## 2 Vibrational Spectroscopy  
Vibrational spectroscopy analyses vibrations within a molecule and the spectrum of vibrational energies or frequencies (in wavenumbers, cm-1) can be used to characterise a molecular structure. Every molecule has a unique spectrum which allows identification of the molecular compound and its abundance in a sample [2]. Figure 1: Definition of sensitivity and specificity. The exact energy required to excite a molecular vibration depends on the masses of the atoms involved in the vibration and the type of chemical bonds between these atoms and may be influenced by molecular structure, molecular interactions and the chemical microenvironment of the molecule. Vibrational energies fall within the mid Infrared (IR) region of the electromagnetic spectrum and are commonly probed using IR absorption spectroscopy. IR spectroscopy is based on the absorption of infrared radiation by the sample and the fact that molecules absorb specific frequencies of the incident light which are characteristic of their structure. A typical cellular FTIR spectrum is shown in Figure 2, together with a list of tentative spectral assignments in Table 1 [3]. The spectrum shown in Figure 2 can be divided into two regions, the high wavenumber region from 2500-4000 cm\({}^{-1}\), corresponding to absorptions due to O-H, N-H, C-H stretching vibrations, and the fingerprint region from 900-1800 cm\({}^{-1}\), which consists of many overlapping bands (see Table 1), and is characteristic of the sample. \begin{table}
\begin{tabular}{c l} \hline
**Wavenumber (cm\({}^{-1}\))** & **FTIR Peak Assignments** \\ \hline
1036 & C-C skeletal stretching (proteins) \\
1080 & PO\({}_{2}\) symmetric stretching (DNA/RNA) \\
1150 & C-C and C-O stretching (proteins) \\
1220–1280 & PO\({}_{2}\) asymmetric stretching (DNA/RNA), Amide III (C-N stretching, N-H bending, proteins) \\
1310 & CH\({}_{2}\) stretching (phospholipids) \\
1450–1470 & CH\({}_{3}\) asymmetric stretching (proteins, lipids) \\
1520–1550 & Amide II (N-H bending, C-N stretching, proteins) \\
1620–1700 & Amide I (C=O stretching, C-N stretching and N-H bending, proteins) \\
2850 & CH\({}_{2}\) symmetric stretching (lipids) \\
2920 & CH\({}_{2}\) asymmetric stretching (lipids) \\
2960 & CH\({}_{3}\) asymmetric stretching (lipids and proteins) \\
3290 & NH asymmetric stretching (proteins) \\
3400 & OH stretching (water) \\ \hline \end{tabular}
\end{table} 
Table 1: Tentative peak assignments [3] for typical FTIR cellular spectrum shown in Figure 2.  
Figure 2: Typical FTIR spectrum from a population of cells. Raman spectroscopy is an optical method based on inelastic light scattering. The sample is illuminated by monochromatic laser light and interactions between the incident photons and molecules in the sample result in scattering of the light. The coupling of the light generates vibrations within the material, which are again characteristic of the chemical structure, and the energy of the scattered light is reduced by an amount equal to the energy of the vibrational energy. Thus, the positions, relative intensities and shapes of the bands in a Raman spectrum carry detailed information about the molecular composition of the sample. A typical cellular Raman spectrum is shown in Figure 3, together with a list of tentative spectral assignments in Table 2[4]. As for Figure 2, the spectrum can be divided into two regions, the high wavenumber region, from 2500-3500 cm\({}^{-1}\), and the fingerprint region, from 400-1800 cm\({}^{-1}\).