Screening And Mutation Breeding Of The Naringinase Producing Strain And The Study Of The Debittering Technology By Naringinase

Naringinase was the extracellular enzyme from Aspergillus niger, Aspergillus oryzae, Penicillium notatum and some other fungi which could hydrolyze naringin to debitterize. It had great potential application in food processing. In our work:1. A new model for screening of naringinase producing strain, plates containing orange meal as the solo carbon and nitrogen resource were used, and a naringinase producing strain A13 was obtained. A13 could secreted 365.31U/mL naringinase in shake flask, and it was identified to be Aspergillus niger;2. A13 was mutated with NTG and the spores were spread on the plates which containing 2.0μg/mL tennecetin, the minimum lethal concentration, to obtain the antidrug mutants. After holo and shake flask screening, one naringinase over-producing strain A-13-62 whose yield was 876.43U/mL in shake flask, 237% higher that of original strain, among 180 anti-tennecetin mutants was got;3. The optimal medium and liquid fermentation condition were determined by One-factor Experimental Design and Orthogonal Experimental Design: bean dregs(containing 30%~40%water)100g, degreasing soybean powder 40g, rhamnose0.2g, CaCl21g, ZnSO41g, MgSO4·7H2O5g, K2HPO41g, distilled water 1000mL, cultured at 30℃, initial pH 5.0, inoculume 10%, 50mL medium in 500mL shake flask, and the yield was 1017U/mL after 96 hr's fermentation in this condition;4. The characterization of the naringinase secreted by A-13-62 was performed: The optimal pH and temperature of the enzymatic reaction was 4.0 and 60℃separately, Mg2+,Ca2+ had great inhibition effect while Zn2+ could promote the enzyme; 5. The process of enzymatic debittering was optimized: at the pH of nature grapefruit juice and 50℃, 8000IU/kg enzyme, after 1 hr's reaction the debittering rate reached to 46.25%, got excellent flavour characteristics.