Long-chain acyl-CoA profiles in cultured fibroblasts from patients with defects in fatty acid oxidation.

Negative chemical ionization (NCI) mass spectrometry was used to quantify the acyl-CoA intermediates present in human fibroblasts growing in media containing the long-chain fatty acid, palmitate. The acyl-CoA intermediates were detected as the N-acyl pentafluorobenzyl glycinates. In fibroblasts from normal individuals only saturated acyl-CoA esters were detected, supporting the concept that the acyl-CoA dehydrogenase reaction is the rate-limiting step of intramitochondrial fatty acid oxidation. In patients with inherited enzymatic defects of intramitochondrial long-chain fatty acid oxidation, there was not a significant increase in the amount of long-chain acyl-CoA compounds, with palmitoyl-CoA amounts similar to those found in controls. However, there was a sharp decrease in the relative amount of lauroyl-CoA and a resultant sixfold elevation in the palmitoyl-CoA:lauroyl-CoA ratio. In contrast, fibroblasts with a defect involving the transport of fatty acids across the mitochondrial membrane, carnitine palmitoyl transferase 1 deficiency, had a fourfold increase in palmitoyl-CoA. Our results suggest that acyl-CoA esters in biological tissues are readily detectable using NCI mass spectrometry. This approach is significantly more sensitive than previous methods for the detection of these important metabolic intermediates, and may prove useful in the study of fatty acid oxidation in both normal and enzyme-deficient tissues.