## 4 Vibrational Spectroscopic Techniques for Non-Invasive Food Authentication
### Raman Spectroscopy for Non-Invasive Food Authentication  
Raman spectroscopy is a highly effective analytical tool for identifying sample matrices, especially for analyzing key compounds like lipids, proteins, and carbohydrates. It is particularly sensitive to trace components, such as microorganisms, that contribute to food spoilage and contamination [99]. Though both IR and Raman spectroscopy are forms of vibrational spectroscopy, they operate based on different molecular interaction mechanisms. While IR absorption spectroscopy measures the absorption of light, Raman spectroscopy relies on the exchange of photon energy between the light and the molecule. The Raman effect occurs when light interacts with the electron cloud of a molecule's bonds, causing the molecule to undergo a transition to a virtual state. As the molecule returns to a lower vibrational energy state, Raman scattering is produced. If the molecule was already in a higher vibrational state, the scattering is referred to as anti-Stokes scattering. In spontaneous Raman spectroscopy, the majority of the incident photons, approximately 99.999%, undergo elastic Rayleigh scattering, which does not provide useful information for molecular analysis because it does not change the molecule's quantum state. Rayleigh scattering is an elastic interaction where light interacts with matter but leaves the material unchanged. Only a very small fraction--around 0.001%--of the incident light produces inelastic Raman scattering, which is the useful signal for molecular characterization. However, since spontaneous Raman scattering is relatively weak, it can be difficult to isolate the Raman signal from the much stronger Rayleigh scattering. To overcome this challenge, various optical components, such as notch filters, laser stop apertures, and tunable filters, are used to suppress Rayleigh scattering and enhance the quality of the Raman spectra. Furthermore, different illumination and sampling techniques are employed to boost the Raman signal. Methods such as SRS, CARS, RRS, and SERS all provide advantages in terms of sensitivity and signal amplification (Table 1), further improving the capabilities of Raman spectroscopy for detailed analysis. Recent advancements in instrumentation have positioned Raman spectroscopy as an increasingly valuable tool in the food industry. One of the notable techniques, CARS spectroscopy, enables label-free chemical imaging with high sensitivity and excellent spatial resolution. However, a limitation of CARS is the presence of a nonresonant electronic background, which can obscure the chemical signals of interest [100]. In contrast, SRS spectroscopy provides enhanced sensitivity and avoids the issue of nonresonant interference, making it particularly advantageous for precise molecular analysis [101]. Both SRS and CARS are commonly applied in the agro-food industry, particularly for the microscopic examination of various products, thereby allowing for the detailed analysis of food components at the molecular level [102]. RRS represents a distinct Raman technique in which the excitation light is carefully aligned with an electronic absorption band of the target molecule. This resonance condition enhances the vibrational signal by promoting electronic excitation, thereby intensifying both Stokes and anti-Stokes scattering features in the resulting spectrum [103]. However, RRS typically requires visible or ultraviolet excitation, which \begin{table}
\begin{tabular}{c c c c} \hline
**Technique** & **Enhancement Factor** & **Food Application** & **Key Limitation** \\ \hline SERS & \(10^{5}\)\(-\)\(10^{6}\) & Pesticides in fruits & Substrate reproducibility \\ CARS & \(10^{3}\)\(-\)\(10^{4}\) & Lipid oxidation in meat & Nonresonant background \\ Resonance Raman & \(10^{2}\)\(-\)\(10^{3}\) & Carotenoids in seafood & UV-induced fluorescence \\ \hline \end{tabular}
\end{table} 
Table 1: Comparison of Raman techniques for food authentication. can lead to interference from fluorescence, complicating the analysis.