U 2001 spectrophotometer

UV-visible (UV-VIS) absorption spectra were recorded with a Hitachi U-2001 spectrophotometer (Hitachi Field Navigator, Tokyo, Japan) in diethyl ether (Et₂O). MS analysis of carotenoids was performed using a Waters Xevo G2S Q-TOF mass spectrometer (Waters Corporation, Milford, CT, USA). ESI-TOF-MS spectra were acquired by scanning from m/z 100 to 1,500 with a capillary voltage of 3.2 kV, cone voltage of 40 eV, and source temperature of 120°C. The ¹H-NMR (500 MHz) spectrum was measured with a Varian Unity Inova 500 spectrometer (Varian Corporation, Palo Alto, CA, USA) in CDCl₃ with TMS as an internal standard. Because of the small amount of carotenoid sample (about 30 μg), ¹H-NMR was measured using a SHIGEMI microtube (sample solution volume 200 μl) (Shigemi Co., Ltd., Tokyo, Japan). The CD spectrum was recorded in Et₂O at room temperature with a Jasco J-500C spectropolarimeter (JASCO Corporation, Tokyo, Japan). Preparative HPLC was performed with a Hitachi L-6000 HPLC intelligent pump and Hitachi L-4250 UV-VIS detector (Hitachi Field Navigator, Tokyo, Japan) set at 450 nm. The column used was a 250 × 4.6 mm i.d. Cosmosil 5C18-II (Nacalai Tesque, Kyoto, Japan).

The evaluation for photoactivity was tested using 0.075 g of the catalyst (ZnO NPs) in 10 mg·L⁻¹ Rhodamine B solution (200 mL) at room temperature. The pollutant solution was prepared by stirring 0.0017 g of Rhodamine B with 200 mL of distilled water for 2 h. Before the experiment, O₂ was passed through the Rhodamine B solution for 60 min to saturate it. In order to fully examine the photocatalytic potential of the synthesized ZnO nanoparticles, the experiments were carried out under irradiation of both visible and UV light. The photoreactor used has four parallel lamps, at a distance of 10 cm over the samples [39 (link),40 ]. The lamps used were 15 W visible lamps of 900 lumens (made by Osram, OSRAM GmbH, Munich, Germany) and blacklight of 368 nm lamps of 830 lumens, made by Sylvania (Sylvania, Wilmington, NC, USA). All the tests were conducted at room temperature and pH equal to 6.4. The degradation of Rhodamine B was inspected, under each light source used. The absorbance of the samples was measured using the Hitachi U-2001 Spectrophotometer (Hitachi, Tokyo, Japan). The evaluation of the ratio of the measured A (absorption at each time) to the initial (A_initial) allowed the determination of the ratio C/C_o [39 (link)]. After a few hours upon irradiation, the degradation can be observed by watching the color progressively turning lighter, until it turns completely transparent.

At the end of the experimental period, photosynthetic pigments in fully developed green leaves from each treatment were extracted using 0.05 g of fresh material in 10 mL of 80% aqueous acetone. After filtering, 1 mL of the suspension was diluted with a further 2 mL of acetone, and chlorophyll a (Chl a) and chlorophyll b (Chl b) contents were determined with a HitachiU-2001 spectrophotometer (Hitachi Ltd., Tokyo, Japan) using three wavelengths (663.2, 646.8 and 470.0 nm). Concentrations of pigments (µg × g of fresh weight⁻¹) were obtained following the method described by Lichtenthaler [36 ].