PHAR 4634 - Chapter 22 Page 2    ## Analysis of Drugs by Visible Spectroscopy

AIM:

1) To determine standard curves for two substances at two wavelengths. Thus four standard curves are to be determined. These are salicylate (reacted with ferric nitrate) at 540 and 500 nm and potassium permanganate at 570 and 540 nm.

2) To exam and confirm Beer's law using the above solutions as examples.

3) To determine the concentration of an `unknown' sample.

BACKGROUND:

According to Beer's law the Absorbance of a solution can be calculated with the formula:

A = a * b * c

where a = extinction coefficient, constant for a given substance, wavelength, and solvent

where b = sample path length, which is fixed by the size of the cell used.

and

where c = concentration of substance.

Beer's law has been found to hold for a fairly wide range of concentrations. Hopefully it will apply in the range you are working in.

Beer's law applies to any wavelength, but you will be working with visible wavelengths. The potassium permanganate is a purple color, and the salicylate forms a purple color after reacting with the ferric nitrate.

METHOD:

Potassium permanganate.

Starting with 4 ml of the 0.05 % solution of potassium permanganate (provided) prepare 40 ml stock solution (a 1 to 10 dilution). Further prepare 10 ml volumes of working solutions 10, 20, 40, 60, 80, and 100 % of the stock solution. Measure the absorbance of each solution at both 570 and 540 nm. Use distilled water as the blank. Prepare the ÔunknownŐ sample and measure the absorbance at 570 and 540 nm.

Sodium salicylate solution.

Starting with 4 ml of the 0.1% solution (provided) prepare 40 ml of a stock solution (a 1 to 10 dilution). Further prepare 10 ml volumes of working solutions 10, 20, 40, 60, 80, and 100% of the stock solution. Take 8 ml of each working solution and add 2 ml of the ferric nitrate solution provided to produce a color. After the color is fully formed measure the absorbance of each solution at 540 and 500 nm. Use 2 ml of the ferric nitrate solution diluted to 10 ml with distilled water as the blank. The ferric nitrate reagent is 1% ferric nitrate in 1% nitric acid.

DATA PRESENTATION:

Prepare standard curve graphs of each solution at the wavelengths used. Calculate the extinction coefficient for each graph. You should have four graphs and four extinction coefficients. Use a path length of 1 cm.

WRITE-UP to be turned in should include:

1. Aims
2. Methods - a brief description of the experiment
3. Results - the raw data
4. Analysis of the data - graphs with linear regression
5. Conclusion - overall conclusion(s) from the experiment    