Improved stable isotope dilution-gas chromatography-mass spectrometry method for serum or plasma free 3-hydroxy-fatty acids and its utility for the study of disorders of mitochondrial fatty acid beta-oxidation

Background: Disorders of fatty acid oxidation (FAO) are difficult to diagno se, primarily because in many of the FAO disorders measurable biochemical i ntermediates accumulate in body fluids only during acute illness. Increased concentrations of 3-hydroxy-fatty acids (3-OH-FAs) in the blood are indica tive of FAO disorders of the long- and short-chain 3-hydroxy-acyl-CoA dehyd rogenases, LCHAD and SCHAD. We describe a serum/plasma: assay for the measu rement of 3-OH-FAs with carbon:chain lengths from C-6 to C-16. Methods: We used stable isotope dilution gas chromatography-mass spectromet ry (GC-MS) with electron impact ionization and selected ion monitoring. Nat ural and isotope-labeled compounds were synthesized for the assay. Results: The assay was linear from 0.2 to 50 mu mol/L for all six 3-OH-FAs. CVs were 5-15% at concentrations near the: upper limits seen in healthy su bjects. In 43 subjects, the medians (and ranges) in mu mol/L were as follow s: 3-OH-C-6, 0.8 (0.3-2.2); 3-OH-C-8, 0.4 (0.2-1.0); 3-OH-C-10, 0.3 (0.2-0. 6); 3-OH-C-12, 0.3 (0.2-0.6); 3-OH-C-14, 0.2 (0.0-0.4); and 3-OH-C-16, 0.2 (0.0-0.5). 3-OH-FAs were increased:in infants receiving formula containing medium chain triglycerides. Two patients diagnosed with LCHAD deficiency sh owed marked increases in 3-OH-C-14 and 3-OH-C-16 concentrations. Two patien ts diagnosed with SCHAD deficiency showed increased shorter chain 3-OH-FAs but no increases in 3-OH-C-14 to 3-OH-C-16. Conclusion: Measuring blood concentrations of the 3-OH-FAs:with this assay may be a valuable tool for helping to rapidly identify deficiencies in LCHA D and SCHAD and may also provide useful information about the status of the FAO pathway. (C) 2000 American Association for Clinical Chemistry.