In the EDTA chelate copper is bonded through both covalent and coordinate bonds and is the center of three locked ring structures, effectively isolating any cupric charge and rendering it totally inactive. If it were not for the competition for copper from the high chloride concentration of the marine environment, it is doubtful that chelated copper would have any effectiveness. Acetic acid with a copper salt, or cupric acetate, is sometimes recommended as a more stable form of copper. This has some merit since acetate does form a complex with copper, but this is only slightly more stable than the chloride complex. Another recommendation has been to buffer acetic acid with tris (trishydroxymethylaminomethane). Although this does not appear to be recommended to stabilize copper, but to act as a buffer, the use of tris is actually one of the better recommendations to stabilize copper that have been made. Tris is an amine compound and it forms complexes with copper in the same manner that ammonia does, rendering the copper quite resistant to magnesium carbonate adsorption, leaving it fully charged, neither sequestered nor inactivated. Although the complex is not totally stable, it represents a remarkable improvement over other types of copper. There are numerous organic compounds that are capable of forming this type of complex with copper, some more effective than others, with varying degrees of toxicity. Their use requires thorough evaluation and testing. Generally, loss of the cupric positive charge, or chelation, or both decrease the effectiveness of copper. One amine-complexede copper product that has hsown excellent results in stability, effectiveness, and low order toxicity to fish is Seachem’s Cupramine™.

What is the mechanism of parasite and fish toxicity for copper and how does this relate to marine copper types? When this question comes up it is generally suggested that copper reacts with sulfhydryl groups, inactivating vital intracellular enzymes and other proteins. Although copper does inactivate sulfnydryl enzymes and binds proteins, it does not seem likely from current knowledge of cellular biochemistry that this is a likely mechanism, Only charged (ionic) copper is effective at the usual recommended concentrations (less than 0.3 ppm copper). Membrane biochemistry suggests it unlikely that charged copper at such low concentrations could pass through cellular membranes sufficiently to cause severe intracellular damage. Body fluid analyses of treated fish show no significant increase of copper during normal treatment. This supports the supposition that ionic copper does not pass through cellular membranes. It seems more likely that ionic copper acts as a membrane poison, binding to membrane components, causing disruption of normal membrane functions, and leading ultimately to osmotic shock. This is suggested by personal observations of apparent swelling and distortion of Cryptocaryon tomites exposed to copper. Also, the secretion response of fish and their respiratory distress when treated with copper is consistent with thi.~ interpretation.

A comparison of the negatively charged chloride-cupric complex (using copper sulfate) and the positively charged amine-cupric complex (using Cupramine™) indicates that the amine complex destrovs tomites more rapidly, or at a lower concentration, than does the chloride complex. When tomites were exposed to 0.2 ppm copper as copper sulfate, or Cupramine™, those exposed to copper sulfate required close to two hours to show evidence of 50% disruption or kill, while those exposed to Cupramine™ required about 45 minutes. Fish treated with copper sulfate showed severe distress after 12 hours at 0.4 ppm and body fluids showed increased copper. Fish treated with Cupramine™ showed severe distress after 12 hours at 0.9 ppm and little increased copper in body fluids. Recovery was also more rapid after stressing with amine-complexed copper. This indicates that positively charged amine-cupric complexes are more effective than the usual negatively charged chloride-cupric complex and that larger or positively charged molecules (organically bound amine copper) are less likely to penetrate membranes (less toxic to fish) than smaller or negatively charged molecules (chloride-cupric complex). This also suggests that, possibly, the positively charged water-cupric complex is the form active against parasites while the negatively charged chloride-cupric complex is more toxic to fish. This difference between amine-complexed copper and chloride-complexed copper probably accounts for varying experiences of success and failure in treating fish with copper, since marine aquaria contain varying amounts of natural chelates and complexing agents that will increase or decrease the effectiveness and toxicity of copper. Amino acids are good examples of such agents, as thev are natural by-products of the biological environment of the aquarium, and all amino acids are powerful complexing agents. Many amine-type organics are also produced by decaying food and other decaying organic matter.

The EDTA-copper chelate passes relatively freely through membranes and fish treated with it show significant elevations of copper in body fluids. Evidence suggests slow deposition of copper in internal organs, although there are no apparent short-term toxic effects. Cryptocaryon tomites treated with 2 ppm EDTA copper show only slight evidence of osmotic shock and the apparent kill rate is at least half that observed with 0.2 ppm ionic copper, although the eventual tomite mortality is high enough to suggest that for chelated copper the toxic mechanism may only partly be osmotic shock, the balance being some type of intracellular poisoning. It can only be assumed that the same type of poisoning takes place in fish, but that mortality does not occur simply because of the greater mass of fish as compared to parasites.

A key factor in the treatment of fish with copper is probability. Success depends on killing susceptible parasites before reinfection can occur. The proportion of parasites killed in a given time is dependent on the copper concentration and the chances of reinfection are inversely proportional to the kill rate and directly proportional to the degree of crowding and the extent of the original infestation. It is important, then, to avoid crowding and to use as high a copper concentration as possible without harming the fish. For copper sulfate, a concentration of less than 0.18 ppm has about a 75% chance of success. If maintained for l0 days. Concentrations between 0.2-0.25 ppm have a 90% chance of success, but are approaching a precarious concentration for fish, about 0.3 ppm. The effectiveness and toxicity for copper sulfate., however, is highly dependent on the pH and organic content of the water. Increasing acidity increases toxicity while increasing organic content decreases toxicity. Amine-complexed copper can be used safely at 0.3-0.5 ppm with virtually 98% chance of success. For heavy infestations of resistant parasites, it can be increased quite safely to 0.7-0.8 ppm. Chelated copper is ineffective at less than 1 ppm and at 2ppm it has about a 70% chance of success with little danger to fish. Chelated copper can be increased to 2.5-3.0 ppm, but this can be dangerous for some fish. The length of exposure for any copper type can be decreased from 10-14 days to 6-8 days, without sacrificing success, if the fish are transferred to another treatment tank after the initial 4 days.

Another area of concern to the marine aquarist is the removal of copper in case of over-dosage or at the end of treatment. Chloride-complexed copper and citrate copper fall out of solution fairly rapidly by themselves with standard bottom filtration. This can be accelerated with carbon filtration. For rapid detoxification, chelating agents have been suggested. If this is done in an emergency, only soluble chelates should be used so that they may be removed by water change later. Insoluble chelates will only precipitate the copper out of solution and leave it in the filter bed where it can cause trouble at a later time. It is unwise to use any kind of copper, or copper associated product, that will deposit copper in the filter bed. Amine-complexed copper can be rapidly removed with carbon filtration. but not with bottom filtration. Chelated copper cannot be removed with either carbon or bottom filtration. Polymeric adsorbents and ion-exchangers have also been ineffective in removing chelated copper. The only way to remove chelated copper is by water change.

Formaldehyde is often recommended in conjunction with copper treatment. It is such a highly reactive substance that commercial 37% solutions actually contain less than 0.1% free formaldehyde, the balance being the reaction product with water, methylene diol. During storage, these solutions generate significant quantities of methanol and formic acid. Despite this, there seems to be sufficient anecdotal evidence to support the use of formaldehyde with copper. Short term exposures ( 1/2 to 1 hour) to about 100 ppm of this powerful irritant is tolerated by fish and seems effective in forcing parasites to detach from fish. When used in a biologically filtered aquarium for an extended time (days), 10-15 ppm have been recommended. This concentration, however, is deleterious to the filter bed and will often result in 30-60% loss of nitrifying capacity. A comparison of the apparent synergistic action of formaldehyde with copper indicates that 2-6 ppm formaldehyde is just as effective as 10-15 ppm and will effect the nitrifying capacity by less than 5%. Such a low concentration of formaldehyde alone has little antibacterial, antifungal, or antiprotozoan activity. What, then, is the mechanism of action for the combination of copper and formaldehyde?