Studies On Inhibition Effects Of Extracts From Sargassum Thumbergii And Bryopsis Plumose On The Xanthine Oxidase

Xanthine oxidase （EC.1.2.3.2） is a kind of nonspecific enzyme, catalyzetransformation of hypoxanthine to xanthine and xanthine to uric acid. Xanthineoxidase play a key role in purine metabolism and it is a significant enzyme in theformation of uric acid. The high content of uric acid in body would inducehyperuricemia, and then gout, cardiovascular disease and renal insufficiency.Researches show that the aetiological agent of hyperuricemia mostly attribute toabnormal activity of xathine oxidase, result in accumulation and deposition of uricacid. In recent years, most of clinical medication for treatment of hyperuricemiaindicate side effect. So the focus of current research is extract efficient xathineoxidase inhibitor from algae.In the study, we choose most common seaweeds by the seaside in Qingdao toresearch xathine oxidase inhibitory——Sargassum thumbergii and Bryopsis plumose.We determine the best extraction condition of the water-soluble extract from the twoalgae through orthogonal test. Using organic solvent extracting method, differentcomponents of two seaweeds are obtained. With kinetic analysis method, theinhibition mechanism and inhibition type are determined. The type of associatedinhibition of water-soluble extraction and allopurinol is determined.We confirm theantioxidant capacity of water-soluble extraction from the two seaweeds. Results are asfollows:1．The best extraction condition of the water-soluble extract from the two algae isdetermined through orthogonal test. The reaction system is5mL volume,using0.24mmol/L xanthine as substrate,the pH of the buffer being6.8, the enzymeconcentration is4μg/mL,and the reaction temperature and time being37℃and1min. First using single factor experiment to confirm the optimum extractioncondition of single factor.And then determine the best extraction condition through the orthogonal experiment. Sargassum thumbergii optimum extraction condition are40℃constant temperature water extraction,7h extracting time,70％concentrationof enthanol,and5min sonicated time. Bryopsis plumose optimum extraction conditionare45℃constant temperature water extraction,4h extracting time,70％concentration of enthanol, and5min sonicated time.2．Determine the inhibition mechanism and inhibition type of every componentextracted from the two seaweeds. Research inhibition effect of ethyl acetatecomponent, water-soluble and ploysccharide from Sargassum thumbergii on thexanthine oxidase. Results: ethyl acetate component have no inhibition effect on theenzyme, the inhibition type of water-soluble effect on XO is anti-competitivereversible inhibition, inhibition constant Ki=0.11, IC₅₀=0.42mg/mL. The inhibitiontype of ploysccharide effect on XO is competitive reversible inhibition, Ki=1.41,IC₅₀=0.69mg/mL. Research inhibition effect of ethyl acetate component, andwater-soluble from Bryopsis plumose. on the xanthine oxidase. Results: both the twocomponent have inhibitory on XO. The inhibition type of water-soluble effect isnoncompetitive reversible inhibition, Ki=1.46, IC₅₀=0.31mg/mL. The inhibition typeof ethyl acetate component effect is competitive reversible inhibition, Ki=0.16,IC₅₀=0.61mg/mL. Both water-soluble components of the two seaweeds havesignificant inhibition activity.3．Determine the type of associated inhibition of water-soluble components ofthe two algaes. Allopurinol is clinical drug for treatment of gout. Results: Cotoxicitycoefficient of allopurinol and water-soluble extracted from Sargassum thumbergii is3.8, the type is additive effect. Cotoxicity coefficient of allopurinol and water-solubleextracted from Bryopsis plumose is-3.2, the type also is additive effect.4．Confirm the antioxidant capacity of water-soluble extraction from the twoseaweeds. The scavenging rate of the every concentration of water-soluble extractfrom Sargassum thumbergii on·OH more than50％, and increased with concentrationof inhibitor,the scavenging rate rise. The scavenging rate of the every concentrationof water-soluble extract from Sargassum thumbergii on O₂^-·, and in a certain range, inhibitor concentration is positively related with scavenging rate. Water-solubleextract from Bryopsis plumose have scavenging effect on·OH, and indicate dosedependent effect, and have no scavenging effect on O₂^-·.