5. EZ NMR S+A

The EZ NMR S+A (Setup and Acquisition) panel is an entirely in-house created panel. This panel provides a simple central interface for acquisition of routine experiments. The panel should be used as a To Do List by following the numbered steps (simply count to ten!).

Notes:

  1. In a multi-user environment it is unknown what the last user left behind (z0, shims, probe tuning, solvent, etc.) and may not be suitable for your sample
  2. The correct solvent is fundamental to the proper setup of the instrument
  3. 20 solvents are available in the drop-down list
  4. Allow the spectrometer time to complete probe tuning, loading standard shims, finding the lock, and gradient shimming before proceeding to the next step. Completion messages are provided and failure to wait will hang the system.

EZ_NMR_SA_panel

Figure 5.15 : EZ NMR S+A panel

5.1 Step 1 - Inserting a sample

sample_on_top_of_magnet
  • click on the Eject button

eject

  • if a sample is in the magnet it will float on a cushion of air to the top of the magnet
  • insert your sample correctly into the spinner (see figure 4.3); use the depth gauge
  • place sample with spinner as shown in the figure to the left; the sample floats on a cushion of air
  • click on the Insert button

insert

  • wait until sample is all the way inside the magnet (the system will turn the air lift off)

Figure 5.16 : Inserting the sample

 

5.2 Step 2 – Select Solvent

solvent drop down
  • select the correct solvent from the drop-down menu
  • seven common solvents will provide automatic referencing
  • the remaining 13 less common solvents below the dashed line may require manual referencing

Figure 5.17 : Solvent drop-down menu

 

5.3 Step 3 – Probe Tuning

  • ProTune or automated tuning on: mr400, i400, u500, ibd5, i600 & v700.
  • NO tuning on m400.

All parameter sets are based on a properly tuned probe. Failure to tune can result in partial or complete loss of NMR signals in your experiment.

Experiments that involve a hetero or X nucleus (where X can be C13, P31, B11, etc.) both the X nucleus and H1 need to be tuned and in that order!

Examples of experiments that need both the X nucleus and H1 tuned: APT, C13[H1] 1D, gHSQC, gHMBC, B11[H1] 1D, H1[P31] 1D, etc. but not F19.

Tuning depends on:  
  • solvent
  • sample volume
  • salt content
  • NMR tube

 

probe_tuning

 

protune_pop

 

Figure 5.18 : Tuning with ProTune. Top - Probe Tuning button, bottom - Tune Probe pop up window

Tune Probe

  • click on 3. Probe Tuning
  • in the Tune Probe pop up window, click on the desired nucleus , H1 is selected as an example, to initiate tuning
  • the following messages will be displayed, please wait for the completion message before continuing

protune_message_1

protune_message_2

  • repeat for second nucleus if needed (e.g. tune C13 and H1 for HM[Q|B]C, HSQC and APT, etc.)

5.4 Step 4 – Load Standard Shims, z0, Lock Power|Gain

Why lock? Every magnet slowly drifts (field drift) to lower magnetic field strength, typically 1 to 5 Hz/hour. To achieve frequency stability over the duration of an experiment (16 hours or more for some 2D, days for 3D, 4D, 5D!), FT spectrometers use the deuterium signal of the solvent as an internal lock. Drift compensation and stability is achieved through comparison of the spectrometer frequency with the lock signal frequency.

The amount of deuterium is quite different in CDCl3, D2O, and CD3OD. Therefore, lock power and lock gain are solvent/spectrometer-dependent (use EZ NMR S+A button 4c: Display z0…. or enter z0 on the vnmr command line to run the z0 macro).

Some solvents like CD3OD have two deuterium signals. Locking on OD is difficult, therefore locking on CD3 is recommended, “much more likely to happen” and assumed throughout these notes. If locked on OD, the spectral window and referencing will be off by ca. 1.5 ppm and gradient shimming may fail.

Loading Standard Shims and Automated Locking step-by-step

load_std_shims_etc Figure 5.19: Load Standard Shims, z0, Lock Power | Gain
  • click on 4. Load Standard Shims, z0, Lock Power | Gain (note that z0 is the Lock Frequency)
  • a standard shim set is loaded and sent automatically to the magnet
  • the system will find the lock frequency (z0) automatically
  • the system automatically turns the lock on, and will provide a message when the lock has been found, please wait for the completion message before continuing
  • clicking on the Lock On button in 5. Lock panel may also be needed to lock to the solvent
  • click on Lock Scan and wait about 10 seconds, if the lock level does not go up or the lock does not look like similar to Fig. 5.24 you may need to add lock gain and/or lock power, adjust the lock phase or lock the sample manually (see the next section for manual locking)
lock_control Figure 5.20 : Lock Control

5.4.1 Locking manually step-by step

lock_far_from_resonance

Figure 5.21 : Lock Scan (far off resonance)

  • change the lock frequency by clicking on the z0 button until a lock signal is observed as a wavy line
  • the middle mouse button toggles 3 sensitivity settings of the z0 button
  • if necessary increase Lock Gain first, and then if necessary increase Lock Power

lock_closer_to_resonance

Figure 5.22 : Lock Scan (off resonance)

  • change z0 to reduce the number of “frequency beats” as shown below

lock_closer_still_to_res

lock_just_about_there

Figure 5.23 : Lock Scan (approaching on-resonance)

  • change z0 until on resonance: the lock signal will be similar to that shown in the figure below, a “plateau”

on_resonance

Figure 5.24 : Lock Scan (on resonance)

  • click on the button Lock On in 5. Lock panel. Further adjustments to z0 are no longer necessary
  • adjust lock phase if necessary to maximize Lock Level then turn off Lock Scan
  • for systems such as the u500, i600, or v700 (VNMRS console) adjust lock phase with the yellow lock line on top, all other instruments (Inova and Mercury+) only have a single yellow lock line

lock_control

Figure 5.25 : Lock Control (on resonance, locked)

If not successful turn Lock Off and/or continue to search for the z0 frequency. Once locked, avoid high lock power which causes saturation, i.e. more power flows into the sample than can be dissipated through relaxation processes resulting in sample heating, poor quality spectra, and potentially no lock.

Use information from z0 macro to estimate suitable values.

General guidelines:

  • If needed, increase lock gain and when necessary increase lock power.
  • The Lock Level should be between 80 and 100 (but not over 100) for a shimmed sample otherwise the lock may be lost under the effect of gradients.

5.5 Step 6 – Gradient Shimming

Purpose of Shimming: optimize the homogeneity of the magnetic field by using shim gradients: Z1(linear), Z2 (squared), X, Y and many more
(There are as many as 28 shims available on some instruments!).

fig_5.26_grad_shim_button

grad_shim_message

  • 6. Gradient Shimming
  • message displayed when gradient shimming starts indicating a temporary change of solvent
  • wait until complete and then turn the spinner on if desired

Figure 5.26 : Gradient shim button and gradient shimming message.

Gradient shimming is an automated shimming protocol that will adjust shim gradients Z1 to Z4 in a very short period of time and is the recommended method of shimming a magnet.

Only if gradient shimming fails (i.e. gradient shimming fails to converge, all peaks within the spectrum are split, etc.), use manual shimming:

  • change Z1 to obtain maximum lock level on “speedometer”
  • change Z2 to achieve the same, then return to Z1
  • repeat until no further improvement can be achieved
  • the middle mouse button changes the sensitivity of the shim and lock buttons (i.e. values of 1,10, or 100 are selectable)
  • if shim changes are large, re-optimize lock phase

shim_panel

Figure 5.27 : Start Tab - Shim panel

5.6 Step 7 – Spinner

Turn spinner on if desired.

spin

Figure 5.28 : Spinner button

Notes:

  1. On most modern NMR spectrometers the improvement seen in magnetic field homogeneity from spinning a sample is very minor.
  2. The system will turn the spinner off for some techniques, i.e. all 2D experiments and selected 1D experiments.

5.7 Step 8 – Select Technique

Common 1D and 2D experiments can be selected as shown in the following Figure:

select_technique

Figure 5.29: EZ NMR Techniques

Note: It is recommended to always record a H1 1D before recording any 2D or heteronuclear experiments. The time requirement for a H1 1D is minimal and this allows one to check

  • if the sample is really worthwhile (purity, concentration, etc.)
  • if the homogeneity of magnetic field (shimming) is what it should be
  • if other adjustments are necessary (spectral width, nt, etc.)

Heteronuclear techniques (Li7[H1] 1D, B11[H1] 1D, F19 1D, Si29[H1] 1D, Si29 gHSQC, Si29 1D, Sn119[H1] 1D, etc.) can be selected from the Heteronuclei drop down menu.

Optionally one can make adjustments to the selected technique prior to starting the experiment. For example a H1 1D:

options

Figure 5.30 : Optional adjustments for selected technique

Calculate & Set nt

Calls macro to calculate number of transients (nt) based on sample concentration which is calculated from user input.

For example, calculating nt for a sample with a solute concentration of 70 mM.

The system will prompt:

What is the molecular weight? 370

How many mg of the compound? 18

The default of 16 scans or nt=16 is set.

Set ppm Range: sw|sw1

Calls a macro to calculate the spectral width (sw and/or sw1) based on user input.

For example setting the sweep width for H1 1D from 8.4 ppm to 1.2 ppm.

The system will prompt:

Enter desired low field (highest ppm) limit: 8.4

Enter desired high field (lowest ppm) limit: 1.2

5.8 Step 9 – GO!

GO

Figure 5.31 : Start the NMR experiment!

Press the very large GO! button to start the experiment. Sit back, relax, your experiment is in progress.

As data becomes available the user can start processing the data. See section 6 for details.

5.9 Step 10 – Auto Save

autosave

Figure 5.32 : Auto Save

Included in the EZ NMR S+A to do list, but optional, is Auto Save.

Auto Save will:

  • save the data for the user in their absence
  • save the data in the current working directory using a standard naming format (see section 7.2 for details on the standard naming format)
  • save the data as long as the experiment is running
  • save the data as long as the instrument does not encounter an error
  • save the data when the experiment finishes
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