| There has been considerable in the development and production of biodegradable polymer to solve the current problem of pollution caused by the continuous use of synthetic polymer of petroleum origin.Poly-β-hydroxybutyrate(PHB) are known to be accumulated as intracellular inclusion in some bacteria,which is a close substitute for the synthetic plastic.The high cost of PHB production has restricted its application.This study investigates the PHB storage behavior of a poly-phosphate accumulating organism(PAO) under different growth conditions in detail.Pure culture studies in batch-growth systems were conducted in shake-flasks,using chemically defined growth media with acetate as the sole carbon source.This batch-growth system with anaerobic-aerobic cycles and varying conditions,similar to enhanced biological phosphorus removal processes(EBPR), was also employed.This paper also highly reports the advances of PHB extraction methods from cell and the PHB analytical methods.Therefore,it is a new way for PHB synthesis,which also develops a novel pathway for the reclamation of residual sludge.The results of this research can be summarized as follows:(1) Firstly,the enrichment and cultivation of poly-phosphate accumulating activated sludge were studied in a set of sequencing batch reactor(SBR) system,under A/O condition. 30 days later,the both removal rates of ammonium and phosphate were higher than 80%.Then, separation 36 strains were isolated from the activated sludge.One of them with high levels of phosphate uptake and release activities,combined poly-P and PHB granules microscopic staining,was confirmed and named LB4.Based on the morphological appearances,physiological and biochemical characteristics, together with analysis of 16S rDNA sequence,the activated-sludge bacterium(LB4) were identified as Acinetobacter baumannii.(2) In this part of the study,the effects of initial pH and temperature on the storage metabolism of A.baumannii,under repeated anaerobic-aerobic cycles on the batch growth, were investigated.Acinetobacter baumannii LB4 was acclimated for a long term with higher initial pH could store more PHB and release more phosphorus in the anaerobic phase,and degraded more PHB and accumulate more polyphosphate in the following aerobic phase.At the same time,the initial metabolism rate of acetate,phosphorus and poly-β-hydroxybutyrate would be faster.The acetate-utilization and PHB-storage behavior of Acinetobacter baumannii LB4 acclimated for a long term with higher temperature was similar to those in higher initial pH. Result also shows that,LB4's PHB storage capability was could be described with the PHB content of the cells with respect to was closely related to phosphorus removal amount.Effects of carbon concentration and nutrient deficiency on the PHB of the batch-growth systems are studied.With 673.6 mg·L-1,1355.4 mg·L-1,2694.6 mg·L-1 and 5393.3 mg·L-1 initial sodium acetate,the maximum PHB content of dry biomass were 12.91%,20.83%, 27.60%and 24.91%respectively,while carbon translating ratios were 41.2%,38.5%,33.3% and 22.0%.During the first 120 min of anaerobic phase,PHB forming rate were 175.2 mg·L-1·h-1,153.8 mg·L-1·h-1,123.9 mg·L-1·h-1 and 114.3 mg·L-1·h-1.The influence of nutrient deficiency on the storage metabolism of A.baumannii under repeated anaerobic-aerobic cycles on the batch growth was not evident.(3) Orthogonal experimental design was applied to optimize the extraction condition PHB from microorganism.The optimized parameters of extracting PHB were as follows:method of extraction strain Acinetobacter baumannii LB4:the extraction temperature 50℃,the extraction time 2 h,the volume ratio of sodium hypochlorite and chloroform 1:1,the liquid volume of chloroform 10 mL,and the dry biomass 100 mg.The extraction yield of PHB under these parameters was 33.2 mg(per 100 mg CDW).Ultraviolet spectrum,infrared spectrum,and elementary analysis were utilized for the characteristics of PHB structure. |
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