Use of Compartmental Modeling and Retinol Isotope Dilution to Determine Vitamin A Stores in Young People with Sickle Cell Disease Before and After Vitamin A Supplementation.

BACKGROUND: Suboptimal plasma retinol concentrations have been documented in US children with sickle cell disease (SCD) hemoglobin SS type (SCD-HbSS), but little is known about vitamin A kinetics and stores in SCD. OBJECTIVES: The objectives were to quantify vitamin A total body stores (TBS) and whole-body retinol kinetics in young people with SCD-HbSS and use retinol isotope dilution (RID) to predict TBS in SCD-HbSS and healthy peers as well as after vitamin A supplementation in SCD-HbSS subjects. METHODS: Composite plasma [13C10]retinol response data collected from 22 subjects with SCD-HbSS for 28 d after isotope ingestion were analyzed using population-based compartmental modeling ("super-subject" approach); TBS and retinol kinetics were quantified for the group. TBS was also calculated for the same individuals using RID, as well as for healthy peers (n = 20) and for the subjects with SCD-HbSS after 8 wk of daily vitamin A supplements (3.15 or 6.29 µmol retinol/d [900 or 1800 µg retinol activity equivalents/d]). RESULTS: Model-predicted group mean TBS for subjects with SCD-HbSS was 428 µmol, equivalent to ∼11 mo of stored vitamin A; vitamin A disposal rate was 1.3 µmol/d. Model-predicted TBS was similar to that predicted by RID at 3 d postdosing (mean, 389 µmol; ∼0.3 µmol/g liver); TBS predictions at 3 compared with 28 d were not significantly different. Mean TBS in healthy peers was similar (406 µmol). RID-predicted TBS for subjects with SCD-HbSS was not significantly affected by vitamin A supplementation at either dose. CONCLUSIONS: Despite differences in plasma retinol concentrations, TBS was the same in subjects with SCD-HbSS compared with healthy peers. Because 56 d of vitamin A supplementation at levels 1.2 to 2.6 times the Recommended Dietary Allowance did not increase TBS in these subjects with SCD-HbSS, further work will be needed to understand the effects of SCD on retinol metabolism. This trial was registered as NCT03632876 at clinicaltrials.gov.

Comparing two simplified questionnaire-based methods with 24-h recalls for estimating fortifiable wheat flour and oil consumption in Mandaluyong City, Philippines.

Information on fortifiable food consumption is essential to design, monitor and evaluate fortification programmes, yet detailed methods like 24-h recalls (24HRs) that provide such data are rarely conducted. Simplified questionnaire-based methods exist but their validity compared with 24HRs has not been shown. We compared two simplified methods (i.e., a household food acquisition and purchase questionnaire [FAPQ] and a 7-day semiquantitative food frequency questionnaire [SQ-FFQ]) against 24HRs for estimating fortifiable food consumption. We assessed the consumption of fortifiable wheat flour and oil using a FAPQ and, for wheat flour only, a 7-day SQ-FFQ and compared the results against 24HRs. The participants included children 12-18 months (n = 123) and their mothers 18-49 years selected for a study assessing child vitamin A intake and status in Mandaluyong City, Philippines. For fortifiable wheat flour, the FAPQ estimated considerably lower mean intakes compared to 24HRs for children and mothers (2.2 vs. 14.1 g/day and 5.1 vs. 42.3 g/day, respectively), while the SQ-FFQ estimated slightly higher mean intakes (15.7 vs. 14.1 g/day and 51.5 vs. 42.3 g/day, respectively). For fortifiable oil, the FAPQ estimated considerably higher mean intakes compared to 24HRs for children and mothers (4.6 vs. 1.8 g/day and 12.5 vs. 6.1 g/day, respectively). The SQ-FFQ, but not the FAPQ, generated useful information on fortifiable food consumption that can inform fortification programme design and monitoring decisions in the absence of more detailed individual-level data. Potential adaptations to improve the FAPQ, such as additional questions on foods prepared away from home and usage patterns, merit further research.

Determination of Vitamin A Total Body Stores in Children from Dried Serum Spots: Application in a Low- and Middle-Income Country Community Setting.

BACKGROUND: The retinol isotope dilution (RID) method has been used to evaluate vitamin A (VA) status in healthy adults and children in low- and middle-income countries (LMIC) and to assess the efficacy of various VA interventions. OBJECTIVE: The study was designed to examine whether dried serum spots (DSS) can be applied to RID when conducting VA total body store (TBS) assessments in community settings. METHODS: Four days after an oral dose of 0.4 mg [13C10]retinyl acetate was administered to Filipino children (12-18 mo), a single blood draw was divided to isolate both serum and plasma. Serum (40 µL) was spotted and dried on Whatman 903 cards and shipped at ambient temperature whereas liquid plasma (LP) was frozen at -80°C and shipped on dry ice. The VA tracer to tracee ratio from DSS and LP was quantified by LC-MS/MS. Comparisons between DSS and LP paired samples (n = 72) were made for [13C10]retinol specific activity (SAp) by Pearson's correlation and for VA TBS by Bland-Altman analysis. RESULTS: The sum of 3 coextracted DSS were required to consistently detect [13C10]retinol above the LC-MS/MS limit of quantitation (LOQ). [13C10]retinol SAp from DSS was highly correlated with SAp from LP (r = 0.945; P < 0.01). A comparison of methods for TBS determination using Bland-Altman analysis indicated agreement with an intraindividual difference of 24.7 µmol (4.6%). Mean total liver reserve (TLR) values from DSS and LP were 1.7 µmol/g (± 0.6 SD) and 1.6 µmol/g (± 0.6 SD), respectively. CONCLUSIONS: VA TBS can be determined from DSS thereby reducing the logistics and cost of maintaining a cold chain by shipping samples at ambient temperature and, thus, making the RID technique more feasible in LMIC community settings. This trial was registered at https://clinicaltrials.gov as NCT03030339.

Reduced plasma carotenoids in individuals suffering from metabolic diseases with disturbances in lipid metabolism: a systematic review and meta-analysis of observational studies.

This review summarises the association between serum carotenoids, serum retinoids and dietary intake outcomes with obesity/overweight and individuals with metabolic diseases with disturbances in lipid metabolism. Observational studies reporting dietary intakes and serum concentrations of carotenoids and retinol were collected from Medline and Web of Science. Mean differences were calculated between "cases" (classified as obese, overweight or having a metabolic disease with disturbances in lipid metabolism; i.e. non-alcoholic fatty liver disease, type 2 diabetes, dyslipidaemia or metabolic syndrome) and "comparator group" (classified as normal weight healthy individuals) and summarised in meta-analyses. Significant summary measures were observed for most serum provitamin A and non-provitamin A carotenoids. Studies reporting total serum carotenoids had shown the greatest decrease (-0.28 µmol/l [-0.33, -0.23], p<.001, I2=62.5%, n = 7). There were no significant summary measures for dietary outcomes, suggesting a physiological role of low serum carotenoids in the development of obesity and associated diseases.

Use of Model-Based Compartmental Analysis and a Super-Child Design to Study Whole-Body Retinol Kinetics and Vitamin A Total Body Stores in Children from 3 Lower-Income Countries.

BACKGROUND: Model-based compartmental analysis has been used to describe and quantify whole-body vitamin A metabolism and estimate total body stores (TBS) in animals and humans. OBJECTIVES: We applied compartmental modeling and a super-child design to estimate retinol kinetic parameters and TBS for young children in Bangladesh, Guatemala, and the Philippines. METHODS: Children ingested [13C10]retinyl acetate and 1 or 2 blood samples were collected from each child from 6 h to 28 d after dosing. Temporal data for fraction of dose in plasma [13C10]retinol were modeled using WinSAAM software and a 6-component model with vitamin A intake included as weighted data. RESULTS: Model-predicted TBS was 198, 533, and 1062 µmol for the Bangladeshi (age, 9-17 mo), Filipino (12-18 mo), and Guatemalan children (35-65 mo). Retinol kinetics were similar for Filipino and Guatemalan groups and generally faster for Bangladeshi children, although fractional transfer of plasma retinol to a larger exchangeable storage pool was the same for the 3 groups. Recycling to plasma from that pool was ∼2.5 times faster in the Bangladeshi children compared with the other groups and the recycling number was 2-3 times greater. Differences in kinetics between groups are likely related to differences in vitamin A stores and intakes (geometric means: 352, 727, and 764 µg retinol activity equivalents/d for the Bangladeshi, Filipino, and Guatemalan children, respectively). CONCLUSIONS: By collecting 1 or 2 blood samples from each child to generate a composite plasma tracer data set with a minimum of 5 children/time, group TBS and retinol kinetics can be estimated in children by compartmental analysis; inclusion of vitamin A intake data increases confidence in model predictions. The super-child modeling approach is an effective technique for comparing vitamin A status among children from different populations. These trials were registered at www.clinicaltrials.gov as NCT03000543 (Bangladesh), NCT03345147 (Guatemala), and NCT03030339 (Philippines).

Intestinal ß-carotene bioconversion in humans is determined by a new single-sample, plasma isotope ratio method and compared with traditional and modified area-under-the-curve methods.

The vitamin A value (bioefficacy) of provitamin A carotenoids is determined by absorption of the carotenoid (bioavailability) and its subsequent conversion to retinol (bioconversion). Here we show that intestinal bioconversion of ß-carotene can be estimated based on analysis of a single plasma sample collected 6 h after subjects ingest a test dose of stable isotope-labeled ß-carotene from the ratio of retinyl esters to retinyl esters plus ß-carotene. Plasma isotope ratio predictions of bioconversion ranged from 50 to- 93% (mean 76%) for 45 healthy young adults with low vitamin A stores. Results were the same as predictions made by a traditional area-under-the-curve method calculated from 0 to- 8 h or a modified area-under-the-curve method calculated from 0 to- 12 h. The modified method may provide better estimates of bioconversion between 8 and 24 h after ingestion of a carotenoid dose when stable isotopes cannot be used due to cost or logistics. Furthermore, because the plasma isotope ratio method requires only one blood sample and no isolation of triglyceride-rich lipoproteins, its use will facilitate estimation of provitamin A carotenoid bioconversion in human subjects and especially children, in whom repeated blood sampling is not feasible.

Association of oily fish intake, sex, age, BMI and APOE genotype with plasma long-chain n-3 fatty acid composition.

n-3 Fatty acids are associated with better cardiovascular and cognitive health. However, the concentration of EPA, DPA and DHA in different plasma lipid pools differs and factors influencing this heterogeneity are poorly understood. Our aim was to evaluate the association of oily fish intake, sex, age, BMI and APOE genotype with concentrations of EPA, DPA and DHA in plasma phosphatidylcholine (PC), NEFA, cholesteryl esters (CE) and TAG. Healthy adults (148 male, 158 female, age 20-71 years) were recruited according to APOE genotype, sex and age. The fatty acid composition was determined by GC. Oily fish intake was positively associated with EPA in PC, CE and TAG, DPA in TAG, and DHA in all fractions (P≤0·008). There was a positive association between age and EPA in PC, CE and TAG, DPA in NEFA and CE, and DHA in PC and CE (P≤0·034). DPA was higher in TAG in males than females (P<0·001). There was a positive association between BMI and DPA and DHA in TAG (P<0·006 and 0·02, respectively). APOE genotype×sex interactions were observed: the APOE4 allele associated with higher EPA in males (P=0·002), and there was also evidence for higher DPA and DHA (P≤0·032). In conclusion, EPA, DPA and DHA in plasma lipids are associated with oily fish intake, sex, age, BMI and APOE genotype. Such insights may be used to better understand the link between plasma fatty acid profiles and dietary exposure and may influence intake recommendations across population subgroups.

A Simple Plasma Retinol Isotope Ratio Method for Estimating ß-Carotene Relative Bioefficacy in Humans: Validation with the Use of Model-Based Compartmental Analysis.

Background: Provitamin A carotenoids are an important source of dietary vitamin A for many populations. Thus, accurate and simple methods for estimating carotenoid bioefficacy are needed to evaluate the vitamin A value of test solutions and plant sources. ß-Carotene bioefficacy is often estimated from the ratio of the areas under plasma isotope response curves after subjects ingest labeled ß-carotene and a labeled retinyl acetate reference dose [isotope reference method (IRM)], but to our knowledge, the method has not yet been evaluated for accuracy.Objectives: Our objectives were to develop and test a physiologically based compartmental model that includes both absorptive and postabsorptive ß-carotene bioconversion and to use the model to evaluate the accuracy of the IRM and a simple plasma retinol isotope ratio [(RIR), labeled ß-carotene-derived retinol/labeled reference-dose-derived retinol in one plasma sample] for estimating relative bioefficacy.Methods: We used model-based compartmental analysis (Simulation, Analysis and Modeling software) to develop and apply a model that provided known values for ß-carotene bioefficacy. Theoretical data for 10 subjects were generated by the model and used to determine bioefficacy by RIR and IRM; predictions were compared with known values. We also applied RIR and IRM to previously published data.Results: Plasma RIR accurately predicted ß-carotene relative bioefficacy at 14 d or later. IRM also accurately predicted bioefficacy by 14 d, except that, when there was substantial postabsorptive bioconversion, IRM underestimated bioefficacy. Based on our model, 1-d predictions of relative bioefficacy include absorptive plus a portion of early postabsorptive conversion.Conclusion: The plasma RIR is a simple tracer method that accurately predicts ß-carotene relative bioefficacy based on analysis of one blood sample obtained at ≥14 d after co-ingestion of labeled ß-carotene and retinyl acetate. The method also provides information about the contributions of absorptive and postabsorptive conversion to total bioefficacy if an additional sample is taken at 1 d.

A Retinol Isotope Dilution Equation Predicts Both Group and Individual Total Body Vitamin A Stores in Adults Based on Data from an Early Postdosing Blood Sample.

BACKGROUND: Retinol isotope dilution (RID) is used to determine vitamin A total body stores (TBS) after an oral dose of a vitamin A stable isotope. The generally accepted prediction equation proposed by Olson's group in 1989 (Furr et al. Am J Clin Nutr 1989;49:713-6) includes factors related to dose absorption and retention, isotope equilibration in plasma compared with stores, catabolism during the mixing period, and the optimal time for measuring plasma isotope enrichment. OBJECTIVES: The objectives were 1) to develop a modified RID equation and identify an earlier sampling time for predicting TBS and 2) to improve prediction in individuals as well as groups. METHODS: To develop a modified RID equation, we used results of model-based compartmental analysis [the Simulation, Analysis and Modeling software (WinSAAM version 3.0.8; http://www.WinSAAM.org)] of plasma [13C10]retinol kinetic data from 32 previously studied, healthy young adults of European ancestry who had moderate vitamin A intakes and who ingested 2.95 µmol [13C10]retinyl acetate. RESULTS: We examined the time dependence of factors in the prediction equation related to absorption/retention (Fa) and isotope equilibration (S) and determined that 4 or 5 d postdosing was the optimal sampling time. TBS calculated by the equation TBS = Fa x S x (1/SAp), where SAp is plasma retinol specific activity (fraction of dose/µmol), were highly correlated with model-predicted TBS (r = 0.95 and 0.96 for 4 and 5 d, respectively; P < 0.001); predictions for individuals were also highly correlated (Rs = 0.94 and 0.94; P < 0.001). CONCLUSION: The equation TBS ≈ 0.5 × (1/SAp) accurately predicted vitamin A TBS in this group of 32 healthy young adults and its individual members with the use of data from 1 blood sample taken 4 d after isotope administration.

Biomarkers of Nutrition for Development (BOND)-Vitamin A Review.

The Biomarkers of Nutrition for Development (BOND) project is designed to provide evidence-informed advice to anyone with an interest in the role of nutrition in health. The BOND program provides information with regard to selection, use, and interpretation of biomarkers of nutrient exposure, status, function, and effect, which will be especially useful for readers who want to assess nutrient status. To accomplish this objective, expert panels are recruited to evaluate the literature and to draft comprehensive reports on the current state of the art with regard to specific nutrient biology and available biomarkers for assessing nutritional status at the individual and population levels. Phase I of the BOND project includes the evaluation of biomarkers for 6 nutrients: iodine, folate, zinc, iron, vitamin A, and vitamin B-12. This review of vitamin A is the current article in this series. Although the vitamin was discovered >100 y ago, vitamin A status assessment is not trivial. Serum retinol concentrations are under homeostatic control due in part to vitamin A's use in the body for growth and cellular differentiation and because of its toxic properties at high concentrations. Furthermore, serum retinol concentrations are depressed during infection and inflammation because retinol-binding protein (RBP) is a negative acute-phase reactant, which makes status assessment challenging. Thus, this review describes the clinical and functional indicators related to eye health and biochemical biomarkers of vitamin A status (i.e., serum retinol, RBP, breast-milk retinol, dose-response tests, isotope dilution methodology, and serum retinyl esters). These biomarkers are then related to liver vitamin A concentrations, which are usually considered the gold standard for vitamin A status. With regard to biomarkers, future research questions and gaps in our current understanding as well as limitations of the methods are described.

Plasma Retinol Kinetics and ß-Carotene Bioefficacy Are Quantified by Model-Based Compartmental Analysis in Healthy Young Adults with Low Vitamin A Stores.

BACKGROUND: Model-based compartmental analysis of data on plasma retinol kinetics after administration of labeled retinol provides unique information about whole-body vitamin A metabolism. If labeled ß-carotene is coadministered, its bioefficacy relative to the retinol reference dose can also be estimated. OBJECTIVES: The objectives were to model plasma retinol kinetics after administration of labeled preformed vitamin A and provitamin A ß-carotene and to determine relative ß-carotene bioefficacy. METHODS: We used the Simulation, Analysis and Modeling software (WinSAAM version 3.0.8; http://www.WinSAAM.org) to analyze previously collected data on plasma [13C10]- and [13C5]retinol kinetics for 14 d after oral administration of 1 mg [13C10]retinyl acetate and 2 mg [13C10]ß-carotene in oil to 30 healthy young adults of European ancestry [13 men, 17 women; mean ± SD age: 24.5 ± 4.2 y; mean ± SD body weight: 65.2 ± 10 kg; mean ± SD body mass index (in kg/m2): 22.5 ± 1.9] with moderate vitamin A intakes. RESULTS: A 6-component model provided the best fit to the data, including compartments for initial metabolism of vitamin A, plasma retinol, and extravascular vitamin A storage. The disposal rate was 6.7 ± 3.1 µmol/d, fractional catabolic rate was 6.0% ± 2.3%/d, and vitamin A stores were 123 ± 71 µmol. Relative ß-carotene bioefficacy, based on the ratio of the areas under the fraction of dose curves calculated by WinSAAM, averaged 13.5% ± 6.02% (retinol activity equivalents = 7.7:1.0 µg). Interindividual variation in relative ß-carotene bioefficacy was high (CV: 44%). CONCLUSIONS: Vitamin A kinetics in these young adults were best described by essentially the same model that had been previously developed by using data for older adults with higher vitamin A stores; differences in parameter values reflected differences in vitamin A status. Estimated ß-carotene bioefficacy was relatively low but similar to previously reported estimates obtained by graphical methods. This trial was registered at the UK Clinical Research Network as UKCRN 7413.

Consumption of Fish Oil Providing Amounts of Eicosapentaenoic Acid and Docosahexaenoic Acid That Can Be Obtained from the Diet Reduces Blood Pressure in Adults with Systolic Hypertension: A Retrospective Analysis.

BACKGROUND: Although many randomized controlled trials (RCTs) have examined the effects of the n-3 (ω-3) fatty acids eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3) on blood pressure (BP) and vascular function, the majority have used doses of EPA+DHA of >3 g/d, which are unlikely to be achieved by dietary manipulation. OBJECTIVE: The objective was to examine, by using a retrospective analysis from a multicenter RCT, the impact of recommended EPA+DHA intakes achievable through diet on systolic and diastolic BPs and microvascular function in adults in the United Kingdom. METHODS: In a double-blind, placebo-controlled RCT, healthy men and women (n = 312) consumed a control oil or fish oil (FO) providing 0.7 or 1.8 g EPA+DHA/d, in random order, each for 8 wk. Fasting BP and microvascular function (using laser Doppler iontophoresis) were assessed and plasma collected for the quantification of markers of vascular function. Participants were retrospectively genotyped for the endothelial nitric oxide synthase (eNOS) rs1799983 variant. RESULTS: No effects of n-3 fatty acid treatment or any treatment × eNOS genotype interactions were evident in the group as a whole for any of the clinical or biochemical outcomes. Assessment of response according to hypertension status at baseline indicated a significant (P = 0.046) FO-induced reduction (mean: 5 mm Hg) in systolic BP, specifically in those with isolated systolic hypertension (n = 31). No dose response was observed. CONCLUSIONS: These findings indicate that in adults with isolated systolic hypertension, daily doses of EPA+DHA as low as 0.7 g show clinically meaningful BP reductions, which, at a population level, could be associated with lower cardiovascular disease risk. Confirmation of findings in an RCT in which participants are prospectively recruited on the basis of BP status is required to draw definite conclusions.

Maximizing the benefits and minimizing the risks of intervention programs to address micronutrient malnutrition: symposium report.

Interventions to address micronutrient deficiencies have large potential to reduce the related disease and economic burden. However, the potential risks of excessive micronutrient intakes are often not well determined. During the Global Summit on Food Fortification, 9-11 September 2015, in Arusha, a symposium was organized on micronutrient risk-benefit assessments. Using case studies on folic acid, iodine and vitamin A, the presenters discussed how to maximize the benefits and minimize the risks of intervention programs to address micronutrient malnutrition. Pre-implementation assessment of dietary intake, and/or biomarkers of micronutrient exposure, status and morbidity/mortality is critical in identifying the population segments at risk of inadequate and excessive intake. Dietary intake models allow to predict the effect of micronutrient interventions and their combinations, e.g. fortified food and supplements, on the proportion of the population with intakes below adequate and above safe thresholds. Continuous monitoring of micronutrient intake and biomarkers is critical to identify whether the target population is actually reached, whether subgroups receive excessive amounts, and inform program adjustments. However, the relation between regular high intake and adverse health consequences is neither well understood for many micronutrients, nor do biomarkers exist that can detect them. More accurate and reliable biomarkers predictive of micronutrient exposure, status and function are needed to ensure effective and safe intake ranges for vulnerable population groups such as young children and pregnant women. Modelling tools that integrate information on program coverage, dietary intake distribution and biomarkers will further enable program makers to design effective, efficient and safe programs.

An LC/MS/MS method for stable isotope dilution studies of ß-carotene bioavailability, bioconversion, and vitamin A status in humans.

Isotope dilution is currently the most accurate technique in humans to determine vitamin A status and bioavailability/bioconversion of provitamin A carotenoids such as ß-carotene. However, limits of MS detection, coupled with extensive isolation procedures, have hindered investigations of physiologically-relevant doses of stable isotopes in large intervention trials. Here, a sensitive liquid chromatography-tandem mass spectrometry (LC/MS/MS) analytical method was developed to study the plasma response from coadministered oral doses of 2 mg [(13)C10]ß-carotene and 1 mg [(13)C10]retinyl acetate in human subjects over a 2 week period. A reverse phase C18 column and binary mobile phase solvent system separated ß-carotene, retinol, retinyl acetate, retinyl linoleate, retinyl palmitate/retinyl oleate, and retinyl stearate within a 7 min run time. Selected reaction monitoring of analytes was performed under atmospheric pressure chemical ionization in positive mode at m/z 537→321 and m/z 269→93 for respective [(12)C]ß-carotene and [(12)C] retinoids; m/z 547→330 and m/z 274→98 for [(13)C10]ß-carotene and [(13)C5] cleavage products; and m/z 279→100 for metabolites of [(13)C10]retinyl acetate. A single one-phase solvent extraction, with no saponification or purification steps, left retinyl esters intact for determination of intestinally-derived retinol in chylomicrons versus retinol from the liver bound to retinol binding protein. Coadministration of [(13)C10]retinyl acetate with [(13)C10]ß-carotene not only acts as a reference dose for inter-individual variations in absorption and chylomicron clearance rates, but also allows for simultaneous determination of an individual's vitamin A status.

Effect of multiple micronutrient-fortified bouillon on micronutrient status among women and children in the Northern Region of Ghana: Protocol for the Condiment Micronutrient Innovation Trial (CoMIT), a community-based randomized controlled trial.

INTRODUCTION: Micronutrient deficiencies are prevalent in West Africa, particularly among women of reproductive age (WRA) and young children. Bouillon is a promising food fortification vehicle due to its widespread consumption. This study aims to evaluate the impact of multiple micronutrient-fortified bouillon cubes, compared to control bouillon cubes (fortified with iodine only), on micronutrient status and hemoglobin concentrations among lactating and non-lactating WRA and young children in northern Ghana. METHODS: This randomized, controlled doubly-masked trial will be conducted in the Kumbungu and Tolon districts in the Northern Region of Ghana, where prior data indicate multiple micronutrient deficiencies are common. Participants will be: 1) non-pregnant non-lactating WRA (15-49 y), 2) children 2-5 y, and 3) non-pregnant lactating women 4-18 months postpartum. Eligible participants will be randomly assigned to receive household rations of one of two types of bouillon cubes: 1) a multiple micronutrient-fortified bouillon cube containing vitamin A, folic acid, vitamin B12, iron, zinc, and iodine, or 2) a control cube containing iodine only. Each participant's household will receive a ration of bouillon cubes every 2 weeks, and households will be advised to prepare meals as usual, using the study-provided cubes. The trial duration will be 9 months for non-pregnant non-lactating WRA and children, and 3 months for lactating women. The primary outcomes will be changes in biomarkers of micronutrient status and hemoglobin among WRA and children and milk micronutrient concentrations among lactating women. Secondary outcomes will include change in prevalence of micronutrient deficiency and anemia; dietary intake of bouillon and micronutrients; inflammation, malaria, and morbidity symptoms; and child growth and development. DISCUSSION: Evidence from this study will inform discussions about bouillon fortification in Ghana and West Africa. TRIAL REGISTRATION: The trial was registered on ClinicalTrials.gov (NCT05178407) and the Pan-African Clinical Trial Registry (PACTR202206868437931). This manuscript reflects protocol version 4 (August 29, 2022).

Scientific opinion on the tolerable upper intake level for preformed vitamin A and ß-carotene.

Following two requests from the European Commission, the EFSA Panel on Nutrition, Novel Foods and Food Allergens (NDA) was asked to deliver a scientific opinion on the revision of the tolerable upper intake level (UL) for preformed vitamin A and ß-carotene. Systematic reviews of the literature were conducted for priority adverse health effects of excess vitamin A intake, namely teratogenicity, hepatotoxicity and endpoints related to bone health. Available data did not allow to address whether ß-carotene could potentiate preformed vitamin A toxicity. Teratogenicity was selected as the critical effect on which to base the UL for preformed vitamin A. The Panel proposes to retain the UL for preformed vitamin A of 3000 µg RE/day for adults. This UL applies to men and women, including women of child-bearing age, pregnant and lactating women and post-menopausal women. This value was scaled down to other population groups using allometric scaling (body weight0.75), leading to ULs between 600 µg RE/day (infants 4-11 months) and 2600 µg RE/day (adolescents 15-17 years). Based on available intake data, European populations are unlikely to exceed the UL for preformed vitamin A if consumption of liver, offal and products thereof is limited to once per month or less. Women who are planning to become pregnant or who are pregnant are advised not to consume liver products. Lung cancer risk was selected as the critical effect of excess supplemental ß-carotene. The available data were not sufficient and suitable to characterise a dose-response relationship and identify a reference point; therefore, no UL could be established. There is no indication that ß-carotene intake from the background diet is associated with adverse health effects. Smokers should avoid consuming food supplements containing ß-carotene. The use of supplemental ß-carotene by the general population should be limited to the purpose of meeting vitamin A requirements.

CD36 and SR-BI are involved in cellular uptake of provitamin A carotenoids by Caco-2 and HEK cells, and some of their genetic variants are associated with plasma concentrations of these micronutrients in humans.

Scavenger receptor class B type I (SR-BI) and cluster determinant 36 (CD36) have been involved in cellular uptake of some provitamin A carotenoids. However, data are incomplete (e.g., there are no data on α-carotene), and it is not known whether genetic variants in their encoding genes can affect provitamin A carotenoid status. The objectives were 1) to assess the involvement of these scavenger receptors in cellular uptake of the main provitamin A carotenoids (i.e., ß-carotene, α-carotene, and ß-cryptoxanthin) as well as that of preformed vitamin A (i.e., retinol) and 2) to investigate the contribution of genetic variations in genes encoding these proteins to interindividual variations in plasma concentrations of provitamin A carotenoids. The involvement of SR-BI and CD36 in carotenoids and retinol cellular uptake was investigated in Caco-2 and human embryonic kidney (HEK) cell lines. The involvement of scavenger receptor class B type I (SCARB1) and CD36 genetic variants on plasma concentrations of provitamin A carotenoids was assessed by association studies in 3 independent populations. Cell experiments suggested the involvement of both proteins in cellular uptake of provitamin A carotenoids but not in that of retinol. Association studies showed that several plasma provitamin A carotenoid concentrations were significantly different (P < 0.0083) between participants who bore different genotypes at single nucleotide polymorphisms and haplotypes in CD36 and SCARB1. In conclusion, SR-BI and CD36 are involved in cellular uptake of provitamin A carotenoids, and genetic variations in their encoding genes may modulate plasma concentrations of provitamin A carotenoids at a population level.

The impact of common gene variants on the response of biomarkers of cardiovascular disease (CVD) risk to increased fish oil fatty acids intakes.

The cardioprotective actions of the fish oil (FO)-derived long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFAs) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been demonstrated, and dose-response relationships have been defined. However, there is a substantial and well-recognized within-population heterogeneity in response to FO, the etiology of which is poorly understood. Genetic variation may influence responsiveness. Here we review the available literature relating to gene variants shown to influence tissue LC n-3 PUFA status and response to FO intervention. From this review we conclude that the available evidence is relatively limited. A number of individual genotype × LC-n3 PUFA × phenotype associations have been described, but few have been investigated in subsequent cohorts or confirmed in independent studies. In the context of a more stratified approach to the provision of dietary advice, there is a need for further research to refine current dietary EPA and DHA recommendations.

Single nucleotide polymorphisms upstream from the ß-carotene 15,15'-monoxygenase gene influence provitamin A conversion efficiency in female volunteers.

ß-Carotene, the most abundant provitamin A carotenoid in the diet, is converted to retinal by ß-carotene 15,15'-monoxygenase (BCMO1). However, ß-carotene absorption and conversion into retinal is extremely variable among individuals, with proportions of low responders to dietary ß-carotene as high as 45%. Recently, 2 common nonsynonymous single nucleotide polymorphisms (SNPs) within the BCMO1 coding region (R267S; rs12934922 and A379V; rs7501331) revealed reduced catalytic activity, confirming that genetic variations contribute to the low responder phenotype. Because 4 SNPs 5' upstream from the BCMO1 gene were recently shown to affect circulating carotenoid concentrations, the current study aimed to investigate the effects of these SNPs on ß-carotene conversion efficiency. Three of the 4 polymorphisms (rs6420424, rs11645428, and rs6564851) reduced the catalytic activity of BCMO1 in female volunteers by 59, 51, and 48%, respectively. The TG-rich lipoprotein fraction retinyl palmitate:ß-carotene ratio was negatively correlated with the G allele of rs11645428 (r = -0.44; P = 0.018), whereas it was positively correlated with the G allele of rs6420424 (r = 0.53; P = 0.004) and the T allele of rs6564851 (r = 0.41; P = 0.028). Furthermore, large inter-ethnic variations in frequency of affected alleles were detected, with frequencies varying from 43 to 84% (rs6420424), 52 to 100% (rs11645428), and 19 to 67% (rs6564851). In summary, a range of SNPs can influence the effectiveness of using plant-based provitamin A carotenoids to increase vitamin A status in at-risk population groups and this effect may vary depending on ethnic origin.