Intestinal permeability and bioaccessibility of a recombinant lectin from Tepary bean ( Phaseolus acutifolius ) using the everted intestine assay

Abstract

Tepary bean (Phaseolus acutifolius) lectins exhibit differential in vitro cytotoxicity and in vivo antitumorigenic effects against colon cancer by oral administration, but their gastrointestinal interactions and digestion have not yet been assessed. This work evaluated the changes of a recombinant Tepary bean lectin (rTBL‐1) through an in vitro/ex vivo gastrointestinal assay. We developed a polyclonal antibody to selectively detect rTBL‐1 by western blot and immunohistochemical analysis. Protein bioaccessibility, apparent permeability coefficient, and efflux ratio showed rTBL‐1 partial digestion and absorption were tested using everted gut sac until 60 min. rTBL‐1 internalization was suggested by immunoblot assays since the lectin was detected in the digestible fraction. The immunohistochemical assay detected rTBL‐1 presence at the apical side of the small intestine, potentially due to the interaction with the intestinal cell membrane. Through in‐silico approach, interactions between rTBL‐1 and some saccharides or derivatives showed high binding affinity to sialic acid (‐6.70 kcal/mol) and N‐acetylglucosamine (‐6.10 kcal/mol). UHPLC‐ESI‐QTOF/MS analysis showed rTBL‐1 presence in the gastric content and the non‐digestible fraction after intestinal simulation conditions. The results indicated that rTBL‐1 is partially resistant to the digestive conditions and interacted with the intestinal membrane, whereas its digestion allowed the absorption or internalization of the protein or the derivative peptides. Further studies will focus on rTBL‐1 protein‐glycans interaction, in the purification of samples at each step of the in vitro digestion for their identification and bioactivity determination, and also to know the rTBL‐1 internalization route in order to determine its pharmacokinetic parameters as an anticancer molecule.