Development and validation of UPLC-TQD/MS method for quantification of six bioactive constituents in Thai herbal Benjakul formula

Abstract

Introduction: Thai herbal Benjakul formula (BKF) is a Thai traditional medicine for balancing and nourishing body elements in unhealthy persons. It is composed of five pungent plants, namely fruits of Piper retrofractum Vahl., roots of Piper sarmentosum Roxb., stems of Piper wallichii (Miq.) Hand.-Mazz., roots of Plumbago indica L., and rhizome of Zingiber officinale Roscoe. There was no efficient method for quantification of the bioactive constituents in BKF using UPLC-TQD/MS. Objective: The objective of the study was to establish a method for quantification of some constituents of BKF, namely pellitorine (PE), piperine (PI), piperlonguminine (PLM), plumbagin (PG), 6-gingerol (GI), and 6-shogaol (SG) using ultra-performance liquid chromatography coupled with a triple quadrupole mass spectrometer (UPLC-TQD/MS) and determine six bioactive constituents amount in Ayurved Siriraj® (AVS) and five brands of BKF randomly selected from the market. Materials and Methods: Six bioactive constituents were determined using UPLC-TQD/MS. The chromatographic separation was performed on a Waters ACQUITY UPLC® HSS T3 column using gradient elution with water and methanol. The method was fully validated in terms of the limit of detection, the lower limit of quantification precision, and recovery. Results: The method was a good linearity (R2 > 0.995) in the range of 1–75 ng/mL for all six bioactive constituents. The precision, accuracy, and recovery are in an acceptable range. The maximum concentrations of PE, PI, PLM, PG, GI, and SG in all samples were 0.342 ± 0.009%, 1.396 ± 0.020%, 0.080 ± 0.001%, 0.055 ± 0.003%, 0.165 ± 0.003%, and 0.175 ± 0.003%, w/w, respectively. Conclusion: This is the first study for quantification of the six standards in BKF using UPLC-TQD/MS. This method is useful in the quality control of BKF and can be used routinely in the manufacturing unit.