In vitro screening and mechanism study of Ganoderma lucidum compounds with protective activity against neuronal oxidative stress injury
[Objective]Screening for compounds in Ganoderma lucidum that had neuroprotective activity and explo-ring their corresponding mechanisms of action,to provide scientific basis for the research and development of functional foods,health products and drugs with the effects of preventing and treating Alzheimer disease(AD).[Method]This study established a cellular model based on β-amyloid protein(Aβ)induced oxidative damage,and screened the antioxi-dant activity of G.lucidum alcohol extracts preliminarily assessed by the 1,1-diphenyl-2-trinitrophenylhydrazine(DPPH)free radical scavenging method,iron ion reduction/antioxidant capacity test,and 2,2-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)ammonium salt(ABTS)free radical scavenging method,and evaluated their neuronal protective activity.Furthermore,the 4 triterpenoids compounds related to antioxidants in G.lucidum alcohol extracts were used to verify their activity and to explore their related mechanisms through surface plasma resonance technology.[Result]The results of in vitro physicochemical primary screening and cellular level screening experiments on DPPH and ABTS free radical scavenging,iron ion reduction/antioxidant capacity indicated that 6 G.lucidum alcohol extracts(108,161,106-2,171,173 and 109)had strong antioxidant protective activity for neurons,among which samples 108,109 and 106-2 showed the best protective effects against oxidative stress damage in rat adrenal pheochromocytoma(PC12)cells.The G.lu-cidum alcohol extracts could exert neuroprotective effects by alleviating cell cycle disruption,inhibiting early cell apopto-sis,enhancing superoxide dismutase(SOD)activity and glutathione(GSH)content,and suppressing the increase of malondialdehyde(MDA)content.The average contents of 4 triterpenoids(ganoderal A,ganoderic acid B,ganoderic acid C2,and ganoderic acid LM2)in G.lucidum alcohol extracts numbered 108,106-2 and 109 were relatively high.Ac-tivity validation demonstrated that these 4 compounds all exhibited obvious reparative effects against cellular oxidative stress damage.Among them,ganoderal A and Aβ1-42 showed strong affinity,with a KD value of 15.62 μmol/L,and could inhibit the binding of Aβ1-42 to RAGE through competitive binding with Aβ1-42.[Conclusion]The antioxidant neuronal ac-tivity of G.lucidum ethanol extracts is achieved through the synergistic effects of 4 compounds:ganoderic acid C2,ganoderic acid B,ganoderic acid LM2 and ganoderal A.Ganoderal A can alleviate or inhibit the cytotoxic effects of Aβ1-42 by competitively binding with RAGE.G.lucidum has the potential for developing into a functional product for the preven-tion and treatment of AD.
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