Reflection Spectroscopy-Assessed Skin Carotenoids Are Sensitive to Change in Carotenoid Intake in a 6-Week Randomized Controlled Feeding Trial in a Racially/Ethnically Diverse Sample.

BACKGROUND: Reflection spectroscopy, utilized by the Veggie Meter, is a less-expensive, noninvasive method to quantify skin carotenoids and is a valid approximation of fruit and vegetable (FV) intake. However, it is unknown to what degree Veggie Meter-assessed skin carotenoid score change is responsive to changes in carotenoid intake. OBJECTIVES: This study aimed to evaluate Veggie Meter-assessed skin carotenoid score response in a 6-wk randomized controlled trial of a carotenoid-containing juice to determine whether the Veggie Meter can be used to detect nutritionally relevant changes in carotenoid intake; and to compare skin and plasma carotenoid responses with the 6-wk trial. METHODS: In this 6-wk trial, participants (n = 162) who self-identified as one of 4 US racial/ethnic groups (25% Black, 25% Asian, 27% non-Hispanic White, 23% Hispanic) were randomized to a control group, receiving negligible carotenoids (177 mL apple juice/d), moderate-dose group, receiving 4 mg total carotenoids/d (177 mL orange-carrot juice/d), or high-dose group, receiving 8 mg total carotenoids/d (355 mL orange-carrot juice/d). Skin carotenoid score and plasma total carotenoid concentrations (α-carotene, ß-carotene, ß-cryptoxanthin, lycopene, lutein, zeaxanthin) were assessed at baseline, 3 wk, and 6 wk (n = 158 completed the trial). Repeated measures linear models were used to examine skin and plasma carotenoids over time and between groups. RESULTS: At 6 wk, participants in the high-dose and moderate-dose groups had significantly higher mean skin carotenoid scores [414.0 (SD = 100.6) and 369.7 (SD = 100.3), respectively] compared with those in the control group [305.2 (100.5)]. In the high-dose group, there was a 42% change in skin carotenoids from baseline (mean = 290.4) to a 6-wk follow-up (increase of 123, 123/290 = 42.4%). There was a 61% change in the plasma carotenoids in the high-dose group. CONCLUSIONS: The Veggie Meter is sensitive to increases in daily carotenoid intake in diverse racial/ethnic groups over 6 wk. CLINICAL TRIALS REGISTRY NUMBER: This trial was registered at clinicaltrials.gov as ID: NCT04056624. Study URL: https://clinicaltrials.gov/ct2/show/NCT04056624.

Pressure-Mediated Reflection Spectroscopy Criterion Validity as a Biomarker of Fruit and Vegetable Intake: A 2-Site Cross-Sectional Study of 4 Racial or Ethnic Groups.

BACKGROUND: Valid biomarkers of fruit and vegetable (FV) intake are needed for field-based nutrition research. OBJECTIVES: To examine criterion-related validity of pressure-mediated reflection spectroscopy as a proxy measure of FV intake, using plasma carotenoids and self-reported FV and carotenoid intake as primary and secondary criterion measures, respectively. METHODS: Healthy adults 18-65 y of age, self-identifying as African American/black (n = 61), Asian (n = 53), white (n = 70), or Hispanic (n = 29), in North Carolina and Minnesota were recruited. Skin carotenoids were assessed via pressure-mediated reflection spectroscopy (Veggie Meter), skin melanin via spectrophotometer, and total plasma carotenoid concentration by HPLC-photodiode array detection. Self-reported carotenoid and FV intake was assessed using a semiquantitative FFQ. Relations between skin carotenoids, plasma carotenoids, FV, and carotenoid intake, with differences by race or ethnicity, age, sex, weight status, cholesterol, and melanin index, were examined by bivariate correlations and adjusted multivariate linear regressions. RESULTS: The overall unadjusted correlation between skin and total plasma carotenoids was r = 0.71 and ranged from 0.64 (non-Hispanic black) to 0.80 (Hispanic). Correlations between skin carotenoids and self-reported FV intake ranged from 0.24 (non-Hispanic black) to 0.53 (non-Hispanic white), with an overall correlation of r = 0.35. In models adjusted for age, sex, racial or ethnic group, and BMI, skin carotenoids were associated with plasma carotenoids (R2 = 0.55), FV (R2 = 0.17), and carotenoid intake (R2 = 0.20). For both plasma carotenoid and FV measures, associations with skin carotenoids did not vary by race, but these relations did differ by skin melanin-those with lower melanin had a lower correlation between skin and plasma carotenoids. CONCLUSIONS: Reflection spectroscopy-assessed skin carotenoids may be a reasonable alternative to measurement of plasma carotenoids, a biomarker used to approximate FV intake.

Carotenoid intake and adipose tissue carotenoid levels in relation to prostate cancer aggressiveness among African-American and European-American men in the North Carolina-Louisiana prostate cancer project (PCaP).

BACKGROUND: Associations between carotenoid intake and prostate cancer (CaP) incidence have varied across studies. This may result from combining indolent with aggressive disease in most studies. This study examined whether carotenoid intake and adipose tissue carotenoid levels were inversely associated with CaP aggressiveness. METHODS: Data on African-American (AA, n = 1,023) and European-American (EA, n = 1,079) men with incident CaP from North Carolina and Louisiana were analyzed. Dietary carotenoid intake was assessed using a detailed-food frequency questionnaire (FFQ), and abdominal adipose tissue samples were analyzed for carotenoid concentrations using high-performance liquid chromatography. Multivariable logistic regression was used in race-stratified analyses to calculate odds ratios (ORs) and 95% confidence intervals (95%CI) comparing high aggressive CaP with low/intermediate aggressive CaP. RESULTS: Carotenoid intake differed significantly between AAs and EAs, which included higher intake of lycopene among EAs and higher ß-cryptoxanthin intake among AAs. Comparing the highest and lowest tertiles, dietary lycopene was associated inversely with high aggressive CaP among EAs (OR = 0.55, 95%CI: 0.34-0.89, Ptrend = 0.02), while an inverse association was observed between dietary ß-cryptoxanthin intake and high aggressive CaP among AAs (OR = 0.56, 95%CI: 0.36-0.87, Ptrend = 0.01). Adipose tissue α-carotene and lycopene (cis + trans) concentrations were higher among EAs than AAs, and marginally significant inverse linear trends were observed for adipose α-carotene (Ptrend = 0.07) and lycopene (Ptrend = 0.11), and CaP aggressiveness among EAs only. CONCLUSIONS: These results suggest that diets high in lycopene and ß-cryptoxanthin may protect against aggressive CaP among EAs and AAs, respectively. Differences in dietary behaviors may explain the observed racial differences in associations. Prostate 76:1053-1066, 2016. © 2016 Wiley Periodicals, Inc.

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.

Circulating Estrogen Metabolites and Risk of Breast Cancer among Postmenopausal Women in the Nurses' Health Study.

BACKGROUND: Estradiol and estrone are well-established risk factors for postmenopausal breast cancer (BC). Experimental evidence suggests that specific estrogen metabolites, produced via irreversible hydroxylation of estrone and estradiol at the 2- or 16-position may independently influence carcinogenesis. METHODS: We performed a nested case-control study of BC (328 BC cases; 639 controls) among postmenopausal women within the Nurses' Health Study (NHS)to examine the role of estrogens and estrogen metabolites (jointly referred to as EM). Plasma concentrations of each EM (unconjugated+conjugated forms) were measured by liquid chromatography-tandem mass spectrometry. Multivariable conditional logistic regression, adjusting for BC risk factors, estimated relative risks (RR) and 95% confidence intervals (CI) of BC across quintiles of individual EM, EM pathways, and pathway ratios. Associations by estrogen/progesterone receptor (ER/PR) status were analyzed by unconditional logistic regression. RESULTS: Estradiol and estrone were strongly associated with increased BC risk [estradiol: RRQ5v.Q1 (95% CI)=2.64 (1.64-4.26), estrone: 2.78 (1.74-4.45); both p-trends<0.001]. The 2-hydroxylation pathway was strongly associated with risk [RRQ5v.Q1=3.09 (1.81-5.27), p-trend<0.001], and remained so after adjusting for unconjugated estradiol [RRQ5v.Q1=2.23 (1.25-3.96), p-trend=0.01]. While the 16-hydroxylation pathway was modestly associated with risk [RRQ5v.Q1=1.62 (1.03-2.54), p-trend=0.01], the association was attenuated after unconjugated estradiol adjustment [RRQ5v.Q1=1.24 (0.77-1.99), p-trend=0.19]. Similar positive associations with the 2-pathway and 16-pathway were observed for ER+/PR+ and ER-/PR- tumors. CONCLUSIONS: In this cohort of postmenopausal women, 2-hydroxylation of estrone and estradiol was associated with increased BC risk, independent of unconjugated estradiol. IMPACT: These results highlight the need to revisit the role of estrogen metabolism in BC etiology and prevention.

Low Xanthophylls, Retinol, Lycopene, and Tocopherols in Grey and White Matter of Brains with Alzheimer's Disease.

BACKGROUND: Oxidative stress contributes to pathogenesis and progression of Alzheimer's disease (AD). Higher levels of the dietary antioxidants- carotenoids and tocopherols- are associated with better cognitive functions and lower risk for AD, and lower levels of multiple carotenoids are found in serum and plasma of patients with AD. Although brains donated by individuals with mild cognitive impairment had significantly lower levels of lutein and beta-carotene, previous investigators found no significant difference in carotenoid levels of brains with AD and cognitively normal brains. OBJECTIVE: This study tested the hypothesis that micronutrients are significantly lower in donor brains with AD than in healthy elderly brains. METHODS: Samples of donor brains with confirmed AD or verified health were dissected into grey and white matter, extracted with organic solvents and analyzed by HPLC. RESULTS: AD brains had significantly lower levels of lutein, zeaxanthin, anhydrolutein, retinol, lycopene, and alpha-tocopherol, and significantly increased levels of XMiAD, an unidentified xanthophyll metabolite. No meso-zeaxanthin was detected. The overlapping protective roles of xanthophylls, carotenes, α- and γ-tocopherol are discussed. CONCLUSION: Brains with AD had substantially lower concentrations of some, but not all, xanthophylls, carotenes, and tocopherols, and several-fold higher concentrations of an unidentified xanthophyll metabolite increased in AD (XMiAD).

Biomarkers of oxidant load and type-specific clearance of prevalent oncogenic human papillomavirus infection: markers of immune response?

Human papillomavirus (HPV) infection is the cause of cervical cancer. Increased production of reactive oxygen species (ROS) maybe the common mechanism through which HPV-cofactors (i.e., smoking and inflammation) influence duration of infections. Biomarkers of total oxidant load may serve as cumulative measures of ROS exposure due to these cofactors. Therefore, we conducted a study evaluating the association between biomarkers of oxidant load and duration of HPV infections, early HPV natural history events. Serum samples were obtained from 444 HPV-positive women in the Ludwig-McGill Cohort Study. Anti-5-hydroxymethyl-2'-deoxyuridine autoantibody (anti-HMdU aAb) and malondialdehyde (MDA) were measured at baseline. Cox-proportional hazard models were used to estimate the probability of clearing any HPV, oncogenic HPV, non-oncogenic HPV and HPV-16 infections. Women with elevated MDA were significantly more likely to clear prevalent oncogenic HPV infections compared to those with lower MDA levels (Adjusted Hazard Ratio (AHR) = 2.7; 95%CI = 1.4-5.1). There did not appear to be an association between elevated MDA and clearance of incident oncogenic HPV infections. Similarly, women with elevated anti-HMdU aAb levels had higher rates of prevalent oncogenic HPV infection clearance (Quartile 3:AHR = 2.2; 95%CI = 1.2-4.4; Quartile 4:AHR = 2.4; 95%CI = 1.2-4.9). Higher levels of oxidant load biomarkers were associated with increased clearance of prevalent HPV infections. However, oxidant load biomarkers measured before incident infections were not associated, suggesting that the elevation of MDA and anti-HMdU aAb may reflect an ongoing effective immune response, such as increased innate immunity. More research focused on the immune responses to HPV and elevated markers of oxidant load is needed.

Multiplexed micronutrient, inflammation, and malarial antigenemia assessment using a plasma fractionation device.

Processing and storing blood samples for future analysis of biomarkers can be challenging in resource limited environments. The preparation of dried blood spots (DBS) from finger-stick collection of whole blood is a widely used and established method as DBS are biosafe, and allow simpler field processing, storage, and transport protocols than serum or plasma. Therefore, DBS are commonly used in population surveys to assess infectious disease and/or micronutrient status. Recently, we reported that DBS can be used with the Q-plex™ Human Micronutrient 7-plex Array (MN 7-plex), a multiplexed immunoassay. This tool can simultaneously quantify seven protein biomarkers related to micronutrient deficiencies (iodine, iron and vitamin A), inflammation, and malarial antigenemia using plasma or serum. Serum ferritin, an iron biomarker, cannot be measured from DBS due to red blood cell (RBC) ferritin content confounding the results. In this study, we assess a simple blood fractionation tool that passively separates plasma from other blood components via diffusion through a membrane into a plasma collection disc (PCD). We evaluated the concordance of MN 7-plex analyte concentrations from matched panels of eighty-eight samples of PCD, DBS, and wet plasma prepared from anticoagulated venous whole blood. The results showed good correlations of >0.93 between the eluates from PCD and DBS for each analyte except ferritin; while correlations seen for plasma/PCD were weaker. However, the recovery rate of the analytes from the PCD were better than those from DBS. The serum ferritin measures from the PCD were highly correlated to wet plasma samples (0.85). This suggests that surveillance for iron status in low resource settings can be improved over the current methods restricted to only measuring sTfR in DBS. When used in combination with the MN 7-plex, all seven biomarkers can be simultaneously measured using eluates from the PCDs.

A multicenter analytical performance evaluation of a multiplexed immunoarray for the simultaneous measurement of biomarkers of micronutrient deficiency, inflammation and malarial antigenemia.

A lack of comparative data across laboratories is often a barrier to the uptake and adoption of new technologies. Furthermore, data generated by different immunoassay methods may be incomparable due to a lack of harmonization. In this multicenter study, we describe validation experiments conducted in a single lab and cross-lab comparisons of assay results to assess the performance characteristics of the Q-plex™ 7-plex Human Micronutrient Array (7-plex), an immunoassay that simultaneously quantifies seven biomarkers associated with micronutrient (MN) deficiencies, inflammation and malarial antigenemia using plasma or serum; alpha-1-acid glycoprotein, C-reactive protein, ferritin, histidine-rich protein 2, retinol binding protein 4, soluble transferrin receptor, and thyroglobulin. Validations included repeated testing (n = 20 separately prepared experiments on 10 assay plates) in a single lab to assess precision and linearity. Seven independent laboratories tested 76 identical heparin plasma samples collected from a cohort of pregnant women in Niger using the same 7-plex assay to assess differences in results across laboratories. In the analytical validation experiments, intra- and inter-assay coefficients of variation were acceptable at <6% and <15% respectively and assay linearity was 96% to 99% with the exception of ferritin, which had marginal performance in some tests. Cross-laboratory comparisons showed generally good agreement between laboratories in all analyte results for the panel of 76 plasma specimens, with Lin's concordance correlation coefficient values averaging ≥0.8 for all analytes. Excluding plates that would fail routine quality control (QC) standards, the inter-assay variation was acceptable for all analytes except sTfR, which had an average inter-assay coefficient of variation of ≥20%. This initial cross-laboratory study demonstrates that the 7-plex test protocol can be implemented by users with some experience in immunoassay methods, but familiarity with the multiplexed protocol was not essential.

Impact of biofortified maize consumption on serum carotenoid concentrations in Zambian children.

Biofortified maize, designed as an intervention strategy to prevent vitamin A deficiency, can provide upwards of 15 µg ß-carotene per g dry weight. Some varieties also have elevated concentrations of other carotenoids. We conducted a cluster randomized, controlled feeding trial in rural Zambia to test the impact of daily consumption of biofortified maize over a 6-month period on vitamin A status. Serum concentrations of retinol and carotenoids were assessed by high-performance liquid chromatography. Data on circulating carotenoids by intervention group in 679 children are reported here. As previously shown, consumption of this ß-carotene-rich maize significantly improved serum ß-carotene concentrations (0.273 vs. 0.147 µmol/L, p < 0.001, in this subset of children). Here we show significant increases in α-carotene, ß-cryptoxanthin, and zeaxanthin (p < 0.001). There was no impact on lutein or lycopene concentrations. Consumption of biofortified maize can have broader implications beyond the control of vitamin A deficiency (Trial registration: NCT01695148).

Assessment of coenzyme Q10 absorption using an in vitro digestion-Caco-2 cell model.

The feasibility of using a coupled in vitro digestion-Caco-2 cell uptake as a model for examining the digestive stability and absorption of coenzyme Q10 (CoQ10) from a variety of commercially available CoQ10 products was examined. The products were first subjected to simulated digestion to mimic their passage through the GI tract to generate micelles containing CoQ10, and the micelle fractions added to monolayers of Caco-2 cells to determine CoQ10 uptake. The data demonstrate enhanced uptake of CoQ10 from formulations containing solubilized forms of CoQ10 and also from a CoQ10-gamma-cyclodextrin complex as compared with pure CoQ10 powder or tablets based on CoQ10 powder. The CoQ10 uptake by the cells was correlated with the extent of micellarization of CoQ10 during simulated digestion. Most of CoQ10 taken up by the cells was converted to ubiquinol either during or following uptake. The data also indicate a correlation between in vitro dissolution of CoQ10 products and uptake of CoQ10 by Caco-2 cells. Thus, this study demonstrates the utility of coupled in vitro digestion-Caco-2 cell model as a cost-effective screening tool that will provide useful information for the optimal design of human trials to assess the bioavailability of CoQ10 and also other bioactive compounds.

Provitamin A-biofortified maize increases serum ß-carotene, but not retinol, in marginally nourished children: a cluster-randomized trial in rural Zambia.

BACKGROUND: Vitamin A deficiency remains a nutritional concern in sub-Saharan Africa. Conventionally bred maize hybrids with high provitamin A carotenoid concentrations may have the potential to improve vitamin A status in maize-consuming populations. OBJECTIVE: We evaluated the efficacy of regular provitamin A carotenoid-biofortified "orange" maizemeal (∼15 µg ß-carotene/g) consumption in improving vitamin A status and reducing vitamin A deficiency in children. DESIGN: This was a cluster-randomized controlled trial in the rural farming district of Mkushi, Zambia. All 4- to 8-y-old children in an ∼400-km(2) area were identified and grouped by proximity into clusters of ∼15-25 children. We randomly assigned clusters to 1) orange maizemeal (n = 25), 2) white maizemeal (n = 25), or 3) a parallel, nonintervention group (n = 14). Children in intervention clusters (n = 1024) received 200 g maizemeal for 6 d/wk over 6 mo; the maizemeal was prepared according to standardized recipes and served in cluster-level kitchens. Staff recorded attendance and leftovers. We collected venous blood before and after the intervention to measure serum retinol, ß-carotene, C-reactive protein, and α1-acid glycoprotein. RESULTS: Intervention groups were comparable at baseline, and vitamin A status was better than anticipated (12.1% deficient on the basis of serum retinol <0.7 µmol/L). Although attendance at meals did not differ (85%), median daily maize intake was higher in white (154 g/d) than in orange (142 g/d) maizemeal clusters. At follow-up, mean serum ß-carotene was 0.14 µmol/L (95% CI: 0.09, 0.20 µmol/L) higher in orange maizemeal clusters (P < 0.001), but mean serum retinol (1.00 ± 0.33 µmol/L overall) and deficiency prevalence (17.1% overall) did not differ between arms. CONCLUSION: In this marginally nourished population, regular biofortified maizemeal consumption increased serum ß-carotene concentrations but did not improve serum retinol. This trial was registered at clinicaltrials.gov as NCT01695148.

A cross-sectional analysis demonstrated the healthy volunteer effect in smokers.

OBJECTIVE: This cross-sectional descriptive analysis sought to determine if a healthy volunteer effect can be demonstrated among smokers selected to participate in a dietary intervention trial. STUDY DESIGN AND SETTING: Body mass index (BMI), body fat, physical activity, dietary intake, and plasma concentration of antioxidant nutrients and carotenoids were assessed cross-sectionally, at the time of enrollment into a dietary intervention trial, among 136 adult smokers. RESULTS: Mean BMI was below national age- and gender-specific averages as was prevalence of overweight and obesity. Physical activity was reported to average 15.4 h/wk. Compared with other sample populations of smokers, our smokers reported lower total fat and cholesterol intakes, higher vitamin C and beta-carotene intakes, and generally equal vitamin E intakes. Plasma ascorbic acid, alpha-tocopherol, alpha- and beta-carotene, and beta-cryptoxanthin concentrations were higher than those of smokers surveyed by NHANES III. CONCLUSION: These findings suggest that a "healthy volunteer effect" can be described among adult smokers. Future dietary intervention trials among smokers should cautiously estimate sample size because smokers electing to participate may report healthier dietary patterns than other smokers. Screening criteria regarding baseline micronutrient status of smokers should be evaluated given that intervention effects may be dependent on overall health status.

Circulating endogenous retinoic acid concentrations among participants enrolled in a randomized placebo-controlled clinical trial of retinyl palmitate.

Retinoids have been studied extensively for their chemopreventive properties. The biological activity of retinoids is acquired through their conversion to retinoic acid (RA). Characterization of endogenous circulating RA concentrations after supplementation with vitamin A over longer time periods has not been done previously. Our investigation was conducted to determine whether vitamin A (retinyl palmitate) supplementation significantly increases circulating RA concentrations of all-trans-, 9-cis-, and 13-cis-RA. Using plasma samples from 41 participants enrolled in a randomized clinical trial of placebo, 25,000, 50,000, or 75,000 IU supplemental retinyl palmitate daily, high-performance liquid chromatography analyses were conducted for concentrations of three RA isomers. Seven plasma samples were analyzed for each participant over a 16-month period. Based on an intention-to-treat analysis, results obtained using linear mixed models showed that supplementation with retinyl palmitate statistically significantly increased concentrations of all three RA isomers from baseline levels. This study suggests that supplementation with retinyl palmitate is an effective means to increase circulating all-trans, 9-cis-, and 13-cis-RA concentrations among humans.

Temporal variation and identification of factors associated with endogenous retinoic acid isomers in serum from Brazilian women.

OBJECTIVE: Retinoids (natural and synthetic derivatives of vitamin A) have cancer chemotherapeutic and chemopreventive activities. Retinoic acid (RA) treatment has been associated with significant regression of preneoplastic lesions. However, serious toxicity associated with some therapies has made long-term chemoprevention in healthy populations unfeasible. Recently, serum RA has been shown to increase in response to oral retinol (vitamin A) supplementation. Here, we assess the variability of circulating RA levels and the lifestyle, demographic, and nutritional factors that explain such variability. METHOD: Total RA concentration and the concentrations of RA isomers (all-trans-RA, 13-cis-RA, and 9-cis-RA) were measured by high-pressure liquid chromatography in serum samples obtained 4 months apart from 502 women participating in the Ludwig-McGill Cohort (Sao Paulo, Brazil). RESULTS: The relative abundance of the three RA isomers was similar for each visit (baseline and month 4), with 13-cis-RA having the highest concentrations followed by 9-cis-RA and all-trans-RA. The within-person variability of total RA and individual isomers was low. Using multivariate logistic regression models (upper tertile versus low/middle tertile of serum RA), we found that age, race, oral contraceptive use, total number of pregnancies, and season of initial blood draw were significantly associated with at least one endogenous RA isomer level. All endogenous RA isomers were positively associated with serum retinol, beta-carotene, and beta-cryptoxanthin levels. CONCLUSION: These results have implications for the design of future epidemiologic studies focused on assessing RA-disease association and intervention studies aimed at modulating RA levels.

Effect of dietary zeaxanthin on tissue distribution of zeaxanthin and lutein in quail.

PURPOSE: The xanthophyll carotenoids (lutein and zeaxanthin) are hypothesized to delay progression of age-related macular degeneration. The quail has a cone-dominant retina that accumulates carotenoids. The purpose of these experiments was to characterize the carotenoid composition of retina, serum, liver, and fat in quail and to determine whether dietary enrichment with zeaxanthin alters zeaxanthin or lutein concentrations in these tissues. METHODS: Quail were fed for 6 months with a commercial turkey diet (T group; n = 8), carotenoid-deficient diet (C- group; n = 8), or a carotenoid-deficient diet supplemented with 35 mg 3R,3'R-zeaxanthin per kilogram of food, (Z+ group; n = 8). Zeaxanthin was derived from Sphingobacterium multivorum (basonym Flavobacterium). Carotenoids in serum, retina, liver, and fat were analyzed by HPLC. RESULTS: As in the primate fovea, the retina accumulated zeaxanthin, lutein, and cryptoxanthin, and preferentially absorbed zeaxanthin (P < 0.005). In contrast, lutein was preferentially absorbed by liver (P < 0.01) and fat (P < 0.0001). In supplemented females, zeaxanthin increased approximately 4-fold in retina, and 74-, 63- and 22-fold in serum, liver, and fat, respectively. In males, zeaxanthin was elevated approximately 3-fold in retina, and 42-, 17-, and 12-fold in serum, liver, and fat, respectively. Birds fed the Z+ diet absorbed a higher fraction of dietary lutein into serum, but lutein was reduced in the retina (P < 0.05). CONCLUSIONS: Xanthophyll profiles in quail mimic those in primates. Dietary supplements of zeaxanthin effectively increased zeaxanthin concentrations in serum, retina, liver, and fat. The robust response to zeaxanthin supplementation identifies the quail as an animal model for exploration of factors regulating delivery of dietary carotenoids to the retina.

Evaluation of Serum Volume Losses During Long-Term Storage.

Aliquots of serum collected in a large case-control study of cervical cancer were stored at -70°C for up to 4 years during implementation of the study. When 500 µL serum aliquots were thawed in preparation for carotenoid and vitamin A assays, volumes were noticeably variable and fell below 500 µL in the majority of the samples. We were concerned about evaporation/sublimation during storage of the samples because loss of water would concentrate the analytes of interest. We evaluated the use of density and sodium ion concentration measurements to confirm its occurrence. We found that serum density was an unreliable indicator of extent of volume loss since the anticipated increases in density due to evaporation were of the same magnitude as inter-individual variation in serum density. In contrast, Na+ concentration is tightly regulated and would rise if water had been lost from the samples. In a representative sample of serum aliquots from the case-control study, 24 of 25 vials contained less than 500 µL of serum. The mean sodium ion concentration (138.1 ± 3.6 mmol/L) was within the normal range for human serum of 136-145 mmol/L, and no correlation was observed between serum volume and Na+ concentration. These results strongly suggest that the observed low volumes were not due to evaporative losses. Instead, the variably low volumes of serum aliquots were probably due to pipetting errors in the initial aliquotting resulting from the use of air-displacement pipettes.

Performance of the CRAFTi portable fluorometer comparing with the HPLC method for determining serum retinol.

OBJECTIVE: To determine the performance of a portable fluorometer for measuring serum retinol (SR) concentration. DESIGN AND METHODS: Serum samples were obtained from 75 factory worker women and 143 school children. SR concentration was quantified using a portable fluorometer ('CRAFTi') and HPLC analysis. RESULTS: SR by HPLC (1.23 ± 0.43 µmol/L) and CRAFTi (1.16 ± 0.46 µmol/L) was significantly correlated. Sensitivity and specificity were 85.3% and 78.0% (cutoff of 1.05 µmol/L). Kappa statistics showed moderate agreement. CONCLUSIONS: CRAFTi portable fluorometer is a promising field-friendly tool for screening vitamin A deficiency.