The toxicity of the antiparasitic mixture FMC is changed by humic substances and calcium

An inexpensive mixture of the therapeutic agents malachite green (MAG), methylene blue (MB) and formalin (known as FMC) is used to control external fungal infections in fish eggs (e.g. Saprolegnia ssp., Achlya ssp.) in aquaculture and ornamental fish breeding. The treatment success as well as the toxicity of the chemicals is determined by the toxicity of each of the substances and synergistic effects among them. Moreover, toxicity and therapeutical index might be affected by water parameters such as the ion content and the presence of humic substances (HS). Four different test waters with combinations of low (0.2 mmol L-1) and high (2 mmol L-1) calcium (Ca), +HS (5 mg L-1 C) or -HS (0 mg L-1 C) were used to determine the toxicity of FMC to early life stages of zebrafish, Danio rerio (Hamilton-Buchanan, 1822, 1823). Thus, the test solutions differed only in the content of dissolved organic carbon, and/or the amount of Ca chloride, and, to a lesser extent, pH. Zebrafish eggs reaching the four- to eight-cell stage were exposed to different concentrations of FMC (10, 15, 22, 33, 50 microliter L-1 FMC) for 144 h. Survival was highest in low Ca/+HS-solution. High Ca and HS did not better protect the eggs than HS alone, so that this group had the lowest survival. Survival of the '-HS-groups' was intermediate. The toxicity of the mixture as well as the interaction with Ca and HS was mainly determined by MAG. MAG, the most toxic part of the mixture, is positively charged and competes with Ca for the negatively charged binding sites of the HS. In presence of high Ca, the binding sites of the HS are filled with Ca-ions. This causes lower binding of MAG to the HS. Thus, MAG binds to the juvenile fish and enhances the toxicity of the mixture. Therefore, the toxicity as well as the therapeutical index may be altered in the presence of HS and the Ca content of the water.