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NMR Sample Preparation

A well-prepared NMR sample is important for acquiring quality spectra.

  1. Choice of Solvent:
    NMR experiments are traditionally done in deuterated solvents, but you can collect usable spectra in non-deuterated solvents (no-D NMR) as well. When selecting an NMR solvent, consider:

    • Solubility
    • The solvent’s chemical shift. Will the residual solvent signal interfere with sample resonances?
    • Literature reports. Chemical shifts are solvent dependent, so if you want to compare your results with a published spectrum, make sure you use the same solvent.
  2. Solvent Purity:
    Many deuterated solvents are available to purchase in the Chemistry stockroom. The “%” value on an NMR solvent bottle refers to the level of deuteration, not purity. If you have a particularly demanding application you may have to purify the solvent. If you have a water-sensitive analyte you must dry the solvent prior to use. We recommend 3 Å, Linde-type, cylindrical molecular sieves. The standard spherical sieves tend to release dust, which can lead to shimming issues. Activate the sieves in a drying oven and cool inside a desiccator before adding them to your solvent. Wrap parafilm around the caps of solvent bottles to stop moisture from getting in. Note: Parafilm is good for keeping water out but it will NOT keep apolar solvents in your tube.
    Be careful when purchasing multiprotic NMR solvents. Deuterated methanol, for example, is available as methanol-d (CH3OD), which is cheap but not an NMR solvent, methanol-d3 (CD3OH), which is an NMR solvent used to prevent deuterium exchange into the analyte, and methanol-d4 (CD3OD), which is the NMR solvent most commonly understood as “deutero methanol”.
  3. Sample Tube:
    Use high-quality 5 mm sample tubes. Anything sold in the stockroom is OK. Anything that costs more than $10 is probably OK, but if you want to buy tubes from another source, please talk to us before ordering. Do NOT use tubes sold as “disposable” or “economy” without checking with us first. Do NOT use tubes of unknown origin. Do NOT use tubes that are not labeled with model and manufacturer.
    NMR tubes are classified based on the type of glass they are made of and mechanical tolerances such as outer diameter, wall thickness, concentricity and camber. Warning: cheap tubes with poor mechanical tolerances may cause catastrophic probe damage. Cheap tubes are also made of lower grade borosilicate glass that has poor thermal properties and may contain larger amounts of paramagnetic metal impurities resulting in broader lines.
    If you want to analyze very small amounts of material (< 500 µg), you should consider using specialized NMR tubes. 5 mm solvent-matched Shigemi tubes or 3 mm NMR tubes can significantly improve sensitivity. Please contact us for details. If your sample is limited by solubility and not availability, you should consider using our 600 MHz 10 mm probe. Please contact us for details.
  4. Sample Volume:
    The height of the solution in the sample tube is very important!

    • The recommended sample height for Bruker spectrometers is 40–50 mm (about 450–500 µL of solution). Using even slightly less solvent or significantly more solvent could result in poor lineshapes. We posted signs on the lab doors where you can check your sample. We also recommend drawing a 40 mm long box on your hood sash with a permanent marker as a quick and easy way to judge sample height.
    • If your sample is shorter than 40 mm, you MUST center it in the depth gauge.
    • NEVER submit a sample that is shorter than:
      • 36 mm on the AV500 and AV501
      • 30 mm on the AV400

      NO EXCEPTIONS! If your sample is too short, go back to your lab and add solvent.

    • Do not use (air-displacement) micropipettes with non-aqueous solutions, such as chloroform! Standard micropipettes were designed specifically for dispensing water-based solutions and rely on the vapor pressure and surface tension of water. They are highly inaccurate and irreproducible with high vapor pressure, low surface tension organic solvents. Use a glass syringe or a positive-displacement micropipette. Plastic syringes may work, but run a sample of just your solvent from the syringe because you may extract additives from the plastic.
  5. Cap Color:
    We use cap colors to sort samples when they are removed from the sample changer. Please make sure you use the correct cap color on standard NMR tubes, unless you want to donate your tubes to a different research group:

    • Abbasov: White/Black
    • Baskin: Pink
    • Coates: Yellow
    • Collum: Magenta/Black
    • Hyster: Magenta
    • Fors: Green
    • Lambert: Sky Blue
    • Milner: Dark Blue
    • Song Lin: Red
    • Schroeder: Black
    • Stache: Green/Black
    • Wilson: Light Blue
    • Other Chemistry: White
    • Engineering: Orange
    • All Others: Purple

    The facility provides caps. Stop by if you need more.

  6. Label your samples with at least your NetID. Labels may be written directly on the cap or the portion of the tube that remains above of the spinner turbine using ultra-fine tip permanent markers. You can remove the marker using acetone or other organic solvents. You may also use adhesive labels but ONLY IF they were made specifically for 5 mm NMR tubes (e.g. Sigma-Aldrich Z220574 , Wilmad WGL-5 or similar). Do NOT use any other adhesive label. Do NOT use any label that is loose or sticks away from the tube. “Flag” labels can jam in the changer or magnet.
  7. Precipitates may or may not affect spectral quality depending on the magnetic susceptibilities of your solution and solid. If a sample that is cloudy or has suspended solids gives spectra with poor lineshapes, consider filtering it through a glass wool plug stuffed into the neck of a Pasteur pipette.

Go to changer rules, go back to safety and rules or return to the main NMR training page.