摘要
中药委陵菜Potentilla chinensis Ser.属多年生草本植物,约有500多种,主要分布在北半球的温带和寒带,只有有少数几种分布在南半球。它又名翻白菜,白头翁,蛤蟆草,天青地白,为蔷薇科(Rosaceae)植物委陵菜Potentilla chinesisSer.的全草,《中国药典》2000,2005年版收载,其性寒,味苦,具有清热解毒,凉血止痢的作用。常用于赤痢腹痛、久痢不止、痔疮出血、痈肿疮毒等。经临床应用调查,天津蓟县中医医院和天津中医药大学附属医院应用委陵菜治疗糖尿病,取得了较好的疗效。本课题组前期研究表明委陵菜降糖作用的主要成分为山萘酚-3-β-D-(6-O-trans-p-桂皮酰基)葡萄糖,称为委陵菜黄酮(trans-tiliroside),其降血糖作用强于盐酸苯乙双胍。该化合物与市售双胍类、磺脲类、α-糖苷酶抑制剂、噻唑烷二酮类以及GLP-1R激动剂肽类药物在结构上存在根本差异,则可能存在不同的作用机制,是具有较高开发价值的活性先导化合物。
为了进一步评价委陵菜黄酮的药用价值和在体内的药效基团,以便于构效关系研究和结构优化,本文进行委陵菜黄酮的初步药代动力学研究。
第一部分研究委陵菜黄酮在动物离体肠组织的吸收情况,通过缚管翻转肠囊法,取SD大鼠的小肠肠段制成肠囊,考察委陵菜黄酮在高、中、低三个浓度下在离体肠组织中的吸收行为。实验结果表明,在一定浓度范围内,随着黄酮溶液浓度的升高,黄酮在小肠内的吸收速率常数增大。吸收过程符合一级消除动力学模型,符合被动转运的特征。同时发现,部分委陵菜黄酮在肠囊内水解成两个分解产物。经高效液相色谱法分析鉴定,这两种分解产物为肉桂酸和山萘酚-3-O-β-D-葡萄糖苷。
第二部分在离体肠囊实验研究的基础上,测定了委陵菜黄酮在大鼠体内的血药浓度-时间曲线。对SD大鼠以尾静脉注射方式给药后,测定其在低、中、高三个给药浓度(分别为10mg/kg、20mg/kg、40mg/kg)下动物体内的血药浓度-时间曲线,并使用药动学软件3P97计算药时相关药动学参数。实验结果表明,在雌雄大鼠体内,黄酮的血药浓度不存在显著性差异。大鼠尾静脉注射10mg/kg、20mg/kg、40mg/kg三个剂量的黄酮对照品溶液后,血浆药时曲线符合三房室模型,分布相半衰期t_(1/2α)分别为103.90min、15.59min、9.49min,消除相半衰期t_(1/2β)分别为30580.98min、2045.70min、894.84min,说明大鼠静注给药后,委陵菜黄酮在体内分布较快但消除慢。
除此之外,我们还对委陵菜黄酮在大鼠体内的分解产物进行了研究。结构分析推测其可能的分解产物后,通过比较大鼠血浆供试液与可能分解产物的HPLC谱图,可知在血液中有检测到这三种物质——肉桂酸、山萘酚与3-O-β-D-葡萄糖苷。
The Chinese folk medicine Potentilla chinensis Bge(Rosaceae) is a perennial herb, and there are about 500 species worldwide,most of which distributed in northern temperate and boreal,only few are in the Southern Hemisphere.It is also called "Fan-bai-cai","Bai-tou-weng","Ha-ma-cao" and "Tian-qing-di-bai",listed in Chinese Pharmacopoeia 2000 and 2005.As a folk medicine,Potentilla chinensis has has been used to reduce swelling and staunch bleeding,and has anti-inflammatory and detoxification properties.In addition,P.chinensis revealed the therapeutic effect for controlling blood glucose and improving glucose tolerance of body in clinic.Our preliminary study showed that the main constituent of its hypoglycemic effect is kaempferol-3-O-β-D-(6-O-trans-p-coumaroyl)glucopyranside,which is named potentilla flavone(trans-tiliroside),and its hypoglycemic effect is better than phenformin hydrochloride.And its structure is different from biguanides, sulfonylureas,α-glycosidase inhibitors,thiazolidinedione,GLP-1R peptide agonist drug which indicates that there is a different anti-diabetic mechanism and is an active leading compound worth studying.
In order to study potentilla flavone's medicinal value and pharmacophore,we do the preliminary pharmacokinetic's study on potentilla flavone to explore its structure-activity relationship and optimize its structure.
During the first part,we studied the absorption of potentilla flavone in animal's intestinal tissue in vitro.Everted-gut technique was utilized to study the absorption properties of potentilla flavone at three different concentration range.The results show that the absorption of potentilla flavone increased by their concentration rising in a definite concentration range,as well as a concentration-depent.And the absorption of potentilla flavone fits to linear absorption model and the passive transport features.In addition,some potentilla flavone was hydrolysed into two hydrolysates.The two main contabolites of potentilla flavone in isolated rat small intestine were determined to be cinnamic acid and kaempferol-3-O-β-D-flavonoid glycoside.
On basis of preliminary study on animal's intestinal tissue in vitro,we studied the blood concentration of potentilla flavone in vivo in second part of this paper.After tail vein injection to SD rats at three different concentration range,we determinate the concentration of potentilla flavone through HPLC and caculate the pharmacokinetic parameters.After intravenous injection of potentilla flavone at the dose of 10,20, 40mg/kg to rats,the average plasma-concentration datum were fitted to three-compartmental models(weight 1/c/c).The distribution half-life time(t_(1/2α)) and elimination half-life time(t_(1/2β)) of the three dose was 103.90 and 30580.98min,15.59 and 2045.70min,9.49 and 894.84min,respectively.All of these indicated the rapid distribution and a slow elimination.
Besides,we do research on potentilla flavone's hydrolysates in vivo.Analysis potentilla flavone's structure and speculate its hydrolysate's possible structure,we compare the HPLC spectrums and make sure that there are three hydrolysatescinnamic, kaempferol and kaempferol-3-O-β-D-flavonoid glycoside.
引文
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