目的: 研究微流成像颗粒分析技术(micro-flow imaging,MFI)在生化蛋白药物(注射用尿激酶)不溶性微粒检测中的应用,同时考察不同企业注射用尿激酶样品中不溶性微粒的现状和2种不同溶剂复溶后不溶性微粒的差异,探讨不溶性微粒的主要来源。方法: 采用注射用水和注射用灭菌生理盐水分别复溶注射用尿激酶,用MFI检测其中的不溶性微粒,进行方法学验证并与药典收载的光阻法比较。结果: MFI测定5 μm和15 μm微粒标准品的微粒计数的准确度(偏差)分别为11.6%和8.5%,精密度(RSD)分别为1.6%和3.8%,粒径的准确度(偏差)分别为2.5%和0.8%,精密度(RSD)分别为0.59%和0.10%;MFI测定样品不溶性微粒结果中,2~10 μm粒径范围的颗粒以蛋白颗粒为主,10 μm粒径以上的颗粒以非蛋白颗粒为主,其中非蛋白颗粒结果与光阻法检测的不溶性微粒结果相似;不同企业生产的注射用尿激酶中不溶性微粒存在较大差异,企业A样品的不溶性微粒相对较多;2种不同溶剂复溶样品后企业A和C不溶性微粒的结果基本无差异,企业B和D不溶性微粒的结果存在较大差异,样品采用注射用灭菌生理盐水复溶后测得的不溶性微粒相对较低。结论: 本文将MFI应用于生化蛋白药物(注射用尿激酶)不溶性微粒的检测,并且为探讨生化蛋白药物的不溶性微粒来源提供新思路;个别企业不溶性微粒结果接近限度,提示该企业需要加强研发,以提升产品质量可控性;同时企业可以根据样品中2~10 μm范围和10 μm粒径以上不溶性微粒的结果,从内源性和外源性成因分析不溶性微粒的主要来源;2种不同溶剂复溶影响部分企业样品的不溶性微粒结果,注射用灭菌生理盐水复溶的不溶性微粒较低,产生上述结果的原因有待进一步研究。

Objective: To evaluate the application of micro-flow imaging(MFI) in the detection of particulate matters in biochemical protein drugs(urokinase for injection), and to investigate the status of particulate matters in urokinase samples for injection from various manufactures, and the differences of particulate matters after resolution with two different solvents, and find out the main origin of particulate matters. Methods: The urokinase for injection was re-dissolved in either sterilized water for injection or sterilized normal saline for injection respectively, and the particulate matters were analyzed by MFI. The MFI methodology was validated, comparing with light obscuration method. Results: For 5 μm and 15 μm particulate standard detected by MFI, the counting accuracy(deviations) of particle count were 11.6% and 8.5%, precision(RSDs) were 1.6% and 3.8%, the accuracy (deviations) were 2.5% and 0.8%, precision(RSDs) were 0.6% and 0.1%. In the results of determination of particulate matters detected by MFI with the particle size of 2-10 μm were mainly proteinaceous, and the partide size of more than 10 μm were mainly non-proteinaceous. The result of non-proteinaceous particulate matters detected by MFI was similar to light obscuration method. The particulate matters in urokinase for injection produced by different manufactures were quite different, and the particulate matters for manufacturer A was more. There was no significant difference in the results of particulate matters after dissolved with two different solvents for samples from manufacturer A and C, and significant difference for samples from manufacture B and D. Particulate matters were less for samples re-dissolved with sterilized normal saline. Conclusion: MFI was first applied to detect particulate matters in biochemical protein drugs (urokinase for injection). It provides new ideas for exploring the source of particulate matters in biochemical protein drugs. The results of particulate matters in individual manufactures were close to the limit, suggesting that the manufactures need to strengthen research and development to improve product quality control. And manufactures can analyze the main sources of particulate matters from endogenous and exogenous cause according to the result of 2-10 μm range and particle size more than 10 μm.Reconstitution with different two solvents affected the results of particulate matters for some samples, and particulate matters are less for samples dissolved with sterilized normal saline. But the cause of the above results need further study.