Temperature calibration

Temperature is an often overlooked parameter when recording NMR spectra. Temperature directly affects molecular dynamics so changing the temperature can change the appearance of an NMR spectrum. Peaks, particularly those of exchangeable protons, move as the temperature is altered. Interconversion of conformers can be slowed or increased with temperature so that peaks can separate or coalesce. This allows determination of thermodynamic parameters, as long as the temperature can be accurately measured. All modern NMR spectrometers provide temperature control, but the systems need to be calibrated.

To determine the temperature of an NMR sample one needs to place a thermocouple inside the tube, but spectra cannot be recorded with the thermocouple present. One can, however, measure the temperature at several settings of the temperature control system, then record spectra of a sample whose peaks are sensitive to temperature at those settings. The figure below shows spectra of methanol recorded at various temperatures.

Plotting the chemical shifts against the measured temperature produces a calibration curve. It is also possible to fit an equation to the calibration data so that the temperature can be readily calculated.

The most commonly used compounds are methanol and ethylene glycol. Methanol is used for lower temperatures (170-320K) and ethylene glycol at higher ones (300-410K). For cryoprobes the best sample appears to be neat methanol-d₄ in a sealed NMR tube¹. This is the standard used at the Skaggs NMR Facility. This standard produces a spectrum with two peaks; one for the methyl group and one for the hydroxyl. As the temperature is increased the hydroxyl peak moves upfield (to the right). Rather than measuring the chemical shift of the peaks (which would require adding a reference compound for accurate results) the distance between the two peaks can be measured. The Bruker TopSpin software provides a macro, calctemp, that given the standard used, will identify the appropriate peaks in the spectrum, measure the chemical shift difference, and report the temperature.

The probes at the NMR Facility are usually calibrated after installation. This allows correction factors to be defined that correct the temperature control system so that the desired temperatures are obtained. Typically, the actual temperature will be within one degree of the setting. If you are doing dynamics experiments and trying to extract quantitative results, however, it is well worth measuring the actual temperature.

References
1. A ¹H‐NMR thermometer suitable for cryoprobes
M. Findeisen, T. Brand, S. Berger
Magn. Reson. Chem. 2007 45(2) 175-178