Federal University of São João del-Rei, Divinópolis, Minas Gerais, Brazil.
Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Biomed Chromatogr. 2021 Mar;35(3):e4999. doi: 10.1002/bmc.4999. Epub 2020 Oct 27.
Midazolam (MDZ) is routinely employed as a marker compound of cytochrome P450 3A (CYP3A) activity. Despite the many HPLC-UV methods described to quantify MDZ in plasma, all of them use acetonitrile (ACN) or a mixture of methanol-isopropanol as organic solvent of the mobile phase. Since the ACN shortage in 2008, efforts have been made to replace this solvent during HPLC analysis. A simple, sensitive, accurate and repeatable HPLC-UV method (220 nm) was developed and validated to quantify MDZ in rat plasma using methanol instead. The method was applied during a herb-drug interaction study involving Maytenus ilicifolia, a Brazilian folk medicine used to treat gastric disorders. Plasma samples were alkalinized and MDZ plus alprazolam (internal standard) were extracted with diethyl ether. After solvent removal, the residue was reconstituted with methanol-water (1:1). The analyte was eluted throughout a C column using sodium acetate buffer (10 mm, pH 7.4)-methanol (40:60, v/v). The precision at the lower limit of quantification never exceeded 19.40%, and 13.86% at the higher levels of quality control standards, whereas the accuracy ranged from -19.81 to 14.33%. The analytical curve was linear from 50 to 2,000 ng/ml. The activity of the hepatic CYP3A enzymes was not affected by the extract.
咪达唑仑（MDZ）通常被用作细胞色素 P450 3A（CYP3A）活性的标记化合物。尽管已经有许多 HPLC-UV 方法用于定量检测血浆中的 MDZ，但它们都使用乙腈（ACN）或甲醇-异丙醇混合物作为流动相的有机溶剂。自 2008 年 ACN 短缺以来，人们一直在努力在 HPLC 分析中替代这种溶剂。本研究开发并验证了一种简单、灵敏、准确和重现性好的 HPLC-UV 方法（220nm），该方法使用甲醇代替 ACN 作为流动相，用于定量检测大鼠血浆中的 MDZ。该方法应用于涉及巴西民间药物 Maytenus ilicifolia 的草药-药物相互作用研究中，该药物用于治疗胃部疾病。将血浆样品碱化，用二乙醚提取 MDZ 和阿普唑仑（内标）。除去溶剂后，用甲醇-水（1:1）重新溶解残留物。分析物通过 C 柱洗脱，使用乙酸钠缓冲液（10mm，pH7.4）-甲醇（40:60，v/v）。定量下限处的精密度从未超过 19.40%，在更高质量控制标准的水平下为 13.86%，而准确度范围为-19.81 至 14.33%。分析曲线在 50 至 2000ng/ml 之间呈线性。肝 CYP3A 酶的活性未受提取物影响。
