The inclusion of whole amaranth flo

The inclusion of whole amaranth flour in the bread formulation significantly increased the amount of phytate from non-detectable values to 2.35 mmol/g (d.m.) for the control sample and 40 g/100 g WAF, respectively. The same tendency was observed in InsP5, and even more in InsP4 and InsP3, which increased significantly with the inclusion of amaranth flour. Phytates are mainly present in outer layers of the grain, and during the breadmaking process endogenous phytate-degrading enzymes with the potential to hydrolyse phytates to InsP5eInsP3 could be active (Sanz-Penella et al., 2008, 2009).The fermentation stage used in the breadmaking process in this study was maintained for 2 h. Consequently, the endogenous phytase could have had enough time to significantly reduce the phytate content present in the amaranth flour. Even so, the amount of InsP6 and lower myoinositol phosphates increased as the whole amaranth flour was introduced in the formulation. The phytate/minerals molar ratios are used to predict the inhibitory effect of InsP6 on the bioavailability of minerals (Ma et al., 2005). The phytate/calcium molar ratio could impair calcium bioavailability in humans at values higher than 0.24. In the case of iron, if the molar ratio is more than 1; whereas if the phytate/Zn molar ratio is higher than 5 the bioavailability of Zn could be less than 50% (Ma et al., 2005). The sample made with 40 g/100 g whole amaranth flour had a phytate/Zn molar ratio value higher than 5 (phytate/Zn: 6.17). This sample and 30WAF both showed phytate/Ca molar ratio values higher than 0.24 (0.95 and 0.43, respectively). The phytate/Fe molar ratio showed values higher than 1 for the 30WAF (1.42) and 40WAF (3.00) samples. The high phytate concentration resulting from the inclusion of a high proportion of whole amaranth flour in the bread formula (30e40 g/100 g) could lead to a mineral bioavailability that is deficient, or at least reduced in the cases of zinc, calcium and iron. Studies carried out with the Caco-2 cell line supported this hypothesis, showing inhibition of the iron bioavailability of samples with 40% flour substitution. Nevertheless, the use of up to 20 g/100 g amaranth flour allowed an increase in iron uptake with regard to the control sample (Sanz-Penella, Laparra, Sanz, & Haros, 2012). Moreover, it must be emphasized that some lower myoinositol phosphates are considered compounds that could perform positive biological functions in the human body such as second messenger, bringing about a range of cellular functions including cell proliferation via intracellular Ca2þ mobilization, particularly InsP3 (Haros et al., 2009; Shi, Azab, Thompson, & Greenberg, 2006).