1. Write your name, date, and use in the login book and then log on to the instrument. This requires a username and password. Use this instrument ONLY with the assistance of a trained user.
2. Login, open VNMRm and select Acqi to Eject the standard in the instrument and put the sample to be tested in. 3. Click on the VNMR icon to open up the NMR program. The VNMR software opens several windows. The Acquisition Status window is found on the upper right hand side and serves as a monitor of the instruments current settings and operations. The main windows on the left are where data is acquired and processed. The top window accepts typed commands and reports messages. Under it are the mouse-driven menu bars. The middle window is for spectral and FID display. The lower window is the text window and is used to display the list of parameters for the current experiment and to print information about a given experiment. The middle window and lower window overlap and can be "flipped" using Flip Button. To get the parameter screen, or type >dg Note the function keys across the top of the keyboard are useful as quick keys in VNMR. The proton homonuclear decoupling procedure is also detailed.
On log in VNRM selects Experiment 1. To go to (or "join") another experiment use the commands: jexp2 (or jexp3, jexp4, etc.). Or use the menu buttons: Main Menu/Workspace/Exp#. If there is an error trying to join experiment (#2 for example) that says the experiment is locked, try: unlock(2). To create new experiments, cexp(#) will create the experiment # specified. Or you can use the menu buttons: Main Menu/Workspace/ Create New. To see a list of all the experiments type: explib
4. Click on the Setup button and choose an appropriate nucleus and solvent. For example, for a proton spectrum in chloroform click on the H1,CDCl3 button, or for a proton spectrum in benzene click on Nucleus, Solvent followed by H1, and Benzene.
5. To allow the Sparc workstation to "talk" directly to the acquisition computer, first open the Acqi window. Click on the Acqi button or type acqi which will open up a smaller new window 'ACQUISITION'. This window allows ejection or insertion of a sample. The magnet should always have a locked sample in it. After finishing replace the reference sample and lock it.
Place the sample in a spinner and check the height with the depth gauge. Always be sure to measure the sample depth with the gauge. Samples of improper depth might break as it descends into the probe.
6.Click on the Lock button to view the deuterium lock signal. If the spectrometer has locked onto the signal the trace should look like a flat-step function and the lock level will be fairly constant and above 30. If it is not locked, try raising the lockpower and or lockgain. The controls for this work just like the shim controls (see below). If that doesn't work, close the window. Type rts(solvent) su (where the solvent = your solvent such as CDCl3, C6D6, toluene, etc.) and start again from the previous step . If there is no lock, ask for assistance. Note: Unix is case sensitive; typed VNMR commands are normally in lower case.
What is Lock -- The deuterium nuclei in a sample are used to maintain a "lock" on the sample. The nuclei are used to monitor and correct for any drift in the magnetic field. If the field "drifts" or changes in strength, the precessional frequency of a nucleus will change accordingly. In a pulsed lock system, the field is monitored by observing the resonance frequency of the deuterium nucleus of the solvent (i.e. D2O, CDCl3, etc.). The resonance frequency of the nucleus is compared to a reference frequency in the spectrometer and any changes are corrected by adjusting Z0. Lock the amplification of the deuterium NMR signal, increases the size of the lock gain signal, but also increases any other signals or noise that may be present. The phase angle used to control the phase of the deuterium NMR signal and phase of the reference signal for the deuterium lock, normally needs very little if any adjustment. The quantity of RF energy used to irradiate the deuterium nucleus, controls power the amplitude of the RF pulse at deuterium frequencies. It must be large enough to produce a signal for the deuterium but still below the saturation limit. If the power is too high, the lock signal may decrease in intensity. 7. "Shimming" is adjusting the magnetic field inside the instrument to be as homogeneous as possible. To shim the field, click on the Shim button. There are two horizontal bar graphs that show the lock level. The bar on the top is a tens unit and the one on the bottom is a ones unit. The same information is also displayed more simply as a number. A current set of shims is always available on the machines if you wish to re-load some starting shims. The shims may be retrieved through the menu or by: rts('current') su. Save shims in your vnmrsys/shims directory by: svs('filename'). Once you feel you have maximized shims, Close the acqi window. Only adjust the parameters listed below, paragraphs 8b and 8c. To adjust a value, simply click on the buttons labeled -1+, -4+, -16+ and -64+. Think of these as fine through coarse controls; clicking on -16+ changes the value 16 times as much as clicking on -1+. Click with the left mouse button to select decrement the value of that parameter or click with the right mouse button to increase the value of the parameter. Shimming is accomplished by adjusting each shim parameter to maximize lock level. It is not the absolute number (these can always be changed using the power and gain), rather the highest level obtainable while having the lock level (bars) on scale. 8. To shim the instrument simply: a. Make sure the lock level is on scale as described above. b. Adjust the Z1C (Z1 coarse) and Z2C (Z2 coarse) shim buttons. Normally they need minimum adjustment of no more than -1+ or -4+ to do this. Maximize the reading on the scale using Z1C and then do the same using Z2C. Iterate this procedure back and forth to maximize the bars/numbers. If the lock level pegs at the top of the scale, reduce the lockpower and/or lockgain until the meter again reads mid-scale (toggle through the parameter list to see these parameters). c. Use the Z1 and Z2 controls to finish shimming (these are about 5x less sensitive than the coarse controls). Normally, use the -16+ button. d. Now maximize the lockphase. e. When finished, adjust the lock power and lock gain so the meter level is somewhere between 40 and 80. f. Click on the Close button to exit the Acquisition window. Never attempt to adjust the other shims such as Z0, Z3, Z4, etc.
9. Click on the top window, type nt=4 and hit return. This sets the number of scans to 4. Make sure the peaks look OK before collecting a full set of data. Typical lock and shim value are given. 10. Type su ga. The spectrometer will collect 4 scans and then display the spectrum (see below on how to manipulate things). Adjust the phase by typing aph and make sure that the peaks look good -- they should be Gaussian in shape and not have humps to either side. Typical problems with shims will give humps or even extra peaks on the side of the real peaks. Fortunately, this problem will show up in all of the peaks of the spectrum, so they are easy to spot.There is also manual phase adjustment.
If the peaks are poorly shaped due to a bad shim, go back and try shimming again. If this still does not help type try using the rts command described above. The rts command will returns shims values and loads in a set of values that should be close to the requisite values for a particular solvent. Ask for help if there are problems. 11. If the trial spectra looks good, select the number of scans (number of transients, nt) to collect. For example, to collect 1 to 16 scans and begin acquisition type nt=16 ga.
12. While scans are being collecting, it is a convenient time to enter text about the sample. Type text('sample information here'). Remember to use the 'single' quotes and (parentheses). Use a double backslash to begin a new line. For example: text('KEB-125A, Crude product from\\ether/pentane, orange oil\\yellow C6D6 solution\\9:55P 09-10-00') will produce: KEB-125A, Crude product from ether/pentane, orange oil yellow C6D6 solution 9:55P 09-10-00 13. When the scan is done type aph f to automatically phase the spectrum and display the full spectrum. 14. There is a button that toggles between Cursor and Box. Select Cursor and click the left and right mouse buttons to get two cursors. Place these on either side of the proton solvent peak and then click on the Expand button. Place one cursor on the center of this peak and type nl rl(x.xxp) where x.xx is a number you need to supply. Look at a list of solvent shifts. Typical values are 7.24p for CDCl3 and 7.15p for C6D6. Do not forget the lowercase "p". 15. Click on the Full button to display the full plot. Use the two cursors to expand the area to be plotted. Alternatively, type in something such as sp=0p wp=8p to display, for example, 0 to 8 ppm automatically. Typing in the values ensures the plot of all spectra is on the same scale and overlay them for easy comparison later. To change the vertical scale click and drag on the spectrum with the middle mouse button. Make sure all the peaks are as high as they can be without being cut off at the top. To set the value of the vertical scale type, for example, vs=200
To display the spectrum on the screen, ds. The spectrum will need to be phased. To phase the spectrum, from the menu use: Main Menu/Display/Interactive/Phase. Use the right mouse button to click on a portion of the spectrum. Hold it down and move the mouse to flatten the spectrum's baseline and makes the peaks symmetrical. The sweep width (sw) of a spectrum may need to be altered and with it, the transmitter offset (tof). Generally the sweep width should not be too large for your sample or too small (results in folded peaks). When using water samples, you should set your tof on your water peak. To do this, set the cursor on the water peak and use movetof. To move your sweep width, enclose your spectrum with the cursors, expand, and use movesw. This command moves the tof to the center of the new sweep width so tof will change. You can also manually reset the sw by using sw=# if you do not want to change the tof and you want the sw exact for a series of experiments.
When referencing a spectrum, the spectrum is often referenced to the solvent frequency. For example, D2O is at 4.67 ppm, standard. So, to reference your spectrum, you would place the cursor on the water peak: nl rl(4.67p) This locates the nearest line more exactly(nl) and references the line (rl) to 4.67ppm.
16. To integrate the spectrum find the button that toggles between No Integral, Part Integral and Full Integral. Select Part Integral to automatically integrate and type cz dc to automatically clear any integral resets and level the baseline. 17. Click the Resets button and then cut (zero) the integrals as needed. Working from left to right, move the mouse and click the left mouse button once where the integral is to be cut. If there is a mistake the right mouse button undoes a cut (or use the cz command to wipe all the cuts). Finally, adjust the height of the integrals by clicking and dragging with the middle mouse button. Make sure the integrals are as tall as possible without being cut off at the top of the page. 18. To plot the spectrum type pl pscale plot which plots the spectrum, scale and parameters, respectively. If there are no inset plots, type page to dump the spooled file to the plotter. For details see print. 19. To generate a peak list first toggle through the Integrals button to get rid of the integrals and then press the th (horizontal threshold) button. Click and drag with the left mouse button to set the height of the peak list threshold. Pick a level that intersects all peaks of interest, but don't put it so low that it picks off every single point in the baseline. Then type printon dll printoff to send the peak list to the printer. dll will list only those peaks that are displayed on the screen and that are above the threshold. For complex spectra it may help to expand several regions and dll them individually. 20. When finished, remove the sample and replace the one that was initially in the instrument. Use the spinner properly.
NEVER PLACE THE SPINNER BACK INTO THE INSTRUMENT WITHOUT A SAMPLE (The spinner will not eject without a sample) 21. Log off the instrument by first selecting the Exit VNMR button and then log out of the Solaris. A confirmation message will come up. Click OK to bring the system back to the login screen. The monitor will automatically go to a screen saver mode. Do not turn the CRT off.
22. Normally spectra are not saved; however, some spectra may be saved. After collecting a spectrum that will be saved, it is possible to add some text: text('my sample, D2O, pH 7.0, date'). To save the data, first make sure you are in the correct directory. Use cd to make sure you are in your home directory and change to a data subdirectory. Then save data using the menu buttons: Main Menu/File/Data/Save/FID or svf('filename'). The filename.fid "file" saved is actually a directory containing: the fid, procpar, a log, and a text file. (Therefore, transferring a .fid file is different than transferring a single file in Unix because it is treated as a directory e.g. use cp -r).
23. Add an array-- Type array and when prompted enter the parameters of the array, including starting points and increments. To see the array type: da (display array) or type: pw=5,10,15,20,25,30 (for example). Use the time macro (time) to find out how long this will take. Sometimes really big arrays and long d1's can create a long experiment. The time macro is very useful when doing 2-D and 3-D experiments. Acquire the spectra with ga. As the spectra come up, use ai dssh to view them together. This command specifies absolute intensity and display a series of spectra horizontally. Use the dssl command to list the array numbers (corresponds to da). If your peaks are coming up all the same height, make sure you use ai because you might be in normalized mode. The vertical position of the spectra may need to be adjusted to see the peaks that become negative. (i. e. vp=50) Look at the spectra and your array numbers and decide where the second null is occurring. Use a second array to narrow down on the 360 null and determine it more accurately. Determine from your array list the best null point and divide by 4. Set your pw to that 90 degree pulse width. Some experiments also require a parameter called pw90. Setting up 2-D experiments set pw90=pw.
24. Finding signal-to-noise ratios -- Measure the intensity of the largest peak in the spectrum. Use the cursor to move onto the line and use nl. Then move the cursor over and enclose an area of noise with two cursors. Then type dsn for display signal-to-noise.
25. Finding digital resolution -- Place the cursor near the maximum of the peak you wish to measure. Use nl dres for nearest line, display digital resolution.
a. Do not switch too quickly between lock and acquisition. b. Try to exit the acq window before starting an experiment. c. Use a pre-acquition delay of 2 seconds (pad=2).
If the computer locks make a note of what process was going on and what other programs might still be active (Are both the keyboard and mouse locked? Is there an hour glass on screen?).
Never turn the computer off with its main switch. First try to close each screen. Use the right mouse button to open a window and select close. Proceed until all windows are closed. As a final resort close the software with the Stop key while holding down the letter "A" key. Then at the OK or "<" prompt type boot and then the Enter key. The system will reboot and start the login software. Do not enter information until the normal sign on screen appears, it will take a few minutes. The proper procedure to turn off the computer is only from root. First seek root then in a terminal window type sync;sync;halt. Wait for the OK. Then to insure handshaking between the magnet and instrument turn the Unity power off. Wait 30 seconds turn the power on and press the green button to start the instrument. Then type boot at the OK prompt and wait for the Solaris login window to appear.
27. Power Failure Procedure -- Turn off the computer and hal box. On the back of the main console the white breaker should be in the up position. Push the green button and hold it down until the system is running and release the green button. Open the right front console door and push the reset button then turn on the hal box and computer. There is no need to reset littleboy.
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