Isotope dilution is currently the most accurate technique in humans to determine vitamin A status and bioavailability/bioconversion of provitamin A carotenoids such as β-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 13C10β-carotene and 1 mg 13C10retinyl acetate in human subjects over a 2 week period. A reverse phase C18 column and binary mobile phase solvent system separated β-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 12Cβ-carotene and 12C retinoids; m/z 547→330 and m/z 274→98 for 13C10β-carotene and 13C5 cleavage products; and m/z 279→100 for metabolites of 13C10retinyl 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 13C10retinyl acetate with 13C10β-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.
β-Carotene, the most abundant provitamin A carotenoid in the diet, is converted to retinal by β-carotene 15,15'-monoxygenase (BCMO1). However, β-carotene absorption and conversion into retinal is ...extremely variable among individuals, with proportions of low responders to dietary β-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 β-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:β-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.
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, I
2
=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.
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.
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.
Composite plasma 13C10retinol 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).
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.
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.
In humans, varying amounts of absorbed β‐carotene are oxidatively cleaved by the enzyme β,β‐carotene 15,15′‐monooxygenase 1 (BCMO1) into two molecules of all‐trans‐retinal. The other carotenoid ...cleavage enzyme β,β‐carotene 9′,10′‐dioxygenase (BCDO2) cleaves β‐carotene at the 9′,10′ double bond forming β‐apo‐10′‐carotenal and β‐ionone. Although the contribution of BCDO2 to vitamin A formation has long been debated, BCMO1 is currently considered the key enzyme for retinoid metabolism. Furthermore, BCMO1 has limited enzyme activity towards carotenoids other than provitamin A carotenoids, whereas BCDO2 exhibits a broader specificity. Both enzymes are located at different sites within the cell, with BCMO1 being a cytosolic protein and BCDO2 being located in the mitochondria. Expression of BCMO1 in tissues other than the intestine has recently revealed its function for tissue‐specific retinoid metabolism with importance in embryogenesis and lipid metabolism. On the other hand, biological activity of BCDO2 metabolites has been shown to be important in protecting against carotenoid‐induced mitochondrial dysfunction. Single‐nucleotide polymorphisms (SNPs) such as R267S and A379V in BCMO1 can partly explain inter‐individual variations observed in carotenoid metabolism. Advancing knowledge about the physiological role of these two enzymes will contribute to understanding the importance of carotenoids in health and disease.
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.
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.
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
C
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
C
retinyl acetate.
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/SA
), where SA
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 (R
= 0.94 and 0.94; P < 0.001).
The equation TBS ≈ 0.5 × (1/SA
) 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.
Replacement of conventional staples with biofortified or industrially fortified staples in household diets may increase maternal breast milk retinol content and vitamin A intakes from complementary ...foods, improving infant total body stores (TBS) of vitamin A.
To determine whether biofortified or industrially fortified maize consumption by Zambian women and their breastfeeding infants could improve milk retinol concentration and infant TBS.
We randomly assigned 255 lactating women and their 9-mo-old infants to a 90-d intervention providing 0 µg retinol equivalents (RE)/d as conventional maize or ∼315 µg RE/d to mothers and ∼55 µg RE/d to infants as provitamin A carotenoid-biofortified maize or retinyl palmitate–fortified maize. Outcomes were TBS, measured by retinol isotope dilution in infants (primary), and breast milk retinol, measured by HPLC in women (secondary).
The intervention groups were comparable at baseline. Loss to follow-up was 10% (n = 230 mother-infant pairs). Women consumed 92% of the intended 287 g/d and infants consumed 82% of the intended 50 g/d maize. The baseline geometric mean (GM) milk retinol concentration was 1.57 μmol/L (95% CI: 1.45, 1.69 μmol/L), and 24% of women had milk retinol <1.05 μmol/L. While mean milk retinol did not change in the biofortified arm (β: 0.11; 95% CI: −0.02, 0.24), the intervention reduced low milk retinol (RR: 0.42; 95% CI: 0.21, 0.85). Fortified maize increased mean milk retinol (β: 0.17; 95% CI: 0.04, 0.30) and reduced the prevalence of low milk retinol (RR: 0.46; 95% CI: 0.25, 0.82). The baseline GM TBS was 178 μmol (95% CI: 166, 191 μmol). This increased by 24 µmol (± 136) over the 90-d intervention period, irrespective of treatment group.
Both biofortified and fortified maize consumption improved milk retinol concentration. This did not translate into greater infant TBS, most likely due to adequate TBS at baseline. This trial was registered at clinicaltrials.gov as NCT02804490.
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. β-Carotene bioefficacy is often estimated from the ratio of the areas under plasma isotope response curves after subjects ingest labeled β-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.
Our objectives were to develop and test a physiologically based compartmental model that includes both absorptive and postabsorptive β-carotene bioconversion and to use the model to evaluate the accuracy of the IRM and a simple plasma retinol isotope ratio (RIR), labeled β-carotene-derived retinol/labeled reference-dose-derived retinol in one plasma sample for estimating relative bioefficacy.
We used model-based compartmental analysis (Simulation, Analysis and Modeling software) to develop and apply a model that provided known values for β-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.
Plasma RIR accurately predicted β-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.
The plasma RIR is a simple tracer method that accurately predicts β-carotene relative bioefficacy based on analysis of one blood sample obtained at ≥14 d after co-ingestion of labeled β-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.
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 β-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 β-carotene from the ratio of retinyl esters to retinyl esters plus β-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.
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