OLE solutions were made by dissolving 15 mg of each crude extract in 15 ml dimethyl sulfoxide (DMSO) (Fine-Chem, Mumbai, India). Then, 0.5 ml of each solution was mixed with 2.5 ml of 0.2 N Folin-Ciocalteu reagent (Sigma-Aldrich, USA) for 5 min at room temperature then 2 ml of sodium carbonate solution (7.5 % in deionized water, w/v) were added. After incubating for two hours at room temperature in a dark place, the absorbance was measured at 760 nm using UV/Visible spectrophotometer (Elico, SL-150, India). The concentrations between 0.01-0.05 mg.ml-1 of gallic acid (Sigma-Aldrich, USA) were used as a standard for the calibration curve. The total phenolic content was expressed as mg gallic acid equivalent.g-1
The peel powder was soaked overnight (or for desired period) at room temperature (30-32°C) for extraction with intermittent shaking. After extraction, it was centrifuged at 2000 rpm for 5 min and filtered through Whatman No. 2 filter paper. The effective volume obtained after centrifugation was noted. 2.3 Determination of radical scavenging activity (RSA) Determination of RSA was carried out according to Murthy, Jayaprakasha, & Singh (2002) using DPPH as stable free radical and butylatedhydroxyanisole (BHA) as standard.
First the solution of 1 mL of extract was added to deionizer water (10 mL), then Folin–Ciocalteu phenol reagents (1.0 mL) added to the mixture. The mixture was left for 5 minutes, and solution of 20% sodium carbonate (2.0 mL) was added to the mixture. This mixture diluted until 50 mL with deionizer water. And after that kept in total darkness room for 1 hour. The mixture absorbance was measured at 750 nm using a spectrophotometer.
Step-III: Synthesis of Cr(II) Complexes: The Schiff's base complexes were synthesized by mixing the Schiff's base (1.5 g) in ethanolic solution of Chromium chloride [CrCl2]. This reaction is refluxed in a waterbath for two hours and their volume were reduced to 70% of it’s original volume and residue was obtained. The coloured product obtained was filtered under suction, washed with ethanol. The product were recrystallized from ethanol. Their yields ranges from 50-55%, the product obtained were light green colour and melting point was 2100C.
Then, the test media is then incubated at 37 ° C, for 18-24 hours. Rinsing reusable instruments The samples were rinsed with 40 ml of pyrogen-free water using a glass beaker that is free from pyrogens. Endotoxin testing using STV A total of 0,2mL from the water obtained from the rinsing was placed in the STV containing LAL reagent and was shaken for 20 to 30 seconds. Then STV was placed in an incubator at 37 ° C for 60 ± 2 minutes. STV was then observed by reversing the reaction tube in one smooth motion.
Stored in light glass bottle. 2.5.2: Formation of PANI(CoFe2O4) 0.1g of Nano material and 50ml of Aniline hydrochloride were mixed in a beaker. Then 50ml of Ammonium peroxydisulphate was added drop wise in reaction mixture with constant stirring below 20 oC. After 24 hours the coated sample was filtered, washed and dried at 60 oC in oven and then grinded into a fine powder in agate
The test tubes with and without enzyme (control) were incubated at 370C for 1 hour. After incubation, the enzyme was denatured by heating the tubes at 1000C for 5 minutes. Both the test treated and untreated samples (control) were assayed for antibacterial activity by agar well diffusion method15. 7. Antibiotic Sensitivity
This diluted solution will be used in the assay as duplicate samples. Then, 1.0mL of standard glycine solution containing (7.5mg/mL) was diluted to 100mL with water using a volumetric flask. This solution contains 1.0µmole/mL of glycine. 8 tubes were set up according to the following protocol and 2.0mL of ninhydrin reagent was then added to each of the 8 tubes and were placed in a boiling water bath for 20 minutes. After 20 minutes, the tubes from the bath was carefully removed, cooled in a beaker of cold water, then 8.0mL of 50% ethanol was added and mixed well.
Kashibai Navale college Of Pharmacy, Kondhwa Bk. Preparation of Leave Extract 50 g of Psidium guajava linn leaves are washed with distilled water and keep in dark room for 4 days and grind into powder form. 2 g of powder dissolved in ethanol, methanol, pet ether of 50 ml of solvent respectively in incubator at 40oc for 5 days. The extract was subsequently filtered and concentrated to dryness. Antibacterial assay The screening was done by disc diffusion method.
2.3. Preparation procedure for DS loaded PC-SA combined beads The ionotropic gelation method was used for the preparation of DS loaded PC-SA bead. The gum (PC) was dissolved in distilled water and initially boiled for 10 minutes, then cool and keep stirring for 24 hours at 400 RPM. Then filtration