[A survey on the current situation of serum vitamin A and vitamin D levels among children aged 2-<7 years of 20 cities in China].

Objective: To investigate serum vitamin A and vitamin D status in children aged 2-<7 years in 20 cities in China. Methods: A cross-sectional study was conducted. A total of 2 924 healthy children aged 2-<7 years were recruited from September 2018 to September 2019 from 20 cities in China, categorized by age groups of 2-<3 years, 3-<5 years, and 5-<7 years. The demographic and economic characteristics and health-related information of the enrolled children were investigated. Body weight and height were measured by professional staff members. The serum vitamin A and vitamin D levels were detected by high-performance liquid chromatography-tandem mass spectrometry. Chi-square test and Logistic regression were applied to analyze the association between vitamin A and vitamin D deficiency and insufficiency as well as their underlying impact factors. Results: The age of the 2 924 enrolled children was 4.33 (3.42, 5.17) years. There were 1 726 males (59.03%) and 1 198 females (40.97%). The prevalences of vitamin A and vitamin D deficiency in enrolled children were 2.19% (64/2 924) and 3.52% (103/2 924), respectively, and the insufficiency rates were 29.27% (856/2 924) and 22.20% (649/2 924), respectively. Children with both vitamin A and vitamin D deficiencies or insufficiencies were found in 10.50% (307/2 924) of cases. Both vitamin A (χ2=7.91 and 8.06, both P=0.005) and vitamin D (χ2=71.35 and 115.10, both P<0.001) insufficiency rates were higher in children aged 3-<5 and 5-<7 years than those in children aged 2-<3 years. Vitamin A and vitamin D supplementation in the last 3 months was a protective factor for vitamin A and D deficiency and insufficiency, respectively (OR=0.68 and 0.22, 95%CI 0.49-0.95 and 0.13-0.40, both P<0.05). The rates of vitamin A and D insufficiency was higher in children with annual household incomes <60 000 RMB than in those with annual household incomes ≥60 000 RMB (χ2=34.11 and 10.43, both P<0.01). Northwest and Southwest had the highest rates of vitamin A and vitamin D insufficiency in children aged 2-<7 yeas, respectively (χ2=93.22 and 202.54, both P<0.001). Conclusions: Among 20 cities in China, children aged 2-<7 years experience high rates of vitamin A and vitamin D insufficiency, which are affected by age, family economic level, vitamin A and vitamin D supplementation, and regional economic level. The current results suggest that high level of attention should be paid to vitamin A and vitamin D nutritional status of preschool children.

Combination of a solid phase extraction and a two-dimensional LC-UV method for the analysis of vitamin D3 and its isomers in olive oil.

The current HPLC methods for the quantification of vitamin D3 (VitD3) and its two isomers previtamin D3 (PreVitD3) and trans-vitamin D3 (trans-VitD3) in olive oil preparations present some limitations mainly due to peak overlapping of the oily matrix components with the compounds of interest. The use of two-dimensional liquid chromatography (2D-LC) with different retention mechanism can reach higher resolving power thus allowing the analysis of complex samples. The present paper proposes a new alternative method including a solid phase extraction sample preparation step and a two-dimensional liquid chromatographic analysis using routine instrumentation, fitting the needs of quality assurance and quality control laboratories of pharmaceutical companies. The extraction protocol was demonstrated to provide a clean-up of the sample and a quantitative recovery of the species of interest. The 2D method proved its suitability in the isolation of vitamins from oil components in the first dimension and the separation and quantification of the analytes in the second dimension thanks to the orthogonal selectivities of phenyl and porous graphitic carbon (PGC) stationary phases. The method was validated following ICH guidelines and possesses an adequate sensitivity to quantify the impurity trans-VitD3 in pharmaceuticals considering the limits imposed by regulatory agencies. The applicability of the phenyl x PGC 2D-LC-UV method to quality control of medicinal products based on VitD3 in olive oil was confirmed by the successful quantification of vitamins in olive oil formulations.

A robust method for simultaneous determination of eight B vitamins in human serum by liquid chromatography tandem mass spectrometry.

The level of vitamin B group in human serum is an important index of human health. Among B vitamins, cyanocobalamin in serum is unstable and its content is extremely low. Rapid and simultaneous detection of multiple B vitamins including cyanocobalamin is a challenge. Herein, we have developed a rapid and stable method that can realize the determination of thiamine, riboflavin, nicotinamide, pantothenic acid, pyridoxic acid, biotin, 5-methyltetrahydrofolate, and cyanocobalamin simultaneously in 6 min. The method was established based on protein precipitation with methanol and then chromatographic separation was achieved using Waters acquity ultra-high-performance liquid chromatography high strength silica T3 column, which was stable and sensitive especially for cyanocobalamin. Limit of quantification, precision, trueness, and matrix effect were validated according to the European Medicines Agency and United States Food and Drug guidelines and Clinical and Laboratory Standards Institute guidelines on bioanalytical method. The limit of quantification for thiamine, riboflavin, nicotinamide, pantothenic acid, pyridoxic acid, biotin, 5-methyltetrahydrofolate, and cyanocobalamin was 0.4, 0.4, 0.8, 2.0, 0.4, 0.1, 0.4, and 0.04 ng/mL separately, respectively. Intra- and interday precisions were 1.1%-12.4% and 2.0%-13.5%, respectively. The relative errors were between 0.3% and 13.3%, and the matrix effects were between 2.6% and 10.4%.

Determination of N-nitroso folic acid in folic acid and multivitamin supplements by LC-MS/MS.

Analysis of N-nitroso folic acid, a nitrosamine impurity found in folic acid, is challenging due to the complex sample matrices. Many of such supplements contain not only a variety of vitamins, including vitamin A, Bs, C, D and E, but also other ingredients such as minerals, docosahexaenoic acid (DHA), glucose syrup, sugar, and herbs. On the other hand, the strength of folic acid is typically low, ranging from 50 µg to 5 mg per unit. In this study, a highly selective and sensitive LC-MS/MS method was developed to accurately quantify N-nitroso folic acid in supplements containing folic acid. The sample was extracted by 0.1% ammonia solution: MeOH (9:1, v/v) containing 5 ng/mL of N-nitroso folic acid-d4 (Isotope internal standard). The quantification was performed by MRM in negative ionization mode. Mobile phases A and B were 0.1% formic acid in deionized water and methanol, respectively. The method was validated and found to have sufficient linearity (R2 > 0.995), accuracy (recovery 83-110%), precision (RSD 3%) and low LOD, LOQ (4 and 10 µg/g respectively, with respect to folic acid). The method was applied to the determination of N-nitroso folic acid in 40 supplements containing folic acid with different strengths and formulation. The content of N-nitroso folic acid was found to be up to 898 ng/unit (1794 µg/g with respect to folic acid). It enabled regulatory actions, such as product recall, to safeguard public health from unsafe products.

Dietary habits in Japanese patients with bullous pemphigoid: low intake of retinol.

BACKGROUND: Bullous pemphigoid (BP) is an autoimmune subepidermal blistering disease. Dietary habits may modulate the pathogenesis of BP. OBJECTIVES: We evaluated dietary habits in Japanese patients with BP and compared their results to those of age- and sex-matched healthy controls. We also examined the relationship between dietary habits versus IgG anti-BP180NC16A antibody or parameters of BP disease area index (BPDAI); cutaneous blisters/erosions, cutaneous urticaria/erythema, and mucosal blisters/erosions. MATERIALS & METHODS: Dietary habits were assessed by the validated, Brief-type self-administered Diet History Questionnaire. Severity of disease was assessed with BPDAI. RESULTS: Patients with BP showed a lower intake of retinol (vitamin A1) and beverages, and a higher intake of seasoning/spices, compared to controls. The bivariate and multivariable logistic regression analysis showed that BP was associated with a low intake of retinol and beverages. There were no significant correlations between IgG anti-BP180NC16A antibody levels and intake of nutrients/foods. The BPDAI score for cutaneous blisters/erosions significantly positively correlated with intake of carbohydrate and negatively with intake of retinol, vitamin A, animal fat, cholesterol, phosphorus, and vitamin B2. The BPDAI score for cutaneous urticaria/erythema significantly negatively correlated with intake of vitamin A. BP patients with mucosal blisters/erosions had a higher intake of cholesterol, n-6 polyunsaturated fatty acid, and eggs, and lower intake of seasoning/spices, compared to patients without BP. CONCLUSION: The supplementation of vitamin A might have prophylactic and/or therapeutic effects on BP.

Rapid simultaneous determination of 7 fat-soluble vitamins in human serum by ultra high performance liquid chromatography tandem mass spectrometry.

Accurate detection of vitamins is critically important for clinical diagnosis, metabolomics and epidemiological studies. However, the amounts of different vitamins vary dramatically in human serum. It is a challenge to achieve simultaneous detection of multiple vitamins rapidly. Herein, we developed and validated a sensitive and specific method using ultra high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) for simultaneous quantification of 7 fat-soluble vitamins (FSVs) across their physiological concentrations in serum for the first time, which was subjected to protein precipitation, liquid-liquid extraction to an organic phase, evaporation to dryness and reconstitution with acetonitrile. In the present procedure, retinol (vitamin A), ergocalciferol (25-OH-D2), cholecalciferol (25-OH-D3), α-tocopherol (vitamin E), phylloquinone (vitamin K1), menatetrenone-4 (MK-4), and menaquinone-7 (MK-7) were detected in one analytical procedure for the first time within 5.0 min by triple quadrupole tandem mass spectrometry. The limit of quantification (LOQ) for vitamin A was 10.0 ng mL-1, LOQs for 25-OH-D2 and 25-OH-D3 were 1.0 ng mL-1, LOQ for vitamin E was 100.0 ng mL-1, and LOQs for vitamin K1, MK-4 and MK-7 were 0.10 ng mL-1, respectively, with a correlation (R2) of 0.995-0.999. Recoveries ranged from 80.5% to 118.5% and the intra-day and inter-day coefficients of variance (CVs) were 0.72-8.89% and 3.2-9.0% respectively. The method was validated according to the European Medicines Agency (EMA) and U.S. Food and Drug guidelines and C62-A on bioanalytical methods, and was used for clinical routine determination.

Wavelength-tailored light-emitting diodes reduce damage to sensory properties of light-exposed milk.

Photooxidation has long been affecting nutrient and sensory quality of fluid milk. Light oxidation starts from the activation of photosensitive compounds, followed by generation of singlet oxygen that reacts with vitamins, proteins, and lipids in milk. It is hypothesized that wavelength-tailored light schemes possessing spectral properties capable of avoiding excitation maxima of common photosensitizers in milk could slow the chemical degradation of light-exposed milk and thus preserve consumer acceptability. A series of 6 consumer tests with sample sizes from 95 to 119 participants tested hedonic responses to fluid milk samples exposed to light of varying wavelength spectra. For milk in clear plastic bottles (polyethylene terephthalate or high-density polyethylene), consumer panels generally liked milk exposed to light-emitting diodes eliminating wavelengths below 520 or 560 nm more than standard white light, or those eliminating other wavelength bands. This higher degree of liking coincided with panelists citing fewer off-flavors or aromas from these samples. Taken together, these observations suggest such light schemes can protect milk from light damage to some extent. Wavelength-tailored light schemes used in this study did not offer effective protection for milk in glass bottles. Dissolved oxygen, color, riboflavin loss, and hexanal content were instrumentally evaluated, but results failed to indicate significant signatures of light damage in milk compared with sensory measures. The appearance of milk bottles illuminated by the slightly greenish or yellowish light were less liked by consumers, suggesting further efforts on consumer education may be necessary if these light schemes were to be installed in retail dairy coolers.

Transcriptome and metabolome profiling identify factors potentially involved in pro-vitamin A accumulation in cassava landraces.

Cassava (Manihot esculenta Crantz) is a predominant food security crop in several developing countries. Its storage roots, rich in carbohydrate, are deficient in essential micronutrients, including provitamin A carotenoids. Increasing carotenoid content in cassava storage roots is important to reduce the incidence of vitamin A deficiency, a public health problem in sub-Saharan Africa. However, cassava improvement advances slowly, mainly due to limited information on the molecular factors influencing ß-carotene accumulation in cassava. To address this problem, we performed comparative transcriptomic and untargeted metabolic analyses of roots and leaves of eleven African cassava landraces ranging from white to deep yellow colour, to uncover regulators of carotenoid biosynthesis and accumulation with conserved function in yellow cassava roots. Sequence analysis confirmed the presence of a mutation, known to influence ß-carotene content, in PSY transcripts of deep yellow but not of pale yellow genotypes. We identified genes and metabolites with expression and accumulation levels significantly associated with ß-carotene content. Particularly an increased activity of the abscisic acid catabolism pathway together with a reduced amount of L-carnitine, may be related to the carotenoid pathway flux, higher in yellow than in white storage roots. In fact, NCED_3.1 was specifically expressed at a lower level in all yellow genotypes suggesting that it could be a potential target for increasing carotenoid accumulation in cassava. These results expand the knowledge on metabolite compositions and molecular mechanisms influencing carotenoid biosynthesis and accumulation in cassava and provide novel information for biotechnological applications and genetic improvement of cassava with high nutritional values.

Effects of supplementation of tropical fruit processing by-products on lipid profile, retinol levels and intestinal function in Wistar rats.

Fruits agro-industrial by-products may have a great variety of bioactive compounds that promote health. Thus, the effects of supplementation with acerola, cashew and guava processing by-products for 28 days on retinol level, lipid profile and on some aspects related to intestinal function in rats were investigated. The animals supplemented with different fruit by-products presented similar weight gain, faecal pH values and intestinal epithelial structures; however, they showed higher moisture and Lactobacillus spp. and Bifidobacterium spp. counts in faeces compared to the control group. Supplementation with the cashew by-product decreased the blood glucose, acerola and guava by-products reduced serum lipid levels and all fruit by-products tested increased serum and hepatic retinol. The results indicated that acerola and guava by-products possess a potential hypolipidemic effect. The three fruit by-products increase the hepatic retinol deposition and the faecal populations of beneficial bacterial groups and modulated aspects of intestinal function. The findings of this study can contribute to sustainable fruticulture and support future clinical studies with the supplementation of by-products.

Quantification of 10 B vitamins in mouse colon by LC-MS/MS: Application on breast cancer mice treated with doxorubicin.

B vitamins play important roles in various physiological processes, including cell metabolism and DNA synthesis. The intestine is critical for the absorption and utilization of B vitamins, but few analytical methods for detecting intestinal B vitamins are currently available. In this study, we developed a novel liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous quantification of 10 B vitamins in mouse colon tissue, including thiamin (B1), riboflavin (B2), nicotinic acid (B3), niacinamide (B3-AM), pantothenic acid (B5), pyridoxine (B6), pyridoxal 5'-phosphate (B6-5P), biotin (B7), folic acid (B9), and cyanocobalamin (B12). The method was thoroughly validated following the U.S. Food and Drug Administration (FDA) guidelines and yielded good results in terms of linearity (r2 > 0.9928), lower limit of quantification (40-600 ng/g), accuracy (88.9-119.80 %) and precision (relative standard deviation ≤ 19.71 %), recovery (87.95-113.79 %), matrix effect (91.26-113.78 %), and stability (85.65-114.05 %). Furthermore, we applied our method to profile B vitamins in the colons of mice with breast cancer after doxorubicin chemotherapy treatment, which revealed that the doxorubicin treatment led to significant colon damage and accumulation of several B vitamins including B1, B2 and B5. We also confirmed the capability of this method for quantifying B vitamins in other intestinal tissues like the ileum, jejunum, and duodenum. The newly developed method is simple, specific, and useful for targeted profiling of B vitamins in mouse colon, with a potential for future studies on the role of these micronutrients in healthy and diseased states.

A Miniaturized Sample Preparation Method for the Determination of Vitamins A and E in Food Products.

A new analytical approach to the simultaneous identification and quantification of vitamins A and E in three representative matrices (Parmesan, spinach, and almonds) was developed. The analyses were based on high-performance liquid chromatography with UV-VIS/DAD detection. The procedure was optimized by a significant reduction in the weight of the tested products and quantities of reagents added during the saponification and extraction stages. A full method validation study was performed for retinol at two concentration levels (LOQ and 200 × LOQ), which showed satisfactory results, with recoveries ranging from 98.8 to 110.1%, and an average CV of 8.9%. Linearity was tested in the range of 1-500 µg/mL and showed the coefficient of determination R2 = 0.999. The satisfactory recovery and precision parameters were achieved for α-tocopherol (LOQ and 500 × LOQ) in the range of 70.6-143.2%, with a mean CV equal to 6.5%. The observed linearity for this analyte in the concentration range of 1.06-532.0 µg/mL was R2 = 0.999. The average extended uncertainties were estimated, using a top-down approach of 15.9% and 17.6% for vitamin E and A, respectively. Finally, the method was successfully applied to determine vitamins in 15 commercial samples.

Recovery of Polyphenolic Compounds and Vitamins from the Stinging Nettle Leaves: Thermal and Behavior and Biological Activity of Obtained Extracts.

Stinging nettle (SN) is an extraordinary plant from the Urticaceae botanical family. It is well-known and widely used in food and folk medicine to treat different disorders and diseases. This article aimed to study the chemical composition of SN leaves extracts, i.e., polyphenolic compounds and vitamins B and C, because many studies ascribed high biological potency to these compounds and their significance in the human diet. Besides the chemical profile, the thermal properties of the extracts were studied. The results confirmed presence of many polyphenolic compounds and vitamins B and C. It also showed that the chemical profile closely correlated with the applied extraction technique. The thermal analysis showed that analyzed samples were thermally stable up to about 160 °C. Thermal degradation of samples UAE, MAE, and MAC took place in four steps, and sample SE in three steps. Altogether, results confirmed the presence of health-beneficial compounds in stinging nettle leaves and indicated the possible application of its extract in pharmaceutical and food industries as both a medicinal and food additive.

From mouse to human: Accessing the biochemistry of vision in vivo by two-photon excitation.

The eye is an ideal organ for imaging by a multi-photon excitation approach, because ocular tissues such as the sclera, cornea, lens and neurosensory retina, are highly transparent to infrared (IR) light. The interface between the retina and the retinal pigment epithelium (RPE) is especially informative, because it reflects the health of the visual (retinoid) cycle and its changes in response to external stress, genetic manipulations, and drug treatments. Vitamin A-derived retinoids, like retinyl esters, are natural fluorophores that respond to multi-photon excitation with near IR light, bypassing the filter-like properties of the cornea, lens, and macular pigments. Also, during natural aging some retinoids form bisretinoids, like diretinoid-pyridiniumethanolamine (A2E), that are highly fluorescent. These bisretinoids appear to be elevated concurrently with aging. Vitamin A-derived retinoids and bisretinoidss are detected by two-photon ophthalmoscopy (2PO), using a new class of light sources with adjustable spatial, temporal, and spectral properties. Furthermore, the two-photon (2P) absorption of IR light by the visual pigments in rod and cone photoreceptors can initiate visual transduction by cis-trans isomerization of retinal, enabling parallel functional studies. Recently we overcame concerns about safety, data interpretation and complexity of the 2P-based instrumentation, the major roadblocks toward advancing this modality to the clinic. These imaging and retina-function assessment advancements have enabled us to conduct the first 2P studies with humans.

Development and application of a SFC-DAD-MS/MS method to determine carotenoids and vitamin A in egg yolks from laying hens supplemented with ß-carotene.

ß-Carotene, a provitamin A carotenoid, can be converted into vitamin A in animals' bodies, and can also be accumulated intactly in many animal products. In this study, supercritical fluid chromatography-tandem mass spectrometry was utilized to determine ß-carotene and different forms of vitamin A in eggs simultaneously. According to the results, ß-carotene contained in yolk reached a plateau after about 2 weeks of supplementation. With an increase in dietary supplement level, the amount of ß-carotene gradually increased, as well as slightly changing the yolk color. Moreover, the contents of retinoids including retinol, retinyl propionate, retinyl palmitate and retinyl stearate were also elevated in yolks with the ß-carotene additive levels; meanwhile, the lutein and zeaxanthin decreased. On the whole, ß-carotene in the diet of laying hens could be partially deposited in egg yolk, and the contents of vitamin A in yolk could be increased due to ß-carotene bioconversion.

Low dietary protein increases vitamin A absorption and deposition into milk in periparturient rats.

Our objective was to study the effect of differing dietary crude protein and vitamin A on retinoid metabolism in a periparturient rat model. Sixty female rats, approximately 21 d before parturition, were fed rations containing either low protein (13%; LP) or high protein (22%; HP) crude protein and either low vitamin A (3 IU/g; LA) or high vitamin A (5 IU/g; HA), yielding treatments HPHA, HPLA, LPHA, and LPLA. Samples were collected at d -14, d +3, and +10 relative to parturition and analyzed for all-trans retinoid acid (RA), 13-Cis RA, and retinol. At d -14, serum all-trans RA concentrations decreased compared to baseline. At both d +3 and d +10, serum retinol increased and liver 13-Cis RA decreased. In the small intestine, 13-cis RA was higher in HPHA than HPLA pre-partum (0.93±0.12 vs. 0.40±0.12 ng/ml, P=0.04). Post-partum, 13-cis RA was lower in high vitamin HPHA and LPHA groups (0.35±0.06 and 0.38±0.06 ng/ml) than in low vitamin A HPLA and LPLA treatments (0.50±0.06 and 1.32±0.06 ng/ml, P<0.01). In rats fed LA diets, TNF-alpha expression tended to be lower in HPLA than LPLA groups on day +3 (0.69±0.34 vs 1.00±0.52, P=0.08), but not day +10 (0.56±0.25 vs. 1.00±0.49 fold change, P>0.10). Retinoids accumulated during pregnancy and were mobilized during lactation. The sequestration of retinoids was increased when dietary protein content was low. Further studies are needed to investigate how retinoid metabolism could be manipulated to improve vitamin A delivery to milk.

A simple method for analysis of vitamin A palmitate in fortified cereal products using direct solvent extraction followed by reversed-phase HPLC with UV detection.

Vitamin A is generally analysed using a time-consuming and possibly detrimental saponification step, followed by extraction and HPLC analysis. We here developed a new method to analyse retinyl palmitate (RP) (also known as vitamin A palmitate) without the need for saponification and validated it in model systems consisting of RP, soy oil and wheat bran, and in RP-fortified cereal products. Two direct solvent extraction protocols using acetone/methanol (7/3, v/v) or chloroform/methanol (1/1, v/v) were tested. After extraction, RP was quantified by reversed-phase HPLC with UV detection. The HPLC method had low limits of detection (0.01 µg/mL) and quantification (0.03 µg/mL). Both extraction protocols showed a good recovery (88-105 %) and intra-and inter-day precision (<5%) for RP extraction from the model systems. The obtained results corresponded to results obtained using a golden standard approach. For complex food matrices like bread and cookies, the chloroform/methanol extraction protocol showed the best performance characteristics.

Exploring the unexplored: A characterization of vitamins and vitamers in white grape musts by high-performance liquid chromatography.

Although prime compounds in yeast metabolism, vitamins in oenology have remained mostly unexplored for decades. Here, a premier characterization of the vitamers in white grape musts has been drawn. A RP-HPLC method has therefore been developed for their direct analysis in musts, allowing for the determination of 19 different vitamers from 8 water-soluble vitaminic groups, including thiamine forms T, TMP and TPP, with LODs between 0.1 and 45.9 µg.L-1 and LOQs between 0.4 and 137.8 µg.L-1. A resulting characterization of 85 grape musts has been drawn from their vitaminic composition. Plus, the use of neither sulfites nor filtration affects the must vitamin content. The method stands as a useful tool for the later determination of yeast requirements, or impact of winemaking products on vitamins. The method has, overall, proven as practical and sensitive, for rapid identification of vitamins and vitamers in musts.

Biotechnological interventions in reducing losses of tropical fruits and vegetables.

Tropical fruits and vegetables are predominantly cultivated in warm climate zones, resulting in cultivar diversity in terms of structure, features, and physiology. These constitute a variety of bioactive ingredients such as vitamins, minerals, phenolic acids, anthocyanins, flavonoids, fatty acids, fiber, and their distinctive appearances attract customers across the world. The global production of fruit and vegetables has been attained a tremendous increase for the last few decades. However, huge losses at pre- and postharvest levels are major constraints in their judicious use. Traditional breeding strategies were used to minimize these losses, but their functionality is limited due to their time and labor intensiveness. Recent biotechnological, computational, and multiomics approaches not only address the losses concern but also aid in boosting crop productivity and nutritional values. This article emphasizes molecular tools that have been used to reduce losses of tropical fruits and vegetables at pre- and postharvest levels.

A nutritional supplement during preconception and pregnancy increases human milk vitamin D but not B-vitamin concentrations.

BACKGROUND & AIMS: Optimal maternal vitamin status during pregnancy and lactation is essential to support maternal and infant health. For instance, vitamin D3 is involved in infant bone development, and B-vitamins are involved in various metabolic processes, including energy production. Through a double-blind randomised controlled trial, we investigated the effects of maternal supplementation from preconception throughout pregnancy until birth on human milk (HM) concentrations of vitamin D3 and B-vitamins. In addition, we aimed to characterise longitudinal changes in milk concentrations of these vitamins. METHODS: Both control and intervention supplements contained calcium, iodine, iron, ß-carotene, and folic acid, while the intervention also contained zinc, vitamins B2, B6, B12, and D3, probiotics, and myo-inositol. HM samples were collected across 4 time points from 1 week to 3 months post-delivery from 158 mothers in Singapore, and 7 time points from 1 week to 12 months from 180 mothers in New Zealand. HM vitamin D was quantified using supercritical fluid chromatography and B-vitamins with mass spectrometry. Potential intervention effects on HM vitamins D3, B2, B6, and B9, as well as other B-vitamin (B1 and B3) concentrations were assessed using linear mixed models with a repeated measures design. RESULTS: Over the first 3 months of lactation, HM 25-hydroxyvitamin D3 concentrations were 20% (95% CI 8%, 33%, P = 0.001) higher in the intervention group, with more marked effects in New Zealand. There were no observed intervention effects on HM concentrations of vitamins B1, B2, B3, B6, and B9. In New Zealand mothers, longitudinally, vitamin D3 concentrations gradually increased from early lactation up to 12 months, while vitamins B1 and B2 peaked at 6 weeks, B3 at 3 weeks, and B6 and B9 at 3 months. CONCLUSIONS: Maternal supplementation during preconception and pregnancy increased HM vitamin D, but not B-vitamin concentrations in lactation. Further studies are required to examine the discrete benefits of vitamin D supplementation starting preconception vs during pregnancy, and to further characterise the effects of supplementation on later offspring health outcomes. CLINICAL TRIAL REGISTRATION: Registered at ClinicalTrials.gov on the 16 July 2015 (identifier NCT02509988); Universal Trial Number U1111-1171-8056. This study was academic-led by the EpiGen Global Research Consortium.

Using follicular fluid metabolomics to investigate the association between air pollution and oocyte quality.

BACKGROUND AND AIM: Our objective was to use metabolomics in a toxicological-relevant target tissue to gain insight into the biological processes that may underlie the negative association between air pollution exposure and oocyte quality. METHODS: Our study included 125 women undergoing in vitro fertilization at an academic fertility center in Massachusetts, US (2005-2015). A follicular fluid sample was collected during oocyte retrieval and untargeted metabolic profiling was conducted using liquid chromatography with ultra-high-resolution mass spectrometry and two chromatography columns (C18 and HILIC). Daily exposure to nitrogen dioxide (NO2), ozone, fine particulate matter, and black carbon was estimated at the women's residence using spatiotemporal models and averaged over the period of ovarian stimulation (2-weeks). Multivariable linear regression models were used to evaluate the associations between the air pollutants, number of mature oocytes, and metabolic feature intensities. A meet-in-the-middle approach was used to identify overlapping features and metabolic pathways. RESULTS: Of the air pollutants, NO2 exposure had the largest number of overlapping metabolites (C18: 105; HILIC: 91) and biological pathways (C18: 3; HILIC: 6) with number of mature oocytes. Key pathways of overlap included vitamin D3 metabolism (both columns), bile acid biosynthesis (both columns), C21-steroid hormone metabolism (HILIC), androgen and estrogen metabolism (HILIC), vitamin A metabolism (HILIC), carnitine shuttle (HILIC), and prostaglandin formation (C18). Three overlapping metabolites were confirmed with level-1 or level-2 evidence. For example, hypoxanthine, a metabolite that protects against oxidant-induced cell injury, was positively associated with NO2 exposure and negatively associated with number of mature oocytes. Minimal overlap was observed between the other pollutants and the number of mature oocytes. CONCLUSIONS: Higher exposure to NO2 during ovarian stimulation was associated with many metabolites and biologic pathways involved in endogenous vitamin metabolism, hormone synthesis, and oxidative stress that may mediate the observed associations with lower oocyte quality.