Summary of Study ST003765

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR002348. The data can be accessed directly via it's Project DOI: 10.21228/M84C2P This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study ID	ST003765
Study Title	1H NMR metabolomics applied to detect metabolic markers of early ostedifferentiation of mesenchymal stem cells from multiple donors
Study Type	1H NMR metabolomics to detect metabolic markers of early ostedifferentiation of human adipose-derived mesenchymal stem cells from multiple donors
Study Summary	In this study, the intra- and extracellular metabolic features accompanying the osteodifferentiation of hAMSC were characterized for 3 independent donors, over a period of 21 days and compared to proliferating cells. By employing untargeted nuclear magnetic resonance (NMR) metabolomics, it was established a reliable donor-independent metabolic signatures (sets of metabolite variations) that may serve as predictive markers of osteodifferentiation, while elucidating particularly responsive metabolic pathways accompanying the process. The integration of endo- and exometabolomic adaptations should facilitate the development of rapid and non-invasive strategies to detect and predict early MSC osteodifferentiation.
Institute	University of Aveiro
Department	CICECO – Aveiro Institute of Materials, Department of Chemistry
Laboratory	Metabolomics Group
Last Name	Gil
First Name	Ana
Address	CICECO - Departamento de Química, Universidade de Aveiro, Campus de Santiago
Email	agil@ua.pt
Phone	+351234370707
Submit Date	2025-01-17
Raw Data Available	Yes
Raw Data File Type(s)	fid
Analysis Type Detail	NMR
Release Date	2025-05-19
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR002348
Project DOI:	doi: 10.21228/M84C2P
Project Title:	1H NMR metabolomics applied to detect metabolic markers of early ostedifferentiation of mesenchymal stem cells from multiple donors
Project Type:	1H NMR metabolomics to detect metabolic markers of early ostedifferentiation of human adipose-derived mesenchymal stem cells from multiple donors
Project Summary:	Mesenchymal stem cells (MSC) are pivotal bioengineering tools that can be used effectively in tissue regeneration. However, their inherent biological variability due to inter-donor and tissue source heterogeneity often limits therapeutic applications. In addition, the lack of standardized user-independent protocols for MSC handling also contributes to this heterogeneity. Although existing assays are invaluable for detecting well-known gene/protein markers, they are often time-consuming, user-dependent and prone to reproducibility issue. The resulting high variability in MSC behavior and differentiation performance calls for improved characterization guidelines, ideally based on new robust markers. Increasing interest has therefore arisen in the development of new strategies to unambiguously monitor and predict MSC behavior, for an effective selection of donors/cells with superior therapeutic quality. MSC metabolomics appears as a valuable tool, mainly applied to: i) profile the endometabolome of different cell types/origins, ii) identify the effects of donor characteristics, and iii) measure metabolic adaptations to differentiation or other culture conditions. The relevance of metabolomics in stem cell research has been reviewed recently, and both intracellular and extracellular metabolomes have been shown to respond to osteodifferentiation, their articulated interpretation having been attempted. In this study, the intra- and extracellular metabolic features accompanying the osteodifferentiation of hAMSC were characterized for 3 independent donors, over a period of 21 days and compared to proliferating cells. By employing untargeted nuclear magnetic resonance (NMR) metabolomics, it was established a reliable donor-independent metabolic signatures (sets of metabolite variations) that may serve as predictive markers of osteodifferentiation, while elucidating particularly responsive metabolic pathways accompanying the process. The integration of endo- and exometabolomic adaptations should facilitate the development of rapid and non-invasive strategies to detect and predict early MSC osteodifferentiation.
Institute:	University of Aveiro
Department:	CICECO – Aveiro Institute of Materials, Department of Chemistry
Laboratory:	Metabolomics Group
Last Name:	Gil
First Name:	Ana
Address:	CICECO - Departamento de Química, Universidade de Aveiro, Campus de Santiago
Email:	agil@ua.pt
Phone:	+351234370707

Subject:

Subject ID:	SU003898
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Age Or Age Range:	donor 1: unknown; donor 2: 27 years-old; donor 3: 42 years-old; osteoblasts donor: 26 years-old.
Gender:	Male and female
Cell Biosource Or Supplier:	donor 1: obtained via abdominoplasty, under an agreement between the University of Aveiro and “Hospital da Luz”, Aveiro, dated 17th February 2023; donor 2: American Type Culture Collection (Lot 70017032, Ref. ATCC PCS-500-011); donor 3: Lonza (Lot 22TL018258, Ref. LSLZPT-5006); osteoblasts donor: Lonza (Lot 19TL217387, Ref. CC-2538)
Cell Passage Number:	For all MSC donors, osteodifferentiation of hAMSC was induced at passages 5 or 6 (for 21 days).

Factors:

Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id	local_sample_id	Sample source	Factor
SA409256	PE_dn1_CTR_day07_03	Cells	PE_dn1_CTR
SA409257	PE_dn1_CTR_day0_02	Cells	PE_dn1_CTR
SA409258	PE_dn1_CTR_day21_03	Cells	PE_dn1_CTR
SA409259	PE_dn1_CTR_day21_02	Cells	PE_dn1_CTR
SA409260	PE_dn1_CTR_day21_01	Cells	PE_dn1_CTR
SA409261	PE_dn1_CTR_day14_03	Cells	PE_dn1_CTR
SA409262	PE_dn1_CTR_day14_02	Cells	PE_dn1_CTR
SA409263	PE_dn1_CTR_day14_01	Cells	PE_dn1_CTR
SA409264	PE_dn1_CTR_day0_01	Cells	PE_dn1_CTR
SA409265	PE_dn1_CTR_day07_02	Cells	PE_dn1_CTR
SA409266	PE_dn1_CTR_day04_03	Cells	PE_dn1_CTR
SA409267	PE_dn1_CTR_day04_02	Cells	PE_dn1_CTR
SA409268	PE_dn1_CTR_day04_01	Cells	PE_dn1_CTR
SA409269	PE_dn1_CTR_day01_03	Cells	PE_dn1_CTR
SA409270	PE_dn1_CTR_day01_02	Cells	PE_dn1_CTR
SA409271	PE_dn1_CTR_day07_01	Cells	PE_dn1_CTR
SA409272	PE_dn1_CTR_day01_01	Cells	PE_dn1_CTR
SA409273	PE_dn1_OI_day14_01	Cells	PE_dn1_OI
SA409274	PE_dn1_OI_day21_03	Cells	PE_dn1_OI
SA409275	PE_dn1_OI_day21_02	Cells	PE_dn1_OI
SA409276	PE_dn1_OI_day21_01	Cells	PE_dn1_OI
SA409277	PE_dn1_OI_day14_03	Cells	PE_dn1_OI
SA409278	PE_dn1_OI_day14_02	Cells	PE_dn1_OI
SA409279	PE_dn1_OI_day07_03	Cells	PE_dn1_OI
SA409280	PE_dn1_OI_day07_02	Cells	PE_dn1_OI
SA409281	PE_dn1_OI_day07_01	Cells	PE_dn1_OI
SA409282	PE_dn1_OI_day04_03	Cells	PE_dn1_OI
SA409283	PE_dn1_OI_day04_02	Cells	PE_dn1_OI
SA409284	PE_dn1_OI_day04_01	Cells	PE_dn1_OI
SA409285	PE_dn1_OI_day01_03	Cells	PE_dn1_OI
SA409286	PE_dn1_OI_day01_02	Cells	PE_dn1_OI
SA409287	PE_dn1_OI_day01_01	Cells	PE_dn1_OI
SA409288	PE_dn2_CTR_day07_03	Cells	PE_dn2_CTR
SA409289	PE_dn2_CTR_day21_03	Cells	PE_dn2_CTR
SA409290	PE_dn2_CTR_day21_02	Cells	PE_dn2_CTR
SA409291	PE_dn2_CTR_day21_01	Cells	PE_dn2_CTR
SA409292	PE_dn2_CTR_day14_03	Cells	PE_dn2_CTR
SA409293	PE_dn2_CTR_day14_02	Cells	PE_dn2_CTR
SA409294	PE_dn2_CTR_day14_01	Cells	PE_dn2_CTR
SA409295	PE_dn2_CTR_day07_02	Cells	PE_dn2_CTR
SA409296	PE_dn2_CTR_day07_01	Cells	PE_dn2_CTR
SA409297	PE_dn2_CTR_day01_02	Cells	PE_dn2_CTR
SA409298	PE_dn2_CTR_day0_02	Cells	PE_dn2_CTR
SA409299	PE_dn2_CTR_day0_01	Cells	PE_dn2_CTR
SA409300	PE_dn2_CTR_day01_01	Cells	PE_dn2_CTR
SA409301	PE_dn2_CTR_day0_03	Cells	PE_dn2_CTR
SA409302	PE_dn2_CTR_day01_03	Cells	PE_dn2_CTR
SA409303	PE_dn2_CTR_day04_01	Cells	PE_dn2_CTR
SA409304	PE_dn2_CTR_day04_02	Cells	PE_dn2_CTR
SA409305	PE_dn2_CTR_day04_03	Cells	PE_dn2_CTR
SA409306	PE_dn2_OI_day07_01	Cells	PE_dn2_OI
SA409307	PE_dn2_OI_day21_01	Cells	PE_dn2_OI
SA409308	PE_dn2_OI_day14_03	Cells	PE_dn2_OI
SA409309	PE_dn2_OI_day14_02	Cells	PE_dn2_OI
SA409310	PE_dn2_OI_day14_01	Cells	PE_dn2_OI
SA409311	PE_dn2_OI_day07_03	Cells	PE_dn2_OI
SA409312	PE_dn2_OI_day07_02	Cells	PE_dn2_OI
SA409313	PE_dn2_OI_day04_03	Cells	PE_dn2_OI
SA409314	PE_dn2_OI_day04_02	Cells	PE_dn2_OI
SA409315	PE_dn2_OI_day04_01	Cells	PE_dn2_OI
SA409316	PE_dn2_OI_day01_03	Cells	PE_dn2_OI
SA409317	PE_dn2_OI_day01_02	Cells	PE_dn2_OI
SA409318	PE_dn2_OI_day01_01	Cells	PE_dn2_OI
SA409319	PE_dn2_OI_day0_03	Cells	PE_dn2_OI
SA409320	PE_dn2_OI_day0_02	Cells	PE_dn2_OI
SA409321	PE_dn2_OI_day0_01	Cells	PE_dn2_OI
SA409322	PE_dn3_CTR_day07_02	Cells	PE_dn3_CTR
SA409323	PE_dn3_CTR_day21_03	Cells	PE_dn3_CTR
SA409324	PE_dn3_CTR_day21_02	Cells	PE_dn3_CTR
SA409325	PE_dn3_CTR_day21_01	Cells	PE_dn3_CTR
SA409326	PE_dn3_CTR_day14_03	Cells	PE_dn3_CTR
SA409327	PE_dn3_CTR_day14_02	Cells	PE_dn3_CTR
SA409328	PE_dn3_CTR_day14_01	Cells	PE_dn3_CTR
SA409329	PE_dn3_CTR_day07_03	Cells	PE_dn3_CTR
SA409330	PE_dn3_CTR_day01_02	Cells	PE_dn3_CTR
SA409331	PE_dn3_CTR_day07_01	Cells	PE_dn3_CTR
SA409332	PE_dn3_CTR_day04_02	Cells	PE_dn3_CTR
SA409333	PE_dn3_CTR_day04_01	Cells	PE_dn3_CTR
SA409334	PE_dn3_CTR_day01_03	Cells	PE_dn3_CTR
SA409335	PE_dn3_CTR_day0_02	Cells	PE_dn3_CTR
SA409336	PE_dn3_CTR_day01_01	Cells	PE_dn3_CTR
SA409337	PE_dn3_CTR_day0_03	Cells	PE_dn3_CTR
SA409338	PE_dn3_CTR_day04_03	Cells	PE_dn3_CTR
SA409339	PE_dn3_CTR_day0_01	Cells	PE_dn3_CTR
SA409340	PE_dn3_OI_day07_03	Cells	PE_dn3_OI
SA409341	PE_dn3_OI_day21_03	Cells	PE_dn3_OI
SA409342	PE_dn3_OI_day21_02	Cells	PE_dn3_OI
SA409343	PE_dn3_OI_day21_01	Cells	PE_dn3_OI
SA409344	PE_dn3_OI_day14_03	Cells	PE_dn3_OI
SA409345	PE_dn3_OI_day14_01	Cells	PE_dn3_OI
SA409346	PE_dn3_OI_day14_02	Cells	PE_dn3_OI
SA409347	PE_dn3_OI_day07_02	Cells	PE_dn3_OI
SA409348	PE_dn3_OI_day04_03	Cells	PE_dn3_OI
SA409349	PE_dn3_OI_day04_02	Cells	PE_dn3_OI
SA409350	PE_dn3_OI_day04_01	Cells	PE_dn3_OI
SA409351	PE_dn3_OI_day01_03	Cells	PE_dn3_OI
SA409352	PE_dn3_OI_day01_02	Cells	PE_dn3_OI
SA409353	PE_dn3_OI_day07_01	Cells	PE_dn3_OI
SA409354	PE_dn3_OI_day01_01	Cells	PE_dn3_OI
SA409355	EXO_dn1_OI_day0_Br	Media	EXO_dn1_Br

Showing page 1 of 3 Results: 1 2 3 Next Showing results 1 to 100 of 237

Collection:

Collection ID:	CO003891
Collection Summary:	For all donors, cryopreserved hAMSC were thawed, plated in culture flasks (T175), expanded in minimum essential alpha medium (α-MEM, Gibco™ 12000063, Waltham, MA, USA) supplemented with 10% v/v heat-inactivated fetal bovine serum (FBS, Gibco 10270106) and 1% v/v antibiotics (penicillin−streptomycin, Gibco 15240062) at 37°C in a humidified 5% CO2 incubator and passaged as described previously (Bispo et al. 2022a, https://doi.org/10.3390/cells11081257). Osteodifferentiation of hAMSC was induced at passages 5 or 6, for 21 days, in T175 culture flasks (for metabolomics) and in 48-well plates (for biochemical assays). An independent experiment was conducted for each of the donors. For metabolomics, cells were seeded at a 0.5 × 106 cells/flask density. After reaching 100% confluence, day 0 (D0) cell samples were collected in triplicate from independent flasks, and media were replaced in the remaining flasks as follows: 15 control (CTR) flasks in standard growth medium and 15 osteoinduced (OI) flasks in medium supplemented with 10 mM β-glycerophosphate (β-GP, Sigma-Aldrich G9422), 50 μg/mL L-ascorbic acid (Sigma A0278) and 10 nM dexamethasone (ACROS Organics 230300010). Media were replaced 2× per week, on days (Di) 0, 4, 7, 11, 14, 18 and 21. Cells were trypsinized and collected in triplicate on D0, D1, D4, D7, D14 and D21. Cell suspensions were filtered through 100 μm pore strainers, centrifuged (300 g, 4 °C, 5 min) and rinsed twice in phosphate-buffered saline (PBS) solution. For intracellular metabolomics (endometabolomics), cell numbers/sample were 3.8-21.1 × 106, 2.3-8.6 × 106 and 1.4-9.1 × 106 for donors 1, 2 and 3, respectively. For extracellular metabolomics (exometabolomics), media samples were collected on D1, D4, D7, D11, D14, D18 and D21 (days when cell collection and/or medium exchange was carried out) and filtered through 40 μm pore strainers to remove cellular debris. Due to contamination and/or technical issues, triplicate samples could not always be retrieved (donor 1: CTR D0 cells n = 2, OI D7 media n = 0; donor 2: CTR D7,D18 media n = 2, OI D11,D21 media n = 1-2, OI D21 cells n = 1). For biochemical assays, cell samples were rinsed 2× with PBS and lysed by osmotic/thermal shock. Collected media and cell samples were stored at − 80 °C. For osteoblast samples (OST), Cryopreserved Clonetics™ normal human osteoblasts (NHOst) from a healthy 26-year-old Caucasian male (Lonza, CC-2538, Lot 19TL217387) were subcultured using ReagentPack™ subculture reagents (Lonza, CC-5034) up to passage 5. Cells were maintained in Clonetics™ OGM™ Osteoblast media (Lonza, CC-3208) with OGM SingleQuot Kit supplements and growth factors (Lonza, CC-4193) for either 6 days (100% confluence) or 13 days, followed by cell collection.
Collection Protocol Filename:	hASCs_Experimental_Procedure.pdf
Sample Type:	Cell and cell media
Storage Conditions:	-80℃

Treatment:

Treatment ID:	TR003907
Treatment Summary:	For all donors, cryopreserved hAMSC were thawed, plated in culture flasks (T175), expanded in minimum essential alpha medium (α-MEM, Gibco™ 12000063, Waltham, MA, USA) supplemented with 10% v/v heat-inactivated fetal bovine serum (FBS, Gibco 10270106) and 1% v/v antibiotics (penicillin−streptomycin, Gibco 15240062) at 37°C in a humidified 5% CO2 incubator and passaged as described previously (Bispo et al. 2022, https://doi.org/10.3390/cells11081257). Osteodifferentiation of hAMSC was induced at passages 5 or 6, for 21 days, in T175 culture flasks (for metabolomics) and in 48-well plates (for biochemical assays). An independent experiment was conducted for each of the donors. For metabolomics, cells were seeded at a 0.5 × 106 cells/flask density. After reaching 100% confluence, day 0 (D0) cell samples were collected in triplicate from independent flasks, and media were replaced in the remaining flasks as follows: 15 control (CTR) flasks in standard growth medium and 15 osteoinduced (OI) flasks in medium supplemented with 10 mM β-glycerophosphate (β-GP, Sigma-Aldrich G9422), 50 μg/mL L-ascorbic acid (Sigma A0278) and 10 nM dexamethasone (ACROS Organics 230300010). Media were replaced 2× per week, on days (Di) 0, 4, 7, 11, 14, 18 and 21. Cells were trypsinized and collected in triplicate on D0, D1, D4, D7, D14 and D21. Cell suspensions were filtered through 100 μm pore strainers, centrifuged (300 g, 4 °C, 5 min) and rinsed twice in phosphate-buffered saline (PBS) solution. For intracellular metabolomics (endometabolomics), cell numbers/sample were 3.8-21.1 × 106, 2.3-8.6 × 106 and 1.4-9.1 × 106 for donors 1, 2 and 3, respectively. For extracellular metabolomics (exometabolomics), media samples were collected on D1, D4, D7, D11, D14, D18 and D21 (days when cell collection and/or medium exchange was carried out) and filtered through 40 μm pore strainers to remove cellular debris. Due to contamination and/or technical issues, triplicate samples could not always be retrieved (donor 1: CTR D0 cells n = 2, OI D7 media n = 0; donor 2: CTR D7,D18 media n = 2, OI D11,D21 media n = 1-2, OI D21 cells n = 1). For biochemical assays, cell samples were rinsed 2× with PBS and lysed by osmotic/thermal shock. Collected media and cell samples were stored at − 80 °C.
Treatment Protocol Filename:	hASCs_Experimental_Procedure.pdf
Treatment Protocol Comments:	After reaching 100% confluence, day 0 (D0) cell samples were collected in triplicate from independent flasks, and media were replaced in the remaining flasks as follows: 15 control (CTR) flasks in standard growth medium and 15 osteoinduced (OI) flasks in medium supplemented with 10 mM β-glycerophosphate (β-GP, Sigma-Aldrich G9422), 50 μg/mL L-ascorbic acid (Sigma A0278) and 10 nM dexamethasone (ACROS Organics 230300010).
Cell Media:	15 control (CTR) flasks in standard growth medium; and 15 osteoinduced (OI) flasks in medium supplemented with 10 mM β-glycerophosphate (β-GP, Sigma-Aldrich G9422), 50 μg/mL L-ascorbic acid (Sigma A0278) and 10 nM dexamethasone (ACROS Organics 230300010).
Cell Media Lastchanged:	Media were replaced 2× per week, on days (Di) 0, 4, 7, 11, 14, 18 and 21.

Sample Preparation:

Sampleprep ID:	SP003904
Sampleprep Summary:	To prepare samples for NMR analysis, intracellular metabolites (endometabolites) were extracted using a methanol-chloroform-water method, as described in Bispo et al. 2022b (https://doi.org/10.3390/cells11233745). Briefly, cell pellets were re-suspended in 1 mL of a cold solution of methanol (Honeywell Riedel-de-Haen 14262) and Milli-Q water (in a 4:1 ratio), transferred to glass tubes containing 150 mg of glass beads (ø = 0.5 mm), and vortexed for 2 min at room temperature (RT 25 °C). Cold chloroform (400 μL, Honeywell Riedel-de-Haen 650471) was then added and vortexed for 2 min at RT, followed by 400 μL of cold chloroform plus 360 μL of cold Milli-Q water with further vortexing for 2 min at RT. After incubating at −20 °C for 10 min, samples were centrifuged (2000 g, 20 min, RT) and the polar extracts (PE) were collected, dried, and stored at − 80 °C until analysis. For NMR analysis, PE samples were re-suspended in 625 μL of 100 mM phosphate buffer (pH 7.4), in D2O (99.9% deuterium, Eurisotop D216) containing 0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid (TSP, in D2O, Sigma-Aldrich 293040) for chemical shift referencing. Extracellular metabolites (exometabolites) were measured in cell media, upon protein-precipitation of blank (Br) and conditioned media as described in Bispo et al. 2022a (https://doi.org/10.3390/cells11081257). Briefly, 600 µL of 100% methanol at – 80 °C were added to microcentrifuge tubes containing 300 µL of medium sample. After incubating at – 20 °C for 30 min, samples were centrifuged (13000 g, 20 min, RT), the supernatant was collected, dried under vacuum and stored at – 80 °C. To prepare samples for NMR analysis, cellular polar extracts were re-suspended in 625 μL of 100 mM phosphate buffer (pH 7.4), in D2O (99.9% deuterium, Eurisotop D216) containing 0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid (TSP, in D2O, Sigma-Aldrich 293040) for chemical shift referencing. For exometabolome analysis, dry media samples were resuspended in 700 µL of the same phosphate buffer, centrifuged (13000 g, 5 min, RT) and the supernatant was collected. A volume of 550 μL from each sample (pH adjusted to 7.4) was transferred into a 5 mm NMR tube.
Sampleprep Protocol Filename:	hASCs_Experimental_Procedure.pdf
Processing Storage Conditions:	-80℃
Extraction Method:	Intracellular metabolites (endometabolites) were extracted using a water/methanol/chloroform method, as described in (Bispo et al., 2022, https://doi.org/10.3390/cells11233745). Extracellular metabolites (exometabolites) were measured in cell media, upon protein-precipitation of blank and conditioned media
Extract Storage:	-80℃
Sample Resuspension:	Cellular polar extracts were re-suspended in 625 μL of 100 mM phosphate buffer (pH 7.4), in D2O (99.9% deuterium, Eurisotop D216) containing 0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid (TSP, in D2O, Sigma-Aldrich 293040) for chemical shift referencing. For exometabolome analysis, dry media samples were resuspended in 700 µL of the same phosphate buffer, centrifuged (13000 g, 5 min, RT) and the supernatant was collected. A volume of 550 μL from each sample (pH adjusted to 7.4) was transferred into a 5 mm NMR tube.
Sample Spiking:	0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid (TSP, in D2O, Sigma-Aldrich 293040) , as a chemical shift reference.

Analysis:

Analysis ID:	AN006180
Laboratory Name:	Metabolomics Group
Analysis Type:	NMR
Analysis Protocol File:	hASCs_Experimental_Procedure.pdf
Acquisition Parameters File:	hASCs_Experimental_Procedure.pdf
Software Version:	TopSpin3.2 and Amix3.9.14
Operator Name:	Daniela Bispo
Detector Type:	NMR
Data Format:	fid, 1r
Results File:	ST003765_AN006180_Results.txt
Units:	ppm

NMR:

NMR ID:	NM000309
Analysis ID:	AN006180
Instrument Name:	NMR 500 Bruker Avance III
Instrument Type:	FT-NMR
NMR Experiment Type:	1D-1H
Spectrometer Frequency:	500.13 MHz
NMR Probe:	TXI probe
NMR Solvent:	100 mM phosphate buffer (pH 7.4), in D2O (99.9% deuterium, Eurisotop D216) containing 0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid (TSP, in D2O, Sigma-Aldrich 293040), for chemical shift referencing.
NMR Tube Size:	5 mm NMR tubes
Shimming Method:	Topshim
Pulse Sequence:	noesypr1d
Water Suppression:	presat
Pulse Width:	90-degree
Chemical Shift Ref Cpd:	3-(trimethylsilyl)propionic-2,2,3,3-d4 acid (TSP-d4)
Temperature:	25
Number Of Scans:	256 scans (exometabolome) and 512 (endometabolome)
Dummy Scans:	8
Acquisition Time:	2.33 s
Relaxation Delay:	4 s
Spectral Width:	7,002.8 Hz
Num Data Points Acquired:	32 k points
Line Broadening:	0.3 Hz
Zero Filling:	128k (exometabolome) or 64 k points (endometabolome)
Apodization:	Exponential
Baseline Correction Method:	Manual
Chemical Shift Ref Std:	0 ppm for TSP-d4