| Inflammation is a very common and important basic pathological process,a rteriosclerosis,hypertension,cancer and other diseases are closely related to inflammation.Therefore,it has important practical significance to develop anti-inf lammatory drugs with high efficiency and low side effect.Arachidonic acid is a precursor to a variety of inflammatory substances.Under normal physiological conditions,arachidonic acid is mainly in the form of phospholipids in the cell membrane,no biological activity;a very small amount of arachidonic acid in free form.When the cell membrane is subjected to various stimuli,the free arachidonic acid is released from the cell membrane phospholipid pool and is converted into a biologically active metabolite through various routes.The cyclooxygenase pathway and the lipoxygenase pathway are two important pathways in the metabolism of arachidonic acid.Epoxidase in vivo in two forms exist,respectively,structural type cyclooxygenase-1 and inducible cyclooxygenase-2,the former involved in maintaining the body's normal physiological function;the latter in normal physiological circumstances Can not be detected,but in inflammatory cells and cancer cells in a large number of expression.5-lipoxygenase is widely present in animals and is not expressed in normal physiologicalconditions and is highly expressed in inflammatory and cancer-related cells.Cyclooxygenase-2/5-lipoxygenase double inhibitor can block two metabolic pathways,inhibit the production of inflammatory mediator prostaglandins and leukotrienes,to achieve good anti-inflammatory effect while avoiding cyclooxygenase inhibitors Triggering side effects.In this study,we used Autodock4.2 to calculate the binding free energy between 5-lipoxygenase and anti-5-lipoxygenase positive inhibitor,and establish the relationship between inhibitor binding free energy(?G)and bioactive p IC50 And the results were compared with the results of this model and molecular docking.The results showed that the anti-5-li poxygenase activity was the best combination of the anti-5-lipoxygenase activity.And the preferred compounds which have a dual inhibitory effect on 5-lipoxygenase and cyclooxygenase-2 are screened out.The preferred compounds with double inhibitory effect were validated by 5-lipoxygenase inhibitor model and colorimetric method,and the reddit sites were screened.The method of reverse phase high performance liquid chromatography(HPLC)was used to study the fingerprints of reddish and anti-inflammatory and anti-inflammatory sites,which provided the basis for the quality control of the medicinal preparations.The results showed that seven anti-5-lipoxygenase inhibitors(luteolin,piclocide,epicatechin,zileutone,lick dragon,CHEMBL367188,2-{2-Oxo-2-[1-T he relationship between free energy(Y)and 5-lipoxygenase inhibitory activity p IC50(X)was as follows: Y =-3.2983-0.72431 X,R2 = 0.93845,p = 0.014,?s = 0.29551,both P and standard errors are within acceptable limits.According to the model and the chemical composition of 123 kinds of Emilia sonchifolia and 5-LOX docking results,33 kinds of anti-5-lipoxygenase compounds were screened from the point of red.The results showed that the eight preferred compounds had double inhibitory effects on 5-lipoxygenase and cyclooxygenase-2,and the eight compounds were Caffeic acid,3,4-dihydroxyphenylacetic acid,4-hydroxyphenylacetic acid,drape,quercetin,luteolin,Tau.muurolol,alphacedrene.It was found that the carboxylic acid compounds had a good inhibitory effect on 5-lipoxygenase in the combination of preferred compound and5-lipoxygenase.Carboxylic acid can be well combined with 5-lipoxygenase,but the aliphatic and aromatic carboxylic acid and 5-lipoxygenase binding mechanism is different.The binding capacity of the aliphatic carboxylic acid to the5-lipoxygenase is mainly provided by the electrostatic force,and the main driving force for the binding of the aromatic carboxylic acid to the 5-lipoxygense van der Waals.Licofelone(ML-3000)was used as a positive control.Six preferred compounds(caffeic acid,3,4-dihydroxyphenylacetic acid,4-Hydroxyphenylacetic acid,rhamnol,quercetin,luteolin)were tested for 5-lipoxygenase inhibitory activity.The half inhibitory concentrations of these six preferred compounds were 0.8871?g/m L,3.613?g/m L,6.01?g/m L,9.132?g/m L,2.201?g/m L,2.105?g/m L,half of the positive inhibitor ML-3000 the inhibitory concentration was 10.851?g/m L.The inhibitory activity of 5-lipoxygenase was determined by colorimetric method with 75% ethanol,petroleum ether,ethyl acetate and n-butanol as solvent,the extraction site of Emilia sonchifolia prepared into 1mg/m L solution.The results showed that,except for the petroleum ether,the rest of the 5-lipox ygenase was inhibited.The inhibition rates of 5-lipoxygenase in the n-butanol Sites were 27.3±1%,58.2±1.8% and 54.7±0.6%,respectively.The effects of ethanol,ethyl acetate and n-butanol on the inhibition rate of 5-lipoxygenase were investigated.The results showed that with the prolongation of ultrasonic time,the effects of ethanol,ethyl acetate Ester site extraction rate gradually increased but anti-5-lipoxygenase activity gradually decreased,n-butanol site al most no effect.The HPLC fingerprints of 10 batches of Emilia sonchifolia and some ethyl acetate sites were further established,and the similarity evaluation was carried out by using the similarity evaluation system of Chinese medicine chromatographic fingerprints 2004 A.The results showed that the Emilia sonchifoliaand acetic acid The similarity of fingerprints at the ester sites was 0.89 and 0.91,respectively.By comparing the liquid retention time of the reference substance,seven peaks of the ethyl acetate site fingerprint were confirmed: the hydroxy benzene acetic acid,caffeic acid,p-hydroxy benzoic acid,quercetin,mignonette element,quercetin,rhamnetin. |
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