Screening Of Anti-Cancer Active Components In Arnebiae Euchroma Based On Affinity Ultrafiltration-LC-MS Technique

The root of Arnebia euchroma(Royle)Johnst.is used as medicine and is mainly distributed in Xinjiang.Modern pharmacology has proved that it has good curative effect on many intractable diseases such as cancer,AIDS and psoriasis,and found that Arnebiae Radix has significant antioxidant,anti-inflammatory,antibacterial,antiviral and anti-cancer activities;At present,there are many studies on the anti-cancer effects of naphthoquinones in Arnebiae Radix.But the anti-cancer effects of Arnebiae Radix the excavation of anti-cancer active components is not enough.Therefore,in this experiment,the chemical composition of Arnebiae Radix was fully characterized by using Arnebiae Radix as the experimental material,and then the LC-MS was used to rapidly screen the anticancer active components in Arnebiae Radix,and verify its activity through molecular docking,to further clarify the basic components of anti-cancer substances in Arnebiae Radix and to clarify these anti-cancer mechanism of active ingredients lays the foundation,and at the same time has important theoretical significance and application value for the development of Arnebiae Radix.The main research contents and results are as follows:(1)The full characterization of the chemical constituents of Arnebiae Radix.This experiment established an effective strategy for the detection of chemical constituents of Arnebiae Radix in negative ion mode based on UHPLC-Q-Exactive Orbitrap MS.A total of 188 compounds were identified by full-scan MS,MS~2databases(mz Vault and mz Cloud),diagnostic fragment ions,retention times,and references,of which 114 were reported for the first time from Arnebiae Radix.The results of this experiment laid the foundation for in-depth understanding of the active ingredients and quality control of Arnebiae Radix.Meanwhile,this strategy can be widely applied to chemical characterization of different samples.(2)Affinity ultrafiltration-LC-MS technique based on estrogen receptor as the target was used to screen the active components of Arnebiae Radix.By optimizing the concentration of crude extract,receptor concentration and incubation time,the optimal affinity-ultrafiltration system for ERαand ERβestrogen receptors was determined.The conditions were as follows:10μL estrogen receptor and5 mg/m L The crude extracts of Arnebiae Radix were incubated at 37°C for 60 mins,and the results showed that 14 compounds with ERαand 10 compounds with ERβwere screened by the affinity ultrafiltration system.(3)The activity was verified by molecular docking technology.The molecular docking evaluation of 21 estrogen receptor specific ligands was carried out by Auto Dock 4.2 software,and the selective ligand compound estradiol was docked to prove the rationality of parameter setting.Finally,potential ligands with better activity are identified,and the possible mechanism of action is understood through binding force and binding mode.It can be seen from the binding energy of the interaction that all show certain binding activities,wherein the binding energies of tryptophan,salvianolic acid J,1-methoxyacetylshikonin,deoxyshikonin and Sativan to ERαare respectively.-9.3,-8.5,-7.9,-7.1,-7.1 Kcal/mol,the binding energies of Rosmarinicacid,Quercetin,Salvianolic acid C,Rutin,and Isoquercitrin to ERβwere-8.4,-8,-7.1,-6.4,-6.3 Kcal/mol,respectively,which further proved that the The compounds screened by the filtration method have good binding activity to estrogen receptors.