Synthesis of mesoporous zinc oxide (ZnO) structures. Mesoporous ZnO samples (3 mg each) were prepared by following a two-step synthetic approach, as previously described [7 (link)]. In the first step, a metallic Zn layer was deposited at room temperature on silicon (Si) substrates (~1 cm2 area) by radio-frequency magnetron sputtering for an overall deposition time of 4 h. Then, thermal oxidation of the Zn-coated Si samples was performed in a muffle furnace at 380 °C (ramp rate 150 °C/h) in air for 2 h. Before Zn deposition, the Si substrates were cleaned in an ultrasonic bath of acetone and ethanol (10 min for each washing cycle) and dried under nitrogen flow.
Gentamicin sulfate (GS) adsorption and release experiments. GS powders were dissolved either in bidistilled water or simulated body fluid (SBF) at room temperature, under continuous stirring conditions (360 rpm) for 30 min. SBF salt solution was prepared according to Kokubo’s protocol [58 ]. Both GS/H2O and GS/SBF uptake solutions had a final concentration of 250 μg/mL. Adsorption of GS was performed at room temperature by soaking the mesoporous ZnO/Si samples for different times (1 h, 2 h, 5 h and 24 h) in a plastic tube filled with the uptake solution (5 mL), under orbital shaking conditions (160 rpm). After GS uptake, all the samples were washed with bidistilled water and air-dried overnight.
GS release experiments were carried out by soaking the GS-ZnO/Si samples in a plastic tube filled with SBF (10 mL), in orbital conditions (160 rpm) at 37 °C for up to seven days. At specific points of time (5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 24 h, 48 h, 72 h, 7 days), 350 μL aliquot was collected from the release solution, centrifuged at 20,000× g for 5 min and analyzed by UV-Vis spectroscopy. The drug release profile was then obtained by considering the characteristic GS absorbance peak at 251 nm. This was compared with a calibration curve obtained from the UV absorbance values at 251 nm of a series of GS dilutions in SBF (from 5 to 1000 μg/mL). The amount of drug at each release time wt was obtained according to the following equation:
where Ct is the concentration of the solution collected at each release time ti (being i an integer = 1, 2, …, n), while Vt is the residual volume of the release solution at that time, i.e., the starting release volume (10 mL) depleted at each time point ti of a fixed volume (ΔVi = 350 μL). The cumulative GS release profile was then obtained according to this equation:
where w0 is the starting amount of GS loaded on the mesoporous matrix.
Sample characterization. The morphology of the samples was evaluated by means of Field-Emission Scanning Electron Microscope (FESEM, Supra®40, Carl Zeiss AG, Oberkochen, Germany). X-ray Diffraction measurements were performed by a Panalytical X’Pert PRO diffractometer in Bragg-Brentano configuration, equipped with a Cu Kα monochromatic radiation (λ = 1.54059 Å) as X-ray source. The chemical composition was investigated by Energy Dispersive Spectroscopy (EDS), using a desktop SEM Phenom XL (Phenom-World B.V. part of Thermo Fisher Scientific, Eindhoven, The Netherlands) equipped with an EDS analyzer. UV-Vis absorbance spectra were collected in the range 200–285 nm, by means of a double-beam Varian Cary 5000 UV-vis-NIR spectrophotometer (Milan, Italy). UV analysis of drug solutions were carried out in a quartz cuvette, with an optical path length of 1 mm, analyzing a volume of 350 μL. All of the UV spectra were background subtracted. IR spectroscopy was carried out with a Nicolet 5700 FTIR Spectrometer (ThermoFisher, Waltham, MA, USA), equipped with a room temperature working L-alanine doped triglycine sulfate (DLaTGS) detector. All of the spectra were background subtracted and acquired with 2 cm−1 resolution and 64 scans accumulation. The release profile for Zn2+ element was determined with an Inductively Coupled Plasma Mass Spectrometer analyzer (ICP-MS, mod. 7500cc, Agilent Technologies, Milan, Italy).
Gentamicin sulfate (GS) adsorption and release experiments. GS powders were dissolved either in bidistilled water or simulated body fluid (SBF) at room temperature, under continuous stirring conditions (360 rpm) for 30 min. SBF salt solution was prepared according to Kokubo’s protocol [58 ]. Both GS/H2O and GS/SBF uptake solutions had a final concentration of 250 μg/mL. Adsorption of GS was performed at room temperature by soaking the mesoporous ZnO/Si samples for different times (1 h, 2 h, 5 h and 24 h) in a plastic tube filled with the uptake solution (5 mL), under orbital shaking conditions (160 rpm). After GS uptake, all the samples were washed with bidistilled water and air-dried overnight.
GS release experiments were carried out by soaking the GS-ZnO/Si samples in a plastic tube filled with SBF (10 mL), in orbital conditions (160 rpm) at 37 °C for up to seven days. At specific points of time (5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 24 h, 48 h, 72 h, 7 days), 350 μL aliquot was collected from the release solution, centrifuged at 20,000× g for 5 min and analyzed by UV-Vis spectroscopy. The drug release profile was then obtained by considering the characteristic GS absorbance peak at 251 nm. This was compared with a calibration curve obtained from the UV absorbance values at 251 nm of a series of GS dilutions in SBF (from 5 to 1000 μg/mL). The amount of drug at each release time wt was obtained according to the following equation:
where Ct is the concentration of the solution collected at each release time ti (being i an integer = 1, 2, …, n), while Vt is the residual volume of the release solution at that time, i.e., the starting release volume (10 mL) depleted at each time point ti of a fixed volume (ΔVi = 350 μL). The cumulative GS release profile was then obtained according to this equation:
where w0 is the starting amount of GS loaded on the mesoporous matrix.
Sample characterization. The morphology of the samples was evaluated by means of Field-Emission Scanning Electron Microscope (FESEM, Supra®40, Carl Zeiss AG, Oberkochen, Germany). X-ray Diffraction measurements were performed by a Panalytical X’Pert PRO diffractometer in Bragg-Brentano configuration, equipped with a Cu Kα monochromatic radiation (λ = 1.54059 Å) as X-ray source. The chemical composition was investigated by Energy Dispersive Spectroscopy (EDS), using a desktop SEM Phenom XL (Phenom-World B.V. part of Thermo Fisher Scientific, Eindhoven, The Netherlands) equipped with an EDS analyzer. UV-Vis absorbance spectra were collected in the range 200–285 nm, by means of a double-beam Varian Cary 5000 UV-vis-NIR spectrophotometer (Milan, Italy). UV analysis of drug solutions were carried out in a quartz cuvette, with an optical path length of 1 mm, analyzing a volume of 350 μL. All of the UV spectra were background subtracted. IR spectroscopy was carried out with a Nicolet 5700 FTIR Spectrometer (ThermoFisher, Waltham, MA, USA), equipped with a room temperature working L-alanine doped triglycine sulfate (DLaTGS) detector. All of the spectra were background subtracted and acquired with 2 cm−1 resolution and 64 scans accumulation. The release profile for Zn2+ element was determined with an Inductively Coupled Plasma Mass Spectrometer analyzer (ICP-MS, mod. 7500cc, Agilent Technologies, Milan, Italy).
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