HPLC ANALYSIS OF INVITRO HEPATIC DEIODINATION PRODUCTS OF THYROID-HORMONES IN THE RAINBOW-TROUT, ONCORHYNCHUS-MYKISS

Reverse-phase HPLC employing five different solvent systems was used to determine the 125I-labeled products formed in rainbow trout by in vitro incubation at 12° of the hepatic microsome fraction with l-thyroxine (T4) labeled with 125I in the outer phenyl ring. Trout were starved for 2 weeks or fed a 2% ration. The only labeled products identified during incubation for 7.5-70 min over a T4 substrate range of 0.03-0.5 nM were 125I- and 3,5,[125I]3′-triiodo-l-thyronine (T3). These products in combination with the parent [125I]T4 accounted for over 96% of the total chromatographic radioactivity. Neither 3,[125I]3′-diiodo-l-thyronine nor 3,[125I]3′,5′-T3 (reverse T3) was detected, suggesting negligible inner-ring deiodination of T4 or T3. Essentially equal production of 125I- and [125I]T3 validated the use of 125I- production as a measure of [125I]T3 generation in assays for hepatic 5′-monodeiodinase activity. However, in some experiments the 125I- level slightly exceeded the [125I]T3 level, indicating that outer-ring deiodination of T3 may occur to a limited degree. In conclusion, the present data for liver support earlier observations from in vivo studies in showing that for trout at 12° deiodination pathways are geared primarily toward T4 outer-ring monodeiodination to produce T3 with undetectable inner-ring deiodination of T3 or T4 and limited outer-ring deiodination of T3. © 1992.