Simultaneous determination of carotenoids and chlorophylls in algae by high performance liquid chromatography

Abstract

An isocratic reversed-phase HPLC method for the simultaneous determination of carotenoids and chlorophylls in algae is presented. Both carotenoids and chlorophylls were separated by using a C18 column and a mobile phase consisting of d ichloromethane/acetonitrile/methanol/water (22.5 : 9.5: 67. 5:0.5 ). The flow rate was 1. 0mL/min. These pigments were detected by a UV-VIS absorbance detector set at 450nm. The average recoveries of these pigments by standard addition method were 99. 1% for lutein,98. 5 % for a- carotene, 99. 4% for β- carotene, 100. 6% for chlorophyll a and 99. 9% for chlorophyll b. The relative standard deviations of these pigments were 2. 4% for lutein, 5. 6% for α- carotene, 6.0% for β-carotene, 4. 1 % for chlorophyll a and 4. 0% for chlorophyll b. The calibration curves for these pigments gave good linearity over a wide range of o-50mg/L (r=0.9992-0. 9999), The retention times were 3.0 min for lutein, 4. 7 min for chlorophyll b, 6. 3 min for chlorophyll a, 14. 1 min for α- carotene, and 14. 7 min for β-carotene. Using this method, carotenoids and chlorophylls were determined in Spirulina Platensis, Chlorella vulgaris and Haematococcus lacustris. Chlorella vulgaris and Haematococcus lacustris samples were ground in a B. Braun homogenizer. Spirulina platensis sample was smashed by supersonic wave, Acetone was used as extractant.