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Nuclear magnetic resonance (NMR) spectroscopy is a powerful, nondestructive technique capable of complete structural and
conformational analysis of complex molecules, quantitative analysis of complex mixtures, and noninvasive measurement of reaction
rates of chemical systems in the test tube and in intact living
organisms, including humans. NMR was discovered simultaneously by two independent laboratories in 1946 (Tl, T2).
Subsequently, high-resolution NMR was quickly developed by
analytical chemists as a powerful technique for the determination
of molecular structure. The fundamentals of NMR technology
are described in specialty texts to which interested readers are
referred (T3-T6). In the first section of this review, we will
provide a brief overview of the basics of NMR spectroscopy
including theory and principles of NMR, instrumentation, technical
limitations, and spectral interpretation methods. At this point we
shall quickly shift to the designation MRS, magnetic resonance
spectroscopy, which has been adopted by clinical proponents to
avoid any negative implications of nuclear.
The development of MRS as a clinical analytical tool has been
spurred on by the widespread use of a related analytical technique,
magnetic resonance imaging (MRI), in clinical medicine. The two
are related since they both utilize the same physical phenomenon,
NMR; MRS emphasizes spectral or chemical information, whereas
MFU emphasizes spatial information. The techniques and applications of MRI will not be addressed in this review. Interested
readers are referred to recent texts and articles (77-23).
MRS is unique in its capability to provide nondestructive in
vivo and in vitro chemical analyses. While significant advances
have been made in in vivo MRS, this has recently been reviewed
elsewhere (TI@. The present review for clinical chemistry
highlights some of the research that has been directed toward in
vitro chemical analysis, namely, physiological fluids, tissue specimens, and tissue extracts. This application of MRS in pharmacology and toxicology is already quite well established; the main areas
of current development are metabolic disorders, organ transplantation, neurological disorders, and cancer. The review covers the
time period of 1990 to the present. Our previous review covered
the literature up to that time ( T l l ) .
THE BASICS OF NMR SPECTROSCOPY
Theory and Principles. Spectroscopy is the measurement
of the frequency dependence of absorption or emission of energy
by a system. NMR refers to the absorption and release of radio
frequency (rf) energy by a nucleus in a magnetic field. Possession
of both charge and spin renders some nuclei magnetic and confers
various properties on them which affect their behavior in an
external magnetic field. One such property is a magnetic moment
@). In an external magnetic field (Bo),the magnetic moment of
a spinning nucleus will precess, or describe a cone, around the
direction of the field. The precessional frequency of a particular
nucleus is proportional to the strength of the magnetic field.
To observe resonance, the nuclei must be irradiated with
electromagnetic (I$radiation, the frequency of which must match
the precessional frequency of the nuclei. The rf energy is then
absorbed by nuclei in the lower energy spin state, raising them
to the higher energy spin state. In actual fact, upward and
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(2-24-T28).
CLINICAL APPLICATIONS OF NMR
SPECTROSCOPY
The analysis of physiological fluids by high-resolution MRS is
a relatively recent application of the NMR phenomenon in clinical
medicine. Despite the fact that MR spectra are rich in information
on endogenous biochemical processes in health and disease, and
that quantitation of metabolites can be readily achieved, the
diffusion of MRS methods into the clinical laboratory remains slow.
While the major technical limitations have been overcome, MRS
of physiological fluids competes with long-established biochemical
methods that are well accepted, highly automated, comparatively
inexpensive, and readily available at most clinical sites. Moreover,
data collection and interpretationis limited by the scarcity of MRS
trained individuals able to exploit fully the wealth of information
in the spectrum of a fluid, tissue, or tissue extract.
A large number of physiological fluids is accessible for MRS
studies in vitro. The first medical applications showing the utility
of H MRS analysis of complex metabolite mixtures involved the
analysis of urine and serum (224,T29,T30).A variety of other
fluids including cerebrospinal fluid (CSF), amniotic fluid, synovial
fluid, sweat, aqueous humor, seminal plasma, saliva, bile, ascites
fluid, and tissue extracts have since been examined. All physiological fluids are not equally available in terms of quantity and
ease of availability (technical difticulties, patient benefit, patient
discomfort, ethical issues). Nevertheless, samples are drawn in
a variety of clinical situations where it would seem prudent to
extract as much information as possible out of as small a specimen
as possible, particularly in situations where a specimen is difficult
to obtain. It is here where MRS can offer distinct advantages over
conventional biochemical analytical techniques: MRS analysis
requires a small sample volume (0.2-0.5 mL), which generally
remains intact during measurement and thus can be used for
subsequent assay by other techniques; the specimen generally
requires no or very little pretreatment; spectra take only a few
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to monitor the severity of erythroblastosis fetalis, and measurements of amniotic fluid a-fetoprotein and acetylcholinesterase are
useful in the prediction of open neural tube defects. Conventional
biochemical tests have various technical limitations, and thus MRS
may be particularly well suited to the analysis of amniotic fluid.
While high-resolution 'H MRS was first used to characterize
amniotic fluid for a variety of components including amino acids,
lactate, and glucose (T52),recent work has generally focused on
two areas, namely, 31PMRS analysis of phospholipid extracts of
amniotic fluid (T52-T54) and quantitation of the constituents of
amniotic fluid and their clinical correlation by 'H MRS (T55T57).
(a) Fetal Lung Maturity. Testing for fetal lung maturity has
traditionally been done by the measurement of the lecithin/
sphingomyelin ( W S ) ratio via numerous thin-layer chromatographic techniques. Novel techniques, such as fluorescence
polarization and lamellar body number counts, have been proposed recently to decrease technical difficulties and increase
turnaround time. In a preliminary study, 600-MHz 'H MR spectra
of untreated amniotic fluid specimens from 66 patients were
analyzed. Linear discriminant analysis was performed to determine how well the peak ratios could predict the fetal maturation
category, as determined by either L/S ratio or fluorescence
polarization. Of 43 third-trimester fluids, 65%were placed in the
correct category-immature, transitional, or mature (T55). While
this is a reasonable prediction of fetal lung maturity, it lacks the
sensitivity and specificity required for a clinical test. However,
the metabolites measured likely do not relate specifically to
pulmonary surfactant; better agreement might be anticipated when
more relevant compounds such as phosphatidylcholine and
phosphatidylglycerol are analyzed. 31P MR spectra of phospholipids in human amniotic fluid have been obtained recently (T52T54), but clinical correlations were not attempted.
(b) Fetomaternal Complications. MRS of human amniotic
fluid yields a wealth of information on chemical content and its
variation with the condition of the mother (T58). To determine
how concentrations of the various metabolites of amniotic fluid
detected by MRS may relate to the clinical status of the fetus and/
or the mother, a number of fetomaternal complications were
studied. No differences in peak intensity ratios were observed
for mothers with gestational diabetes or in cases of fetal trisomy
21 where the spectra were generally normal in appearance (T55).
Amniotic fluid from mothers with preeclampsia, on the other hand,
showed differences in peak intensity ratios for choline, succinate
and acetate. Linear discriminant analysis correctly distinguished
all cases (n = 5) of open spina bifida where the MRS spectra were
markedly altered: lactate, glutamate, and acetate concentrations
were significantly increased. New peaks, previously not detected
in normal amniotic fluid were found, and other peaks normally
present were absent (T55). Resonances observed at 6-8 ppm in
the MR spectrum of amniotic fluid have also been observed in
the same region in MR spectra of human urine. It has been
suggested that these resonances might be useful as markers for
fetal renal output (T56). In addition, many low molecular weight
compounds in amniotic fluid have been reported to be of clinical
importance: amino acid elevations have been reported in central
nervous system disorders (T59); glucose concentrations have
been used in the diagnosis of intraamniotic infection (2'60);lactic
acid has been associated with fetal acidosis (7'59). Thus, the
ability of MRS to provide a high-resolution spectrum with the
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