血凝仪-APTT法测定肝素的效价

目的: 建立一种基于血液凝固分析仪测定活化部分凝血活酶时间(APTT)的肝素生物测定法(简称血凝仪-APTT法)。 方法: 将高、中、低浓度的肝素钠标准品和供试品分别与血浆混和,用血液凝固分析仪测定各含药血浆的活化部分凝血活酶时间(APTT),结果进行对数转换后,按量反应平行线3.3法计算供试品的效价。 结果: 对数凝血时间与肝素钠的浓度在0.7~3 IU·mL^-1范围内线性关系良好(r=0.9965);当供试品的测得效价与估计效价的比值在80%~120%范围内时,血凝仪-APTT法的测定结果准确可靠,重复性(RSD=1.45%,n=6)及中间精密度(RSD=1.61%,n=6)良好。以兔全血法、羊血浆法和血凝仪-APTT法分别测定同一批肝素钠供试品的效价(IU·mg^-1,n=3),结果依次为207.56±6.87,205.59±3.62,202.79±2.08,无显著性差异(P>0.05);兔全血法和血凝仪-APTT法的可信限率(%,n=3)分别为8.51±0.77和2.63±0.21,具有显著性差异(P<0.01)。使用不同仪器测定样品效价的结果基本一致。 结论: 血凝仪-APTT法与兔全血法和羊血浆法的效价测定结果一致,可信限率比兔全血法更低。该法操作简便,客观准确,重复性好,精密度高,适用于不同仪器,可以用于肝素钠生物效价的测定。

Objective: To establish a new method for heparin assay based on the activated partial Thromboplastin time by coagulometer (abbreviated as coagulometer-APTT method). Methods: Prepare the heparin standard and the article of high, middle and low concentration, mixed them with plasma separately, and determinate the APTT of the drug-containing plasma by coagulometer. Results: Within the range of 0.7-3 IU·mL^-1, the heparin sodium concentration and the log clotting time shows a good linear relationship (r=0.9965); within the range of 80% -120% of the ratio of P_T and estimated potency of the test article, coagulometer-APTT method was accurate and reliable with good reproducibility (RSD=1.45%, n=6) and intermediate precision (RSD=1.61%, n=6). The results (IU·mg^-1, n=3) of rabbit blood method, sheep plasma method and coagulometer-APTT method were 207.56±6.87, 205.59±3.62 and 202.79±2.08, no significant differences (P> 0.05); the FL% (%, n=3) of rabbit blood method and coagulometer-APTT method were 8.51±0.77 and 2.63±0.21, with significant difference (P<0.01). The results of coagulometer-APTT method by different coagulometers were basically consistent. Conclusion: Coagulometer-APTT method coincided with rabbit blood method and sheep plasma method in the potency result, and its FL was lower than the rabbit blood methods. The results demonstrated that coagulometer-APTT method is simple, objective, accurate, reproducible, precise and applying to different instruments. It could be used for the assay of heparin.

1 ChP(中国药典).2010.Vol Ⅱ(二部):Appendix(附录)Ⅻ D:108
2 USP 33-NF 28.2010.Vol 2:3316
3 JPⅩⅤ. 2006. Heparin sodium: 721
4 BP. 2008. Vol Ⅳ:Appendix ⅩⅣ J.,A340
5 BP. 2008. Vol Ⅳ:Appendix ⅩⅣ J,A340
6 ChP(中国药典).2010.Vol Ⅱ(二部):Appendix(附录)ⅩⅣ,130
7 QIAO De-shui(乔德水),MAO Pu(毛璞),GU Ke-xiu(顾克秀).Determination of heparin potency by coagulometer(肝素钠效价的血凝度计测定法).Chin J Pharm(中国医药工业杂志),1993,24(7):317
8 WANG Bi-song(王碧松),LI Na(李娜),XIAO Yuan(肖渊),et al.The differences in the biological assay of heparin on different sources plasma(不同来源试验系应用于肝素钠生物效价测定的差异考察).Chin J Pharm Anal(药物分析杂志),2010,30(4):691
9 SONG Yuan-zhi(宋愿智),ZHAO Wei(赵威),LI Bo(李波).Heparin determining method & current condition of research(肝素钠测定方法及研究近况).Chin New Med(中国新医药),2003,2(8):39
10 HE Hang-yong(贺航咏),YANG Yuan-Huan(杨媛华).The clinical application of 4 coagulations(凝血4项的临床应用).Chin J Med(中国医刊),2008,43(1):15