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March 21, 2009.
Indirect chemical shift referencing for heteronucleus

Chemical shifts are relative terms, depending on what the reference is. While the choice of such areference can be arbituary, TMS has be widely accepted as The reference for proton and carbon. For other nuclei, the reference is less agreed upon in the literature. For example, ammonia gas (NH3) has been traditionally regarded as the reference for nitrogen. Partly due to the difficulty in using ammonia gas in solution NMR, IUPAC recently recommended MeNO2 reference, which is about 380ppm downfield from ammonia. On the other hand, it is not always convient to locate a good referencing compound or desired at all to add it to the precious sample. Hence many users struggle to reference their spectra for heteronuclei other than proton or carbon. It might be also confusing to read literatures which either use a secondary reference by declaring its chemical shift out-right or citing some older literature (DISLAIMER: I have nothing against older literatures).

It turns out only a proton reference is needed for heteronuclear chemical shift referencing of almost any compound, which may not contain a proton at all: the chemical shift for a heteronucleus can be found easily through the proton chemical shift, its absolute frequency, the heteronuclear frequency and the lock solvent's chemical shift. Here's the rationale: 1. let's define a reference compound(a) for proton and another reference compound(b). 2. imagine both compounds are added to the sample of interest. 3. we can find their chemical shifts (of course both should be zero) and their abosolute frequencies. Since the chemical shift is independent of the magnetic field (assuming high field), the ratio between those two frequencies should be indifferent to field. In fact, that frequency ratio should be the gyromagnetic ratio. With gyromagnetic ratio as an intrinsic property, the frequency ratio of two references should be a constant. Once that is a constant is measured with sufficient accuracy and authority, it in turn can be used to calculate the heteronucler(0 ppm) frequency based on the proton (0 ppm) frequency. Then the heternuclear spectrum is referenced.

A excel spreadsheet is made for indirect chemical shift referencing based on proton chemical shift and frequency. Contact me if you need further information.


March 26, 2009
Is NMR repsponse genuinely linear?

We have generally regarded NMR as a linear system (see Ernst et al. Principle of 1D and 2D NMR). This is something that a NMR spectroscopist can brag about: NMR signal is strictly linearly proportional to the concentration (or total transverse magnetization) and sine of the excitation angle (ignoring RF inhomogeneity). The question is, is the slope really strictly indifferent to the concentration? If so, a solvent signal (at 100 Molar for water, for example) can serve easily as an internal concentrationon concentration reference to quantitize an analyte in micromole or minimolar range concentration.

The answer is yes but you have to create conditions to overcome the detrimental effects of RF inhomogeneity and radiation damping in reality (see Mo & Raftery "Solvent Signal as an NMR Concentration Reference" Anal. Chem (2008), 80:9835-9839). Furthermore, you have to be aware of some potential complications including conformational or chemical exchanges that may result in multiple resonances for a single nucleus. In most situations, we can assume those complications do not apply. Then we can focus on radiation damping and RF inhomogeneity.

First, radiation damping might be severe if the observed signal is very strong. A typical example is radiation damping of water in a highly protonated aqueous sample. Care has to be taken to reduce the transverse magnetization and the total equilibrium z magnetization. Otherwise, the linear relationship between signal size and sine of the excitation angle may break down. Hence significant errors in quantitation may occur when large excitation angles are used to observe water signal as a solvent concentration reference.

Next time I will give a simple method to find out if the excitation angle is proper for quantitation purpose.


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Last updated: March 16, 2009