尖吻蝮(Deinagkistron acutus)生长过程不同时期排毒量及蛇毒组分的变化

对人工培育的4个年龄段尖吻蝮蛇毒和野生成体尖吻蝮蛇毒进行了聚丙烯酰胺凝胶电泳分析.结果显示3龄以前尖吻蝮蛇毒蛋白电泳图谱,无论是在分带、泳动率及相应组分的量等方面,均呈现一定的差异,但随着尖吻蝮蛇龄的增长,蛇毒蛋白电泳图谱与野生成蛇蛇毒蛋白电泳图谱越来越接近.3龄尖吻蝮生长发育达到性成熟,其蛇毒蛋白电泳图谱与野生成蛇趋于一致.蛇毒组分的变化,提示了尖吻蝮的生长发育进程.     

尖吻蝮（Deinagkistrodon acutus）生长过程不同时期排毒量及蛇毒组分的变化

对人工培育的4个年龄段尖吻蝮蛇毒和野生成体尖吻蝮蛇毒进行了聚丙烯酰胺凝胶电泳分析。结果显示：3龄以前尖吻蝮蛇毒蛋白电泳图谱,无论是在分带、泳动率及相应组分的量等方面,均呈现一定的差异,但随着尖吻蝮蛇龄的增长,蛇毒蛋白电泳图谱与野生成蛇蛇毒蛋白电泳图谱越来越接近。3龄尖吻蝮生长发育达到性成熟,其蛇毒蛋白电泳图谱与野生成蛇趋于一致。蛇毒组分的变化,提示了尖吻蝮的生长发育进程。     

尖吻蝮蛇毒的聚丙烯酰胺凝胶电泳，冬眠特异蛋白的发现

目的 比较冬眠期、冬眠前后、活动期尖吻蝮蛇毒的组分.方法 聚丙烯酰胺凝胶电泳.结果 发现了冬眠期尖吻蝮蛇毒中有一个特异性增强的蛋白质成分,进入冬眠几天后,该成分就可以检测到,出眠几天后及活动期蛇毒中不可见.结论 该成分应该与尖吻蝮冬眠密切相关,暂且称其为"冬眠特异蛋白".     

尖吻蝮蛇毒对小鼠肾脏损伤的研究

尖吻蝮(五步蛇)Agkistrodonacutus是我国特有的毒蛇,尖吻蝮蛇毒除引起局部出血肿胀和坏死等症状外,临床常伴有肾功能衰竭或尿毒症的症状。为给临床治疗提供依据,本实验对尖吻蝮蛇毒引起损伤后肾脏的改变,用小鼠作了试验观察,材料与方法取体重20克左右的小白鼠。每只注入40微克的尖吻蝮蛇毒,先将粗蛇毒溶子生理盐水中,使每0.1毫升生理盐水含     

武夷山区不同月份尖吻蝮蛇毒的毒力比较

尖吻蝮 Dienagkistrotrodon acutus(Guenther)属于蝰科 viperidae 蝮亚科 crolalinae 尖吻蝮属Dienagkistrodon 是一种剧毒蛇,也是一种重要的药用动物.主产我国南方山区,为进一步做好蛇伤防治,我们对武夷山区不同月份的尖吻蝮蛇毒的毒力进行了比较研究。     

尖吻蝮自然越冬洞穴的结构观察

为了解决尖吻蝮在人工饲养条件下难以安全越冬的问题 ,在黄山地区对尖吻蝮的自然越冬进行了数年的调查 ,陆续观察了 2 3个越冬洞穴的结构。初步总结了尖吻蝮自然越冬的生态 ,并用数据和图示方式较详细地报道了尖吻蝮自然越冬洞穴的构造 ,介绍了人工饲养过程中保证尖吻蝮安全越冬的管理方法。     

蛇油化学成分的研究

蛇油的应用在我国有悠久的历史,李时珍在《本草纲目》中记载了蛇油的功效:蛇脂“绵裹塞耳聋,亦傅肿痛”,又“摩着物皆透也”。民间常用蛇油治疗冻疮、皲裂、湿疹及水火烫伤。蛇油的现代研究,已见文献及产品。作者以尖吻蝮蛇油为原料制成了“黄山蛇油膏”,用于临床治疗冻疮显效率753%,总有效率94%,治疗手足皲裂显效率90%,总有效率98%。为深入研究蛇油成分,更好地开发蛇油制品,作者对尖吻蝮蛇油及以混合蛇油为原料制成的“黄山蛇油膏”的主要脂肪酸进行了定性、定量分析。1　实验材料尖吻蝮蛇油　取成体尖吻蝮10条,放血致死,取蝮腔内脂肪,洗净…     

尖吻蝮蛇凝血酶进入国家一类新药临床实验阶段

由中国科学院昆明动物研究所肖昌华研究员主持的“注射用尖吻蝮蛇凝血酶”研究项目 ,经国家药品监督管理局审查 ,符合国家一类新药审批的有关条件 ,批准于 2 0 0 3年 4月进入Ⅰ期临床实验 (批件号 :2 0 0 3L0 14 30 )。尖吻蝮蛇凝血酶为我国特产的尖吻蝮 (Agkistrodonacutus)蛇     

尖吻蝮蛇毒去纤酶及其临床应用研究

Ottyang,C.等(1967)从尖吻蝮(Agkistrodon acutus)蛇毒中分离纯化得到凝血酶样成分,分子量33500,由17种263个氨基酸组成,具有体内去纤作用,但未见用于临床的报道。杉原久义等(1978)也从尖吻蝮蛇毒中分离纯化得到了一个促凝组分,分子量52000,具有精氨酸酯酶和凝血酶活性,未见体内血液学效应及临床应用的报道。我们从1974年起,对尖吻蝮蛇毒凝血酶样酶进行7较为系统的研究,并研制成“去纤酶注射液”,于1978年首先用于临床,1981年鉴定通过。这是我国最先研制并用于临床的蛇毒酶制剂。它由四个凝血酶样同工酶组成,其分子量分别是33500,68000,19500,30000;它们均由17种氨基酸组成,但组成     

中国蝮蛇头骨形态比较及分类学讨论

目前,我国蝮蛇某些种类的分类仍存在不同见解,对蝮蛇类群的分类主要集中在外部形态的比较,而较少涉及内部结构的研究.因此,本文对我国13个地区的8种(亚种)共26号蝮蛇的头骨进行了比较解剖.以头骨特征为依据,对岩栖蝮Agkistrodon saxatilis、乌苏里蝮A.ussurensis、蛇岛蝮A.shedaoensis、短尾蝮A brevicaudus、秦岭蝮A.qinlingensis等的有效性及分类地位进行了讨论;同时对尖吻蝮Deinagkistrodon acutus的骨骼形态与其他种类也进行了比较,认为它们在谱系上应代表两个不同的类型,将它们分隶于蝮属Agkistrodon和尖吻蝮属Deinagkistrodon是合理的.还以头骨形态特征为依据对8种蝮蛇编制了检索表.     

五步蛇毒对小鼠免疫功能的影响

目的探索五步蛇毒对正常小鼠免疫功能的影响。方法将KM小鼠随机分为5组,分别为空白组、五步蛇毒安全范围内低剂量组(30 mg/kg)、五步蛇毒安全范围内中剂量组(60 mg/kg)、五步蛇毒安全范围内高剂量组(120 mg/kg)(以下简称五步蛇毒低、中、高剂量组)以及西洋参组(30 mg/kg),每组10只。各组小鼠每日定时灌胃1次,连续14天。观察小鼠的呼吸、精神状态等生命体征。取小鼠脾脏和胸腺组织称重后,计算小鼠脾脏、胸腺指数;制备脾细胞悬液,采用CCK-8法检测小鼠脾淋巴细胞增殖率,碳粒廓清法检测小鼠吞噬系数α,中性红法检测小鼠腹腔巨噬细胞吞噬率;同时采用二硝基氟苯诱导小鼠迟发型变态反应,检测小鼠耳廓肿胀度。结果小鼠的吞噬系数α在五步蛇毒高剂量组最高(5.56±0.46),高于其余四组,差异均有统计学意义(均P<0.05)。小鼠腹腔巨噬细胞吞噬率(%)在五步蛇毒中剂量组和西洋参组中较高,分别为(119.07±19.31)%、(124.79±24.93)%,差异有统计学意义(均P<0.05)。小鼠的B淋巴细胞增殖率(%)在五步蛇毒中剂量组和西洋参组中较高,分别为(105.95±14.59)%、(108.14±10.58)%,差异有统计学意义(均P<0.05)。小鼠的T淋巴细胞增殖率(%)在五步蛇毒中、高剂量组中较高,分别为(119.30±20.07)%、(110.85±23.13)%,差异有统计学意义(均P<0.05)。与空白组比较,五步蛇毒中、高剂量组和西洋参组均能显著提高小鼠的耳廓肿胀度,差异有统计学意义(均P<0.05)。与空白组比较,五步蛇毒各剂量组和西洋参组均能显著提高小鼠脾脏指数(均P<0.05)。与空白组比较,各实验组胸腺指数均无明显差异(均P>0.05)。结论口服低于120 mg/kg的五步蛇毒对正常小鼠的免疫功能具有一定的增强作用。     

Age-related Variation in Snake Venom： Evidence from Two Snakes （Naja atra and Deinagkistrodon acutus） in Southeastern China

In this study we explored electrophoretic profiles, enzymatic activities and immunoreactivity of neonate and adult venoms from two snakes （Naja atra and Deinagkistrodon acutus） coexisting in southeastern China. Age-related variation in electrophoretic profiles was found in both species and proteolytic and fibrinogenolytic activity was higher in neonate than adult venoms. Neonate D. acutus venom had higher 5＇ nucleotidase, PLA2, hyaluronidase and gelatinolytie activity, but lower esterolytic activity, than adult venom. Neonate and adult D. acutus venoms showed identical phosphomonoesterase, LAO and fibrinolytic activities. Neonate N. atra venom had higher phosphomonoesterase and LAO activity, but lower 5＇ nucleotidase, PLA2, hyaluronidase and Ache activities than adult venom. Neonate and adult N. atra venoms showed similar gelatinolytic activity. Further, age-dependent immunoreactivity was found in both species, and cross-reactions between homologous venoms and antiserums were closely related to venom composition. We speculate that age-related variation in venom characteristics is possibly driven by evolutionary forces associated with ontogenetic shifts in dietary habits, competition and predation pressure.     

墨旱莲对4种蝮蛇毒引起的炎症和出血的影响

目的探讨墨旱莲提取液对短尾蝮蛇毒、蛇岛蝮蛇毒、白眉蝮蛇毒及尖吻蝮蛇毒所致的炎症和出血的影响。方法应用短尾蝮蛇毒、蛇岛蝮蛇毒、白眉蝮蛇毒及尖吻蝮蛇毒所致大鼠足跖肿胀的致炎模型,观察墨旱莲提取液对蛇毒所致大鼠足跖肿胀的影响。墨旱莲提取液分别与不同蛇毒混合,给小鼠腹部皮下注射,观察其对蛇毒引起的小鼠皮下出血的影响。结果墨旱莲提取液15g／kg连续2次灌胃给药,对短尾蝮蛇毒、蛇岛蝮蛇毒、白眉蝮蛇毒或尖吻蝮蛇毒所致大鼠足跖肿胀的急性炎症造模和短尾蝮蛇毒棉球肉芽肿的慢性炎症造模(20g／kg)均有明显的抑制作用,对这些蛇毒引起的小鼠皮下出血也能明显抑制。结论墨旱莲提取液对短尾蝮蛇毒、蛇岛蝮蛇毒、白眉蝮蛇毒及尖吻蝮蛇毒引起的炎症和出血均有明显的抑制作用。     

尖吻蝮幼蛇就地和异地人工养殖研究

目的研究尖吻蝮（Deinagkistrodon acutbus）幼蛇在人工养殖过程中的技术难关,探讨尖吻蝮幼蛇生长缓慢的原因,以提高幼蛇的成活率和生长率。方法采用湖南省永州市野生尖吻蝮种蛇产卵孵化的幼蛇,在原产地永州与非原产地广西南宁进行幼蛇的人工饲养对比实验,观察在不同地域人工饲养的尖吻蝮幼蛇的生长速度。结果尖吻蝮幼蛇在本地及异地人工饲养均获得成功,尤其以南宁人工饲养的幼蛇最为明显,2010年9月饲养至2011年8月,幼蛇均重从（13．8±1．8）g增长至（198．8±80．6）g,最重的个体体重达350g。两地饲养的幼蛇均可自行捕食中华蟾蜍、老鼠及活小鸡、鸭苗,仅用1年时间将野生尖吻蝮子代幼蛇人工饲养成功。结论尖吻蝮幼蛇在本地及异地人工饲养是可行的。     

乌梢蛇血清的抗出血因子:一个有前途的抗蛇毒药物原料

用柱层析和聚丙烯酰胺凝胶盘状电泳法,从乌梢蛇血清中分离纯化了一个抗出血因子。用SDS-聚丙烯酰胺凝胶电泳法测得其分子量大约为65 kD;测定了五种蝮亚科蛇毒(尖吻蝮、竹叶青蛇、原矛头蝮、哈扑和短尾蝮)的最小出血剂量和乌梢蛇血清中抗出血因子对这五种蛇毒的抗出血活性;还测定了七种蛇毒(除上述五种毒蛇外,还包括圆斑蝰和银环蛇)的半数致死量,以及抗出血因子对中毒小鼠的治疗作用。结果显示:从乌梢蛇血清中提纯的抗出血因子的抗蛇毒活性,不仅可以抵抗它的捕食者尖吻蝮的蛇毒,而且还可以抵抗具出血活性的其它蛇毒;但它对不具出血活性的银环蛇毒的致死抑制作用不明显。该抗出血因子不仅在体外实验表现出强的中和出血毒素的活性,而且在体内实验中亦表现出对中毒小鼠良好的治疗作用,因而可能成为新的抗蛇毒药物的有前途的原料。乌梢蛇血清对血循毒的中和能力的获得,可能归因于尖吻蝮与乌梢蛇之间捕食与被捕食相互作用的关系。     

Characterization of the cDNA encoding a BPI/LBP homologue in venom gland of the hundred-pace snake Deinagkistrodon acutus

Bactericidal/permeability-increasing protein(BPI)and LPS-binding protein(LBP)play an important role in host defence.Current evidence shows that BPI/LBP may be widely existed in different cells and tissue types of animals.A full-length cDNA clone encoding a BPI/LBP homologue(dBPI),1757 bp in size,was characterized in venom gland of the hundred-pace snake Deinagkistrodon acutus.Its deduced amino acid sequence of 417 residues had 13.8%-21.5% identity to BPI like 1(BPIL1)and BPI like 3(BPIL3)of other animals.Co...     

Molecular phylogeography of endangered sharp-snouted pitviper (Deinagkistrodon acutus; Reptilia, Viperidae) in Mainland China

Using phylogenetic and population genetic approaches, the present study reports the phylogeographic structure of the sharp-snouted pitviper (Deinagkistrodon acutus), a threatened snake species with commercial and medicinal importance in China. The entire mitochondrial ND2 gene (NADH dehydrogenase subunit 2) sequences of 86 individuals of D. acutus from 14 localities across its range in China were determined. Based on the results of phylogenetic analyses, distribution of diagnostic sites, haplotype network, and AMOVA hierarchical analysis, an east-west division of the whole D. acutus population could be observed. Geographically, a line formed by a lake, river, and mountain chain (the Poyang Lake, Gan River to the southern end of the Wuyi Mountains), results in vicariance and approximately vertically splits the range into two and the whole population into two main lineages (western and eastern). The bifurcating tree suggested generally west to east dispersal trend. The data fit the isolation by distance (IBD) model well. Star-like clusters in haplotype network, significantly negative values of Fs statistics, and unimodal mismatch distributions all suggest recent demographic expansions in four areas. The results show that isolation, dispersal, bottleneck, and expansion jointly constitute the history of D. acutus. In a haplotype network, the excessive predominance of central haplotypes, few medium-frequency haplotypes, predominance (73.1%) of the singletons among the derived haplotypes, most of which are connected to the central haplotype by only one mutational step, unsymmetrical campanulate unimodal curve of mismatch distributions and leftwards shift of the peaks, all suggest that the whole D. acutus population is a young population with low genetic diversity. Based on the data, the first priority for conservation action should be given to the Huangshan unit.     

响应面法优化提取虎耳草总酚及其抑制五步蛇毒PLA2酶活性的相关性

为了探讨虎耳草总酚（TP）与抑制五步蛇毒中磷脂酶A₂（PLA₂）之间的关系,本研究通过单因素试验及Box-Behnken试验对虎耳草TP的提取条件与抑制五步蛇毒中PLA₂活性进行优化,考察各试验因素对超声辅助提取虎耳草TP得率及其对五步蛇毒PLA₂活性抑制率（PIR）的作用,并对虎耳草TP含量与PIR的相关性进行分析。结果表明,响应面法提取虎耳草TP和PIR的最佳工艺条件为：粒径为60目,料液比为1∶50,超声功率为250 W,超声时间为1.8 h。此优化工艺条件下虎耳草TP得率为（8.69±0.46） mg·g^-1,PIR为（40.91±0.21）%;相关性分析结果显示虎耳草TP含量与PIR具有极显著正相关（P<0.01）,说明TP可能为虎耳草抑制五步蛇毒中PLA₂活性的物质基础。     

Purification, cloning and sequence analyses for pro-metalloprotease-disintegrin variants from Deinagkistrodon acutus venom and subclassification of the small venom metalloproteases.

Acidic and basic hemorrhagic metalloproteases were purified from the venom of Deinagkistrodon acutus (from Fujian Province, China) using gel filtration and anion exchange on FPLC and reversed-phase HPLC. Their hemorrhagic activities and N-terminal sequences were characterized. Extensive screening of the venom gland cDNA after PCR amplification resulted in the identification and sequencing of a total of seven cDNA clones encoding the multidomain precursors of six acidic and one alkaline low molecular mass metalloproteases. Two of the precursors contain a processable disintegrin domain. Disintegrins of 5 kDa were also purified from the venom. The partial amino-acid sequences and molecular masses determined by electrospray ionization mass spectrometry of the purified proteins specifically match those deduced from two of the cDNA sequences. Moreover, phylogenetic analyses based on 30 complete sequences of low molecular mass venom metalloproteases revealed that they may be classified into three functional subtypes: acidic hemorrhagins, basic and moderate hemorrhagins, and nonhemorrhagic enzymes. Subtype-specific amino-acid substitutions in the C-terminal regions of the enzymes were highlighted to explore the structure-activity relationships of the enzymes.