Measurement of the unbound fraction of long chain nonesterified fatty acids (NEFA) in serum: methodological considerations.
Long-chain nonesterified fatty acids (NEFA) are extensively bound to albumin; knowledge of their unbound concentrations is important in evaluating the numerous biologic effects attributed to these compounds. We measured the unbound fraction of five long-chain NEFA in serum using the equilibrium partition of 14C-NEFA between heptane and aqueous phases. Commercial 14C-NEFA preparations gave non-linear estimates of unbound fraction with serum dilution, consistent with the presence of polar tracer impurities, but 14C-NEFA purified by alkaline ethanol extraction gave an approximately linear relationship between unbound fraction and serum dilution over a 4096-fold range of dilution, provided that pH of the aqueous phase remained stable. Mean unbound percentages were: myristic acid 0.0066, linolenic acid 0.0019, arachidonic acid 0.0017, oleic acid 0.00078 and palmitic acid 0.00061. These data suggest that some previous studies appear to have overestimated the free fraction of long-chain NEFA at physiological albumin concentrations by at least one order of magnitude.
Lim
C F
CF
Ewen Downie Metabolic Unit, Department of Medicine, Alfred Hospital, Melbourne, Vic., Australia.
Wynne
K N
KN
Stockigt
J R
JR
D001095:Arachidonic Acids / Q000097:blood; D002021:Buffers; D005230:Fatty Acids, Nonesterified / Q000097:blood*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D008041:Linoleic Acids / Q000097:blood; D009226:Myristates / Q000097:blood; D009829:Oleic Acids / Q000097:blood; D010169:Palmitic Acids / Q000097:blood
D001095:Arachidonic Acids; D002021:Buffers; D005230:Fatty Acids, Nonesterified; D007202:Indicators and Reagents; D008041:Linoleic Acids; D009226:Myristates; D009829:Oleic Acids; D010169:Palmitic Acids