Hyphenated NMR in Drug Analysis and Drug Metabolism.
The new equipment installed autumn 2002 at the Royal Danish School
of Pharmacy greatly facilitates the use of NMR in drug metabolism
and drug analysis, as the greatly enhanced sensitivity compared with
equipment yields substantially better detection limits. Furthermore,
the 1 mm probe installed and tested on the AV 400 instrument during
2005, extends the detectionlimit of the AV 400 downto the sub-nanomolar
My research will
mainly be centred on the following subheadings.
1. Drug metabolism studies.
When studying drug metabolism identification and quantification of
and parent compounds in complex biological fluids is inevitable. A
of separation and spectroscopic techniques is therefore needed to work
and obtain reliable results.
Identification of drug metabolites using LC-NMR-MS (Liquid
coupled with Nuclear Magnetic Resonance and Mass Spectroscopy) is very
when both NMR and MS data are available for the different metabolites.
the Solid Phase Extraction (SPE) equipment installed with the new
will make it possible to obtain useful spectra at substantially lower
Especially the fact that one can be fairly sure, that the NMR and MS
are from the same compound facilitates reliability of analysis. Our
is, that this is a crucial point. The possibility of using smaller
improves detection limits as well as making it affordable to use
organic solvents for the Liquid Chromatography which in turn allows
to some important regions of the spectra otherwise obscured by solvent
Chemometric techniques (subheading 4) will further enhance the spectra
The actual compounds analysed are chosen in collaboration with
at the Royal Danish School of Pharmacy.
Refs. 3, 8, 9, 13, 16, 19, 22, 23,29, 32, 35, 36, 37,40, 45, 46, 48,
A Ph.D.-project (Michael Lauridsen) was started medio 2004 concerning
metabonomic studies of inflammatory diseases (Morbus Crohn, Colitis
Ulcerosa, and Rheumatoid Arthritis and Osteoarthritis) in collaboration
with the Department of Medicinal Chemistry and the Parker Institue at
A collaboration concerning identification of biomarkers for
fruit- and vegetable ingestion is in progress with the Danish
Institute for Food and Veterinary Research.
3. Chiral resolution of NMR spectra of drugs and drug metabolites
NMR can’t distinguish between enantiomeric compound in itself, but by
chiral cosolutes (cyclodextrins eg.) or chiral solvents the solutes in
form complexes that are diastereomeric, such that enantiomers can be
an identified, and enantiomeric excess determined, which is
for drugs. Specifically we plan to examine the use of Solid Phase
coupled with HPLC and NMR in such a way, that an enantiomer or
pair is trapped on an SPE-column during HPLC and eluated with a chiral
into the NMR, where identity is established at the same time as
excess. Some interesting solvents can be found in the literature,
solvents with DMSO-like properties which is better suited for most
Refs. 14, 51-54.
4. Drug analysis.
Both analysis of impurities in drugs and analysis of complex drugs or
obtained from natural sources such as plants are highly amenable to
analysis. Identity of drug impurities are very important. Knowledge of
compounds in complex plant extracts is very important in order to
the quality of the drugs or in order to develop new drugs bases on
from natural sources.
As of 2005 an industrial Ph.D. project under DRA in collaboration with
Novo Nordisk, KVL, an dSchool of Pharmacy University College London was
started (Carsten Ravn): Increased
and physico-chemical knowledge of pharmaceutical solid dosage
design using fast near infrared spectroscopy and chemical imaging
technologies”.(NIR chemical imaging in pharmaceutical formulation
design). The project is implemented with an intention of developning
and evaluating the techniques for Process Analtyical Technology.
Refs. 1, 2, 4, 5, 6, 7, 10, 11, 15, 17, 20, 21, 24, 25, 26, 27, 30, 31,
34, 38, 39, 41, 42, 43, 44, 47.
5 and 6 will largely be performed as an aid to 1, 2, 3, and 4
5. Extraction of NMR spectra and spectral features using chemometric
Chemometrics can be defined as the use of mathematical and statistical
with the object of getting the most out of Your data. The application
chemometric techniques to analysis of complex matrices is a rapidly
area of analytical chemistry, also in the area of complex biological
Using chemometric techniques spectra can be extracted from
run in “screening mode”. An NMR-chromatogram can be subjected to factor
techniques in such a way, that individual spectra of an improved
(compared to extracting slices) can be obtained using the complete
in the NMR-chromatogram. A spectrum and its separate chromatogram is
for each discernible species in the chromatogram.
Spectra of the pure compounds can even be obtained in cases of
peaks, which makes it possible to obtain information otherwise obscured
solvent peaks, as long as the solvent peak doesn’t move during the
or the solvent signal doesn’t change multiplicity and linewidth.
The improved stability (less drift, better baseline in the spectra) of
new NMR-equipment is an important aspect of the future development of
methods, as the chemometric techniques can’t separate artifacts created
instability of the instrument from “real” changes of features of
See Reprint of latest poster “Moore or less automatic extraction of
from NMR-chromatograms presented at the 23.rd Danish NMR-meeting
at Hotel Scandic, Roskilde, Denmark.
Lecture at the 22. Danish NMR meeting at Ledernes Kursuscenter, Odense,
“Screening of plant- and other extracts by means of an old NMR
(keywords: LC NMR, Chemometrics, Plant Extracts)
6. Determination of physical parameters of drugs and drug
using NMR and other spectroscopi methods.
Physical parameters such as pK-values, solubilities, complexation
etc. are very important for separations techniques, drug formulations
Using NMR it is possible in many cases to determine these parameters
at low concentrations and for complex mixtures. Interaction of drugs
and excipients with metal-ions, using UV, IR, NIR, Raman, NMR, a.o
Refs 1, 18.
Link to References
Last update: 2/3-2007, cc