## 3 Results and Discussion  
Figure 2 compares the Raman spectra of riboflavin measured on the gold nanoisland SERS substrate described above, on a pure glass substrate as a reference and in solid form. All spectra were recorded with the same measurement conditions. The spectra were recorded with 633 nm excitation, as in the case of riboflavin, a higher SERS enhancement was obtained with this excitation wavelength. The following characteristic peaks of riboflavin can be detected in the spectra: high-intensity bands at 1174 and 1222 cm\({}^{-1}\) corresponding to the C-N bending vibrations of the uracil ring and the C-CH\({}_{3}\) bond of the benzene ring, respectively. The peak at 1402 cm\({}^{-1}\) can be assigned to C-C vibrations and the peak at 1496 cm\({}^{-1}\) can be assigned to the -CH\({}_{3}\) group. Furthermore, the peak appearing as a shoulder at 1530 cm\({}^{-1}\) and the high-intensity band at 1562 cm\({}^{-1}\) can be linked to the vibration of the C-N bond [24; 31; 33]. Figure 3 shows the Raman spectra of R6G, recorded similarly to those of riboflavin in Figure 2. A comparison of the spectra shows that using gold nanoisland substrate and the SERS process, the main bands of rhodamine 6G appear with a higher intensity than in other samples. The spectra shown were recorded with 532 nm excitation, since in the case of R6G a higher degree of enhancement was achieved with this laser. In the SERS spectrum, the C-C vibration of the aromatic ring appears at 1650, 1599, and 1363 cm\({}^{-1}\). The band at 1312 cm\({}^{-1}\) can be related to the stretching vibration of the C-O-C group of the carbon skeleton, and the peaks at 1189 and 773 cm\({}^{-1}\) are the vibrations of the -CH\({}_{3}\) group. At 614 cm\({}^{-1}\), the vibration of the C-C-C group appears at a high intensity [34]. In our study, the magnitude of the SERS enhancement was calculated based on the enhancement factor (\(EF\)) [35] defined as follows:  
\[EF=\frac{I_{SERS}/N_{SERS}}{I_{NR}/N_{NR}}, \tag{1}\]  
where \(I_{SERS}\) is the intensity of the signal obtained during the SERS measurement; \(I_{NR}\) is the intensity of the signal obtained during the normal Raman (reference) measurement; and \(N_{SERS}\) and \(N_{NR}\) are the average number of molecules absorbed on the surface during the SERS and normal Raman measurements, respectively [35]. For riboflavin, the magnitude of SERS EF was calculated from the intensity of the peak at \(\sim\)1575 cm\({}^{-1}\), corresponding to the vibration of the C-N bond. For R6G, the EF value was calculated from the intensity of the peak at \(\sim\)1650 cm\({}^{-1}\), arising from the vibrations of the C-C bond of the aromatic ring. Figure 4 compares the surface-enhanced Raman spectra measured on a given gold nanoisland sample with two excitation wavelengths. The main peaks of riboflavin can be detected in both spectra in the Figure; however, the degree of the enhancement of the Raman signal is more pronounced while using the 633 nm laser. The EF value for the 532 nm laser was found to be 9, while, for 633 nm excitation, it was 42. Figure 3: Raman spectra of rhodamin 6G measured on gold nanoisland SERS and pure glass substrates, and in solid form. The same comparison for the other analyte, i.e., R6G, is provided in Figure 5. The characteristic peaks of R6G can be detected for both excitations. In terms of the enhancement, it is better with 532 nm excitation; the EF for R6G is 110 for the 532 nm excitation and only 14 for 633 nm. Figure 4: Surfaced-enhanced Raman signal of riboflavin measured with two excitation wavelengths. Figure 5: Surfaced-enhanced Raman signal of rhodamine 6G measured with two excitation wavelengths. In the case of gold nanoisland substrates, a higher enhancement would be expected for 532 nm excitation, since the magnitude of the SERS enhancement is influenced by the plasmon wavelength of the substrate. We know that the LSPR peak of the active surface must fall close to the wavelength of the excitation source [12].