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BACKGROUND: Limited data are available on vitamin A kinetics and total body stores (TBS) in women. Such information can be obtained using compartmental modeling and retinol isotope dilution (RID). OBJECTIVES: Objectives were to apply population-based (super-subject) modeling to determine retinol kinetics in nonpregnant Ghanaian women of reproductive age and to use RID to predict TBS in the group and its individuals. METHODS: Women (n = 89) ingested a dose of [2H6]retinyl acetate and blood samples (3/woman) were collected from 6 h to 91 d, with all participants sampled at 14 d, about half at either 21 or 28 d, and each at one other time. Composite data (plasma retinol fraction of dose; FDp) were analyzed using Simulation, Analysis and Modeling software to obtain kinetic parameters, TBS, and other state variables as well as model-derived values for the RID composite coefficient FaS. The latter were used in the RID equation TBS (μmol) = FaS × 1/SAp (where SAp is plasma retinol specific activity) to predict TBS at various times. RESULTS: Model-predicted TBS was 973 μmol (n = 87). Geometric mean RID-predicted TBS was 965, 926, and 1006 μmol at 14, 21, and 28 d, respectively, with wide ranges [for example, 252-3848 μmol on day 14 (n = 86)]; TBS predictions were similar at later times. Participants had a mean 2 y of vitamin A in stores and estimated liver vitamin A concentrations in the normal range. Model-predicted vitamin A disposal rate was 1.3 μmol/d and plasma recycling number was 37. CONCLUSIONS: Super-subject modeling provides an estimate of group mean TBS as well as group-specific values for the RID coefficient FaS; the latter can be used to confidently predict TBS by RID for individual participants in the group under study or in similar individuals at 14 d or more after isotope ingestion. TRIAL REGISTRATION NUMBER: Trial is registered (NCT04632771) at https://clinicaltrials.gov.

Background: Limited data are available on vitamin A kinetics and total body stores (TBS) in women. Such information can be obtained using compartmental modeling and retinol isotope dilution (RID). Objectives: Objectives were to apply population-based (“super-subject”) modeling to determine retinol kinetics in nonpregnant Ghanaian women of reproductive age and to use RID to predict TBS in the group and its individuals. Methods: Women (n = 89) ingested a dose of [2H6]retinyl acetate and blood samples (3/woman) were collected from 6 h to 91 d, with all participants sampled at 14 d, about half at either 21 or 28 d, and each at one other time. Composite data (plasma retinol fraction of dose; FDp) were analyzed using Simulation, Analysis and Modeling software to obtain kinetic parameters, TBS, and other state variables as well as model-derived values for the RID composite coefficient FaS. The latter were used in the RID equation TBS (μmol) = FaS × 1/SAp (where SAp is plasma retinol specific activity) to predict TBS at various times. Results: Model-predicted TBS was 973 μmol (n = 87). Geometric mean RID-predicted TBS was 965, 926, and 1006 μmol at 14, 21, and 28 d, respectively, with wide ranges [for example, 252–3848 μmol on day 14 (n = 86)]; TBS predictions were similar at later times. Participants had a mean 2 y of vitamin A in stores and estimated liver vitamin A concentrations in the normal range. Model-predicted vitamin A disposal rate was 1.3 μmol/d and plasma recycling number was 37. Conclusions: Super-subject modeling provides an estimate of group mean TBS as well as group-specific values for the RID coefficient FaS; the latter can be used to confidently predict TBS by RID for individual participants in the group under study or in similar individuals at 14 d or more after isotope ingestion. Trial registration number: Trial is registered (NCT04632771) at https://clinicaltrials.gov.

BackgroundYoung children exposed to high-dose vitamin A supplements (VAS) and vitamin A (VA)-fortified foods may be at risk of high VA intake and high VA total body stores (TBS).ObjectivesTBS and estimated liver VA concentration were compared among children with adequate or high VA intake and different timing of exposure to VAS, and associations between estimated liver VA concentrations and biomarkers of VA toxicity were examined.MethodsChildren 12-18 mo of age (n = 123) were selected for 3 groups: 1) retinol intake >600 µg/d and VAS within the past mo, 2) retinol intake >600 µg/d and VAS in the past 3-6 mo, and 3) VA intake 200-500 µg retinol activity equivalents (RAE)/d and VAS in the past 3-6 mo. Dietary intake data were collected to measure VA intakes from complementary foods, breast milk, and low-dose, over-the-counter supplements. TBS were assessed by retinol isotope dilution, and VA toxicity biomarkers were measured. Main outcomes were compared by group.ResultsMean (95% CI) VA intakes excluding VAS were 1184 (942, 1426), 980 (772, 1187), and 627 (530, 724) µg RAE/d, in groups 1-3, respectively; mean VA intake was higher in groups 1 and 2 compared with group 3 (P 1 µmol/g liver). There was no evidence of chronic VA toxicity, suggesting that the liver VA cutoff value should be re-evaluated. This trial was registered at www.clinicaltrials.gov as NCT03030339.

BackgroundThe 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.ObjectiveThe 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.MethodsFour 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.ResultsThe 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

We review recent work in which model-based compartmental analysis has been applied to data for theoretical human subjects in order to study questions related to vitamin A kinetics and metabolism. Using model simulations in this way, one can validate experimental designs, evaluate or improve vitamin A assessment methods, study the influence of perturbations on assessment methods, and/or advance information related to retinol kinetics. We also provide some information on the rationale for assigning physiologically appropriate values for specified characteristics [e.g., plasma retinol concentration, vitamin A total body stores (TBS), vitamin A intake] to hypothetical individuals, and in addition, we outline how one might first select an appropriate compartmental model for whole-body vitamin A metabolism and then specify physiologically reasonable values for the associated retinol kinetic parameters. In the studies discussed here, the Simulation, Analysis, and Modeling software was used to simulate responses in key model compartments for hypothetical subjects so that model predictions could be compared to assigned values or projected outcomes. For example, in the case of the retinol isotope dilution (RID) method that is used to assess vitamin A status, application of this approach has provided a way to evaluate the accuracy of TBS predictions under different steady state and non-steady state conditions, thus increasing confidence about the validity of RID results obtained in the field. Although datasets for theoretical subjects have been used to evaluate protocols in pharmacokinetics, to our knowledge, other nutrition researchers have not previously used approaches such as those described here. Our results to date indicate that this strategy has the potential to provide useful information related not only to vitamin A but perhaps to other nutrients as well. [ABSTRACT FROM AUTHOR]

Background: As currently applied, the paired retinol isotope dilution (RID) test, which is used to assess the impact of a vitamin A intervention on vitamin A total body stores (TBS), requires 2 doses of stable isotope-labeled vitamin A., Objectives: The objectives of this study were to evaluate use of a single isotope dose (4 μmol) to assess TBS by RID before and after intervention in theoretical children with low/moderate TBS., Methods: We selected 6 theoretical children with assigned values for TBS ranging from 82 to 281 μmol. Using Simulation, Analysis and Modeling software, we simulated the variable [plasma retinol specific activity (SA p )] and coefficients (Fa and S) used in the RID equation TBS (μmol) = FaS × 1/SA p in both the unsupplemented steady state at day 14 postdosing and during the subsequent 4 mo without or with vitamin A supplementation [2.8 μmol retinol/d (801 μg retinol activity equivalents/d)]., Results: Fraction of dose in plasma on day 150 compared with day 14 was similar in the unsupplemented and supplemented conditions [geometric mean, 32% (range, 20%-48%) and 30% (20%-48%), respectively] and simulated values for FaS were similar under the 2 conditions. After 2 and 4 mo of daily vitamin A supplementation with 2.8 μmol/d, TBS was 78% and 128% higher, respectively, than without supplementation., Conclusions: Results indicate that the paired RID method can successfully be done using a single 4 μmol dose of stable isotope. Furthermore, because values for the RID coefficient FaS were similar in the unsupplemented and vitamin A-supplemented conditions, these results in theoretical children indicate that FaS determined by population ("super-subject") modeling of steady state vitamin A kinetic data could be used to predict TBS by RID after a vitamin A intervention in individuals from the same or a similar group., (Copyright © 2024 American Society for Nutrition. Published by Elsevier Inc. All rights reserved.)

Background: Vitamin A status may influence the choice of a blood sampling time for applying the retinol isotope dilution (RID) equation to predict vitamin A total body stores (TBS) in children.Objectives: We aimed to identify time(s) after administration of labeled vitamin A that provide accurate estimates of TBS in theoretical children with low or high TBS.Methods: We postulated 2- to 5-y-old children (12/group) with low (<200 μmol) or high TBS (≥700 μmol) and used compartmental analysis to simulate individual subject values for the RID equation TBS =   FaS/SAp (Fa, fraction of dose in stores; S, retinol specific activity in plasma/in stores; SAp, retinol specific activity in plasma). Using individual SAp and group geometric mean FaS values from 1-28 d, we calculated individual and group mean TBS and compared them to assigned values.Results: Mean TBS was accurately predicted for both groups at all times. For individuals, predicted and assigned TBS were closest when the CV% for FaS was low [12-14%; 4-13 d (low), 12-28 d (high)]. The mean percentage error for TBS was <10% from 2-19 d (low) and 7-28 d (high). Predicted TBS was within 25% of assigned TBS for ≥80% of children from 3-23 d (low) and 9-28 d (high). Within groups, RID tended to overestimate lower TBS and underestimate higher TBS.Conclusions: Using a good estimate for FaS, accurate RID predictions of TBS for individuals will be obtained at many times. If vitamin A status is low, results indicate that early sampling (e.g., 4-13 d) is optimal; if vitamin A status is high, sampling at 12-28 d is indicated. When vitamin A status is unknown, sampling at 14 d is recommended, or a super-subject design can be used to obtain the group mean FaS at various times for RID prediction of TBS in individuals. [ABSTRACT FROM AUTHOR]

Background: An optimal blood sampling time for application of the retinol isotope dilution (RID) method for predicting vitamin A total body stores (TBS) (i.e., vitamin A status) has not been established.Objectives: Objectives were to identify sampling times that provide accurate estimates of TBS by RID in groups and individuals by applying compartmental modeling to data for theoretical adults and children.Methods: We selected previously generated hypothetical adults and children (20 per group) that had a wide range of assigned values for TBS and vitamin A kinetic parameters. We used the Simulation, Analysis and Modeling software to simulate individual kinetic responses; then we calculated geometric mean values for the RID equation coefficients and each individual's plasma retinol specific activity at various times, using those values to predict group mean and individual subject TBS. Predicted values for TBS were compared with assigned values.Results: Accurate estimates of group mean TBS were obtained at all sampling times from 1 to 30 d in both adults and children. For individuals, correlations between RID-predicted TBS and assigned values increased with time in the adults (R2 = 0.80 at day 14, 0.96 at day 21, and 0.99 at day 28); a similar trend was observed for the children, with R2 = 0.82 at day 7 and increasing to 0.97 at days 21 and 28 (P < 0.001 for all comparisons).Conclusions: Although no single, unique time provided the most accurate prediction of TBS for all individuals within these groups, applying the RID method at 21 or 28 d yielded predictions that were within 25% of assigned values for 90% or 95% of adults, respectively; corresponding values for children were 80% from 10 to 20 d, and 85% at 21 and 28 d. For most subjects, early times (<14 d for adults and <10 d for children) provided less accurate predictions. [ABSTRACT FROM AUTHOR]

Background: Retinol isotope dilution (RID) and model-based compartmental analysis are recognized techniques for assessing vitamin A (VA) status. Recent studies have shown that RID predictions of VA total body stores (TBS) can be improved by using modeling and that VA kinetics and TBS in children can be effectively studied by applying population modeling ("super-child" approach) to a composite data set.Objectives: The objectives were to model whole-body retinol kinetics and predict VA TBS in a group of Mexican preschoolers using the super-child approach and to use model predictions of RID coefficients to estimate TBS by RID in individuals.Methods: Twenty-four healthy Mexican children (aged 3-6 y) received an oral dose (2.96 μmol) of [13C10]retinyl acetate in corn oil. Blood samples were collected from 8 h to 21 d after dosing, with each child sampled at 4 d and at 1 other time. Composite data for plasma labeled retinol compared with time were analyzed using a 6-component model to obtain group retinol kinetic parameters and pool sizes. Model-predicted TBS was compared with mean RID predictions at 4 d; RID estimates at 4 d were compared with those calculated at 7-21 d.Results: Model-predicted TBS was 1097 μmol, equivalent to ∼2.4 y-worth of VA; using model-derived coefficients, group mean RID-predicted TBS was 1096 μmol (IQR: 836-1492 μmol). TBS at 4 d compared with a later time was similar (P = 0.33). The model predicted that retinol spent 1.5 h in plasma during each transit and recycled to plasma 13 times before utilization.Conclusions: The super-child modeling approach provides information on whole-body VA kinetics and can be used with RID to estimate TBS at any time between 4 and 21 d postdose. The high TBS predicted for these children suggests positive VA balance, likely due to large-dose VA supplements, and warrants further investigation. [ABSTRACT FROM AUTHOR]

Retinol isotope dilution (RID) is a well-accepted technique for assessing vitamin A status [i.e., total body stores (TBS)]. Here, in an effort to increase understanding of the method, we briefly review RID equations and discuss their included variables and their coefficients (i.e., assumptions that account for the efficiency of absorption of an orally administered tracer dose of vitamin A, mixing of the dose with endogenous vitamin A, and loss due to utilization). Then, we focus on contributions of another technique, model-based compartmental analysis and especially the "super-person" approach, that advance the RID method. Specifically, we explain how adding this modeling component, which involves taking 1 additional blood sample from each subject, provides population-specific estimates for the RID coefficients that can be used in the equation instead of values derived from the literature; using model-derived RID coefficients results in improved confidence in predictions of TBS for both a group and its individuals. We note that work is still needed to identify the optimal time for applying RID in different groups and to quantify vitamin A absorption efficiency. Finally, we mention other contributions of modeling, including the use of theoretical data to verify the accuracy of RID predictions and the additional knowledge that model-based compartmental analysis provides about whole-body vitamin A kinetics. [ABSTRACT FROM AUTHOR]

Background: Biofortification of staple crops with β-carotene is a strategy to reduce vitamin A deficiency, and several varieties are available in some African countries. β-Cryptoxanthin (BCX)-enhanced maize is currently in field trials. To our knowledge, maize BCX bioavailability has not been assessed in humans. Serum retinol 13C content and xanthophyll concentrations are proposed effectiveness biomarkers for biofortified maize adoption. Objective: We determined the relative difference in BCX and zeaxanthin bioavailability from whole-grain and refined BCX-biofortified maize during chronic feeding compared with white maize and evaluated short-term changes in 13C-abundance in serum retinol. Design: After a 7-d washout, 9 adults (mean ± SD age: 23.4 ± 2.3 y; 5 men) were provided with muffins made from BCX-enhanced whole-grain orange maize (WGOM), refined orange maize (ROM), or refined white maize (RWM) for 12 d in a randomized, blinded, crossover study followed by a 7-d washout. Blood was drawn on days 0, 3, 6, 9, 12, 15, and 19. Carotenoid areas under the curve (AUCs) were compared by using a fixed-effects model. 13C-Abundance in serum retinol was determined by using gas chromatography/combustion/isotope-ratio mass spectrometry on days 0, 12, and 19. Vitamin A status was determined by 13C-retinol isotope dilution postintervention. Results: The serum BCX AUC was significantly higher for WGOM (1.70 ± 0.63 μmol ⋅ L−1 ⋅ d) and ROM (1.66 ± 1.08 μmol ⋅ L−1 ⋅ d) than for RWM (−0.06 ± 0.13 μmol ⋅ L−1 ⋅ d; P < 0.003). A greater increase occurred in serum BCX from WGOM muffins (131%) than from ROM muffins (108%) (P ≤ 0.003). Zeaxanthin AUCs were higher for WGOM (0.94 ± 0.33) and ROM (0.96 ± 0.47) than for RWM (0.05 ± 0.12 μmol ⋅ L−1 ⋅ d; P < 0.003). The intervention did not affect predose serum retinol 13C-abundance. Vitamin A status was within an optimal range (defined as 0.1–0.7 μmol/g liver). Conclusions: BCX and zeaxanthin were highly bioavailable from BCX-biofortified maize. The adoption of BCX maize could positively affect consumers’ BCX and zeaxanthin intakes and associated health benefits. This trial is registered at www.clinicaltrials.gov as NCT02800408. [ABSTRACT FROM AUTHOR]

Background: Worldwide, an estimated 250 million children <5 y old are vitamin A (VA) deficient. In Mexico, despite ongoing efforts to reduce VA deficiency, it remains an important public health problem; thus, food-based interventions that increase the availability and consumption of provitamin A-rich foods should be considered.Objective: The objectives were to assess the VA equivalence of 2H-labeled Moringa oleifera (MO) leaves and to estimate both total body stores (TBS) of VA and plasma retinol kinetics in young Mexican children.Methods: β-Carotene was intrinsically labeled by growing MO plants in a 2H2O nutrient solution. Fifteen well-nourished children (17-35 mo old) consumed puréed MO leaves (1 mg β-carotene) and a reference dose of [13C10]retinyl acetate (1 mg) in oil. Blood (2 samples/child) was collected 10 times (2 or 3 children each time) over 35 d. The bioefficacy of MO leaves was calculated from areas under the composite "super-child" plasma isotope response curves, and MO VA equivalence was estimated through the use of these values; a compartmental model was developed to predict VA TBS and retinol kinetics through the use of composite plasma [13C10]retinol data. TBS were also estimated with isotope dilution.Results: The relative bioefficacy of β-carotene retinol activity equivalents from MO was 28%; VA equivalence was 3.3:1 by weight (0.56 μmol retinol:1 μmol β-carotene). Kinetics of plasma retinol indicate more rapid plasma appearance and turnover and more extensive recycling in these children than are observed in adults. Model-predicted mean TBS (823 μmol) was similar to values predicted using a retinol isotope dilution equation applied to data from 3 to 6 d after dosing (mean ± SD: 832 ± 176 μmol; n = 7).Conclusions: The super-child approach can be used to estimate population carotenoid bioefficacy and VA equivalence, VA status, and parameters of retinol metabolism from a composite data set. Our results provide initial estimates of retinol kinetics in well-nourished young children with adequate VA stores and demonstrate that MO leaves may be an important source of VA. [ABSTRACT FROM AUTHOR]

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 [ 13 C 10 ]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., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Retinol isotope dilution (RID) methodology provides a quantitative estimate of total body vitamin A (VA) stores and is the best method currently available for assessing VA status in adults and children. The methodology has also been used to test the efficacy of VA interventions in a number of low-income countries. Infections, micronutrient deficiencies (eg, iron and zinc), liver disease, physiological age, pregnancy, and lactation are known or hypothesized to influence the accuracy of estimating total body VA stores using the isotope dilution technique.Objective: Our objectives were to review the strengths and limitations of RID methods, to discuss what is known about the impact of various factors on results, and to summarize contributions of model-based compartmental analysis to assessing VA status.Methods: Relevant published literature is reviewed and discussed.Results: Various equations and compartmental modeling have been used to estimate the total body VA stores using stable isotopes, including a newer 3-day equation that provides an estimate of total body VA stores in healthy adults. At present, there is insufficient information on absorption of the isotope tracer, and there is a need to further investigate how various factors impact the application of RID techniques in field studies.Conclusions: Isotope dilution methodology can provide useful estimates of total body VA stores in apparently healthy populations under controlled study conditions. However, more research is needed to determine whether the method is suitable for use in settings where there is a high prevalence of infection, iron deficiency, and/or liver disease. [ABSTRACT FROM AUTHOR]

Background: Stable isotope techniques using 13 C to assess vitamin A (VA) dietary sources, absorption, and total body VA stores (TBSs) require determination of baseline 13 C abundance. 13 C-natural abundance is approximately 1.1% total carbon, but varies with foods consumed, supplements taken, and food fortification with synthetic retinyl palmitate., Objectives: We determined 13 C variation from purified serum retinol and the resulting impact on TBSs using pooled data from preschool children in Burkina Faso, Cameroon, Ethiopia, South Africa, Tanzania, and Zambia and Zambian women., Methods: Seven studies included children (n = 639; 56 ± 25 mo; 48% female) and one in women (n = 138; 29 ± 8.5 y). Serum retinol 13 C-natural abundance was determined using GC-C-IRMS. TBSs were available in 7 studies that employed retinol isotope dilution (RID). Serum CRP and α 1 -acid-glycoprotein (AGP) were available from 6 studies in children. Multivariate mixed models assessed the impact of covariates on retinol 13 C. Spearman correlations and Bland-Altman analysis compared serum and milk retinol 13 C and evaluated the impact of using study- or global-retinol 13 C estimates on calculated TBSs., Results: 13 C-natural abundance (%, median [Q1, Q3]) differed among countries (low: Zambia, 1.0744 [1.0736, 1.0753]; high: South Africa, 1.0773 [1.0769, 1.0779]) and was associated with TBSs, CRP, and AGP in children and with TBSs in women. 13 C-enrichment from serum and milk retinol were correlated (r = 0.52; P = 0.0001). RID in children and women using study and global estimates had low mean bias (range, -3.7% to 2.2%), but larger 95% limits of agreement (range, -23% to 37%)., Conclusions: 13 C-natural abundance is different among human cohorts in Africa. Collecting this information in subgroups is recommended for surveys using RID. When TBSs are needed on individuals in clinical applications, baseline 13 C measures are important and should be measured in all enrolled subjects., (Copyright © 2023. Published by Elsevier Inc.)

Background: Vitamin A (VA) assessment is important for targeting public health programs. Retinol isotope dilution (RID) is a sensitive method to estimate total body VA stores (TBSs) and total liver reserves (TLRs), but the impact of subclinical inflammation on RID is unclear., Objective: We determined the association between TBSs and TLRs, estimated by RID, and inflammation among preschool children without clinical infection in Burkina Faso, Cameroon, Ethiopia, South Africa, and Tanzania., Methods: Five studies (n = 532; 47.9 ± 8.3 mo; 49.0% male) included 13 C-RID and measurement of inflammation markers, CRP, and α 1 -acid glycoprotein (AGP). Spearman correlations were used to evaluate TBSs and TLRs with inflammation biomarkers. Wilcoxon and Kruskal-Wallis tests were used to compare TBSs and TLRs by inflammation categories [normal vs. elevated CRP (>5 mg/L) or AGP (>1 g/L)] and inflammation stage [reference, incubation (elevated CRP), early convalescence (elevated CRP and AGP), and late convalescence (elevated AGP)]., Results: Complete data were available for 439 children. Median (Q1, Q3) TLRs ranged from 0.12 (0.07, 0.18) μmol/g in Ethiopia to 1.10 (0.88, 1.38) μmol/g in South Africa. Elevated CRP ranged from 4% in Burkina Faso to 42% in Cameroon, and elevated AGP from 20% in Tanzania to 58% in Cameroon. Pooled analysis (excluding Cameroon) showed a negative correlation between TBSs and AGP (ρ = -0.131, P = 0.01). Children with elevated AGP had higher probability of having lower TBSs (probability = 0.61, P = 0.002). TBSs differed among infection stages (P = 0.020). Correlations between TLRs and CRP or AGP were not significant., Conclusions: No indication of systematic bias in RID-estimated TLRs was found due to subclinical inflammation among preschool children. The inverse relationship between TBSs and AGP may reflect decreased stores after infection or an effect of inflammation on isotope partitioning. Further research should investigate potential confounding variables to improve TBS-estimate validity., (Copyright © 2022. Published by Elsevier Inc.)

Background: Biomarkers of micronutrient status are needed to best define deficiencies and excesses of essential nutrients. Objective: We evaluated several supporting biomarkers of vitamin A status in Zambian children to determine whether any of the biomarkers were consistent with high liver retinol stores determined by using retinol isotope dilution (RID). Design: A randomized, placebo-controlled, biofortified maize efficacy trial was conducted in 140 rural Zambian children from 4 villages. A series of biomarkers were investigated to better define the vitamin A status of these children. In addition to the assessment of total-body retinol stores (TBSs) by using RID, tests included analyses of serum carotenoids, retinyl esters, and pyridoxal-5'-phosphate (PLP) by using high-pressure liquid chromatography, retinol-binding protein by using ELISA, and alanine aminotransfer-ase (ALT) activity by using a colorimetric assay. Results: Children (n = 133) were analyzed quantitatively for TBSs by using RID. TBSs, retinyl esters, some carotenoids, and PLP differed by village site. Serum carotenoids were elevated above most nonintervened reference values for children. α -Carotene, β-carotene, and lutein values were >95th percentile from children in the US NHANES III, and 13% of children had hypercarotenemia (defined as total carotenoid concentration >3.7 µmol/L). Although only 2% of children had serum retinyl esters >10% of total retinol plus retinyl esters, 16% of children had .5% as esters, which was consistent with high liver retinol stores. Ratios of serum retinol to retinol-binding protein did not deviate from 1.0, which indicated full saturation. ALT activity was low, which was likely due to underlying vitamin B-6 deficiency, which was confirmed by very low serum PLP concentrations. Conclusions: The finding of hypervitaminosis A in Zambian children was supported by high circulating concentrations of carotenoids and mildly elevated serum retinyl esters. ALT-activity assays may be compromised with co-existing vitamin B-6 deficiency. Nutrition education to improve intakes of whole grains and animal-source foods may enhance vitamin B-6 status in Zambians. This trial was registered at clinicaltrials.gov as NCT01814891. [ABSTRACT FROM AUTHOR]

Impact Statement: Vitamin A (VA) deficiency and hypervitaminosis A have been reported in groups of people worldwide. Conventional biomarkers of VA deficiency (e.g. serum retinol concentration, dose response tests) are not able to distinguish between sufficiency and hypervitaminosis A. Retinol isotope dilution (RID) predictions of VA status have been validated in humans and animal models from deficiency through toxicity; however, RID during life stages with unique issues related to isotopic tracing, such as infancy and lactation, requires further evaluation. This study investigated RID in piglets and lactating sows as models for human infants and women. In piglets, RID successfully determined VA deficiency (confirmed with liver analysis), and that the tracer mixes quickly. Conversely, in lactating sows, although serum and milk enrichments were similar, traditional RID equations overestimated VA stores, likely due to losses of tracer and higher extrahepatic VA storage than predictions. These data inform researchers about the challenges of using RID during lactation.

Plants that undergo C 4 photosynthesis, such as maize, are enriched in the stable isotope of carbon ( 13 C) compared with other dietary plants and foods. Consumption of maize that has been biofortified to contain elevated levels of provitamin A carotenoids (orange maize) increased the abundance of 13 C in serum retinol of Mongolian gerbils. We evaluated this method in humans to determine if it has potential for further use in intervention effectiveness studies. A random subset of samples from a two-month randomized controlled feeding trial of rural three- to five-year old Zambian children were used to determine the impact of orange maize intake on serum carotenoid concentrations ( n = 88) and 13 C-natural abundance in serum retinol ( n = 77). Concentrations of β-cryptoxanthin (a xanthophyll provitamin A carotenoid) and the dihydroxy xanthophylls lutein and zeaxanthin, which do not have vitamin A activity, were elevated in children consuming orange maize compared with those consuming a white maize control ( P < 0.001), while β-carotene was not different ( P > 0.3). Furthermore, 13 C natural abundance was higher after two months' intervention in the orange maize group compared with the white maize group ( P = 0.049). Predictions made from equations developed in the aforementioned gerbil study estimated that maize provided 11% (2-21%, 95% confidence interval) of the recent dietary vitamin A to these children. These results demonstrate that orange maize is efficacious at providing retinol to the vitamin A pool in children through provitamin A carotenoids, as monitored by the change in 13 C enrichment, which was not reflected in serum β-carotene concentrations. Further effectiveness studies in countries who have adopted orange maize should consider determining differences in retinol 13 C-enrichment among target groups in addition to profiling serum xanthophyll carotenoids with specific emphasis on zeaxanthin. Impact statement Maize biofortified with provitamin A carotenoids (orange) has been released in some African markets. Responsive and sensitive methods to evaluate dissemination effectiveness are needed. This study investigated methods to evaluate effectiveness of orange maize consumption using serum from Zambian children fed orange maize for two months. Many varieties of orange maize contain higher amounts of the xanthophyll carotenoids in addition to β-carotene compared with typical varieties. This study uniquely showed higher concentrations of the maize xanthophylls lutein, zeaxanthin, and β-cryptoxanthin in children who consumed orange maize compared with white. Furthermore, maize is a C 4 plant and is therefore naturally enriched with 13 C. Higher 13 C was detected in the serum retinol of the orange maize consumers with no change in serum β-carotene concentration suggesting preferential bioconversion to retinol. The combined analyses of serum zeaxanthin specifically and 13 C-natural abundance of retinol could prove useful in effectiveness studies between orange maize adopters and non-adopters.