Rhodamine 6g dye

Solutions of several analytes were used to investigate the quality and sensitivity of SERS substrates. First, rhodamine 6 G dye (Sigma-Aldrich, product number 252433) was used. Isopropanol R6G solutions were prepared with concentrations in the 10⁻⁹–10⁻⁵ M range. Next, we used L-tryptophan (Sigma-Aldrich, product number T8941) as a model organic compound with a low molecular weight, whose electronic resonance (280 nm) is far from the plasmonic resonance of Au films, to test the applicability of the developed SERS substrates for detection of biologically relevant molecules. Finally, isopropanol solutions of Bovine Serum Albumin (BSA) (MP Biomedicals, product number 02152401) and keratin hydrolysates were chosen as a test biomacromolecular objects. Isopropanol solution of tryptophan was prepared of 99% Sigma-Aldrich L-tryptophan (Product number T8941). Keratin water solution was prepared from a human nail as it was described in^{18 (link)}. To apply the solutions to the substrate, the 50 μl Discovery micropipette was used.10 μl of solution was applied to all substrates. Surface area of substrates was roughly 0.25 cm². Prepared substrates are sensitive to the choice of solvent, see Fig. S1.

Analytical grade NaOH and H₂O₂ (30%) were obtained from Merck (Merck, Germany) and zinc salts (ZnSO₄·7H₂O, Zn(OAc)₂·2H₂O, and Zn(NO₃)₂·6H₂O) and Rhodamine 6G dye were obtained from Sigma Chemicals (USA) and used as received without further purification. All the reagent solutions were prepared in double-distilled water and the glassware used in the experiments were properly cleaned and dried in a hot-air oven at 180 °C for 2 h before use. Sugarcane press mud generated from the filtration of sugarcane juice was procured from TR Solvents Pvt. Ltd.