SUMMARYA 3H+-release method has been developed for the assay of P-hydroxylation of the adrenolytic drug mitotane. P-3H-rnitotane was synthesized by the reduction of 142-chlorophenyl)-l-(4-chlorophenyl)-2,2,2-trichloroethane by an aluminium-HgZC12 couple in the presence of 3Hz0. For P-hydroxylation of mitotane, the jH+-release assay is more efficient and sensitive than a method utilizing 14C-mitotane and chromatographic separation of metabolites by HPLC. The 3H+-release assay has been used to evaluate the ability of adrenal tumors to metabolize mitotane via the Phydroxylation route.

Two novel cholesteryl ether derivatives were synthesized and radioiodinated: (1) [125I]cholesteryl m-iodobenzyl ether (125I-CIBE) and (2) [125I]cholesteryl 12-(m-iodophenyl)dodecyl ether (125I-CIDE). These radioiodinated ethers were incorporated into low-density lipoprotein (LDL) by incubating the compounds (solubilized in saline with Tween-20) with isolated LDL or with whole plasma. Such LDL preparations were taken up by cultured fibroblasts in a receptor-dependent manner similar to that of radioiodinated LDL. Upon injection into guinea pigs, 125I-CIBE-labeled guinea pig LDL cleared from the plasma similarly to radioiodinated guinea pig LDL. The primary sites of 125I-CIBE uptake were the adrenal and the liver, and the compound was stable to both hydrolysis and deiodination over 24 h. In summary, 125I-CIBE and 125I-CIDE, like previously described tritiated cholesteryl ethers, appear to be potentially useful tracers of cholesteryl ester uptake. Moreover, these radioiodinated probes have the advantage of being more easily quantitated in tissue samples as well as being detectable by noninvasive scintigraphic imaging.

Two different methods were evaluated for incorporating [125I]cholesteryl iopanoate ([125I]CI), a non-hydrolyzable cholesteryl ester analog, into LDL. The first procedure was an organic solvent delipidation-reconstitution procedure (R[125I-CI]LDL) while the second involved incubation of detergent (Tween-20)-solubilized [125I]CI with whole plasma (D[125I-CI]LDL). R[125I-CI]LDL behaved similar to native LDL in vitro, but was markedly different in vivo, apparently due to a heterogeneity in particle size. D[125I-CI]LDL, however, was metabolized normally both in vitro and in vivo. These results, combined with the residualizing nature of [125I]CI, demonstrate that D[125I-CI]LDL is appropriate for tracing LDL uptake in vivo.

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