Common solvent impurities

Please see OM 2010, 29, 2176 and JOC 1997, 62, 7512.

NMR HowTos

Please see NMR at The University of Chicago (or google “your question + Topspin”) for information concerning

• simple 1D NMR
• 90 deg pulse calibration
• spin-lattice relaxation time (T1) measurement
• DQF-COSY setup
• NOESY setup
• VT-NMR setup

How to Integrate NOESY peaks manually on Topspin3.2

(by Rebecca Black, 3/12/2015)

1.    Open NOESY spectrum and click “Process” tab at top

2.    CHOOSE SCAN OF INTEREST

a.    Click “Advanced” tab at top

b.    In pull down menu, choose “extract rows/ columns (slice)”

c.    In window that opens, check circle for “interactive slice/ column display” and close

d.    Click the square for “scan rows” (figure is a spectrum and up.down arrown)

e.    Cursor over cross peak and 1D spectrum of a particular scan will appear with the diagonal cross peak up and NOE signals down

f.     Find a row scan that has visually highest integration of both positive and negative peaks and remember the scan number (i.e. 33 of 1024 in bottom of spectrum window)

3.    SAVE 1D Spectrum

a.    Click “Advanced” tab at top

b.    In pull down menu, choose “extract rows/ columns (slice)”

c.    This time, check  “extract a spectrum row” and type in the row number of scan from step 7 and “destination PROCNO” (this will bring up the 1D spectrum and save it within the NOESY experiment)

4.    Integrate 1D Spectrum

a.    For first scan, take integrals like normal 1D experiment. Before saving/closing integration menu, right click diagonal cross peak and calibrate to equal 100 and close.

b.    Before save/close, click the floppydishA square and “save regions to ‘intrng’”. This will save the integrations of this 1D slice of the 2D experiment and you can then use this to calibrate the rest of the 1D slices so the integrations are comparable.

c.    For subsequent 1D slices after the first integrals are saved, right click on an integral in subsequent peaks and apply “use lastscale for calibration”.

How to dry deuterated solvents

P₂O₅ as drying agent

CD₂Cl₂, C₆D₅Cl, C₆D₅Br, CDCl₃, ClD₂CCD₂Cl, Cl₂DCCDCl₂

Place P₂O₅ in round bottom flas in glovebox. P₂O₅ should be a white, free-flowing powder.
Outside glovebox, pour solvent over P₂O₅. Place solvent under a static N₂ pressure. Stir for 1-2 days, the longer the better.
Degas: Freeze-pump-thaw solvent 3 times.
Freeze-pump-thaw: Place liquid N₂ bath around flask (N₂ level BELOW the level of the solvent). Once solvent is frozen solid, open vessel to vacuum. When flask is under full vacuum, seal flask and warm to RT.
Vacuum transfer degassed solvent into a dried storage container. Label the container. Take a reference spectrum with an internal standard, e.g. C₆Me₆, to determine the amount of residual water.

Na/benzophenone as drying agent

benzene-d₆, toluene-d₈

In glovebox, place Na/benzophenone in round bottom flask with stir bar and attach 180 deg adapter.
Add the solvent in the box. Attach to line. freeze-pump-thaw 3 times.
Stir under static vaccum for several hours.
Attach hot vacuum transfer flask to line and evacuate. Let flask cool to RT under a dynamic vacuum. Once at RT, seal flask.
Vacuum transfer dried solvent from Na/benzophenenone to empty, dried storage flask. Label flask.

 THF-d₈

In glovebox, place Na/benzophenone in round bottom flask with stir bar and attach 180 deg adapter.
Attach flask to line and evacuate. Once flask is evacuated, seal flask.
In separate Schlenk vessel, add desired amount of THF-d₈. freeze-pump-thaw 3 times.
Vacuum transfer degassed THF-d₈ to flask containing Na/benzophenone.
Stir under static vaccum for several hours.
Attach hot vacuum transfer flask to line and evacuate. Let flask cool to RT under a dynamic vacuum. Once at RT, seal flask.
Vacuum transfer dried solvent from Na/benzophenenone to empty, dried storage flask. Label flask.

CaH₂ and 4Å molecular sieves as drying agent

Attention: molecular sieves have to be dried at >150 deg in the vacuum for at least 12 h.

DMSO-d₆, CD₃CN

In glovebox place CaH₂ in round bottom flask with stir bar and attach 180 deg adapter.
Pour solvent quickly in round bottom flask outside glovebox. Place flask under N₂ atmosphere.
Freeze-pump-thaw 3 times. Warm to room temperature and stir for 12 h under static vacuum.

For DMSO perform two more freeze-pump-thaw procedures after stirring.
In glove box place 4Å molecular sieves in a dry storage flask (Schlenk flask with stir bar in the case of CD₃CN).
Attach to line and place under vacuum.
Vacuum transfer solvent to storage flask. These solvents are stored over molecular sieves.
For CD₃CN stir for 12 h and vacuum transfer to another storage flask with 4Å molecular sieves.

Acetone-d₆

In glovebox place 4Å molecular sieves in round bottom flask with stir bar and attach 180 deg adapter.
Pour solvent quickly in round bottom flask outside glovebox. Place flask under N₂ atmosphere.
Freeze-pump-thaw 3 times. Warm to room temperature and stir for 12 h under static vacuum.
In glovebox place more 4Amolecular sieves in a dry storage flask.
Attach to line and place under vacuum.
Vacuum transfer solvent to storage flask. Acetone-d₆ is also stored over molecular sieves.

Cost per NMR sample

(based on a 0.5 mL sample volume)

HMQC on Bruker 500-2

Calibrate the 90 deg pulse for ¹H and ¹³C.
Determine T₁ for ¹H
rpar HMQC.BBI
eda: set SW to the smallest window possible. F2 channel for ¹H NMR window, F1 channel for ¹³C NMR window. Use default AQ value in the parameter file. Set O1P value as ¹H O1P, O2P value as ¹³C O1P. Set TD for F2 channel as 1 k, TD for F1 channel as 256.
set P1 and D1 to 1.27*T1 as determined for ¹H NMR, set P3 to 13C NMR P1 value. Double check the power level, i.e. PL1= 0, PL2= -3.
choose NS in multiples of 4, and Dummy Scan DS as 16.
Optimize D7: Set starting value of d7 = 400ms. Enter acqu to enter acquisition window, enter gs to start go setup routine. Adjust the value of d7 to reach the minimum internsity. Set d7 as that value and type STOP and click the RETURN button.
Enter rga to optimize the receiver gain.
check the required experiment time with expt. If the time is too long, either reduce NS or the number of experiments in F1 (eda – TD(F1)=128-512)
Start experiment: ZG. The spectrum accumulated so far can be viewed any time by xfb.

Phasing:
xfb (can be entered anytime during spectrum acquisition to check the spectrum achieved so far, but MUST be entered at the end of data acquisition. Each experiment increases the resolution in F1. Comparable to tr in 1D.)
The standard setting is to display a magnitude spectrum and phasing is neither required nor possible.
To display a phase sensitive spectrum, type edp and change phase correction from mc to pk for F2 and F1. Enter RSER 1, then SINM, then FT, you should see a normal 1D ¹H NMR spectrum. Manually phase the spectrum as you would any 1D spectrum EXCEPT that when you are finished, click “RETURN” and select “Save as 2D & return”. Then click the “2D” button to return to 2D spectrum. Enter xau calcphinv to determine phase correction in F1, then xfb again. At this point, the spectrum should be correctly phased.

Plotting:
rpar standard2d: Load only the plot parameters (clicking on plot)
edg: Adjust the values as needed, refer also to the bruker manual.
Zoom the desired region, click on Def-Plot, plot.

¹³C coupled HMQC
eda: Change pulse sequence to invbtp.hein. This is the standard HMQC pulse sequence, only that decoupling on ¹³C is turned off during data acquisition.
record and phase spectrum like normal HMQC.
Remember: Since acquisition takes place on ¹H, the HMQC spectrum contains all homonuclear H-H coupling. Determination of coupling constants is thus only advisable, if the ¹H resonance appears as a singlet.

Homodecoupled ¹H spectra

rpar std1hhd.
set PL14 120.
choose desired SW, AQ, D1. Record normal spectrum, correct phasing. Since the receiver is turned on and off during acquisition to allow homonuclear decoupling, the signal to noise ratio is much worse than in normal experiments. Don’t be surprised.
Copy the parameters to a new experiment with edc
Decide which signal you want to decouple and set O2P to the desired frequency in ppm.
Choose the power of the decoupling pulse, typically PL14 70, never smaller than 54. Setting pl14 to 120 means no power at all. Decreasing this number increases the power of the decoupling pulse by a factor of 2 for approx. 6db. Never ever use a value less than 54, since this might damage the probe head!!
record the spectrum using normal zg – efp sequence. The decoupled peak should more or less vanish.
Adjust PL14 and O2P until the optimal result is obtained. Never set PL14 smaller than 54! You could also play around with AQ and D1.
The best results are obtained with small AQ (0.5s), no D1 (0), NS > 8 and O2P set 0.01 ppm upfield of the desired peak. It will not be possible to decouple broad multiplets or even triplets with big coupling constants completely.
For complicated spin systems and/or overlapping peaks, record one spectrum with identical settings, but PL14 120 and O2P 15, and use the DUAL DISPLAY mode to recognize changes in the spectrum.

Temperature Calibration Plots for NMR spectrometers