Raman Spectroscopy and Microspectroscopy

Raman spectroscopy is an inelastic light scattering experiment for which the intensity depends on the amplitude of the polarizability variation associated with the molecular vibration under consideration. The polarizability variation is represented by a second-rank tensor, oiXyZ, the Raman tensor. Information about orientation arises because the intensity of the scattered light depends on the orientation of the Raman tensor with respect to the polarization directions of the electric fields of the incident and scattered light. Like IR spectroscopy, Raman 

For orientation measurements, this tensor also needs to be expressed in the coordinate system OXYZ, axrz, using the matrix transformation u.xyz = Oaxyz / where O is a matrix whose elements are the direction cosines of the coordinate axes and J is its transposed matrix [44]. 

The theory of orientation measurements by linearly polarized Raman spectroscopy has been developed in detail by Bower in 1972 [44]. The Raman intensity, I, is given by 

For a uniaxial sample, there are five independent quantities (a,yap( ) or (a ) given 

An approach simpler than the general method has been used by Bower and coworkers [52,53] and has proved to be useful to determine the order parameters of uniaxially oriented polymers. This approach, called cylindrical method, requires a smaller number of spectra that are easily measurable, but it assumes that the Raman tensor associated with the vibrational mode under study is cylindrical. In such case, at — a2 — a so that the depolarization ratio R so of an 

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Also the infrared microspectroscopy (IR) is a vibrational spectroscopy, but it presents some differences with respect to Raman spectroscopy and also provides different information. In infrared spectroscopy the sample is radiated with infrared light, whereas in Raman spectroscopy a monochromatic visible or near infrared light is used. In this way, the vibrational energy... 

Fig. 8.1 Raman microspectroscopy as combination of surface-enhanced Raman spectroscopy and confocal microscopy...

Raman spectroscopy s sensitivity to the local molecular enviromnent means that it can be correlated to other material properties besides concentration, such as polymorph form, particle size, or polymer crystallinity. This is a powerful advantage, but it can complicate the development and interpretation of calibration models. For example, if a model is built to predict composition, it can appear to fail if the sample particle size distribution does not match what was used in the calibration set. Some models that appear to fail in the field may actually reflect a change in some aspect of the sample that was not sufficiently varied or represented in the calibration set. It is important to identify any differences between laboratory and plant conditions and perform a series of experiments to test the impact of those factors on the spectra and thus the field robustness of any models. This applies not only to physical parameters like flow rate, turbulence, particulates, temperature, crystal size and shape, and pressure, but also to the presence and concentration of minor constituents and expected contaminants. The significance of some of these parameters may be related to the volume of material probed, so factors that are significant in a microspectroscopy mode may not be when using a WAl probe or transmission mode. Regardless, the large calibration data sets required to address these variables can be burdensome. 

Wright of Advanced Micro Devices discusses the use of Raman microspectroscopy to measure the integrity of a film on semiconductor wafers during manufacture in US patent 6,509,201 and combined the results with other data for feed-forward process control [181]. Yield is improved by providing a tailored repair for each part. Hitachi has filed a Japanese patent application disclosing the use of Raman spectroscopy to determine the strain in silicon semiconductor substrates to aid manufacturing [182]. Raman spectroscopy has a well established place in the semiconductor industry for this and other applications [183]. 

Written by an international panel of experts, this volume begins with a comparison of nonlinear optical spectroscopy and x-ray crystallography. The text examines the use of multiphoton fluorescence to study chemical phenomena in the skin, the use of nonlinear optics to enhance traditional optical spectroscopy, and the multimodal approach, which incorporates several spectroscopic techniques in one instrument. Later chapters explore Raman microscopy, third-harmonic generation microscopy, and nonlinear Raman microspectroscopy. The text explores the promise of beam shaping and the use of a broadband laser pulse generated through continuum generation and an optical pulse shaper. 

A major advantage of Raman spectroscopy in the chemical characterization of pollen is its applicability in situ. The pollen grains in this study were investigated without prior purification or extraction procedures, which has been the case in most other investigations on pollen composition so far [45, 46, 57, 58]. Using Raman microspectroscopy on sections of snap-frozen samples, the chemical composition and very likely also their ultrastructure remain unaltered, and the co-localization of individual components could be studied in the context of pollen micromorphology, simultaneously in very short... 

Beyond imaging, the combination of CRS microscopy with spectroscopic techniques has been used to obtain the full wealth of the chemical and the physical structure information of submicron-sized samples. In the frequency domain, multiplex CRS microspectroscopy allows the chemical identification of molecules on the basis of their characteristic Raman spectra and the extraction of their physical properties, e.g., their thermodynamic state. In the time domain, time-resolved CRS microscopy allows the recording of the localized Raman free induction decay occurring on the femtosecond and picosecond time scales. CRS correlation spectroscopy can probe three-dimensional diffusion dynamics with chemical selectivity. 

A series of advances over the past decade have made CRS microscopy a highly sensitive tool for label-free imaging and vibrational microspectroscopy that is capable of real-time, non-perturbative studies of complex biological samples based on molecular Raman spectroscopy. In particular, biomedical applications where fluorescent labeling of small molecules represents a severe pertur-... 

Microspectroscopy Raman spectroscopy is a scattering technique and hence specimens do not need to be fixed or sectioned resulting in collecting spectra from a very small volume (<1 pm in diameter) leading to identification of species present in that volume. Raman spectroscopy, therefore, offers several advantages for microscopic analysis, particularly, suitable for microscopic... 

Fiber stress determination is of major importance for the modeling of composites and it is now weU established that Raman microspectroscopy, with its main advantage being its nondestructive nature, makes it possible in composites. First, Galiotis and Young demonstrated that Raman spectroscopy is an excellent method to follow the deformation of aramid and carbon fibers. This is a result of variation in the (stretching) vibrational wave number, as a consequence of the anharmonicity of the interatomic bonds. The relationship linking Raman wave number shifts (Av) to the tensile strain (Ae) is hnear, Av = TAe. 

As described in the following sections, IR and Raman spectroscopy, using modem Fourier transform techniques such as IR microspectroscopy, offer excellent analytical tools for the burgeoning field of combinatorial chemistry. 

A general advantage of Raman spectroscopy is the extended spectral range, and the Stokes shift (2) in FT-Raman spectra is usually recorded from 3500 to 50 cnr1. Like IR internal reflection spectroscopy (see Sec. Ill) and IR microspectroscopy (see Sec. IV), Raman spectroscopy is a nondestructive technique requiring little or no sample preparation. 

Techniques that apply to in situ analysis of the dosage form, its precursor granulations, or powders are discussed. Applications of solid-state NMR, FTIR microspectroscopy, visual and scanning electron microscopy, Raman spectroscopy, NIR analysis, thermal techniques, mass spectrometry, and imaging techniques are presented. 

These FT-Raman and CCD-Raman spectrometers revolutionized Raman spectroscopy such that, within the space of about five years, about ten different Raman spectrometers based on multiplex and multicharmel technologies had been introduced commercially [32, 33]. Several of the CCD-Raman spectrometers were either designed for, or could be readily modified for, microspectroscopy. Although FT-Raman microspectrometers have been reported (e.g.. Ref [34]), they have not proved very popular for three reasons ... 

Additional techniques such as FT-IR microspectroscopy, IR and Raman spectroscopy, NMR, and energy dispersive x-ray, in conjunction with scanning electron microscopy, inductively coupled plasma, etc., may also be utilized to provide additional pieces of information toward the comprehensive analysis of materials and the identification of unknowns. Although HSM may not be a technique that all laboratories require, it is clear that the technique can provide valuable information for the visual confirmation of thermal transitions. 

Raman spectroscopy was also reported to be useful for studying the molecular nature of human SC (Williams etal., 1992a). Recently, Raman microspectroscopy is being used to characterize the Upid domains of SC (Percot and Lafleur, 2001). Combined with confocal microscopy, it is useful as a noninvasive in vivo optical method to measure molecular concentration profiles in the skin (Caspers et al., 2001), which allows measuring percutaneous absorption along with detailed information about the molecular composition of the skin with high spatial resolution (Caspers et al., 2003). 

In principle, Raman spectroscopy is a microtechnique [161) since, for a given light flux of a laser source, the flux of Raman radiation is inversely proportional to the diameter of the laser-beam focus at the sample, i.e., an optimized Raman sample is a microsample. However, Raman microspectroscopy able to obtain spatially resolved vibrational spectra to ca. 1 pm spatial resolution and using a conventional optical microscope system has only recently been more widely appreciated. For Raman microspectroscopy both conventional [162] and FT-Raman spectrometers [ 163], [ 164] are employed, the latter being coupled by near-infrared fiber optics to the microscope. 

The methodology of Raman spectroscopy is rapidly becoming matured. Here, we describe two classes of new Raman spectroscopies, Raman microspectroscopy and low-frequency Raman spectroscopy, that are considered to be important for future applications to ionic liquid studies. 

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