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MB Sample ID: SA218430

Local Sample ID:1024028927
Subject ID:SU002365
Subject Type:Fish
Subject Species:Oncorhynchus mykiss

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Subject:

Subject ID:SU002365
Subject Type:Fish
Subject Species:Oncorhynchus mykiss

Factors:

Local Sample IDMB Sample IDFactor Level IDLevel ValueFactor Name
1024028927SA218430FL0264676_weeksSample time
1024028927SA218430FL0264670,1% ß-glucanTreatment

Collection:

Collection ID:CO002358
Collection Summary:For the metabolite and microbiota analyses, 10 fish from the control group were collected on day 0 and 10 fish from each group (control, 0.1, 1.0, 5.0% β-glucan) at week 6. Fish were euthanized by immersion in an overdose (200 mg∙L-1) of MS-222 (cat no. A5040, Sigma-Aldrich). The tail was cut, and blood was sampled from the vena caudalis using Na-heparinised 25 mL and 50 mL capillary pipettes (Hirschmann Laborgeräte, Germany). Blood samples were centrifuged at 3,000 g for 10 min at 4 °C, and serum was isolated and stored at -80 °C. The intestine was aseptically sampled and immediately immersed in RNAlater® (Sigma-Aldrich) and transferred to -20 °C.
Sample Type:Blood (serum)
Storage Conditions:-20℃

Treatment:

Treatment ID:TR002377
Treatment Summary:A commercial pelleted (1.5 mm) trout feed (INICIO 917, BioMar A/S) based on a mixture of fish and plant protein and containing 47% crude protein, 20% crude lipid, 18% carbohydrates (NFE), 1.2% fibre, 8.5% ash, and 1.1% phosphorus was used for preparation of the diets. A β-1,3;1,6-glucan (purity: 81.6%; mean particle size: 37.7 µm; supplementary table S2) purified from yeast (Saccharomyces cerevisiae) (Biorigin, Brazil) was supplemented to this diet at varying doses: 0 g, 1 g, 10 g, and 50 g β-glucan were added to 1 kg of INICIO 917, respectively, during continuous mixing and sealed to the pellets by spraying with 30 mL organic, cold-pressed rapeseed oil (Gyldenmark), resulting in a control diet without β-glucan and three experimental diets of 0.1%, 1.0%, and 5.0% Wt/Wt β-glucan.

Sample Preparation:

Sampleprep ID:SP002371
Sampleprep Summary:Sample preparation for MetIDQ p180 Kit measurement Solvents: Acetonitril (Merck KGaA, Darmstadt, Germany hypergrade for LC-MS) Water MiliQ, Extracting agent - ACN / H2O (1:1) Equipment 4 steel balls size M Eppendorf Tubes 2mL Tissue slicer (Rettberg, Germany) Centrifuge (Sigma) Work steps Approximately 100 mg of sample and 4 steel balls of size M into each tube. Per mg of sample 5 µL of extracting agent was added. Shake the samples for 10 minutes at 30 Hz in the tissue slicer and centrifuge for 2 minutes at 14000 rpm. 10µL of supernatant was used for the targeted analytics. For blood sample analysis 10 µL plasma was taken. Kit reparation The analysis was performed using the validated MetIDQ p180 Kit and described in Siskos et al. [1]. Data processing was carried out with the provided quantitation method Kit (Biocrates Life Sciences AG, Innsbruck, Austria). 1 Siskos AP, Jain P, Römisch-Margl W, Bennett M, Achaintre D, Asad Y, et al. Interlaboratory Reproducibility of a Targeted Metabolomics Platform for Analysis of Human Serum and Plasma. Anal Chem 2017;89:656-65.
Sampleprep Protocol Filename:Sample_preparation.pdf

Combined analysis:

Analysis ID AN003723
Analysis type MS
Chromatography type Reversed phase
Chromatography system UPLC (Waters Acquity, Waters Corporation, Milford, USA)
Column Agilent Zorbax Eclipse XDB C18 (100 x 3.0mm,3.5um)
MS Type ESI
MS instrument type Triple quadrupole
MS instrument name ABI Sciex 5500 QTrap
Ion Mode POSITIVE
Units µM

Chromatography:

Chromatography ID:CH002757
Chromatography Summary:LC - Instrument Parameters AbsoluteIDQ® p180 Kit (Biocrates Life Science AG, Innsbruck, Austria) System: UPLC (Waters Acquity, Waters Corporation, Milford, USA) Column: Agilent, Zorbax Eclipse XDB C18, 3.0 x 100 mm, 3.5 µM, Agilent Waldbronn, Germany Precolumn: Security Guard, Phenomenex, C18, 4 x 3 mm; Phenomenex, Aschaffenburg, Germany Materials: Water MiliQ, Methanol (Merck KGaA, Darmstadt, Germany, hypergrade for LC-MS) Acetonitril (Merck KGaA, Darmstadt, Germany hypergrade for LC-MS) Ammonium Acetate (Honeywell - Fluka, Seelze, Germany) Formic Acid (Honeywell, Fluka, Seelze, Germany) Running Solvent: 5mM ammonium acetat in methanol FIA: B: Mix running solvent and MiliQ water 1:1 Gradient Table FIA Time (min) Flow Rate (mL/min) %A %B Curve Initial 0.030 0.0 100.0 Initial 1.60 0.030 0.0 100.0 6 2.40 0.200 0.0 100.0 6 2.80 0.200 0.0 100.0 6 3.00 0.030 0.0 100.0 6
Methods Filename:LC_parameters_amino_acids.pdf
Instrument Name:UPLC (Waters Acquity, Waters Corporation, Milford, USA)
Column Name:Agilent Zorbax Eclipse XDB C18 (100 x 3.0mm,3.5um)
Flow Gradient:FIA Time (min) Flow Rate (mL/min) %A %B Curve Initial 0.030 0.0 100.0 Initial 1.60 0.030 0.0 100.0 6 2.40 0.200 0.0 100.0 6 2.80 0.200 0.0 100.0 6 3.00 0.030 0.0 100.0 6
Flow Rate:0.03ml/min
Solvent A:100% methanol; 5mM ammonium acetate
Solvent B:50% methanol/50% water; 5mM ammonium acetate
Chromatography Type:Reversed phase

MS:

MS ID:MS003471
Analysis ID:AN003723
Instrument Name:ABI Sciex 5500 QTrap
Instrument Type:Triple quadrupole
MS Type:ESI
MS Comments:MS - Instrument Parameter AbsoluteIDQ® p180 Kit (Biocrates Life Science AG, Innsbruck, Austria) MS -Q-Trap 5500 Sciex Ion Source: Turbo Spray Curtian Gas: 20 CAD Gas: Medium Ion Spray Voltage: 5500 V Temperature: 500°C Ion Source Gas 1: 40psi Ion Source Gas 2: 50 psi
Ion Mode:POSITIVE
Analysis Protocol File:MS_amino_acids.pdf
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