Attenuated total reflection-Fourier transform infrared imaging of large areas using inverted prism crystals and combining imaging and mapping

Attenuated total reflection-Fourier transform infrared (ATR-FT-IR) imaging is a very useful tool for capturing chemical images of various materials due to the simple sample preparation and the ability to measure wet samples or samples in an aqueous environment. However, the size of the array detector used for image acquisition is often limited and there is usually a trade off between spatial resolution and the field of view (FOV). The combination of mapping and imaging can be used to acquire images with a larger FOV without sacrificing spatial resolution. Previous attempts have demonstrated this using an infrared microscope and a Germanium hemispherical ATR crystal to achieve images of up to 2.5 mm x 2.5 mm but with varying spatial resolution and depth of penetration across the imaged area. In this paper, we demonstrate a combination of mapping and imaging with a different approach using an external optics housing for large ATR accessories and inverted ATR prisms to achieve ATR-FT-IR images with a large FOV and reasonable spatial resolution. The results have shown that a FOV of 10 mm x 14 mm can be obtained with a spatial resolution of approximately 40-60 microm when using an accessory that gives no magnification. A FOV of 1.3 mm x 1.3 mm can be obtained with spatial resolution of approximately 15-20 microm when using a diamond ATR imaging accessory with 4x magnification. No significant change in image quality such as spatial resolution or depth of penetration has been observed across the whole FOV with this method and the measurement time was approximately 15 minutes for an image consisting of 16 image tiles.     

Applications of attenuated total reflection infrared spectroscopic imaging to pharmaceutical formulations

This paper demonstrates an approach to obtain chemical images of pharmaceutical tablets using attenuated total reflection infrared (ATR-IR) spectroscopy. FT-IR images with different fields of view and spatial resolution have been obtained using a combination of different ATR accessories. FT-IR imaging with the diamond ATR accessory and micro-ATR imaging technique have been compared. With the diamond ATR imaging accessory, compaction to a tablet can be performed and the chemical image measured in situ. It has been found that the diamond ATR imaging accessory gives information on the overall distribution of different components in a tablet while the micro-ATR imaging technique provides a closer look at the tablet with 4-microm spatial resolution. Low-concentration components down to 0.5% have been detected by the micro-ATR method. Both experimental and commercial systems are studied in this paper.     

Fourier transform infrared imaging of human hair with a high spatial resolution without the use of a synchrotron

The cross-section of a human hair has been imaged for the first time using the micro attenuated total reflection (ATR) Fourier transformed infrared (FT-IR) method in combination with a focal plane array (FPA) detector. A rigorous approach was applied to determine the spatial resolution, namely, measuring the distance over which the band absorbance changes from 95 to 5% of the maximum absorbance when passing through a sharp interface. The measured value for IR transmission was approximately 16 microm, while the value obtained using ATR imaging was approximately 5 microm. The enhanced spatial resolution achieved by this method allows the medulla of the hair (approximately 8 microm in diameter) to be imaged clearly without the need for a synchrotron source. The spatial resolution of transmission and ATR imaging is compared, and advantages of ATR imaging are discussed.     

Chemical imaging with variable angles of incidence using a diamond attenuated total reflection accessory

A new development in Fourier transform infrared (FT-IR) imaging using a diamond attenuated total reflection (ATR) imaging accessory in a novel manner that allows the angle of incidence to be varied in order to obtain images from subsurface layers of different thickness is introduced. Chemical images of samples from the same area but with different depths of penetration are obtained by changing the angle of incidence as well as using different spectral bands at different wavenumbers. Changes in the angle of incidence with this accessory were made possible by taking advantage of the relatively large numerical aperture employed by the original imaging optics. This arrangement allowed us to introduce an additional movable aperture in the optical design to restrict the angle of incidence to certain values. Two samples have been studied, one for the calibration of the angle of incidence while the other demonstrates the capability of obtaining three-dimensional (3D) information using this approach. Advantages of this new approach include the relatively high spatial resolution (it can spatially resolve features as small as 12 mum without a microscope) and no change in the imaging area and sampling area during manipulation of the angle of incidence.     

New opportunities in micro- and macro-attenuated total reflection infrared spectroscopic imaging: spatial resolution and sampling versatility

New opportunities exist to obtain chemical images using attenuated total reflection infrared (ATR-IR) spectroscopy. This paper shows the feasibility of obtaining FT-IR images with a spatial resolution of at least 3-4 microm using a Ge ATR objective coupled with an infrared microscope. The improved spatial resolution compared to FT-IR images obtained by the transmission method is due to the high refractive index of the ATR crystal, which gives a high numerical aperture and hence, a higher spatial resolution. FT-IR imaging with a conventional diamond ATR accessory has been investigated. This is the first time that FT-IR imaging is reported using such a versatile accessory based on a diamond ATR crystal. These results showed that a spatial resolution up to 13 microm can be achieved without the use of infrared microscope objectives. One advantage of the diamond element is that it allows pressure to be applied and hence, good contact to be obtained over the whole field of view.     

Analysis of Gibbs monolayer adsorbed at the toluene/water interface by UV-visible partial internal reflection spectrometry

Gibbs monolayers of lipophilic tetraphenylporphyrinatomanganese(III) and hydrophilic diacid of meso-tetrakis(4-sulfonatopheny)porphyrin adsorbed at the liquid-liquid interface have been analyzed by UV-visible external reflection (ER) and partial internal reflection (PIR) spectra measured at different angles of incidence. The angle-dependent ER and PIR spectra over the Brewster angles (thetaERB and thetaIRB) have readily been measured at the toluene/water interface. As preliminarily expected in our previous study, the present study has first proved that the reflection-absorbance of UV-visible PIR spectra quantitatively agrees with the theoretical calculations for the Gibbs monolayer over thetaIRB. In addition, it has also been proved that the absorbance of the PIR spectra is greatly enhanced in comparison to that of the ATR spectra. The enhancement is caused by an optical effect in the monolayer sandwiched between two phases of toluene and water that have different but refractive indices close to each other. This optical enhancement requires an optically perfect contact between the phases, which is difficult to prepare for a solid-solid contact. At the liquid/liquid interface, however, an ideal optical contact is easily realized, which makes the enhancement as much as the theoretical expectation. The PIR spectrometry will be recognized to be a new high-sensitive analytical tool to study Gibbs monolayer at the liquid/liquid interface.     

High-resolution ab initio three-dimensional x-ray diffraction microscopy

Coherent x-ray diffraction microscopy is a method of imaging nonperiodic isolated objects at resolutions limited, in principle, by only the wavelength and largest scattering angles recorded. We demonstrate x-ray diffraction imaging with high resolution in all three dimensions, as determined by a quantitative analysis of the reconstructed volume images. These images are retrieved from the three-dimensional diffraction data using no a priori knowledge about the shape or composition of the object, which has never before been demonstrated on a nonperiodic object. We also construct two-dimensional images of thick objects with greatly increased depth of focus (without loss of transverse spatial resolution). These methods can be used to image biological and materials science samples at high resolution with x-ray undulator radiation and establishes the techniques to be used in atomic-resolution ultrafast imaging at x-ray free-electron laser sources.     

Attenuated total internal reflection infrared microscopy of multilayer plastic packaging foils

Multilayer plastic foils are important packaging materials that are used to extend the shelf life of food products and drinks. Fourier transform infrared (FT-IR) spectroscopic imaging using attenuated total internal reflection (ATR) can be used for the identification and localization of different layers in multilayer foils. A new type of ATR crystal was used in combination with a linear array detector through which large sample areas (400 x 400 microm(2)) could be imaged with a pixel size of 1.6 microm. The method was tested on laminated plastic packing materials containing 5 to 12 layers. The results of the identification of the different materials using ATR-FT-IR were compared with differential scanning calorimetry (DSC) and the layer thickness of the individual layers measured by ATR-FT-IR was compared with polarized light microscopy (LM) and scanning electron microscopy (SEM). It has been demonstrated that individual layers with a thickness of about 3 microm could be identified in multilayer foils with a total thickness ranging from 100 to 150 microm. The results show a spatial resolution of about 4 microm (measured at wavenumbers ranging from 1000 to 1730 cm(-1)), which is about a factor of two better than can be obtained using transmission FT-IR imaging. An additional advantage of ATR is the ease of sample preparation. A good correspondence was found between visible and FT-IR images. The results of ATR-FT-IR imaging were in agreement with those obtained by LM, SEM, and DSC. ATR-FT-IR is superior to the combination of these techniques because it delivers both spatial and chemical information.     

Characterization of pollen by vibrational spectroscopy

Classification, discrimination, and biochemical assignment of vibrational spectra of pollen samples belonging to 43 different species of the order Pinales has been made using three different vibrational techniques. The comparative study of transmission (KBr pellet) and attenuated total reflection (ATR) Fourier transform infrared (FT-IR) and FT-Raman spectroscopies was based on substantial variability of pollen grain size, shape, and relative biochemical composition. Depending on the penetration depth of the probe light, vibrational techniques acquire predominant information either on pollen grain walls (FT-Raman and ATR-FT-IR) or intracellular material (transmission FT-IR). Compared with the other two methods, transmission FT-IR obtains more comprehensive information and as a result achieves superior spectral identification and discrimination of pollen. The results strongly indicate that biochemical similarities of pollen grains belonging to the same plant genus or family lead to similar features in corresponding vibrational spectra. The exploitation of that property in aerobiological monitoring was demonstrated by simple and rapid pollen identification based on relatively small spectral libraries, with the same (or better) taxonomic resolution as that provided by optical microscopy. Therefore, the clear correlation between vibrational spectra and pollen grain morphology, biochemistry, and taxonomy is obtained, while successful pollen identification illustrates the practicability of such an approach in environmental studies.     

Spectroscopic investigations on the depth-dependent degradation gradients of aged triterpenoid varnishes

The depth-profiles of accelerated aged triterpenoid dammar and mastic varnishes, which had been uncovered by optimized KrF excimer laser ablation (248 nm, 25 ns), were examined by ultraviolet-visible (UV/VIS) spectrophotometry and attenuated total reflection (ATR) Fourier transform infrared (FT-IR) spectroscopy. The results reflect both the optical properties at the bulk (UV/VIS) and the surface (ATR/FT-IR) and establish the presence of degradation gradients across the depth of the degraded natural resin films. It is demonstrated that the UV part of the ambient radiation is absorbed by the deteriorated surface and that the optical densities of the aged varnish decrease as a function of depth. The absorbed UV wavelengths at the surface and the depth-wise reduction of light intensity result in a depth-dependent decrease in carbonyl absorbance and an increase in C-H bending vibration modes of methylene species pointing to structural modification in the triterpenoid mixture across depth.     

Novel attenuated total reflection Fourier transform infrared microscopy using a gem quality diamond as an internal reflection element

A novel technique for attenuated total reflection Fourier transform infrared (ATR FT-IR) spectral acquisition by an infrared microscope with a gem-quality faceted diamond as an internal reflection element (IRE) is introduced. Unlike conventional IREs, the novel diamond IRE has a sharp tip configuration instead of a flat tip configuration. Light at normal incidence was coupled into the diamond while the transflected radiation from the diamond was collected through the table facet by the built-in 15x Cassegrainian objective. The number of reflections in the novel diamond IRE equals two. The evanescent field generated under total internal reflection at the pavilion facet was exploited for ATR spectral acquisition of materials attached to the IRE. The observed ATR spectra were compared to those obtained via a traditional zinc selenide IRE.     

Double-pass oblique-incidence interferometer for the inspection of nonoptical surfaces

Interferometric testing of the flatness or straightness of ground surfaces of metals, granite, ceramics, and glasses has always proved difficult because of the extremely low specular reflection from these optically rough surfaces. At oblique angles of incidence the specular reflection coefficient of nonoptical surfaces increases rapidly, typically from near 0.01 at 75 degrees up to, say, 0.50 at 85 degrees and near 1.00 at 90 degrees angle of incidence. By taking advantage of this enhanced reflection at oblique incidence and the brightness and spatial coherence of visible gas lasers, it becomes practical to use interferometric inspection of nonoptical surfaces. A novel double-pass oblique-incidence interferometer is described which enables testing of large pieces having a specular reflection as low as 1%.     

Cr /sc multilayers for the soft-x-ray range

We have systematically investigated ultrathin Cr/Sc multilayers (nanolayers), using tunable soft-x-ray synchrotron radiation. The multilayers were optimized for use either in normal incidence or at 45 degrees at photon energies around the 2p-absorption edges of Sc (399 eV) and Cr (574 eV), respectively. They were sputter deposited on Si wafers or on thin Si(3)N(4)-membrane support structures for use in reflection and in transmission, respectively, as polarizing and phase-retarding elements in a polarimeter. The performance theoretically expected with respect to reflection/transmission and energy resolution has been confirmed experimentally: A value of 7% for the normal-incidence peak reflectance at 395 eV was measured as well as a pronounced minimum in transmission for certain incidence angles and energies below the respective absorption edges, indicating significant phase-shifting effects.     

Attenuated total internal reflection infrared microspectroscopic imaging using a large-radius germanium internal reflection element and a linear array detector

The number of techniques and instruments available for Fourier transform infrared (FT-IR) microspectroscopic imaging has grown significantly over the past few years. Attenuated total internal reflectance (ATR) FT-IR microspectroscopy reduces sample preparation time and has simplified the analysis of many difficult samples. FT-IR imaging has become a powerful analytical tool using either a focal plane array or a linear array detector, especially when coupled with a chemometric analysis package. The field of view of the ATR-IR microspectroscopic imaging area can be greatly increased from 300 x 300 microm to 2500 x 2500 microm using a larger internal reflection element of 12.5 mm radius instead of the typical 1.5 mm radius. This gives an area increase of 70x before aberrant effects become too great. Parameters evaluated include the change in penetration depth as a function of beam displacement, measurements of the active area, magnification factor, and change in spatial resolution over the imaging area. Drawbacks such as large file size will also be discussed. This technique has been successfully applied to the FT-IR imaging of polydimethylsiloxane foam cross-sections, latent human fingerprints, and a model inorganic mixture, which demonstrates the usefulness of the method for pharmaceuticals.     

Thickness measurement of nanoscale polymer layer on polymer substrates by attenuated total reflection infrared spectroscopy

For the first time, attenuated total reflection (ATR)-Fourier transform infrared (FT-IR) spectroscopy was utilized to measure the thickness (d0) of a nanoscale polymer layer on polymer substrate with significant credibility. First, a mathematical formula, A/A0 = 1 - 2d0/ d(p), was derived based on a self-defining subsection function (where d(p) was defined as depth of penetration of ATR and A and A0 were defined as the absorption band area of the characteristic functional group only contained in bulk substrate with a thin polymer layer attachment and the same group in blank substrate, respectively). On the mathematical model, through changing incidence angles, a series of values of A (A0) and corresponding d(p) were obtained, and when plotting A/A0 versus 2/d(p), d0 was obtained as the slope. With polystyrene (coating)/olypropylene (substrate) as a model system, we obtained the relevant values (d0). Comparing the results with the values of practical coating thickness (calculation and TEM observation), we found that this method was able to characterize well the thickness of a thin polymer layer on a polymer substrate in the range from 10 to 110 nm. Errors in the measurement were given and analyzed. Furthermore, this method was well applied in the thickness measurement of a polyacrylamide graft layer on a polypropylene film surface. The effect of pressure in the ATR technique on the coating thickness measurement was also discussed. In comparison with other methods such as XPS, SEM, TEM, and AFM, this approach based on a universal ATR technique was very convenient and fast. This method is expected to widen the application of the ATR-FT-IR technique and stimulate the further development of many fields such as surface self-assembly and surface functionlization.     

Light profile microscopy based on Raman and wavelength resolved luminescence contrast

Light profile microscopy based on contrast from wavelength resolved Raman and luminescence measurements is demonstrated experimentally for the first time. A Raman/multispectral light profile microscope (RMSLPM) has been constructed based on a line profiling geometry in which the sample is irradiated with a tightly focused laser beam (of ten micrometers radius or less) behind a polished view surface and the resulting line image is dispersed over the wavelength using an imaging spectrograph. The instrumentation developed in this laboratory has a spectral resolution approaching 10 cm(-1) and an (actual) depth independent spatial resolution of 6-8 times the Rayleigh diffraction limit, limited at present by optical aberrations and alignment. The technique has the potential to image at approximately twice the Rayleigh diffraction limit. The spectral signatures reconstructed from a variety of common industrial polymers show excellent agreement with reference spectra from the literature, and may be used to identify individual layers in depth images of unknown materials. RMS-LPM image data based on luminescence contrast have also been used to provide concentration depth profiles of additives and degradation products in injection molded samples of high-density poly(ethylene) (HDPE).     

Improving the performance of the MWPC X-ray imaging detector by means of the multi-step avalanche technique

X-ray imaging systems based on conventional MWPC technology and artificial delay line readout techniques have been developed at RAL for several applications over a period of some eight years. Successful as this effort has been, it is now perceived that very limited scope exists for the further improvement of the imaging capability of the standard MWPC design. Attention has therefore been turned to the possibility of exploiting the multi-step avalanche (MSA) system of electron multiplication in this context. Results from a prototype system are presented which show spatial resolution better than that achieved in our MWPC systems with no wire artefacts in the image. The facility for controlling the effective depth of the detector electronically is also demonstrated so offering some alleviation of the parallax problem common to all gaseous X-ray detectors.     

Micro-attenuated total reflection spectral imaging in archaeology: application to Maya paint and plaster wall decorations

Fourier transform infrared (FT-IR) attenuated total reflection (ATR) imaging has been successfully used to identify individual mineral components of ancient Maya paint. The high spatial resolution of a micro FT-IR-ATR system in combination with a focal plane array detector has allowed individual particles in the paint to be resolved and identified from their spectra. This system has been used in combination with micro-Raman spectroscopy to characterize the paint, which was found to be a mixture of hematite and silicate particles with minor amounts of calcite, carbon, and magnetite particles in a sub-micrometer hematite and calcite matrix. The underlying stucco was also investigated and found to be a combination of calcite with fine carbon particles, making a dark sub-ground for the paint.     

Analysis of counterfeit pharmaceutical tablet cores utilizing macroscopic infrared spectroscopy and infrared spectroscopic imaging

Advantages and limitations of analyzing authentic and counterfeit pharmaceutical tablets with both macro (nonimaging) attenuated total internal reflection Fourier transform infrared (ATR-FT-IR) spectroscopy and micro ATR-FT-IR spectroscopic imaging have been evaluated. The results of this study demonstrated that micro ATR imaging was more effective for extracting formulation information (sourcing), whereas a macro ATR approach was better suited for counterfeit detection (screening). More importantly, this study demonstrated that a thorough analysis of the counterfeit core can be achieved by combining the results of both techniques.     

Attenuated total reflection infrared spectroscopy: an efficient technique to quantitatively determine the orientation and conformation of proteins in single silk fibers

Polarized attenuated total reflection (ATR) infrared spectroscopy is an efficient technique to determine the orientation and conformation of a large variety of samples, but it is more difficult to apply to very small specimens such as silk fibers. The Golden Gate single-reflection ATR accessory that uses diamond as an ATR element and a focalized beam turns out to be highly efficient to study quantitatively the orientation and conformation of a single silk fibroin filament of the silkworm Bombyx mori that is about 10 mum in diameter. For orientation measurements, rotating the sample instead of the electric field greatly simplifies the theoretical analysis and keeps the penetration depth of the infrared radiation constant. A sample holder that can be fitted on the ATR accessory has thus been developed to allow accurate rotation of the sample and to obtain spectra with a low, non-damaging, and reproducible pressure on the fiber. To validate the method, spectra have been recorded as a function of the angle theta between the fiber axis and the polarization of the incident radiation. The data have been fitted following the cosine square dependency of the absorbance with respect to the angle theta. The procedure has been applied to the spectral components of the amide I bands, as determined from spectral decomposition. Multiple angle measurements turn out to be quite useful to correct systematic angle errors and validate the accuracy of the curve-fitting parameters of the band decomposition. By using the calculated dichroic ratio, a parameter of -0.46 +/- 0.01 has been calculated for the antiparallel beta-sheets and -0.04 +/- 0.02 for the remaining structures. From the orientation-insensitive spectrum A(0), the amount of beta-sheets has been estimated to 49 +/- 3%. The results obtained from only two measurements with the electric field of the incident radiation parallel and perpendicular to the fiber axis has demonstrated that ATR spectroscopy can be used routinely in quantitative studies of the molecular orientation and conformation of macromolecules.