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 > 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 Human Milk Metabolic Profiling Using Biocrates MxP® Quant 500 Kit

2021 ◽  
Vol 5 (Supplement_2) ◽  
pp. 874-874
Author(s):  
Daniela Hampel ◽  
Setareh Shahab-Ferdows ◽  
Gilberto Kac ◽  
Lindsay Allen
Keyword(s):  
Fatty Acids ◽  
Human Milk ◽  
Small Molecules ◽  
Metabolic Profiling ◽  
National Council ◽  
Funding Sources ◽  
Upper Limit ◽  
Limit Of Quantitation ◽  
Gates Foundation ◽  
Comparability Of Results

Abstract Objectives Comprehensive metabolic profiling of human milk is a useful tool for examining its composition and relationship to maternal and infant metabolism and status. Methods A pooled milk sample was used to evaluate the Biocrates MxP® Quant 500 kit for human milk metabolomics (107 small molecules and 523 lipids) using an ABSciex 5500QTRAP mass-spectrometer in LC-MS/MS and flow injection analysis (FIA) mode. Additionally, milk from Brazilian mothers (A: 2–8, B: 28–50, C: 88–119d postpartum, ntotal = 25) was analyzed. Results 424 of the 630 assay metabolites were detected in the milkpool above the limit of quantitation (LOQ); 31 metabolites were below the lower limit of quantitation (LLOQ), while 7 were above the upper limit (ULOQ), mostly free fatty acids including arachidonic acid, docohexaenoic acid, and eicosapentaenoic acid. Bile acids were only detected below LLOQ. Concentrations measured in 5 different sample volumes (2–20μL, n = 10) showed satisfactory reproducibility (CV 3.7–20.0%) for 458 metabolites. Acceptable intraday variation (80–120%, 6 replicates) was achieved for 409 metabolites when spiking with 3 levels of QC-standards, but only for 127 metabolites after dilution (1:2 and 1:5). However, 396 metabolites revealed a good intraday variation when only considering the 1:2-dilution. Conclusions The MxP Quant® 500 kit was successfully employed for human milk providing data for over 400 metabolites in 10μL milk. and thus offers ability to use the same assay for both human plasma and milk, enhancing comparability of results by reducing analytical bias and increasing our ability to study milk as a biological system. 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 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 Human Milk Metabolomics Using Biocrates AbsoluteIDQ® p180 Kit Assay (OR06-04-19)

2019 ◽  
Vol 3 (Supplement_1) ◽  
Author(s):  
Daniela Hampel ◽  
Setareh Shahab-Ferdows ◽  
Muttaquina Hossain ◽  
M Munirul Islam ◽  
Tahmeed Ahmed ◽  
...  
Keyword(s):  
Amino Acids ◽  
Human Milk ◽  
Biogenic Amines ◽  
Human Plasma ◽  
Analytical Techniques ◽  
Milk Composition ◽  
Funding Sources ◽  
Carry Over ◽  
Gates Foundation ◽  
Student’S T

Abstract Objectives Targeted metabolomics are commercially available for human plasma, but not for human milk. However, metabolite analyses could provide a novel and efficient approach to understanding human milk composition and relationships to maternal and infant status. Methods Pooled human milk was used to evaluate and validate the Biocrates AbsoluteIDQ® p180 kit for human milk metabolomics (40 acylcarnitines, 42 acids/biogenic amines, 91 phospholipids, 15 sphingolipids, sum of hexoses) using an ABSciex 5500QTRAP mass spectrometer in LC-MS/MS and flow injection analysis (FIA) mode. In a feasibility study, milk collected <6 mo lactation from A) Bangladeshi healthy mothers (BMI >18.5, n = 12) and from B) mothers with stunted infants (HAZ-score < −2; n = 13) were analyzed. Results 120 of the detectable 188 assay metabolites were found in the pooled milk, including all of the sphingolipids and amino acids. Additional internal standards (IS) were prepared for lysine and some biogenic amines for higher accuracy. Higher amounts of glutamate, taurine, and putrescine in milk required higher levels of calibrators than for plasma in LC-MS/MS mode. For metabolites of low abundance diluted calibrators (0.25 and 0.5) were added. FIA provided results for 94 of 146 metabolites above LOD without any carry-over. Metabolite recoveries (levels) varied between 64.1 and 127.0%. Intra-assay variations (6 replicates) for all detectable metabolites ranged between 3.4 and 18.4%. Human milk given to healthy compared to malnourished infants was higher in the amino acids citrulline, glutamate, glycine, and phenylalanine, and carnitine, while histamine and dodecanoylcarnitine were lower (Student's t-test, P < 0.05 for all). Conclusions The AbsoluteIDQ® p180 can be used for human milk application and thus allows the application of the same assay for both human plasma and milk, enhancing comparability of results by reducing analytical bias due to different analytical techniques. Funding Sources Bill & Melinda Gates Foundation (OPP1148405 & OPP1164613), USDA/ARS Intramural Project (5306–51,530-019–00).

RADIOCARBON IN MEXICO: FROM PROPORTIONAL COUNTERS TO AMS

Radiocarbon ◽  
2021 ◽  
pp. 1-7
Author(s):  
Corina Solís ◽  
Efraín Chávez ◽  
Arcadio Huerta ◽  
María Esther Ortiz ◽  
Alberto Alcántara ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
High Voltage ◽  
Accelerator Mass Spectrometry ◽  
Science And Technology ◽  
National Council ◽  
University Of Arizona ◽  
Technological Developments ◽  
High Voltage Engineering ◽  
The University

ABSTRACT Augusto Moreno is credited with establishing the first radiocarbon (14C) laboratory in Mexico in the 1950s, however, 14C measurement with the accelerator mass spectrometry (AMS) technique was not achieved in our country until 2003. Douglas Donahue from the University of Arizona, a pioneer in using AMS for 14C dating, participated in that experiment; then, the idea of establishing a 14C AMS laboratory evolved into a feasible project. This was finally reached in 2013, thanks to the technological developments in AMS and sample preparation with automated equipment, and the backing and support of the National Autonomous University of Mexico and the National Council for Science and Technology. The Mexican AMS Laboratory, LEMA, with a compact 1 MV system from High Voltage Engineering Europa, and its sample preparation laboratories with IonPlus automated graphitization equipment, is now a reality.

scholarly journals Differences in the Concentration and Composition of Human Milk Components Are Related to Variation in Milk and Infant Fecal Microbiomes

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 1056-1056
Author(s):  
Ryan Pace ◽  
Janet Williams ◽  
Kimberly Lackey ◽  
Mark McGuire ◽  
Michelle McGuire ◽  
...  
Keyword(s):  
Human Milk ◽  
Microbial Communities ◽  
Infant Health ◽  
Hypervariable Region ◽  
Fecal Microbiota ◽  
National Institutes Of Health ◽  
Funding Sources ◽  
Infant Feces ◽  
Microbe Interactions ◽  
Foundation Award

Abstract Objectives Profiles of human milk oligosaccharides (HMO) and milk/infant fecal microbiota vary globally. However, associations between and among HMO, other milk-borne factors (e.g., lactose, protein), and milk/infant fecal microbiomes have not been well-investigated. Here we tested the hypothesis that variations in milk lactose, protein, and HMO concentrations are associated with variations in the structure of milk and infant fecal microbial communities. Methods Milk/infant fecal samples from 357 maternal-infant dyads collected as part of the INSPIRE study from 11 geographically/culturally diverse sites located in eight countries (Ethiopia, The Gambia, Ghana, Kenya, Peru, Spain, Sweden, and USA) were analyzed. DNA was extracted and bacterial 16S rRNA V1V3 hypervariable region amplified/sequenced for microbiome analysis. HMO, lactose, and protein profiles were generated from HPLC and spectrophotometric assays. Results Milk and infant feces share many of the same abundant bacterial genera, while also containing unique bacterial communities. Community states type (CST) analyses indicate both sample types group into a relatively small number of discrete communities characterized by enrichment of specific taxa (e.g., Streptococcus, Bifidobacterium). Concentrations of milk lactose and protein varied by population/CST. Additionally, variation in the microbial community structure of milk and infant feces was associated with concentrations of total/individual HMO, lactose, and protein. Conclusions Similar to HMO concentrations, milk lactose and protein vary globally. Variations in milk and infant fecal microbial communities are associated with those of milk lactose, protein, and HMO concentrations. Given these results, as well as prior data on the influence of other environmental variables (e.g., pumped vs. direct breastfeeding), additional longitudinal studies are needed to better understand this complex network of maternal-infant-microbe interactions with respect to environmental factors and how differences impact postnatal maternal-infant health. Funding Sources National Science Foundation (award 1,344,288), National Institutes of Health (R01 HD092297), and USDA.

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.

scholarly journals Effect of Fat on Protein Estimation in Milk and Its Correlation with Lactose in Different Milk Types: A Small-Scale Study

2020 ◽  
Author(s):  
Sweekruthi A. Shetty ◽  
Melissa F. Young ◽  
Sunita Taneja ◽  
Kannan Rangiah
Keyword(s):  
Sample Preparation ◽  
Human Milk ◽  
Internal Standard ◽  
Cow Milk ◽  
Selected Reaction Monitoring ◽  
Small Scale ◽  
Milk Samples ◽  
Protein Estimation ◽  
Animal Milk ◽  
Fat Removal

Background: Estimation of macronutrients like protein and lactose is important to assess the quality of milk. To estimate these two macronutrients, ten raw milk samples obtained from each group of different animals (cow, goat, buffalo), ten pasteurized cow milk and ten human milk samples were analysed. Methods: Bicinchoninic acid (BCA) method was used to estimate protein from different milk samples. Four different sample preparation protocols were compared to check the effect of fat on BCA based protein estimation: dilution (D), fat removal-protein precipitation (FR and PP), fat removal-dilution (FR and D) and dilution-fat removal (D and FR). For lactose quantification, ultrahigh-performance liquid chromatography-mass spectrometry-selected reaction monitoring (UHPLC-MS/SRM) method was developed and validated using 13C6 lactose as internal standard (ISTD).Result: Among these four different protocols, D and FR method showed consistent data for total protein content in animal milk (cow-3.16%, goat-3.21%, buffalo-3.81%, pasteurized-2.98%) and FR and PP showed consistent data in human milk samples (1.2%). Though BCA method is simple to use, proper sample preparation protocol has to be applied prior to protein estimation to avoid the interference due to fat or lactose. In case of lactose, inter-day validation showed the accuracy ranging from 97.13 to 100.54%, coefficient of variation varying between 0.1 to 1.53%, correlation R2=0.999. Lactose is in the range of 4.1 to 4.8% in animal milk and 6.6% in human milk samples. The internal ratio of lactose/protein (1.28 to 1.55 in animal milk and 5.33 in human milk) will be useful to differentiate human milk from animal milk type and to assess the milk quality.

scholarly journals Pushing Nanomaterials past the Kilogram Scale—a Targeted Approach Integrating Scalable Microreactors, Machine Learning and High-Throughput Analysis

2020 ◽  
Author(s):  
Nicholas Jose ◽  
mikhail Kovalev ◽  
Eric Bradford ◽  
Artur Schweidtmann ◽  
Hua Chun Zeng ◽  
...  
Keyword(s):  
Machine Learning ◽  
High Throughput ◽  
Scale Up ◽  
Process Efficiency ◽  
High Yield ◽  
Synthesis Methods ◽  
High Throughput Analysis ◽  
Throughput Analysis ◽  
Highly Active ◽  
Technological Advances

Novel materials are the backbone of major technological advances. However, the development and wide-scale introduction of new materials, such as nanomaterials, is limited by three main factors—the expense of experiments, inefficiency of synthesis methods and complexity of scale-up. Reaching the kilogram scale is a hurdle that takes years of effort for many nanomaterials. We introduce an improved methodology for materials development, combining state-of-the-art techniques—multi-objective machine learning optimization, high yield microreactors and high throughput analysis. We demonstrate this approach by efficiently developing a kg per day reaction process for highly active antibacterial ZnO nanoparticles. The proposed method has the potential to significantly reduce experimental costs, increase process efficiency and enhance material performance, which culminate to form a new pathway for materials discovery.

scholarly journals Frequency of Breakfast and Meals in Relation to Weight Change in a Cohort of Mexican Women (P18-031-19)

2019 ◽  
Vol 3 (Supplement_1) ◽  
Author(s):  
Claudia Martínez ◽  
Eduardo Ortíz-Panozo ◽  
Adriana Monge ◽  
Mario Flores-Aldana ◽  
Martín Lajous
Keyword(s):  
Weight Gain ◽  
Weight Change ◽  
Protective Role ◽  
Self Report ◽  
National Council ◽  
Mexican Women ◽  
American Institute ◽  
Funding Sources ◽  
Important Relationship ◽  
Day Of The Week

Abstract Objectives To evaluate weight change between 2008 and 2011 in relation to the frequency of breakfast and meals. Methods Data were obtained from 60,946 women from the Mexican Teacher's cohort. Lifestyle, diet, and anthropometric data were obtained by self-report. Frequency of breakfast and meals were categorized (0, 1–3, 4–6, or 7 day/week; 1–2, 3–4, or >5 times/day). We used linear and logistic regression to analyze weight change continuous or dichotomized as significant weight change (>5 kg). Models were adjusted for sociodemographic, dietary and lifestyle factors. Results Women who ate breakfast daily obtained an odds ratio (OR) of 0.95; (95% confidence interval (CI) 0.88, 1.03) versus women who did not have breakfast any day of the week (reference category). As the frequency of breakfast increased, the tendency to gain weight increased (P-trend = 0.07). Daily breakfast decreased the chance of gaining 5 kg in women with BMI 18–24.9 m/kg2, (OR 0.86; CI 0.75, 0.99). In this group of women, the odds of gaining 5 kg in 3 years decreased as the weekly breakfast frequency increased (P-trend < 0.02). Women who consumed 5 meals a day gained 300 g more than women who consumed 3–4 meals a day (CI = .170 g, .480 g). Weight gain was greater as the number of meals per day increased (P-trend < .0001). Women who had 5 or more meals a day were 22% more likely gain 5 kg than women who had 3–4 meals a day (OR 1.22; CI 1.14, 1.31). This trend increased as the frequency of meals increased (P-trend < .0001). Conclusions The effect of breakfast is not clear, however it seems to have a protective role. The number of meals showed an important relationship with weight gain among Mexican women. Funding Sources This work was supported by the American Institute for Cancer Research, National Council of Science and Technology and Ministry of Health Mexico.

scholarly journals Harmonization of Maternal Nutrition Trials – Finding and Creating Similarities in Protocols and Outcomes

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 1738-1738
Author(s):  
Alison Gernand ◽  
Yemane Berhane ◽  
Nita Bhandari ◽  
Ranadip Chowdhury ◽  
Fyezah Jehan ◽  
...  
Keyword(s):  
Data Analysis ◽  
Gestational Age ◽  
Maternal Nutrition ◽  
Time Frame ◽  
Protein Supplementation ◽  
Fetal Biometry ◽  
Funding Sources ◽  
Biospecimen Collection ◽  
Gates Foundation ◽  
Meta Analyses

Abstract Objectives Public health and clinical recommendations should be based on results from multiple studies, however trials often have outcomes that are not defined in the same way. This project aims to harmonize selected protocols, outcome definitions, and data analysis across five randomized trials of antenatal balanced energy-protein supplementation being conducted in Burkina Faso, Ethiopia, India, Nepal, and Pakistan. Methods Harmonization efforts include a range of activities from reviewing detailed protocols, biospecimen collection plans, data dictionaries, and data analysis plans to proposing best practices and acceptable practices based on field limitations. Most studies have not begun or are early in enrollment, an ideal time frame to make changes. A two-day workshop of lead investigators, content experts and advisors will be held in late February, and harmonization activities will continue thereafter. Results All studies are examining anthropometry at birth as a primary outcome, however the timing of birth measurements (hours since birth) and types of measurements taken differ across trials. All studies are estimating gestational age by ultrasound measurements, but the gestational age at ultrasound differs (in part due to differences in timing of antenatal care by country) as well as the number of fetal biometry measures. Finally, stillbirth is a key outcome across trials, but initial definitions had slight differences that will now be harmonized. We are also able to add new, important maternal and child health outcomes to each trial that will have the same protocols from inception (e.g., microbiome). Conclusions Efforts thus far have resulted in communication between study investigators, consideration of improved protocols, and addition of new outcomes to collect across all sites. Further results are forthcoming after the February workshop, which will include documentation of how much definitions vary across studies and the challenges of standardization. We expect the harmonization process to improve overall reporting within each study and provide opportunities for better meta-analyses. Funding Sources The Bill and Melinda Gates Foundation.

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