Analysis On Characteristics And Bioactivity In Organic Selenium From Se-enriched Shiitake Mushrooms

Selenium（Se）is a necessary trace element in animals,and the loss of selenium can cause a variety of diseases.Dietary Se supplementation is a safe and effective way to supplement Se.Use the plant’s own biotransformation function to convert exogenous inorganic selenium into organic selenium,so as to cultivate excellent quality selenium-rich crops.In this study,Se-enriched shiitake mushrooms were cultivated by means of basswood cultivation,the absorption and conversion effect and biological activity of shiitake mushrooms on Se were studied.Se in Se-enriched shiitake mushrooms were analyzed by microwave closed digestion-hydride atomic fluorescence spectroscopy（HG-AFS）method,and the antioxidant and scavenging free radicals of Se-containing compounds such as selenoprotein and selenopolysaccharides in Se-enriched shiitake mushrooms were detected by ultraviolet-visible spectroscopy（UV）.Se-containing organic biomacromolecule compounds in Se-enriched shiitake mushrooms were quantitatively and qualitatively analyzed by high performance liquid chromatography-mass spectrometry（HPLC-ESI-MS）.The results are as follows:In the experimental concentration range,with the increase of Se application gradient,Se-enriched shiitake mushrooms cultivated under different Se concentration conditions can grown.The yield of Se-enriched shiitake mushrooms is related to the amount of Se applied,and the appropriate amount of Se can increase the yield of shiitake mushrooms,and the highest yield group is 1.9 times that of the blank group.And the Se content in Se-enriched shiitake mushrooms is analyzed by microwave digestion-HG-AFS quorum,and the Se content of Se-enriched shiitake mushrooms in the lyophilized group is up to 121.18 mg/kg,which is 216 times compared with the control group.The sun-dried Se-enriched shiitake mushroom could reach up to 36.21mg/kg,which was nearly 90 times compared with the control group.It is found that the Se content of lyophilized shiitake mushrooms is greater than that obtained by sun-dried treatment,and the amount of exogenous Se application is positively correlated with the Se-enriched capacity of shiitake mushrooms.Shiitake mushrooms do have an enriching effect on Se.Four different types of extractants of Na OH,H₂O,Tris-HCL and HCL were selected and different extraction conditions were set for the analysis of soluble Se in Se-enriched shiitake mushrooms,and it was found that the Se content of the four dissolved states changed positively correlated with the increase of Se concentration,of which the alkali-soluble extraction effect was significantly better than the other three extractants,and under the conditions of temperature 50℃,the extraction time was 2 h,and the material-liquid ratio was 1:30,the alkali-soluble extraction effect was the best.Based on the nature that organic biological macromolecules such as proteins are easily soluble in alkaline solution,it can be inferred that the organic selenium in selenium-rich shiitake mushrooms accounts for a large amount,and the organic selenium in shiitake mushrooms is more easily migrated and transformed in the weak alkaline medium,which is effectively absorbed by human body and has high bioavailability.The extraction conditions of soluble selenoproteins in selenium-rich shiitake mushrooms were optimized by two methods:univariate and response surface analysis.It was found that the leaching effect was the best when the temperature of Na OH leaching solution was 40℃,the leaching time was 2 h,and the ratio of material to liquid was 1:25.The content of soluble selenoproteins and protein Se in Se-enriched Lentinus edodes was positively correlated,and both increased with the increase of Se concentration.Under the optimal conditions,the highest content of soluble selenoproteins in Se-enriched shiitake mushroom samples was 51312 mg/kg,and the content of selenium in soluble selenoproteins could reach 22.54 mg/kg.It can be seen that the increase of Se concentration promotes the synthesis of soluble selenoproteins to a certain extent,and increases the combination of protein and Se,indicating that Substantial increase in protein Se concentration.H₂O was used as a extraction agent to extract soluble selenopolysaccharides,and the extraction effect of selenopolysaccharides was best when the extraction time was2.5 h,the temperature was 50℃,and the material-to-liquid ratio was 1:25 according to the results of univariate and response surface.With the increase of Se concentration,the content of soluble selenopolysaccharides and polysaccharide Se in Se-enriched shiitake mushrooms was positively correlated,and the content of soluble selenopolysaccharides was 56561 mg/kg under optimal conditions,and the Se content of selenopolysaccharides was up to 33.91 mg/kg.It can be known that the increase of Se concentration in Se-enriched shiitake mushrooms can promote the synthesis of soluble selenopolysaccharides in their bodies,promote the combination of Se and polysaccharides,and greatly increase the concentration of polysaccharide Se.The increase of Se concentration could promote the synthesis of soluble selenopolysaccharides in its shiitake mushroom,promote the combination of Se and polysaccharide,and the concentration of Se in selenpolysaccharides was improved.The contents of soluble selenopolysaccharides were all slightly greater than those of soluble selenoproteins,and the contents of selenium in selenopolysaccharides were all slightly greater than those of selenoproteins,indicating that the increase of selenium content in selenium-enriched shiitake mushroom was more favorable to the production of selenopolysaccharides,and the organic selenium in the form of selenopolysaccharides in selenium-enriched shiitake mushroom was higher than that in selenoprotein form.Firstly,selenopolyaccharides and selenoproteins from shiitake mushrooms were extracted in the experimental concentration range and under the optimal extraction conditions.Then the reducing power and free radical scavenging ability of selenopolysaccharide and selenoprotein were tested.Thus,the antioxidant activities of sodium selenite with selenopolyaccharides and selenoproteins were studied comparatively.The results show that the reducing power and ability to scavenging free radicals of organic Se such as selenopolysaccharides and selenoproteins in Se-rich shiitake mushrooms are greater than those of inorganic Se,and the reducing power and ability to scavenor free radicals under the same concentration of Se are greater than those of selenoproteins.When the Se content is the largest,selenopolysaccharides and selenoproteins activities are the strongest,which have obvious guiding effects on improving the nutritional quality of shiitake mushrooms and producing Se-rich shiitake mushrooms with strong antioxidant activity.Provide technical support for Se-rich shiitake mushrooms as functional health foods.HPLC-ESI-MS combined technology was used to qualitatively and quantitatively analyze the Se amino acids in Se-enriched shiitake mushrooms.The MRM mode was selected for quantitative analysis,and the results showed that Se-enriched technology could effectively increase the content of Se Cys₂,Se-Met and Se-Me Se Cys in shiitake mushrooms,and the maximum amounts were up to 3937.45μg/kg、8732.95μg/kg and 58.13μg/kg respectively.At the same time,by analyzing a large number of liquid mass maps,three unknown Se-containing compounds and their fragmented ions in Se-rich shiitake mushrooms were qualitatively analyzed and identified.And the results showed that their possible structures were:selenomethylmethionine(C₆H₁₄NO₂Se),N-acetyl-selemethionine(C₇H₁₃NO₃Se)and Se-5methyl-selenosine selenium oxide(C₁₂H₁₈N₅O₃Se).This conclusion fills that the molecular structure of organic selenium in selenium-rich shiitake mushrooms and the study of the mechanism,lays the theoretical foundation,and provides important scientific data as a superior selenium supplement.