Chemical Constituents And Bioactivitives Of Ilex Kudingcha

Ilex kudingcha C.J.Tseng is one of the species in the Ilex genus of the Aquifoliaceae family,which is mainly produced in Guangxi,Hainan and other places.Its dry leaves are cool,bitter,slightly sweet,and have the effect of mind refreshing and thirst relieving.According to reports,Ilex kudingcha mainly contains saponins,flavonoids,polyphenols,polysaccharides and volatile oil and other components.Ilex kudingcha has a variety of pharmacological activities,mainly in the aspects of hypoglycemic,hypolipidemic,anti-bacterial and anti-inflammatory.In this paper,the chemical constituents of the Ilex kudingcha were separated and purified by modern chromatographic technique,17 compounds were isolated from the ethyl acetate part?A part?of Ilex kudingcha leaves.Combined with the physicochemical properties and spectral data,a total of 14 structures were identified,of which ten triterpenoids:Kudinoside D?1?,Kudinoside H?2?,Kudinoside LZ₁₁?3?,Ilekudinoside O?4?,Ilekudinoside P?5?,3a,23-dihydroxyursan-12-en-28-acid?6?,Rotungenoside?7?Ulmoidol?8?,Ursolic Acid?9?,Oleanic acid?10?;Two flavonoids:Kaempferol-3-O-?-D-glucoside?11?,Kaempferol-3-O-?-D-rutinoside?12?;two phenolics:Chlorogenic acid?13?Isochlorogenic acid A?14?.Compound 6 was isolated from this plant for the first time.In this paper,the components of Ilex kudingcha were analyzed,and the experimental conditions of the analytical method were investigated and optimized.The results showed that Ilex kudingcha contains mainly saponins,flavonoids and polyphenols,and the content of polyphenols and flavonoids in the ethyl acetate part is higher than that in n-butanol part?B part?.The inhibitory effect of Ilex kudingcha leaves on xanthine oxidase?XO?had been studied in vitro and vivo.In vitro,inhibition of XO activity screening showed that the inhibitory effect of Ilex kudingcha leaves on the ethyl acetate part was better than n-butanol part,IC50 is 7.64 mg/mL.It is speculated that it may be related to the higher content of polyphenols and flavonoids in the ethyl acetate part of Ilex kudingcha.The model of hyperuricemia in mice induced by urokinase inhibitor potassium oxonate was used to study the effect of the ethyl acetate part of Ilex kudingcha leaves on serum XO activity in mice by spectrophotometry,serum uric acid levels in mice were determined by phosphotungstic acid method.The results showed that compared with the model group,serum uric acid levels in each group at the ethyl acetate part decreased,among which,the serum uric acid level in group was extremely significantly?P<0.01?,and the uric acid level in the middle group was significantly?P<0.05?.Compared with the model group,the serum XO levels in each group at the ethyl acetate part were decreased,and the XO activity in the high and middle groups at the ethyl acetate part was extremely significant?P<0.01?.Therefore,it is speculated that the decrease in serum uric acid levels in mice is related to the suppression of XO activity.This paper was conducted on chlorogenic acid,Isochlorogenic acid A,kaempferol-3-O-?-D-glucoside?Flavonoids A?,kaempferol-3-O-?-D-rutinoside?Flav-onoids B?preliminary discussion of the interaction mechanism with XO.The results of fluorescence spectra show that the chlorogenic acid and Isochlorogenic acid A can quench the endogenous fluorescence of XO at 298K and 310K,and the quenching type is a combination of static quenching and dynamic quenching.The binding constant is chlorogenic acid.>Isochlorogenic acid A,with 1 binding site;The thermodynamic equation indicates that the interaction between chlorogenic acid and XO are hydrogen bond and van der Waals force,and the interaction between Isochlorogenic acid A and XO is dominated by hydrophobic interaction force.The combination of kaempferol-3-O-?-D-glucoside,kaempferol-3-O-?-D-rutinoside and XO is a spontaneous process,which is mainly based on static quenching,accompanied by a certain dynamic quenching,and the binding constant kaempferol-3-O-?-D-glucoside>kaempferol-3-O-?-D-rutinoside has one binding site;thermodynamic equations indicate that The main interaction between kaempferol-3-O-?-D-glucoside and XO is hydrogen bonding and Van der Waals force.The main interaction between kaempferol-3-O-?-D-rutinoside and XO is the typical electrostatic force.Circular dichroism spectroscopy studies showed that the?-helix content of the four compounds combined with XO increased and the?-sheet content decreased.This may be due to the binding of the compound molecule to XO and affecting the hydrogen bond network structure of the protein.Furthermore,the?-helix acts as the main structure,its increased content may make the secondary structure of XO more compact,which is not conducive to the formation of active centers of the enzyme and the entry of the substrate into the active center,and thus the two types of compounds affect the activity of XO.Based on the binding constants,the steric hindrance of the drug molecule may greatly reduce its ability to bind to XO.In this paper,the chemical constituents and biological activities of Ilex kudingcha were studied.The results showed that 14 compounds were isolated and identified at the ethyl acetate part,and XO inhibitory activity was better at the ethyl acetate part,it can significantly inhibit XO activity and decrease uric acid levels in hyperuricemic mice.At the same time,the interaction mechanism of chlorogenic acid,Isochlorogenic acid A,kaempferol-3-O-?-D-glucoside and kaempferol-3-O-?-D-rutin-oside and XO was studied.This article not only provides the theoretical basis for the chemical constituents of Ilex kudingcha,but also the development and utilization of uric acid lowering drugs.