School of Molecular and Microbial Biosciences, University of Sydney, Sydney, NSW, Australia.
Biomacromolecules. 2009 Mar 9;10(3):638-44. doi: 10.1021/bm8014413.
A (1)H NMR method is presented that monitors the initial and later stages of in vitro enzymatic digestion of starch suspensions. It allows, for the first time to our knowledge, the accurate analysis of the initial 5% of the extent of hydrolysis. This is significant because rapidly digested starch produces glucose that determines the blood glucose concentration immediately after ingestion of food. The two key hydrolytic enzymes, alpha-amylase and amyloglucosidase, showed clear systematic deviation from Michaelis-Menten kinetics as the starch or wheat flour substrate that was used changed its character during the reaction. Estimates of Michaelis-Menten parameters for amyloglucosidase and alpha-amylase were successfully found by analyzing two stages of digestion separately. The Michaelis-Menten constants for purified starch were (6.4 +/- 0.8) and (1.1 +/- 0.3) g dL(-1) (% w/v), respectively; and the maximum velocities of glucose release by amyloglucosidase, and short oligoglucosides and glucose by alpha-amylase were (1.9 +/- 0.4) x 10(-2) and (1.6 +/- 0.2) x 10(-2) mmol L(-1) s(-1) for the first stage of digestion, and (9.0 +/- 1.0) x 10(-3) and (4.7 +/- 1.4) x 10(-3) mmol L(-1) s(-1) for the second stage, giving a ratio of the two V(max) values of 2.1 and 3.4, respectively.
一种 (1)H NMR 方法被提出，用于监测淀粉悬浮液体外酶解的初始和后期阶段。据我们所知，它首次允许准确分析水解程度的最初 5%。这是重要的，因为快速消化的淀粉会产生葡萄糖，这会立即影响食物摄入后血糖浓度。两种关键的水解酶，α-淀粉酶和淀粉酶，在淀粉或小麦粉底物在反应过程中改变其性质时，明显表现出与米氏-门坦动力学的系统偏差。通过分别分析两个消化阶段，成功地找到了淀粉酶和α-淀粉酶的米氏-门坦参数估计值。纯化淀粉的米氏-门坦常数分别为（6.4 +/- 0.8）和（1.1 +/- 0.3）g dL(-1)（% w/v）；淀粉酶释放葡萄糖、短寡糖和α-淀粉酶释放葡萄糖的最大速度在第一阶段分别为（1.9 +/- 0.4）x 10(-2)和（1.6 +/- 0.2）x 10(-2)mmol L(-1) s(-1)，在第二阶段分别为（9.0 +/- 1.0）x 10(-3)和（4.7 +/- 1.4）x 10(-3)mmol L(-1) s(-1)，两个 V(max)值的比值分别为 2.1 和 3.4。
