κ-卡拉胶寡糖酶解制备工艺优化

Technology optimization for enzymatic preparation of κ-carrageenan oligosaccharides

为改进卡拉胶寡糖的制备工艺,本研究以κ-卡拉胶为底物,采用酶法制备κ-卡拉胶寡糖,以还原糖生成量为评价指标,对酶解条件进行优化,并采用薄层色谱法及质谱对酶解产物进行分析。结果显示酶解反应最优工艺条件为:底物浓度12 g/L、p H7.0、反应温度40℃、振荡速率100 r/min,经优化后的酶解反应更完全,还原糖生成量由0.91 mg/m L提高至1.61 mg/m L,提高了77%,酶解卡拉胶反应的Km值为171.96 mg/m L,最大反应速度Vmax为0.925 mg·m L^(-1)·min^(-1)。最优工艺验证实验和20 L放大实验结果一致,经薄层色谱及质谱分析酶解24 h后的反应主产物为二糖和四糖。

In this study, K- carrageenan oligosaccharides were prepared by enzymatic hyrolysis method to improve the technology.Hydrolysis conditions were optimized with the characteristic signs of reducing sugars content, and the optimal conditions for K-carrageenan oligosaeeharides preparation were obtained as substrate concentration 12 g/L, pHT.0, reaction temperature 40 ℃ and shaking speed 100 r/min,respectively.Compared with the reducing sugars yield of the initial conditions, the yield of the optimal conditions increased from 0.91 mg/mL to 1.61 mg/mL,whieh was increased by 77%.The Km and V values of the enzyme hydrolysis reaction was 171.96 mg/mL and 0.925 mg· mL^-1 · min-1 , respectively. Then, the result of optimal process validation test could consistent with that of 20 L enlarge experiment.At last,the enzymatic hydrolysis products were analyzed by thinlayer chromatography and mass spectrometry, the result showed that the main products included tetrasaccharide and hexasaceharides in 24 h reaction.