NMR Experiments

General Information | Proton-detected | Carbon-detected
Phosphorus-detected | Fluorine-detected

General Information

Sensitivity

• Higher field gives better S/N, so 600>500>400 MHz.
  
  
• Probes optimized for a particular nucleus give the best signal to noise ratio (S:N) for that nucleus.
  
  
  • The cryoprobe on the 500 MHz is an indirect detection probe with the 1H electronics cooled to reduce thermal noise, and is thus most sensitive to proton.
    
    
  • The most commonly used probe on the 400 MHz is the QNP. The QNP (Quad Nuclear Probe) has 4 channels for running direct observation of 1H, 13C, 19F and 31P, as well as many 2D experiments involving those nuclei.
    
    
  • Other probes are also available for all three systems.

Resolution and Dispersion

• Higher field spreads out the signal better, makes coupling patterns simpler, separates different signals better. Again, 600>500>400>300 MHz.
  
  
• Linewidths in Hz are similar whatever the field, so the linewidths in ppm are smaller at the higher field strengths, unless a frequency-dependent line broadening mechanism interferes.

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Proton-detected experiments

• Availability:  H1 experiments are available on all instruments.
  
  
• 1D Proton Experiment
  
  
  • Default Parameters
    
    
    • ~3 second acquisition time
      
      
    • ~2 second relaxation delay
      
      
    • Spectral width -4ppm to 16ppm
      
      
    • 16 scans
      
      
    • Total time: 1 minute
  • Recommended concentration:  0.1 mM
• Variations on the 1D experiment include:
  
  
  • 1D NOE or selective NOE (difference or gradient)
    
    
  • 1D COSY or Selective COSY (gradient)
    
    
  • T1 determination
• 2D COSY - Proton-proton correlation experiment
  
  
  • Gives information about pairs of protons that are J-coupled. This usually indicates that the protons are on adjacent carbons, e.g., 3-bonds away (though protons further apart may in some cases be J-coupled).
    
    
  • Default Parameters
    
    
    • 2 second relaxation delay
      
      
    • 1024 complex points in t2 and 128 increments in t1
      
      
    • 4 scans per increment (no PFG) / 1 scan per increment (PFG)
      
      
    • Total time: 20 minutes (no PFG) / 5 minutes (PFG)
  • Recommended Concentration:  at least 10 mM
    
    
  • Variations on COSY
    
    
    • DQF-COSY
      
      
      • phase-sensitive experiment
        
        
      • diagonal peaks are narrower
        
        
      • less sensitive
    • TOCSY - total correlation spectroscopy
      
      
      • gives correlations for all protons within a spin system
    • NOESY - Proton-proton through-space interactions via NOE
      
      
      • Gives information about pairs of protons that are close in space (<5 A apart)
        
        
      • Default Parameters
        
        
        • 1.5 second relaxation delay
          
          
        • 1024 complex points in t2 and 256 increments in t1
          
          
        • 2-8 scans per increment
          
          
        • Total time: 1 - 5 hours.
          
          
        • Recommended Concentration:  at least 10 mM
      • ROESY - rotating frame NOE
        
        
        • Compounds of molecular weight ~1000-2000
          
          
        • Exchange peaks are opposite sign from NOE peaks
      • HMQC - Heteronuclear Multiple Quantum Correlation experiment
        
        
        • Gives information about strong proton-carbon J-couplings. A strong proton-carbon J-coupling indicates that the proton is directly bonded to the carbon. This experiment gives information that is identical to HETCOR, but because it is proton-detected, it is more sensitive than the standard HETCOR especially on indirect-detection probes.
          
          
        • Default Parameters
          
          
          • 1.5 second relaxation delay
            
            
          • 1024 complex points in t2 and 128 increments in t1
            
            
          • 4 scans per increment (no Pulsed Field Gradient) / 1 scan per increment (PFG)
            
            
          • Total time: 20 minutes (no PFG) / 5 minutes (PFG)
        • Recommended Concentration:  at least 50 mM
      • HMBC - Heteronuclear Multiple Bond Correlation experiment
        
        
        • Gives information about weak proton-carbon J-couplings. A weak proton-carbon J-coupling indicates that the proton is two, three, or four bonds away from the carbon. This experiment gives information about which protons are near to (but not directly bonded to) different carbons. This experiment (in conjunction with the HMQC) can give an enormous amount of information about molecular structure, since the long range proton-carbon correlations can include quaternary carbons, in addition to protonated carbons.
          
          
        • Default Parameters
          
          
          • 1.5 second relaxation delay
            
            
          • 1024 complex points in t2 and 128 increments in t1
            
            
          • 4 scans per increment (no PFG) / 4 scan per increment (PFG)
            
            
          • Total time: 20 minutes (no PFG) / 20 minutes (PFG) (NOTE: PFG HMBC experiments usually have less t1 noise than non-PFG HMBC experiments.)
        • Recommended Concentration:  at least 50 mM

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  Carbon-detected experiments

  • Availability
    
    
    • Probes optimized for X-nuclei give better S/N. The direct detection probes are either QNP or Broadband Observe.  The cryoprobe cannot observe 13C directly at all.
      
      
    • Sensitivity is approximately 5700 times less than for proton, so adequate S/N for a reasonable length experiment (10 minutes, 256 scans) requires a concentration of about 50 mM.
  • 1D Carbon experiment
    
    
    • Default Parameters
      
      
      • 0.8 second acquisition time with proton decoupling
        
        
      • 2.0 second relaxation delay (with NOE enhancement)
        
        
      • Spectral width -10ppm to 225ppm
        
        
      • 256-1024 scans (depending on concentration)
        
        
      • Experiment time:  approximately 1 hour for 1024 scans
        
        
      • Recommended Concentration:  at least 50 mM
    • Special Considerations
      
      
      • Increase d1 for carbons with long T1's (quaternary, carbonyls)
    • DEPT experiment
      
      
      • Gives information about the number of protons bonded to each carbon.
        
        
        
      • Default Parameters
        
        
        
        • 0.8 second acquisition time with proton decoupling
          
          
          
        • 2.0 second relaxation delay (with NOE enhancement)
          
          
          
        • Spectral width:  -10 ppm to 225 ppm
          
          
          
        • 128 scans per spectrum (2 spectra total)
          
          
          
        • Total time:  10 minutes
          
          
          
        • Recommended concentration:  100 mM
      • HETCOR - Proton-Carbon correlation experiment
        
        
        
        • Gives information about strong proton-carbon J-couplings.  A strong proton-carbon J-coupling indicates that the proton is directly bonded to the carbon.  This experiment gives information that is identical to the HMQC experiment, but because it is carbon-detected it is less sensitive than the HMQC.  It does give higher resolution in the carbon dimension, and is often used if regions of the carbon spectrum are crowded.
          
          
          
        • Default Parameters
          
          
          
          • 2.0 second relaxation delay
            
            
            
          • 512 complex points in t2 and 128 increments in t1
            
            
            
          • 4 scans per increment
            
            
            
          • Total time:  30 minutes  (Note that the time required for HETCOR is approximately 6 times the time required for HMQC)
            
            
            
          • Recommended concentration:  100 mM

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  Phosphorus-detected experiments

  • Availability
    
    
    • The 400 MHz NMR can run 31P without changing any cables.  The QNP probe doesn't need any adjustments to observe 31P.
      
      
    • The BBO probe will need to be tuned to 31P before acquisition.
      
      
    • Sensitivity is approximately 15 times less than for proton, so adequate S/N for a reasonable length experiment (10 minutes, 256 scans) requires a concentration of > 0.1mM.
  • 1D Phosphorus experiment
    
    
    • Default Parameters
      
      
      • 0.8 second acquisition time with proton decoupling
        
        
      • 3.0 second relaxation delay (d1; with NOE enhancement)
        
        
      • Spectral width -100ppm to 250ppm
        
        
      • 32 scans
        
        
      • Total time: ~2 minutes
        
        
      • Recommended Concentration:  0.1 mM

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  Fluorine-detected experiments

  • Availability
    
    
    • The 600 MHz NMR with the H/F probe gives the best sensitivity for 19F.
      
      
    • The 400 MHz with QNP probe is well-suited for direct observe 19F experiments.  The QNP probe requires no recabling or additional setup to observe 19F directly.
      
      
    • Sensitivity is approximately the same as for proton, so adequate S/N can be obtained with concentration > 0.1mM.
  • 1D Fluorine experiment
    
    
    • Default Parameters
      
      
      • 0.8 second acquisition time
        
        
      • 3.0 second relaxation delay
        
        
      • Spectral width 150ppm to -200ppm
        
        
      • 64 scans
        
        
      • Total time: 2 minutes
        
        
      • Recommended Concentration:  0.1mM
    • 1D Fluorine observe, 1H decouple
      
      
      • Setup is exactly like direct observe 19F, only F2 is 1H and uses Waltz 16 decoupling.

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