Citrulline test

Cyclic citrullinated peptide A circular peptide (a ring of amino acids) containing the amino acid citrulline. Autoantibodies directed against cyclic citrullinated peptide provide the basis for a test of importance in rheumatoid arthritis. 

In 1930 Wada isolated citrulline from watermelon. It was an obvious candidate as an intermediate in the urea cycle. When tested by Krebs it was catalytic only at high concentrations. Citrulline is not normally detectable in liver. 

The linearity of the AAA was tested with an aqueous mixture. In general the assay was linear up to 2500 pmol/1, for glutamine even 5000 pmol/1. Some amino acids were tested at a lower level (cystine, taurine, citrulline) and were found to be linear up to 625-1250 pmol/1. 

The large concentrations of acyl phosphatase present in most tissues led to the belief that it prevented the demonstration of acetyl-P biosynthesis and accumulation in animal tissues. We purified extensively, some years ago, what- we first thought to be a specific carba-myl-P phosphatase. This enzyme interested us as a tool for better understanding the mechanism of citrulline synthesis however, on testing for specificity, we found that it attacked acetyl-P just as readily as carbamyl-P. 

The product of the stoichiometric reaction of acetyl-P with ornithine, catalyzed by ornithine transcarbamylase, has been shown unequivocally to be 6-acetylornithine the transcarbamylases from rat liver, frog liver, and bacteria, however, even though yielding the same product, appear to differ in their ratios of activity with carbamyl-P and acetyl-P (Table n). While it is possible that the synthesis of 6-acetylornithine is catalyzed by other enzymes (16), the different ratios may be due to species differences we know now that the ratios of activity with carbamyl-P and acetyl-P of all ornithine transcarbamylases thus far tested remain constant with purification. Further, the ratio of citrulline to acetylornithine formation does not change with a number of treatments, such as heat inactivation of preparations containing orni-... 

The efficacy of the spin trapping agent PBN and the antioxidant vitamin E was tested to suppress increase in NO and hpid peroxidation and prevent neurodegeneration of pyramidal neurons in the CAl hippocampal area in the model of KA-induced excitotoxicity. Vitamin E or PBN alone did not alter basal citrulline and F4-NeuroPs levels or dendritic arborization. However, vitamin E and PBN suppressed KA-induced increases in citrulline and cerebral and neuronal markers of oxidative damage, p2-IsoPs and F4-NeuroPs, respectively (Figures 42.8 and 42.9). 

Diagnosis is based upon hyperammonaemia, which is detectable either spontaneously or after the oral intake of proteins - or, most obviously, following intravenous infusion of amino acids. The respective amino acids are increased in the serum prior to the disturbed metabolic reaction. Argininosuccinate is only detectable in the urine. A striking feature in these patients is their thin, brittle hair. With defective ornithine transcarbamylase, there is an increase in orotic acid, uridine and uric acid in the urine, while the respective citrulline concentration is decreased. The determination of OTC activity in liver tissue verifies the diagnosis and facilitates a genomic analysis. (172) The allopurinol test can be applied for the identification of heterozygosity (or the mild form of OTC deficiency). The liver shows steatosis, portal inflammation and portal fibrosis. 

Fearon, W. R. The Carbamid Diacetyl Reaction A Test for Citrulline. 

Fearon W.R. 1939. The carbamide diacetyl reaction a test for citrulline. Biochem.. 33 902-907. Butler A.R. and Hussain 1.1981. Mechanistic studies in the chemistry of urea. Part 8. Reactions of urea, 1-methylurea, and 1,3-dimethylurea with some acyloins and butane-2,3-dione (diacetyl) in acid solution. /. Chem. Soc. Perk. Trans. II2 310-316. 

FIGURE 48.8 Ipsilateral cerebral citrulline concentrations following ICV KA with or without vitamin E or PBN pretreatment. Brains from mice exposed to KA were collected SOmin post-injection (n > 5 for each group). One-way ANOVA had p < 0.001 with Bonferroni s multiple comparison tests significant for KA versus control, vitamin E + KA, or PBN + KA treatment. 

The test is given by urea and substituted ureas, of which citrulline is the only example known to occur in ordinary proteins. 

Since the preparation of this edition has required the re-writing of about three-quarters of the entire book, it hardly seems necessary to specify more than the principal alterations. Solutions and Colloidal Systems, Steroids, Pigments (Pyrrole Derivatives, Caro-tinoids, Flavins), Tissue Respiration, and Internal Environment are now made the subject of special chapters. New analytical methods include the citrulline reaction for proteins and the diffusion test for acetone. Owing to the kindness of Dr. E. J. King I have been able to reprint three of his micro-methods for blood analysis. 

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