Chlorophyll-a & Phaeopigments
Chlorophyll a is cold-extracted in a 90% acetone solution for ~24 hours. Chlorophyll and phaeopigments are then measured fluorometrically using an acidification technique. The method used today is based on those developed by Yentsch and Menzel (1963), Holm-Hansen et al. (1965) and Lorenzen (1967). Note that concentrations of 'phaeopigments' are not a good measure of Chl a degradation products present in the sample since Chl b present in the sample will be measured as 'phaeopigments'.
Complete methods of chlorophyll-a determination can be found here.
Seawater samples of a known volume are filtered (<=10 psi) onto GF/F filters. These filters are then placed into 10 ml screw-top culture tubes containing 8.0 ml of 90% acetone. After a period of 24 to 48 hours, the fluorescence of the samples are read on a fluorometer. Once the pre-acidified reading is saved, the sample is acidified to degrade the chlorophyll to phaeopigments (i.e. phaeophytin) and a second reading is measured & recorded. These readings, prior to and after acidification, are then used to calculate concentrations of both chlorophyll a and 'phaeopigment'.
Chlorophyll bottles are rinsed three times with sample prior to filling. The bottles are volumetrically calibrated so air bubbles should be eliminated from the sample. The sensitivity of the fluorometric method allows for sample bottles of ~50 to 250 ml.