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At‐line LC‐QTOF‐MS micro‐fractionation of Derris scandens (Roxb.) Benth, coupled to radioassay for the early identification of PDE5A1 inhibitors
Author(s) -
Bhandari Samjhana,
Nuengchamg Nitra,
Chaichamg Nattiya,
Seasong Tongchai,
Ingkaninan Kornkanok,
Temkitthawon Prapapan
Publication year - 2019
Publication title -
phytochemical analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 72
eISSN - 1099-1565
pISSN - 0958-0344
DOI - 10.1002/pca.2895
Subject(s) - chemistry , chromatography , fractionation , high performance liquid chromatography , mass spectrometry , column chromatography
Abstract Introduction Chromatographic techniques coupled with bioassays are popularly used for the detection of bioactive compounds in natural products. In this study phytochemicals responsible for showing Phosphodiesterase type 5 (PDE5) inhibitory activity in Derris scandens were studied using at‐line method. Objective The objective of this study was to develop an at‐line liquid chromatography quadrupole time‐of‐flight mass spectrometry (LC‐QTOF‐MS) micro‐fractionation method for rapid separation and identification of PDE5A1 inhibitors in 95% ethanolic extract of D. scandens . Methodology Initially, the correlation between LC‐MS and PDE5A1 inhibitory activity was studied using three concentrations of 1:1 mixture of sildenafil and derrisisoflavone A; PDE5A1 inhibitors. The mixture was separated by high‐performance liquid chromatography (HPLC) column and the eluent was split into two flows in the ratio of 1:9. The major part was collected in a 96‐well plate, in each well consecutively every 30 s. The minor part was fed into an electrospray ionisation (ESI)‐QTOF‐MS system. After subsequent solvent removal, the collected micro‐fractions were subjected to radioassay to determine PDE5A1 inhibition. Results The result showed, PDE5A1 inhibitory activities of the micro‐fractions were observed in a dose response manner and found to be in agreement with an off‐line study. Similarly, 95% ethanolic extract of D. scandens was subjected to the at‐line LC‐QTOF‐MS micro‐fractionation developed, resulting in separation and tentative identification of 25 compounds with PDE5A1 inhibitory activity. Most of the compounds contained prenylated isoflavone skeleton. Additionally, the active micro‐fractions also showed selectivity on PDE5A1 over PDE6 and PDE1B. Conclusion Our results demonstrated that the at‐line coupled LC‐QTOF‐MS micro‐fractionation with PDE5A1 inhibitory assay is a valuable tool for identifying PDE5A1 inhibitors from complex extracts.