Synthesis, analysis and characterization of deuterated analogues of vitamin A

Abstract

<p>Several deuterated analogues of vitamin A have been synthesized by two methods. The aldehyde, 20, 20, 20-('2)H(,3) retinaldehyde, was prepared by the approach of Tanis et al. (Tetrahedron Lett. 1978, 10, 869-872) based on a modification of Wittig's directed aldol condensation. Four more deuterated analogues of vitamin A were synthesized based on the use of a Wittig-Horner reaction and deuterium exchange for the incorporation of deuterium. These analogues included 20,20,20-('2)H(,3) retinal, 14,20,20,20-('2)H(,4) retinol, 12,14,20,20,20-('2)H(,5)-retinol and 10,19,19,19-('2)H(,4) retinol. The last compound contains deuterium with 99% deuterium in the cited positions. The deuterated analogues were characterized by NMR, UV, HPLC, and mass spectral analysis;These deuterated analogues were synthesized for use in an isotope dilution assay for vitamin A in humans. The majority of the technical details needed for such an analysis were worked out, including the extraction, quantitation, purification and recovery of the vitamin from plasma. The quantitative mass spectral measurement of the ratio of deuterated to nondeuterated labeled vitamin A has been refined to levels of 50-100ng standard vitamin A derivatives. Introduction of the sample into the ionization source was via gas chromatography. Conditions for efficient chromatography were necessarily worked out, including the columns, packings and temperature programs which gave a maximum response. Derivatization methodology was also examined, including the selection of reagents and conditions that give maximum yields of TMS-retinol in the 0.5-2ug range;Analysis of each step in the proposed method reveals a yield of 16% from plasma. Because of the isomerization of vitamin A and losses during GC/MS analysis, however, the method could not be effectively applied to small (1ml) plasma samples.</p>