Study On Biological Characteristics And Digestive Enzyme Activities Of ’JinHua’ Fungi From Fuzhuan Brick Tea

The earliest Fuzhuan Brick Tea which was a compressed tea manufacturedby in southern Shaanxi in early Ming dynasty.There was a kind of golden funguswhich was called ‘Jin Hua’ Fungi growing in the brick body.The quality andquantity of ‘Jin Hua’ Fungi is taken as the factor of judging the quality ofFuzhuan Brick Tea.‘Jin Hua’ Fungi was attentted in recent years because of it′spowerful function. This study takes ‘Jin Hua’ Fungi as the spawn which wasextracted from Fuzhuan Brick Tea in Shaanxi, analysis it′s biologicalcharacteristics.Through the optimization test was determined the most suitablesolid and liquid cultivate conditions. Under the optimum cultivate conditionsdetermined the production of acid protease activity, alkaline protease activity,neutral protease activity, amylase activity and lipase activity. The experimentanalysis results show that ‘Jin Hua’ Fungi has dramatically activity of producingneutral and alkaline protease. And then study the culture medium optimization ofneutral and alkaline protease. The results of this study are as follows:（1）‘Jin Hua’ Fungi was separated from Fuzhuan Brick Tea in Shaanxi,observed the colonies, mycelia and reproductive character of ‘Jin Hua’ Fungiwhich living on PDA medium, CZG（5%NaCl） medium, CZG（14%NaCl）medium, CZ medium, CZ（20%of sucrose） medium, CZG（5%NaCl） medium,andidentified fungi in the molecular level by DNA sequencing.The conclusion is：the‘Jin Hua’ Fungi separated from Fuzhuan Brick Tea in Shaanxi was identified asEurotium cristatum.（2） Take dry weight of mycelia pellets as an indicator, investigated theoptimize cultivate conditions of liquid culture of Eurotium cristatum. firstly,inspect different carbon sources: sucrose, ammonium nitrate, urea, peptone, yeastextract, soybean powder, and different contents of the best carbon source:0.5%,1%,1.5%,2%,2.5%,meanwhile,inspect different nitrogen sources: sucrose, cornflour, glucose, lactose, soluble starch, and different contents of the best nitrogen source:2%,4%,6%,8%,10%were also. Secondly, inspect inoculation amount:1%,5%,10%, liquid medium volume:12%,16%,20%, rotating speed:110r/min,140r/min,150r/min, pH:5.0,5.5,6.0, influence on the liquid fermentation.Theoptimal liquid culture conditions were determined as:when the nitrogen sourcewas2.5%soybean powder, carbon source was6%sucrose, inoculums size was10%, liquid volume was16%, rotating speed was150r/min, initial pH was5.0,and the dry weight of mycelia pellets of Eurotium cristatum could reach0.32%.Select three better liquid culture conditions and determinat the activity ofproducing digestive enzymes. A: inoculums size was10%, liquid volume was16%, rotating speed was150r/min, and initial pH was5.0. B: inoculums size was10%, liquid volume was16%, rotating speed was110r/min, and initial pH was5.5. C: inoculums size was10%, liquid volume was16%, rotating speed was150r/min, and initial pH was5.0. The determination results: three conditionshave not activity of producing acid protease and lipase. In the three conditions,the activity of neutral protease respectively is5U/mL,4U/mL,8U/mL, theactivity of alkali protease respectively is4U/mL,2U/mL,6U/mL and the activityof amylase respectively is U/mL,5U/mL,7U/mL.（3）Take spore number of Eurotium cristatum as an indicator, investigatedthe optimize cultivate conditions of Solid State Fermentation of Eurotiumcristatum:study4mL、7mL、10mL、13mL、16mL whater were added in10g wheatbran，and Study different culture temperature of24℃,28℃,32℃,37℃Theeffecton Sporulation on sporulation. Determine the optimum cultivationconditions as follows: added water to10mL and culture temperature of28℃. Inthis condition, the sporulation could reach8.0×108/g.Eurotium cristatum’s ability of producing protease, amylase and lipase wasstudied. The results showed that, when Eurotium cristatum was fostered for5days, the ability of producing amylase could achieve254.3U/g, and the ability ofproducing lipase could achieve11.4U/g for6days. When Eurotium cristatumwas fostered for7days, the activity of producing neutral protease was160.2U/g,and the activity of producing alkaline protease was117.8U/g.When it wasfostered for4.5days, the activity of producing acid protease was22.6U/g. Allthe results showed that Eurotium cristatum mainly produced neutral and alkalineprotease, and the activity of them all reached the biggest at7days.Optimized the culture medium for Eurotium cristatum producing neutral and alkaline proteasein different carbon source: sucrose, lactose, maltose, soluble starch, corn flour,and different nitrogen source: nitric acid amine, urea, specially designed, yeastpowder, soybean powder, different inorganic salt: MgSO₄·7H₂O, MnSO₄·H₂O,ZnSO₄·7H₂O, CaCl₂, FeSO₄·7H₂O. Results indicated that the optimal carbonsource was lactose, the optimal nitrogen source was bran, and the optimalinorganic salt was sodium chloride. With the optimal culture medium, theactivity of neutral protease could be up to237.0U/g and alkali protease is up to235.8U/g.The Eurotium cristatum’s liquid fermentation conducted bacteriostaticexperiment to Escherichia coli and StapHylococcus aureus. The test resultsproved that the ‘Jin Hua’ Fungi fermentation liquid have bacteriostatic functionto Escherichia coli and StapHylococcus aureus. The diameter of antibacterialcircle is12mm and10mm.