Development and initial evaluation of a novel method for assessing tissue-specific plasma free fatty acid utilization in vivo using (R)-2-bromopalmitate tracer

We describe a method for assessing tissue-specific plasma free fatty acid (FFA) utilization in vivo using a non- b -oxidizable FFA analog, (9,10- 3 H)-(R)-2-bromopalmitate ( 3 H-R-BrP). Ideally 3 H-R-BrP would be transported in plasma, taken up by tissues and activated by the enzyme acyl-CoA synthetase (ACS) like native FFA, but then 3 H- labeled metabolites would be trapped. In vitro we found that 2-bromopalmitate and palmitate compete equivalently for the same ligand binding sites on albumin and intestinal fatty acid binding protein, and activation by ACS was stereoselec- tive for the R-isomer. In vivo, oxidative and non-oxidative FFA metabolism was assessed in anesthetized Wistar rats by infusing, over 4 min, a mixture of 3 H-R-BrP and (U- 14 C) palmitate ( 14 C-palmitate). Indices of total FFA utilization ( R * f ) and incorporation into storage products ( R fs 9 ) were de- fined, based on tissue concentrations of 3 H and 14 C, respec- tively, 16 min after the start of tracer infusion. R * f , but not R fs 9 , was substantially increased in contracting (sciatic nerve stimulated) hindlimb muscles compared with contralateral non-contracting muscles. The contraction-induced increases in R * f were completely prevented by blockade of b -oxida- tion with etomoxir. These results verify that 3 H-R-BrP traces local total FFA utilization, including oxidative and non-oxi- dative metabolism. Separate estimates of the rates of loss of 3 H activity indicated effective 3 H metabolite retention in