scholarly journals High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application

2021 ◽  
Vol 7 ◽  
Author(s):  
Daniela Hampel ◽  
Setareh Shahab-Ferdows ◽  
Ngoc Nguyen ◽  
Gilberto Kac ◽  
Lindsay H. Allen
Keyword(s):  
Human Milk ◽  
Matrix Effects ◽  
Rapid Assessment ◽  
Water Soluble ◽  
Signal Loss ◽  
High Throughput Analysis ◽  
Infant Outcomes ◽  
Breastfed Infants ◽  
Minimal Sample ◽  
High Concentrations

Choline and related metabolites are key factors in many metabolic processes, and insufficient supply can adversely affect reproduction and fetal development. Choline status is mainly regulated by intake, and human milk is the only choline source for exclusively breastfed infants. Further, maternal status, genotype, and phenotype, as well as infant outcomes, have been related to milk choline concentrations. In order to enable the rapid assessment of choline intake for exclusively breastfed infants and to further investigate the associations between milk choline and maternal and infant status and other outcomes, we have developed a simplified method for the simultaneous analysis of human milk choline, glycerophosphocholine, phosphocholine, and the less abundant related metabolites betaine, carnitine, creatinine, dimethylglycine (DMG), methionine, and trimethylamine N-oxide (TMAO) using ultraperformance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS). These analytes have milk concentrations ranging over 3 orders of magnitude. Unlike other recently described LC-based methods, our approach does not require an ion-pairing reagent or high concentrations of solvent modifiers for successful analyte separation and thus avoid signal loss and potential permanent contamination. Milk samples (10 μl) were diluted (1:80) in water : methanol (1:4, v:v) and filtered prior to analysis with an optimized gradient of 0.1% propionic acidaq and acetonitrile, allowing efficient separation and removal of contaminants. Recovery rates ranged from 108.0 to 130.9% (inter-day variation: 3.3–9.6%), and matrix effects (MEs) from 54.1 to 114.3%. MEs were greater for carnitine, creatinine, and TMAO at lower dilution (1:40, p < 0.035 for all), indicating concentration-dependent ion suppression. Milk from Brazilian women (2–8, 28–50, and 88–119 days postpartum, ntotal = 53) revealed increasing concentration throughout lactation for glycerophosphocholine, DMG, and methionine, while carnitine decreased. Choline and phosphocholine were negatively correlated consistently at all three collection time intervals. The method is suitable for rapid analysis of human milk water-soluble forms of choline as well as previously not captured related metabolites with minimal sample volumes and preparation.

scholarly journals Variability of Water-Soluble Forms of Choline Concentrations in Human Milk during Storage, after Pasteurization, and among Women

Nutrients ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 3024 ◽  
Author(s):  
Sara Moukarzel ◽  
Alejandra M. Wiedeman ◽  
Lynda S. Soberanes ◽  
Roger A. Dyer ◽  
Sheila M. Innis ◽  
...  
Keyword(s):  
Human Milk ◽  
Infant Development ◽  
Sole Source ◽  
Individual Variability ◽  
Water Soluble ◽  
Total Water ◽  
Breastfed Infants ◽  
The Stability ◽  
Stability Of Water ◽  
And Storage

Choline is critical for infant development and mother’s milk is the sole source of choline for fully breastfed infants until six months of age. Human milk choline consists to 85% of water-soluble forms of choline including free choline (FC), phosphocholine (PhosC), and glycerophosphocholine (GPC). Donor milk requires safe handling procedures such as cold storage and pasteurization. However, the stability of water-soluble forms of choline during these processes is not known. The objectives of this research were to determine the effect of storage and pasteurization on milk choline concentration, and the diurnal intra- and inter-individual variability of water-soluble choline forms. Milk samples were collected from healthy women who were fully breastfeeding a full-term, singleton infant <6 months. Milk total water-soluble forms of choline, PhosC, and GPC concentrations did not change during storage at room temperature for up to 4 h. Individual and total water-soluble forms of choline concentrations did not change after storage for 24 h in the refrigerator or for up to one week in the household freezer. Holder pasteurization decreased PhosC and GPC, and thereby total water-soluble choline form concentrations by <5%. We did not observe diurnal variations in PhosC and total water-soluble forms of choline concentrations, but significant differences in FC and GPC concentrations across five sampling time points throughout one day. In conclusion, these outcomes contribute new knowledge for the derivation of evidence-informed guidelines for the handling and storage of expressed human milk as well as the development of optimized milk collection and storage protocols for research studies.

scholarly journals High-Throughput Analysis of Water-Soluble Choline and Related Metabolites in Human Milk

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 1171-1171
Author(s):  
Daniela Hampel ◽  
Setareh Shahab-Ferdows ◽  
Ngoc Nguyen ◽  
Gilberto Kac ◽  
Allen Lindsay
Keyword(s):  
Sample Preparation ◽  
Human Milk ◽  
Water Soluble ◽  
Process Efficiency ◽  
National Council ◽  
High Throughput Analysis ◽  
Standard Curve ◽  
Funding Sources ◽  
Gates Foundation ◽  
Acquity Uplc

Abstract Objectives Choline and related metabolites play important roles in metabolic processes. Inadequate provision of these nutrients to the exclusively breast-fed infant can negatively impact its healthy growth and development. Methods We developed an UPLC-MS/MS method for analyzing choline (Cho), phospho-choline (PCho), glycerophospho-choline (GPCho), total choline (tCho = Cho + PCho + GPCho), betaine, carnitine, creatinine, dimethyl glycine (DMG), methionine, and trimethylamine N-oxide (TMAO) in human milk. Results Optimized results were obtained using a Phenomenex Luna Silica (2) column, 100 × 2 mm, 3 µm, and a gradient of 0.1% aqueous propionic acid (A) and acetonitrile (B) from 60% to 90% A over 2 min (Waters ACQUITY UPLC I-Class - SCIEX 4500TQ mass spectrometer). Sample preparation required only 5–10µL of milk, diluted 1:80 in methanol/water, 4:1, v/v, prior to analysis. Quantification was done using isotopically labeled internal standards and an external standard curve. Pooled human milk used for method validation showed recovery rates from 108–131% for all analytes, and an overall process efficiency from 54 to 114%. All standard curves revealed good linearity (r &gt; 0.999). Milk from apparently healthy Brazilian mothers (1–120days pp) revealed large concentration ranges within and between analytes (IQR, mg/L): Cho 10.3, 20.3; GPcho 48.7, 101; PCho 134, 221; tCho 120, 166; betaine 0.25, 0.53; carnitine 3.35, 5.06; creatinine 2.92, 3.90; DMG 0.26, 0.54; methionine 0.47, 0.90. 63% of the milk samples reached the tCho value used for the Adequate Intake (1–6 mo). Conclusions Our newly implemented method enabled the simultaneous analysis of water-soluble forms of choline and related metabolites in human milk in minute amounts of sample, and requiring only minimalistic sample preparation. Funding Sources Bill & Melinda Gates Foundation (OPP1148405), USDA/ARS Intramural Project (5306–51,530-019–00), and CNPq (Brazilian National Council for Science and Technology).

scholarly journals Development, characterization and first deployment of an improved online reactive oxygen species analyzer

2018 ◽  
Vol 11 (1) ◽  
pp. 65-80 ◽  
Author(s):  
Jun Zhou ◽  
Emily A. Bruns ◽  
Peter Zotter ◽  
Giulia Stefenelli ◽  
André S. H. Prévôt ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Detection Limit ◽  
Atmospheric Aerosols ◽  
Half Life ◽  
Matrix Effects ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Wood Combustion ◽  
Storage Duration ◽  
High Concentrations

Abstract. Inhalation of atmospheric particles is linked to human diseases. Reactive oxygen species (ROS) present in these atmospheric aerosols may play an important role. However, the ROS content in aerosols and their formation pathways are still largely unknown. Here, we have developed an online and offline ROS analyzer using a 2′,7′-dichlorofluorescin (DCFH) based assay. The ROS analyzer was calibrated with H2O2 and its sensitivity was characterized using a suite of model organic compounds. The instrument detection limit determined as 3 times the noise is 1.3 nmol L−1 for offline analysis and 2 nmol m−3 of sampled air when the instrument is operated online at a fluorescence response time of approximately 8 min, while the offline method detection limit is 18 nmol L−1. Potential interferences from gas-phase O3 and NO2 as well as matrix effects of particulate SO42− and NO3− were tested, but not observed. Fe3+ had no influence on the ROS signal, while soluble Fe2+ reduced it if present at high concentrations in the extracts. Both online and offline methods were applied to identify the ROS content of different aerosol types, i.e., ambient aerosols as well as fresh and aged aerosols from wood combustion emissions. The stability of the ROS was assessed by comparing the ROS concentration measured by the same instrumentation online in situ with offline measurements. We also analyzed the evolution of ROS in specific samples by conducting the analysis after storage times of up to 4 months. The ROS were observed to decay with increasing storage duration. From their decay behavior, ROS in secondary organic aerosol (SOA) can be separated into short- and long-lived fractions. The half-life of the short-lived fraction was 1.7 ± 0.4 h, while the half-life of the long-lived fraction could not be determined with our uncertainties. All these measurements showed consistently that on average 60 ± 20 % of the ROS were very reactive and disappeared during the filter storage time. This demonstrates the importance of a fast online measurement of ROS.

scholarly journals NMR Structure and Action on Nicotinic Acetylcholine Receptors of Water-soluble Domain of Human LYNX1

2011 ◽  
Vol 286 (12) ◽  
pp. 10618-10627 ◽  
Author(s):  
Ekaterina N. Lyukmanova ◽  
Zakhar O. Shenkarev ◽  
Mikhail A. Shulepko ◽  
Konstantin S. Mineev ◽  
Dieter D'Hoedt ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Nmr Structure ◽  
Water Soluble ◽  
Gpi Anchor ◽  
Nicotinic Acetylcholine ◽  
Brain Functions ◽  
The Central Nervous System ◽  
High Concentrations

Discovery of proteins expressed in the central nervous system sharing the three-finger structure with snake α-neurotoxins provoked much interest to their role in brain functions. Prototoxin LYNX1, having homology both to Ly6 proteins and three-finger neurotoxins, is the first identified member of this family membrane-tethered by a GPI anchor, which considerably complicates in vitro studies. We report for the first time the NMR spatial structure for the water-soluble domain of human LYNX1 lacking a GPI anchor (ws-LYNX1) and its concentration-dependent activity on nicotinic acetylcholine receptors (nAChRs). At 5–30 μm, ws-LYNX1 competed with 125I-α-bungarotoxin for binding to the acetylcholine-binding proteins (AChBPs) and to Torpedo nAChR. Exposure of Xenopus oocytes expressing α7 nAChRs to 1 μm ws-LYNX1 enhanced the response to acetylcholine, but no effect was detected on α4β2 and α3β2 nAChRs. Increasing ws-LYNX1 concentration to 10 μm caused a modest inhibition of these three nAChR subtypes. A common feature for ws-LYNX1 and LYNX1 is a decrease of nAChR sensitivity to high concentrations of acetylcholine. NMR and functional analysis both demonstrate that ws-LYNX1 is an appropriate model to shed light on the mechanism of LYNX1 action. Computer modeling, based on ws-LYNX1 NMR structure and AChBP x-ray structure, revealed a possible mode of ws-LYNX1 binding.

Water-Soluble Vitamin D in Human Milk: A Myth

PEDIATRICS ◽  
1982 ◽  
Vol 70 (3) ◽  
pp. 499-499
Author(s):  
Joseph A. Little
Keyword(s):  
Vitamin D ◽  
Human Milk ◽  
Clinical Observation ◽  
Water Soluble ◽  
University Of Wisconsin ◽  
Water Soluble Vitamin ◽  
The University ◽  
Breast Fed

I was happy to read the report from Greer et al.1 This biochemically confirms the clinical observation that the majority of children who develop clinical rickets are breast-fed. During the ten years, 1970-1979, I have seen six cases of clinical rickets. All of these infants were breast-fed.2 The only exception has been the rickets of prematurity. I should like to commend, again, the report from the Department of Pediatrics and Biochemistry of the University of Wisconsin.

Diversification of a fucosyllactose transporter within the genus Bifidobacterium

2021 ◽  
Author(s):  
Miriam N. Ojima ◽  
Yuya Asao ◽  
Aruto Nakajima ◽  
Toshihiko Katoh ◽  
Motomitsu Kitaoka ◽  
...  
Keyword(s):  
Human Milk ◽  
Binding Proteins ◽  
Heterologous Gene Expression ◽  
Metagenomic Data ◽  
Dependent Manner ◽  
Human Milk Oligosaccharides ◽  
Milk Oligosaccharides ◽  
Formula Milk ◽  
Breastfed Infants ◽  
Strain Dependent

Human milk oligosaccharides (HMOs), which are natural bifidogenic prebiotics, were recently commercialized to fortify formula milk. However, HMO-assimilation phenotypes of bifidobacteria vary by species and strain, which has not been fully linked to strain genotype. We have recently shown that specialized uptake systems, particularly for the internalization of major HMOs (fucosyllactose (FL)), are associated with the formation of a bifidobacteria-rich gut microbial community. Phylogenetic analysis has revealed that FL transporters have diversified into two clades harboring four clusters within the Bifidobacterium genus, but the underpinning functional diversity associated with this divergence remains underexplored. In this study, we examined the HMO-consumption phenotypes of two bifidobacterial species, Bifidobacterium catenulatum subspecies kashiwanohense and Bifidobacterium pseudocatenulatum , which both possess FL binding proteins that belong to phylogenetic clusters with unknown specificities. Growth assays, heterologous gene expression experiments, and HMO-consumption analysis showed that the FL transporter type from B. catenulatum subspecies kashiwanohense JCM 15439 T conferred a novel HMO-uptake pattern that includes the complex fucosylated HMOs (lacto- N- fucopentaose II and lacto- N- difucohexaose I/II). Further genomic landscape analyses of FL transporter-positive bifidobacterial strains revealed that H-antigen or Lewis antigen-specific fucosidase gene(s) and FL transporter specificities were largely aligned. These results suggest that bifidobacteria have acquired FL transporters along with the corresponding gene sets necessary to utilize the imported HMOs. Our results provide insight into the species- and strain-dependent adaptation strategies of bifidobacteria to HMO-rich environments. Importance The gut of breastfed infants is generally dominated by health-promoting bifidobacteria. Human milk oligosaccharides (HMOs) from breastmilk selectively promote the growth of specific taxa such as bifidobacteria, thus forming an HMO-mediated, host-microbe symbiosis. While the co-evolution of humans and bifidobacteria has been proposed, the underpinning adaptive strategies employed by bifidobacteria require further research. Here, we analyzed the divergence of the critical fucosyllactose (FL) HMO transporter within Bifidobacterium . We have shown that the diversification of the solute-binding proteins of the FL-transporter led to uptake specificities of fucosylated sugars ranging from simple trisaccharides to complex hexasaccharides. This transporter and the congruent acquisition of the necessary intracellular enzymes allows for bifidobacteria to import different types of HMOs in a predictable and strain-dependent manner. These findings explain the adaptation and proliferation of bifidobacteria in the competitive and HMO-rich infant gut environment and enable accurate specificity annotation of transporters from metagenomic data.

scholarly journals Characteristics of aerosol pollution during heavy haze events in Suzhou, China

2016 ◽  
Vol 16 (11) ◽  
pp. 7357-7371 ◽  
Author(s):  
Mi Tian ◽  
Huanbo Wang ◽  
Yang Chen ◽  
Fumo Yang ◽  
Xiaohua Zhang ◽  
...  
Keyword(s):  
Gas Phase ◽  
Inorganic Ions ◽  
Weather Conditions ◽  
Water Soluble ◽  
Chemical Components ◽  
Formation Mechanisms ◽  
Visibility Impairment ◽  
Aerosol Pollution ◽  
Haze Formation ◽  
High Concentrations

Abstract. Extremely severe haze weather events occurred in many cities in China, especially in the east part of the country, in January 2013. Comprehensive measurements including hourly concentrations of PM2.5 and its major chemical components (water-soluble inorganic ions, organic carbon (OC), and elemental carbon (EC)) and related gas-phase precursors were conducted via an online monitoring system in Suzhou, a medium-sized city in Jiangsu province, just east of Shanghai. PM2.5 (particulate matter with an aerodynamic diameter of 2.5 µm or less) frequently exceeded 150 µg m−3 on hazy days, with the maximum reaching 324 µg m−3 on 14 January 2013. Unfavorable weather conditions (high relative humidity (RH), and low rainfall, wind speed, and atmospheric pressure) were conducive to haze formation. High concentrations of secondary aerosol species (including SO42−, NO3−, NH4+, and SOC) and gaseous precursors were observed during the first two haze events, while elevated primary carbonaceous species emissions were found during the third haze period, pointing to different haze formation mechanisms. Organic matter (OM), (NH4)2SO4, and NH4NO3 were found to be the major contributors to visibility impairment. High concentrations of sulfate and nitrate might be explained by homogeneous gas-phase reactions under low RH conditions and by heterogeneous processes under relatively high RH conditions. Analysis of air mass trajectory clustering and potential source contribution function showed that aerosol pollution in the studied areas was mainly caused by local activities and surrounding sources transported from nearby cities.

scholarly journals High concentrations of coarse particles emitted from a cattle feeding operation

2011 ◽  
Vol 11 (16) ◽  
pp. 8809-8823 ◽  
Author(s):  
N. Hiranuma ◽  
S. D. Brooks ◽  
J. Gramann ◽  
B. W. Auvermann
Keyword(s):  
United States ◽  
Particulate Matter ◽  
Chemical Properties ◽  
Field Measurements ◽  
The United States ◽  
Water Soluble ◽  
Size Distributions ◽  
Cattle Feeding ◽  
High Concentrations ◽  
Organic Particles

Abstract. Housing roughly 10 million head of cattle in the United States alone, open air cattle feedlots represent a significant but poorly constrained source of atmospheric particles. Here we present a comprehensive characterization of physical and chemical properties of particles emitted from a large representative cattle feedlot in the Southwest United States. In the summer of 2008, measurements and samplings were conducted at the upwind and downwind edges of the facility. A series of far-field measurements and samplings was also conducted 3.5 km north of the facility. Two instruments, a GRIMM Sequential Mobility Particle Sizer (SMPS) and a GRIMM Portable Aerosol Spectrometer (PAS), were used to measure particle size distributions over the range of 0.01 to 25 μm diameter. Raman microspectroscopy was used to determine the chemical composition of particles on a single particle basis. Volume size distributions of dust were dominated by coarse mode particles. Twenty-four hour averaged concentrations of PM10 (particulate matter with a diameter of 10 μm or less) were as high as 1200 μg m−3 during the campaign. The primary constituents of the particulate matter were carbonaceous materials, such as humic acid, water soluble organics, and less soluble fatty acids, including stearic acid and tristearin. A significant fraction of the organic particles was present in internal mixtures with salts. Basic characteristics such as size distribution and composition of agricultural aerosols were found to be different than the properties of those found in urban and semi-urban aerosols. Failing to account for such differences may lead to errors in estimates of aerosol effects on local air quality, visibility, and public health.

scholarly journals Characteristics, seasonality and sources of carbonaceous and ionic components in the tropical Indian aerosols

2011 ◽  
Vol 11 (2) ◽  
pp. 3937-3976 ◽  
Author(s):  
C. M. Pavuluri ◽  
K. Kawamura ◽  
S. G. Aggarwal ◽  
T. Swaminathan
Keyword(s):  
Organic Carbon ◽  
Atmospheric Aerosols ◽  
Atmospheric Transport ◽  
Indian Subcontinent ◽  
Ionic Species ◽  
Water Soluble ◽  
Carbonaceous Aerosols ◽  
Mass Fractions ◽  
High Concentrations ◽  
Ionic Components

Abstract. To better characterize South and Southeast Asian aerosols, PM10 samples collected from tropical Chennai, India (13.04° N; 80.17° E) were analyzed for carbonaceous and water-soluble ionic components. Concentration ranges of elemental carbon (EC) and organic carbon (OC) were 2.4–14 μg m−3 and 3.2–15.6 μg m−3 in winter samples whereas they were 1.1–2.5 μg m−3 and 4.1–17.6 μg m−3 in summer samples, respectively. Concentration of secondary organic carbon (SOC) retrieved from EC-tracer method was 4.6 ± 2.8 μg m−3 in winter and 4.3 ± 2.8 μg m−3 in summer. SO42- (8.8 ± 2.5 μg m−3 and 4.1 ± 2.7 μg m−3 in winter and summer, respectively) was found as the most abundant ionic species (57% on average, n = 49), followed by NH4+ (15%) > NO3− > Cl− > K+> Na+ > Ca2+ > MSA− > Mg2+. The mass fractions of EC, organic matter (OM) and ionic species varied seasonally, following the air mass trajectories and corresponding source strength. Based on mass concentration ratios of selected components and relations of EC and OC to marker species, we found that biofuel/biomass burning is the major source of atmospheric aerosols in South and Southeast Asia. The high concentrations of SOC and WSOC/OC ratios (ave. 0.45; n = 49) as well as good correlations between SOC and WSOC suggest that the secondary production of organic aerosols during long-range atmospheric transport is also significant in this region. This study provides the baseline data of carbonaceous aerosols for southern part of the Indian subcontinent.

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