Novel in situ visualisation of rat intestinal absorption of polyphenols via matrix-assisted laser desorption/ionisation mass spectrometry imaging

Abstract

Matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) is presently used in physiological evaluations for visualisation of targets in organs. In the present study, MALDI-MSI was used as a visualisation technique to investigate the intestinal absorption of polyphenols. Nifedipine/phytic acid-aided MALDI-MSI was performed to visualise theaflavin-3′- O -gallate (TF3′G) and epicatechin-3- O -gallate (ECG) in the rat jejunum for 50-µM, 60-min transport experiments. Non-absorbable TF3′G was successfully visualised at the apical region, whereas absorbable ECG was detected throughout the rat jejunum. MALDI-MSI was also performed to determine the transport routes of the target metabolites. Signals corresponding to TF3′G and ECG in the membranes were diminished following treatment with inhibitors targeting the monocarboxylic acid transporter and organic anion transporting polypeptides. Enhanced visualisation of TF3′G was achieved by inhibiting efflux routes. Our findings demonstrated that the present MALDI-MSI can provide critical spatial informations on intestinal absorption of targets, by which TF3′G and ECG were incorporated into intestinal tissues, followed by efflux back to the apical compartment. In addition, MALDI-MSI analyses suggested that TF3′G was resistant to phase II metabolism during the influx/efflux processes, whereas ECG was susceptible to methylation and sulphation reactions. In conclusion, inhibitor-aided MALDI-MSI could serve as a powerful in situ visualisation technique for verifying intestinal transport routes and investigating the metabolism of penetrants.

Detection and Visualization of Food-derived Polyphenols by Matrix-assisted Laser Desorption/Ionization Mass Spectrometry Imaging

“…To elucidate the possible absorption route(s) of polyphenols, phloretin (an MCT inhibitor), (57) estrone-3-sulfate (an OATP inhibitor), (55) and wortmannin (a transcytosis inhibitor) (58) have been used in intestinal ECG transport experiments. (53) The significant reduction in the level of ECG distribution in the intestinal membrane in the presence of phloretin and estrone-3-sulfate clearly demonstrated the involvement of MCT and OATP in the intestinal transport of ECG, respectively [ Fig. 4(a)], was reported by Vaidyanathan and Walle.…”

“…4(b)]. (53) Thus, these visualized results demonstrated for the first time that TF3'G can be incorporated into intestinal tissues through MCT or OATP followed by efflux back to the apical compartment, and this discovery will help elucidate the mechanism of the regulatory effect of TF3'G on the expression of PepT1 and TJ proteins. Such observation of tissue incorporation of nonabsorbable polyphenols is due to the great advantage of in situ tissue visualization by MALDI-MS imaging, which indicates the potential use of the present MS imaging technique for the elucidation of intestinal transport mechanisms of substrates including absorbable and nonabsorbable polyphenols.…”

“…3(c)]. (53) The involvement of intestinal membrane uptake transporters such as monocarboxylic transporter (MCT) and organic anion-transporting polypeptide (OATP) in the cellular uptake of polyphenols has been reported. (48,(54)(55)(56) Therefore, the elucidation of intestinal absorption routes for polyphenols is important.…”

“…Thus, the inhibitor-aided MALDI-MS imaging technique is a powerful and novel analytical tool for the direct analysis of the intestinal absorption of analytes without the pretreatment steps, such as extraction and separation, required in LC and LC-MS analyses. (53) Inhibitor-aided intestinal MALDI-MS imaging has also been used to investigate the absorption of nonabsorbable theaflavin-3'-O-gallate (TF3'G), (47) which can regulate the intestinal expression of peptide transporter 1 (PepT1) and tight-junction (TJ)-related proteins (claudin-1, ZO-1, and occludin) in order to inhibit the intestinal transport of peptides and TJmediated substrates. (62,63) Interestingly, although TF3'G was not absorbed, the apical-specific localization of TF3'G on an intestinal membrane after Ussing chamber transport was clearly visualized [ Fig.…”

“…5(b)]. (53) Considering the results of a previous study in which TFs and their phase-II metabolites were not detected in human urine after the oral administration of 1000 mg TF extract, TFs seem to be stable against phase-II metabolism in the intestinal tissue. (71) Moreover, MALDI-MS imaging has also been applied in the in vivo study of polyphenol metabolism, in which sulfate and glucuronide metabolites of EGCG were visualized in liver and kidney tissues after its oral administration.…”