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Updated: May 8, 2026
The Caco-2 Cell Bioassay for Measurement of Food Iron Bioavailability
Published on: April 28, 2022
Bioavailability of Sodium Iron Ethylenediaminetetraacetate, Ferrous Fumarate, and Ferrous Sulfate in Corn Flour Using
Jiejia Zhang1, Brian L Lindshield1
1School of Health Sciences, College of Human Health and Sciences, Kansas State University, Manhattan, KS, United States.
Background:
The static digestion/Caco-2 cell model has been widely employed as a bioassay to assess iron bioavailability. However, there are many methodological differences in how it is utilized. Thus, the feasibility of different chamber systems and iron treatment options was examined.
Objectives:
To investigate iron bioavailability of INFOGEST digesta compared with supernatant in Caco-2 model with Glahn's dual-chamber system (dialysis membrane), an alternative dual-chamber system (insert membrane), or a triple-chamber system (insert membrane + dialysis membrane). To determine the bioavailability of precooked corn flour fortified with sodium iron ethylenediaminetetraacetate (NaFeEDTA), ferrous fumarate, ferrous sulfate (FeSO4), and their combinations.
Methods:
Precooked corn flour fortified with NaFeEDTA, ferrous fumarate, or FeSO4 were digested following the INFOGEST method. The iron solubility of supernatant was analyzed, and Caco-2 cells were treated with iron digesta or supernatant in the dual-chamber or the triple-chamber systems for 2 h, respectively. The ferritin concentrations were quantified to compare systems. Finally, the bioavailability of NaFeEDTA, ferrous fumarate, and FeSO4 combinations was measured using Glahn's dual-chamber system.
Results:
Iron solubility of NaFeEDTA was significantly higher than ferrous fumarate and FeSO4. The combinations of NaFeEDTA and fumarate/FeSO4 at the 4:9 weight ratio had significantly higher iron solubility than combinations of NaFeEDTA and fumarate/FeSO4 at the 2:11 weight ratio. Ferritin concentrations in Glahn's dual-chamber system were significantly higher than the alternative dual-chamber and the triple-chamber systems. The combination of NaFeEDTA and FeSO4 at a 2:11 weight ratio had significantly higher ferritin concentrations than the other combination samples.
Conclusions:
The Caco-2 cell model with Glahn's dual-chamber system and INFOGEST digesta was the best treatment and chamber system. NaFeEDTA had significantly higher bioavailability than FeSO4, but not ferrous fumarate. The combination of NaFeEDTA and FeSO4 at a 2:11 weight ratio was the most promising iron treatment ratio.
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