Partition Coefficient Of Acetaminophen

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To determine the partition coefficient of acetaminophen, 500 µl of 1-octanol and 500 µl of 1 mg/mL acetaminophen were pipetted into a 1.5 ml tube, vortexed vigorously for 90 minutes, and then centrifuged for 1 minute. The two solutions, 1-octanol and the acetaminophen diluted in water, mimic the properties of the phospholipid bilayer and shows to what extent the acetaminophen diffuses into cells. After centrifuging, 100 µl of the aqueous layer was collected to determine the concentration of acetaminophen in the aqueous layer by doing a calorimetric assay. This portion of the experiment was done twice, one for an aqueous solution at pH 6 and one for an aqueous solution at pH 12. For the calorimetric assay, a standard curve was generated by doing…show more content…
6 mL of 2X bovine plasma was combined gently with 6 mL of 1.0 mg/mL acetaminophen in a 15 mL conical tube and then incubated for 30 minutes at 37°C. During the incubation period, a serial dilution was prepared with acetaminophen which was exactly similar to dilution prepared in the experiment described above except in this case PBS was used to dilute the sample and each sample was made to 200 µL. After incubation of the acetaminophen/plasma mixture, 10 mL of the sample was added to a dialysis tube and the ends of the tube were tied via mint wax dental floss. The tube was then submerged in a 50 mL conical tube containing 25 mL of PBS for 90 minutes at 37°C. Following this, 200 µl of the plasma from the dialysis tube was pipetted and mixed with 1.8 ml trichloroacetic acid in a 2 ml tube and centrifuged for 2 minutes. After centrifugation, 1.0 mL of the supernatant was pipetted into a new 2.0 ml tube and labeled as the plasma. 100 ml of each of the standards prepared was also transferred to a new 2.0 ml tube along with 100 ml buffer, which was collected from the conical tube, and these samples were mixed with 900 µl of trichloroacetic acid. Next 500 µl of sodium nitrite was added to all the samples including the plasma and then they were incubated for 10 minutes at 37°C. Then, 50 µl of NaOH was added to each of the samples and…show more content…
The volume of acetaminophen given to each rabbit was determined by the weight of the rabbit in kg times the 250 mg/kg of acetaminophen in a single dose, divided by the concentration of the stock acetaminophen in mg/mL. Rabbit #1, which weighed 3.07 kg, was given acetaminophen via IV. Rabbit #2, which weighed 3.19 kg, was given acetaminophen via IP. Rabbit #3, which weighed 2.96 kg, was given acetaminophen via PO. Blood sample was taken via a syringe, from an indwelling catheter in the dorsal central artery, and transferred to a Vacutainer serum separating tube at t = 0 (before the drug was administered) and then at 5, 15, 30, 60, 90, and 120 minute marks (after the drug was administered). The 7 blood sample were then transferred to 2.0 mL tubes and centrifuged at 2000 rpm for 5 minutes and then 200 µL of each of those individual samples were transferred to another set of 2.0 mL tubes and mixed with 1.8 mL of TCA and centrifuged again. Finally 1.0 mL of the supernatant of these samples was transferred to another set of 2.0 mL tubes and then mixed with 500 µL of sodium nitrate. Another set of standards for the acetaminophen was prepared in this experiment. Starting with 1 mg/ml of acetaminophen, 100 µl of 500, 250, 125, 62.5, 31.25, 15.625, and 0 µg/ml were prepared. All
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    determining the partition coefficient, shows an increased absorbance reading as the concentration increases and this is shown in Figure 1. The R2 value generated from the trend-line of the points plotted is 0.9808, therefore giving us an accurate equation of y = 1.392x – 0.0486 to determine the concentration of acetaminophen in the aqueous phase at pH 6 and 12. Plugging in the absorbance value of 1.225, at pH 12, for y in the equation gives us a value of 0.927 mg/ml for x. To determine the partition coefficient

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