After 30 minutes, tubes were taken out and kept in ice-cold water for 30 minutes. These were centrifuged at 3000 rpm for 15 minutes. The absorbance of the supernatant was read at 540 nm at room temperature against appropriate blank. Blank consist of 1 ml distilled water, 0.5 ml of 30% TCA, and 0.5 ml of 0.8% TBA. TBARS values were expressed as n moles malonaldehyde (MDA)/mg protein.
After the evaluation of stomach for ulcers, the gastric mucosa of glandular portion was scrapped with the help of two glass slides, weighed (100 mg) and homogenized in 1 mL of a 0.15 M, ice cold potassium chloride (KCl) solution and centrifuged at 3,000 RPM for 10 minutes (REMI centrifuge). 1 mL of suspension was taken from the above tissue homogenate in test tube and 0.5 mL of 30% w/v TCA (trichloroacetic acid) was added to it, followed by 0.5 mL of 0.8% w/v TBA (thiobarbituric acid) reagent. The tubes were then covered with aluminium foil and kept in water bath for 30 minutes at 80 °C. After 30 minutes, tubes were taken out and kept in ice-cold water for 30 minutes and centrifuged at 3000 rpm for 15 minutes (R-BC DX REMI centrifuge). The absorbance of the supernatant was read in spectrophotometer (UV-1601, SHIMADZU) at 540 nm against blank.
The equipment used for preparation the extraction unit are shown in Figure 1. The pH of aqueous solution containing 10 μg L−1 of Cd (II) and 50 μg L−1 of Pb (II) was adjusted with HNO3 and NaOH (0.1 M). Then 2 mL of this solution was poured into the syringe. Afterward, 2 mg of NDNPG was added to the extraction unit and immersed in an ultrasonic bath for 60 s to disperse the NDNPG in the sample solution and increase the contact surface between the adsorbent and solution. After sonication, NDNPG was separated from sample solution easily by pressing the plunger, the sample solution from the syringe came out and NDNPG adsorbent remain in the syringe.
While under the UV light the mixtures appeared a purple colour. The R_fValue Starting material: R_F=2.6/4.5 = 0.58 The sum: R_F=2.55/4.5 = 0.57 The reaction mixture: R_F=2.9/4.5 = 0.64 The final product: R_F=1.9/4.5 = 0.42 Table of calculations Benzaldehyde Acetone Sodium Hydroxide Ethanol Dibenzalacetone Co-efficient 2 1 1 1 1 Volume (ml) 4.04 1.5 30 Grams used (g) 4.2 1.185 4 0.95 2 Formula weight (g/mol) 106.12 58.08 40 46.07 234.29 Moles 0.0396 0.02 0.1 0.02 Moles/ Coefficient 0.02 0.02 0.1 0.02 Density 1.04 0.79 (1) Calculations Mass Impure compound mass: 2.02g Final compound mass: 2.88g Dibenzalactone = (4.2×234.29×1)/(106.12×2) = 4.64g (theoretical
Cells were disrupted using RLT buffer and the lysate was homogenized by centrifugation through a QIAshradder spin column for 2 min at full speed. The flowthrough was mixed with 70% ethanol (equal volume as the RLT buffer used), transferred into an RNeasy spin column and centrifuged at 17000 g for 1 min. after having the flowthrough discard, RW1 buffer was added to the column, centrifuged at 17000 g for 1 min and the flowthrough was discard. The spin column was washed twice with RPE buffer under at 17000 g centrifugation for 2 min. The column was centrifuged empty at full speed for 1 min to remove any residual ethanol.
Commercial TiO2 P25 was obtained from Evonik. Ultrapure water (18MΩ.cm-1) was used throughout the whole experiments. 2.2. Synthesis of photocatalysts The TiO2 nanoparticles were prepared by the sol-gel method described below: 3.9 ml of TiCl4 was slowly added into 10 milliliter of absolute ethanol in reaction vesel, this reaction performed under fume hood at 0°C with vigorous stirring due to exothermic reaction,the high volatilityof TiCl4and also therelease of hydrogen chloride. Then, water was added dropwise during the mixing process.
Finally, the sample was placed in Rheometer which was maintained under constant temperature (46° C) and shear rate (34 s-1)for 150 minutes (2 hours 30 minutes). The Aton Parr Rheometer was selected for conducting the experiment. The Sensor Probe (SN33439) was 38.719 mm in diameter and 60.012 mm in length. The radius of the cup used for storing the crude oil sample was 21.000 mm with the measuring gap of 1.641 mm. The cone of the probe was making an angle of 120°.The volume of the crude sample filled in the Rheometer cup was 58 ml.
2.5mL of ferric chloride solution was added to 2.5mL of the pure extract. Occurrence of dark blue or greenish-black color indicates presence of tannins. Test for Anthocyanin. 1.25mL of Sodium hydroxide solution was added to 2.5mL of the extract. Formation of blue or green precipitates indicates presence of anthocyanin.
For ascorbate peroxidase assay extraction 241 buffer was supplemented with 1.0 mM ascorbic acid. The homogenate was centrifuged at 242 15,000×g for 15 min at 40C, and the supernatant was used as a crude enzyme extract. 243 Spectrophotometric determinations were performed using UV visible spectrophotometer 244 (UV-1700, Shimadju, Japan). 245 2.11.2. Estimation of superoxide dismutase (SOD) activity 246 SOD activity was estimated by its ability to catalyse NBT to formazan at 560nm 247 according to the method of Beyer and Fridovich (40).