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Abstract
Humans are dependent on folate uptake from exogenous sources through intestinal absorption. Plant-based foods appear to be the richest source of folate. Pinto bean (Phaseolus vulgaris L.), was chosen for this study because of its abundance in Nebraska and is a commonly consumed dry bean. In vitro digestion coupled to the Caco-2 cell culture model was used to assess and evaluate the permeability of dietary 5-methyltetrahydrofolate (5–methyl–THF) from pinto beans. Two models were used to estimate absorption of 5-methyl-THF: folate-depleted and folate-sufficient cell monolayers. Confluence and integrity of Caco-2 cell monolayers grown on inserts were validated with the use of transepithelial electrical resistance measurements and red phenol exclusion assay. In vitro digestion method was applied to estimate the bioaccessible 5-methyl-THF from pinto beans. The bean extract was applied to the apical side of Caco-2 cell monolayers. Samples were collected in a solution, containing 2% (v/v) ascorbic acid. The differentiated Caco-2 cell monolayers grown on inserts were treated with increasing concentrations of: 5-methyl-THF (190 μM); 5-methyl-THF+2.5%, v/v legume extract (193 μM); 5-methyl-THF+5%, v/v legume extract (202 μM); 2.5%, v/v legume extract (4 μM); or 5%, v/v legume extract (7 μM) for 80 min at 37°C without agitation. Transport experiments were performed under “sink” conditions. The permeability coefficients were determined in the apical-to-basolateral direction. 5-methyl-THF was transported across the intestinal mucosa with permeability coefficients of 1.03–1.79×10–5 cm/s in folate-depleted and with permeability coefficients of 0.99–1.32×10 –5 cm/s in folate-sufficient cell monolayers. The cumulative fractions of transported 5-methyl-THF increased linearly for both folate-depleted and folate-sufficient cell monolayers. The results of these studies demonstrate that 5-methyl-THF is bioaccessible from pinto bean extract and taken up by the intestinal Caco-2 cells. The experimental conditions were established for the use of in vitro digestion/Caco-2 cell culture model to assess intestinal absorption of 5–methyl–THF from pinto beans. The in vitro digestion method mimics the in vivo digestion process and provides an estimation of bioavailable 5-methyl-THF from pinto beans, which mimics the human gastrointestinal physiology.
