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Using a PCR instrument to hydrolyze polysaccharides for monosaccharide composition analyses
Meng Zhang1, Yiran Zhang2, Xuexiao Ma2
1Systems Biology & Medicine Center for Complex Diseases, Affiliated Hospital of Qingdao University, Qingdao, 266003, China.
Carbohydrate Polymers
|June 2, 2020
Summary
A novel polymerase chain reaction (PCR) instrument enables rapid monosaccharide analysis of complex polysaccharides. This method streamlines structural characterization, offering a faster alternative for polysaccharide research.
Area of Science:
- Carbohydrate Chemistry
- Analytical Chemistry
- Biochemistry
Background:
- Monosaccharide composition analysis is crucial for polysaccharide structural characterization.
- Traditional acid hydrolysis methods can be time-consuming and labor-intensive.
- Modernizing hydrolytic procedures is essential for efficient polysaccharide analysis.
Purpose of the Study:
- To develop a modernized, rapid, and efficient method for monosaccharide composition analysis of polysaccharides.
- To adapt a polymerase chain reaction (PCR) instrument for acid hydrolysis of polysaccharides.
- To optimize and validate the PCR instrument-based hydrolysis assay for various polysaccharide types.
Main Methods:
- Utilized a polymerase chain reaction (PCR) instrument for simultaneous acid hydrolysis of up to 96 samples in 30 minutes.
- Optimized hydrolytic conditions for complex (fucoidan), basic (chitosan), and acidic (PSS) polysaccharides using orthogonal L9 (3^4) experiments.
- Measured hydrolysis loss ratio for monosaccharide standards.
- Combined hydrolysis and 1-phenyl-3-methyl-5-pyrazolone (PMP) labeling for analysis within 90 minutes using High-Performance Liquid Chromatography (HPLC).
- Confirmed assay reliability using HPLC coupled with Mass Spectrometry (MS).
Main Results:
- Achieved rapid monosaccharide product generation from polysaccharides using a PCR instrument.
- Optimized hydrolysis conditions for diverse polysaccharide types.
- Demonstrated high efficiency with hydrolysis and PMP labeling completed in 90 minutes.
- Obtained Relative Standard Deviation (RSD) values less than 5% via HPLC analysis.
- Validated the assay's reliability through HPLC-MS analysis.
Conclusions:
- The PCR instrument-based hydrolysis assay provides a rapid and efficient method for monosaccharide composition analysis.
- This assay is suitable for a wide range of polysaccharides, including complex, acidic, and basic types.
- The modernized approach significantly reduces analysis time compared to traditional methods, facilitating structural characterization.
