Department of Pharmaceutical Chemistry, College of Pharmacy, Taif University, Al-Hawiah 21974, Taif, Saudi Arabia.
>Pharmaceutical Analytical Chemistry Department, Faculty of Pharmacy, Beni-Suef University, Alshaheed Shehata Ahmad Hegazy St., 62514 Beni-Suef, Egypt.
J AOAC Int. 2020 Jun 1;103(3):747-754. doi: 10.1093/jaocint/qsz015.
Amiloride hydrochloride (AM) is a potassium sparing diuretic, while hydrochlorothiazide (HCZ) is the protype of thiazide diuretics. The combining of the studied drugs exhibits a synergistic effect. Moreover, HCZ prevents the potassium depletion side effect caused by AM.
Two accurate and precise simultaneous chromatographic separation methods were promoted and investigated to quantify AM, HCZ, official impurities of HCZ (cholorothiazide and salamide), and the official impurities of AM (methyl 3, 5-diamino-6-chloropyrazine-2-carboxylate).
The components of the quintuple mixture were quantified by two methods. The first method was high-performance thin layer chromatography (HPTLC), where exemplary separation was achieved on silica gel HPTLC F254 plates at the stationary phase using ethyl acetate-ethanol-ammonia solution (8 + 2 + 0.2, v/v) as a developing system. Scanning of bands at 273 nm was done. The second method was a reversed-phase chromatography (RP-HPLC) method using C18 (4.6 × 100 mm) column and mobile phase comprising 0.1% phosphoric acid solution-acetonitrile (90 + 10, v/v) with UV determination at 273 nm. Adjustment of the flow rate at 1 mL/min and pH at 3.6 was performed.
Regarding RP-HPLC, optimum separation of the quintuple mixture was achieved within just five minutes. According to HPTLC, symmetric and sharp peaks were separated on the resulted chromatogram. Validity of the introduced methods was investigated by applying international conference on harmonization (ICH) guidelines.
The methods were successfully applied for assays of the studied drugs in their pure and tablet forms. No significant difference was revealed through application of statistical comparison between results of the suggested methods and those of the reported method regarding both accuracy and precision.
盐酸阿米洛利（AM）是一种保钾利尿剂，而氢氯噻嗪（HCZ）是噻嗪类利尿剂的原型。研究药物的组合表现出协同作用。此外，HCZ 可防止 AM 引起的钾耗竭副作用。
提出并研究了两种准确、精密的同时色谱分离方法，以定量分析 AM、HCZ、HCZ 的官方杂质（氯噻嗪和磺胺酰胺）以及 AM 的官方杂质（甲基 3,5-二氨基-6-氯哒嗪-2-羧酸酯）。
使用两种方法对五重混合物的成分进行定量分析。第一种方法是高效薄层色谱法（HPTLC），在固定相上使用硅胶 HPTLC F254 板，展开系统为乙酸乙酯-乙醇-氨溶液（8+2+0.2，v/v），实现了典型的分离。在 273nm 处扫描条带。第二种方法是反相色谱（RP-HPLC）法，使用 C18（4.6×100mm）柱，流动相为 0.1%磷酸溶液-乙腈（90+10，v/v），在 273nm 处进行紫外检测。调整流速为 1mL/min，pH 值为 3.6。
对于 RP-HPLC，只需五分钟即可实现五重混合物的最佳分离。根据 HPTLC，在得到的色谱图上分离出对称且尖锐的峰。通过应用国际协调会议（ICH）指南验证了所提出方法的有效性。
该方法成功地应用于研究药物的纯品和片剂的测定。通过应用统计学比较，所提出的方法与已报道的方法在准确度和精密度方面均无显著差异。
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