The Use of Datasets for Theoretical Subjects to Validate Vitamin A-Related Methods and Experimental Designs.

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.

Influence of Vitamin A Status on the Choice of Sampling Time for Application of the Retinol Isotope Dilution Method in Theoretical Children.

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.

Use of Model-Based Compartmental Analysis and Theoretical Data to Further Explore Choice of Sampling Time for Assessing Vitamin A Status in Groups and Individual Human Subjects by the Retinol Isotope Dilution Method.

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.

Development of a Compartmental Model to Investigate the Influence of Inflammation on Predictions of Vitamin A Total Body Stores by Retinol Isotope Dilution in Theoretical Humans.

BACKGROUND: Inflammation, both acute and chronic, is associated with reductions in the synthesis of retinol-binding protein (RBP) and the concentration of retinol in plasma. Consequently, it is currently recommended that the retinol isotope dilution (RID) method not be used to estimate vitamin A total body stores (TBS) in subjects with inflammation. OBJECTIVES: To apply compartmental analysis to study the effects of inflammation on hepatic apo-RBP input, plasma retinol pool size, and RID-predicted TBS in theoretical subjects with known steady state values for these parameters. METHODS: We selected 6 previously generated hypothetical subjects who ingested a dose of stable isotope-labeled vitamin A (day 0). Starting with each subject's published steady state model for retinol tracer kinetics, we developed a parallel model for unlabeled retinol and RBP that incorporated links between these entities and tied liver retinol secretion to RBP availability. Then we perturbed the steady state model by initiating chronic or acute inflammation on day 0 or acute inflammation on day 3 or 9 and simulating results for RBP, plasma retinol, and TBS. RESULTS: Chronic inflammation led to substantial reductions in RID-predicted TBS for at least 2 weeks after tracer administration. In contrast, acute inflammation induced on day 0 or 3 resulted in less dramatic impacts on TBS (37% or 20% reduction, respectively, from steady state levels, with levels rebounding by 14 days). When inflammation was induced 9 days after administration of the tracer, the effects on predicted TBS were minimal. Overall, for acute inflammation initiated at 0, 3, or 9 days, accurate predictions of TBS were obtained by 2 weeks. CONCLUSIONS: Compartmental analysis can be applied in the novel way described here to study the influence of perturbations such as inflammation on predictions of vitamin A status using RID. Such an approach has potential value for studying other perturbations and different nutrients.

A Compartmental Model Describing the Kinetics of ß-Carotene and ß-Carotene-Derived Retinol in Healthy Older Adults.

BACKGROUND: Descriptive and quantitative information on ß-carotene whole-body kinetics in humans is limited. OBJECTIVES: Our objective was to advance the development of a physiologically based, working hypothesis compartmental model describing the metabolism of ß-carotene and ß-carotene-derived retinol. METHODS: We used model-based compartmental analysis (Simulation, Analysis and Modeling software) to analyze previously published data on plasma kinetics of [2H8]ß-carotene, [2H4]ß-carotene-derived retinol, and [2H8]retinyl acetate-derived retinol in healthy, older US adults (3 female, 2 male; 50-68 y); subjects were studied for 56 d after consuming doses of 11 µmol [2H8]ß-carotene and, 3 d later, 9 µmol [2H8]retinyl acetate in oil. RESULTS: We developed a complex model for labeled ß-carotene and ß-carotene-derived retinol, as well as preformed vitamin A, using simulations to augment observed data during model calibration. The model predicts that mean (range) ß-carotene absorption (bioavailability) was 9.5% (5.2-14%) and bioefficacy was 7.3% (3.6-14%). Of the absorbed ß-carotene, 41% (25-58%) was packaged intact in chylomicrons and the balance was converted to retinol, with 58% (42-75%) transported as retinyl esters in chylomicrons and 0-2% by retinol-binding protein. Most (95%) chylomicron ß-carotene was cleared by the liver. Later data revealed differences in the metabolism of retinyl acetate- versus ß-carotene-derived retinol; data required that both ß-carotene and derived retinol be recycled from extrahepatic tissues (e.g. adipose) in HDL. Of total bioconversion [73% (47-99%)], 82% occurred in the intestine, 17% in the liver, and 0.83% in other tissues. CONCLUSIONS: Our model advances knowledge about whole-body ß-carotene metabolism in healthy adults, including the kinetics of transport in all lipoprotein species, and suggests hypotheses to be tested in future studies, such as the possibility that retinol derived from hepatic conversion over a long period of time might contribute to plasma retinol homeostasis and total body vitamin A stores.

Vitamin A Absorption Efficiency Determined by Compartmental Analysis of Postprandial Plasma Retinyl Ester Kinetics in Theoretical Humans.

BACKGROUND: Better methods are needed for determining vitamin A absorption efficiency in humans to support development of dietary recommendations and to improve the accuracy of predictions of vitamin A status. OBJECTIVES: We developed and evaluated a method for estimating vitamin A absorption efficiency based on compartmental modeling of theoretical data on postprandial plasma retinyl ester (RE) kinetics. METHODS: We generated data on plasma RE and retinol kinetics (30 min to 8 h or 56 d, respectively) after oral administration of labeled vitamin A for 12 theoretical adults with a range of values assigned for vitamin A absorption (55-90%); we modeled all data to obtain best-fit values for absorption and other parameters using Simulation, Analysis, and Modeling software. We then modeled RE data only (16 or 10 samples), with or without added random error, and compared assigned to predicted absorption values. We also compared assigned values to areas under RE response curves (RE AUCs). RESULTS: We confirmed that a unique value for vitamin A absorption cannot be identified by modeling plasma retinol tracer kinetics. However, when RE data were modeled, predicted vitamin A absorptions were within 1% of assigned values using data without error and within 12% when 5% error was included. When the sample number was reduced, predictions were still within 13% for 10 of the 12 subjects and within 23% overall. Assigned values for absorption were not correlated with RE AUC (P = 0.21). CONCLUSIONS: We describe a feasible and accurate method for determining vitamin A absorption efficiency that is based on compartmental modeling of plasma RE kinetic data collected for 8 h after a test meal. This approach can be used in a clinical setting after fasting subjects consume a fat-containing breakfast meal with a known amount of vitamin A or a stable isotope label.

Dietary Iron Repletion Stimulates Hepatic Mobilization of Vitamin A in Previously Iron-Deficient Rats as Determined by Model-Based Compartmental Analysis.

BACKGROUND: Iron deficiency can result in hyporetinolemia and hepatic vitamin A (VA) sequestration. OBJECTIVES: We used model-based compartmental analysis to determine the impact of iron repletion on VA metabolism and kinetics in iron-deficient rats. METHODS: At weaning, Sprague-Dawley rats were assigned to either a VA-marginal diet (0.35 mg retinol equivalent/kg) with adequate iron (35 ppm, control group [CN]) or reduced iron (3 ppm, iron-deficient group [ID-]), with an equivalent average body weight for each group. After 5 wk, n = 4 rats from each group were euthanized for baseline measurements of VA and iron indices, and the remaining rats (n = 6 CN, n = 10 ID-) received an intravenous injection of 3H-labeled retinol in an emulsion as tracer to initiate the kinetic study. On day 21 after dosing, half of the ID- rats were switched to the CN diet to initiate iron repletion, referred to as the iron-repletion group (ID+). From the time of dosing, 34 serial blood samples were collected from each rat over a 92-d time course. Plasma tracer and tissue tracee data were fitted to 6- and 4-compartment models, respectively, to analyze the kinetic behavior of VA in all groups. RESULTS: Our mathematical model indicated that ID- rats exhibited a nearly 6-fold decrease in liver VA secretion and >4-fold reduction in whole-body VA utilization, compared with CN rats, whereas these perturbed kinetic behaviors were notably corrected in ID+ rats, close to those from the CN group. CONCLUSIONS: Iron repletion can remove the inhibitory effect that iron deficiency exerts on hepatic mobilization of VA and restore retinol kinetic parameters to values similar to that of never-deficient CN rats. Together with improvements in iron and VA indices, our results suggest that restoration of an iron-adequate diet is sufficient to improve VA kinetics after a previous state of iron deficiency.

The "Super-Child" Approach Is Applied To Estimate Retinol Kinetics and Vitamin A Total Body Stores in Mexican Preschoolers.

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.

Better Predictions of Vitamin A Total Body Stores by the Retinol Isotope Dilution Method Are Possible with Deeper Understanding of the Mathematics and by Applying Compartmental Modeling.

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.

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).

Addition of Vitamin A Intake Data during Compartmental Modeling of Retinol Kinetics in Theoretical Humans Leads to Accurate Prediction of Vitamin A Total Body Stores and Kinetic Parameters in Studies of Reasonable Duration.

BACKGROUND: Mathematical modeling of theoretical data has been used to validate experimental protocols and methods in several fields. OBJECTIVES: We hypothesized that adding dietary vitamin A intake data as an input during compartmental modeling of retinol kinetics would lead to accurate prediction of vitamin A total body stores (TBS) at 2 specified study lengths and would reduce study duration required to accurately define the system. METHODS: We generated reference values for state variables (including TBS and intake) and kinetic parameters for 12 theoretical individuals (4 each of children, younger adults, and older adults) based on modeling plasma retinol tracer data for 365 d. We compared TBS predictions using data to 28 d (children) or 56 d (adults) without and with intake included in the model to reference values for each subject. Then, by truncating data sets from 365 d, we determined the shortest study duration required to accurately define the system without and with inclusion of vitamin A intake. RESULTS: Reference values for TBS ranged from 30 to 3023 µmol. Study durations of 28 and 56 d were sufficient to accurately predict TBS for 6 of the 12 subjects without intake; adding intake resulted in accurate predictions of TBS for all individuals. When intake was not included as a modeling input, durations of 35-310 d were required to define the system; inclusion of intake data substantially reduced the time required to 10-42 d. CONCLUSIONS: Inclusion of vitamin A intake as additional data input when modeling vitamin A kinetics allows investigators to accurately predict TBS and define the vitamin A system in studies of reasonable length (4 wk in children and 8 wk in adults). Because it is generally possible to obtain estimates/measures of intake, including such data increases confidence in model predictions while also making studies more feasible.

Inclusion of Vitamin A Intake Data Provides Improved Compartmental Model-Derived Estimates of Vitamin A Total Body Stores and Disposal Rate in Older Adults.

BACKGROUND: Sampling times and study duration impact estimates of kinetic parameters and variables including total body stores (TBS) and disposal rate (DR) when compartmental analysis is used to analyze vitamin A kinetic data. OBJECTIVE: We hypothesized that inclusion of dietary intake (DI) of vitamin A as an additional input would improve confidence in predictions of TBS and DR when modeling results appear to indicate that studies are not long enough to accurately define the terminal slope of the plasma retinol isotope response curve. METHODS: We reanalyzed previously published data on vitamin A kinetics monitored over 52 d in 7 US and 6 Chinese adults (means: 56 y, BMI 26.6 kg/m2, 38% males), adding an estimate for vitamin A intake [2.8 µmol/d (mean RDA)] as an input during application of the Simulation, Analysis and Modeling software. RESULTS: Use of a model with 1 extravascular compartment (1 EV), as in the original analysis, resulted in predictions of vitamin A intake that were higher than physiologically reasonable; inclusion of intake data in a model with 2 extravascular compartments (2 EV DI) resulted in more realistic estimates of intake and DR. Specifically, predictions of DR by the 2 EV DI (versus 1 EV) model were 2.10 compared with 12.2 µmol/d (US) and 2.21 compared with 5.13 µmol/d (Chinese). Predictions of both TBS [2056 compared with 783 µmol (US) and 594 compared with 219 µmol (Chinese)] and days of vitamin A stores [981 compared with 64 d (US) and 269 compared with 43 d (Chinese)] were higher using the new approach. CONCLUSIONS: Inclusion of vitamin A intake as additional data input when modeling vitamin A kinetics can compensate for less-than-optimal study duration, providing more realistic predictions of vitamin A TBS and DR. This approach advances the application of compartmental analysis to the study of vitamin A and, potentially, other nutrients.

A Population-Based (Super-Child) Approach for Predicting Vitamin A Total Body Stores and Retinol Kinetics in Children Is Validated by the Application of Model-Based Compartmental Analysis to Theoretical Data.

BACKGROUND: Public health nutritionists need accurate and feasible methods to assess vitamin A status and to evaluate efficacy of interventions, especially in children. The application of population-based designs to tracer kinetic data is an effective approach that reduces sample burden for each child. OBJECTIVES: Objectives of the study were to use theoretical data to validate a population-based (super-child) approach for estimating group mean vitamin A total body stores (TBS) and retinol kinetics in children and to use population-based data to improve individual TBS predictions using retinol isotope dilution (RID). METHODS: We generated plasma retinol kinetic data from 6 h to 56 d for 50 theoretical children with high vitamin A intakes, assigning values within physiologically reasonable ranges for state variables and kinetic parameters ("known values"). Mean data sets for all subjects at extensive (n = 36) and reduced (n = 11) sampling times, plus 5 data sets for reduced numbers (5/time, except all at 4 d) and times, were analyzed using Simulation, Analysis and Modeling software. Results were compared with known values; population RID coefficients were used to calculate TBS for individuals. RESULTS: For extensive and reduced data sets including all subjects, population TBS predictions were within 1% of the known value. For 5 data sets reflecting numbers and times being used in ongoing super-child studies, predictions were within 1-17% of the known group value. Using RID equation coefficients from population modeling, TBS predictions at 4 d were within 25% of the known value for 66-80% of subjects and reflected the range of assigned values; when ranked, predicted and assigned values were significantly correlated (Rs  = 0.93, P < 0.0001). Results indicate that 7 d may be better than 4 d for applying RID in children. For all data sets, predictions for kinetic parameters reflected the range of known values. CONCLUSION: The population-based (super-child) approach provides a feasible experimental design for quantifying retinol kinetics, accurately estimating group mean TBS, and predicting TBS for individuals reasonably well.

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.

Retinol Isotope Dilution Is Applied during Restriction of Vitamin A Intake to Predict Individual Subject Total Body Vitamin A Stores at Isotopic Equilibrium.

BACKGROUND: Retinol isotope dilution (RID) equations are used to determine vitamin A status and the efficacy of vitamin A intervention programs. Recent work related to RID methods has focused on modifying the "Olson equation" to improve the accuracy of predictions of vitamin A total body stores (TBS) in individual subjects. OBJECTIVE: We investigated the hypothesis that short-term restriction of vitamin A intake would result in accurate RID prediction of vitamin A TBS in individuals. METHODS: We applied model-based compartmental analysis to a 6-component model derived from published retinol kinetic studies on 12 individuals with a wide range of vitamin A stores and determined vitamin A TBS in the steady state. Then we simulated the impact of eliminating or strictly limiting vitamin A intake at the time of isotope administration, while maintaining plasma retinol homeostasis, on retinol specific activity in plasma (SAp; fraction of dose/µmol retinol) and stores, and we calculated TBS using the simplified RID equation TBS = 0.75 × 1/SAp, where the fractional absorption of tracer was set at 0.75 and SAp was simulated 5 d after dosing. RESULTS: When vitamin A intake was zero or strictly limited (0.25 µmol/d), mean TBS predicted by the equation at 5 d after dose administration divided by TBS determined by using the model was 1.00 (range: 0.959-1.04) or 1.02 (range: 0.983 - 1.06), respectively. CONCLUSIONS: By eliminating or strictly limiting vitamin A input, isotopic equilibrium was reached by 5 d. At isotopic equilibrium, SAp is the same as that in the body's exchangeable vitamin A pools; under these conditions, SAp may be measured at any time from 5 d on and used to calculate TBS.

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.

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.

Direct and indirect vitamin A supplementation strategies result in different plasma and tissue retinol kinetics in neonatal rats.

Many questions remain regarding vitamin A (VA) supplementation of infants. Herein we compared direct oral VA supplementation of the neonate and indirect treatment through maternal dietary VA (M-VA) treatment on VA status and kinetics in neonatal rats. Treatments included direct VA combined with retinoic acid (RA) [D-VARA; VA (6 mg/kg) + 10% RA, given orally to neonates on postnatal day (P)2 and P3] and indirect VA supplementation through increased M-VA, compared with each other and oil-treated neonates. [(3)H]retinol was administered orally to all neonates on P4. Plasma and tissue [(3)H]retinol kinetics were determined from 1 h to 14 days post-dosing. D-VARA versus placebo dramatically increased liver and lung retinol, but only in the first 8-10 days. In M-VA neonates, liver and lung VA increased progressively throughout the study. Compartmental modeling of plasma [(3)H]retinol showed that both D-VARA and indirect M-VA reduced retinol recycling between plasma and tissues. Compartmental models of individual tissues predicted that D-VARA stimulated the uptake of VA in chylomicrons to extrahepatic tissues, especially intestine, while the uptake was not observed in M-VA neonates. In conclusion, indirect maternal supplementation had a greater sustained effect than D-VARA on neonatal VA status, while also differentially affecting plasma and tissue retinol kinetics.

Evaluation of the "“Olson equation"”, an isotope dilution method for estimating vitamin A stores.

Isotope dilution methods have been successfully used to estimate vitamin A status in human populations as well as to evaluate the impact of vitamin A interventions. The most commonly applied isotope dilution method is the retinol isotope dilution technique, which is based on the 1989 "“Olson equation"” for estimating total body vitamin A stores (sometimes equated to liver vitamin A) after an oral dose of labeled vitamin A. The equation relies on several factors related to absorption and retention of the dose, the equilibration of label in plasma vs. liver, and timing of a blood sample for measurement of labeled vitamin A. Here, the assumptions underlying these factors are discussed, and new results based on applying model-based compartmental analysis [specifically, the Simulation, Analysis and Modeling software (WinSAAM)] to data on retinol kinetics in humans are summarized. A simplification of the Olson equation, in which plasma tracer is measured 3 days after administration of the oral dose and several factors are eliminated, is presented. The potential usefulness of the retinol isotope dilution technique for setting vitamin A requirements and assessing vitamin A status in children, as well as the confounding effects of inflammation and likely variability in vitamin A absorption, are also discussed.

Retinol kinetics in unsupplemented and vitamin A-retinoic acid supplemented neonatal rats: a preliminary model.

Vitamin A (VA) metabolism in neonates is virtually uncharacterized. Our objective was to develop a compartmental model of VA metabolism in unsupplemented and VA-supplemented neonatal rats. On postnatal day 4, pups (n = 3/time) received 11,12-[(3)H]retinol orally, in either oil (control) or VA combined with retinoic acid (VARA) [VA (∼6 mg/kg body weight) + 10% retinoic acid]. Plasma and tissues were collected at 14 time points up to 14 days after dose administration. VARA supplementation rapidly, but transiently, increased total retinol mass in plasma, liver, and lung. It decreased the peak fraction of the dose in plasma. A multi-compartmental model developed to fit plasma [(3)H]retinol data predicted more extensive recycling of retinol between plasma and tissues in neonates compared with that reported in adults (144 vs. 12-13 times). In VARA pups, the recycling number for retinol between plasma and tissues (100 times) and the time that retinol spent in plasma were both lower compared with controls; VARA also stimulated the uptake of plasma VA into extravascular tissues. A VARA perturbation model indicated that the effect of VARA in stimulating VA uptake into tissues in neonates is both dramatic and transient.