and results are as follows:(1)One salt-tolerant PHA synthetic strain was screened and identified.This strain was selected from the sewage leachate treatment system according to the synthetic PHA yield.Furthermore,the growth curve of the strain was determined by turbidity method.The results showed that the lag phase was between 0 to 2 h,logarithmic phase was between 2 to8 h,stable phase was between 8 to 16 h,and a decay period was after the 16_thhours.16S r DNA molecular biotechnology was used and showed that the strain belonged to Bacillus cereus and the strain name was Bacillus cereus strain HY-3.(2)The fermentation conditions of PHA synthetic bacteria were optimized.The effects of different fermentation conditions on the synthesis of PHA by PHA synthetic bacteria,such as salt content,inoculum amount,carbon to nitrogen ratio,fermentation time and initial p H value,were studied.The optimal fermentation process conditions were finally determined.The results showed that when the salt content was 0.5%,the inoculum amount was 3%,the carbon to nitrogen ratio was 20:0.5,the fermentation time was 48h,and the initial p H value was 7.PHA yield was up to 165.2 mg/L under this optimal fermentation conditions.(3)The ability and kinetics of synthesis of PHA by salt-tolerant bacteria with different VFAs carbon source were studied.Firstly,the strain HY-3 was cultured with acetic acid,propionic acid,butyric acid,and valeric acid as carbon sources,respectively.According to cell growth and product synthesis,the synthesis of PHA by acetic acid was more better.Then,the strain HY-3 was cultured with the simulated mixed acid and the actual mixed acid as the carbon source,and then the synthesis of PHA was carried out,and the fermentation kinetic model of the simulated mixed acid system was established.There were no difference between them.The cell growth and carbon source consumption were the same under the two carbon sources,indicating that the experiment with simulated acid as carbon source has a guiding effect on the actual VFA as fermentation carbon source.The results showed that the model had a good fitting effect and could predict the experimental data,optimize the fermentation process.And the large-scale continuous fermentation could use it.Model-related parameters indicated that the addition of substrate during the pre-fermentation phase would promote cell growth and increase product yield.(4)Based on metabolomics,the synthesis mechanism of PHA was studied.The strain HY-3 was cultured at a concentration of 0.5 g/L and 5.0 g/L using acetic acid as a carbon source.The metabolites of the log phase,stationary phase and decay phase of the strain were analyzed by gas chromatography-mass spectrometry(GC-MS).The data were analyzed using principal component analysis(PCA),metabolite load method(PLS-DA),metabolic pathway effects,and microbial metabolic network analysis.metabolites had significant metabolic differences at different growth stages and different carbon source concentrations.Trihydroxybutyrate(3-HB),threonine,serine,malic acid,and pyruvate were potential metabolic markers that could affect PHB synthesis.