Exploring extraction/dissolution procedures for analysis of starch chain-length distributions
Exploring extraction/dissolution procedures for analysis of starch chain-length distributions
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DOI：
10.1016/j.carbpol.2014.08.001
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发表时间：
2014-12-19
影响因子：
11.2
通讯作者：
Gilbert, Robert G.
中科院分区：
文献类型：
--
作者：
Wu, Alex Chi;Li, EnPeng;Gilbert, Robert G.
分析淀粉链长分布(CLDs)对于了解淀粉生物合成-结构-性质关系具有重要意义。它是通过分析淀粉α-(1->；6)糖苷键在酶促脱支过程中释放的线性葡聚糖链的数量分布来获得的，用于后续的荧光团辅助碳水化合物电泳法(Face)或尺寸排除层析(SEC)等技术的表征。目前文献中对CLD确定前的分支化进行的前处理涉及不同的方案，这可能会产生人工结果。本文研究了两种广泛使用的淀粉溶解方法，一种是含有0.5%(w/w)溴化锂的二甲基亚砜(DMSO)，另一种是在100℃下用碱性水溶液(即NaOH)溶解淀粉。对不同结构的大麦淀粉的CLD进行了FACE和SEC的分析。(1)NaOH处理，即使在稀释浓度下，在聚合度较高时也会导致显著的降解，导致定量上不正确的CLD结果，导致支链淀粉和直链淀粉链变长。(2)经NaOH处理后，CLD的支链淀粉和直链淀粉组分中的某些特征减少为凸起或缺失。(3)稀碱条件下支链淀粉溶解不完全，高估了CLD中直链淀粉的含量。这些结果表明，用二甲基亚砜-溴化锂溶解淀粉是减少伪影的首选方法。提出了一种改进的用于Face和SEC分析淀粉CLD的前处理方法。(C)2014爱思唯尔有限公司。保留所有权利。
The analysis of starch chain-length distributions (CLDs) is important for understanding starch biosythesis-structure-property relations. It is obtained by analyzing the number distribution of the linear glucan chains released by enzymatic debranching of starch alpha-(1 -> 6) glycosidic bonds for subsequent characterization by techniques such as fluorophore-assisted carbohydrate electrophoresis (FACE) or size-exclusion chromatography (SEC). Current literature pretreatments for debranching prior to CLD determination involve varying protocols, which might yield artifactual results. This paper examines the two widely used starch dissolution treatments with dimethyl sulfoxide (DMSO) containing 0.5% (w/w) lithium bromide (DMSO-LiBr) at 80 degrees C and with aqueous alkaline (i.e. NaOH) solvents at 100 degrees C. Analyses by FACE with a very high range of degree of polymerization, and by SEC, of the CLD of barley starches with different structures show the following. (1) The NaOH treatment, even at a dilute concentration, causes significant degradation at higher degrees of polymerization, leading to quantitatively incorrect CLD results in longer amylopectin and in amylose chains. (2) Certain features in both amylopectin and amylose fractions of the CLD reduced to bumps or are missing with NaOH treatment. (3) Overestimation of amylose chains in starch CLD due to incomplete amylopectin dissolution with dilute NaOH concentration. These results indicate starch dissolution with DMSO-LiBr is the method of choice for minimizing artifacts. An improved pretreatment protocol is presented for starch CLD analysis by FACE and SEC. (C) 2014 Elsevier Ltd. All rights reserved.