Development of a Bioassay for Phytochemicals Using Daphnia pulex

Abstract

The freshwater cladoceran Daphnia pulex was explored as an alternative to Artemia salina for the biological screening of phytochemicals. This paper reports on the results of screening 27 compounds, and comparisons are made with screens using Artemia salina. The effect of miniaturisation of the assay system was investigated. Petri-dish based tests were performed for the 27 compounds using 10 daphnids at each of 5 concentrations (1 - 1000 mg l^-1). Potassium dichromate was used as a control and the number immobile after 24 hours were counted. Results were expressed as EC₅₀ values. The active compounds (EC₅₀ < 25 mg l^-1) comprised mainly adrenoceptor agonist alkaloids, while the cytotoxins vincristine and colchicine, and several antibiotics were relatively non-toxic towards Daphnia. The method was validated by comparative bioassay of A. belladonna fractions. Overall, this screen compared favourably with those based on Artemia. Daphnia were sensitive to a wide range of biologically active molecules including CNS-stimulants, anti-malarials, narcotics and anti-spasmodics, and the sensitivity was broader than previously seen with Artemia. The screen is perhaps limited by the lack of sensitivity to cytotoxins, but this might be overcome by altering the test criteria.

Key words

Bioassay - Daphnia pulex - phytochemicals - screen - biological activity

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