Abstract
A fast and sensitive reverse phase ion-pair chromatographic method was reported for the separation and quantification of 4-aminophenylacetic acid (Actarit) and its related compounds. The compounds were separated using a Kromasil C18 column (5 μm, 4.6 × 25cm) with a mobile phase containing 70% methanol and 1% Tetrabutylammonium bromide in water flowing at 1.0 mL/min, and were detected using a UV detector operating at 245nm. Actarit was completely separated from its related compounds that included its synthetic starting materials and degradations. The retention times were 3.1, 4.0, 9.4, 21.4, and 25.8min for blank solvent, 4-aminophenylacetic acid, Actarit, 4-nitrophenylacetonitrile, and 4-nitrophenylacetic acid, respectively. Three known and one unknown compound (retention time 13.7minute) were detected after Actarit was boiled in acidic, neutral, and basic conditions, each for two hours. The linear response between peak area (A) and Actarit concentration (c) over the range of 0.5 - 2.5 μg/mL was The detection limit at signal-to-noise ratio two was 5.08 ng/mL. The recovery was 98.64 – 101.72% (n=5) (within day) and 99.60 – 101.86% (n=5) (day to day). The relative standard deviations were 0.87 – 1.77% (n=5) (within day) and 0.51 – 1.92% (n=5) (day-to-day), respectively. Good correlation was found between this HPLC method and a spectrophotometric method when measuring the contents of Actarit in three batches of commercial tablets. The contents were 97.32, 99.20, and 97.58% when measured by this HPLC method, and 97.76, 100.43, and 98.02% by a spectrophotometric method. Good separation, linearity, recovery, and accuracy implied that the chromatographic method is suitable for quantitative analysis of Actarit.
Published Version
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