Transport of 5-methyltetrahydrofolic Acid Across the Rat Jejunum

Abstract

The intestinal transport of 5-methyltetrahydrofolic acid was investigated using various in vitro preparations of rat jejunum. The transport of 5-methyltetrahydrofolic acid using the everted sac preparation was linear with increasing incubation time at the mucosal 5-methyltetrahydrofolic acid concentration of 10-5M. Tissue uptake was appreciable  whether there was any concentration gradient between the mucosal and serosal medium or not. Total transport studies carried out at mucosal concentrations of 5-methyltetrahydrofolic acid ranging from 10-7M to 10-4M failed to show any saturation of uptake.  When the tissue uptake and serosal transfer of 5-methyltetrahydrofolic acid were considered separately at the aforesaid concentrations no saturation could be observed. Absence of saturation kinetics, low temperature coefficient and low serosal to final mucosal 5-methyltetrahydrofolic acid concentration ratio fail to advocate, in any way a carrier mediated transport mechanism for 5-methyltetrahydrofolic acid. Both 5-formyltetrahydrofolic acid and 2,4-dinitrophenol (DNP) were found to depress the tissue uptake and serosal transfer of 5-methyltetrahydrofolic acid if present in the mucosal medium. 5-formyltetrahydrofolic acid may compete for the brush-border binding site and thus reduce tissue uptake of 5-methyltetrahydrofolic acid but the reduced serosal transfer in presence of 5-formyltetrahydrofolic acid is difficult to explain. DNP by inhibiting proton formation may lead to a rise in the pH of the microclimate with a corresponding fall in the concentration of permeable zwitterion at the surface of the epithelial cell. It was found that the radioactive mucosal efflux from the everted sacs of rat jejunum preloaded with the labelled d-methyltetrahydrofolic acid was stimulated in the presence of mucosal unlabelled 5-methyltetrahydrofolic acid, folic acid, methotrexate and 5-formyltetrahydrofolic acid. Serosal transfer and tissue content of the labelled compounds from the sacs preloaded with the labelled 5-methyltetrahydrofolic acid were lower in presence of mucosal 5-methyltetrahydrofolic acid than in its absence. The uptake of 5-methyltetrahydrofolic acid by preparations of isolated jejunal mucosal cells was also studied.  At pH 7.0 all the uptake takes place within 3 minutes and very little or none in the following 3 minutes and 5 minutes after that. It appears that there is a rapid binding of 5-methyltetrahydrofolic acid to cell surface protein rather than transport into cell. At pH 5.0 there was again a rapid uptake within the first 3 minutes followed by a slower but significant uptake in next 3 minutes. This latter uptake represents transport into the cell. But there is no uptake after 6 minutes indicating saturation.  Thus in terms of total uptake at 6 minutes or uptake between 3 and 6 minutes the uptake is greater at pH 5.0 than at pH 7.0. A similar situation may be found at 27°C. Histochemical studies showed that these cells had lost the glycocalyx thus indicating its importance for the transporting of 5-methyltetrahydrofolic acid into the cell.  At pH 5.0 ebout. 15% of 5-methyltetrahydrofolic acid is present as neutral zwitterion and this is sufficient to allow transport into the cell to occur without the glycocalyx. The results of this study have been discussed in the perspective of the previous studies on 5-methyltetrahydrofolic acid and the parent compound folic acid and it has been suggested that 5-methyltetrahydrofolic acid is converted to the neutral zwitterion in an acid microclimate at the mucosal surface of the jejunum prior to transport and then transported by passive diffusion and also to some extent by solvent drag.