Summary of Study ST000121
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 PR000108. The data can be accessed directly via it's Project DOI: 10.21228/M8GW2Z This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST000121 |
| Study Title | Impact of anesthesia and euthanasia on metabolomics of mammalian tissues: studies in a C57BL/6J mouse model |
| Study Type | Anesthesia effect study |
| Study Summary | We examined the effect of several commonly-used methods of anesthesia and euthanasia for collection of skeletal muscle, liver, heart, adipose and serum of C57BL/6J mice. The data revealed tissue-specific impacts of different anesthesia and euthanasia strategies. Based on these findings, we present a more optimal collection strategy mammalian tissues and recommend that rodent tissues intended for metabolomics studies be collected under anesthesia rather than post-euthanasia. |
| Institute | University of Michigan |
| Department | Internal Medicine |
| Laboratory | Evans Lab / Burant Lab |
| Last Name | Evans |
| First Name | Charles |
| Address | 6321 Brehm Tower, 1000 Wall Street, Ann Arbor MI 48105 |
| chevans@umich.edu | |
| Phone | 734-232-8177 |
| Submit Date | 2014-11-21 |
| Num Groups | 6 |
| Total Subjects | 49 |
| Raw Data Available | No |
| Raw Data File Type(s) | d |
| Analysis Type Detail | LC-MS |
| Release Date | 2015-02-01 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR000108 |
| Project DOI: | doi: 10.21228/M8GW2Z |
| Project Title: | Impact of anesthesia and euthanasia on metabolomics of mammalian tissues: studies in a C57BL/6J mouse model |
| Project Type: | Anesthesia effect study |
| Project Summary: | We examined, for the first time using untargeted and targeted metabolomics, the effect of several commonly-used methods of anesthesia and euthanasia for collection of skeletal muscle, liver, heart, adipose and serum of C57BL/6J mice. The data revealed tissue-specific impacts of different anesthesia and euthanasia strategies. Based on these findings, we present a more optimal collection strategy mammalian tissues and recommend that rodent tissues intended for metabolomics studies be collected under anesthesia rather than post-euthanasia. |
| Institute: | University of Michigan |
| Department: | Internal Medicine |
| Laboratory: | Evans lab / Burant lab |
| Last Name: | Evans |
| First Name: | Charles |
| Address: | 6321 Brehm Tower, 1000 Wall St. |
| Email: | chevans@umich.edu |
| Phone: | 734-232-8177 |
| Funding Source: | National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), National Institutes of Health |
Subject:
| Subject ID: | SU000140 |
| Subject Type: | Animal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Genotype Strain: | C57BL/6J |
| Age Or Age Range: | 20 weeks |
| Weight Or Weight Range: | 26.8 +/- 2.2 g |
| Gender: | Male |
| Animal Animal Supplier: | Jackson Labs (Bar Harbor, ME) |
| Animal Housing: | 8 / cage |
| Animal Light Cycle: | 12 h / 12 h light/dark |
| Animal Feed: | Standard chow, ad lib. |
| Animal Water: | Ad lib. Access |
| Species Group: | Mammals |
Factors:
Subject type: Animal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Tissue | Anesthesia / Euthanasia method |
|---|---|---|---|
| SA006504 | F14 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006505 | F16 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006506 | F17 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006507 | F13 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006508 | F15 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006509 | F12 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006510 | F09 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006511 | F10 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006512 | F11 | Epididymal white adipose (fat) | Carbon Dioxide |
| SA006513 | F23 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006514 | F24 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006515 | F22 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006516 | F18 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006517 | F25 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006518 | F19 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006519 | F20 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006520 | F21 | Epididymal white adipose (fat) | Cervical Dislocation |
| SA006521 | F40 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006522 | F35 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006523 | F37 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006524 | F38 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006525 | F41 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006526 | F39 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006527 | F34 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006528 | F36 | Epididymal white adipose (fat) | Continuous isoflurane |
| SA006529 | F05 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006530 | F06 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006531 | F07 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006532 | F04 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006533 | F08 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006534 | F03 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006535 | F01 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006536 | F02 | Epididymal white adipose (fat) | Isoflurane overdose |
| SA006537 | F28 | Epididymal white adipose (fat) | Ketamine |
| SA006538 | F26 | Epididymal white adipose (fat) | Ketamine |
| SA006539 | F29 | Epididymal white adipose (fat) | Ketamine |
| SA006540 | F27 | Epididymal white adipose (fat) | Ketamine |
| SA006541 | F30 | Epididymal white adipose (fat) | Ketamine |
| SA006542 | F33 | Epididymal white adipose (fat) | Ketamine |
| SA006543 | F32 | Epididymal white adipose (fat) | Ketamine |
| SA006544 | F31 | Epididymal white adipose (fat) | Ketamine |
| SA006545 | F46 | Epididymal white adipose (fat) | Pentobarbital |
| SA006546 | F47 | Epididymal white adipose (fat) | Pentobarbital |
| SA006547 | F49 | Epididymal white adipose (fat) | Pentobarbital |
| SA006548 | F45 | Epididymal white adipose (fat) | Pentobarbital |
| SA006549 | F43 | Epididymal white adipose (fat) | Pentobarbital |
| SA006550 | F42 | Epididymal white adipose (fat) | Pentobarbital |
| SA006551 | F48 | Epididymal white adipose (fat) | Pentobarbital |
| SA006552 | F44 | Epididymal white adipose (fat) | Pentobarbital |
| SA006553 | G17 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006554 | G13 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006555 | G12 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006556 | G11 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006557 | G09 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006558 | G16 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006559 | G15 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006560 | G14 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006561 | G10 | Gastrocnemius (skeletal muscle) | Carbon Dioxide |
| SA006562 | G18 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006563 | G23 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006564 | G24 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006565 | G22 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006566 | G21 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006567 | G19 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006568 | G20 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006569 | G25 | Gastrocnemius (skeletal muscle) | Cervical Dislocation |
| SA006570 | G34 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006571 | G39 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006572 | G40 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006573 | G38 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006574 | G37 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006575 | G35 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006576 | G36 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006577 | G41 | Gastrocnemius (skeletal muscle) | Continuous isoflurane |
| SA006578 | G08 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006579 | G02 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006580 | G01 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006581 | G03 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006582 | G04 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006583 | G06 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006584 | G07 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006585 | G05 | Gastrocnemius (skeletal muscle) | Isoflurane overdose |
| SA006586 | G30 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006587 | G31 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006588 | G29 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006589 | G28 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006590 | G26 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006591 | G27 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006592 | G32 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006593 | G33 | Gastrocnemius (skeletal muscle) | Ketamine |
| SA006594 | G47 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006595 | G48 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006596 | G49 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006597 | G42 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006598 | G46 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006599 | G45 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006600 | G43 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006601 | G44 | Gastrocnemius (skeletal muscle) | Pentobarbital |
| SA006602 | H16 | Heart | Carbon Dioxide |
| SA006603 | H17 | Heart | Carbon Dioxide |
Collection:
| Collection ID: | CO000124 |
| Collection Summary: | 4 tissues plus blood serum were collected and immediately following euthanasia or anesthesia induction, frozen in liquid nitrogen and stored at -80C until analysis |
| Sample Type: | Skeletal muscle (gastrocnemius), Liver, Heart, Epididymal white adipose (fat), blood serum |
| Collection Method: | Surgical dissection |
| Collection Time: | As soon as possible following anesthesia/euthanasia (all tissues collected within 3 minutes) |
| Storage Conditions: | -80 C |
Treatment:
| Treatment ID: | TR000142 |
| Treatment Summary: | Mice were prepared or tissue collection by one of 6 modes of anesthesia or euthanasia, then tissues were rapidly surgically collected and frozen |
| Animal Anesthesia: | One of 6 methods: 1) Cervical dislocation, 2) Carbon dioxide inhalation, 3) Isoflurane overdose, 4) continuous isoflurane, 5) ketamine, 6) pentobarbital |
| Animal Fasting: | 5 hours |
| Animal Endp Euthanasia: | All tissue collection were terminal studies; in methods 1-3, animals were euthanized prior to tissue collection; in methods 4-6, animals were under deep anesthesia while tissues were collected. |
Sample Preparation:
| Sampleprep ID: | SP000137 |
| Sampleprep Summary: | Frozen tissue samples were pulverized in a liquid nitrogen-chilled mortar and pestle, then sonicated in cold extraction solvent (7:2:1 methanol:chloroform:water) at an approximate ratio of 30 mg tissue (wet mass) /1 mL of solvent. After allowing samples to rest on ice 5 minutes, they were centrifuged to pellet precipitated protein and cell debris; the supernatant was placed in autosampler vials for direct LC-MS analysis. For serum, extraction solvent (1:1:1 methanol:acetonitrile:acetone) was added to serum at a 4:1 solvent:serum ratio, vortexed to mix, then centrifuged and supernatant recovered for LC-MS analysis. |
| Sampleprep Protocol Filename: | PROTOCOLS_mouse_anesthesia.pdf |
| Processing Method: | Cryo-pulverization + sonication in extraction solvent |
Chromatography:
| Chromatography ID: | CH000135 |
| Chromatography Summary: | Polar metabolites were analyzed by HILIC-MS using an aminopropyl stationary phase |
| Instrument Name: | Agilent 1200 |
| Column Name: | Phenomenex Luna NH2 (150 x 2.1mm,3um) |
| Column Pressure: | 150 bar |
| Column Temperature: | 25 C |
| Flow Gradient: | 80-0% B over 20 minutes, hold 4 minutes, return to 80%B |
| Flow Rate: | 0.25 mL/min |
| Sample Injection: | 25 uL |
| Solvent A: | 100% water; 5 mM ammonium acetate, pH 9.9 |
| Solvent B: | 100% acetonitrile |
| Analytical Time: | 32 min |
| Chromatography Type: | HILIC |
Analysis:
| Analysis ID: | AN000203 |
| Laboratory Name: | Michigan Regional Comprehensive Metabolomics Resource Core |
| Analysis Type: | MS |
| Analysis Comments: | NEGATIVE |
| Instrument Name: | Agilent 6220 TOF MS |
| Software Version: | Masshunter 4.0 |
| Operator Name: | Charles Evans |
| Detector Type: | Time-of Flight |
| Chromatography ID: | CH000135 |
| Num Factors: | 30 |
| Num Metabolites: | 128 |
| Units: | Peak area |