Development of a quantitative analysis method for the 12 marker compounds in Palmijihwang-hwan, a herbal formula, using a reversed-phase C18 column and an amino column by HPLC

Abstract

The aim of this study was to determine quantitatively the 12 marker compounds in Palmijihwang-hwan (PJH) using a high performance liquid chromatography-photodiode array detector. The separation of marker compounds was performed on a reversed-phase C₁₈ column for gallic acid, 5-hydroxymethylfurfural (5-HMF), morroniside, loganin, paeoniflorin, mesaconitine, benzoic acid, coumarin, cinnamic acid, cinnamaldehyde, and paeonol, and an amino (NH₂) column for allantoin. The correlation coefficient of marker compounds was ≥0.9993, which means good linearity. The limit of detection and limit of quantification values were in the ranges from 0.01–0.25 μg mL⁻¹ and 0.02–0.78 μg mL⁻¹, respectively. The within-day precision was 0.06–2.95% and the between-day precision was 0.10–4.44% over five consecutive days. The recovery of marker compounds ranged from 97.20 to 106.36%, with RSD values <3.5%. The repeatability was <2.1% of the RSD value. The quantification results indicated that the quantities of the 12 marker compounds differed between a water extract and commercial granules of PJH. Cinnamaldehyde and paeonol were particularly difficult to determine in commercial granules because of its low LOQ. The evaluation of the Pearson coefficient and principal component analysis showed a clear discrimination between a PJH water extract and commercial granules of PJH. The analytical method established was precise, accurate, and reproducible for evaluating the quality of PJH.