Qiu Jinshu, Lee Hans, Zhou Chengfeng
Department of Analytical Science, Amgen Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA.
J Chromatogr A. 2005 May 6;1073(1-2):263-7. doi: 10.1016/j.chroma.2004.10.045.
A simple and sensitive HPLC method for quantitative determination of guanidine in high salt and protein matrices was developed. The HPLC system consisted of an Agilent 1100 pump with an online degasser, a UV detector, an autosampler, and Dionex CS 14 cation-exchange guard (4 mm x 50 mm) and analytical (4 mm x 250 mm) columns. The mobile phase was 3.75 mM methanesulfonic acid (MSA) with a flow rate of 1 mL/min. The other analysis parameters were: 50 microL injection volume, 195 nm UV detection, and 21 min runtime. The limit of quantitation (LOQ) for guanidine HCl was determined to be 0.25 mg/L and the standard curve ranged from 0.25 mg/L to 10 mg/L. Sample preparation was required for the samples containing high protein concentrations. Proteins were removed by centrifuging a sample in a 30 K NanoSep centrifugal filter at 15,300 x g for 20 min. The method could determine guanidine accurately in sample matrices containing up to 200 mM sodium ion or up to 50 mM potassium ion. The method can be used for clearance testing of guanidine in biopharmaceutical products.
开发了一种用于在高盐和蛋白质基质中定量测定胍的简单灵敏的高效液相色谱法。高效液相色谱系统由配备在线脱气机的安捷伦1100泵、紫外检测器、自动进样器以及戴安CS 14阳离子交换保护柱（4 mm×50 mm）和分析柱（4 mm×250 mm）组成。流动相为3.75 mM甲磺酸（MSA），流速为1 mL/min。其他分析参数为：进样体积50 μL、紫外检测波长195 nm、运行时间21 min。盐酸胍的定量限（LOQ）确定为0.25 mg/L，标准曲线范围为0.25 mg/L至10 mg/L。对于含有高蛋白浓度的样品需要进行样品制备。通过在30 K NanoSep离心过滤器中以15300×g离心样品20分钟来去除蛋白质。该方法可以在含有高达200 mM钠离子或高达50 mM钾离子的样品基质中准确测定胍。该方法可用于生物制药产品中胍的清除测试。
