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Simultaneous measurement of chlorophyll and astaxanthin in Haematococcus pluvialis cells by first-order derivative ultraviolet-visible spectrophotometry.

A first-order derivative spectrophotometric method has been developed for the simultaneous measurement of chlorophyll and astaxanthin concentrations in Haematococcus pluvialis cells. Acetone was selected for the extraction of pigments because of its good sensitivity and low toxicity compared with other organic solvents tested; the tested solvents included acetone, methanol, hexane, chloroform, n-propanol, and acetonitrile. A first-order derivative spectrophotometric method was used to eliminate the effects of the overlaping of the chlorophyll and astaxanthin peaks. The linear ranges in 1D evaluation were from 0.50 to 20.0 microg x ml(-1) for chlorophyll and from 1.00 to 12.0 microg x ml(-1) for astaxanthin. The limits of detection of the analytical procedure were found to be 0.35 microg x ml(-1) for chlorophyll and 0.25 microg x ml(-1) for astaxanthin. The relative standard deviations for the determination of 7.0 microg x ml(-1) chlorophyll and 5.0 microg x ml(-1) astaxanthin were 1.2% and 1.1%, respectively. The procedure was found to be simple, rapid, and reliable. This method was successfully applied to the determination of chlorophyll and astaxanthin concentrations in H. pluvialis cells. A good agreement was achieved between the results obtained by the proposed method and HPLC method.