柠檬苦素诱导塔宾曲霉UA13高产柠檬苦素转化酶的研究

Limonoid-induced high production of limonin invertase in Aspergillus tabin UA13

以柠檬苦素为代表的三萜化合物是柑橘类水果在加工过程中产生“后苦味”的主要原因,采用酶法转化使柠檬苦素的含量降低是目前较适合果汁加工业的方法。该研究以实验室前期分离并经等离子体(atmospheric room temperature plasma,ARTP)诱变获得的塔宾曲霉UA13为出发菌株,采用柠檬苦素作为诱导物对其进行产酶诱导,并对菌株的发酵产酶工艺和酶的底物亲和性进行研究。结果表明,在1 L发酵罐培养基中,接种量10%(体积分数)、初始pH值5.0、搅拌转速250 r/min、30℃的最适条件下发酵,柠檬苦素转化酶活力最高可达48.52 U/mL。酶对柠檬苦素的底物亲和性研究结果表明,柠檬苦素转化酶的K m值为0.622 mmol/L,说明诱导后的菌株UA13所产酶对柠檬苦素的亲和力增强。同时研究结果显示,柠檬苦素转化酶具有较好的耐热性和耐酸性,在柑橘果汁加工业中具有非常好的应用前景。

Limonoid compounds existed in citrus are the major contributors to the bitter taste of citrus fruits.Limonin,the most representative limonoid compounds,produce an extraordinarily bitter taste after processing of fresh fruits without obvious bitter taste.The enzymatic method is suitable for removal of bitter components in citrus processed products nowadays.In this study,limonin is applied to induce the limonin invertase production in an Aspergillus tabin strain UA13,which was previously isolated in our laboratory and bred by atmospheric room temperature plasma(ARTP)mutagenesis.Furthermore,the fermentation and enzyme production were optimized and the substrate's affinity for the enzyme was investigated.The results demonstrated that the activity of limonin invertase by A.tabin UA13 reached up to 48.52 U/mL in 1 L fermentor at optimized conditions with 9.0 mg/L limonin induction,10%(v/v)inoculum size,initial pH 5.0,250 r/min stirring speed and 30℃fermentation temperature.The results of limonin’s affinity for the enzyme displayed that the K m value of limonin converting enzyme was 0.622 mmol/L.This indicated that the enzyme-substrate affinity of limonin invertase produced by the A.tabin was significantly increased after limonin induction.Moreover,with high heat resistance and acid resistance,limonin invertase in this study is commercially potential and has prospective applications in the citrus processing industry for juice and concentrate.