The objective of this current study is to establish a single method for potency and related proteins analysis of human insulin formulations using reverse-phase high performance liquid (RP-HPLC) chromatography technique which was validated and verified for the potency analysis in insulin formulations. Chromatographic separation was achieved using an octadecylsilane (C-18) stationary phase and a mobile phase composed of 55% (v/v) buffer (0.2 M sodium sulfate in water, {pH 2.3}) and 45% (v/v) acetonitrile. Detection was performed by UV detector at 214 nm with a flow rate of 1 ml/min and an injection volume of 20 µL, at 40°C. Currently there are separate methods available in Indian Pharmacopoeia for analysis of Potency and Related proteins in human insulin. We have validated a single method where quantitation of potency and related proteins can be performed in the same run. The method validation exhibited linearity over the concentration range of 0.08–4.5 mg/ml (r=0.999) with limit of detection of 0.094 mg/ml The accuracy of the method was 99–102.8%. Thus, it is proposed that both potency and related proteins in insulin formulations can be precisely evaluated using a single run thus saving the time and cost for quality analysis of insulin preparations both at manufacturing and regulatory laboratories which in turn will increase the market availability of such standard quality insulin preparations for public health use.

中文翻译：

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少即是多：验证综合 rh-胰岛素分析的单一方法

本研究的目的是建立一种使用反相高效液相 (RP-HPLC) 色谱技术对人胰岛素制剂的效力和相关蛋白质进行分析的单一方法，该方法已针对胰岛素制剂的效力分析进行了验证和验证。使用十八烷基硅烷 (C-18) 固定相和由 55% (v/v) 缓冲液（0.2 M 硫酸钠水溶液，{pH 2.3}）和 45% (v/v) 乙腈组成的流动相实现色谱分离。使用 UV 检测器在 214 nm 处进行检测，流速为 1 ml/min，进样量为 20 µL，温度为 40°C。目前，印度药典中有单独的方法可用于分析人胰岛素的效力和相关蛋白质。我们已经验证了一种可以在同一运行中对效力和相关蛋白质进行定量的单一方法。方法验证在 0.08–4.5 mg/ml (r=0.999) 的浓度范围内表现出线性，检测限为 0.094 mg/ml。该方法的准确度为 99–102.8%。因此，建议使用单次运行即可精确评估胰岛素制剂中的效力和相关蛋白质，从而节省制造和监管实验室对胰岛素制剂进行质量分析的时间和成本，从而增加此类制剂的市场可用性。供公共卫生使用的标准质量胰岛素制剂。