2-ethylhydracrylic aciduria in short/branched-chain acyl-CoA dehydrogenase deficiency: application to diagnosis and implications for the R-pathway of isoleucine oxidation.

Isolated excretion of 2-methylbutyrylglycine (2-MBG) is the hallmark of short/branched-chain acyl-CoA dehydrogenase deficiency (SBCADD), a recently identified defect in the proximal pathway of L-isoleucine oxidation. SBCADD might be underdiagnosed because detection and recognition of urine acylglycines is problematic. Excretion of 2-ethylhydracrylic acid (2-EHA), an intermediate formed in the normally minor R-pathway of L-isoleucine oxidation, has not previously been described in SBCADD. Samples from four patients with 2-MBG excretion were analyzed by gas chromatography-mass spectrometry for urine organic acids, quantification of 2-MBG, and chiral determination of 2-methylbutyric acid. Blood-spot acylcarnitines were measured by electrospray-tandem mass spectrometry. Mutations in the ACADSB gene encoding SBCAD were identified by direct sequencing. SBCADD was confirmed in each patient by demonstration of different ACADSB gene mutations. In multiple urine samples, organic acid analysis revealed a prominent 2-EHA peak usually exceeding the size of the 2-MBG peak. Approximately 40-46% of total 2-methylbutyric acid conjugates were in the form of the R-isomer, indicating significant metabolism via the R-pathway. If, as generally believed, SBCAD is responsible for R-2-MBG dehydrogenation in the R-pathway, 2-EHA would not be produced in SBCADD. Our observation of 2-ethylhydracrylic aciduria in SBCADD implies that a different or alternative enzyme serves this function. Increased flux through the R-pathway may act as a safety valve for overflow of accumulating S-pathway metabolites and thereby mitigate the severity of SBCADD. Awareness of 2-ethylhydracrylic aciduria as a diagnostic marker could lead to increased detection of SBCADD and improved definition of its clinical phenotype.