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目的:对具有广谱抑菌活性的东北林蛙(Rana dybowskii)重组抗菌肽dybowskin-1ST进行热稳定性、酸碱稳定性、局部过敏反应(Arthus反应)及溶血活性分析,为进一步摸清重组肽的理化特性及其应用提供科学依据。方法:将重组抗菌肽dybowskin-1ST分别置于不同环境温度(25～100℃)及不同酸碱环境(pH2～pH12)中处理4 h,观察其对金黄色葡萄球菌(Staphylococcus aureus)及大肠杆菌(Escherichia coli)的抑菌环直径,确定其稳定性; 用不同浓度梯度的重组dybowskin-1ST进行溶血试验和局部过敏反应(Arthus反应)。结果:1.重组dybowskin-1ST在25～100℃温度及pH2～pH12环境下均有抑菌活性,其中,最适环境温度为25℃; 重组dybowskin-1ST在pH3～pH7范围对Staphylococcus aureus抑菌活性较强(抑菌环直径为12.77～13.17 mm),在pH3～pH8范围对Escherichia coli抑菌性较强(抑菌环直径为12.63～13.37 mm)。2.溶血试验结果显示,重组dybowskin-1ST对家兔红细胞的溶血率在0.14%～4.4%; 小鼠Arthus氏试验观察其腹部皮下重组dybowskin-1ST注射部位红肿不明显,无出血、坏死等炎性反应。结论:实验结果显示重组dybowskin-1ST具有较好稳定性、低溶血性及低致敏性,为其应用提供科学依据。

Cyanea nozakii Kishinouy e (C. nozakii), a giant cnidarian of the class Scyphomedusae, order Semaeostomeae and family Cyaneidae, is widely distributed in the East China Sea, the Yellow Sea and the Bohai Sea, and is abundant from late summer to early autumn. Venom produced by C. nozakii during mass agglomerations can contaminate seawater resulting in death of the halobios and seriously damage commercial fisheries. Swimmers and fishermen commonly suff er painful stings from this jellyfish, resulting in local edema, tingling, breathing difficulties, depressed blood pressure and even death. Such effects arise from the complex mixture of biologically active molecules that make up jellyfish venom. In the present study, the hemolytic activity of venom from tentacles of C. nozakii and factors aff ecting its activity were assayed. The HU50 ( defined as the amount of protein required to lyse 50 % of erythrocytes) of the venom against dove and chicken erythrocytes was 34 and 59 μg/mL, respectively. Carboxylmethyl chitosan and glycerol could increase hemolytic activity at concentrations greater than 0.06% and 0.2 mol/L, respectively.

The present work is the first report of the biochemical characterization of the venom from nematocysts of the jellyfish Rhopilema esculentum Kishinouye. The nematocysts were isolated by autolysis and centrifugation and separated by flow cytometry. Four types of nematocysts were identified: mastigophores, euryteles, and atrichous and holotrichous isorhiza. SDS-PAGE and amino acid analyses demonstrated that most of the proteins in the nematocyst extract were between 10 kDa and 40 kDa, and that glutamic acid was the main amino acid. A hemolytic activity assay showed that the activity of the nematocyst venom (RNV) was strongest in Tris-HCl buffer (50 mmol/L, pH 7.8, 5% glycerol, 0.5 mmol/L EDTA, 0.1 mol/L NaCl). The hemolytic activity was related to protein concentration and the HU50 against chicken erythrocytes was 0.91 mcg/mL. [PUBLICATION ABSTRACT]