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Rainbow trout (250 g) were maintained at 15 degrees C for 3 months on a low ascorbic acid diet, given [1-14C]ascorbic acid by gavage, then fed the NAS/NRC requirement 12 times per week. Total urine, fecal water and branchial water were collected daily from five fish placed in metabolism chambers for four successive 5-day periods. Tissue samples were analyzed for 14C, ascorbic acid (C1) and ascorbate-2-sulfate sulfate (C2). Excretion analysis indicated t1/2 = 42 days. After 20 days, the feeding schedule was changed to 3 times per week. Fish fed 14C were sampled after 1, 2, 3 and 4 months. The half-life in each organ except brain was inversely proportional to the dietary level of ascorbate. Concentrations of C1 and C2 in the various tissues reflected dietary intake of vitamin C. Total C (CT = C1 + C2) levels were maintained in the liver even with the low vitamin C diet. Estimates of body pool for C1 are 27-29 mg/kg. At the higher ascorbate intake CT was 92-114 mg/kg, but decreased by 34% at the lower feeding rate to 51-62 mg/kg. Data indicate that there are two or more body pools that include a store of C2, which is readily interconverted in metabolizing tissues to and from C1. Since air and water stable C2 is antiscorbutic for fish, it is the preferred form of ascorbate for fish feeds.

Although most vertebrate animals synthesize L-ascorbic acid (C1), some animal species lack the ability to produce L-gulonolactone oxidase and are thus dependent upon a dietary source of vitamin C. Fish are unique among this latter group in that they store a chemically stable form of vitamin C and appear to metabolize this compound differently from other vitamin C-requiring organisms. Ascorbate-2-sulfate (C2) contributes to total body stores of ascorbate, but the commonly used assays for ascorbate concentrations in tissues and body fluids do not generally measure C2. An HPLC assay distinguishes between and measures both C1 and C2. Modification of the less exact but commonly used DNPH method can provide adequate data to estimate total vitamers C, C1, and (by difference) C2. Since vitamin C is a required component of feed for salmonids, catfish, eels, shrimp and carp, use of C2 in feed formulation would provide a bioavailable form of ascorbate which is heat and water stable at pH 4-13.

a potentially fatal condition resulting S from inadequate hydroxylase activity, and causing reduced collagen formation, has been known and feared since ancient times. Its prevention and cure were associated with consumption of fresh fruitsespecially citrusbut it was not until 1928 that the antiscorbutic factor was identified. Although most vertebrate animals synthesize L-ascorbic acid (AA), several species (primates, guinea pigs, fish, bats, insects and some birds) lack the ability to produce L-gulonolactone oxidase and are thus dependent upon a dietary source of vitamin C. Fish are unique among these animals in that they have a system to store a chemically stable form of vitamin C which appears to allow metabolism of this compound differently from other vitamin C-requiring organisms.McCay and Tunison' first observed scoliosis and lordosis (broken back syndrome) in brook trout fed formalin-preserved meat. Since these signs took a year to develop they were not then recognized as being related to vitamin C deficiency. These malformations were later identified as indications of scurvy and trout and salmon were added to the list of scurvy-susceptible animal^.^,^ More recently, other species of fish requiring dietary ascorbic acid have been identified and supplementary vitamin C is necessary in all aquaculture rations. Signs of the deficiency in fish are reduced growth, anorexia and lethargy, followed by structural deformation of skeleton and cartilage, hemorrhage and anemia.4 Wild fish consume insects, Barbee W. Tucker, Ph. D., is Assistant Professor in the Department of Nutrition Sciences and John E. Halver, Ph. D., is Professor of Fisheries at the School of Fisheries (WH-lo), University of Washington, Seattle, WA 98195.

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