Research On Polyhydroxyalkanoate Synthesized With Activated Sludge Mixed Bacterial

Polyhydroxyalkanoates (PHA) are a class of polymer which can be synthesized by many species of microorganisms under imbalanced nutrient conditions, PHA accumulated in the microbial metabolism plays a carbon source, energy, adjust intracellular ion balance. PHA's physical characteristics are similar to chemical plastics such as polypropylene, and no harm to environment, PHA can be completely degraded in the natural condition, it is expected to replace chemical plastic, and gives good potential alternatives to completely solve the white pollution further. Currently, there are many companies using pure bacteria and pure culture fermentation to produce commercial PHA, but the high cost prevented the large-scale application of PHA.To reduce PHA production costs, a approach is use of mixed bacteria, such as activated sludge instead of pure bacteria, using organic waste, food waste instead of glucose, fructose, etc. as the substrate, can greatly reduce the cost of PHA production, but research shows that, PHA could synthesized by too many microorganisms, not all strains are suitable for mixed culture product PHA , some PHA synthesis process is relatively complicated, not easy to operate and control the conditions, the domestication time and efficiency was not satisfactory. The purpose of this research topic is to acclimate excess sludge synthesize PHA by low molecular fatty acid, find a simple and feasible for PHA synthesis methods, simplify procedures and steps, while maximizing the yield of PHA.With acetate as sole carbon source, sludge or excess sludge from urban sewage treatment plants secondary settling tank is inoculated to reactor, study the way of different start and acclimated sludge to synthesize PHA. The results found that, running first by anaerobic - aerobic for 7-15 days, the activity of the sludge was restored, then the ratio of COD: N or COD: P, was gradually increased, N or P element concentration reduced on a certain gradient (10% -20%), after 10-15 days of acclimation, PAOs or GAOs which Synthesize PHA under anaerobic conditions were gained. when the first starting phase changed to complete aerobic condition for 7-10 days,and then anaerobic anoxic condition for acclimation, the concentration of N or P was reduced meantime, after 7-10 days, PHA synthesized under aerobic conditions in activated sludge was gained. In terms of a single carbon source, through a different start, domestication of control was achieved; PHA synthesis of two different strains could be control and selection. In the basis of domestication, using acetate as sole carbon source, the ratio of nutrients (COD:N, COD:P) on PHA synthesis were adjusted, the results show that with carbon-nitrogen or carbon-phosphorus ratios increased, PHA yield increased, too. The proportion of both nutrients stimulate microbial synthesis of PHA, and also affects the maximum PHA yield of cells; for the synthesis of PHA under anaerobic conditions in the sludge,of the COD:N=125:1 and COD:P=250:1, the maximum PHA accumulations in cell dry weight were 57% and 26% respectively; PHA synthesized under aerobic conditions, the maximum amount of PHA synthesis of cell dry weight in COD:N=200:1 and COD:P=500 were 61% and 33% respectively. Limiting nitrogen source was batter to the PHA yield than limiting the phosphorus content. The experiment also found that under anaerobic conditions the constituted of the PHA are PHB and PHV, PHV PHA proportion accounted for 10% to 20%; PHA synthesized under aerobic conditions is almost PHB. Both PHA production reactors appears sludge bulking after about 35 days of operation, when phosphorus is restricted, non-filamentous sludge bulking was founded, when nitrogen source is restricted, filamentous sludge bulking occurred, then the PHA synthesis capacity lost.Experiment with different temperatures and different pH conditions was studied, the synthesis of PHA under the conditions of phosphorus limited shows a maximum yield in 19℃; while in N-limited conditions, the PHA yielded max at 16℃. No matter what nutrients are limited, it indicated that a lower temperature is suitable for PHA synthesis. In pH affect study, without control of pH conditions, pH ranged of 8 ~ 9, PHA synthesis is better, when pH is lowered to 7.0 substrate utilization capacity and PHA synthesis decreased, when pH is lowered to 5.0, the synthesis of PHA capacity significantly reduced. Neutral and alkaline environment is conducive to PHA synthesis.In order to simulate the real wastewater contained molecule organic acid, use brown sugar acidification wastewater as a carbon source to synthesize PHA, adjust the pH of fatty acid wastewater and then import to reactor, the experiment showed that not only can the rich carbon source to increase the total amount of PHA, but also makes the PHA monomer composition changes, PHV ratio was dramatically increased; if the water without pH adjustment import into the PHA synthesis reactor directly, it will have some impact on the synthesis of Panther substrate utilization rate and the maximum PHA production rate decreased by 46% and 58%, respectively; analysis from the PHA monomer product, PHB yield is affected greater, the efficiency in the use of substrate is decreased, either. When adjusting the proportion of nutrient ratio of fatty acid wastewater to make a condition that carbon is excess to nitrogen or phosphorus, PHA accumulation will get a promotion, the proportion of substrate for PHA synthesis increased by more than 100%. An unbalanced diet is to stimulate PHA accumulation ratio is still the best external factors, partially offset by the adverse effects of lower pH.Base on the PHA synthesis study, using 5% NaClO on 2800mg/L of sludge can be broken to achieve better results. Designed a set including the purge evaporation, solvent absorption, emission security under laboratory conditions bottle PHA extraction apparatus, using chloroform dissolved PHA, using NMP absorbed chloroform, synchronization of the PHA precipitation and chloroform solvent recovery; The PHA extraction was white flakes or powder, the impurities was residual fragments of the activated sludge, for about 20% dry cell weight of the PHA in activated sludge, after extraction, the product obtained over 60-70% in purity.After the pilot production of domesticated PHA, screening a strain with a high PHA synthesis, named as S1, its fermentation experiments showed that the strain could accumulated PHA at a higher ratio of carbon to phosphorus, carbon-phosphorus ratio is the main factors affecting the PHA yield, which is consistent with reactor acclimation results; batch fermentation experiments showed that, in the COD:P=400, the cell dry weight reached 4.78g/L at the same time the accumulation of PHA also reached to 51.2%. Cell growth and PHA accumulation was simultaneous. 16s rDNA sequencing showed that, S1 bacteria was closely related to Thauera sp.