In the round-bottom flask (100 mL), we placed p-aminobenzoic acid (1.2 g) and ethanol (12 mL). We swirled the mixture until the solid dissolved completely. We used Pasteur pipet to add concentrated sulfuric acid (1.0 mL) to the flask. We added boiling stone and assembled the reflux. Then, we did reflux for 75 minutes.
Finally the hexane layer was washed with 100 ml water and then evaporated to dryness with a vacuum rotary evaporator. The concentrated extract was then passed through a chromatographic column (30 cm x 10 mm i.d) containing 2 g florisil (lower) and 1 g sodium sulphate (upper) which is pre wetted with hexane: acetone (1:1). OCPs were eluted with 25 ml hexane: acetone (1:1).The solvent was evaporated using rotary evaporator and final volume was adjusted to 5 ml, which is used for GC analysis. All the sediments were analyzed for HCH and
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.
The incubation mixture contained 2.5 ml of 1.2% (w/v) fibrin, 2.5 ml of 100 mM Tris–HCl buffer, 10 mM CaCl2 (pH 7.8), and 20 µg of enzyme. The incubation was carried out at 37°C for 30 min, and the reaction was stopped by adding 5 ml of 110 mM trichloroacetic acid containing 220 mM sodium acetate and 330 mM acetic acid. This reaction mixture was centrifuged at 3,000×g for 5 min, and the absorbance of the trichloroacetic acid (50 mM) soluble product was determined at 275 nm. One unit of fibrinolytic enzyme activity was defined as the amount of enzyme required to liberate 1 µg of L-tyrosine per minute at 37°C. The total protein determination was performed as described by Lowry et al.
First, 50 mL of the sample was placed into a 250 mL Erlenmeyer flask, and onto a stirring plate. Then, the pH of the solution was measured and adjusted to be within the range of 4 and 6, using nitric acid and sodium hydroxide. After the pH was optimal for the experiment, a single mL of indicator- acidifier reagent was added to the sample. Then, 50 mL of mercuric nitrate was place into a burette and titrated with the sample until the color of the solution turned from blue to purple. The volume of titrant used for the reaction to reach endpoint was recorded.
Shake c) Drain the lower aqueous layer through the stopcock into the same 250 ml beaker in which the solution had been prepared in steps above. d) Pour the upper solvent layer through the neck of the funnel into a clean 125 ml Erlenmeyer flask. Return the aqueous solution from the 250 ml beaker to the separatory funnel. Add another fresh 20 ml of solvent to the funnel and again extract the aqueous solution as you did in b)
4- Set up reflux system using a clean and dry condenser . 5- Place the flask on the hot plate and heat the reaction for 45 minutes - 1 hour . 6- When the reflux is over , remove magnetic stirrer and allow the reaction to cool to room temperature . 7- Add 20 ml of ice water to a separating funnel
0,1 gr of kaffir lime oil nanocapsule is weighed and diluted to 100 ml using aquadest, taken as much as 1 ml (100x dilution) to put in the reaction tube then 1ml of saturated NaCO3 solution is added to the test tube and incubated for 10 min at room temperature. Then 0.5 ml of the folinciocalteu (Chemix CV, Yogyakarta) reagent and 7.5 ml aquadest were added, the mixture homogenized using vortex and then incubated for 30 min at room temperature under dark environmental conditions. Absorbance of the sample was then measured using a UV-vis spectrophotometer at a wavelength of 770 nm. The total phenol content of the sample was interpreted to be equivalent to gallic acid based on the standard curve of obtained gallic
Fill the fourth well with 90ml dh20 to reach 100ml.Move 10 ml of the fourth well to the fifth well. FIll the fifth well with 90 ml of dh20 to reach 100ml. Start with 1% solution for Fluoride 100 ml and move 10ml of the first well into the next. Fill the well with 90ml dh20 to reach 100ml. move 10 ml of the second well to the third well.
32 100 μL of afore-prepared sample solution and the mixed reference standard were diluted 100 times with ethyl acetate. 50 μL of these dilution solutions were separated on the TLC plate coated with SNISG. The plate was developed with petroleum ether: ethyl acetate (4:1) and the movement of solvent was usually controlled at 1 cm from the upper edge. After completion, the plate was dried until no solvent smell remained. It was sprayed with an ethanol solution containing 10% sulfuric acid, and heated at an infra-red drier until obvious color came up, as shown in Fig.2 (B.ab).
Then, the pipet was rinsed with distilled water. The bulbs were then attached to the pipette; filling and dispensing water were practiced using both bulbs. Furthermore, the 250-mL beaker was weighed, and its mass was recorded. After that, the Erlenmeyer flask was filled with 100 mL of distilled water. The temperature was recorded.