【作者】 霍星华；

【导师】 赵宝玉；

【作者基本信息】 西北农林科技大学 ， 临床兽医学， 2008， 硕士

【摘要】 镰形棘豆(Oxytropis falcate Bunge)藏药中称为“达夏”,系豆科(Leguminosae)棘豆属(Oxytropis)多年生草本植物。具有广阔的药用开发前景。对于镰形棘豆的化学成分的报道较多,但对其毒性的研究则未见报道。本试验运用化学成分的系统预试和薄层色谱技术,初步确定镰形棘豆中化学成分种类。通过致突变试验、亚慢性毒性试验及生殖试验来对镰形棘豆的毒性进行研究和评价,以证明镰形棘豆具有跟疯草植物一样的毒性提供有力的论据,也为镰形棘豆药物的开发提供更加深入的资料。研究获得以下结果:1.镰形棘豆化学成分的预试验及生物碱薄层色谱分析利用系统预试和专项预试相结合对镰形棘豆所含的化学成分进行了预试验,结果表明镰形棘豆中含有酚类、鞣质、生物碱、黄酮及其苷类、糖、多糖及其苷类、氨基酸及多肽、皂甙、有机酸、甾体和三萜类物质、挥发油;不含蒽醌及其苷类、内酯、香豆素及其苷类、强心甙、氰甙及脂肪族硝基化合物。对镰形棘豆生物碱进行系统提取,通过选择适宜的展开系统对各萃取段生物碱薄层色谱分析,结果表明,镰形棘豆中主要是吲哚里西啶类生物碱,且以大极性成分为主,同时也含有少量的中、小极性生物碱。氯仿部分有6种生物碱,乙酸乙酯部分有11种生物碱,正丁醇部分有5种生物碱。经与苦马豆素标准品对照,镰形棘豆各萃取部分均含有苦马豆素,并从正丁醇部分分离得到28 mg苦马豆素,提取率0.00187%。2.镰形棘豆正丁醇提取物致突变试验选用苦马豆素含量最高的正丁醇萃取物,配制成800 mg/kg、1 000 mg/kg、2 000 mg/kg、4 000 mg/kg四个梯度的水溶液,运用骨髓微核试验与精子畸形试验进行致突变性评价,结果表明,镰形棘豆正丁醇萃取物引起小鼠骨髓嗜多染红细胞的微核率分别为2.625‰、4.625‰、6.50‰和12.75‰;及精子畸形率为1.44%、1.99%、3.40%和3.475%;且存在剂量-效应关系,也就是说低浓度的时候对机体是安全的,剂量在2 000 mg/kg以下是安全范围,不引起微核及精子畸形发生率的升高;而高浓度时会引起突变发生率的显著升高,具有致突变性。3.镰形棘豆全草亚急性毒性试验通过给小鼠饲喂添加了镰形棘豆全草的饲料,对小鼠的临床症状观察,血液常规系数的检测,外周血T细胞及腹腔巨噬细胞吞噬功能的检查、内脏组织器官的剖检,研究镰形棘豆全草对小鼠的影响。结果表明,镰形棘豆全草对白细胞数及血红蛋白有一定的影响,而对红细胞、淋巴细胞的影响不显著。实验组小鼠的外周血T淋巴细胞ANAE阳性率分别为8.4%、15%和18.5%,与对照组比较差异显著(P<0.05),而对巨噬细胞的非特异性吞噬功能的影响则是低含量有促进作用;而高含量则有抑制作用,试验组巨噬细胞吞噬指数分别为0.86、0.47和0.32。镰形棘豆全草还能使小鼠各种器官细胞出现空泡变性。4.镰形棘豆全草生殖毒性试验镰形棘豆全草饲料喂养小鼠进行三段生殖毒性试验,结果显示镰形棘豆对怀孕动物体增重的有着显著的影响,草粉含量越高怀孕动物体增重的差异越显著(P<0.05),引起胚胎、骨骼和内脏畸形机率越大,说明了镰形棘豆与其它棘豆属疯草一样,具有损伤器官,使雄性动物减少甚至丧失性活动,致母畜流产,产弱胎、畸胎等生殖毒性。更多还原

【Abstract】 Oxytropis falcate Bunge also named Da Xia in Tibetan medical herd which is perennial herb that belongs to Oxytropis of Leguminosae. It has broad prospects for development of medicinal . The chemical composition of Oxytropis falcate Bunge have been reports more, but its toxicity study haven’t been reported. This research applied the chemical constituents of systemic pretesting and TLC techniques to identify types of chemical constituents from Oxytropis falcate Bunge. Through mutagenesis experiments, sub-chronic toxicity test and three of the reproductive test to research and evaluation the toxicity to prove that the Oxytropis falcate Bunge like locoweed providing strong support, as well as the development of drugs to provide more in-depth information. The experimental results were given as follows.1. Pre-experiment on Chemical Composition and TLC alkaloids analysis of Oxytropis falcate BungeThe chemical constitutes was preliminary tested though systematic and special pre-experiment. The results show that plenty of Phenols, tannins, alkaloids, flavones and its glycosides, saccharides polysaccharides and glycosides, amino acids and opypeptides, saponins, organic acids, sterides and triterpenes, volatile oils exist in Oxytropis falcate Bunge, does not contain anthraquinone and its glycoside, lactone, coumarin and its glycoside, cardiotonic glycoside, cyanophoric glycoside and aliphatic series nitrocompound.Through systematic extraction of alkaloids and selection eligible development system, TLC was used to analyze alkaloids that extracted by different solvent. The results indicated that the major alkaloids in Oxytropis falcate Bunge is indolizidine compounds, furthermore, there are minimum, neutral and maximum polarity indolizidine alkaloids and the maximum polarity alkaloids hold the first place. The extractum extracted by chloroform contains 6 species alkaloids, The fraction of EtOAc contains 11 species alkaloids, The extractum extracted by n-butanol contain 5 species alkaloids. In three extracted fractions of Oxytropis falcate Bunge all contain swainsonine compared with standard preparation, and obtain 28 mg Swainsonine from n-Butanol Extractive, extraction rate 0.00187%. 2. The Mutagenicity Test of n-Butanol Extractive of Oxytropis falcate Bunge in miceThe mutagenicity of three gradient aqueous solution 800 mg/kg, 1 000 mg/kg, 2 000 mg/kg, 4 000 mg/kg of n-Butanol extractive from Oxytropis falcate Bunge were evaluated by PCE micronucleus test and sperm abnormality test. The result showed that n-butanol extractant from Oxytropois falcate Bunge induced the micronucleus rate were 2.625‰, 4.625‰, 6.50‰and 12.75‰and sperm abnormality rate were 1.44%, 1.99%, 3.40% and 3.475%, moreover have a dose-response raletionship. It means that low-concentration extractant from Oxytropis falecate Bunge, which safety range was below 2 000 mg/kg, have not mutagenic effect and genetic toxity to micronucleus cells and sperms in mice, but high-concertration have mutagenic effect .3.Subacute Toxicity Test of Oxytropis falcate Bunge in MiceThis study fed mice on feedstuff which added Oxytropis falcate Bunge powder, passed through period of time to mice clinical symptom observation, the blood convention coefficient examination, detectioning peripheral blood T-lymphocyte and peritoneal macrophage swallow function, autopsy, studies the effect of Oxytropis falcate Bunge to mice. Result showed that white cell number and the hemoglobin have the certain influence, but the lymphocyte and granular cell’s influence were not different. The peripheral blood T-lymphocyte ANAE positive stain of trial groups were 8.4%, 15% and 18.5%, compared to control group has remarkable difference(P<0.05). This study observed that the peritoneal macrophage swallow function was enhanced with low Oxytropis falcate powder content foodstuff, but it was inhibit with high content, phagoindex of macrophagus were 0.86, 0.47 and 0.32. Furthermore, cytoplasmic vacuolation was observed in parenchymal cell of varoius organs that sample from Oxytropis falcate Bunge feeding mouse.4.Oxytropis falcate Bunge reproductive toxicity testOxytropis falcate Bunge feeding mice three reproductive toxicity test results showed that the Oxytropis falcate Bunge pregnancy weight gain of animals have a marked impact, the higher the plant contented the pregnancy mouse body weight gain more significant difference (P<0.05), from embryos, bone and offal deformity greater probability that the Oxytropis falcate Bunge with other Oxytropis grass, with organ damage, or even reduce male’s sexual activity, to the dams abortion, fetal weak production, teratogenic, and other reproductive toxicity.更多还原