Optical techniques microspectroscopy

Cherry RJ. New Techniques of Optical Microscopy and Microspectroscopy, CRC Press, Boca Raton, FL, 1991. 

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. 

Raman microspectroscopy results from coupling of an optical microscope to a Raman spectrometer. The high spatial resolution of the confocal Raman microspectrometry allows the characterization of the structure of food sample at a micrometer scale. The principle of this imaging technique is based on specific vibration bands as markers of Raman technique, which permit the reconstruction of spectral images by surface scanning on an area. 

A third class of sampling geometries involves Raman microscopy and closely related Raman imaging techniques. Combination of a Raman spectrometer with a modified optical microscope permits spectra to be obtained from very small sample regions, down to less than 1 pm laterally and a few microns in depth (Fig. 6.3). Raman microspectroscopy is a term generally used to describe this spatially resolved technique in which spectra are obtained... 

The technique of FT-IR internal ATR has been developed to the point that, today, ATR mirror lenses are available for an IR microscope. Furthermore, a newly developed, dedicated diamond internal reflection instrument, the lUuminatIR (Smith s Detection, Shelton, CT, USA) has now joined the ranks of microspectroscopy. This instrument incorporates a small, horizontally mounted diamond, on the surface of which is placed the material to be examined. In this way, the material is in optical contact with the diamond, and is held in place by a shaft pressing down from above. In this case, the radiation enters from beneath the instrument at an appropriate angle, and internally reflected rays are subsequently collected. The specimen is illuminated from beneath with a near-IR source that is detected and displayed on a video screen. With this optical arrangement, it is possible to locate a particular part of the material in the field of view and to interrogate it Such an arrangement is particularly user friendly, and indeed it is mostly used by... 

Recrystallization experiments frequently yield crystals having different shapes and morphologies which are not necessarily different polymorphs. For example, Figs 8.2 and 8.3 show crystals of p-estradiol with distinctly different shapes but are, in fact, the same polymorph. The morphology differences are due to different crystallization solvents. It is important then to have some microscopical technique that allows one to distinguish between polymorphs. Optical crystallography, thermal microscopy and microspectroscopy have this ability. 

Peters, R., and Scholz, M. (1991). Fluorescence photobleaching techniques. In New Techniques of Optical Microscopy and Microspectroscopy (R. J. Cherry, ed.), pp. 199-228. Macmillan, London. 

FTIR microspectroscopy is a microanalytical technique, which interfaces an FTIR spectrometer to an optical microscope. Regions of interest in the sample are spatially isolated using the microscope s apertures. It enables the IR spectrum of sampling regions down to about 10 pm resolution to be taken. Consequently, FTIR microscopy is ideal for compositional mapping and analysis of heterogeneous samples whose domain sizes are in the tens of micrometre range. 

These examples cQso illustrate the difference in spatial resolution and contrast mechanisms between optical and infrared microscopies. While optical microscopy is capable of higher spatial resolution, its discrimination is limited to a difference in the average of a property of materials, namely refractive index, unless specially labeled to detect a property of the label. Infrared microspectroscopy derives its contrast mechanism from the intrinsic composition of the material but suffers from a poorer spatial resolution. A judicious use of the two complementary techniques is often required to achieve good characterization. While the example above illustrated the detection of differences, FTIR microspectroscopy can also be used to determine homogeneity. For example, compositional differences in a PP-PE film could not be detected between the surface and up to 500 pm into the bulk of the sample [61]. 

Industrial Analysis with Vibrational Spectroscopy 5 Ionization Methods in Organic Mass Spectrometry 6 Quantitative Millimetre Wavelength Spectrometry 7 Glow Discharge Optical Emission Spectroscopy A Practical Guide 8 Chemometrics in Analytical Spectroscopy, 2nd Edition 9 Raman Spectroscopy in Archaeology and Art History 10 Basic Chemometric Techniques in Atomic Spectroscopy 11 Biomedical Applications of Synchrotron Infrared Microspectroscopy 12 Microwave Induced Plasma Analytical Spectrometry 13 Basic Chemometric Techniques in Atomic Spectroscopy, 2" Edition... 

This technique has been used for the identification of undispersed particles in plastic applications. It is particularly useful for resolving customer complaints as well as process development problems. As a general rule particles 20 microns or larger can be analyzed by FT-IR spectroscopy. The approach is to focus the IR beam onto the particle of interest using the microscope, and then scan the FT-IR spectra several times (100 scans), either in transmission or reflectance mode. The sample is then moved slightly to another position and the microscope is focused on a portion of the sample without a defect. The FT-IR spectrum of this part of the sample is recorded in exactly the same way as that of the defective part. The spectrum of the non-defective part is then subtracted from spectrum of the defective part of the sample. The difference spectrum is then used to identify the spot or particle in the defective part. Optical microscopy is often used together with FT-IR microspectroscopy to aid in selecting the area of interest to be analyzed. 

On-line/in-line technology for monitoring extrusion processes, including FTIR microscopy, near-IR spectroscopy and optical microscopy was reviewed [500]. Several reviews describe uFTIR applications to polymers [458,501]. Line map applications of /U.FTIR have been discussed [491]. A recent review [502] refers to a large number of FTIR mi-crospectroscopic studies as an important source of structural and spatial information for polymer-based articles. A monograph describes applications of FTIR microspectroscopy to polymers [393]. ASTM E 334 (1990) describes the general techniques of infrared microanalysis. 

The future of Raman microspectroscopy is probably imaging and optical near-field nano-Raman spectroscopy [529], cfr. Chp. 5.5.2. While conventional laser Raman spectroscopy samples 10 g (mm ), /zRS handles 10 g (nm ) and near-field Raman spectroscopy 10 g (nm ). Mobile Raman microscopy (MRM) allows in situ Raman analysis [530]. One can expect further developments in the field of NIR multichannel Raman spectroscopy with the advent of 2D array detectors offering extended response in the NIR. With these 2D sensors it wiU become possible to apply in the NIR region the powerful techniques already developed in the visible, such as confocal line imaging techniques or multisite remote analysis with optical fibres. 

---