Uv 1900i

UV spectrum method was conducted to clarify the metal chelating ability of 2-(4-(benzyloxy)-5-(hydroxyl) phenyl) benzothiazole derivatives. Test compound solution (1.0 ml, 75 µM in methanol) and metal ion solution (1.0 ml, 75 µM in methanol) were added to a vial. The vial was placed at room temperature for 2 h. Then, the mixture was removed from the vial to a quartz cell. After transfer, the absorbance of the mixture at 200–800 nm was measured by using UV spectrophotometer (Shimadzu UV-1900i).
As the Fe²⁺ played important role in the pathogenetic process of PD, the stoichiometry of 3h-Fe²⁺ complex was detected by using molar ratio method. In detail, test compound solution (1.0 ml, 75 µM in methanol) was incubated with Fe²⁺ solution (1.0 ml, 7.5–187.5 µM in methanol) at 37 °C for 30 min. Then, the mixture was removed from the vial to a quartz cell, and the absorbance of mixture at 200–800 nm was measured by using UV spectrophotometer (Shimadzu UV-1900i).

The blood compatibility test was conducted following the protocols described in previously published literature.^{39,40 (link)} Fresh rabbit blood obtained from the Laboratory Animal Center of Zhejiang University was centrifuged to separate erythrocytes. The obtained erythrocytes were then washed three times with PBS and diluted to 5% (v/v). The sample was cut into 0.5 cm × 0.5 cm squares and placed into a 2 mL centrifuge tube containing 1.6 mL of PBS. Subsequently, 0.2 mL of 5% (v/v) erythrocyte suspension was added. 0.2 mL of 5% (v/v) erythrocyte suspension was added to 1.6 mL of PBS and ultrapure water, respectively, to serve as negative and positive controls. After incubation at 37 °C for 1 h and centrifugation at 1000 rpm for 10 min, the supernatant was collected, and its absorbance at 540 nm was measured using an ultraviolet spectrophotometer (Shimadzu, UV-1900i). Finally, the hemolysis rate was calculated using the following formula:where Abs(sample) is the absorbance of the experimental group, Abs(positive) is the absorbance of the positive control group, and Abs(negative) is the absorbance of the negative control group.

Fecal microbial genomic DNA was extracted using FastDNA SPIN Soil Kit (MP Biomedicals, Santa Ana, CA). The DNA concentration and purity were determined by UV-vis spectrophotometer UV-1900i (Shimadzu Corporation, Tokyo, Japan), and its integrity was determined by 0.8% agarose gel electrophoresis. The DNA templates extracted above were amplified using universal primers in the V3-V4 region of the bacterial 16S rRNA gene (338F: 5′-ACTCCTAC GGGAGGCAGCA-3′ and 806R: 5′-GGACTACHVGGGTWTCTAAT-3′). The 20 μl reaction system was as follows: DNA 10 ng, 2.5 mmol/L dNTPs 2 μl, 5 × FastPfu buffer 4 μl, BSA 0.2 μl, FastPfu polymerase 0.4 μl, 5 mol/L primers 0.8 μl, adding ddH₂O supplemented the reaction system. The PCR conditions were 95°C predenaturation for 3 min, 98°C denaturation for 20 s, 58°C annealing for 15 s, and 72°C was extended for 20 s, totaling 30 cycles. Finally, 72°C was maintained for 5 min. The PCR amplification products were detected using a 2% agarose gel electrophoresis and then recovered using a DNA gel extraction kit (Tiangen Biotech, Beijing, China). Sequencing libraries were constructed with the amplified fragments, and then, qualified sequencing libraries were sequenced using the Novaseq 6000 platform (Illumina, CA, USA).

From the group of pigment compounds chlorophyll a (Chl_a), chlorophyll b (Chl_b), total chlorophylls (TCh), and total carotenoids (TCA) content were determined according to the method described by Holm (48 (link)) and Wettstein (49 (link)). For the extraction of pigments from nettle alcoholic extracts, 5 g ± 0.01 of the extract was weighed, and a total of 15 ml of acetone (p.a.) was added three times. After each addition of acetone, the samples were homogenized using a laboratory homogenizer (IKA, UltraTurrax T-18, Staufen city, Germany). The final solution was filtered through Whatman filter paper and transferred to a 25 ml volumetric flask filled with acetone to the mark. Absorbance was measured spectrophotometrically (Shimadzu UV 1900i, Duisburg Germany) at 662, 644, and 440 nm using acetone as a blank. Holm–Wettstein equations were used to quantify chlorophyll and carotenoid content according to Equation (2 (link)), while the final content was expressed in μg/g.

DPPH assay was performed as described by Thaipong et al. (2006) [47 (link)]. First, a stock solution was prepared by dissolving 24 mg DPPH in 100 mL methanol. Then, 24 mL of stock solution was mixed with 90 mL methanol to reach an absorbance of 1.1 ± 0.02 units at 515 nm, thus obtaining the working solution.
One hundred fifty microliters of the extract were transferred into a 16-mL glass bottle with a rubber stopper; 2850 μL of DPPH working solution was then added and vortexed (Vortex V-1 Plus, Biosan Ltd., Riga, Latvia). The mixture was kept in the dark, at room temperature, for 1 h. The absorbance value of the mixture was read at 515 nm against methanol using a double-beam UV-VIS spectrophotometer (UV-1900i, Shimadzu Scientific Instruments, Inc., Columbia, MD, USA). The blank was prepared with methanol (instead of extract) and treated identically to the test sample. The absorbance value of the blank was subtracted from that of the extract. Samples were tested in triplicate. For the calibration curve preparation, Trolox was used in concentrations from 25 to 800 µmol/L. Results were expressed in µM Trolox equivalent (TE)/g dry weight (dw).

Life weight, carcass weight, and meat chemical analyses were performed at the study end (17-week-old), with two quails/experimental units used after 8 h of fasting. Quails were slaughtered according to halal procedures, then scalded, defeathered, and eviscerated [24 ]. Meat chemical composition analyses were performed on breasts and thighs. Samples were taken separately and stored at −20°C. Meat was removed from bones and homogenized and analyzed for moisture, crude protein, and crude fat content according to Association of Official Agricultural Chemists method [25 ]. Cholesterol content was determined using an enzymatic colorimetric method; samples were saponified using methanolic potassium hydroxide and Fluitest cholesterol kit (DSI, Indonesia) was used for assay. Results were recorded at 500 nm using a spectrophotometer (Shimadzu, UV1900I, EUROPA GmbH, Germany).