Unexpected retention behavior of baicalin: Hydrophilic interactionlike properties of a reversed-phase column

Magda, Balázs and Szabó, Pál Tamás and Imre, Timea and Márta, Zoltán (2015) Unexpected retention behavior of baicalin: Hydrophilic interactionlike properties of a reversed-phase column. Journal of Pharmaceutical and Biomedical Analysis, 111. pp. 119-125. ISSN 0731-7085

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Abstract

tThe original aim of this study was to develop a method for the determination of baicalin from membranevesicles. The unconventional chromatographic separation (“inverse gradient elution” on a reversed phasecolumn) was due to a lucky chance, which is detailed and discussed in this study. The validated liquidchromatography-tandem mass spectrometry (LC–MS/MS) method is proved to be sensitive, rapid andselective. Chromatographic separation was performed on a Zorbax SB-C8 column (250 mm × 4.6 mm,i.d.; 5 �m) with 0.1% formic acid in water and methanol by linear gradient elution. Quantification ofbaicalin was determined by multiple reaction monitoring (MRM) mode using electrospray ionization(ESI). The calibration curve was linear (r = 0.9987) over the concentration range from 1 to 1000 nM.The coefficient of variation and relative error of baicalin for intra- and inter-assay at three quality con-trol (QC) levels were 2.0–10.2% and −6.1 to 6.7%, respectively. The lower limit of quantification (LLOQ)for baicalin was 1 nM (0.446 ng/ml), without preconcentration of the sample. This method was subse-quently applied to vesicular transport assays of baicalin in membrane vesicles successfully. The developedmethod can open up new area of research in the chromatographic separation of flavonoids and theirglucuronides.

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