Our results showed that 90 days of supplementation with 30 mg/day of elemental iron as either ferrous sulfate or iron bis-glycinate chelate had a positive effect on raising ferritin concentration in school-age children with low iron stores. This effect was seen one week post-supplementation and was still present 6 months after supplementation. Furthermore, the bis-glycinate chelate compound was more efficient for maintaining higher ferritin concentration 6 moths after supplementation than ferrous sulfate. Persistence of the observed effect for this long period after supplementation may be explained by the effect of a diet that may provide enough bioavailable iron to maintain iron status once the initial deficit was overcome. Actually, the effect of iron bis-glycinate chelate may be larger where individuals consume a typical Mexican diet (as do families with low incomes, such as the ones included in this study), which includes phytic acid from cereals (i.e., maize) and legumes (i.e., beans). This compound binds iron, reducing its bioavailability for intestinal absorption.
In a longitudinal study about iron deficiency due to consumption of low bioavailable iron diet in Africa by schoolchildren with normal iron status after a fortification intervention, after 15 months the authors found a mean change in total body iron stores of −142 mg, and estimated a 2% mean iron absorption of dietary iron. Hemoglobin decreased by 12 g/l, and 75% of the cohort had deficit in tissue iron. The authors concluded that low iron bioavailability from a legume- and cereal-based diet is a cause of iron deficiency anemia in children in rural Africa . In our study, there were no differences in the odds of having low iron storages between the two iron compounds at either evaluation period. This could mean that the higher ferritin concentration at 6 months post-supplementation in the bis-glycinate chelate group versus ferrous sulfate group had no clinical implication.
Our findings concurred with other studies carried out on pregnant women, adolescents, school-age children, and preschool children in which both ferrous sulfate and iron bis-glycinate chelate were found to be equally effective [20, 33]; however, in those studies, the effects 6 months post-supplementation had not been evaluated. Results of a systematic review and meta-analysis on iron supplementation on children, that included 32 studies including 7,089 children aged 5–12 years; 31 of these studies were conducted in lower- or middle-income countries  and showed that iron supplementation improves hematologic and non-hematologic outcomes. Iron supplementation decreased the prevalence of anemia by about 50% and reduced the prevalence of iron deficiency by 79%; there was also evidence of a benefit on cognitive performance in children with anemia at baseline . The increase in hemoglobin or ferritin concentrations is related to iron status at baseline, dose of iron, and length of time of supplementation [3, 34, 35]. However, most studies have focused on treating anemia more than on treating the iron deficiency to prevent iron deficiency anemia.
In a supplementation study carried out in Brazil that compared administration of ferrous sulfate (40 mg of iron per week) versus iron bis-glycinate chelate (3.8 mg per week as bis-glycinate chelate-enriched cookies) for 8 weeks on schoolchildren with anemia, the authors reported a significant increase in Hb concentration but no significant difference in the inter-group comparison. No effect was observed on serum ferritin for either intervention, but children with depleted iron stores (ferritin <15 ng/mL) at the beginning of the intervention showed increased serum ferritin concentration, although no difference between treatments was observed .
Ribeiro and Sigulem supplemented anemic children between 6 and 36 months with 5 mg/kg/d of iron as bis-glycinate chelate  and found a significant increase in the Hb concentration; however, ferritin concentration was not measured. Pineda and Ashmead supplemented children with anemia and malnutrition between 6 to 36 months of age with ferrous sulfate versus iron bis-glycinate chelate offering 5 mg/kg/d for 28 days. This study showed a significant increase in Hb and ferritin concentration in both supplemented groups. The difference in Hb concentration between the two groups post-intervention was not statistically significant; however, the difference in the serum ferritin concentration was significantly higher in the group that received iron bis-glycinate chelate . In summary, these studies show a positive effect of supplementation with ferrous sulfate and of supplementation or food fortification with ferrous bis-glycinate chelate on treating anemia, but we found no published study that focused on preventing iron deficiency anemia in schoolchildren population that included the bis-glycinate chelate iron compound.
As mentioned, iron bis-glycinate chelate has been used for food enrichment and for supplementation. Due to its chemical structure (consisting of a covalently bounded iron molecule to an organic ligand, in this case glycine), this form of iron can be partially resistant to the action of enzymes and to the binding action of substances naturally present in food such as metals, dietary fiber, phytates, and phenols, with which the iron can form insoluble compounds. In addition, because of the amino acid bonding, there is less direct exposure of iron to the gastrointestinal mucosa cells; this can reduce local toxicity and side effects attributable to iron compounds present in the intestinal lumen, like abdominal pain [18, 19, 33, 37, 38]. In various supplementation studies, correlations have been noted between the doses of iron given and/or the chemical formula used and side effects. The most common side effects associated with iron supplementation affect the gastrointestinal tract, manifesting as constipation, nausea, diarrhea, and vomiting [1, 3, 36, 38].
Coplin et al. compared the tolerability of ferrous sulfate and iron bis-glycinate chelate in a study. They found that both ferrous sulfate and iron bis-glycinate chelate had equivalent therapeutic efficiency at a similar dosage (50 mg), but the side effects were greater (37% vs. 21%) in the group that received ferrous sulfate . Gastrointestinal complaints and symptoms as constipation, nausea, vomiting, and diarrhea have been observed commonly in women consuming high doses of supplemental iron. The frequency and intensity of these complains are related to the amount of elemental iron released in the stomach [1, 39]. In malaria areas, additional iron may exacerbate malaria infection. For theses reasons, iron supplementation programs should be supervised and planned according to target the population .
In a systematic review, Low et al. evaluated the benefits and safety of daily iron supplementation in school-aged children. There was no difference in the frequency of children with gastrointestinal upset, constipation, vomiting, or diarrhea between children who received iron supplementation and children in the placebo group. The authors concluded that iron supplements appeared to be well tolerated, but the safety dates were limited: only 6 of 32 studies reported safety outcomes . In our study, there were few gastrointestinal side effects with either treatment, so there was no need to suspend supplementation. Moreover, there were no differences in the observation of gastrointestinal side effects between the two supplementation groups. This lack of significant side effects may have been due to the low dose of iron given (30 mg/d).
We recognize some caveats of our study. In terms of design, a more comprehensive description of iron status in the body would include other indicators of iron status, such as serum transferrin, serum transferring receptors, and total serum iron. Likewise, indicators of infection as C reactive protein and α1-glycoprotein have been evaluated in some studies on iron supplementation. However, serum ferritin has been considered a good indicator of iron storage and is useful in evaluating interventions response at a population level .
While serum concentrations of this protein can be increased during inflammatory and infectious processes as well as in response to trauma, children participating in our study received anti-helminthic prophylaxis at the beginning of the study, and the blood sample was obtained only if the children did not have a cold, fever, or any other common symptom of infection, so we consider that the use of serum ferritin in this study was a good indicator of iron storage as well as a good indicator of intervention response at the population level, as has been suggested . In terms of implementation, the main constraint of our study was related to the high attrition rate to effect evaluation at 6 months after supplementation. However, the only difference between participants who completed the study and the ones missing data during follow-up was age, and the results of our multivariate analyses were adjusted by age as soon as each evaluation of iron status was done. Therefore, we do not think that this attrition reflected a selection bias that could substantially bias our results.