NMR TUTORIAL

 

The Parts of the NMR

 

BASIC OPERATIONS

 

 

ADVANCED OPERATIONS

 

 

 

 

 

 

The Parts of the NMR

 

The nuclear magnetic resonance spectrometer (usually abbreviated NMR) has three basic parts:

  1. There are two computers which allow you to control the other parts. The main computer (nmr1) sits on the desk and is usually the only thing you will ever be touching. The acquisition computer (nmr2) is on the floor by your right knee. It’s also called Jill and she will occasionally say things.
  2. The spectrometer is the large, beige box on wheels to the right of the computers. It has two doors on its front. The spectrometer generates the radiofrequency waves that irradiate the sample. You should not open the doors or touch anything inside them or the back of the spectrometer.
  3. The magnet is the large, silver cylinder with three black legs. The sample is inserted in the top of the magnet. You should not touch any parts of the magnet, especially the bottom part. There is a small, beige box next to the magnet that indicates the sample state (loaded or ejected) and shows the pulses of radiation.

 

 

 

NMR Sample Preparation

 

You cannot just put a pure liquid or solid into the NMR. There are a series of steps you need to go through to prepare your sample.

 

  • If you have a liquid you need to analyze, put one or two drops of it into a clean NMR tube that you get out of the oven. Add about three (3) fingers worth of a deuterated solvent (usually chloroform-d also called CDCl3) to the tube. Cap the tube and you’re ready to load. The NMR will NOT work if you do not use a deuterated solvent!
  • If you have a solid, it’s easier to put the solid into a small test tube first. Use about as much solid as your pinky fingernail at the end of a spatula. Add as much deuterated solvent as you would for a liquid sample (you’ll get used to the correct amount), dissolve the solid, and transfer the solution to an NMR tube.

 

Deuterated solvents are very expensive so try not to waste them.

 

The NMR computer is fairly easy to use—the Delta software that runs everything works very similarly to a Windows program. Usually when you get on the NMR computer there will be four windows open.

 

DO NOT CLOSE THESE WINDOWS!!!

 

  1. Delta—informs you what’s going on when you are running an experiment, allows you to open old spectra, and enables you to connect to the spectrometer through a spectrometer control window
  2. Spectrometer Control—shows you experiments that are running, allows you to open new experiments, and gives access to a new sample control window
  3. Sample Control window (nmr2.centre.edu)—allows you to load/eject a sample, and lock/shim on a sample
  4. Experiment window (single_pulse.exp for 1H NMR and single_pulse_dec.exp for 13C NMR)—gives you opportunity to make changes to experimental parameters

 

If an NMR spectrum is open you can close it.

 

 

There are four things you have to do to obtain an NMR spectrum on your sample:

 

  1. Load the sample
  2. Lock and shim the sample
  3. Run the experiment on the sample
  4. Modify the appearance of and print the spectrum

 

 

 

 

SAMPLE CONTROL WINDOW

 

 

 

 

 

 

 

Load the Sample

 

  1. Locate the sample control window, nmr2.centre.edu, shown above. The Sample State should be loaded (the green down arrow with an L is lit and the flask is blue).

 

 

Click ONCE on the  to eject the previous sample. When you do this, the  will empty and the green bar will disappear and the red bar will appear. At the same time, the Spinner icon  will fall down and the  will be lit. You will see the sample tube pop up in the magnet (it’s pushed up by a cushion of air).

 

DO NOT PLACE THE SPINNER IN THE MAGNET WITHOUT A TUBE IN IT!

 

When you place the NMR tube into the spinner, make sure the spinner is pulled all the way apart. Use the depth gauge to position the tube to the right height in the spinner. Try not to touch the bottom of the spinner assembly or the bottom of your NMR tube.

 

Place the sample/spinner assembly into the magnet where it will float on the cushion of air. Go back to the computer and click ONCE on the  icon to load the sample. The  will fill blue again and the   will hop up.

 

Lock and Shim the Sample

 

After you have loaded the sample and it is spinning, you need to lock the instrument on the correct solvent. Go to the Solvent area and scroll to the correct solvent and click on it. Usually, we will be using CHLOROFORM-D (CDCl3).

 

After you have selected the solvent, you must lock the spectrometer on it by going to the Lock Control area and clicking ONCE on autolock . You will see the red bars below the autolock turn yellow:

 

 

When the instrument has locked on the solvent, these bars will turn green:

 

 

After the instrument has locked, you need to shim the magnet so that the sample gets the maximum irradiation. Go over to the shim control box next to the monitor and press the Z1-Z2 button. Then turn the Z1 knob either clockwise or counterclockwise (it doesn’t matter which way you turn it to start). As you turn the knob you will hear a clicking sound. You are looking for the biggest lock signal number possible. If you turn the Z1 knob one way and the lock signal goes down, then you need to turn the knob the other way. Don’t turn the knob too fast or slow, just a nice steady motion. When you have gotten the number as high as possible with the Z1 knob, switch to the Z2 knob and repeat the process. There is no magic number to shoot for, just the biggest you can get. If nothing seems to be happening, look at the sheet posted with the latest shim numbers and turn the knobs to those numbers and shim from there. Do not touch the Z3-Z4 button.

 

 

SHIM BOX

 

 

Run the Experiment

 

Locate the experiment window. If you are running a 1H NMR look for single_pulse.exp and if you are doing 13C look for single_pulse_dec.exp. The experiment window is divided into sections by color. One software quirk is that the cursor arrow must be in the box in order to change the value.

 

In the green section, change the filename to something that you can find later. Use a different filename for each sample (don’t use your name for every sample). DO NOT USE A BACKSLASH (/) IN YOUR FILENAME, JUST NUMBERS, LETTERS, AND UNDERSCORES. The computer will automatically save each spectrum you run. Click once on the auto_gain box to put a red check mark in it (this may already be checked from the previous person).

 

 

 

TOP OF THE EXPERIMENT WINDOW

 

In the pink section, make sure the solvent is the same as the one in the sample control window. Do not touch any other things in this section.

 

 

The purple section controls such things as the width of your spectrum and how many scans you are going to run. You usually only need to run 4 scans on the sample (50 for 13C) and the default sweep width is usually OK. The default 1H spectrum is centered at 5 ppm and runs 7.5 ppm to either side (a total width of 15 ppm). If you suspect that you have a signal outside that range, then set the sweep width accordingly. Don’t change any of the other values. Do not change any values inside the yellow section.

 

 

Once you have changed the filename and other values, click ONCE on . You will hear Jill ask you to check your filename and a box will appear:

 

 

Click on the  button and the experiment will start running. You can check to make sure your experiment is running by going to the spectrometer control window. You should see one experiment listed with an asterisk (*) by it. The asterisk indicates that experiment is running. If there are more than one experiment listed than you or the person before you hit  more than once. Just highlight the experiment that you want to delete from the queue and click . Some other boxes will come up, just answer OK to them. You can highlight all the experiments and stop them all and then just go back and submit your experiment again. 

 

 

SPECTROMETER CONTROL WINDOW SHOWING TWO EXPERIMENTS IN THE QUEUE

 

You can follow the course of the experiment in the Delta window. It should look something like this:

 

 

DELTA WINDOW SHOWING AN EXPERIMENT RUNNING

 

If the window does not look like this, there is a problem with the instrument and you should consult an instructor. Your spectrum should come up in a few minutes, if not, ask an instructor.

 

Manipulating the Spectrum

 

When the spectrum comes up, it will not completely fill the screen. You should maximize it by clicking on the square in the upper right corner (just like any normal windows program). The window will be called 1D Processor: your filename. Below is a typical 1H NMR spectrum of ethylbenzene (the filename is ethyl and it’s version 1) :

 

 

INITIAL APPEARANCE OF ETHYL.1

 

Zoom Tool

 

You will not use the majority of buttons on the screen. The first thing you should do is to zoom in on the portion of the spectrum you are interested in, in the case of ethyl.1, from ~8 ppm to ~0 ppm. To zoom, you need the zoom tool bar which you can access by hitting the F1 key on the keyboard. If the tool bar does not appear, click on the  which is in the upper right of the spectrum. The tool bar will appear in the upper right of the spectrum as shown below. The default tool is the zoom tool,  (the other buttons do things you shouldn’t need). Move the cursor into the spectrum, click and hold the left mouse button to start the zoom rectangle, size the rectangle by moving the mouse, and release the mouse button to complete the zoom. If you make a mistake, just hit the Backspace key on the keyboard. Sometimes, solvent signals are much larger than the signals from your sample. In those cases, you don’t have to zoom on the top part of the solvent signal, just zoom to the height of your highest sample signal. If you need to get back to your original spectrum, hit the Home key on the keyboard.

 

Once you have zoomed on the correct portion of your spectrum, you need to integrate your signals.

 

 

 

ETHYL.1 WITH ZOOM TOOLBAR AND ZOOM TOOL SELECTED

 

Integrate the Spectrum

 

If you are running a 13C NMR spectrum, you will not need to integrate the spectrum. You should always integrate a 1H NMR spectrum. To access the integration tool bar, hit F8 on the keyboard. The integration tool bar has several keys you need.

 

 

 

  Is the tool used to set the integrals. When you integrate a signal you are measuring the area of that signal, so the integral must start to the left of the signal and end to the right of it. To set an integral, move the cursor to the gray area below the spectrum. Click and hold the left mouse button somewhere to the left of the signal you are integrating. A small blue square will appear. Move the cursor to the right and you will see a second blue square appear that is connected to the first by a blue line. The second square will move along the green line of the spectrum. A yellow integral line will appear as you integrate the signal. Keep moving until you reach the end of the signal and release the mouse button. Repeat this process for the other signals you want to integrate. Red integral lines will be present for signals you have already integrated. The computer will assign integral values that will seem meaningless to you until you normalize the values (see  below). The computer arbitrarily thinks the largest signal should have an area of 1 and scales the others accordingly. You will learn that this practice does not reflect reality. If you know a signal is due to the solvent or some other impurity, don’t integrate it.

 

 

ETHYL.1 SHOWING TWO INTEGRALS, BEFORE NORMALIZATION

 

 Is the tool used to delete any integrals that you don’t need or if you made a mistake on a signal. You must click on the red or yellow integral line, not the signal itself or the integral value.

 

 Is the tool for selecting an integral. You must click on the integral line, not the signal or integral value. When you need to normalize your integral values, click on an integral line that you are fairly confident you know the value of. For instance, if you know a signal comes from a CH3 group, then you know its integration should be 3. When you select an integral, its line and number will turn yellow. Press the Ctrl and N keys simultaneously and you will get the Integral Normal Value window:

 

 

The default value is 1. Change the value to what you think the integration for that signal should be. Remember to move the mouse cursor into the window. If your integral values are not close to whole numbers, try to normalize a different integral or try a different number. If you see an M at the end of the integral value it means that number is multiplied by 0.001. So, 84.5M is really 0.0845. If one of your integral areas is this low, it’s probably an impurity since you must have a whole number of protons in a molecule.

 

Print the Spectrum

 

You are now almost ready to print. If you are happy with what appears on the screen, click ONCE on the  button at the top of the window. If you need to zoom in on a different portion of the spectrum than what is displayed, go back to the zoom tool and do it. The integrals will vanish. You can get them back by hitting F8 before you print.

 

Eject the Sample

 

When your spectrum has printed, close []the spectrum (don’t minimize it!). Go back to the Sample Control window and eject  your sample. You must always leave a sample loaded in the magnet! If someone else needs to run a sample after you, just hand them the spinner. If you are the last to use the instrument for awhile, load the tube with the yellow cap. Be sure you load it! Don’t just leave it floating over the magnet. Take your sample back to the lab, discard the liquid in the halogentated waste container, clean it using the NMR tube cleaner, and put the tube in the oven.

 

ADVANCED OPERATIONS

 

How do I retrieve a spectrum?

 

If you need to see an old spectrum again for some reason, it’s easy to get to it. Go to the Delta window and click and hold on Processors. Scroll to Process 1D and let go the mouse button. A blank spectrum window will appear. Click once on  and you should get a window with all the filenames. Scroll down to yours (the ALL CAPS filenames are listed first) and highlight it. To the right you will see a series of numbers which are the different versions of that file. Highlight the highest number first and click OK. Your spectrum should appear, although it takes some time. If you get an FID (see below) instead of a spectrum, you need to process the FID. To do that, click once on  and you will get a new window. You are looking for the process list called std_proton_autophase.list if you are processing a 1H spectrum or std_carbon_autophase.list for 13C. Highlight the process list and click OK. That window will disappear and you’ll be back in the spectrum. Click once on  and wait a minute for the spectrum to appear.

 

 

 

AN UNPROCESSED FID

 

 

 

Hopefully you won’t have to do anything outlined below, but just in case…

 

What if the experiment window isn’t there?

 

If no experiment window is present (or if the wrong one is there), you need to open one. First make sure it hasn’t been minimized. Otherwise, go to the Spectrometer Control window and click on . You will then get a new window that looks like:

 

 

If the experiment you need isn’t listed, click ONCE on  and you will get a new window with a list of experiments. Scroll down to the one you need (single_pulse.exp for 1H or single_pulse_dec.exp for 13C) and highlight it. Click the OK button and the experiment window will come up.

 

 

What if the Sample Control Window isn’t there?

 

Always make sure the window you need hasn’t been minimized. If the Sample Control Window has been closed, go to the Spectrometer Control window and click ONCE on  and the Sample Control Window should come up.

 

What if the Spectrometer Control Window isn’t there?

 

If the Spectrometer Control Window has been closed inadvertently, go to the Delta Window and click ONCE on  and the Spectrometer Control Window should appear. You will need to connect to the spectrometer by highlighting nmr2-FREE-Centre College and clicking ONCE on .

 

What if the sample isn’t spinning?

 

You will know the sample isn’t spinning if the Current value is 0[Hz] or far off from the Target value of 15[Hz]. If the sample isn’t spinning, eject it first. Make sure the spinner is pulled all the way apart. If it is, take out the NMR tube and wipe the bottom of the wide part of the spinner with a kimwipe. Put the spinner back together, re-insert the tube, and load the sample again. If it still doesn’t spin, consult an instructor.

 

 

What if the experiment is not running?

 

The usual reason the experiment won’t be running is because you forgot to hit OK when Jill asked you to check your sample ID. Go to the Spectrometer Control Window to see if there is an experiment in the queue. If not, go back to the Experiment Window and click ONCE on . If there is an experiment listed in the queue, but no * by it, highlight the experiment and click ONCE on OK. It should start running. If that doesn’t work, consult an instructor.