An improved phenol-sulfuric acid method for the quantitative measurement of total carbohydrates in algal biomass

Total carbohydrate analysis with phenol‑sulfuric acid method is the simplest method among many colorimetric and modern instrument analytical methods. It has great potential to be used as a rapid screening method for characterization of algal biomass for algae-based biofuel research. The reproducibility and accuracy of the conventional phenol‑sulfuric method remain to be improved when applied directly to algal biomass. In the present study, several important factors that influenced the colorimetric determination were characterized and methods to eliminate the influences were further developed. Results indicated that system temperature and the addition mode of sulfuric acid and phenol were the key parameters which affected the color development and reproducibility of the conventional methods. A new design using the premixed reagent replacing the individual phenol and sulfuric acid solution solved this problem ideally. The color inferences caused by algal pigments including chlorophylls and carotenoids were quantitatively analyzed. Solutions for reducing overestimation of total carbohydrates were also addressed. In addition, hydrolysis conditions for algae biomass such as type of acids, temperature, and time were also investigated and optimized in this study. The new modified phenol‑sulfuric acid method has been shown to be more reproducible and reliable than the conventional methods for the analysis of total carbohydrates from algal biomass.