Page 380 - Academic Press Encyclopedia of Physical Science and Technology 3rd Analytical Chemistry

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Encyclopedia of Physical Science and Technology EN0011A-541 July 25, 2001 17:27

Organic Chemistry, Compound Detection 483

from a heated, linear capillary nozzle toward the surface tube that is electrically biased with respect to an entrance
of a slowly rotating germanium disc. The microdrops are orifice of a MS. The electrode shapes, geometries, and
constrained by a heated gas sheath that also rapidly va- potentials are important for penetrating a stable spray of
porizes the solvent, leaving a particle stream of solute droplets; however, many different experimental configu-
material that collects on the disc surface. Instrumentation rations can be used. Two widely held views of the mech-
based on this technology is used for a wide variety of ap- anism for ESI are that solvent-free molecular ions are
plications in polymer, pharmaceutical, and environmental formed either after a series of droplet fissions, followed
laboratories. by solvent evaporation from the ion, or by ion evaporation
The recent development of electrospray ionization from the droplet surface.
(ESI) and matrix-assisted laser desorption-ionization
(MALDI) sources for creation of macromolecular ions
C. Fourier Transform Raman
has brought about a renaissance in the field of mass spec-
Spectroscopy (FT-RS)
trometry (MS).
It is now common to determine high molecular weights Raman spectroscopy (RS) has been known for more than
such as proteins to within a few Daltons. 60 years. It is during the past decade that the organic
MS-based methods for sequencing are proliferating chemist has included RS as an additional tool for struc-
and replacing or complementing traditional biochemical ture determination. The Raman spectrum provides com-
methods for many applications. Mounting evidence sug- plementary information to the IR spectrum in that both
gests that some conformational properties of biomolecules are vibrational spectra. Often the weakest signals in the IR
in solution are preserved during the ionization process and spectrum are the strongest signals in the Raman spectra.
persistoverthetransiencetimethationsexistinmassspec- Modern Raman spectrometers operate with a monochro-
trometers. matic coherent laser light source. Sampling handling is
In the past 5 years a new field involving structural much easier in RS than in IR spectroscopy. The sample
studies of biomolecular ions has emerged. Studies of can be dissolved in a number of solvents since their Raman
biomolecular ions in the gas phase provide informa- absorptions are more limited than in the IR. Solid samples
tion about the nature of conformation in the absence of can also be examined as powders or crystals. Liquids can
solvent: be placed in small quartz or Pyrex glass capillaries, and the
The structural studies of protein ions have been grouped scattered radiation is observed through the open end of the
into two categories: chemical probes, in which structural capillary.Samplescanalsobeexaminedwithoutremoving
characteristics are deduced by monitoring the products them from their bottles. If several spectroscopic analyses
of ion-molecule reactions, and physical probes such as are to be performed on a small amount of an organic sam-
measurements of dissociation patterns and measurements ple, then the RS is obtained first because it is totally nonde-
of cross sections. structive and does not require dilution in solvents, as in the
An important factor in folding of proteins involves caseofUVorNMR.Certaingroupfrequenciessuchaspri-
solvent–molecule interactions. Proteins appear to form mary amines, alkynes, and nitriles are strongly absorbed
physiologically relevant conformations in the presence of in the RS but are difficult to observe in the IR spectrum.
a minimum hydration shell. Raman spectroscopy has a number of experimental advan-
Recent studies have considered structural changes that tages over IR: (a) Window problems hardly exist, if visi-
occur when proteins are lyophilized by examining anhy- ble or near IR lasers are used as sources; glass is a robust
drous powder and thin films. Techniques such as FTIR window material with excellent transmission; (b) since
spectroscopy, Raman spectroscopy, and H-D exchange transmission through the sample is not involved, very lit-
NMR methods have shown structural changes upon re- tle preparation is required in RS; (c) water, which has an
moval of solvent. extremely strong IR spectrum, produces only a weak RS
When the proteins are heated, they denature, making and is the solvent of choice in biochemical and aqueous
them less volatile; the molecules decompose rather than studies.
vaporize. ESI has offered a simple, efficient means of Raman spectra are often plagued by the problem of flu-
producing gaseous, anhydrous biomolecules for study by orescence when conventional shorter (visible) wavelength
MS. Thus, anhydrous proteins also fold and unfold, and is used. It was estimated that from 1966 until 1989 less
new conformations are established in a grossly simpli- than 20% of the samples studied gave suitable spectra
fied environment whose only intramolecular constraints with visible laser Raman dispersive spectrometers. Even
are relevant. so, considerable time is wasted in obtaining a reason-
Charged droplets containing molecular ions are initially able Raman spectrum. Since 1987 Near Infrared Fourier
formed by pumping a solution through a narrow capillary Transform Raman Spectroscopy (NIR-FTRS) has been