川芎及其中成药抗凝血作用测定方法的研究

目的以延长活化部分凝血活酶时间(APTT)为指标,建立定量测定川芎抗凝血作用的方法,评价川芎及其中成药的质量。方法川芎先后用乙醇和水定量提取,以总提取物为供试品,体外测定延长APTT值。家兔心脏取血,制备血浆,加入APTT试剂后,测定凝结时间。以APTT延长率为抗凝血活性的评价指标,用阿魏酸钠标定川芎的延长APTT活性。根据量反应平行线法(2.2)计算川芎的抗凝血活性。并测定了9份川芎药材、饮片及中成药的抗凝血活性。结果阿魏酸钠和川芎总提取物均具有显著的抗凝血活性,且可靠性检验结果成立。阿魏酸钠的给药质量浓度(1~5 mg/m L)与其APTT延长率呈良好的线性关系(r=0.995 5)。供试品重复测定抗凝血活性的RSD为9.34%(n=6),可信限率为15.98%(n=6)。不同川芎样品的抗凝血活性不同,5份川芎药材的抗凝血效价分别为5.431~7.620 U/g,川芎饮片及川芎酒炙饮片分别为5.910、3.017 U/g,通脉颗粒和血府逐瘀丸分别为14.516、29.035 U/g。结论建立的方法能准确测定川芎药材、饮片及其中成药的抗凝血活性并评价其质量。

Determination of anticoagulating bioactivity in Chuanxiong Rhizome and related Chinese patent medicines

Objective To activate partial thromboplastin time (APTT) indicating the coagulation state in intrinsic coagulation system and the efficacy of anticoagulant drugs, and develop a bioassay method to quantify anticoagulating bioactivity in Chuanxiong Rhizome and related Chinese patent medicines for quality assessment. Methods Chuanxiong Rhizome powder was extracted in ethanol and water, respectively. The mixed extract was used as sample to quantify the level of extended APTT in vitro. The blood was taken from the heart of rabbit. The agglomerating time was determined after adding APTT reagent in plasma. The prolongation rate of APTT was chosen as biomarker for anticoagulating bioactivity. Sodium ferulate was chosen as reference. The amount of anticoagulating bioactivity was quantified in Chuanxiong Rhizome extract by the Amount reaction of parallel line analysis (2.2) method. Moreover, the amounts of anticoagulating bioactivity were quantified in the nine Chuanxiong Rhizome samples including Chuanxiong Rhizome raw materials, decoction pieces, and related Chinese patent medicines. Results Both sodium ferulate and Chuanxiong Rhizome extract showed significant anticoagulating bioactivity (P < 0.01). The reliability test for quantifying the level of anticoagulating bioactivity in solidum ferulate and Chuanxiong Rhizome extract was passed through, and the measured value was valid. The correlation coefficient was 0.995 5 (n=5) between the concentration of solidum ferulate in the range of 1-5 mg/mL and their rates of anticoagulating bioactivity. The RSD for the amounts of anticoagulating bioactivity was 9.34% (n=6) by six replicated tests with the confidence limit rate of 15.98% (n=6). The amounts of anticoagulating bioactivity were significant difference among various types of Chuanxiong Rhizome samples, i.e. 5.431-7.620 U/g for five Chuanxiong Rhizome raw materials, 5.910 and 3.017 U/g for Chuanxiong decotion pieces and processed slice with yellow wine, 14.516 and 29.035 U/g for Tongmai Granules and Xuefu Zhuyu Pills. Conclusion The developed method can accurately quantify the level of anticoagulating bioactivity in Chuanxiong Rhizome raw materials, decoction pieces and related Chinese patent medicines, and assess their quality.