Abstract

A novel analytical method, based on high-performance liquid chromatography with a UV (HPLC-UV) detection system for the sensitive detection of a genotoxic impurity (GTI) 5-amino-2-chloropyridine (5A2Cl) in a model active pharmaceutical ingredient (API) tenoxicam (TNX), has been developed and validated. The HPLC-UV method was used for the determination of GTI 5A2Cl in API TNX. The compounds were separated using a mobile phase composed of water (pH 3 adjusted with orthophosphoric acid): MeOH, (50:50: v/v) on a C₁₈ column (150 × 4.6 mm i.d., 2.7 μm) at a flow rate of 0.7 mL min⁻¹. Detection was carried out in the 254 nm wavelength. Column temperature was maintained at 40°C during the analyses and 10 μL volume was injected into the HPLC-UV system. The method was validated in the range of 1–40 μg mL⁻¹. The obtained calibration curves for the GTI compound was found linear with equation, y = 40766x − 1125,6 (R² = 0.999). The developed analytical method toward the target compounds was accurate, and the achieved limit of detection and limit of quantification values for the target compound 5A2Cl were 0.015 and 0.048 μg mL⁻¹, respectively. The recovery values were calculated and found to be between 98.80 and 100.03%. The developed RP-HPLC-UV analytical method in this research is accurate, precise, rapid, simple and appropriate for the sensitive analysis of target GTI 5A2Cl in model API TNX.

中文翻译：

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使用HPLC-UV系统测定活性药物成分中潜在的遗传毒性杂质5-氨基-2-氯吡啶

摘要

一种新型分析方法，基于具有UV（HPLC-UV）检测系统的高效液相色谱法，用于灵敏检测模型活性药物成分中的遗传毒性杂质（GTI）5-氨基-2-氯吡啶（5A2Cl）（ API）tenoxicam（TNX）已开发和验证。HPLC-UV法用于测定API TNX中的GTI 5A2Cl。在C ₁₈色谱柱（150×4.6 mm内径，2.7μm）上，使用由水（用正磷酸调节的pH 3）：MeOH（50:50：v / v）组成的流动相分离化合物0.7 mL min ^-1。在254nm波长下进行检测。在分析过程中，柱温保持在40°C，并将10μL体积注入HPLC-UV系统。在1–40μgmL ^-1的范围内验证了该方法。发现所获得的GTI化合物的校准曲线与方程y = 40766x − 1125,6（R ²  = 0.999）成线性关系。建立的针对目标化合物的分析方法准确，目标化合物5A2Cl的检测限和定量值的极限分别为0.015和0.048μgmL ^-1， 分别。计算回收率值，发现回收率在98.80和100.03％之间。本研究中开发的RP-HPLC-UV分析方法准确，精确，快速，简单，适合于API TNX模型中目标GTI 5A2Cl的灵敏分析。