Studies On The Extraction Of ALDH From Xuefeng Dry Yeast And Enzymatic Properties Of The ALDH

Aldehyde dehydrogenase （ALDH） is one of the key enzymes involved in acetaldehydemetabolism in the human body. ALDH can catalyze the oxidation of a variety of aliphatic andaromatic aldehydes to generate the corresponding acid oxidation. As a result, it reduces thetoxicity of aldehydes in the body. Currently, ALDH was mainly extracted from the plant andanimal cells by using chromatograph which is high cost and low yield. Animal and plant cellscontain small amounts of ALDH and it is difficult for large-scale production. With theincreased demand of ALDH, we will be focused on how to find an alternative which containslarge amounts of ALDH, simple and efficient ways to obtain large amounts of the purifiedaldehyde dehydrogenase. As microbial culture is easy and quick, the extraction of ALDHfrom microorganism will have great prospects.In this study, Xuefeng instant yeast （high and low sugar）, brewer’s yeast waste, AngelYeast tablets, Caihong instant yeast, Saf-instant yeast were chosed to extract ALDH. Theresults showed that the content of ALDH in the Xuefeng （low sugar） instant yeast is thehighest. Ultrasonication, repeated freeze-thaw method, chemical osmosis and grindingmethod were selected to extract ALDH from Xuefeng （low sugar） instant yeast.The resultsshowed that the ALDH activity of crude enzyme solution extracted by ultrasonication was thehighest. Various influencing factors of ultrasonication method were discussed and theoptimum conditions for extraction were as follows: ratio of solid to liquid was1:4, handlingcapacity was20ml, pH of extraction buffer was8.0, ultrasonic power was300W,ultrasonication time was16minuts. In the optimum conditions, the ALDH activity of crudeenzyme solution was up to3.56U/mL.In this study, salt precipitation, column chromatography and aqueous two-phaseextraction method were used to purify crude enzyme solution of ALDH. The results showedthat the recovery of ALDH was up to71.77%, purification fold was up to2.200by using saltprecipitation to purify crude enzyme solution of ALDH. the recovery of ALDH was up to31.31%, purification fold was up to2.705by using column chromatography. The recovery ofALDH was up to70.00%and purification fold was up to2.069by using aqueous two-phaseextraction. ALDH solutions extracted from Xuefeng （low sugar） instant yeast had a commonprotein band with the standard by polyacrylamide gel electrophoresis, so the weight of theALDH monomer molecular was about57kDa.Parts of the enzymatic properties of ALDH extracted from Xuefeng （low sugar） instantyeast were also investigated in this study. The results showed that the optimum reaction pH and temperature were8.0and25℃. It was suitable for ALDH to preserve in phosphatebuffer of pH6.5and the optimum preservation temperature was25℃, which also keptALDH with high activity. Various metal ions affected the activity of ALDH extracted fromXuefeng （low sugar） instant yeast varies. Using the activity of ALDH plus k+as a contrastexperiment, Al³⁺and Fe³⁺had a strong inhibition of ALDH activity, Zn²⁺and Cu²⁺had thestrongest inhibition, Ca²⁺had a certain inhibition, Mg²⁺had little inhibition. Na⁺strengthenssalting-out, so it had an inhibition of ALDH activity. The ALDH had an extensive substratespecificity, and propionaldehyde was the best substrate for the ALDH reaction, followed byacetaldehyde, formaldehyde reactivity was very small.