A rapid, simple questionnaire to assess gastrointestinal symptoms after oral ferrous sulphate supplementation

BACKGROUND: Oral iron supplementation is often associated with rapid onset of gastrointestinal side-effects. The aim of this study was to develop and trial a short, simple questionnaire to capture these early side-effects and to determine which symptoms are more discriminating. METHODS: The study was a double-blind placebo-controlled randomized parallel trial with one week treatment followed by one week wash-out. Subjects were randomized into two treatment groups (n = 10/group) to receive either ferrous sulphate (200 mg capsules containing 65 mg of iron) or placebo, both to be taken at mealtimes twice daily during the treatment period. Subjects completed the questionnaires daily for 14 days. The questionnaire included gastrointestinal symptoms commonly reported to be associated with the oral intake of ferrous iron salts (i.e. nausea, vomiting, heartburn, abdominal pain, diarrhoea, and constipation). RESULTS: Seventy five per cent of participants reporting the presence of one or more symptoms in the first week of the study were in the ferrous sulphate group. In the second week of the study (i.e. wash-out), 67% of the participants reporting one or more symptom(s) were in the ferrous sulphate group. In the first week of the study (treatment) the number of symptoms reported by participants in the ferrous sulphate group (mean ± SEM = 6.7 ± 1.7) was significantly higher than that for participants in the placebo group (1.2 ± 0.5) (p = 0.01). In the second week of the study (wash-out) the number of symptoms reported by participants in the ferrous sulphate group (4.6 ± 2.0) appeared higher than for participants in the placebo group (1.0 ± 0.7) although this did not reach significance (p = 0.12). Events for which the gastrointestinal symptom questionnaire was most discriminatory between ferrous sulphate and placebo groups were: heartburn, abdominal pain and the presence of black stools (all p ≤ 0.03). CONCLUSIONS: A tool for the detection of commonly-occurring side effects should not require large study numbers to be effective. With just 10 subjects per group (iron or placebo), this simple questionnaire measures gastrointestinal side-effects associated with oral iron (ferrous sulphate) supplementation, and would be appropriate for use in intervention studies or clinical trials. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02146053 (21/05/2014)

Development of the Caco-2 Model for Assessment of Iron Absorption and Utilisation at Supplemental Levels

Caco-2 cells may be typically used as a first step to investigate the bioavailability of different dietary and fortificant forms of iron (Fe) at low levels (< 10 µM) in tissue culture medium (TCM). Whether this model is suitable with supplemental levels of Fe (ca. 200 µM in TCM) is not clear and neither, therefore, is the choice of reference iron compound under those conditions as a ‘positive control’. Here we show that with 200 ?M iron in TCM (serum-free MEM), Fe(II) sulphate precipitates and while high levels of ascorbic acid can prevent this, it is to the detriment of the Caco-2 cell monolayer and/or it adversely affects the pH of the TCM. Adjusting the pH of TCM to account for this issue again leads to Fe precipitation, which is detectable as both a true precipitate (~ 50%) and a nano-precipitate in suspension (~20%). In contrast, Fe(III) maltol which, clinically, appears less toxic to the intestinal mucosa than Fe(II) sulphate, retains solubility at supplemental levels in cell culture medium, without adversely affecting pH or the Caco-2 cell monolayer. Moreover, the iron is also well utilized by the cells as assessed through ferritin formation. Thus Caco-2 cells may also provide a model for screening iron uptake and utilisation at supplemental levels through the cellular generation of ferritin although care must be taken in ensuring (i) appropriate TCM conditions (e.g. pH and chemical speciation of the iron) (ii) monolayer integrity (i.e. the assay response is not an artefact of toxicity) and (iii) that an appropriate reference material is used (e.g. Fe:maltol at 1:5 ratio)

Development of the Caco-2 Model for Assessment of Iron Absorption and Utilisation at Supplemental Levels

Caco-2 cells may be typically used as a first step to investigate the bioavailability of different dietary and fortificant forms of iron (Fe) at low levels (< 10 µM) in tissue culture medium (TCM). Whether this model is suitable with supplemental levels of Fe (ca. 200 µM in TCM) is not clear and neither, therefore, is the choice of reference iron compound under those conditions as a ‘positive control’. Here we show that with 200 ?M iron in TCM (serum-free MEM), Fe(II) sulphate precipitates and while high levels of ascorbic acid can prevent this, it is to the detriment of the Caco-2 cell monolayer and/or it adversely affects the pH of the TCM. Adjusting the pH of TCM to account for this issue again leads to Fe precipitation, which is detectable as both a true precipitate (~ 50%) and a nano-precipitate in suspension (~20%). In contrast, Fe(III) maltol which, clinically, appears less toxic to the intestinal mucosa than Fe(II) sulphate, retains solubility at supplemental levels in cell culture medium, without adversely affecting pH or the Caco-2 cell monolayer. Moreover, the iron is also well utilized by the cells as assessed through ferritin formation. Thus Caco-2 cells may also provide a model for screening iron uptake and utilisation at supplemental levels through the cellular generation of ferritin although care must be taken in ensuring (i) appropriate TCM conditions (e.g. pH and chemical speciation of the iron) (ii) monolayer integrity (i.e. the assay response is not an artefact of toxicity) and (iii) that an appropriate reference material is used (e.g. Fe:maltol at 1:5 ratio)

A nano-disperse ferritin-core mimetic that efficiently corrects anemia without luminal iron redox activity

AbstractThe 2-5nm Fe(III) oxo-hydroxide core of ferritin is less ordered and readily bioavailable compared to its pure synthetic analogue, ferrihydrite. We report the facile synthesis of tartrate-modified, nano-disperse ferrihydrite of small primary particle size, but with enlarged or strained lattice structure (~2.7Å for the main Bragg peak versus 2.6Å for synthetic ferrihydrite). Analysis indicated that co-precipitation conditions can be achieved for tartrate inclusion into the developing ferrihydrite particles, retarding both growth and crystallization and favoring stabilization of the cross-linked polymeric structure. In murine models, gastrointestinal uptake was independent of luminal Fe(III) reduction to Fe(II) and, yet, absorption was equivalent to that of ferrous sulphate, efficiently correcting the induced anemia. This process may model dietary Fe(III) absorption and potentially provide a side effect-free form of cheap supplemental iron.From the Clinical EditorSmall size tartrate-modified, nano-disperse ferrihydrite was used for efficient gastrointestinal delivery of soluble Fe(III) without the risk for free radical generation in murine models. This method may provide a potentially side effect-free form iron supplementation

Influence of simulated gastrointestinal conditions on particle-induced cytotoxicity and interleukin-8 regulation in differentiated and undifferentiated Caco-2 cells

Novel aspects of engineered nanoparticles offer many advantages for optimising food products and packaging. However, their potential hazards in the gastrointestinal tract require further investigation. We evaluated the toxic and inflammatory potential of two types of particles that might become increasingly relevant to the food industry, namely SiO2 and ZnO. The materials were characterised for their morphology, oxidant generation and hydrodynamic behaviour. Cytotoxicity and interleukin-8 mRNA and protein expression were evaluated in human intestinal Caco-2 cells. Particle pretreatment under simulated gastric and intestinal pH conditions resulted in reduced acellular ROS formation but did not influence cytotoxicity (WST-1 assay) or IL-8 expression. However, the differentiation status of the cells markedly determined the cytotoxic potency of the particles. Further research is needed to determine the in vivo relevance of our current observations regarding the role of particle aggregation and the stage of intestinal epithelial cell differentiation in determining the hazards of ingested particles