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Unlabelled: In the human gastrointestinal tract, trypsin and mucin may affect the absorption of heme iron. However, these interactions have not been well-established. We determined the effect of trypsin and mucin on heme iron absorption in humans., Design: Twenty-eight apparently healthy females participated in two studies (14 per study). Study A evaluated the effect of trypsin on iron bioavailability. Subjects ingested 100 mg trypsin and 1.7 g mucin on 5 mg heme iron bioavailability on days 1, 2, 14, and 15, respectively. In study B, which assessed the effect of mucin on heme iron bioavailability, the subjects ingested hemin, hemin plus mucin, hemoglobin (Hb), and Hb plus mucin, on days 1, 2, 14, and 15, respectively., Results: In study A, the geometric means ± 1 SD of heme iron absorption were 5.1 % (3.1-8.3), 2.9 % (1.6-5.1), 7.3 % (4.1-13.1), and 6 % (2.7-13) for hemin, hemin plus trypsin, Hb plus trypsin, and Hb plus mucin plus trypsin, respectively. In study B, the geometric means ± 1 SD of heme iron absorption were 16.4 % (10.5-25.7), 13.1 % (9.0-18.9), 13.7 % (9.0-20.7), and 11.8 % (7.6-18.3) for hemin, hemin plus mucin, Hb, and Hb plus mucin, respectively. The ratio increased when Hb plus trypsin was ingested and decreased when hemin plus trypsin was ingested. There were no differences in other ratios with respect to the ratio on day 1 (P < 0.05)., Conclusion: Trypsin is the only human gastrointestinal protein that evaluated the affects of heme iron absorption. However, this effect depends on how heme iron is ingested.