Researchers propose new enzyme-based criteria to verify polysaccharide purity

Natural polysaccharide — complex carbohydrates found in plants, fungi, and marine organisms — are increasingly valued for their therapeutic potential, from immune regulation to gut health. However, there is a growing concern in the field is that many polysaccharides that are reported to have elaborate, novel structures may in fact be simple physical mixtures of different polymers. Current gold-standard purity tests often fail to catch these mixtures, leading to incorrect structural conclusions and unreliable biological data. Ultimately, this raises a fundamental question: are researchers actually studying what they think they are?

In a study published in the KeAi journal Glycoscience & Therapy, a joint research team from Hong Kong Baptist University and Northeast Normal University pit this concern to the test and proposed a solution.

The researchers created deliberate "mixture models" using polymer reference standards. The first mimicked a mixture of two different homopolysaccharides of the same molecular weight, while the second represented a heteropolysaccharide scenario. When assessed by conventional techniques — high-performance gel permeation chromatography (HPGPC), monosaccharide composition analysis, methylation analysis, and 1D/2D NMR — both mixtures deceptively appeared as single, pure substances. Worse still, standard structure elucidation based on the mixed data produced complicated and incorrect structures.

To that end, the team applied an "enzyme diagnostic criteria": they selected specific enzymes predicted to cleave the individual polymers in each mixture and monitored the change in HPGPC peaks after enzymatic digestion. The hidden mixtures were unmasked in both models, demonstrating the superior reliability of the enzyme-based approach over existing methods.

"Current purity criteria are not rigorous enough to identify whether a polysaccharide sample is truly a single entity or a mixture," explains Professor Quanbin Han, corresponding author of the study. "By incorporating diagnostic enzymes, we can verify purity upfront, before the sample goes deeper into structure analysis. That way, any further biological evaluation or functional study is built on a reliable, pure sample-we’re not chasing artifacts from hidden mixtures."

As interest in polysaccharide-based therapeutics and nutraceuticals continues to surge, the stakes for getting purity right have never been higher: impurities can confound bioactivity assays, compromise clinical reproducibility, and slow the translation of promising candidates into real-world treatments.

Against this backdrop, the study provides a validation framework that can be readily adopted by polysaccharide research laboratories worldwide.  The authors hope their findings will encourage the field to adopt enzyme-based verification as a standard step in the polysaccharide characterization workflow, ensuring that the polysaccharides entering preclinical pipelines are exactly what they are.

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Contact the author:

Quanbin Han, School of Chinese Medicine, Hong Kong Baptist University, Hong Kong, China.   simonhan@hkbu.edu.hk;

Yifa Zhou, School of Life Sciences, Northeast Normal University, Changchun, China. zhouyf383@nenu.edu.cn.

The publisher KeAi was established by Elsevier and China Science Publishing & Media Ltd to unfold quality research globally. In 2013, our focus shifted to open access publishing. We now proudly publish more than 200 world-class, open access, English language journals, spanning all scientific disciplines. Many of these are titles we publish in partnership with prestigious societies and academic institutions, such as the National Natural Science Foundation of China (NSFC).