NMR Analysis for 13C with the Unity 400

The ¹³Carbon NMR spectrum of an organic compound provides information concerning:

The major differences between ¹³C-NMR in comparison to ¹H-NMR spectra include:

Before logging on -- Open the login book and enter the user name along with the date and time. Check to make certain no one is currently using the instrument, enter the password for the log-on computer. If the instrument is currently being used, check with the person to see if it is possible to log them off and log in. Normally if a user forgets to logoff the instrument is locked.

Logging in -- Move the mouse to see the login window. Type the login name and when prompted, the password. In the login process, a series of programs start. Wait until the Front Panel us displayed:

The most important applications are on the front panel:  vnmr (an icon with a blue spectrum on a black background), and the File Manager (an icon with an open filing cabinet). To open an application click on it once. Additional clicks will open multiple copies of VNMR which will not operate.

Click on the vnmr icon on the Front Panel once. Wait until the Acqi button shows up at the right of the first row of buttons, and the Acquisition Status-window opens.

Menu-driven operation. The most important commands can be called by clicking on the buttons in various menus. However, it does not give access to the full range of commands. The Main Menu button is on the first row of buttons at any time. Click on it to retrieve the second row of buttons on the Main Menu. These buttons are arranged in the normal sequence for obtaining a plot of a spectrum. In any menu, the buttons on the second row are defined by clicking the Help button.

Command mode. Type commands in the Input Window. The input window must be selected before typing a command.

Selecting the Workspace -- NMR spectra are obtained in experiments. An experiment is actually a directory containing files of the text associate with the sample, the file with the parameters for acquisition and data processing, the FID file, and the spectrum file after transformation of the FID. Each user has his own experiments, which means data in an experiment will be the same, no matter what the other users have done. The data in an experiment is lost if the set up command is given, if another FID is retrieve, by starting an acquisition, or by delete the experiment. At the first log in there os one experiment. To create more, click Workspace - Create New. Do not create more experiments that needed at a time. They occupy disk space. Use the Delete button to erase the unnecessary ones. To see the text associated with each experiment click Library. To move to (join) experiment 3, click the Exp 3 button, or type jexp3 in the input window.

Setting the Parameters -- From the Main Menu click Setup. The setup copies standard parameters file into the parameters file for the current experiment. First, specify the nucleus and the solvent. For the five solvents listed as buttons, appropriate lock parameters can be retrieved. There are buttons defined for proton or carbon in d-chloroform. All the frequencies are set based on the lock frequency. Select the solvent correctly or the spectral window will be improperly set and some region of the spectrum will show up folded. Or select the parameters by typing in the input window, e.g. setup(‘C13',’CDCl3'). To display the parameters at any time, type in the input window dg (display group parameters). Not all the parameters are listed by dg. To inquire about the value of a parameter, type the parameter name followed by question mark, e.g. gain?. To modify the value of a parameter, type parameter_name=new_value. Note that text values should be in quotes, e.g. gain=’n’.

1. Remove standard and Insert sample into the magnet

2. Set up Parameters

3. Retrieve shims

4. Tune, only if necessary

5. Manual lock

6. Manual shim

7. Now change parameter nt, then type ga to collect data

8. Process and print out data

9. To Save Click Data, then type svf and file name

10. Plot Data

11. Remove Sample and put Lock Standard Back

12. Log off the Computer

1. Remove the sample from the top of the magnet by holding onto the top of the spinner. Remove the previous sample from the spinner with a firm pull on the top end and store it in a safe place. Insert the sample by typing commands e and i.

To Insert a Sample into the Magnet -- Push the sample tube through the spinner using a twisting motion to avoid breaking the sample tube. The bottom of the sample should be inserted into the wide end of the spinner first. Adjust the spinner's position using the gauge. Wipe the outside of the tube and the spinner with a soft tissue. When moving the sample/spinner assembly, always hold the top of the spinner to avoid repositioning of the spinner. Never insert a sample unless the bore of the probe is known and there is a sample in the spinner.

• Place the sample tube and spinner into the transfer tube opening in the center of the magnet. Make sure that there is enough air to support the sample before letting go of the sample tube and spinner. Type 'i' so that the sample will be lowered into the probe. Listed for two click sounds (or a click and then a cluck) which indicate if the sample tube sits in the right position.

• If there is no "click" of the sample resting in the probe or the spin indicator light does no stay on (it will blink until the sample is spinning at the desired rate), try ejecting and inserting again. Cover the transfer tube opening in the center of the magnet briefly with one hand and wait for the spinner light to turn on.

Note -- Always leave a sample in the magnet with the lock on at a level of 30% to 70%. The purpose of using a lock solvent is to minimize magnetic field drift.

2. Set up parameters by clicking setup, then select the nucleus and solvent, or by typing rtp, then answer carbon 13: C13. If the solvent is correct then type su.

3. Retrieve shim set by typing rts, plus the shim file name, such as CDCl₃, then su.

• Type rts, at the prompt, enter CDCl₃, and then su. This will load a standard shim settings of the using probe. This is a good start to shim a sample.

• Once a good shim setting has been established for a sample type, it is generally possible to switch between different nucleus without further shimming.

4. Tune. Only if necessary, details are given at the end of these instructions.

5. Manual lock . Or try autolock (type lock) if manual lock is difficult.

• Enter 'acqi' to open the acquisition window. Turn the lock control off. The lock signal appears as a sine wave if the lock signal is off-resonance, and as a dc signal if the lock signal is on-resonance. Adjust Z0 until the lock signal is on resonance.

• Increase the lock power to 20 and the lock gain to 30 or higher until there is sine wave signal. Adjust Z0 to get a dc signal.

• Adjust the lock power and lock gain until the dc signal is from 30% to 70% of the full screen.

• Turn the lock on. Adjust the phase so that the dc signal is highest at the above setting of the lock power and lock gain.

6. Manual shim. Or try autoshim (type shim) if manual shim is difficult.

• The screen will show a digital lock amplitude and two horizontal lock signal bar graphs. The top one shows the total lock signal, the bottom one shows either 0-20%, 20-40%, 40-60%, 60-80%, or 80-100% of the full scale, using different colors for each range.

    (A)  Adjust each of the following to maximize lock signal- Z1C, Z2C, Z1, Z2. Normally only adjust Z1C and Z2C.

     (B)  Increase Z1 by 20 units, maximize lock signal with Z2.

     (C)  If level is higher or the same, repeat step (B). If it is lower go to step (D).

     (D)  Decrease Z1 by 20 units, maximize lock signal with Z2.

     (E)  If level is higher or the same, repeat step (D). If it is lower, increase Z1 by 20 units and maximize lock signal with Z2 and the sample is shimmed.

Continue the steps until signal is maximized.

7. Now change parameter nt, then type ga to collect data.

8. Process and print out data.

Type command 'wft aph' if go was used instead of ga. The following are a few possible commands to further process the spectrum if needed: to display the scale click Dscale or type dscale if the scale is in hertz, type axis=’p’ to view the full spectrum in the full screen f full, to adjust the vertical scale use the middle mouse button directly on the spectrum or type vs=value wanted, to zoom in on the region within the cursors left-click on the left and right-click on the right of a region in the spectrum then click  Expand to change the limits of the spectrum to specific values, for example 1.0 ppm and 6.0 ppm: sp=1.0p wp=5.0p. For baseline correction type dc, or bc to reference a peak, for example, set acetone to 2.05 ppm (in organic solvent) or 2.225 ppm (in D₂O) click as near as possible to the center of the acetone peak, nl rl(2.225p).

For peak-picking
dpf displays the peak frequencies
 th  allows threshold level adjustment with the left mouse button.

9. Data are not normally saved but it possible to save fid in a data directory. Click Data, then type svf and file name.

10. Plot Data -- Click Main Menu/ Display/Plot/Plot/Scale/other options/Page

Page is the command that sends the information to the plotter. Every option selected before page will be included in the plot.

11. Put Lock Standard Back

• Take out the sample and insert the lock sample back into the magnet.

• Reset lock for the lock standard.

• If somebody is waiting, insert his/or her sample instead of the lock sample. Make sure the sample is spinning. In return, the person will log the user off.

12. Log off the Computer

First close down VNMR and then exit the software. Reverse the logon procedure.

Tuning -- NOTE: Ask for help of someone who has done it before! Never tune the probe unless without training.

1. Insert the sample.

2. Setup 1D proton parameters (make sure tn=’H1’, dn=’C13’ and correct solvent). Type su. Now machine knows CHAN1 for H1 and CHAN2 for C13.

3. Disconnect the cable from the 1H preamplifier and connect it to the PROBE J5321 port on the TUNE INTERFACE panel.

4. Press the CHAN + button so that the readout is channel 1 for proton. Make sure the red indicator light is not flashing before proceeding . If it is flashing, check the connections just made with the two cables.

5. Tune the probe using the appropriate stick (note that the red stick always for proton). The upper, smoother part is the tune and the lower, rougher part is the match. As the probe gets closer to being tuned, the
number on the INTERFACE display will decrease.

6. Press the ATTEN buttons until the readout is 8. Continue tuning the probe until the displayed number is less than 10.

7. If the numbers do not change easily (or no response) when turning the rod, there must be something wrong. Find someone who can help. Do not force the tuning rod or turn it too far.

8. Disconnect the tuning function by pressing the CHAN buttons until the readout is 0 (otherwise acquisition will not be possible).

Reconnect the rf cables to their original positions. The red indicator light should turn off.

Repeat the process for tuning C13, by choosing CHAN 2, and C13 cable connections.