| 罗非鱼肌肉乙酰胆碱酯酶的纯化、性质及农药敏感性研究 |
| Purification and Characteristics of Muscle Acetylcholinesterase of Oreochromis aurea and Its Sensitivity to Insecticides |
| 投稿时间：2011-02-09 修订日期：2011-02-09 |
| DOI： |
| 中文关键词:罗非鱼 乙酰胆碱酯酶 生物化学标记 |
| 英文关键词:Oreochromis aurea acetylcholinesterase biochemical marker |
| 基金项目:广东省科技计划项目(2011B030800021) |
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| 中文摘要: |
| 鱼体内乙酰胆碱酯酶(AChE)活性的变化是水体环境农药污染的重要生物化学标记。以奥利亚罗非鱼(Oreochromis aurea)肌肉组织为材料,研究了乙酰胆碱酯酶的分离纯化方法、性质及杀虫剂对酶的抑制作用。结果表明,当肌肉组织的Triton X-100抽提物采用(NH4)2SO4分级盐析、Sephadex G-100凝胶过滤和DEAE-纤维素DE23离子交换柱层析的分离纯化步骤后,可获得纯化的乙酰胆碱酯酶,酶的比活力达到21.34U/mg,纯化倍数118,回收率22.08%。多因素正交试验分析确定的酶反应条件为反应体系pH7.5,反应温度40℃,反应时间15min,底物浓度7mmol/L。纯化酶水解碘化硫代乙酰胆碱的速度快,Km值为0.229mmol/L,存在过量底物抑制现象。4种杀虫剂(敌敌畏、辛硫磷、三唑磷、灭多威)对罗非鱼肌肉AChE的IC50值均小于5×10-6mol/L,有机磷类杀虫剂对罗非鱼脑AChE的抑制能力高于氨基甲酸酯类,抑制能力分别为:敌敌畏>辛硫磷>三唑磷>灭多威。4种农药对罗非鱼肌肉中AChE的抑制具有很好的剂量-效应关系,相关系数均在0.93以上，表明罗非鱼肌肉AChE可以作为水体环境中农药污染的生物化学标记。 |
| 英文摘要: |
| The decline of enzymatic activity of acetylcholinesterase (AChE ) is a significant biochemical indicator to agricultural chemical pollution in aquatic environment. Muscular tissues of Oreochromis aurea were tested on to study purification methods，characteristics of acetylcholinesterase(AChE)and its sensitivity to insecticides. The result indicated AChE was purified from the Triton X一100 extract by(NH4)2SO4 fractional precipitation，SephadexG-100 gel filtration and DEAE-cellulose through DE23 ion exchange chromatography，with the specific activity of this purified AChE was 21.34 U/mg，purification fold 118，and recovery rate 22. 08%.The optimal condition of AChE activity determined by multi-factor orthogonal method :pH value of the reaction system was 7.5 ; temperature was 40℃; reaction time was l0 min; concentration of substrate was 7 mmol/L. This purified acetylcholinesterase (AChE) hydrolyzed agent transport and communication interface(ATCI)at high rate with Michaelis constant(Km)0.229 mmol/L，there being inhibition of excess substrate. Four insecticides(dichlorvos，phoxim，triazo-phos，and methomyl) exhibited strong inhibitions on this AChE with IC50 values less than 5 x 10一6mol/L. Compared to carbamate insecticide，the inhibition potential of organophosphate was higher，and the inhibition potential was: dichlorvos>phoxim>triazophos>methomyl. The inhibition of four insecticides(dichlorvos，phoxim，triazo-phos，and methomyl)on muscle acetylcholinesterase(AChE) of 0. aurea showed a good dose-effect relation，with their correlation coefficients all above 0. 93.The above results suggest that it is possible to use muscle AChE in 0.aurea as a biochemical marker to monitor the contamination of insecticides in aquatic environment. |
| 丁运华,吴晓敏,方俊彬.2012.罗非鱼肌肉乙酰胆碱酯酶的纯化、性质及农药敏感性研究[J].水生态学杂志,33(1):120-126. |
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