NMR Sample Preparation
The following guidelines are
offered so that you can make as perfect a sample as
possible, the first time. The three steps of sample
preparation are concentration, clarity, and sample
volume. Paying attention to these three factors when you
are preparing your sample will eliminate wasted time and
materials. What is important to recognize is that if
your sample is poorly prepared the resulting spectrum
will likely be lower in resolution than what you might
expect. What is also important is that if you get a
poor quality spectrum with 1 scan, increasing the number
of scans will usually not improve it.
Concentration
Optimally, one wants a sample of sufficient
concentration for the particular spectrum required. For
1H nmr spectra (1-dimension and COSY spectra) usually
10-20mg of material dissolved in a total volume of 0.6mL
of solvent will be sufficient for a good quality
spectrum requiring anywhere from 1-12 scans. Strictly
speaking, a deuterated solvent is not necessary because
the Eft does not lock on the solvent signal. A
deuterated solvent does help to eliminate background
peaks arising from the solvent.
Since C13 is about
5000 times less sensitive than 1H, one will require
significantly more sample. If the sample being measured
is a liquid it is permissible to measure the liquid
directly without a solvent. If the sample is a solid,
then 200mg dissolved in a minimum volume of 0.6mL
solvent should provide a good spectrum in about a
minute. If only 100mg of material is available, then
this concentration will provide a good spectrum in about
4 minutes. Remember that in order to double the signal
to noise ratio, the measurement time will increase by 4
times.
Clarity
Once you have the sample in solution, you should
check it for suspended particles such as bits of filter
paper or other fibers or powders. Hold the tube up to a
bright light a look for diffractions off the suspended
particles. The suspended particles will cause
broadening of the lines and give poorer resolution
spectra. To remove the suspended particles, filter the
sample through a pipette with a small plug of glass wool
packed into it. When preparing the pipette containing
the glass wool, be sure that the glass wool is not
packed too loosely nor is the plug of glass wool too
large (see photo). After filtering the sample through
the glass wool, the sample should be free of particulate
matter.
Quantity
The NMR detector is situated so that a volume
of about 50-80uL is detected about 15mm from the bottom
of the tube. It is therefore important that the volume
of the sample be kept to a minimum, no more than 0.6mL
of sample volume. If one uses more than 0.6mL of
solvent, the sample will be more dilute than desired and
the data acquisition may require more time because of a
dilute solution. Samples lower in volume are also
undesirable because the meniscus of the sample will
distort the magnetic field and the close the meniscus is
to the detector coil, the more difficult the shimming or
field optimization of the sample.
Sample Labeling
If you are measuring more than one sample, it
is wise to label the tube using a Sharpee marker on the
top of the tube or on the tube cap or a small label. Do
not use a label which has a flap because the sample
needs to spin when it is in the magnet and this can
interfere with good spinning.