Uv 1700 pharmaspec

The Folin-Ciocalteu assay is widely used to assess the total polyphenol content of samples [35 (link)]. This assay relies on the transfer of electrons from phenolic compounds to phosphomolybdic/phosphotungstic acid complexes in an alkaline medium to form blue complexes that are detected spectroscopically at 700 nm. This assay was used to assess the antioxidant activity of the spinach samples by adding 200 μL of each extracted sample to 500 μL of Folin-Ciocalteu reagent and leaving it for 5 min at room temperature. Following this, 500 μL of 10% sodium carbonate solvent in ultrapure water was added to each sample, mixed by vortex, and left for 1 h at room temperature. Color development was then determined at 700 nm using a spectrophotometer (UV-1700 PharmaSpec; Shimadzu). We measured antioxidant activity by two additional methods: Oxygen Radical Antioxidant Capacity (ORAC) assay (Cell BIOLABS, INC, CA, USA) and Trolox Equivalent Antioxidant Capacity (TEAC) assay (Cayman Chemical Company, MI, USA). In ORAC assay, samples and standards were measured with a fluorescent microplate reader (Fluoroskan Ascent FL, Thermo Fisher Scientific, USA) at 37°C with an excitation wavelength of 480 nm and an emission wavelength of 520 nm. In TEAC assay, color development measured at 405 nm using a spectrophotometer (UV-1700 PharmaSpec, Shimadzu).

Serum total antioxidant capacity of experimental rats was assessed using a modified Trolox equivalent antioxidant capacity (TEAC) assay described by Katalinic et al. [25 (link)] and Chan et al. [18 (link)]. ABTS^•+ stock solution was prepared by reacting 7 mM of ABTS with 2.45 mM of potassium persulfate. After 18 h of incubation in the dark at room temperature, the stock solution was diluted with phosphate buffer saline to the absorbance of 0.70 ± 0.02 at 734 nm (PharmaSpec UV-1700, Shimadzu, Kyoto, Japan). Subsequently, 50 μL of diluted serum was reacted with 950 μL of adjusted ABTS^•+ solution for 10 min, and the absorbance was measured at 734 nm (PharmaSpec UV-1700, Shimadzu, Kyoto, Japan). Trolox was used as standard, and the serum total antioxidant capacity of experimental rats was expressed as mg Trolox equivalent antioxidant capacity (TEAC)/mL serum.

LC-MS analysis was performed with Agilent Technologies^® 6130 Quadrupole LC/MS (Santa Clara, CA, USA). UV–vis spectra were obtained using Shimadzu^® UV-1700 PharmaSpec (Kyoto, Japan). IR spectra were determined using Bruker^® Alpha FT-IR (Billerica, MA, USA). NMR spectra of the compounds were acquired on a Bruker^® Avance III spectrometer operating at 600 MHz for ¹H and 150 MHz for ¹³C in deuterated chloroform (Billerica, MA, USA). HRESIMS analysis of compounds 1–6 were performed using Agilent^® 6530 Accurate Mass Q-TOF (Santa Clara, CA, USA), while the HRESIMS data of turcicasulphide (7) were acquired on a Thermo Scientific-Q Exactive^® (Waltham, MA, USA). Optical rotation data were acquired using a Rudolph Analytical Autopol V Plus^® in dichloromethane (Hackettstown, NJ, USA). A Buchi rotary evaporator was used to evaporate the solvent of the extract (Buchi, Flawil, Switzerland). A Sephadex LH-20 (Sigma Chem. Co. 25–100 µm) (GE Healthcare, Chicago, IL, USA) column (5 × 100 cm) was used for the initial fractionation. A Gilson^® PLC 2050 was used for the further purification of the compounds (Saint-Avé, France). Hexane, dichloromethane, methanol, and acetonitrile (Merck, Darmstadt, Germany) were used during the chromatographic analyses.