Summary MATERIALS AND METHODSThe abnormal metabolites 3-hjdroxypropionic acid (1.6-4.0 SUBJECT mglday) and methylcitric acid (3.7-5.8 mglday) were identified and quantitated in the urine of a patient in whom biotin-responAt the times the urine samples were collected for analysis, the sive 3-methylcrotonylglycinuria and deficiency of 3-methylcro-patient J R was 21 or 23 months of age. His weight was 11 kg tonyl-CoA carboxylase had previously been documented. The and his height 81.6 cm. His level of development was normal for level of excretion of these metabolites was in the lower range of his age. H e was receiving 0.5 mg/day biotin, and a protein intake those found in patients with propionic acidemia in whom there is of about 3.2-3.6 g/kg.day. The patient was in excellent condia deficiency of propionyl-CoA carboxylase. The activity of this tion and not ketotic or acidotic. Specimens were obtained with enzyme in fibroblasts derived from the patient and grown in informed consent. media low in biotin was 4% of normal. This is in the range of ANALYSIS OF URINE patients with propionyl-CoA carboxylase deficiency. Documented deficiency in this patient of two carboxylases, both of Urinary organic acids were analyzed quantitatively by liquid which contain biotin, suggests that the primary defect is in the partition chromatography on H2S04-hydrated silicic acid eluted metabolism of biotin.with a gradient of t-amyl alcohol in chloroform. The eluted acids were continuously titrated by mixing with sodium o-nitrophenol (16, 21) and the titration of the indicator was recorded spectrophotometrically by the increase in absorbance at 350 nm. Some Speculation analyses were performed at twice the usual flow rate to shorten ~h~ deficiency of two mitochon~rial carboxylases in a patient the time of analysis to 3.5 hr. Equivalents of acid were calculated suggests the presence of a fundamental defect in either the from peak areas relative to pcak areas of standards. Fractions transport of biotin or in the holocarboxylase synthetase that obtained from liquid partition chromatography were analyzed attaches biotin covalently to both carboxylases. further by gas liquid chromatography (GLC) and mass spectrometry (GCMS) of the trimethylsilyl (TMS) derivatives on Dexsil 300 using an H P 402 gas chromatograph and an LKB 9000 gas chromatograph-mass spcctromcter as previously dc-3-Methylcrotonylglycinuria is a rare inborn error uf leucine scribed (21). Rctcntion times on GLC were expressed as metabolism which is characterized by excessive excretion of 3-methylcne unit (MU) values by co-injection of even-numbered methylcrotonylglycine and 3-hydroxyisovaleric acid. The second alkancs. Acids cocluted on the liquid partition chromatography patient reported differed from the first (10, 20) in that he was system were quantitated by GLC. severely ketoacidotic, and he responded both clinically and biochemically to biotin (14). Biotin is a cofactor of 3-methylcro-ENZYME ASSAY ton~l-CoA carboxylase. Deficiency of 3-meth~lcroton~l-CoA Fibroblasts derived f...

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