骆驼蓬总碱对草地贪夜蛾离体细胞系和幼虫血细胞的毒性作用  

Toxicity effects of total alkaloids of Peganum harmala against in vitro cultured Sf9 cells and larvae blood corpuscles of Spodoptera frugiperda

作　　者：李金梅[1,2] 木魁 姜双林[1,2] 李登坤 李文雍[1,2] 马同富[1] 李永民[1] 

机构地区：[1]阜阳师范学院生物与食品工程学院,安徽阜阳236037 [2]胚胎发育与生殖调节安徽省重点实验室,安徽阜阳236037

出　　处：《西北农林科技大学学报（自然科学版）》2017年第4期127-133,共7页Journal of Northwest A&F University(Natural Science Edition)

基　　金：安徽省自然科学基金面上项目(1308085MC39);安徽省教育厅高校自然科学基金重点项目(KJ2013A207);安徽省省级学科建设重大项目(2014-28-A12)

摘　　要：【目的】研究骆驼蓬总碱对草地贪夜蛾幼虫的杀虫作用机理,为此类生物碱作为新型植物杀虫剂的开发应用提供理论依据。【方法】从骆驼蓬成熟种子中提取骆驼蓬总碱,采用MTT法测定0,1,5,10和20μg/mL骆驼蓬总碱对草地贪夜蛾(Spodoptera frugiperda)卵巢细胞系(Sf9)的毒性;测定5μg/mL骆驼蓬总碱在处理10,30,50,70和90min时对Sf9细胞Na^+和K^+吸收的影响,并测定5μg/mL骆驼蓬总碱在处理1,2,3,4,5d后对葡萄糖吸收的影响;最后采用叶碟法(叶碟在20μg/mL骆驼蓬总碱中浸3s)和注射法(2μg/头)分别测定处理1,8,16,24和48h时骆驼蓬总碱对草地贪夜蛾4龄幼虫血细胞数量的影响。【结果】10和20μg/mL骆驼蓬总碱处理草地贪夜蛾Sf9细胞24h后,对细胞的抑制率分别为62.04%和83.25%;骆驼蓬总碱处理后8,16,24和48h,其对草地贪夜蛾Sf9细胞的LC50分别为35.32,23.21,14.87和8.98μg/mL,骆驼蓬总碱对Sf9细胞的毒性有时间滞后效应。5μg/mL骆驼蓬总碱处理Sf9细胞后10min,Sf9细胞对Na^+、K^+的吸收速率明显增加,处理后30min Sf9细胞对Na^+、K^+的吸收逐渐减弱;处理后3dSf9细胞对葡萄糖的吸收迅速增加,处理后4dSf9细胞对葡萄糖的吸收趋于停止。以叶碟法和注射法处理草地贪夜蛾4龄幼虫,8h后幼虫血细胞数量显著降低,但随时间延长幼虫血细胞数量又逐渐增加。【结论】骆驼蓬总碱对草地贪夜蛾卵巢细胞系Sf9有明显的毒杀活性,且对Sf9细胞中Na^+、K^+和葡萄糖的吸收具有抑制作用;对草地贪夜蛾4龄幼虫血细胞数量也有明显的抑制作用。[Objective] This study investigated the insecticidal mechanism of total alkaloids of Peganum harmala （TAPH） against larvae of Spodoptera frugiperda to provide basis for the development and apc plication of alkaloids as a new plant insecticide. [Method] TAPH was isolated from mature seeds of P. harmala and its cytotoxicity （0,1,5,10,and 20μg/mL） against S. frugiperda Sf9 cells was detected u-sing MTT method. The effect of 50μg/mL TAPH treatment of 10,30,50,70 and 90 min on absorption of Na＋ and K＋ by Sf9 cells was measured,and the effect of 50μg/mL TAPH treatment of 1,2,8,4, 5 d on the absorption of glucose was determined. Finally,leaf disc method （leaves disc in 20 t^g/mL TAPH dip- ping for 3 s） and injecting test （2/1g/head TAPH） were used to detect the effect on blood corpuscle of 4thinstar larvae of S. frugiperda. [Result] After 24 h treatment with 20 gg/mL and 10μg/mL TAPH, the inhibition rates were 83.25% and 62.04 %, respectively. The LC50 to Sf9 cells after treatment of 8,16,24 and 48 h were 35.32,23.21, 14, 87 and 8.98μg/mL,respectively. After 10 min treatment with TAPH,the absorption of Na＋ and K＋ increased rapidly, and the absorption decreased after 30 min. After 3 d treatment,the absorption of glucose increased rapidly, while the absorption stopped after 4 days. Treated by leaf disc and injection methods, the cell amounts of the 4th instar larvae of S. frugiperda decreased significantly, and recovered gradually as time extended. [Conclusion] The total alkaloids had obvious toxicity a- gainst Sf9 cells. The absorption of Na＋ , K＋ , and glucose and the number of blood corpuscle of 4th-instar larvae of S. frugiperda were significantly inhibited.

关 键 词：骆驼蓬 生物总碱 草地贪夜蛾 卵巢细胞系 细胞毒性 

分 类 号：Q965[生物学—昆虫学]