Comparison of three methods for Chlorophyll determination: Spectrophotometry and Fluorimetry in samples containing pigment mixtures and spectrophotometry in samples with separate pigments through High Performance Liquid Chromatography

DOS SANTOS¹, A.C.A., CALIJURI², M.C., MORAES², E.M., ADORNO², M.A.T., FALCO, P.B.² , CARVALHO², D.P., DEBERDT², G.L.B. & BENASSI², S.F.

¹ Faculty of Biology. University of Santo Amaro. R. Enéas de Siqueira Neto, 340. São Paulo, S.P. andrecas@terra.com.br

² University of São Paulo. School of Engineering at São Carlos. Department of Hydraulic and Sanitation. Laboratory of Biotoxicology in Continental Waters and Sewage (BIOTACE) and Laboratory of Anaerobic Processes, Av. Trabalhador Sãocarlense, 400. São Carlos, S.P. calijuri@sc.usp.br

ABSTRACT: Chlorophyll concentration is one of the most used forms for the determination of phytoplanktonic organisms biomass. Despite the fact that this substance concentration varies due to both, the cells physiological state and species composition, it permits a faster evaluation of the natural phytoplanktonic organisms biomass, than microscopic methods. Therefore it is a useful tool in scientific studies and in hydric resources monitoring. Because of the wide utilization, some methods have been considered to determine the chlorophyll concentration, with different extraction and quantification forms. This research compares different forms of chlorophyll a concentration quantification: spectrophotometry, fluorimetry and spectrophotometry after high performance liquid chromatography separation (HPLC). From two environments (Lobo and Salto Grande Reservoir), 120 samples had been used to compare these methods. Although they have a very great correlation, two of them, spectrophotometric and fluorimetric, had overestimated chlorophyll concentration. This overestimate was more significant in the Lobo reservoir.

Key-words: chlorophyll, phytoplankton, spectrophotometry, fluorimetry, HPLC.