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I am on Bruker 400

I am doing...

First things to do

Locking

Tuning

Shimming

Making a new experiment in the workspace

Proton 1D

Carbon 1D

Complex NMR experiments: 2D, selective, etc. (separate HTML page)

Using Temperature Control

Current Temperature Calibrations

Data Processing

DOSY

Finishing my work

Known glitches of Bruker 400 in Stepan

Solvent reference data

Troubleshooting

• Topspin frozen. Correct protocol for force-quitting Topspin

   YOU CAN DO THE ACTIONS LISTED BELOW ONLY IF YOU WERE DIRECTLY INSTRUCTED BY NMR CENTER STAFF !!!

Restarting spectrometer

Restarting the sample changer

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First things to do

1. Log in

• Log into workstation
• start Topspin software
  
  

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2. Insert your sample

• Bruker 400 is equipped with an automatic sample changer Bruker SampleCase 24
  
  
  Before you use the changer you must read and view SampleCase Training

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Locking

At this point, your sample is the probe.

1. Load the default shimset
   • Issue: rsh
   • Find bbfo-latest in the dialog window
   • Click Read button

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2. Issue: lock
3. Select your solvent in the dialog window

Wait for the lock system to finish locking (see the status messages)

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Tuning

NOTE: If your solvent is toluene-d8 (Tol) and 'atma' fails with an error, follow these instructions: System will not tune when lock solvent is toluene-d8

Tuning system needs to know what nucleus you are going to work with. Follow these steps:

• Right-click on the appropriate dataset in the data browser of Topspin
  • If you only need to use proton, any proton-detected experiment is OK
  • if you will use both proton and carbon during you session - join a carbon 1D experiment because it uses both channels and they both will be tuned.
    
    
• Click "Display"

Automatic Tuning

• issue atma
• Watch the screen: you need to see how the tuning system successfully shifts the wobble curve minimum to the center line of the screen.
• Once it exits it is done.

Manual Tuning

Manual tuning mode allows you to see what channels are tuned, inspect the result of tuning and adjust if necessary

• issue atmm
• move the mouse cursor onto the Atma Control window and wait till the cursor stops rotating (during this time the computer talks to the motors in the probe - it is a slow communication)
• The lowest-gamma nucleus (13C) is automatically selected and a "wobble curve" - the probe response at different frequencies is shown.
• Select Optimize:Start to start automatic tuning
  
  
  
  
• The good tuning result should is when center of the wobble curve at the center line
  
  
  
  
• Click on 1H radio button to start tuning proton
  
  
  
  If the wobble curve is very much off (probe was tuned to a previous solvent that was very different) you have two choices
  1. Click Optimize:Start and let it optimize automatically
  2. (A little faster, applies to Stepan 400) Click 18 times on [>>>] button in Tuning section to bring a resonance closer to the center
     
     
     
     and then select Optimize:Start

  The result is the same:
  
  
  
  
  NOTE: The automatic tuning result is completely acceptable for most routine work. However, for best sensitivity you may want to adjust Tuning and Matching manually to make the resonance sharper.

• Finish tuning by choosing File:Exit and click Close button in a dialog.

Probe is now tuned and ready for experiments.

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Shimming

Shimming is required to achieve good resolution in NMR spectra.

• issue topshim in the experiment you intend to run
  
  
• Run a 1D 1H on you sample and look some multiplet (best - if it is a solvent multiplet like DMSO at 2.5ppm, etc.).
  
  
• Typically, you have multiplets with about 2 Hz splitting resolved to about 50% height.
  
  
• If reolution is not so good, you may try topshim optoff
  
  
• if resolution is still bad, your tube may be not of adequate quality - try shimming a different sample to check for that
  

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Proton 1D

• Spectral width, SW, FIrst you should seCenter of the spectrum, O1 is set to wherever

• Relevant parameters to adjust (click on Acqpars, "Square wave" symbol):
  • Relaxation delay, D1 is between 1 and 3 seconds
  • Acquisition time, AQ is from 2 to 4 seconds. If your signal dies off very fast, you may want to reduce AQ because after signal is gone receiver is only collecting noise, which reduces signal-to-noise ratio in the processed spectrum

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Carbon 1D

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Using Temperature Control

IMPORTANT NOTE: If you need to use temperature OUTSIDE of 20-40 Deg C range,
you must be trained on the "Variable Temperature" protocol: contact NMR center staff to schedule training.

For all Variable Temperature tutorials and instructions see Variable-temperature NMR operation

The instrument is equipped with manual BCU-1 refrigerated chiller and liquid N2 chiller.

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Current Temperature Calibrations on Stepan 400

• Black curve: calibration with ethylene glycol (KVT_Glycol_3ramps_July_15_2020)
  • Dataset for plotting (column 1, VTU setting; column 2, Real T): VTU_realT.KVT_Glycol_3ramps_July_15_2020.400.extended.txt
    
    
• Red curve: calibration with a commercial methanol standard (KVT_methanol_3ramps_July_16_2020)
  • Dataset for plotting (column 1, VTU setting; column 2, Real T): VTU_realT.KVT_methanol_3ramps_July_16_2020.400.extended.txt
    
    
• Green line: 1:1 line

Stepan 400: Overview (see zoomed regions next)

Stepan 400: Low-temperature branch (obtained with LN2 chiller)

Stepan 400: Same data for extrapolation

Stepan 400: Low-temperature kink on a curve (no-trust region)

Stepan 400: Mid-range region

Stepan 400: High-temperature kink on a curve (no-trust region)

Stepan 400: High-temperature branch

IMPORTANT: Make sure that your LN2 chiller is disconnected and BCU-1 is in "Flush" mode!

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Data Processing

Calibrating the chemical shift axis using a reference spectrum in 1D

• Open a spectrum that contains a peak of a reference compound
• Zoom around the reference peak
• issue .cal on Topspin command line
• Left-click in a center of the reference peak

• Enter reference chemical shift in ppm and hit OK
  NOTE: For tables of chemical shifts you may look at UW Chemistry or SpectraBase or some other reputable source.
  
  Now the chemical shift axis in this experiment is calibrated:
  
  
  
  
  
  
• You can transfer this calibration to a different experiment if it was acquired on the same spectrometer.
  
  Open Procpar tab of the calibrated experiment and copy SR value to a clipboard
  
  
  
  
• Next, you display the target experiment and enter this value in its Procpar, SR field.
  
  
• Hit Enter and check your Spectrum tab: it will show a properly calibrated 1D.
  

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DOSY Experiments

For detailed protocol see  DOSY on Bruker 400 and 500

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Finishing my work

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Known glitches of Bruker 400 in Stepan

Probe cannot tune. ATMA error "Channel cannot be wobbled with this probe (Probe [1H] cannot be selected on [1H] channel)

Sometimes ATMA will fail with a nonsensical message saying that "1H cannot be tuned on 1H channel". This is a glitch we are working with Bruker on resolving. It appears to happen at all times when the lock solvent is Tol, but may happen with other solvents too. Current workaround is this as follows. Any time you need to issue atma, you have to

• unlock first (issue 'lock off'),
• run atma, and
• lock back to your solvent after atma is done.

For example, the following is a basic sequence of commands for NMR acquisition with Tol (note how atma is flanked by lock commands):

1. rsh
   - select 'bbfo-latest'
2. lock Tol
3. lock off
4. atma
5. lock Tol
6. topshim
7. rga
8. zg

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YOU DO THIS ONLY IF YOU WERE DIRECTLY INSTRUCTED BY NMR CENTER STAFF !!!

    Restarting spectrometer

To restart the Bruker 400 MHz system, follow these steps:

1. Restart the workstation:
1. Terminate Topspin
2. Log out
3. Choose "Restart" to reboot the computer

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2. Open the spectrometer cabinet (pull and turn the handle)
   
   
   
   
3. Locate all system switches according to this picture:
   
   
      
   
   
4. Side view of the console with the red reset button and Post Code display:
   
   
   
   

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5. Turn off power to the NMR system
1. AQS3 - push red button, flip power switch to OFF (see the image above)
2. Amplifiers: turn power OFF
3. BSMS: turn power OFF
4. turn Main Power Switch: OFF (push in red section)
5. Ignore

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6. IMPORTANT: Eject current sample with 'sx ej'. If changer does not respond, you still proceed with next steps.

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7. Wait till the computer workstation boots up and reaches the login screen

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8. Boot up the NMR system
1. Turn Main Power switch: ON (push in green section)
2. Turn AQS3 power: ON
3. wait for 1 minute to see the Post Code display increase the code from zero to around 94-98 (exact value does not matter)
4. Turn Amplifier power: ON
5. Turn BSMS power: ON
6. Turn Sample Express power: ON (push the button on the front of the sample changer)
7. wait for 1-2 minutes for all errors on amplifier and BSMS to clear and for the Sample Express to finish initialization and turn on a green light (sample will be ejected and re-inserted automatically).
8. close the spectrometer cabinet

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9. Login to the workstation

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10. launch Topspin

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11. join a proton experiment

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12. issue "ii restart"
    (ignore the "status line active" message)

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13. Load a standard shimset: "rsh bbfo-latest"

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14. Proceed with locking (CDCl3)

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15. join experiment with proton and carbon and issue "atma"
    NOTE: Startup of atm module takes long time (3-5 minutes) but it continually outputs messages in the status line indicating it is not just frozen.

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16. issue "topshim "

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17. adjust gain: issue 'rga'

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18. Run test 1D 1H on CDCl3 standard: you should see undeuterated trace CHCl3 in one scan.

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    At this point, spectrometer is ready for use.

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   Restarting the sample changer

For instructions on resetting SampleCase, please, see SampleCase tutorial

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