Study On Anti-melanin Synthesis And Anti-Photoaging Activity Of Brown Algae Phlorotannins

Phlorotannins are a class of secondary metabolites that only exist in marine brown algae.In recent years,phlorotannins have attracted much attention due to their extensive biological activity.In this study,phlorotannins were extracted from Sargassum fusiforme and Sargassum thunbergii,and the antioxidant activity,anti-aging activity,anti-melanin production and photoprotective effects of phlorotannins were studied,so as to provide theoretical support for the application of brown algae polyphenols in cosmetics.Firstly,we take the S.fusiforme as raw materials,in 40%absolute ethanol and temperature of 70℃ extraction twice to get two brown algae crude extract,and the crude materials were purified by XDA-7 to get S.fusiforme phlorotannins(SFPR).Simultaneously extract the crude fraction of S.fusiforme with ethyl acetate to get ethyl acetate phase polyphenol(SFPE)and water phase polyphenol(SFPW).The antioxidants,anti-aging and anti-tyrosinase activities of four phlorotannins:SFPA,SFPR,SFPE and SFPW were tested in vitro.In vitro antioxidant results showed that SFPR and SFPE have powerful antioxidant activity.The anti-aging activities of four polyphenols were tested with elastase and collagenase.The results indicated that SFPA,SFPR and SFPE can effectively inhibit the activity of elastase;all four phlorotannins have strong inhibition of collagenase activity,of which SFPR has the strongest inhibitory activity.The inhibitory effects of SFPR and SFPE on mTYR monophenolase and diphenolase were measured with tyrosinase as the object.The results show that both phlorotannins can significantly inhibit the activity of mTYR,the monophenolase IC50 is 2.022 and 3.20μg/mL,and the diphenolase IC50 is 3.892,0.8895 μg/mL,respectively.Secondly,the inhibition mechanism and type of inhibition of bisphenolase by SFPR and SFPE were studied.SFPR is a mixed reversible inhibition,its inhibition constant(KI)is 5.54μg/mL,and the dissociation constant KIS is 11.25μg/mL.The inhibition mechanism of SFPE on diphenolase is irreversible inhibition.Based on the inhibition of mTYR activity in vitro,the impact of SFPR on melanin production in mouse B16F10 cells was studied.The results show that SFPR can effectively inhibit the melanin content and the activity of tyrosinase in cells.Further research shows that SFPR down-regulates MITF expression by inhibiting the CREB signaling pathway,thereby inhibiting the expression of TYR family proteins,thereby inhibiting melanin synthesis in B16F10 cells.Secondly,we get phlorotannins isolated from Sargassum thunbergii(STPE).We have demonstrated that STPE has a photoprotective effect on UVB-induced photoaging damage in L929 cells.STPE increases the activity of SOD in L929 cells after UVB radiation.The concentration of STPE at 40μg/mL can significantly reduce the content of MDA in cells and effectively relieve cell damage caused by ultraviolet radiation.At the same time,STPE can effectively reduce the content of ROS in cells and relieve DNA damage.Further,we studied the impact of STPE on UVB-induced mRNA expression of inflammatory factors and MMPs in L929 cells.The results indicated that STPE inhibited the expression of inflammatory factors.However,STPE has no significant inhibitory effect on the expression of MMPs.In addition,we studied the repair impact of STPE on zebrafish skin damage.The results show that STPE has a significant repair effect on zebrafish skin damage and can effectively remove excess ROS in zebrafish.The above results indicate that STPE has a significant anti-photoaging effect,and the mechanism is related to scavenging oxygen free radicals.Finally,we evaluate the effect of SFPR and STPE on the activity of 3D skin model was determined by MTT method.The results show that 100 mg/mL SFPR and STPE has no irritation to the skin.