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7 protocols using pentafluorobenzyl bromide

1

Isotopically Labeled Compounds Analysis

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All solvents were GC/MS and/or HPLC grade, purchased from Sigma-Aldrich. Pentafluorobenzyl bromide was purchased from Sigma-Aldrich. L-[5,5,5-2H3]Leucine and [3,4,methyl-13C3]HMB were purchased from Cambridge Isotopes.
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2

Comprehensive Lipid Extraction and Quantification

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Acetonitrile (ACN) was HPLC-grade and purchased from Acros Organics (Geel, Belgium). Methanol HPLC-grade (MeOH), Dichloromethane (CH2Cl2), Ammonium Formate (>99%) (AF), Boron trifluoride-methanol solution 14% (BF3-MeOH), Heptane, Ethyl acetate (EtOAc), potassium hydroxide (KOH), pentafluorobenzyl bromide (PFB-Br), and diisopropylethylamine (DIPEA), iodoacetamide, ammonium bicarbonate, trifluoroacetic acid, and trypsin was supplied by Sigma Aldrich Chemicals Co. (Saint Quentin Fallavier, France), acetic acid (AA) from Honeywell Fluka. Ultrapure water (18.2 MΩ) was obtained from a milliQ apparatus from Millipore (Guyancourt, France).
Internal synthetic standards of phospholipids (PL: PE 12:0/12:0, PC 13:0/13:0, PS 12:0/12:0), Ceramides (Cer: Cer d18:1/15:0), sphingomyelins (SM: SM d18:1/12:0), and sphingosine (So: So d17:1) and sphinganine (Sa: Sa d17:0) were purchased from Avanti Polar Lipids (Alabaster, AL, USA). Synthetic internal standard PI 16:0/17:0 was supplied by J. Clark (Cambridge). Synthetic internal standards for neutral lipid (LN: stigmasterol, cholesteryl heptadecanoate, glyceryl trinonadecanoate) and for free FAs (FFA: heptadecanoate) and total FAs (TFA: glyceryl triheptadecanoate, glyceryl trinonadecanoate) were purchased from Sigma Aldrich (St Quentin Fallavier, France).
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3

Characterization of Chlorinated Phenols

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All chemicals were of the highest quality available. 2,4,5‐T and the other phenoxyacetic acid derivatives were obtained from Dr. Ehrenstorfer (Augsburg, Germany), Sigma‐Aldrich (Steinheim, Germany) or Acros Organics (Geel, Belgium). The 2‐, 3‐ and 4‐monochlorophenol (MCP) derivatives were obtained from Laborchemie (Apolda, Germany), Merck (Darmstadt, Germany) and Jenapharm (Jena, Germany) respectively. The dichlorophenols (DCP) were obtained from Applichem (Darmstadt, Germany), Merck and Dr. Ehrenstorfer. Deuterated standards and pentafluorobenzylbromide were purchased from Sigma, and N‐methyl‐N‐trimethylsilyl‐heptafluorobutyramide (MSHFBA) was purchased from Macherey‐Nagel (Düren, Germany).
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4

Sensitive Metabolite Quantification

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Ammonium formate solution, 5-sulfosalicilic acid, glacial acetic acid, dimethyl sulfoxide, N,N-diisopropylethylamine (DIPEA) and pentafluorobenzyl bromide (PFB) were purchased from Sigma-Aldrich (St Louis, MO, USA). Optima liquid chromatography/mass spectrometry grade methanol, acetonitrile (ACN), 2-propanol (IPA) and water were purchased from Fisher Scientific (Pittsburgh, PA, USA). 2-(2-pyridyl) ethyl functionalized silica gel 100 mg/1 ml tubes were obtained from Supelco Analytical (Bellefonte, PA, USA). Streptomycin and penicillin were purchased from Invitrogen. Charcoal-stripped FBS was from Gemini Bioproducts (West Sacramento, CA, USA). [13C315N1]-calcium pantothenate was purchased from Isosciences (King of Prussia, PA, USA).
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5

Synthesis and Analysis of Isotopically Labeled Compounds

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Sodium nitrate, sodium 15N‐nitrate, and pentafluorobenzyl bromide were purchased from Sigma Aldrich (Steinheim) each at the highest purity available. Acetone (GC grade), toluene (GC grade), and water were purchased from Merck (Darmstadt). Bovine serum was purchased from Fiebig Animalblood Products (Idstein‐Niederauroff).
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6

Metabolic Profiling of Cell Cultures

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Rotenone, 4,4,4-trifluoro-1-(2-thienyl)-1,3-butanedione (TTFA), 3-nitropropionic acid (3-NPA), malonate, 2,6-dichlorophenolindophenol, decylubiquinone, 2-(4,5-dimethyl-2-thiazolyl)-3,5-diphenyl-2H-tetrazolium bromide (MTT), phenazine methosulfate (PMS), carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP),[13C5,15N1]glutamate, methoxyamine HCl, diisopropylethylamine (DIPEA), pentafluorobenzyl bromide, 1,1,1,3,3,3-hexafluoro-2-propanol, dimethyl sulfoxide (DMSO), 2′,7′-dichlorofluorescin diacetate, N-acetylcysteine, propidium iodide (PI), succinate, [13C4]succinate, [13C3]lactate, [13C6]citrate, and [13C5,15N2]glutamine were purchased from Sigma-Aldrich. LND was purchased from Santa Cruz Biotechnology. Optima LC-MS grade water, methanol, acetonitrile, and isopropanol were purchased from Thermo Fisher Scientific (Waltham, MA). [13C4]Fumarate and glutathione ([13C2,15N1]glycine) were purchased from Cambridge Isotope Laboratories (Tewksbury, MA).
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7

Synthesis of Tetraphenylethylene Derivatives

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Sodium azide (99%), pentafluorobenzyl bromide (99%), acetone, n-hexane -gradient grade for gas chromatography, sodium tetraborate, copper (II) chloride, sodium thiosulphate (VI), sodium chloride, hydrochloric acid, tetraphenylethylene, sodium hydroxide -analytical grade were purchased from Sigma-Aldrich (Germany).
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