Preparation, Purification And In Vivo Antioxidant Activity Of Ginkgo Pepitdes From Ginkgo Nuts

Ginkgo nuts were used as raw material and peptides were prepared from Ginkgo nutswith Neutral protease and Flavor enzyme in turn. In the single-factor experiments ofNeutral protease, four conditions, namely temperature, pH, enzyme concentration andsubstrate concentration, were optimized in term of degree of hydrolysis (DH). On that base,an orthogonal experiment (L9(34)) was conducted to optimize the hydrolysis conditions ofNeutral protease. The results showed that substrate concentration, reaction temperature andinitial pH were the most significant variables that influenced DH. The highest DH wasobtained when reaction temperature, initial pH, enzyme amount and substrate concentrationwere45℃,7.0,1.8mg/mL and0.02g/mL, respectively. Later, the resulting solution wasfurther hydrolyzed with Flavor enzyme. Initial pH, temperature, time and enzymeconcentration were optimized in term of DH. Finally, an orthogonal experiment (L9(34))was conducted to obtain the optimal enzymatic hydrolysis conditions of Flavor enzyme,and enzyme concentration, initial pH, temperature and time were used as parameters. Theresults indicated that the DH was greatly improved when enzyme concentration, reactiontemperature, initial pH, and hydrolysis time were9.0mg/mL,50℃,7.0and6h,respectively. The DH was found to be25.03％at optimized conditions.The ginkgo peptides prepared by Neutral protease and Flavourzyme hydrolysis thenwere subjected to ultrafiltration with different molecular weight cutoff (MWCO)membranes (2.5,10and20kDa) for fractionation and purification. Four peptide groupswith molecular weight of>20,10～20,2.5～10and <2.5kDa were obtained finally.Yields of each group were0.69、0.13、3.67and2.74%(w/w), respectively. The solubleprotein yield was8.76%. In addition, the peptide group with molecular weight of10～20kDa showed the highest foaming capacity and foam stability. However, the group of2.5～10kDa exhibited the highest oil-absorption capacity comparing with the others.The in vivo anti-oxidative capacity of ginkgo peptides was also investigated. Theresults indicated that the peptide group with molecular weight of <2.5kDa had the bestperformance of increasing the activities of total antioxidant capacity (T-AOC), glutathioneperoxidase (GSH-PX), superoxide dismutase (SOD), inhibition of hydroxyl radical anddecreasing malondialdehyde (MDA) levels in blood, liver, kidney and heart, comparingwith the control group and vitamin C group. The anti-oxidative capacity wasdose-dependent. Meanwhile, the peptide group with molecular weight of <2.5kDa showedthe best immunostimulating property as well. The findings suggested that ginkgo peptides possess strong in vivo antioxidant capacities and are potential to be developed as healthfood.