The Caco-2 cell line is used by many investigators as a model of the intestinal epithelium to study nutrient uptake and transport. Our goal was to create an awareness of inherent variabilities in the Caco-2 cell line which may influence their suitability as a model or their application to specific problems. To study the influence of passage on the model, cultures were monitored from passage 20 to 109. Transepithelial electrical resistance (TEER) and sucrase activity (measured in 21-day-old cultures) increased through about passage 36. TEER values declined after about passage 60; sucrase remained elevated but variable. Cells at passage 22, 33, and 72 were grown simultaneously for 24 days. Older-passaged cells reached plateau phase sooner. Before Day 15, passage 72 cells had higher TEER and lower permeability to 14C-mannitol than passages 22 and 33; however, after Day 15 all passages showed similar permeability. On Day 21, passage 72 cells had significantly lower alkaline phosphatase activity than did the other passages. Electron microscopy did not reveal any major morphological differences between the passages; however, it did show that some areas of cells grown on membranes were not monolayers but were several cells thick with varied morphology. Investigation of the formation of these multilayered areas showed them to be an inherent part of cell growth under the conditions used. These results emphasize the inherent variability in Caco-2 cell models and emphasize the need to monitor closely the culture characteristics during growth and differentiation under specific experimental conditions.

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Foods are fortified with elemental forms of iron to reduce iron deficiency. However, the nutritional efficacy of current, commercially produced elemental iron powders has not been verified. We determined the bioavailability of six commercial elemental iron powders and examined how physicochemistry influences bioavailability. Relative biological value (RBV) of the iron powders was determined using a hemoglobin repletion/slope ratio method, treating iron-deficient rats with repletion diets fortified with graded quantities of iron powders, bakery-grade ferrous sulfate or no added iron. Iron powders were assessed physicochemically by measuring iron solubility in hydrochloric acid at pH 1.0 and 1.7, surface area by nitrogen gas adsorption and surface microstructure by electron microscopy. Bioavailability from the iron powders, based on absolute iron intake, was significantly less than from FeSO4 (100%; P < 0.05) with the following rank order: Carbonyl (64%; Ferronyl, U.S.) > Electrolytic (54%; A-131, U.S.) > Electrolytic (46%; Electrolytic Iron, India) > H-Reduced (42%; AC-325, U.S.) > Reduced (24%; ATOMET 95SP, Canada) > CO-Reduced (21%; RSI-325, Sweden). Solubility testing of the iron powders resulted in different relative rankings and better RBV predictability with increasing time at pH 1.7 (R2 = 0.65 at 150 min). The prediction was improved with less time and lower pH (R2 = 0.82, pH 1.0 at 30 min). Surface area, ranging from 90 to 370 m2/kg, was also highly predictive of RBV (R2 = 0.80). Bioavailability of iron powders is less than bakery-grade ferrous sulfate and varies up to three times among different commercial forms. Solubility at pH 1.0 and surface area were predictive of iron bioavailability in rats.