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Rapidly and accurately screening histidine decarboxylase inhibitors from Radix Paeoniae alba using ultrafiltration–high performance liquid chromatography/mass spectrometry combined with enzyme channel blocking and directional enrichment technique

Guizhou Hu,Lu Wang,Xinqi Li,Jin Qi

Histidine Decarboxylase (HDC), an unique enzyme responsible for the synthesis of histamine, which is an important mediator in allergy. Inhibition of HDC activity to decrease histamine production is one way to alleviate allergic symptoms. Traditional Chinese medicines (TCMs) with reported anti-allergy effect is one of important source to search for natural HDC inhibitor. Ultrafiltration combined with high–performance liquid chromatography/mass spectrometry (UF–HPLC/MS) is an effective method for screening HDC inhibitor from TCMs. Nevertheless, false-positive and false-negative results caused by the non-specific binding and the neglection of the trace active compounds are major problems in this method. In this study, an integrated strategy that combined UF–HPLC/MS with enzyme channel blocking (ECB) technique and directional enrichment (DE) technique was developed to seek natural HDC inhibitors from Radix Paeoniae alba (RPA), and at the same time, to reduce false-positive and false-negative results. HDC activity was detected to determine the validity of the screened compounds by RP–HPLC–FD in vitro . Molecular docking was applied to assay the binding affinity and binding sites. As a result, three compounds were screened from low content components of RPA after the DE. Among them, two non-specific compounds were eliminated by ECB, and the specific compound was identified as catechin, which has obvious HDC inhibition activity with IC 50 0.52 mM. Furthermore, gallic acid (IC 50 1.8 mM) and paeoniflorin (IC 50 >2 mM) from high content components of RPA were determined having HDC inhibitory activity. In conclusion, the integrated strategy of UF–HPLC/MS combined with ECB and DE technique is an effective mode for rapid and accurate screening and identification of natural HDC inhibitors from TCMs.