Abstract
Tepary bean (Phaseolus acutifolius) lectins exhibit differential in vitro and in vivo biological effects, but their gastrointestinal interactions and digestion have not yet been assessed. This work aimed to evaluate the changes of a recombinant Tepary bean lectin (rTBL-1) through an in vitro and ex vivo gastrointestinal process. A polyclonal antibody was developed to selectively detect rTBL-1 by Western blot (WB) and immunohistochemical analysis. Everted gut sac viability was confirmed until 60 min, where protein bioaccessibility, apparent permeability coefficient, and efflux ratio showed rTBL-1 partial digestion and absorption. Immunoblot assays suggested rTBL-1 internalization, 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. The in silico 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). The ultra-high-performance liquid chromatography–electron spray ionization–quantitative time-of-flight coupled to mass spectrometry (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 partially resisted 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 purification of digestion samples should be conducted to identify intact rTBL-1 protein and digested peptides to assess their physiological effects.
Highlights
Some natural lectins have been investigated due to the fact of their value as phytotherapeutic agents providing health benefits [1,2]
Our results showed a decrease in the gut sac weight, indicating basolateral-apical movement that increased between 30 and 60 min with a low reversion at 90 min (Figure 1C)
For the first time, we developed a polyclonal antibody against the recombinant Tepary bean (Phaseolus acutifolius) lectin that allowed its selective monitoring through the in vitro gastrointestinal digestion using viable ex vivo tissues
Summary
Some natural lectins have been investigated due to the fact of their value as phytotherapeutic agents providing health benefits [1,2]. Lectins from the Tepary bean (Phaseolus acutifolius) have gained interest due to the fact of its lower toxicity grade than other legume lectins [3]. Tepary bean lectin fraction (TBLF), obtained through molecular weight exclusion chromatography, has shown in vitro differential cytotoxic effects on cancer cell lines, where colon cancer cells were the most sensible ones [4]. TBLF induces apoptosis in human HT-29 colorectal cancer cell death and causes cell-cycle changes such as a G0/G1 phase arrest. The results indicated lectin-induced intestinal atrophy, pancreatic hypertropia, and decreased body weight gain; the animals, recovered after a two-week resting period. TBLF can negatively affect the intestinal permeability, decreasing protein digestibility, and changes in the occludin distribution [8]
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