Flow scintillation counting of 14C-labeled microalgal photosynthetic pigments

Photopigment radiolabcling, a useful method for measuring thein situ carbon-specific growth rates of microalgae, is basedon the determination of synthesis rates of chemosystematic (i.e.specific for microalgal phylogenetic groups) chlorophylls andcarotenoids using photosynthetically assimilated ¹⁴C as a radiotracer.The reliability of this method depends on accurate measurementsof the ¹⁴C-specific activity of individual photopigments. Typically,photopigments are separated by high-performance liquid chromatography(HPLC) with fraction collection of individual peaks, followedby further purification and standard scintillation counting.To simplify analyses, we evaluated in-line flow scintillationcounting to determine its applicability and reliability formeasuring the activity of radio-labeled photopigments. Incubationswere conducted using both pure cultures and natural phyto-planktonsamples. The radiochemical purity of photopigments was determinedby extract acidification (10% HC1) to transform chlorophyllsinto degradation products. Purity was also checked by comparingabsorbance spectra with purified standards. Although ¹⁴C-labeledcolorless compounds are a common feature in radiograms, thesecompounds do not co-elute with photopigments using our HPLCprotocol. Flow scintillation counting, coupled with a highlyselective HPLC protocol, provides an efficient, reliable andfeasible alternative to fraction collection/repurification methodsfor measuring the ¹⁴C-specific activity of microalgal photosyntheticpigments.