Pablo Miranda, Analía Castro, Luisina Rodríguez Grassi, Danilo Davyt, Marcelo Hill, Mercedes Segovia, Alvaro W Mombrú, Helena Pardo
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引用次数: 0
摘要
根据 ICH 分析验证指南,我们报告了一种测定离子通道调节剂 Bay K8644 的普通 C18 柱稳定性指示等度反相高效液相色谱法。质谱法还初步鉴定了两种主要的强制降解产物和一种次要杂质。流动相为 50/50 乙腈/缓冲液混合物,流速为 2 mL/min。Bay K8644 的平均保留时间为 3.030 分钟。在 274 nm 波长下,0.10-1.40 μg/mL 范围内线性关系良好(r = 0.9998)。检测和定量的分析限分别为 16.56 ± 1.04 纳克/毫升和 55.21 ± 3.48 纳克/毫升。准确度和精密度研究显示结果良好(95-105%)。温度和流速的±3%变化评估了鲁棒性。为评估 Bay K8644 的稳定性,采用了五种不同的压力条件。只有碱性和光解处理产生了降解产物,这两种情况都在 4 分钟的总运行时间内得到了正确解析。总之,我们开发了一种快速、简便、灵敏、准确、精确、可靠且具有稳定性指示的方法来检测/定量 Bay K8644,并初步确定了其主要杂质/强制降解产物的特征。
Reverse Phase HPLC Methodology for the Determination of Bay K8644.
Following ICH guidelines for analytical validation, we report a common C18 column stability indicating isocratic reverse phase high performance liquid chromatography method for the determination of the ion channel modulator Bay K8644. Two main forced degradation products and a minor impurity were also tentatively identified by Mass Spectrometry. The mobile phase consisted of a 50/50 acetonitrile/buffer mixture at a flow rate of 2 mL/min. Mean retention time for Bay K8644 was 3.030 minutes. Excellent linearity (r = 0.9998) was achieved in the range 0.10-1.40 μg/mL at 274 nm wavelength. Analytical limits were 16.56 ± 1.04 ng/mL for detection and 55.21 ± 3.48 ng/mL for quantitation respectively. Accuracy and precision studies showed good results (95-105%). Robustness was assessed by varying ±3%, both temperature and flow rate. Five different stress conditions were applied to assess Bay K8644's stability. Only basic and photolytic treatments yielded degradation products, both correctly resolved in a total runtime of 4 minutes. In conclusion, we developed a fast, simple, sensitive, accurate, precise, reliable and stability indicating method for detecting/quantifying Bay K8644, and tentatively characterized its main impurities/forced degradation products.
期刊介绍:
The Journal of Chromatographic Science is devoted to the dissemination of information concerning all methods of chromatographic analysis. The standard manuscript is a description of recent original research that covers any or all phases of a specific separation problem, principle, or method. Manuscripts which have a high degree of novelty and fundamental significance to the field of separation science are particularly encouraged. It is expected the authors will clearly state in the Introduction how their method compares in some markedly new and improved way to previous published related methods. Analytical performance characteristics of new methods including sensitivity, tested limits of detection or quantification, accuracy, precision, and specificity should be provided. Manuscripts which describe a straightforward extension of a known analytical method or an application to a previously analyzed and/or uncomplicated sample matrix will not normally be reviewed favorably. Manuscripts in which mass spectrometry is the dominant analytical method and chromatography is of marked secondary importance may be declined.