Guidelines | BioDiscovery Institute

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Guidelines

General GC/LC-MS Preparation Guideline

The following guidelines should be followed when you prepare your samples for mass spectrometry.

Should you have any questions on what may or may not be used, please contact the BAF manager (Jeanchristophe.Cocuron@unt.edu).

LC-MS/MS Sample Preparation Guideline

Rules for LC-MS sample preparation

  • Samples MUST BE filtered to a 0.22 micron filter or 3 kDa Amicon filtering device prior to the submission to BAF.
  • Samples MUST BE completely dissolved prior to the injection through the mass spectrometer or LC-MS system.
  • For ESI and APCI analysis, please submit at least 50 µL or up to 100 µmol/L of biological sample solutions, and at least 5 mg of material for dried samples (standard or biological sample).
  • The liquid samples should be submitted in polypropylene vials or deactivated glass vials with PTFE or polypropylene-lined screw caps. Use a small volume insert (300 or 500 µL) if a limited amount of sample is available. For dried material, submit samples in 2 mL screw cap tubes.
  • All sample tubes/vials MUST BE labeled with a sample ID, requester name, and must be accompanied by the sample submission form and a form (Word or Excel document) listing the samples that will be analyzed.

Rules for LC/MS Solvents and Modifiers

Safe solvents (volatile, low molecular weight protic):

  • Acetonitrile
  • Methanol
  • Ethanol
  • Isopropanol
  • Water

Safe mobile phase modifiers:

  • Acetic acid (up to 1.0 %)
  • Formic acid (up to 0.1 %)
  • Ammonium hydroxide (up to 0.1 %)
  • Ammonium formate (salt concentration = 10 mM or less)
  • Ammonium acetate (salt concentration = 10 mM or less)

DO NOT USE

  • Nonvolatile salts (Tris, phosphates, citrates, and HEPES) can deposit in the source and plug capillaries.
  • Surfactants/Detergents such as SDS, CHAPS, PEG, Tween, and Triton that suppress electrospray ionization.
  • Inorganic acids (Sulfuric, nitric, and hydrochloric acids), which are corrosive and destructive for the LC/MS system.
  • DMSO, DMF, THF, acelate, and glycerol.
  • Trifluoroacetic acid (TFA) suppresses positive-ion electrospray at levels exceeding 0.01 %. Commonly used alternatives to high concentrations of TFA are mixtures of either 1 % acetic or 0.1 % formic acid with 0.025% TFA.
  • Hydrocarbon solvents (hexane and benzene) cannot be used for ESI. Triethylamine (TEA) yields an intense [M+H]+ ion at m/z 102 and suppresses positive-ion electrospray of less basic ions.

GC-MS Sample Preparation Guideline

Rules for GC/MS Solvents

Safe solvents (high volatility) for sample preparation:

  • Acetone
  • Acetonitrile
  • Benzene
  • Ethers
  • Hexanes
  • Methanol
  • Methylene chloride

DO NOT USE/INJECT

  • Water, DMSO, DMF because they will damage the inlet of the GC system as well as the column. Therefore, DO NOT RESUSPEND your samples in water, DMSO or DMF.
  • Metals, strong acids or bases, salts, oligomers, and polymers should not be injected through the GC-MS system. These compounds will damage the inlet of the GC, the column, and the EI source used for the ionization of your metabolites.

Rules for GC-MS sample preparation

  • Try to minimize the use of "plasticizers" (tips, tubes…) when you perform your sample extraction and derivatization. This will help in minimizing contaminant peaks on your GC-MS chromatogram. Pipette tips can be replaced by Hamilton syringes.
  • Samples MUST BE filtered to a 0.22 micron filter or a 3 kDa Amicon filtering device prior to the submission to BAF.
  • Samples MUST BE volatile and completely dissolved in a non-polar solvent prior liquid injection through the GC-MS system.
  • The molecular weight from your metabolite(s) of interest has to be under 1,100 amu.
  • For liquid injection analysis, please submit at least 50 µL or up to 1000 µmol/L of derivatized biological sample solution, and at least 5 mg of material for dried samples (standard or biological sample).
  • For headspace or solid phase micro-extraction (SPME) injection analysis, liquid, wet/dried samples can be prepared directly in 20 mL screw cap vials and run through the GC-MS system as they are.
  • The liquid samples should be submitted in polypropylene vials or deactivated glass vials with PTFE or polypropylene-lined screw caps. Use a small volume insert (300 or 500 µL) if a limited amount of sample is available. For dried material submit samples in 2 mL screw cap tubes.
  • All sample tubes/vials MUST BE labeled with a sample ID, requester name, and must be accompanied by the sample submission form and a form (Word or Excel document) listing the samples that will be analyzed.

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