Secondary Referencing with Unified Chemical Shift Scale

H. Zhou updated Aug 2015

Background

For referencing chemical shift of any nucleus, including 1H, 19F and X-nuclei, we follow one of several methods recommended by IUPAC whenever possible (proposed in 2001 and updated in 2008). IUPAC recommends the use of a "Unified Chemical Shift Scale" and that ALL chemical shifts be referenced against the 1H signal from very dilute TMS (tetramethylsilane, ~1% or less) in chloroform, either directly or indirectly. The TMS reference is called the "primary reference". You can imagine that all nuclei share the same frequency scale ranging from 0 MHz to 500 MHz (on a 500Mhz spectrometer). In principle, only ONE referencing point is needed for all nuclei. This is the foundation of "unified chemical shift scale" concept.

This unified scale is simply the ratio (Xi) of the resonance frequency of any nucleus relative to the 1H frequency of TMS, and was supposed to replace the traditional chemical shifts (in ppm) all together. But, this reporting method has not seen widespread use. For historic reasons, each nucleus and research field used their own 0ppm reference compound for a particular nucleus (imagine discrete zero ppm points in different regions of the frequency scale). Traditional, individual nucleus 0ppm reference is called "secondary reference" following the introduction of the unified scale (see below). Most chemical shift referencing is now done under the umbrella of the "unified chemical shift" scale but using the ppm referencing against traditional reference compounds.

To continue to use such historic referencing compounds and also to comply with other IUPAC recommendations, IUPAC summarized tables of the frequency ratios of each nucleus in its historic reference compound (i.e P31 in H3PO4, see Tables 1 and 2) over that of TMS 1H signal. This ratio (Xi), measured precisely and field independent, allows indirect establishment of the zero ppm point of traditional (secondary) reference compound once the resonance frequency of the TMS 1H signal (primary) is known. The chemical shifts of all other reference compounds can be calculated based on the calculated TMS 1H frequency using the pre-measured Xi value. In fact, based on a similar principle, if the frequency of the deuterium lock signal (2H) (or any nucleus in general) is known, the ppm value of any other nucleus can also be calculated using the Xi values.

Referencing using the pre-determined Xi ratios along with any resonance with a known chemical shift value in ppm dramatically simplifies chemical shift referencing with traditional reference compounds. This "secondary referencing" method is not super-precise in the traditional sense but it is sufficient for most work conducted here, unless a strict comparison of shift values across multiple samples is needed. In that case, extreme care has to be taken regardless what reference method is used.

Description of Referencing Method in Publication

If a direct, internal or external reference compound is used, this should be stated.

In most other cases, we use the "secondary referencing" method above. The description needs to be clear about how the referencing is done (modify as needed):

Recommended Method

For most applications: If a more accurate chemical shift reference is required:
Note TMS has a melting point of -99C and a low boiling point of 27C. Collect a 1H spectrum at ~25C and reference your spectrum against the TMS 1H signal. Outside this temperature range, you could still use the reference done within the above temperature range, by using ppm values of peaks from your compounds. Alternatively, but less precise, try to use the lock signal from the solvent to roughly reference your spectrum (with the command setref, see below), or choose to have other internal or external reference standards.

If super accurate chemical shift values are desired, you must consider and correct a number of factors that affect chemical shift referencing for each method, such as temperature, solvent, bulk magnetic susceptibility, sample geometry, etc. The truth is that  such occasion arises only rarely and no matter what you do, referencing contains uncertainty once the sample condition deviates from well-known conditions.

More Details on Chemical Shift Referencing Procedures

Use Residual Solvent 1H or Solvent 13C Signal (rough referencing)
This should work sufficiently well for most situations unless a very accurate referencing is needed. Simply locate the residual 1H or solvent 13C signal and set the chemical shifts to the known values.

1H Referencing with Internal TMS (<1%)


For All Nuclei (including 1H, 19F, and X-nucleus)

setref

CAUTION: For some deuterated solvents, there are multiple deuteron peaks. This may cause reference errors with the through-solvent method because the lock position may be one of several positions. In this case, for reliability, you can transfer the reference of a properly referenced 1H spectrum to any other spectrum with the command mref (see below).

mref(source_exp#,target_exp#)

In some situations, you may obtain more accurate X-nucleus referencing by using a carefully referenced 1H spectrum (using an internal TMS signal for example) to indirectly reference all other nuclei. However, if the 1H referencing is rough (such as from solvent residual signal or lock signal), you might as well just use the referencing by the lock (setref) method for the other nucleus.

reff1, reff2