Qualitative analysis of the major constituents in traditional oriental prescription bang-poong-tong-sung-san by liquid chromatography/ ultraviolet detector/ion-trap time-of-flight mass spectrometry

초록

An advanced and reliable high performance liquid chromatography (HPLC)/ultraviolet detector (UV)/ion-trap timeof- flight (IT-TOF) mass spectrometry was developed for the simultaneous quantification of 19 marker compounds in Bangpoong- tong-sung-san (BPTS), a traditional oriental prescription. Various parameters affecting HPLC separation and IT-TOF detection were investigated, and optimized conditions were identified. The separation was achieved on a Capcell PAK C18 column (1.5 mm × 250 mm, 5 μm particle size) using a gradient elution of acetonitrile and water containing 0.1% formic acid at a flow rate of 0.1 mL/min. The column temperature was maintained at 40°C and the injection volume was 2 μL. IT-TOF system was equipped with an electrospray ion source (ESI) operating in positive or negative ion mode. The optimized electrospray ionization parameters were as follows: ion spray voltage, +4.5 kV (positive ion mode), or -3.5 kV (negative ion mode); drying gas (N2), 1.5 L/min; heat block temperature, 200°C. Automatic MSn (n = 1~3) analyses were carried out to obtain structural information of analytes. Elemental compositions and their mass errors were calculated based on their accurate masses obtained from a formula predictor software. The marker compounds in BPTS were identified by comparisons between MSn spectra from standards and those from extracts. Moreover, the libraries of MS2 and MS3 spectra and accurate masses of parent and fragment ions for marker compounds were constructed. The developed method was successfully applied to the BPTS extracts and identified 17 out of 19 marker compounds in the BPTS extracts.

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