Extraction and quantification of pigments from a marine microalga: a simple and reproducible method

The pigment content in microalgae is a specific feature of each species. Its evaluation is essential as an indirect measure of cell growth, as well as a parameter to check the trophic level of waters. Several methods can be found in literature, most of them based on spectrophotometric analyses, referred as a good and practical tool for chlorophyll evaluation. A comparison of different methods to quantify pigments was done, using as selection criteria the simplicity, the amount of pigments identified and the time required for the analysis. Several factors were tested to maximize the yield of pigments extraction, such as: i) the solvent to be used, ii) the cell wall disruption technique, iii) the extraction time and iv) the use of different empirical correlations. The marine microalga Nannochloropsis gaditana was the biological material used for this study because chlorophyll is the most abundant pigment in this strain. Methanol was shown to be the most suitable solvent to extract chlorophyll from this strain, using the Lichtenthaler correlation [1], after 24 hours of extraction. Furthermore the lysis ability of the solvent and the use of additional cell wall disruption techniques favour the extraction yield. The freezing/unfreezing technique with liquid N2 was found to be more efficient than the use of ultrasounds for 15 min. To extract more polar pigments, such as carotenoids, hydrophilic solvents like methanol play the main role in extraction, which is not so influenced by the use of physical or mechanical disruption methods.