HPLC of Analgesic
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University Of Arizona *
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326
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Medicine
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Apr 3, 2024
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Abdallah Alazhari HPLC of Analgesics Abstract
Two over-the- counter medicines were analyzed by using High-Pressure Liquid Chromatography (HPLC) to measure the amount of acetaminophen,aspirin, and caffeine in the pill. Standards were made in the dilutions from a mixed standard set provided that contained the three analgesics. The measured times for each peak was acetaminophen at 1.49 minutes, caffeine 1.48 minutes, and aspirin at 4.46 minutes. The area under the curve of each peak was not measured, so the maximum value at the peak, and the baseline of the peak were taken in order to find the concentration of each analgesic.
Methods
Three pure standards of caffeine, acetaminophen, and aspiring were given to directly inject into the HPLC machine in order to measure their absorbance. The mixed standard contained the following concentrations in ppm: 3029.4 for acetaminophen, 3091. for aspirin, and
1061.1 for caffeine. From here, a set of dilutions were made in the following ratios: 1 to 10, 1 to 50 and 1 to 100. This was done volumetrically pipetting the mixed standards into a volumetric flask and filling it with nano pure water. Two unknown brands of analgesics were given in the tablet form. The averages of the tablets were
taken, and 1 tablet per unknown was grinded into a fine powder, and mixed in nanopure until a homogenous solution was obtained. Measured for sample A-1 was 0.0237g of fine powder, while 0.0231g of fine powder for sample A-2 were recorded. The solutions were then transferred into a plastic bottle, and filtered with a 0.45
μ M
syringe into a 20mL glass vial, to be ready for HPLC analysis
Results and Discussion Chromatograms of pure and mixed standards, as well as the unknowns were taken. Peaks will be labeled in order to show each specific component. Image 1:
Chromatogram of mixed standards (1 to 10 dilution) with their respective peaks labeled. Acetaminophen is showing a peak at 1.550 minutes, caffeine at 2.00 minutes, and aspirin at 4.456 minutes. Graph 1: Calibration curves for each of the standards obtained from the dilutions. From these equations, will be able to determine the concentration of the unknown, as well as the amount of mg per tablet for each analgesic. Image 2
: Chromatogram of unknown A-2, with acetaminophen having a peak at 1.503m with an area of 76.1568, caffeine showing a peak at 2.000m with an area
of 7972. 1204, and aspirin showing its peak at 4.405 with an area of 8048.2962.
From these images and using the calibration curves from
graph 1, we are able to make the following table in order
to determine the concentration in ppm, and convert to mg per tablet. The following table was made determining the concentrations of each analgesics respective to their sample. Analgesic
Sample A-
1 Area Concentratio
n (ppm) Sample A-
2 Area Concentration
(ppm) 2
Acetamin
ophen
6633.4326
4.047
76.1758
-1.800
Caffeine 8107.4094
2.431
7972.1204
2.386
Aspirin 14763.2244
4.006
8048.62
1.497
Table 1: Concentration of each analgesic with their unknown sample. Unfortunately, the concentrations that were calculated seem too low and incorrect to be contained in a tablet, especially the unknown for acetaminophen. This may be due to the standards not being prepared properly, or using the correct concentrations to set up the calibration curves. However, from these values we are able to obtain a value for milligrams per tablet. Analgesic A-1 concentratio
n A-1 mg per tablet A-2 Concentratio
n
A-2 mg per tablet Acetamin
ophen
4.047
11.699
-1.8
4.966
Caffeine 2.431
7.027
2.386
6.583
Aspirin 4.006
11.580
1.497
4.130
Table 2: Analgesics with their respective concentration in ppm, as well as their quantities in milligrams per tablet. These values, unfortunately, seem to be too low to
be sold over the counter or have any effect. Conclusion
An over the counter medicine was given to us and analyzed using HPLC. Given three pure standards of acetaminophen, aspirin, and caffeine, we were able to create calibration curves of absorbance versus concentration to be able to find the amount of each analgesic in milligrams per tablet. The amount of each analgesic in a tablet is as follows: 76.41mg of acetaminophen, 607.48mg of aspirin, and 41.4mg of caffeine. These amounts fall in the range of what is an acceptable amount of each analgesic in medication, and there were no contaminants.
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