Retinal is not formed in vitro by enzymic central cleavage of .beta.-carotene

Abstract

Rat intestinal mucosa was prepared and incubated with beta-carotene by the procedure of Goodman and Olson [Goodman, DeW. S., & Olson, J.A. (1969) Methods Enzymol. 15, 462-475] to determine beta-carotene cleavage activity. A new detection system for the reaction products of the described enzyme beta-carotene 15,15'-dioxygenase (EC 1.13.11.21) employs solvent extraction of retinoids and carotenoids followed by high-performance liquid chromatography separation and photometric detection of the pigments. It has not detected any newly formed retinal or other retinoids in the intestinal protein preparations from normal or vitamin A deficient rats. The latter were chosen as a possible source of more active enzyme preparations. With corresponding blank samples subjected to identical conditions of incubation but without added protein, small amounts of beta-apocarotenals could be detected. They were previously reported as cleavage products of beta-carotene [Ganguly, J., & Sastry, P.S. (1985) World Rev. Nutr. Diet. 45, 198-220] but are clearly not formed as a result of an enzymatic reaction. The failure to detect in vitro enzymatic central or random cleavage of the beta-carotene molecule in extracts of rat intestinal mucosa emphasizes the need to reevaluate the existing theory of conversion of beta-carotene into vitamin A.

Enzymatic conversion of all-trans-beta-carotene to retinal by a cytosolic enzyme from rabbit and rat intestinal mucosa.

“…(iii) Although it is not absolutely essential, it is important to fractionate the BCC enzyme activity from other interfering activities, such as the retinal reductase or the oxidase, to characterize the product and estimate its true activity. It is possible that these interfering activities far exceeded the BCC activity in the previous studies (21,24), presumably because of very low BCC enzyme activity to start with due to reasons listed above. Thus, the present study in vitro fully confirms the in vivo observations (1-7) that (3-carotene is the precursor of vitamin A in biological systems and that retinal is the true product of BCC even in in vitro systems.…”

“…This was extended by a number of workers (10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20) using different species as the source ofthe enzyme. However, a more recent report (21) claimed that it could not duplicate the original work (8,9,15) and hence raised the possibility that retinal may not be the true product of (3-carotene conversion to vitamin A in vitro, although it did not question the validity of the formation of vitamin A from ,(-carotene in vivo.…”

Substrate Specificity of Purified Recombinant Human β-Carotene 15,15′-Oxygenase (BCO1)

“…Carotenoids are relatively easy to oxidize, and even no-enzyme incubations carried out for relatively long periods (1 h) with ␤-carotene produces random cleavage products (23). The intestine contains other enzymes that could potentially oxidize carotenoids (reviewed by Hansen and Maret (23)).…”

Vitamin A: Physiological and Biochemical Processing

“…The enzymatic mechanisms responsible for intestinal conversion of {3carotene to retinol have been a subject of recent controversy (23,43,59). Work by Dmitrovski (23) indicates that carotene may be cleaved both central ly, as originally reported by Goodman, Olson, and collaborators (37,38,73), and peripherally by two separate enzymes.…”