小菜蛾Plutella xylostella羧酸酯酶、酰胺酶及乙酰胆碱酯酶的研究

研究结果表明,小菜蛾(Plutella xylostella)乙酰胆碱酶、酰胺酶的最适pH值分别为8.0和8.5。pH值对闳酸酯酶活力影响较小,酰胺酶和羧酸酯酶反应速度在反应开始60钟内,仍在线性范围,乙酰胆碱酯酶反应速度在反应开始的8分钟内呈线性关系。羧酸酯酶、酰胺酶和乙酰胆碱酯酶的Vmax分别为551.81(nmol/mg,mln)、5.25×10~(-4)(μmol/mg.min)和20.36(nmol/mg.min);km分别为5.52×10~(-5)(mol)、1.18×10~(-3)(mol)和8.33×10~(-3)(mol)。     

正交试验法确定烟草甲成虫羧酸酯酶反应的最适条件

应用正交表L25(56)设计正交试验,研究了羧酸酯酶质量浓度、体积分数浓度、反应体系的pH值、反应温度和反应时间等5个因素对烟草甲成虫羧酸酯酶活性检测的影响,并从试验组合中选出最佳条件.在进行极差分析、方差分析和多重比较后,选择各因素的最优水平,得到了烟草甲成虫羧酸酯酶活性测定的最适条件,即酶的质量浓度为1头/mL、底物的体积分数为5.0×10-4 mo1/L,pH值为7.5,温度为42℃,时间为10 min.     

2种拟青霉代谢产物对烟蚜乙酰胆碱酯酶和羧酸酯酶的影响

 以粉拟青霉菌的sw03032菌株和玫烟色拟青霉菌的sw03085为研究对象,比较其代谢产物的杀蚜虫活性和对烟蚜乙酰胆碱脂酶和羧酸酯酶活性的影响.结果表明2种真菌发酵液含有能毒杀蚜虫的毒素物质.且毒素粗提物浓度越大,杀蚜活性越强.经玫烟色拟青霉菌不同浓度毒素粗提液处理的蚜虫乙酰胆碱酯酶比活力分别比对照(CK2)降低了70.4%～29.8%.不同浓度玫烟色拟青霉菌毒素粗提液处理12h后,蚜虫羧酸酯酶比活力分别比对照(CK2)降低79.5%～44.4%.用不同浓度粉拟青霉菌毒素粗提液处理烟蚜12h后,乙酰胆碱酯酶和酸酸酯酶活性分别降低了38%～18.6%和51.6%～59.5%.玫烟色拟青霉菌的sw03085菌株代谢产物对烟蚜的伤害活性明显较粉拟青霉菌sw03032菌株代谢产物高.     

3种杀虫剂对杨树羧酸酯酶的抑制效果

研究了酶浓度、底物浓度、反应体系pH、反应温度、反应时间5个因素对美洲黑杨羧酸酯酶(CarE)活性测定的影响,得出测定杨树CarE的最适反应条件为:酶稀释10倍,底物浓度为3×10-4mol/L,pH为7.0,温度30℃,时间15 min。利用试验得到的反应最适条件分析了氧化乐果、吡虫啉和氟虫腈3种内吸性药剂对杨树CarE活性的影响。结果表明:400、1 600、3 200和6 400mg/L氧化乐果能明显抑制杨树CarE活性,其中6 400 mg/L氧化乐果抑制作用最强。50、100、200、400和800 mg/L吡虫啉处理杨树48 h,对杨树CarE均没有显著影响;当处理96 h时,400和800 mg/L吡虫啉能显著抑制CarE活性。而400 mg/L氟虫腈处理杨树48 h,800 mg/L氟虫腈处理杨树96 h,都能显著抑制杨树CarE活性。     

Effects of dietary nickel on detoxification enzyme activities in the midgut of <Emphasis Type="Italic">Spodoptera litura</Emphasis> Fabricius larvae

Nickel accumulated in midugt of Spodoptera litura Fabricius could induce the expression of metallothionein, one of the most important detoxification proteins in organisms. In the present study, the effects of dietary nickel on the activities of detoxification enzymes, such as carboxylesterase （CarE） and giutathione S-transferase （GST） in the midgut of S. litura larvae have been studied to get an understanding of the detoxification mechanisms of S. litura larvae to excessive nickel. Results showed that CarE activities in the midgut of the 5th instar larvae decreased at lower levels of nickel （≤5 mg/kg）, while increased with increasing nickel doses at higher levels of nickel （≥10 mg/kg） exposure in successive 3 generations. CarE activities of the 6th instar larvae were also characterized as inhibited at low levels of nickel exposure, and improved at higher levels in the 1st generation. CarE activities of 6th instar larvae in the 2nd and 3rd generations were all lower than that in control. However, GST activities in the midgut of the 5th and 6th instar larvae all increased with increasing nickel doses （1 --20 mg/kg） in diets.     

Toxicity of Diphenyl-pentenone Against Three Aphids （Homoptera： Aphididae） and Its Effects on the Esterases and Glutathione S-transferase

The most provocative aspect of this study is its original findings on the toxicity of diphenyl-pentenone (1,5-diphenyl-2-penten-l-one, DP) against the three kinds of aphids (Maerosiphum granarium Kirby, Lipaphis erysimi (kaltenbaeh), Schizaphis aurantiae (Fosc.) and the inhibitory effects of DP againstdetoxification enzyme system of the aphid (S aurantiae). The result of bioassay in the laboratory shows that the product has strong contact activity and very good antifeeding activity, with higher efficacy than anabasine and nicotine, two botanical aphidicides. The median lethal concentrations of the three products against the apterous adult (Mgranarium) are 175.85, 217.23 and 245.22 mg/L, respectively, at 24h of pest treatment. DP is inferior to methomyl in contact assay but superior in antifeeding activity assay against the three aphides. DP has strong inhibitory effects on nonspecific esterases, carboxylesterase of aphid (S aurantiae), but it can not inhibit AchE. DP also has strong inhibitory effects on glutathione Stransferase of the aphid.     

Novel substrates for the kinetic assay of esterases associated with insecticide resistance

Naphthalene thioesters were synthesized as substrates for a continuous, non- disruptive kinetic assay of general carboxylesterase activity. The continuous nature of the assay is based on the production of a soluble dianion chromophore from the reaction of naphthalene thio with 5,5-dithiobis (2-nitrobenzoic acid). Applications with 1- and 2-naphthalene thioacetates demonstrated their use in a fast accurate kinetic microassay of esterase activity, using porcine carboxylesterase as a model. These novel esters proved to be useful as substrates for the spectrophotometric assay of insecticide-resistance in two aphid species and may be applicable to other esterasebased diagnostic procedures.     

褐飞虱羧酸酯酶基因cDNA片段的克隆及表达分析

利用反转录多聚酶链式反应(RT-PCR)技术克隆了褐飞虱羧酸酯酶基因编码区的cDNA片段,并进行了序列测定.结果表明,所克隆到的cDNA片段长度为396 bp,经BLAST查找比对发现,该片段所编码的氨基酸序列与来自铜绿蝇、家蝇、沟鼠、黑腹果蝇、线虫和埃及伊蚊的羧酸酯酶的片段存在高度同源性.Northern杂交分析显示,在褐飞虱取食抗性水稻后,羧酸酯酶基因表达水平明显升高.以上结果表明,羧酸酯酶基因的表达受抗性水稻的诱导,该基因在有毒化学物质解毒及增强褐飞虱对抗性水稻的耐受性方面可能起着重要作用.     

Effects of dietary nickel on detoxification enzyme activities in the midgut of Spodoptera litura Fabricius larvae

Nickel accumulated in midugt of Spodoptera litura Fabricius could induce the expression of metal-lothionein,one of the most important detoxification proteins in organisms.In the present study,the effects of dietary nickel on the activities of detoxification enzymes,such as carboxylesterase(CarE) and glutathione S-transferase(GST) in the midgut of S.litura larvae have been studied to get an un-derstanding of the detoxification mechanisms of S.litura larvae to excessive nickel.Results showed that CarE activities in the midgut of the 5th instar larvae decreased at lower levels of nickel(≤5 mg/kg),while increased with increasing nickel doses at higher levels of nickel(≥10 mg/kg) exposure in suc-cessive 3 generations.CarE activities of the 6th instar larvae were also characterized as inhibited at low levels of nickel exposure,and improved at higher levels in the 1st generation.CarE activities of 6th instar larvae in the 2nd and 3rd generations were all lower than that in control.However,GST activities in the midgut of the 5th and 6th instar larvae all increased with increasing nickel doses(1―20 mg/kg) in diets.