Study On Separation And Purification, Structure, Organic Se Speciation, And Antioxidant Activity Of Tea Selenoproteins

Selenoproteins are not only the predominant form of organic selenium,but also the mainly biological and functional speciation of the essential trace element of selenium,which possess numerous bioactivities,such as antioxidant,anticancer,and immunity enhancement.But so far there has yet been no report both at home and abroad regarding the study on organic seleniu species and antioxidant activity of tea selenoproteins(TSPs),extracted from Se-enriched tea.In this paper,TSPs were studied in terms of their extraction technology,structure,organic selenium speciation,and antioxidant activity.The main results of this project are as follows:(1)The separation and purification of TSPs:(1)TSPs were extracted by ultrasonic-assisted extraction with different extracting solvent(water,salt,ethanol and alkali),and the yield of TSPs by ultrasonic-assisted extraction using salt was 44.67%.Though slightly below to the yield of protein by alkali extraction and ethanol extraction,TSPs extracted by salt extraction own superior apparent property and higher antioxidant activity;(2)The optimal extraction conditions of ultrasonic-assisted extraction using salt were optimized by single factor experiment and response surface method as following: the concentrate of NaCl 0.75 mol/L,liquid-solid ratio 40 mL/g,ultrasonic time 120 min,extracting temperature 50℃.In this condition,the yield of protein was 63.68%.(2)The structure of TSPs: the structure of TSPs fractions(TSPs1、TSPs2、TSPs3)purified by DEAE Sepharose FF was studied by UV spectrum,IR spectrum,and circular dichroism spectrum.The results show that: I the UV spectrum of TSPs owns strong peak at 211 nm and weak acromion at 280 nm,which is the characteristic absorption peak of selenoproteins;II circular dichroism spectrum of TSPs shows that the major secondary structure of TSPs1、TSPs2 were random coil and α-helix respectively,and the major secondary structures of TSPs3 were α-helix and β-sheet.(3)The organic speciation of TSPs: the organic speciation of TSPs was studied by HPLC-ICP-MS and the results show that: I the organic speciation of TSPs consist of SeCys and SeMet,and SeCys is the predominant form of organic selenium in TSPs;II as the supplement of organic selenium,crude TSPs are superior to purified TSPs.(4)The antioxidant activity of TSPs: I TSPs possesses strong antioxidant activity,and their order: salt-extract TSPs > water-extract TSPs > ethanol-extract TSPs > alkali-extract TSPs;II TSPs exhibit obvious antioxidant activity,the antioxidant activity of TSPs is superior to that of normol tea proteins,and the intensity of antioxidant activity is in direct proportion to its content of selenium;III mechanism of antioxidant activity: the antioxidant activity of TSPs derived from its antioxidant element of selenium and high content of antioxidant amino acids.This project pave the way for the study and development of functional food and Se supplementation using TSPs as raw materials and play an important role in preventing and alleviating the deficiency of Se in our country.