5-氨基乙酰丙酸对中华稻蝗的毒性作用及相关酶活性的影响研究
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摘要
中华稻蝗(Oxya chinensis)属于直翅目(Orthoptera)、斑腿蝗科(Acridoidea)、稻蝗属(Oxya Serville),主要分布在非洲、澳大利亚、亚洲,在我国大多数地区均有分布。其食性比较专一,对禾本科植物为害严重,是水稻产区的重要害虫之一。近年来,由于对传统有机磷农药抗药性的提高和农业生态环境条件破坏的影响,中华稻蝗的发生较为频繁,给农业生产带来了严重为害。目前在中华稻蝗的防治中,主要仍是采用化学杀虫剂防治的手段,但是长期大量使用化学农药,引发了对人、畜健康和土壤水体等生态环境的严重污染,而且中华稻蝗已对常用杀虫剂产生了一定程度的抗药性。因此,研究和开发新型环境友好型杀虫剂进行农业害虫的合理防治具有重要的理论和实践意义。本文研究了光敏杀虫剂5-氨基乙酰丙酸(ALA)对中华稻蝗的急性毒性作用并对其生化机制进行了初步探讨。
本文以中华稻蝗为材料,采自太原市晋源区或取卵室内孵化。用5-氨基乙酰丙酸以不同施药方式处理各龄期的若虫,测定药物的急性毒性作用及对虫体内酶活性的影响,从而探讨药物的毒杀效果和其生化作用机制。
用不同浓度ALA以注射法(CK,250mM,450 mM,750 mM,1000 mM)处理中华稻蝗,在施药48h后(先经14h暗光饲养再给予3h光照,然后于室内饲养至48h)试虫出现明显死亡,同时测定ALA处理组中华稻蝗的乙酰胆碱酯酶(AChE)、谷胱甘肽S-转移酶(GSTs)和谷胱甘肽过氧化物酶(GPx)活性并比较分析。生化研究结果表明,随着5-氨基乙酰丙酸浓度的升高,处理组雌、雄虫体内AChE活性分别较对照组有显著下降(P＜0.05);处理组GPx活性也较对照组有显著下降(P＜0.05);同时,处理组雌、雄虫体内GSTs活性分别较对照组有显著升高(P＜0.05)。由以上结果分析得出,ALA对雌、雄中华稻蝗均有明显的毒性效应且雌、雄中华稻蝗对ALA的敏感性相似;ALA可引起AChE和GPx光失活并能激活GSTs。
基于以上研究结果,确定ALA对中华稻蝗的确有显著的毒杀作用并对虫体内部分酶活性有影响,继而本文又开展了进一步研究,以寻找合理的处理浓度、处理方式以及ALA的毒性作用与中华稻蝗龄期的关系。用不同浓度ALA以药膜法(CK,0.00597 mM,0.0597 mM,0.597 mM,5.97 mM)处理不同龄期(一龄,二龄,三龄,四龄)中华稻蝗,处理后先经17h-20h暗光饲养后再给予2h光照,光照过程中各龄期试虫均表现出离草、扒壁、骚动等明显的药物毒性反应,之后陆续开始出现倒地死亡。同时分体段(头、胸、腹)测定虫体内谷胱甘肽S-转移酶(GSTs)和酯酶的活性。毒杀作用结果表明,以药膜法施药可以使ALA在比注射法降低10~2-10~3的浓度下同样达到很高的致死率。生化测定结果显示,随ALA浓度的升高,一龄到四龄各个龄期处理组的GSTs活性均较对照组呈显著升高(P＜0.05),且GSTs活性升高趋势显示,龄期越大变化越显著,其中四龄虫处理组的头、胸、腹三个体段GSTs活性都表现出显著增加,一龄虫只有腹部有显著升高(P＜0.05)。酯酶活性(三种底物α-NA、α-NB、β-NA)在一龄到四龄虫体内都没有表现出显著的变化趋势。由上述结果分析得出,在GSTs和酯酶这两类代谢解毒酶系中,GSTs可被ALA诱导激活,在加速对ALA的代谢以及缓解由ALA毒性作用所引发的氧化损伤中起一定作用;而酯酶几乎没有参与到与ALA相关的生化反应中。
本文通过研究ALA对中华稻蝗的毒杀作用及生化影响,表明卟啉杀虫剂ALA在低浓度下对中华稻蝗各个龄期都有十分显著的毒杀作用,其毒杀效果与施药剂量、施药方式和光照密切相关,证实ALA在中华稻蝗治理中确是一种高效、有开发潜力的新型杀虫剂。本文从生化水平初步探讨了ALA对中华稻蝗体内相关酶活性的影响,但由于ALA对昆虫毒理机制比较复杂,有待进一步对ALA主要影响的代谢酶系以及光氧化损伤的组织分布开展进一步的系统研究。
Oxya chinensis(Thunberg)(Orthoptera:Acridoidea) is a common species of Oxya Serville,Mainly distributes in Africa,Australia,Asia widely and abundantly distributes in most regions of China.The grasshopper have specific diet and have serious damage to gramineous plants.They are an important agriculture pest to crops.In recent years,due to the increasing resistance to traditional organophosphorus insecticides and the break of the ecological environment and agricultural conditions,the occurrence of O. chinensis is more frequent.At present,for the control of O.chinensis,we mainly use chemical control,but long-lasting use of chemical insecticides have caused serious pollution to people's and animals' health and ecological environment.Meanwhile,the grasshopper have a certain degree of resistance to these pesticides.Therefore,research and development of new environment-friendly pesticides to control agricultural pests have an important theoretical and practical significance.The acute toxicity of photosensitive pesticides 5-aminolevulinic acid(ALA) to O.chinensis and its biochemical mechanism were discussed in this paper.
Various instars nymphs of O.chinensis from Jinyuan district of Taiyuan, Shanxi province were treated with different methods,then measured acute toxicity of 5-aminolevulinic acid and its effect to enzyme activities on O. chinensis,for discussing the toxicity and the biochemical mechanism of the drug.
The nymphs of O.chinensis were treated with different doses of ALA (250 mM;450 raM;750 mM;1000 mM).After treated for 48 h(the first 14 h was dark-keeping,then given 3h light and kept indoors for 48h),the test insects appeared significant death and the activities of acetylcholinesterase (ACHE),glutathione S-transferase(GSTs) and glutathione peroxidase(GPx) were determinated.Biochemical studies showed that with increasing dose of ALA,the activities of ACHE and GPx in treatments significantly declined in female and male compared to control(P＜0.05).Meanwhile,the GSTs activities of female and male O.chinensis in treatments remarkably increased compared to control(P＜0.05).The results obtained from the above analysis indicated that ALA had obviously toxic effect on O.chinensis and the sensitivity of female and male to ALA was similar.Moreover,ALA caused the photoinactivation of AChE and GPx,and activated GSTs.
Based on the above findings,the fact was determined that ALA indeedly played a significant role on poisoning the insects and affect the activities of some enzymes of O.chinensis.The paper carried out a further study to find not only a more reasonable application method on the concentration of ALA and the treatment approach,but also the relationship between toxicity and the instar.The glutathione S-transferase(GSTs) and esterase activities of the head,chest and abdomen of the insects were measured.
Various instar nymphs of O.chinensis were treated with different doses of ALA(0.00597 mM;0.0597 mM;0.597 mM;5.97 mM) using film analysis. After treated with ALA,the insects first kept in dark for 17-20 h,then given 2 h light.During irradiation,the insects showed some obvious toxical reaction to drug,as left from the grass,climbed onto the wall of glass bottle and restlessness,then lay on the ground and died in succession.Toxicity results showed that the concentration of ALA was 100-1000 times lower using application of film bioassays than injection method when reached the same mortality.Biochemical studies showed that with increasing dose of ALA,the activities of GSTs in treatments of every instars nymphs significantly enhanced compared to control(P＜0.05).Moreover the increasing trend of GSTs activities indicated that the older the nymphs were the more significant changes the GSTs activities had.The GSTs activities of fourth-instar nymphs in head,chest and abdomen all appeared signicicant increase;its of first-instar nymphs showed significant rising only in abdomen(P＜0.05). Esterase activities(three substrates:α-NA,α-NB,β-NA) did not showed signicicant changes in trends in every instar nymphs.Based on the above resultes,we analyzed that concerded to GSTs and Esterase these two kinds of metabolism and detoxification enzymes,GSTs could be ALA-induced activitated and played a role on speeding up the metabolism of ALA and reduced toxic effects of oxidative damage caused by ALA.Esterase almost no involved in the ALA-related biochemical metabolism.
The studies on the toxicity and biological effects of ALA on O.chinensis indicated that porphyrin insecticides indeedly showed obvious toxicity at very low concemtrate to various instar nymphs,furthermore its toxic effects correlated with application concerntrate,application method and illumination. ALA was confirmed as a highly efficient and development-potential new insecticide on control of O.chinensis.This paper made an elementary studies on the effects of ALA to some conrrelative enzymes of O.chinensis. However,because of the complexity of the toxicological mechanisms of ALA,it is expected that further systemic researches will be made on the application of ALA and the metabolic enzymes mainly impacted,as well as the distribution of oxidative,damaging tissues.
本文以中华稻蝗为材料,采自太原市晋源区或取卵室内孵化。用5-氨基乙酰丙酸以不同施药方式处理各龄期的若虫,测定药物的急性毒性作用及对虫体内酶活性的影响,从而探讨药物的毒杀效果和其生化作用机制。
用不同浓度ALA以注射法(CK,250mM,450 mM,750 mM,1000 mM)处理中华稻蝗,在施药48h后(先经14h暗光饲养再给予3h光照,然后于室内饲养至48h)试虫出现明显死亡,同时测定ALA处理组中华稻蝗的乙酰胆碱酯酶(AChE)、谷胱甘肽S-转移酶(GSTs)和谷胱甘肽过氧化物酶(GPx)活性并比较分析。生化研究结果表明,随着5-氨基乙酰丙酸浓度的升高,处理组雌、雄虫体内AChE活性分别较对照组有显著下降(P＜0.05);处理组GPx活性也较对照组有显著下降(P＜0.05);同时,处理组雌、雄虫体内GSTs活性分别较对照组有显著升高(P＜0.05)。由以上结果分析得出,ALA对雌、雄中华稻蝗均有明显的毒性效应且雌、雄中华稻蝗对ALA的敏感性相似;ALA可引起AChE和GPx光失活并能激活GSTs。
基于以上研究结果,确定ALA对中华稻蝗的确有显著的毒杀作用并对虫体内部分酶活性有影响,继而本文又开展了进一步研究,以寻找合理的处理浓度、处理方式以及ALA的毒性作用与中华稻蝗龄期的关系。用不同浓度ALA以药膜法(CK,0.00597 mM,0.0597 mM,0.597 mM,5.97 mM)处理不同龄期(一龄,二龄,三龄,四龄)中华稻蝗,处理后先经17h-20h暗光饲养后再给予2h光照,光照过程中各龄期试虫均表现出离草、扒壁、骚动等明显的药物毒性反应,之后陆续开始出现倒地死亡。同时分体段(头、胸、腹)测定虫体内谷胱甘肽S-转移酶(GSTs)和酯酶的活性。毒杀作用结果表明,以药膜法施药可以使ALA在比注射法降低10~2-10~3的浓度下同样达到很高的致死率。生化测定结果显示,随ALA浓度的升高,一龄到四龄各个龄期处理组的GSTs活性均较对照组呈显著升高(P＜0.05),且GSTs活性升高趋势显示,龄期越大变化越显著,其中四龄虫处理组的头、胸、腹三个体段GSTs活性都表现出显著增加,一龄虫只有腹部有显著升高(P＜0.05)。酯酶活性(三种底物α-NA、α-NB、β-NA)在一龄到四龄虫体内都没有表现出显著的变化趋势。由上述结果分析得出,在GSTs和酯酶这两类代谢解毒酶系中,GSTs可被ALA诱导激活,在加速对ALA的代谢以及缓解由ALA毒性作用所引发的氧化损伤中起一定作用;而酯酶几乎没有参与到与ALA相关的生化反应中。
本文通过研究ALA对中华稻蝗的毒杀作用及生化影响,表明卟啉杀虫剂ALA在低浓度下对中华稻蝗各个龄期都有十分显著的毒杀作用,其毒杀效果与施药剂量、施药方式和光照密切相关,证实ALA在中华稻蝗治理中确是一种高效、有开发潜力的新型杀虫剂。本文从生化水平初步探讨了ALA对中华稻蝗体内相关酶活性的影响,但由于ALA对昆虫毒理机制比较复杂,有待进一步对ALA主要影响的代谢酶系以及光氧化损伤的组织分布开展进一步的系统研究。
Oxya chinensis(Thunberg)(Orthoptera:Acridoidea) is a common species of Oxya Serville,Mainly distributes in Africa,Australia,Asia widely and abundantly distributes in most regions of China.The grasshopper have specific diet and have serious damage to gramineous plants.They are an important agriculture pest to crops.In recent years,due to the increasing resistance to traditional organophosphorus insecticides and the break of the ecological environment and agricultural conditions,the occurrence of O. chinensis is more frequent.At present,for the control of O.chinensis,we mainly use chemical control,but long-lasting use of chemical insecticides have caused serious pollution to people's and animals' health and ecological environment.Meanwhile,the grasshopper have a certain degree of resistance to these pesticides.Therefore,research and development of new environment-friendly pesticides to control agricultural pests have an important theoretical and practical significance.The acute toxicity of photosensitive pesticides 5-aminolevulinic acid(ALA) to O.chinensis and its biochemical mechanism were discussed in this paper.
Various instars nymphs of O.chinensis from Jinyuan district of Taiyuan, Shanxi province were treated with different methods,then measured acute toxicity of 5-aminolevulinic acid and its effect to enzyme activities on O. chinensis,for discussing the toxicity and the biochemical mechanism of the drug.
The nymphs of O.chinensis were treated with different doses of ALA (250 mM;450 raM;750 mM;1000 mM).After treated for 48 h(the first 14 h was dark-keeping,then given 3h light and kept indoors for 48h),the test insects appeared significant death and the activities of acetylcholinesterase (ACHE),glutathione S-transferase(GSTs) and glutathione peroxidase(GPx) were determinated.Biochemical studies showed that with increasing dose of ALA,the activities of ACHE and GPx in treatments significantly declined in female and male compared to control(P＜0.05).Meanwhile,the GSTs activities of female and male O.chinensis in treatments remarkably increased compared to control(P＜0.05).The results obtained from the above analysis indicated that ALA had obviously toxic effect on O.chinensis and the sensitivity of female and male to ALA was similar.Moreover,ALA caused the photoinactivation of AChE and GPx,and activated GSTs.
Based on the above findings,the fact was determined that ALA indeedly played a significant role on poisoning the insects and affect the activities of some enzymes of O.chinensis.The paper carried out a further study to find not only a more reasonable application method on the concentration of ALA and the treatment approach,but also the relationship between toxicity and the instar.The glutathione S-transferase(GSTs) and esterase activities of the head,chest and abdomen of the insects were measured.
Various instar nymphs of O.chinensis were treated with different doses of ALA(0.00597 mM;0.0597 mM;0.597 mM;5.97 mM) using film analysis. After treated with ALA,the insects first kept in dark for 17-20 h,then given 2 h light.During irradiation,the insects showed some obvious toxical reaction to drug,as left from the grass,climbed onto the wall of glass bottle and restlessness,then lay on the ground and died in succession.Toxicity results showed that the concentration of ALA was 100-1000 times lower using application of film bioassays than injection method when reached the same mortality.Biochemical studies showed that with increasing dose of ALA,the activities of GSTs in treatments of every instars nymphs significantly enhanced compared to control(P＜0.05).Moreover the increasing trend of GSTs activities indicated that the older the nymphs were the more significant changes the GSTs activities had.The GSTs activities of fourth-instar nymphs in head,chest and abdomen all appeared signicicant increase;its of first-instar nymphs showed significant rising only in abdomen(P＜0.05). Esterase activities(three substrates:α-NA,α-NB,β-NA) did not showed signicicant changes in trends in every instar nymphs.Based on the above resultes,we analyzed that concerded to GSTs and Esterase these two kinds of metabolism and detoxification enzymes,GSTs could be ALA-induced activitated and played a role on speeding up the metabolism of ALA and reduced toxic effects of oxidative damage caused by ALA.Esterase almost no involved in the ALA-related biochemical metabolism.
The studies on the toxicity and biological effects of ALA on O.chinensis indicated that porphyrin insecticides indeedly showed obvious toxicity at very low concemtrate to various instar nymphs,furthermore its toxic effects correlated with application concerntrate,application method and illumination. ALA was confirmed as a highly efficient and development-potential new insecticide on control of O.chinensis.This paper made an elementary studies on the effects of ALA to some conrrelative enzymes of O.chinensis. However,because of the complexity of the toxicological mechanisms of ALA,it is expected that further systemic researches will be made on the application of ALA and the metabolic enzymes mainly impacted,as well as the distribution of oxidative,damaging tissues.
引文
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