Thummala Veera Raghava Raju, Lanka Mohana Krishna
Se Pu. 2015 Oct;33(10):1051-8. doi: 10.3724/sp.j.1123.2015.04043.
A sensitive, stability-indicating gradient reverse phase ultra performance liquid chromatographic method has been developed for the quantitative estimation of nebivolol impurities in active pharmaceutical ingredient (API) and pharmaceutical formulation. Efficient chromatographic separation was achieved on an Acquity BEH C18 column (100 mm x 2.1 mm, 1.7 μm) with mobile phase of a gradient mixture. The flow rate of the mobile phase was 0.18 mL/min with column temperature of 30 degrees C and detection wavelength of 281 nm. The relative response factor values of (R*)-2-( benzylamino)-1-((S*)-6-fluorochroman-2-yl) ethanol ((R x S*) NBV-), (R)-1-((R)-6-fluorochroman-2-yl)-2-((S)-2-((S)-6-fluoro-chroman-2-yl)-2-hydroxyethyl-amino) ethanol ((RRSS) NBV-3), 1-(chroman-2-yl)-2-(2-(6-fluorochroman-2-yl)-2-hydroxyethyl amino) ethanol (monodesfluoro impurity), (S)-1-((R)-6-fluorochroman-2-yl)-2-((R)-2 (S*)-6-fluoro-chroman-2-yl)-2-hydroxyethylamino) ethanol hydrochloride ((RSRS) NBV-3) and (R*)-1-((S*)-6-fluorochroman-2-yl)-2-((S*)-2-((S*)-6-fluoro-chroman-2-yl)-2-hydroxyethylamino) ethanol ((R* S* S* S*) NBV-2) were 0.65, 0.91, 0.68, 0.92 and 0.91 respectively. Nebivolol formulation sample was subjected to the stress conditions of acid, base, oxidative, hydrolytic, thermal, humidity and photolytic degradation. Nebivolol was found to degrade significantly under peroxide stress condition. The degradation products were well resolved from nebivolol and its impurities. The peak purity test results confirmed that the nebivolol peak was homogenous and pure in all stress samples and the mass balance was found to be more than 98%, thus proving the stability-indicating power of the method. The developed method was validated according to International Conference on Hormonization (ICH) guidelines with respect to specificity, linearity, limits of detection and quantification, accuracy, precision and robustness.
已开发出一种灵敏的、指示稳定性的梯度反相超高效液相色谱法，用于定量测定活性药物成分（API）和药物制剂中奈必洛尔的杂质。在Acquity BEH C18柱（100 mm×2.1 mm，1.7μm）上，以梯度混合的流动相实现了高效的色谱分离。流动相流速为0.18 mL/min，柱温为30℃，检测波长为281 nm。（R*）-2-（苄基氨基）-1-（（S*）-6-氟苯并二氢吡喃-2-基）乙醇（（R×S*）NBV-）、（R）-1-（（R）-6-氟苯并二氢吡喃-2-基）-2-（（S）-2-（（S）-6-氟苯并二氢吡喃-2-基）-2-羟乙氨基）乙醇（（RRSS）NBV-3）、1-（苯并二氢吡喃-2-基）-2-（2-（6-氟苯并二氢吡喃-2-基）-2-羟乙氨基）乙醇（单去氟杂质）、（S）-1-（（R）-6-氟苯并二氢吡喃-2-基）-2-（（R）-2（S*）-6-氟苯并二氢吡喃-2-基）-2-羟乙氨基）乙醇盐酸盐（（RSRS）NBV-3）和（R*）-1-（（S*）-6-氟苯并二氢吡喃-2-基）-2-（（S*）-2-（（S*）-6-氟苯并二氢吡喃-2-基）-2-羟乙氨基）乙醇（（RSSS）NBV-2）的相对响应因子值分别为0.65、0.91、0.68、0.92和0.91。对奈必洛尔制剂样品进行了酸、碱、氧化、水解、热、湿度和光解降解等强制条件试验。发现奈必洛尔在过氧化物胁迫条件下会显著降解。降解产物与奈必洛尔及其杂质得到了很好的分离。峰纯度测试结果证实，在所有强制样品中奈必洛尔峰均均匀且纯净，质量平衡大于98%，从而证明了该方法的指示稳定性的能力。所开发的方法根据国际协调会议（ICH）指南在特异性、线性、检测限和定量限、准确度、精密度和稳健性方面进行了验证。
