Summary of Study ST002355

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 PR001512. The data can be accessed directly via it's Project DOI: 10.21228/M8812G 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.

Study ID	ST002355
Study Title	Stool short chain fatty acid (SCFA) levels in peanut allergy
Study Summary	Prior evidence supports differential levels of short chain fatty acids in the stool of human beings with allergy and murine models of allergy. Here we performed a targeted study of selected short chain fatty acid levels in stool samples collected from children with allergy risk factors. Sample processing included homogenization of stool samples, inclusion of internal standards, and derivitization for liquid chromatography tandem mass spectrometry.
Institute	Icahn School of Medicine at Mount Sinai
Last Name	Bunyavanich
First Name	Supinda
Address	1 Gustave L. Levy Pl, New York, NY 10029
Email	supinda.bunyavanich@mssm.edu
Phone	Stool metabolite levels in individuals with peanut allergy were measured.
Submit Date	2022-11-08
Raw Data Available	Yes
Raw Data File Type(s)	mzXML
Analysis Type Detail	LC-MS
Release Date	2023-12-08
Release Version	1

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Sample Preparation:

Sampleprep ID:	SP002450
Sampleprep Summary:	Stool samples were weighed into bead blaster tubes with zircon beads and homogenized in 100% methanol to a final concentration of 10mg/mL including deuterated SCFA internal standards Chun et al. p.22 (CDN Isotopes). 46 Control samples were created by combining sample homogenates into a 1mL of pooled sample (pooled control) or 1mL of methanol (instrument blank). A portion of the pooled control was then subjected to three successive rounds of extraction to deplete SCFAs but retain the insoluble particles to generate null matrix material. A standard curve for the cocktail of SCFAs was prepared in null matrix, and these samples were extracted side-by-side with the study samples and sequence blanks (either methanol or null matrix). The resulting SCFA extracts were then derivatized as described47 for liquid chromatography tandem mass spectrometry (LCMS) with methanol being used instead of acetonitrile. Samples were analyzed with a QExactive HF coupled to a Dionex Ultimate 3000. A Waters BEH C18 column (2.1 x 150mm, 1.7µm) was used with Buffer A as 0.1% formic acid in water, and Buffer B as 0.1%formic acid in acetonitrile. A gradient elution was used (15%B to 55%B in 9 min, 200µL/min) and the mass spectrometer was operated in negative ion mode (HESI, -3.5kV). The order of acquisition was randomized and several control blocks including blanks and external standards were interspersed throughout the run to assess instrument performance and quality control of the derivatization protocol. All analytes were corrected to their respective internal standard, and the resulting ratio was interpolated against the matrix-controlled standard curve to quantify the level of SCFA in each sample. The limit of detection was determined from null-matrix samples for each analyte, and this limit was imputed for any point falling below the limit of detection. The interpolated values were used for the downstream analyses, and the imputed ratio threshold values were used for samples below the limit of detection.

Sampleprep ID:

SP002450

Sampleprep Summary:

Stool samples were weighed into bead blaster tubes with zircon beads and homogenized in 100% methanol to a final concentration of 10mg/mL including deuterated SCFA internal standards Chun et al. p.22 (CDN Isotopes). 46 Control samples were created by combining sample homogenates into a 1mL of pooled sample (pooled control) or 1mL of methanol (instrument blank). A portion of the pooled control was then subjected to three successive rounds of extraction to deplete SCFAs but retain the insoluble particles to generate null matrix material. A standard curve for the cocktail of SCFAs was prepared in null matrix, and these samples were extracted side-by-side with the study samples and sequence blanks (either methanol or null matrix). The resulting SCFA extracts were then derivatized as described47 for liquid chromatography tandem mass spectrometry (LCMS) with methanol being used instead of acetonitrile. Samples were analyzed with a QExactive HF coupled to a Dionex Ultimate 3000. A Waters BEH C18 column (2.1 x 150mm, 1.7µm) was used with Buffer A as 0.1% formic acid in water, and Buffer B as 0.1%formic acid in acetonitrile. A gradient elution was used (15%B to 55%B in 9 min, 200µL/min) and the mass spectrometer was operated in negative ion mode (HESI, -3.5kV). The order of acquisition was randomized and several control blocks including blanks and external standards were interspersed throughout the run to assess instrument performance and quality control of the derivatization protocol. All analytes were corrected to their respective internal standard, and the resulting ratio was interpolated against the matrix-controlled standard curve to quantify the level of SCFA in each sample. The limit of detection was determined from null-matrix samples for each analyte, and this limit was imputed for any point falling below the limit of detection. The interpolated values were used for the downstream analyses, and the imputed ratio threshold values were used for samples below the limit of detection.