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 diagnose, primarily because in many of the FAO disorders measurable biochemical intermediates accumulate in body fluids only during acute illness. Increased concentrations of 3-hydroxy-fatty acids (3-OH-FAs) in the blood are indicative of FAO disorders of the long- and short-chain 3-hydroxy-acyl-CoA dehydrogenases, LCHAD and SCHAD. We describe a serum/plasma assay for the measurement of 3-OH-FAs with carbon chain lengths from C(6) to C(16).


METHODS
We used stable isotope dilution gas chromatography-mass spectrometry (GC-MS) with electron impact ionization and selected ion monitoring. Natural and isotope-labeled compounds were synthesized for the assay.


RESULTS
The assay was linear from 0.2 to 50 micromol/L for all six 3-OH-FAs. CVs were 5-15% at concentrations near the upper limits seen in healthy subjects. In 43 subjects, the medians (and ranges) in micromol/L were as follows: 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 showed marked increases in 3-OH-C(14) and 3-OH-C(16) concentrations. Two patients diagnosed with SCHAD deficiency showed increased shorter chain 3-OH-FAs but no increases in 3-OH-C(14) to 3-OH-C(16).


CONCLUSIONS
Measuring blood concentrations of the 3-OH-FAs with this assay may be a valuable tool for helping to rapidly identify deficiencies in LCHAD and SCHAD and may also provide useful information about the status of the FAO pathway.
Jones
P M
PM
University of Texas Southwestern Medical Center, Department of Pathology, and Children's Medical Center of Dallas, TX 75235, USA. PJONES@CHILDMED.DALLAS.TX.US
Quinn
R
R
Fennessey
P V
PV
Tjoa
S
S
Goodman
S I
SI
Fiore
S
S
Burlina
A B
AB
Rinaldo
P
P
Boriack
R L
RL
Bennett
M J
MJ
D015094:3-Hydroxyacyl CoA Dehydrogenases / Q000172:deficiency*; D005227:Fatty Acids / Q000378:metabolism*; D005230:Fatty Acids, Nonesterified / Q000097:blood*; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D006880:Hydroxy Acids / Q000097:blood*; D007201:Indicator Dilution Techniques; D007223:Infant; D007231:Infant, Newborn; D008052:Lipid Metabolism, Inborn Errors / Q000097:blood / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism*
D005227:Fatty Acids; D005230:Fatty Acids, Nonesterified; D006880:Hydroxy Acids; D015094:3-Hydroxyacyl CoA Dehydrogenases