圆褐固氮菌G-3菌株聚羟基烷酸（PHA）的提取研究

【作者】 彭菊芳；

【导师】 董兆麟；

【作者基本信息】 西北大学 ， 微生物， 2001， 硕士

【摘要】 本论文对圆褐固氮菌G—3菌株聚羟基烷酸（PHA）的八种提取方法进行了优化研究。结果表明：（1）氯仿提取分为两种，温和搅拌法：用30倍体积的氯仿在50℃下与细胞温和作用2h，再加入30倍体积的水作用1h，得到的PHA纯度为98.7％，回收率为78％，分子量为1.4×10⁶；剧烈搅拌法：30倍体积的氯仿在20℃下先将细胞温和处理10min，再剧烈作用15min，可得纯度为99％的PHA，回收率为89.6％，分子量为1.47×10⁶；（2）NaClO法中，采用40％NaClO在20℃下处理细胞30min，所得PHA纯度为93.6％，回收率为91.1％，分子量为0.54×10⁶；（3）NaClO—CHCl₃法：30倍体积的CHCl₃和30％（V/V）NaClO共同处理细胞1h，得PHA纯度为96.5％，回收率为91％，分子量为1.15×10⁶；（4）SDS法：30g/l细胞在50℃下经7g/lSDS处理12min，得到的PHA纯度为89.2％，回收率为90％，分子量为1.04×10⁶；（5）SDS—NaClO法：30g/l细胞在50℃下先经7g/lSDS处理12min，再用30％NaClO作用3min，得PHA纯度为97.5％，回收率为87.1％，分子量为0.83×10⁶；（6）SDS—EDTA法：30g/l细胞在50℃下用7g/lSDS和10g/lEDTA共同作用15min，得PHA纯度为97.75％，回收率为89％，分子量为1.1×10⁶；（7）酸法：0.2N HCl在100℃下作用50g/l细胞1h，得到的PHA纯度为95％，回收率为90.3％，分子量为0.905×10⁶；（8）碱法：50g/l细胞在65℃下用0.15N NaOH处理1h，得PHA纯度为96.2％，回收率为92％，分子量为1.01×10⁶。对上述各方法作综合分析比较，氯仿法中，剧烈搅拌较温和搅拌大大缩短了提取时间，而且成本也有大幅下降，更适合于实验室的提取研究工作。在化学试剂法中，SDS—EDTA法操作简单，价格低廉（成本约为￥0.092/gPHA），所得PHA纯度、回收率、分子量都令人较满意，是本实验室开辟的一种新的PHA提取方法。本文还就提取得到的PHA的理化性质及加工性能作了研究，结果证明，PHA的各种技术质量指标完全符合国际上目前采用的质量标准。 另外，本论文对PHA的脱色作了初步探索，实验就物理方法和化学方法针对氯仿提取细胞后得到的PHA有机相进行了脱色处理。表明，物理脱色较化学脱色效果明显，其中活性炭脱色效果最显著。更多还原

【Abstract】 Eight methods for the recovery of polyhydroxyalkanoate(PHA) from the Azotobacter chroococcum G-3 were used to optimize extraction and digestion conditions.Various chemical reagents such as chloroform(CHCl3), sodium hypochlorite(NaClO), SDS, EDTA, HC1, NaOH and various methods were examined for their ability to extract PHA and digest non-PHA cellular materials(NPCM).The result showed that (1) the PHA recovery of 78% at the purity of 98.7% was obtained with the mild mix process by CHC13 and the PHA recovery of 89.6% at the purity of 99% was obtained with the severe mix process by CHC13. (2) the purity and recovery of PHA were 93.6%, 91.1% when the cells were treated for 30 min by 40%(V/V) NaClO. (3) the PHA recovery was 91% and the purity was 96.5% when CHC13 and NaCIO were used together. (4) the SDS mix process made the purity and /ecovery of PHA 89.2% and 90%. (5) the PHA recovery of 87.1% at the purity of 97.5% was obtained when the cells were treated by 0.7(w/v)% SDS for 12 min and then by 30% NaCIO for 3 min. (6) the PHA recovery of 89% at the purity of 97.75% was obtained with the mutual mix process by SDS and EDTA. (7) the purity and the recovery of PHA were 95%, 90.3% with the digestion process by HC1. (8) the PHA recovery of 92% at the purity of 96.2% was obtained with the digestion by NaOH. In addition, physical and mechanical properties of PHA was studied. The result certified the recovered PHA product was coincident with the international quality standard in PHA engineering material for high-performance mechanical applications.This paper was also studied primarily on the decolor of PHA. Physical and chemical methods were all used to decolor the organic phase after the recovery by CHC13 from the cells. The result showed that the effect of chemical methods was not more obvious than that of physical methods and acticarbon decolor was a better method for the decolor of PHA.更多还原