Production Of Ellagic Acid From Pomegranate Husk By Enzyme

Ellagic acid is a kind of natural polyphenolic compounds, widely distributing in various nuts and baccas, which is a dimeric ramification of gallic acid, polyphenolic lactone. Ellagic acid have been associated with anticancer, antivirus, anti-oxidant and advanced whitening activities. In recent years, the attention to ellagic acid and application of ellagic acid are more and more extensively. The dissertation focused on the technique about the production of ellagic acid by enzyme (tannase) using pomegranate husk tannin (an ellagitannin) as the substrate.The main research items and results are as follows:(1) By investigating the effects of carbon sources, pomegranate husk tannin concentration, incubation time,tween-80concentration, pH, temperature on tannase acivity. Based on the Aspergillus niger, the optimum culture medium and cultivation condition were obtained. The optimum medium was listed as follows:Surose10g/L, NaNO32g/L, KH2PO41g/L, K2HPO41g/L, KCl0.5g/L, MgSO40.5g/L,FeSO40.01g/L, tannins3.3%,tween-806ml/L, initial pH4.0,6days. (2) Considering the effect of reaction pH, time and concentration of tannase on the ellagic acid production, we determined the optimum enzymatic reaction condition. The result was shown as follow:15ml tannase fluid (65U/ml) was added into50ml solution with6.7%(V/V)tannins, the reaction was done with PH4.5and temperature at30℃for48h.(3) A new method for purification of ellagic acid has been developed. The ellagic acid produced by enzyme methods is purified. According to the character of ellagic acid, which can be dissolved in alkalescent liquid and separated out in acidic liquid, the preliminary process conditions is established:Crude ellagic acid (22.3%) is dissolved in solution of NaOH (32g/L), and then stirred. Remove the precipitation by centrifuge, adjust the supernatant solvent to pH4.8with1.5%HCl. The precipitation, namely the product, is collected by centrifuge and washed once with water then dried. The product is yellow and green with61.3%purity.(4) Preliminary study was done on the property of the tannase produced from the Asperyillus niger. Using PG as the substrate, the optimal pH value of tannase is5.0, the most suitable reaction temperature is60℃.In the pH range of5.0-6.0, the tannase activity is stable. Under the70℃for30min, the70%tannase activity is left.(5) The effect of additives on tannase activity was studied. Ba2+, OH-, CO32-activated tannase activity; Ag+, Cu2+, Mn2+, Zn2+, Ca2+, Fe2+little inhibited enzyme activity; Co2+, Al3+, EDTA, DMSO, β-Mercaptoethanol more inhibited tannase activity; Pb2+competitively inhibited tannase activity; Inorganic nitrogen sources and surfactants have no inhibition on tannase activity.