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Vibrational Spectroscopy: From Theory to Applications - ebook

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Vibrational Spectroscopy: From Theory to Applications - ebook

Compendium of IR and Raman spectroscopy!
Vibrational spectroscopy is one of the fundamental tool widely employed in the physico-chemical, material, natural, medical and pharmacological sciences.
The book describes basic techniques of Fourier-Transform Infrared (FTIR) and Raman scattering spectroscopies. The textbook is complemented by an extensive set of experiments which can be conducted by broadly available as well as advanced instrumentation.
After discussion of present theoretical approaches, instrumentation, data handling that explain the phenomenon of vibrational spectroscopy, the book reports new and exciting experiments and applications of the many fascinating spectroscopic effects. They can serve as a basis of several laboratory courses in the field of optical spectroscopy, physical chemistry as well as in specialised panels of medical chemistry, materials science and analytics in the broadest sense. Each exercise is preceded by a description of fundamentals required to understand a research problem. Lab practicals illustrate the application of the given vibrational spectroscopy technique as well as provide a detailed method to solve research problems, which can be introduced to a research, industrial and quality control laboratories.
The authors of the book are specialists in the field of infrared and Raman spectroscopy, for years introducing new courses for a wide panel of the educational offer and conducting research using vibrational spectroscopy.
The manual is designed for students of chemistry, environmental protection, biophysics, medical and life sciences universities. Undoubtedly it will be also helpful in analytical laboratories.

Kategoria: Chemia
Zabezpieczenie: Watermark
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ISBN: 978-83-01-18893-1
Rozmiar pliku: 9,0 MB

FRAGMENT KSIĄŻKI

Vibrational spectroscopy. From theory to practice
Editor Kamilla Malek

AUTHORS

Barańska Małgorzata, Prof., PhD, DSc
Faculty of Chemistry, Jagiellonian University in Krakow

Bukowska Jolanta, Prof., PhD, DSc
Faculty of Chemistry, University of Warsaw

Chmura-Skirlińska Antonina, PhD
Jagiellonian Centre for Experimental Therapeutics (JCET), Jagiellonian University in Krakow

Chruszcz-Lipska Katarzyna, PhD
AGH University of Science and Technology, Faculty of Drilling, Oil and Gas

Czamara Krzysztof, MSc
Faculty of Chemistry, Jagiellonian University in Krakow

Dybaś Jakub, MSc
Faculty of Chemistry, Jagiellonian University in Krakow

Gąsior-Głogowska Marlena, PhD, Eng.
Faculty of Fundamental Problems of Technology, Wroclaw University of Science and Technology

Jaworska Aleksandra, PhD
Faculty of Chemistry, Jagiellonian University in Krakow

Kaczor Agnieszka, PhD, DSc
Faculty of Chemistry, Jagiellonian University in Krakow

Kochan Kamila, MSc
Faculty of Chemistry, Jagiellonian University in Krakow

Królikowska Agata, PhD
Faculty of Chemistry, University of Warsaw

Lipiński Piotr, F.J., PhD
Mossakowski Medical Research Centre, Polish Academy of Sciences

Majzner Katarzyna, PhD
Faculty of Chemistry, Jagiellonian University in Krakow

Malek Kamilla, PhD, DSc
Faculty of Chemistry, Jagiellonian University in Krakow

Marzec Katarzyna M., PhD
Jagiellonian Centre for Experimental Therapeutics (JCET), Jagiellonian University in Krakow

Miśkowiec Paweł, PhD
Faculty of Chemistry, Jagiellonian University in Krakow

Oleszko Adam, MSc, Eng.
Faculty of Fundamental Problems of Technology, Wroclaw University of Science and Technology

Pacia Marta Z., MSc
Faculty of Chemistry, Jagiellonian University in Krakow

Rode Joanna, PhD
1. Institute of Nuclear Chemistry and Technology 2. Institute of Organic Chemistry, Polish Academy of Sciences

Ryguła Anna, PhD
Jagiellonian Centre for Experimental Therapeutics (JCET), Faculty of Chemistry, Jagiellonian University in Krakow

Staniszewska-Ślęzak Emilia, MSc
Faculty of Chemistry, Jagiellonian University in Krakow

Wiercigroch Ewelina, MSc
Faculty of Chemistry, Jagiellonian University in Krakow

Wróbel Tomasz, PhD
Faculty of Chemistry, Jagiellonian University in Krakow

Zając Grzegorz, MSc
Faculty of Chemistry, Jagiellonian University in Krakow4

Special Raman techniques

4.1. Resonance Raman scattering spectroscopy

Katarzyna M. Marzec, Jakub Dybaś

4.1.1. Resonance versus normal Raman scattering and fluorescence

Similar to normal Raman scattering (NR) Resonance Raman Scattering (RRS) can also be described as the two single-photon process. However, the difference appears in the energy of the excitation line. In the case of NR, such energy allows for the transition of the photon to the virtual state, which is far below the first electronic state. When the energy of virtual state corresponds to the energy of electronic excited state of a specific chromophoric group(s) in a molecule, the resonant enhancement is observed (Fig. 4.1.1.). The second part of the process is the same as in the case of NR: emission of the photon with the same (Rayleigh), lower (Stokes) or higher energy (anti-Stokes). The presence of the additional electronic transition in the case of RRS causes the strong enhancement (by a factor of 10³-10⁶) of specific bands originating from the chromophore group in Raman spectrum .

Fig. 4.1.1. Comparison of a simple diatomic energy levels for the normal Raman, resonance Raman and fluorescence spectra

The pre-resonance effect, which corresponds to the situation when the exciting line is close enough to the electronic excited state and also leads to bands enhancement, could also be observed.

The difference between RRS and fluorescence can be seen on the level of excited electronic state. The lifetime of this excited state for RRS is around 10⁻¹⁴ s, while for fluorescence, it may vary between 10⁻⁸-10⁻⁵ s. Moreover, the fluorescence spectrum is observed when the excited state molecule decays via non-radiative transitions (vibrational relaxation) from the discrete vibrational level of the excited electronic state to the lowest vibrational level of the excited state (which is not observed in RRS). Subsequently, this process is followed by the emission of radiation. Weak NR signals may be overwhelmed by fluorescence signals, as fluorescence is characterized by a longer excited state lifetime. This situation is observed not only for fluorescent molecules excited with specific wavelengths (in the range of visible light), but also for many complex samples where the signal is coming from the components’ matrix . As an example we can present the autofluorescence of elastic lamina fibers when radiated with a laser wavelength of 532nm, even though the main components of this aorta structure (elastin and collagen) are not fluorescent molecules at this wavelength .

To eliminate the fluorescence interference in Raman spectra, different procedures may be carried out, starting with the use of different laser wavelengths as an excitation source. To obtain the Raman spectrum of some fluorescent proteins, a laser may be used to irradiate samples for some time before Raman measurement in order to cause the photon-induced destruction of the chromophore. Such a phenomenon is known as photobleaching and was previously used to obtain Raman spectra of proteins or biological samples. Fluorescence effects may also be reduced with the use of confocal Raman systems. In such laser scanning confocal instruments samples are penetrated only in a specific plane (the signal is not collected from the whole volume of the sample), reducing the fluorescent signal from potential contaminations. Secondly, in such conditions, the sample is excited to a high enough point to reach fluorophore saturation (molecules are in the excited state). As a consequence, an increase in the excitation wavelength produces an increase of Raman signal and a reduction in fluorescence emission.

4.1.2. Phenomenon of Resonance Raman scattering

As previously described in Chapter 2, the NR transition moment must have values different from zero (formula 2.3), which is determined by the change of polarizability during the transition between vibrational states, from initial m to final n (see Fig. 4.1.1.). Moreover, it was also postulated that the intensity of a normal Raman band is given by the equation 2.4, where ν_(mn)= ν_(osc).

, (4.1)

In the case of RRS, α_(mn) will represent the change of polarizability α during the transition between the m→e→n states, where e represents the electronic excited state (see Fig. 4.1.1.). That is why, the polarizability tensor α_(mn) in RRS depends on the frequencies (ν_(me) and ν_(en)), as well as on the electric transition dipole moments (M_(me) and M_(en)) which correspond to the energy differences between m→e→n states.

In NR, the sample is irradiated with an exciting line which energy is much smaller than that of electronic transition, so ν₀<<ν_(me). Contrary to NR, in RRS, ν₀ approaches ν_(me), which also has an impact on the increase of the α_(mn) value, and consequently on the significant increase of the intensity (I_(mn)) of the Raman band at ν₀ – ν_(mn.) The intensity of resonance Raman scattering can be expected to be orders of magnitude greater than normal Raman scattering when ν₀ approaches ν_(me).

This shows that compared to non-resonant NR, even components at low concentrations may be detected and analysed with the use of the proper excitation wavelength, which proves the high sensitivity of this technique. Using RRS, it is possible to analyse samples even with nanomolar concentrations . This also explains that to properly understand the observed RRS profile of a sample it is useful to know the UV-Vis absorption spectrum of the sample. To obtain RRS, a given sample is irradiated with an exciting line which coincides with the wavelength of an electronic transition of the sample. That is why the UV-Vis profile allows us to choose an exciting line which corresponds to the electronic transition of specific sample chromophore.

Fig. 4.1.2. The model of the UV-Vis absorption spectrum of molecule X containing two chromophoric groups A and B with two major absorption bands with the maxima at λ_(A) and λ_(B). Exciting the Raman spectra at λ_(A) nm and λ_(B) nm results in two different resonantly enhanced Raman spectra of chromophore A and B, respectively. All parts of the molecule contribute to the non-resonant Raman spectrum excited at λ_(C) nm

Let’s take the theoretical molecule X, containing two chromophoric groups A and B, which has an absorption spectrum with two maxima at wavelengths λ_(A) and λ_(B). The theoretical model of UV-Vis spectrum of molecule X is presented in Fig. 4.1.2. To selectively enhance the vibrations of the chromophore A in a complex spectrum of the molecule, the exciting line has to have ν₀ near to ν_(A )(which corresponds properly to λ_(A)) _(.) Vibrations of chromophore B will be enhanced when the laser wavelength will be equal to λ_(B). On the other hand, if the exciting line λ_(C) is used, the non-resonant Raman spectrum which comes from all parts of molecule will be observed.

For this reason, the UV–Vis electronic absorption spectrum of the studied compound, which shows the allowed electronic transitions will help us to choose the best laser wavelength in order to observe the vibrations of specific chromophore. By changing the excitation wavelength, the different RRS of the same molecule may be obtained and give us information about specific parts of the molecule. Such selective enhancement suggests a high specificity of the RRS technique.

The detailed information about UV-Vis spectrum may provide additional information about the RRS origin. The quantitative description of RRS scattering theory was provided by Albrecht et al., who showed how RRS intensity can arise from several mechanisms, mainly from A-term (Franck–Condon) and B-term (Herzberg– –Teller vibronic coupling) .

In type A or Franck–Condon scattering, only totally symmetric modes are enhanced in RRS. Such mechanism is observed for many compounds. As an example we can include here the RR spectra of TiI₄ and NH₃ obtained by 514.5 and 216.8 nm excitations, respectively . Non–Condon dependence of the electronic transition moment upon the vibrational coordinate is possible in B–term enhancement, where both symmetric and non-symmetric fundamentals can be enhanced. However, the magnitude of B-term enhancement of symmetric vibrations is lower than that of A-term enhancement. B-term enhancement will dominate only for non-symmetric vibrations . Such RR scattering involves vibronic coupling between the two allowed excited electronic transitions. This mechanism is observed for metalloproteins being excited with the laser wavelength which corresponds to the electronic transitions of the Q band of the UV-Vis spectrum. If the enhancement of fundamentals cannot occur via A or B-terms, as transition is rigorously forbidden at the equilibrium geometry, then C-term enhancement of overtones and combinations modes may occur .

4.1.3. Application and potential of RRS

As we proved above, RRS is characterized by high sensitivity and selectivity in comparison with NR, which gives RRS technique the advantage in many analytical studies. Similar to NR, RRS allows for the study of samples in the gaseous, liquid and solid state.

In art history, archaeology and forensics RRS is successfully used to study the composition of different pigments and dyes. It is also known as a non-invasive and non-destructive method of assessing the distribution and concentration of various biomolecules inside plant and animal tissues. It has been applied to study the carotenoid status in human skin, as a biomarker of fruit/vegetable intake . A single RRS skin measure allowed for the classification of inter-individual variability in skin carotenoid status and to identify factors associated with the biomarker in this population . It is also possible to differentiate various retinoid fractions from a mixture with the use of this technique utilizing different excitation wavelengths. Upon excitations with different wavelengths it was possible to differentiate lutein, violaxanthin, β-carotene and 9-cis neoxanthin . The use of the 532 nm excitation laser line, which allows observation of the pre-resonance Raman spectrum of retinols, was also used to study the distribution of vitamin A component in liver and lung tissues .

Resonance Raman spectroscopy has long been applied to monitor the molecular dynamics of different metalloproteins, among which the most common is hemoglobin . This highly symmetrical and chromophoric heme prosthetic group provides strong resonance enhancement, especially when the excitation wavelength is in resonance with the intense electronic transitions cantered at ~400 nm (Soret), 525 nm (Q_(v) or α band) and 575 nm (Q₀ or β band) . Moreover, peptide chains of heme proteins may also be studied with this technique as they exhibit transitions below 250 nm. RRS was successfully used not only for standard hemoporphirins, but also for the detection, analysis and visualization of 2D and 3D distributions of heme in both cells and tissues . RRS provides excellent signal–to–noise ratio spectra with very high reproducibility from single erythrocytes, which allows for the study of various hemopathies .

As in case of heme proteins, peptide chains of other proteins, or protein–drug interactions, may also be studied with the use of excitation sources below 260 nm. The use of RRS in such deep UV was successfully applied in order to investigate DNA, RNA and nucleic acid components . The use of RRS in such deep UV is mainly used in the bioanalytical and life science fields, however it is also useful to study solid catalysts and heterogeneous catalytic reactions .

Because of the effects of this vibrational technique, information about the electronic structure of a studied sample can be obtained. This makes RRS a very useful technique in nanotechnology and materials science in order to study and characterize structures of such materials as carbon nanotubes, graphite, graphene and others .

4.1.4. Instrumentation

As already presented and described in Chapter 2, for RRS detection the standard Raman instrumentation may be applied. As mentioned before, selective resonance Raman enhancement of specific chromophores of molecules may be obtained by changing the excitation wavelength. That is why tunable lasers, in which the wavelength can be altered within a specific range, are commonly applied to this technique. To provide positive identification, even with higher than RRS sensitivity and selectivity, RRS is successfully used in combination with liquid chromatography and SERS (surface-enhanced resonance Raman scattering, SERRS). Tip-enhanced Raman spectroscopy (TERS), which is a variation of SERS, may also use the resonance effect (TERRS) and is a promising technique for future nanoanalysis .

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