| As the research object,based on the wildness around the wildness saponin extraction,purification,antioxidant activity,inhibit enzyme activity,cell biological activity in vitro and liquid chromatography-mass spectrometry(UPLC/Q-TOF-MS)saponins of wildness for chemical composition analysis on the specific research work and so on,for the further development and utilization of wildness and related byproducts.In the process of extracting saponin from Tricholoma hazelloides,smilax saponin was used as the standard to analyze the content of saponin from Tricholoma hazelloides.The technological conditions of extracting saponin from Tricholoma hazelloides by reflux method,ultrasonic method and microwave method were optimized by single factor test and response surface test.The optimal conditions of reflux extraction were as follows: ethanol concentration of 75.05 %,solid-liquid ratio of 1:28.25 g/m L,extraction time of 78.04 min,the theoretical extraction rate was34.48 %,the actual extraction rate was 34.52 %.The optimal extraction conditions were as follows: ethanol concentration of 84.62%,solid-liquid ratio of 1:27.22 g/m L,extraction time of 13.06 min.The theoretical extraction rate was 22.90%,and the actual extraction rate was 22.91%.The optimal extraction conditions of saponin from mushroom were as follows: ethanol concentration 57.52 %,solid-liquid ratio 25 g/m L,extraction time 20.00 min,extraction power 369.75 W.The theoretical extraction rate was 34.29 %,and the actual extraction rate was 34.61 %.The results of the three extraction methods showed that the extraction rate of reflux method was higher than that of microwave method and ultrasonic method.At the same time,reflux extraction method in industrial production,simple operation,simple equipment,more suitable for mass production.In the study on the purification of saponin from hazelnut mushroom,Macroporous resin was used to purify the hazelnut mushroom.The recovery and purity of saponin from hazelnut mushroom were taken as the indexes.The purification process was optimized by single factor test and response surface test,and the optimal conditions of purification process were: The analytical liquid volume was55.85 m L,the adsorption liquid concentration was 5.46 mg/m L,the adsorption liquid volume was 30 m L,and the recovery rate was 56.37 %.The purity of saponins increased from 34.51 % to 63.50 %.By measuring the antioxidant activity of saponin from hazelleaf,it was found that when the concentration of vitamin C reached 1.25 mg/m L,the scavenging rate of ·OH free radical reached 40.13 %,33.51 % and 29.09 %,respectively.When the concentration reached 5 mg/m L,the total antioxidant capacity reached 0.577,0.494 and 0.555,respectively.When the concentration reached 10 mg/m L,the reducing power reached 0.460,0.381 and 0.304,respectively.When the concentration reached25 mg/m L,the DPPH clearance rate reached 93.51 %,95.35 % and 93.27 %,respectively.The inhibitory enzyme activity of the purified saponins of Corylus hazelloides was determined,and acarbose and vitamin C were used as positive controls.The test results were as follows: when the concentration of saponin from hazel mushroom reached 25 mg/m L,the inhibitory rates of saponin from hazel mushroom onα-glucosidase were 38.73 %,36.38 % and 35.45 %,respectively.The inhibition rates of amylase were 52.58 %,63.10 % and 65.08 %,respectively.The inhibition rates of tyrosinase were 55.14 %,57.48 % and 66.82 %,respectively.The inhibition rates of lipase were 96.33 %,84.49 % and 88.96 %,respectively.Cytotoxicity(MTT)test and apoptosis test were performed on the purified saponins from hazelnut mushroom,and lung cancer cell A549 and osteoma cell MG63 were selected as recipient cells.The results showed that when the concentration of hazelnut mushroom reached 400 μg/m L,the inhibition rate of the saponins extracted from hazelnut mushroom by reflux method reached 93.26 % and88.88 %,respectively.The inhibition rate of saponins extracted by ultrasonic method reached 91.53 % and 84.31 %,respectively.The inhibitory rate of saponins extracted by microwave was 92.01 % and 83.29 %,respectively.Apoptosis rate of human lung cancer cell A549 and myeloma MG63 was 70.00 % and 61.10 %,respectively,induced by the saponins from hazelnut mushroom extracted by reflow method.The apoptotic rate of human lung cancer cell A549 and myeloma MG63 was 68.80 % and58.00 %,respectively.The apoptotic rate of human lung cancer cell A549 and myeloma MG63 was 61.60 % and 61.80 %,respectively.Using liquid chromatography-mass spectrometry(UPLC/Q-TOF-MS)to analyze the chemical constituents of the purified saponins from Tricholoma hazelloides,18 chemical constituents were identified from the saponins extracted by reflux method,including 12 sesquiterpenoids,1 adenosine,4 sterols and 1 purine.Twenty-two chemical constituents were identified from the saponins extracted by ultrasonic method,including 14 sesquiterpenes,3 adenosine compounds,3 sterols,1 purine compound and 1 diterpene triterpene compound.Eight chemical constituents were identified from the saponins extracted by microwave method,including 12 sesquiterpenoids,1 adenosine,4 sterols and 1 purine. |
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