Page 464 - Tandem Techniques

Page 448

and likely to be fruitless. Under such conditions, sample size, column diameter, and the shape and size
of the NMR cell itself, may need to be changed to obtain greater sensitivity.

Employing 750 MHz Spectrometer in LC/NMR Tandem Systems

As the development of NMR spectrometers has continued, stronger and stronger magnetic fields have
become attainable and, as a consequence, the sensitivity of the spectrometer has increased which, in
turn, has rendered the LC/NMR tandem instrument more practical. Sidelmann et al. [6] coupled the
liquid chromatograph to a 750 MHz NMR spectrometer to examine the positional isomers of 6,11-
dihydro-11-oxodibenz[b,e]oxepin2-acetic acid. NMR spectroscopy is perhaps the only simple technique
for examining this type of problem. The details of the chemistry need not concern us here, but the
sensitivity and spectra quality that was obtained are both interesting and exciting.

The chromatograph was fitted with a Bruker LC22C pump and an LC33 Variable wavelength UV
detector operated at 200 nm. The column was a Spherisorb ODS-2, 25 cm long, 4.6 mm I.D., and
packed with 5 µm particles. The mobile phase employed was acetonitrile : 0.2 M potassium phosphate
(pH 7.4) : deuterium oxide (21:10:69) at a flow rate of 1 ml/min. and an excellent separation was
obtained. The NMR spectrometer was the Bruker DMX 750 MHz model fitted with a H flow probe.
1
The results were obtained from 1.5 µl sample of a 0.15% solution of the solutes. The position of all the
groups were unambiguously identified and it is probable that the tandem combination of liquid
chromatography and high resolution NMR was only really effective way of examining this chemical
system.

Sidelmann et al. [7] also employed a LC/NMR tandem system to characterize the positional isomers
and anomers of 2-3- and 4-fluorobenzoic acid glucuronides in equilibrium mixtures. The apparatus was
essentially that described previously, and the NMR chromatogram they obtained was by directly
coupled continuous-flow 750 MHz-LC-NMR of the O-(3-fluorobenzoyl)-D-glucopyranuronic acid
mixture The NMR