Developing a high-throughput microassay for large samples of fungal polysaccharides

Abstract

The phenol–sulfuric acid method is routinely used for polysaccharide quantification. However, it represents a tedious procedure at a tube-scale and there is still no reliable procedure at the micro-scale for large sample analysis. By studying systematically the influence of the detecting conditions at the microplate scale, a colorimetric microassay has been developed with higher sensitivity, higher reproducibility, and higher absorbance response than the previous established protocol by Tatsuya Masuko and coworkers. As the key innovations, the influence of the vibration and the sequence of the chemical addition on the detection was elucidated for the first time at microplate scale. The microassay has been validated through the detection of large samples of polysaccharides generated from Phellinus sp under submerged cultures at the microplate scale. Moreover, its sensitivity, linear response, precision, reproducibility and feasibility were further validated through the detection of large samples containing various unknown amounts of polysaccharides from Phellinus sp. This study would contribute to the development of a reliable detection method for large samples of polysaccharides in a microplate format as an in-process analysis for the characterization of critical product/process attributes.