4-Nitrophenol, hydrochloroauric acid trihydrate (HAuCl4 · 3H2O), sodium borohydride, and (+)-catechin hydrate were purchased from Sigma-Aldrich (St. Louis, MO, USA). Carbon-coated copper grids (carbon type-B, 300 mesh) were purchased from Ted Pella (Redding, CA, USA). The RTESP AFM probe (MPP-11100-10, premium high-resolution tapping mode silicon probe) was obtained from Bruker Nano (Santa Barbara, CA, USA). Mica (grade V-1, 25 mm × 25 mm length, 0.15 mm thick) was purchased from SPI Supplies Division of Structure Probe (West Chester, PA, USA). All the other reagents were of analytical grade. The UV-visible spectra were recorded using a Shimadzu UV-2600 with a quartz cuvette (Shimadzu Corporation, Kyoto, Japan). The HR-TEM images were acquired with a JEM-3010 (JEOL, Tokyo, Japan) operated at 300 kV. The AFM images were obtained using a Dimension® Icon® (Bruker Nano, Santa Barbara, CA, USA) operated under tapping mode. The sample-loaded mica and copper grids were dried in a 60°C oven overnight before the analyses. The FE-SEM images were collected in a JSM-7100 F SEM using an accelerating voltage of 15 kV (JEOL). ICP-MS analysis was performed in an ELAN 6100 (Perkin-Elmer SCIEX, Waltham, MA, USA). The ICP-MS samples were prepared using centrifugation. The centrifugation of catechin-AuNPs was performed at 12,300 × g for 40 min, and the supernatant containing the unreacted Au3+ was used for ICP-MS analysis. The total concentration of Au3+ of the catechin-AuNPs solution was also measured using ICP-MS. The average value of the three measurements was used to determine the yield. For HR-XRD analyses, the catechin-AuNP solution was centrifuged at 12,300 × g for 40 min to remove the supernatant. The pellet was pooled and freeze-dried. The freeze-dried samples were prepared with a FD5505 freeze dryer (Il Shin Bio, Seoul, Korea). A Bruker D8 Discover high-resolution X-ray diffractometer (Bruker, Karlsruhe, Germany) equipped with a CuKα radiation source (λ = 0.1541 nm) was used in the range of 20° to 90° (2θ scale).
The stock solutions of HAuCl4 · 3H2O (0.5 mM) and catechin (0.5 mM) were prepared using deionized water. Then, 600 μL of HAuCl4 · 3H2O (0.5 mM) was placed in a 5-mL glass vial with 200 μL of deionized water, and catechin (0.5 mM, 200 μL) was subsequently added to this solution. The reaction mixture was then further incubated under ambient temperature (26°C) for 1 h. The synthesis of gold nanoparticles was monitored through the acquisition of UV-visible spectra.
To evaluate the catalytic activity of the catechin-AuNPs, the reduction of 4-NP to 4-AP in the presence of NaBH4 was performed. The catalytic reduction of 4-NP was conducted in aqueous solution under ambient temperature (26°C), and UV-visible spectra were measured in a quartz cuvette. The 4-NP solution (899.9 μL, 0.15 mM) was mixed with deionized water (450.1 μL). Then, freshly prepared NaBH4 (1.65 mL, 5.5 mM) was added. To this reaction mixture, 1 mL of freshly synthesized catechin-AuNPs was added. UV-visible spectra were recorded at a time interval of 5 min in the range of 200 to 700 nm.