Development and validation of a RP-HPLC method for simultaneous determination of cimetidine, metoprolol tartrate and phenol red for intestinal perfusion studies

IF 2.8 3区医学 Q2 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography B Pub Date : 2025-02-01 DOI:10.1016/j.jchromb.2025.124449

Meryem Ermis , Fatma Kir , Selma Sahin

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Abstract

A new reversed phase high-performance liquid chromatography (RP-HPLC) method, with a short analysis time and easy to apply, was developed for the simultaneous detection of cimetidine (CIM), metoprolol tartrate (MT) and phenol red (PR) for use in intestinal perfusion studies. The analysis was performed with phosphate buffer (pH 5.0, 12.5 mM)-acetonitrile mixture as mobile phase and C₁₈ column (Inertsil ODS-3; 5 µm, 4.6 × 250 mm) as stationary phase. Gradient analysis conditions were used and the acetonitrile ratio in the mobile phase varied from 10 to 50% in 10 min. Total run time for analysis was 10 min and the injection volume was 20 µL. Detection of compounds was performed at 207 nm. Under optimum HPLC conditions, retention times were 4.03 min for CIM, 6.99 min for MT and 8.49 min for PR. The method was validated according to ICH Q2 (R1) guideline for specificity, linearity, sensitivity, precision, accuracy, stability and robustness. Developed method was linear and determination coefficients of the calibration curves were 0.9993, 0.9991 and 1.0 for CIM, MT and PR, respectively. The limits of quantification were 6.20, 2.78 and 0.45 μg/mL for CIM, MT and PR, respectively. The precision and accuracy values of the developed analytical method met the ICH Q2 (R1) limits. The applicability of the method was demonstrated by preliminary in-situ intestinal perfusion studies. In conclusion, samples obtained from in-situ intestinal perfusion studies performed to examine the absorption/permeability of CIM, MT, and PR can be analyzed with the developed HPLC method.

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肠道灌注研究中同时测定西咪替丁、酒石酸美托洛尔和酚红的反相高效液相色谱方法的建立与验证。

建立了一种分析时间短、易于应用的反相高效液相色谱（RP-HPLC）方法，用于肠道灌注研究中同时检测西咪替丁（CIM）、酒石酸美托洛尔（MT）和酚红（PR）。以磷酸缓冲液(pH 5.0, 12.5 mM)-乙腈混合物为流动相，C18色谱柱(Inertsil ODS-3；5µm, 4.6 × 250 mm)作为固定相。采用梯度分析条件，流动相乙腈比为10 ~ 50%，分析时间为10 min，总运行时间为10 min，进样量为20µL。在207 nm处检测化合物。在最佳HPLC条件下，CIM的保留时间为4.03 min， MT的保留时间为6.99 min， PR的保留时间为8.49 min。方法的特异性、线性度、灵敏度、精密度、准确度、稳定性和鲁棒性按照ICH Q2 （R1）指南进行验证。所建立的方法是线性的，CIM、MT和PR的标定曲线决定系数分别为0.9993、0.9991和1.0。CIM、MT和PR的定量限分别为6.20、2.78和0.45 μg/mL。所建立的分析方法精密度和准确度符合ICH Q2 （R1）的要求。初步的原位肠道灌注研究证实了该方法的适用性。综上所述，通过原位肠道灌注研究来检测CIM、MT和PR的吸收/渗透性，所获得的样品可以用所开发的HPLC方法进行分析。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Journal of Chromatography B 医学-分析化学

CiteScore

5.60

自引率

3.30%

发文量

306

审稿时长

44 days

期刊介绍： The Journal of Chromatography B publishes papers on developments in separation science relevant to biology and biomedical research including both fundamental advances and applications. Analytical techniques which may be considered include the various facets of chromatography, electrophoresis and related methods, affinity and immunoaffinity-based methodologies, hyphenated and other multi-dimensional techniques, and microanalytical approaches. The journal also considers articles reporting developments in sample preparation, detection techniques including mass spectrometry, and data handling and analysis. Developments related to preparative separations for the isolation and purification of components of biological systems may be published, including chromatographic and electrophoretic methods, affinity separations, field flow fractionation and other preparative approaches. Applications to the analysis of biological systems and samples will be considered when the analytical science contains a significant element of novelty, e.g. a new approach to the separation of a compound, novel combination of analytical techniques, or significantly improved analytical performance.