BACKGROUND: Previous studies showed that pre- and probiotics may enhance iron absorption. Probiotics combined with prebiotics (synbiotics), including human-identical milk oligosaccharides (HiMOs), are commonly added to infant and follow-up formula (FUF). Whether these additions enhance iron absorption from iron-fortified commercial milk formula is uncertain.
OBJECTIVE: We determined the effect of adding 1) a synbiotic [galacto-oligosaccharides [GOSs] + Limosilactobacillus reuteri (L. reuteri)] or 2) the HiMO 2\'-fucosyllactose (2\'FL) to iron-fortified FUF on iron absorption in young Thai children.
METHODS: In a randomized, controlled, single-blinded (participants) crossover study, 82 Thai children aged 8-14 mo were enrolled to consume single servings (235 mL) of FUF with isotopically labeled ferrous sulfate (2.2 mg iron) with 1) the synbiotic (400 mg/100 mL GOS and L. reuteri DSM 17938), 2) the 2\'FL (100 mg/100 mL), and 3) without synbiotic and 2\'FL (control) in random order and a 3-d washout period between administrations. Fractional iron absorption [FIA (%)] was assessed by measuring erythrocyte incorporation of isotopic labels 14 d (n = 26) and 28 d (n = 76) after consumption of the last test FUF.
RESULTS: Median (IQR) FIA from iron-fortified FUF with the synbiotic [8.2 (5.2, 12.9)%] and with 2\'FL [8.4 (5.5, 14.1)%] did not differ from the control FUF [8.1 (4.8,14.7)%] (synbiotic compared with control, P = 0.24; 2\'FL compared with control, P = 0.95). FIA from all FUF did not differ when measured after 14 and 28 d of erythrocyte incorporation (Time, P = 0.368; FUF, P = 0.435; Time × FUF, P = 0.937). Fecal pH and hemoglobin were negatively associated with FIA.
CONCLUSIONS: In young Thai children, the addition of a synbiotic (GOS + L. reuteri) or 2\'FL to iron-fortified FUF did not impact FIA from a single serving. The study was registered at clinicaltrials.gov as NCT04774016.

Obtaining valid estimates of nutrient intake in infants is currently limited by the difficulties of accurately measuring human milk intake. Current methods are either unsuitable for large-scale studies (i.e., the gold standard dose-to-mother stable isotope technique) or use set amounts, regardless of known variability in individual intake.
This cross-sectional study aimed to develop equations to predict human milk intake using simple measures and to carry out external validation of existing methods against the gold standard technique.
Data on human milk intake were obtained using the dose-to-mother stable isotope technique in 157 infants aged 7-10 mo and their mothers. Predictive equations were developed using questionnaire and anthropometric data (Model 1) and additional dietary data (Model 2) using lasso regression. Bland-Altman plots and intraclass correlation coefficients (ICC) also assessed the validity of existing methods (FITS and ALSPAC studies).
The strongest univariate predictors of human milk intake in infants of 8.3 mo on average (46% female) were infant age, infant body mass index (BMI), number of breastfeeds a day, infant formula consumption, and energy from complementary food intake. Mean [95% confidence interval (CI)] differences in predicted versus measured human milk intake [mean (SD): 762 (257) mL/day] were 0.0 mL/day (-26, 26) for Model 1 (ICC 0.74) and 0.5 mL/day (-21, 22) for Model 2 (ICC 0.83). Corresponding differences were -197 mL/day (-233, -161; ICC 0.32) and -175 mL/day (-216, -134; ICC 0.41) for the methods used by FITS and ALSPAC, respectively.
The Human Milk Intake Level Calculation provides substantial improvements on existing methods to estimate human milk intake in infants aged 7-10 mo, while utilizing data commonly collected in nutrition surveys. Although further validation in an external sample is recommended, these equations can be used to estimate human milk intake at this age with some confidence. This clinical trial was registered at http://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=379436) as ACTRN12620000459921.

Although extensive research has been carried out on the occurrence of mercury (Hg) in biota, bioaccumulation and tissue distribution of Hg in songbirds have not been well characterized. In the present study, Hg was investigated in insects and barn swallows (Hirundo rustica) to explore the bioaccumulation characteristics of Hg. Hg in swallow feathers and tissues including muscle, liver, and bone was investigated to determine the tissue distribution of Hg. The concentrations of Hg were 1.39 ± 1.01 μg/g, 0.33 ± 0.09 μg/g, 0.47 ± 0.10 μg/g, and 0.23 ± 0.09 μg/g in feather, muscle, liver, and bone samples, respectively. The trophic magnification factor of Hg in swallows and insects was higher than 1. However, the Hg concentrations in swallow feathers were not significantly correlated with stable isotope values of carbon or nitrogen, which implies the complex food sources and exposure processes of Hg for swallows. Feathers had significantly higher concentrations of Hg than liver, muscle, and bone samples (p < 0.01 for all comparisons). Feather, muscle, bone, and other organs had fractions of 64.4 ± 11.9%, 6.07 ± 2.06%, 20.0 ± 8.19%, and 9.56 ± 2.96% in total body burden of Hg in swallows. Hg in feathers contributed more than half of Hg in the whole body for most swallow individuals. Swallows may efficiently eliminate Hg by molting, and the excretion flux of Hg and other contaminants via molting deserves more investigation.

Protein recommendations for older adults are based on nitrogen balance data from young adults. Physiological studies using the indicator amino acid oxidation method suggest they need 30% to 50% more protein than current recommendations. We herein present glutathione (GSH) as a physiological estimate of protein adequacy in older adults.
The objective was to measure GSH kinetics in response to varying protein intakes in a repeated-measures design in healthy adults aged ≥60 y using the precursor-product method.
Sixteen healthy older adults (n = 8 male and n = 8 female; body mass index ≤30 kg/m2) were studied. Each received 4 of 6 protein intakes in random order (0.66, 0.8, 0.9, 1.1, 1.3 and 1.5 g⋅kg-1⋅d-1). At each intake level, participants underwent isotope infusion studies of 7 h duration following a 3-d adaptation to the test level of protein. On the fourth day, GSH fractional (FSR) and absolute synthesis (ASR) rates were quantified by measuring the incorporation of U-[13C2-15N]glycine into GSH at isotopic steady state. A mixed-effect change-point regression model was used to determine a breakpoint in FSR and ASR. Secondary outcomes included plasma concentrations of oxidative stress markers, homocysteine, 5-L-oxoproline (5-OP), and urinary sulfate. The effect of secondary outcomes on GSH kinetics was analyzed using a joint linear mixed-effect model and Tukey\'s post hoc test.
A protein intake of 1.08 g⋅kg-1⋅d-1 (95% confidence interval [CI]: 0.83, 1.32; Rm2 = 0.207; Rc2 = 0.671; P < 0.001) maximized GSH FSR. There was no effect of protein intake on concentrations of erythrocyte GSH, plasma homocysteine, oxidative stress markers, or 5-OP (P > 0.05). Protein intake had a positive effect on urinary sulfate excretion (P < 0.0001).
A protein intake of 1.08 g⋅kg-1⋅d-1 from a high-quality protein maximized GSH synthesis in adults ≥60 y. This lends support to data suggesting a requirement higher than the current recommendation. This study was registered at clinicaltrials.gov as NCT02971046.

Clinical and biochemical vitamin B12 (B12) deficiency is lower than anticipated in vegetarians. Extraileal absorption, such as from the colon, as well as reduced daily excretion, may be adaptive mechanisms to maintain B12 homeostasis with marginal intakes.
To measure the absorption of B12 from the small and large intestine, and its daily rate of excretion from the body, using a [13C]-cyanocobalamin tracer.
Oral B12 bioavailability was measured over 12 h after administration of [13C]-cyanocobalamin tracer (2.5 μg) in normal participants. The colonic B12 bioavailability was evaluated by direct instillation of [13C]-cyanocobalamin (5 μg) into the ascending colon. Bioavailability was calculated from 2-compartmental modeling of the tracer appearance in plasma. The excretion rate of B12 was measured from [13C]-cyanocobalamin elimination from the body over 4 wk after oral dosing (5 μg).
The oral B12 bioavailability (n = 11) was 63% ± 10% measured over 12 h. A late absorption peak, accounting for 12% of the absorption, was observed after an average lag time of 8.7 h from dosing. The colonic B12 bioavailability (n = 10) was 7% ± 5% over 4 h. The daily B12 excretion rate (n = 4) was 0.7 ± 0.2 μg/d. The minimum daily requirement of B12 in these participants was derived at 1 μg /d.
B12 is absorbed in the human colon. This observation confirms the potential contribution of the colon in daily B12 nutriture, and along with a possible lower requirement, could explain the absence of clinical deficiency in populations with marginal B12 intakes.
This study was registered in Clinical Trials Registry of India (CTRI) with the registration number CTRI/2018/04/012957, available from https://ctri.nic.in/Clinicaltrials/showallp.php?mid1=49319&EncHid=&userName=029108.

The Japanese flying squid (Todarodes pacificus) is an important cephalopod in the northwest Pacific Ocean. In this study, the proostracum of the gladius of T. pacificus samples collected by Chinese squid fishing vessels in the East China Sea and the Sea of Japan in August and December 2018 were continuously cut, and stable isotope values of the cut fragments were used to analyze the migration path and feeding ecology of T. pacificus. The results showed that when the proostracum grew to 120 mm from the distal end, T. pacificus began to migrate. In the East China Sea, T. pacificus migrated to low latitudes and nearshore areas, and the trophic level of their food showed no large changes during migration. In the Sea of Japan, T. pacificus migrated to high latitudes and offshore areas, and the trophic level of their food showed a decreasing trend during migration. There was no significant difference in migration or feeding ecology between females and males, but the competitive capacity of the females may be stronger than that of the males. The results provided a scientific basis for the scientific management and development of T. pacificus resources.

BACKGROUND: Official stable isotope databases, based on the analysis of (D/H)I ethanol , (D/H)II ethanol , δ13 Cethanol and δ18 Owater of wine, are an indispensable tool for establishing the limits beyond which the mislabeling or the addition of sugar and/or water in wine production can be detected. The present study investigates, for the first time, whether the use of hybrid varieties instead of European Vitis vinifera for wine production can have an impact on the stable isotope ratios.
RESULTS: The analyses were performed by isotope ratio mass spectrometry and site-specific natural isotope fractionation by nuclear magnetic resonance, in accordance with the official methods of the International Organization of Grapes and Wine. The comparison shows the tendency of some stable isotope ratios of hybrid varieties, in particular (D/H)I , to deviate from the regional averages of the V. vinifera samples. Notably, Baron, Monarch and Regent showed significantly different values at one of the two sampling sites. Particularly high δ13 C values characterize Helios compared to other hybrid varieties.
CONCLUSIONS: For the first time, and from an isotopic point of view, the present study investigates the wine obtained from hybrid varieties, showing that further attention should be paid to their interpretation, on the basis of the database established according to the European Regulation 2018/273. © 2022 Society of Chemical Industry.

Dietary methyl donors (e.g., choline) support the activity of the phosphatidylethanolamine N-methyltransferase (PEMT) pathway, which generates phosphatidylcholine (PC) molecules enriched in DHA that are exported from the liver and made available to extrahepatic tissues.
This study investigated the effect of prenatal choline supplementation on biomarkers of DHA status among pregnant participants consuming supplemental DHA.
Pregnant participants (n = 30) were randomly assigned to receive supplemental choline intakes of 550 mg/d [500 mg/d d0-choline + 50 mg/d deuterium-labeled choline (d9-choline); intervention] or 25 mg/d (25 mg/d d9-choline; control) from gestational week (GW) 12-16 until delivery. All participants received a daily 200-mg DHA supplement and consumed self-selected diets. Fasting blood samples were obtained at baseline, GW 20-24, and GW 28-32; maternal/cord blood was obtained at delivery. Mixed-effects linear models were used to assess the impact of prenatal choline supplementation on maternal and newborn DHA status.
Choline supplementation (550 vs. 25 mg/d) did not achieve a statistically significant intervention × time interaction for RBC PC-DHA (P = 0.11); a significant interaction was observed for plasma PC-DHA and RBC total DHA, with choline supplementation yielding higher levels (+32-38% and +8-11%, respectively) at GW 28-32 (P < 0.05) and delivery (P < 0.005). A main effect of choline supplementation on plasma total DHA was also observed (P = 0.018); its interaction with time was not significant (P = 0.068). Compared with controls, the intervention group exhibited higher (P = 0.007; main effect) plasma enrichment of d3-PC (d3-PC/total PC). Moreover, the ratio of d3-PC to d9-PC was higher (+50-67%; P < 0.001) in the choline intervention arm (vs. control) at GW 20-24, GW 28-32, and delivery.
Prenatal choline supplementation improves hepatic DHA export and biomarkers of DHA status by bolstering methyl group supply for PEMT activity among pregnant participants consuming supplemental DHA. This trial is registered at www.clinicaltrials.gov as NCT03194659.

Ferrous ammonium phosphate (FAP) is an iron salt that has been developed for the fortification of food matrices sensitive to color and flavor changes. The objective of the study was to measure iron absorption from FAP in young children and compare it to a previous evaluation of FAP in young women. A double-blind randomized crossover study with two parallel arms was used to evaluate the iron absorption from FAP added to reconstituted milk powder in comparison to that from ferrous sulfate (FeSO4) and ferric pyrophosphate (FePP). Iron absorption was measured in 39 children aged 3- to 6-years-old using erythrocyte incorporation of stable Fe isotopes (57Fe, 58Fe). The geometric mean iron absorption in iron replete children from FAP, FeSO4 and FePP from milk was 8.3%, 7.6% and 2.1%, respectively. Iron absorption from FAP and FeSO4 fortified milk was not significantly different (p = 0.199); however, it was significantly higher than from FePP fortified milk (p < 0.001). Iron bioavailability from FAP and FePP relative to FeSO4 (relative bioavailability (RBV)) was 110% and 33%, respectively. The RBV of FAP (110%) in iron replete children was higher than previously reported RBV (71%) in mainly iron deficient women. The difference in iron status between the children and women in the respective studies may explain the different RBV values and is discussed.

Nitrate (NO3-) pollution in water bodies has received widespread attention, but studies on nitrogen transformation and pollution risk assessment are still limited, especially in rare earth mining areas. In this study, surface and groundwater samples were collected from the largest rare earth mining site in southern China, and analyzed for the hydrochemical and stable isotopic characteristics. The results showed that the NO3- concentrations ranged from 1.61 to 453.11 mg/L, with 35% of surface water and 53.3% of groundwater samples exceeding the WHO standard (i.e., 50 mg/L). Health risk assessment showed that 31.4% of the water samples had a moderate to high non-carcinogenic risk, and the high-risk areas were concentrated in rare earth mining regions. Additionally, adults were more vulnerable to the non-carcinogenic health risks than children. The high variability of δ15N-NO3- (from -6.43 to 17.09‰) and δ18O-NO3- (from -7.91 to 22.79‰) showed that NO3- was influenced by multiple nitrogen sources and transformation processes. Hydrochemistry and isotopic evidence further indicated that NO3- was primarily influenced by nitrification and hydraulic connection between surface and groundwater. The results of the Bayesian mixing model showed that about 70% of NO3- originated from mine drainage and soil N in the rare earth mining area, while more than 90% of NO3- originated from fertilizer, soil N, and manure and sewage in rural and urban areas in the middle and downstream. This study suggests reducing anthropogenic nitrogen discharge (e.g., leaching agents and fertilizer inputs) as the primary means of NO3- pollution control with biogeochemical processes (e.g., denitrification) to further reduce its pollution.