An incubation system containing rat jejunoileum was used to determine whether analogues of cyanocabalamin (CNCbl) inhibited the in vitro absorption of CNCbl. The naturally occurring analogues "pseudo B12," "factor A," and the nonnaturally occurring analogue desdemethyl B12, were produced by guided biosynthesis with Propionibacterium arabinosum. Factor B was produced by acid cleavage of cyanocabalamin. Inhibition of Co57-CNBbl uptake was demonstrated for cold CNCbl and for desdemethyl B12, although statistical significance was achieved only for cold CNCbl at 40X and 100X the molar quantity of the radiolabeled CNCbl. However, a trend towards progressive inhibition of specific CNCbl binding was shown as increasing concentrations of CNCbl or desdemethyl B12 were used. No inhibition of specific CNCbl binding could be demonstrated for any of the naturally occurring cobamides tested.