Zhang Meng, Zhang Yiran, Ma Xuexiao, Liu Xiangping, Niu Min, Yao Ruyong, Zhang Lijuan
Systems Biology & Medicine Center for Complex Diseases, Affiliated Hospital of Qingdao University, Qingdao, 266003, China.
Shandong Institute of Orthopaedics and Traumatology, Affiliated Hospital of Qingdao University, Qingdao, 266003, China.
Carbohydr Polym. 2020 Jul 15;240:116338. doi: 10.1016/j.carbpol.2020.116338. Epub 2020 Apr 22.
Monosaccharide composition analysis after acid hydrolysis is the first step towards structural characterization of the polysaccharides. To modernize the hydrolytic procedure, we used a polymerase chain reaction (PCR) instrument to accomplish the task, which allows to generate monosaccharide products from up to 96 samples simultaneously within 30 min. Fucoidan, chitosan and propylene glycol alginate sodium sulfate (PSS) were chosen as representatives of complex, basic and acidic polysaccharides to optimize the hydrolytic conditions, respectively, through the orthogonal L9 (3) experiments. The hydrolysis loss ratio for monosaccharide standards were also measured. Using this assay, the hydrolysis plus 1-phenyl-3-methyl-5-pyrazolone (PMP) labeling of the monosaccharide products could be accomplished in 90 min with the RSD values less than 5 % based on HPLC analysis. We further confirmed the reliability of the assay by HPLC coupled MS analysis. In conclusion, PCR instrument-based hydrolysis assay is suitable for monosaccharide composition analysis of complex, acidic and basic polysaccharides.
酸水解后的单糖组成分析是多糖结构表征的第一步。为了使水解过程现代化，我们使用聚合酶链反应（PCR）仪器来完成这项任务，该仪器能够在30分钟内同时从多达96个样品中生成单糖产物。分别选择岩藻聚糖、壳聚糖和藻酸丙二醇酯硫酸钠（PSS）作为复杂多糖、碱性多糖和酸性多糖的代表，通过正交L9（3）实验来优化水解条件。还测定了单糖标准品的水解损失率。使用该方法，基于高效液相色谱（HPLC）分析，单糖产物的水解加1-苯基-3-甲基-5-吡唑啉酮（PMP）标记可在90分钟内完成，相对标准偏差（RSD）值小于5%。我们通过HPLC联用质谱分析进一步证实了该方法的可靠性。总之，基于PCR仪器的水解分析适用于复杂多糖、酸性多糖和碱性多糖的单糖组成分析。
