## 3 Results and Discussion  
It is worth noting that the SERS substrate obtained by AgNPs deposition resulted in a smooth surface without defects. Moreover, the scanning electron microscope micrograph (Figure 1b) showed a uniform surface of small nanoparticles (\(\sim\)150 nm) with homogeneous  
Figure 2: Diagram of the home-made cooling system. size distribution as result of the \"silver mirror\" reaction. The precision of the mean particle size, expressed as %RSD, for three independent substrates was 14.2%. First, the weight of meat was optimized to obtain a significant fraction of the extracted lipids, since the amount of fat deposited on the SERS substrate was crucial. On the one hand, a drop of lipid extract was completely evaporated after laser heating. On the other hand, when a thick layer (8 mm) was applied, no SERS effect was observed at all. Finally, a layer of about 3 mm of fat was used, which corresponds to about 12 g of meat. The next parameter to be optimized was the measurement temperature. No significant differences were found when comparing fresh and oxidized samples at room temperature. Taking into account the results obtained from our previous work [35], when using the cooling device at \(-2\)\({}^{\circ}\)C, better definition of the relevant bands and notorious differences in intensity appeared in the oxidized sample, and so, further analyses were performed at low temperature. Although the effect of temperature on SERS was poorly explored, our findings were consistent with a work [37] suggesting that the dielectric constant of noble metal nanoparticles is influenced by electron-phonon scattering and electron-electron scattering, with EF being favored at low temperatures. In addition, laser power (ranging from 3.5 to 9.5 mW), exposure time, and number of exposures were adjusted by sequential optimization. The selected conditions were as follows: 5 mW of laser power, 30 s of exposure time, and 20 exposures. Finally, the results of area and height of the spectral bands were collected. Since significant differences were obtained only in the case of areas, this parameter was used in subsequent analyses. An analysis of the photographs of meat samples showed that active packaging containing oregano extract prolonged the shelf life of meat. Oregano extract is a well-known antioxidant that was widely studied for active packaging purposes [10; 38; 39; 40; 41; 42; 43]. Therefore, it was chosen as antioxidant agent to inhibit the degradation of lipids in meat. Figure 3 shows the aspect of meat samples stored under refrigeration after 5 days. LDPE-packaged samples started to lose color due to myoglobin oxidation. This process was shown to be associated with lipid oxidation [4]. Figure 3: Aspect of meat samples stored at 5 \({}^{\circ}\)C after 5 days, where: **(a)** samples from conventional LDPE packaging, **(b)** samples from active packaging. In this work, the Raman spectrum of fat extracted from meat stored in active packaging was collected and compared with the spectrum of fat extracted from meat stored in conventional LDPE packaging. Figure 4 shows the spectrum of the extracted lipids at 0 days, highlighting the major Raman shifts associated with fat oxidation in the SERS spectrum. As can be seen, the SERS spectrum showed a much improved signal-to-noise ratio compared to conventional Raman. Raman shifts corresponding to the oxidation of lipids present in oils and fats were extensively described in the scientific literature [44; 45; 46; 47; 48]. The most relevant detected shifts related to lipid degradation are briefly described in Table 1.  
Three replicates of each sample were measured and three measurements of each replicate were performed in the case of SERS. The RSD% values for the measurements of lipids extracted from ground pork in LDPE and in active packaging, were less than 29% for LDPE and 15% for active packaging. The SERS bands showed strong enhancement with respect to the ordinary Raman spectrum.