Carbohydrate analysis by a phenol-sulfuric acid method in microplate format.

Area of Science:

• Biochemistry
• Analytical Chemistry

Background:

• The phenol-sulfuric acid method is a widely used, reliable colorimetric technique for carbohydrate analysis.
• Previous versions required significant sample amounts (50-450 nmol), limiting its use with precious samples.
• A scaled-down version existed but required 10-80 nmol of sugars.

Purpose of the Study:

• To modify and optimize the phenol-sulfuric acid method for high-throughput analysis using 96-well microplates.
• To increase sensitivity and economize reagent consumption.
• To develop a faster, more convenient method for carbohydrate quantification.

Main Methods:

• Adaptation of the phenol-sulfuric acid assay to a 96-well microplate format.
• Optimization of reaction conditions for enhanced sensitivity and a broader linear range.
• Elimination of shaking and covering steps to simplify the procedure.

Main Results:

• The modified method demonstrates a longer linear range (1-150 nmol for mannose) and excellent sensitivity.
• High-throughput capability suitable for analyzing large numbers of samples.
• Reduced reagent usage and a simplified protocol requiring less than 15 minutes.

Conclusions:

• The optimized microplate-based phenol-sulfuric acid method offers a sensitive, rapid, and economical approach for carbohydrate analysis.
• This method is highly suitable for analyzing precious samples and processing large sample volumes, such as chromatographic fractions.