The minimum inhibitory concentration of abietic acid was studied by broth micro dilution method using 96-well microtitre plates.9 Test compound was dissolved in DMSO (1%) with the addition of Tween-80(0.5%) and diluted in Muller Hinton Broth to get a concentration range of 100-1000μg/mL. The solution was then two fold diluted in Muller Hinton Broth (100 μL), inoculated with bacterial strains and then incubated at 37°C for 24 h. The bacterial growth was measured as turbidity with a Cyberlab microplate reader at 405 nm. The minimum inhibitory concentration was defined as the lowest concentration of test compound that inhibits the growth of the test bacteria. DMSO assayed as the negative control at a concentration of 1% did not inhibit any of the strains tested. All tests were assayed in triplicate in three independent experiments and median values were used for MICs calculation.
3) Centrifuged at 2500 rpm for 12 mins. Upper hexane layer (supernatant) was transferred carefully into another test tube. 4) Evaporated the hexane under a stream of grade 1 nitrogen gas and added 100 µl of methanol to the residue left and vortexed for 1 min. 5) Injected 100 µl of extract in HPLC vials and closed properly. Standard curves and calculations- Retinol was quantified from standard curves peak area for each vitamin.
MATERIALS AND METHODS Bacterial strains and culture conditions Two S. aureus strains were used in the present study; S. aureus 8325-4 (SigB-) and SH1000 representing a SigB+.strain. Overnight cultures were grown in Luria Broth (LB) at 37°Cwith shaking at 150 rpm. Exposure to antibiotics was carried out as detailed below. Antibiotics Ciprofloxacin were purchased from Sigma-Aldrich CO. 10 mg/ml stock solution of antibiotic were prepared freshly with 0.1N HCl and stored at -20°C. During the experiment we diluted with sterile water 1:10 and 1:100 depending on the different drug concentration.
Two ml of washed pack cells and 250 unit of FVIII were put into a dialysis bag (Sigma). Dialysis bag presoaked for 4 h in 50 ml PBS buffer containing 5 mM glucose. The bag was placed in a bea¬ker equipped with a magnetic stir bar. The beaker filled with 100 ml of hypoosmotic buffer containing 0.0451 mM KH2PO4/KOH, 0.1 mM MgCl2, 0.22 mM glucose, and 0.2 mM of adenosine triphosphate; pH 7.45. Dialysis was
The fresh intestine was collected, which was cut through lumen. The microsphers equivalent to 100 mg of the drug was taken on mucosal side of the intestine, was placed b/n donor and receptor compartment; in such a way that mucosal side is facing towards donor compartment. One ml of phosphate buffer pH7.4 was added to donor compartment, the receptor compartment is filled with 25 ml of phosphate buffer pH7.4. The rate of drug permeation was studied by using collection of receptor fluid at regular time intervals, and analysed for drug
Subsequently extracted by Microwave at power level 70 for 16 minutes and then filtered. The filtrate obtained, was added 10% HCl (until pH 2-3). Then do the bleaching with NaOCl diluted with water 1: 1 to white. Then converted to sodium alginate by adding 20 g of Na2CO3 and stirred in a mixer. The resulting solution is then etched with ethanol to form sodium alginate fibers.
This has to be prepared fresh. Procedure : (a) Preparation of calibration curve :(i) Pippet portions of standard NiSO4 solution into 100 ml volumetric flask. Use a series from 50 to 250 µg Ni. (ii) Add 25 ml 1.0 N HCl in 5 ml bromine water. (iii) Cool with cold running tap water and add 10 ml concentrated Ammonium hydroxide.
Top agar was dispensed into 50ml test tubes and then autoclaved for 15 minutes at 121℃ as well. Stored the top agar at -5℃ refrigerator when it cooled down. Preheated top agar for 20 minutes to liquefy before use. The optimal dispense temperature for top agar was 45℃. 3.3.4 Preparation of CaCl2 solution The molecular weight of CaCl2 was 111 g/mole.
Analysis of oxidative rancidity Thiobarbituric acid reactive substance (TBARS) was determined by the modified methods of Buege and Aust 1978. Five grams of sample was weighed into 50mL test tube, homogenized with 15mL of deionized distilled water using the polytron homogenizer for 10 seconds at high speed. 1mL of homogenized sample was transferred into a disposable test tube 13 x 100 mm butylated hydroxyanisole (50µl, 10%). Tricholoroacetic acid (2mL) was added. The mixture was vortexed and then incubated in a boiling water bath for 15min to develop colour.
0.825 g Na2WO4H2O (2.5 mmol), desired amounts of acidic ligand and 44.5 mL 30% hydrogen peroxide (440 mmol) placed in an 100 mL flask, stirred for 10–15 min to get a clear solution at room temperature. Then 10.5 mL cyclohexanone (100 mmol) added into it without stopping stirring. Continue stirring the reaction mixture at a reflux temperature for 8 h. After completion of the reaction, the reaction mixture cooled in the refrigerator for 12 h. Then white crystalline product obtained after filtration, washed with an ice water and dried in the air. The product weighted and the isolated yield of the adipic acid calculated which is based on the cyclohexanone that was put in the reaction flask. Mechanism: Experiment No.