【摘要】：湖北釘螺是日本血吸蟲的唯一中間宿主,藥物滅螺能有效預(yù)控血吸蟲病流行。NiC是WHO唯一指定、我國常用的化學(xué)滅螺藥物,但存在水溶解度低、靶標(biāo)/點的選擇毒性差及價格昂貴等技術(shù)經(jīng)濟(jì)難點。因此,需要緊密結(jié)合預(yù)控血吸蟲病的國家重大技術(shù)需求研發(fā)新型滅螺藥物。本文以苯胺NiC和滅螺植物水溶性甾體皂甙/生物堿為供試藥物,以釘螺和財魚為靶標(biāo),探討藥物對其蛋白質(zhì)表達(dá)和酶活力的影響,這為新型滅螺藥物研發(fā)的先導(dǎo)物和特定靶點分選提供依據(jù)。 1、釘螺軟體和財魚肝臟GOT酶催化最適條件的研究 通過L25(56)實驗及極/方差分析,測得釘螺軟體/財魚肝臟勻漿液GOT酶催化最適條件分別在酶液濃度0.2/0.4mg/mL、底物濃度5.0/5.0μmol/L、反應(yīng)體系pH7.5/8.0、反應(yīng)時間60/20min及反應(yīng)溫度45/45℃時,其轉(zhuǎn)氨基速率最高。提示物種間GOT酶催化動力學(xué)不同。這為滅螺藥物對其活性影響研究提供實驗依據(jù)。 2、滅螺藥物溫孵/浸泡對釘螺和財魚的GOT酶活力影響 2.1藥物溫孵離體對釘螺軟體的GOT酶活力 濃度效應(yīng)：梯度質(zhì)量數(shù)的NiC和吉祥草、黃果茄及盾葉薯蕷水提取物,分別與酶溫孵20min,在其0.25、0.5、0.5及2.0mg/L的酶活力誘導(dǎo)率最高,分別達(dá)到19.3、23.7、18.9及11.52%,提示釘螺GOT酶對吉祥草甾體生物堿的敏感性最高。 2.2 NiC溫孵離體財魚肝臟的GOT酶活力 濃度效應(yīng)：梯度質(zhì)量數(shù)的NiC分別與酶溫孵30min,0.125mg/L NiC誘導(dǎo)率高達(dá)39.94%。 時間效應(yīng)：0.25mg/L NiC與酶經(jīng)時溫孵,40min溫孵酶活力誘導(dǎo)率高達(dá)44.86%。 2.3 NiC染毒財魚肝臟的GOT酶活力 梯度質(zhì)量的NiC浸泡染毒財魚,1.5mg/L時財魚死亡時間最短3.5h,GOT酶活力誘導(dǎo)率最大113.99%。 提示NiC誘導(dǎo)GOT酶活力可能是其致死財魚的靶點之一 3、NiC染毒對財魚肝臟蛋白質(zhì)表達(dá)影響 0.5mg/L NiC染毒財魚48h,取肝臟,運用2-DE技術(shù)及PDQuest 8.0.1軟件分離分析,篩選出處理組有10個蛋白表達(dá)上調(diào)。這為NiC毒魚機制及新型滅螺藥物可能作用的特定靶蛋白提供一定的依據(jù)。
[Abstract]:Oncomelania hupensis (Oncomelania hupensis) is the only intermediate host of Schistosoma japonicum (Schistosoma japonicum). The technical and economic difficulties such as poor toxicity and high price of target / point selection. Therefore, it is necessary to develop new snail control drugs closely in combination with the national major technical requirements for schistosomiasis control. The effects of aniline NiC and water soluble steroidal saponins / alkaloids of snail killing plants on protein expression and enzyme activity of Oncomelania hupensis and Oncomelania hupensis were studied in this paper. This provides a basis for the development of novel snail-killing drugs and the selection of specific targets. 1. The optimum conditions of GOT enzyme catalytic activity of Oncomelania molluscum and liver of Oncomelania hupensis were studied by L25 (56) experiment and pole / variance analysis. The results showed that the optimum conditions of GOT catalytic activity in the liver homogenate of Oncomelania hupensis were 0.2 渭 g / L and 5.0 / 5.0 渭 mol / L, respectively, and the reaction system pH7.5/8.0, was obtained at the concentration of 0.2 渭 mol / L and 5.0 渭 mol / L, respectively. When the reaction time 60/20min and reaction temperature were 45 鈩，

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