The purposes of this study were to determine the location of beta-carotene dioxygenase (EC 1.13.11.21) activity within the rat gastrointestinal tract, within the villus and within enterocytes, and to identify the metabolites produced in each intestinal fraction. In Wistar female rats, maximal activity was detected in the cytosol (74-93% of the total cellular activity) of mature functional enterocytes harvested from the jejunum (67% of the intestinal activity). The specific activity, expressed in pmol of retinoids/(h x mg protein) rose from 49 +/- 3 in the stem cells to 199 +/- 12 in the mature functional cells (P < 0.05). Thus the intestinal beta-carotene cleavage activity might be regulated during the enterocyte maturation process. By using HPLC with diode array and radioactive detectors, retinal, and in the presence of NAD+, retinoic acid, were identified as the only metabolites produced. No beta-12'-, 10'-, and 8'-apo-carotenals were detected, even when various enzyme sources were tested. These results suggest that the major, if not the sole, pathway for the formation of vitamin A from beta-carotene in the rat intestine is central cleavage.

SummaryIn this paper we report the synthesis of all-trans Retinoic acid, Retinal, Retinol specifically labelled with I4C at position 7 and all-trans Beta-carotene labelled at positions 7 and 7', with more than 98 % radiochemical purity. All products were obtained with about 15 % overall yield from I4C sodium cyanide.

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Carbonation of phenyl lithium with 14CO2 followed by reduction of [14CO2] benzoic acid, led to [α‐14C] benzyl alcohol 3, the oxidation of which afforded the [α‐14C] benzaldehyde 4. The latter, then condensed with pivoylamide 6 (trimethyl acetamide‐N‐(2‐lithio‐4chlorophenyl) gave 5‐chloro‐2 (t‐Butylamide)‐α‐phenyl‐[‐α‐14C] benzyl alcohol 7 which was then oxidized to ketone 8 which it self were subsequently hydrolysed to 5‐chloro‐2‐amino‐[‐α‐14C]benzophenone 9. Grignard reaction of 9 with methylmagnesium iodide gave 5‐chloro‐2‐amino‐α‐methyl‐α‐phenyl‐[‐α‐14C] benzyl alcohol 10. Treatment of the alcohol 10 with ethylthioisocyanate, followed by cyclisation led to 6‐chloro‐2‐(ethylamino)‐4‐methyl‐4‐phenyl‐[‐4‐14C]‐4H‐3,1 benzoxazine 12. The latter was acidified to give rise to Etifoxine hydrochloride 13 in an overall yield of about 31% based on [14C] barium carbonate. © 1997 John Wiley & Sons, Ltd.