|MadSci Network: Chemistry|
It is quite possible, and in fact it is the easiest way to find the acid dissociation constants for these substances. Here is a procedure that is fairly simple. It assumes that you have fairly large quantities of material available. It can be scaled down as required: (1) Make up a large amount of stock solution of the anthocyanin at a fairly low concentration, so that when you measure its spectrum, the highest peak has an absorbance (optical density) of about 1 in the cells you are going to use. (2) pipette 25 mL of stock solution into a 50 mL volumetric flask, and make up the solution with 0.1 M hydrochloric acid. This will produce a fairly pure acid form of the dye. Measure the spectrum of the acid form of the dye. (3) pipette 25 mL of stock solution into a 50 mL volumetric flask, and make up the solution with 0.1 M sodium hydroxide. This will produce the pure basic form of the dye. Measure the spectrum of the basic form of the dye. (4) examine the two spectra, and find a wavelength where one form of the dye has an absorption peak, and the other form has a much lower absorption. It is not a bad idea to use two such wavelengths as a cross-check. (5) carefully remeasure the optical densities of the two solutions with the colorimeter set to this wavelength (these wavelengths). (6) you must now try to make up a solution whose dye concentration is the same, and whose absorbance is equal to (Aacid + Abase)/2. To do this you must pipette 25 mL of stock solution into a 50 mL volumetric flask, and add buffer solution to make it up to the volume. Adjust for a colour halfway between the two forms by eye by adding bits of, say, acetic acid and concentrated sodium acetate solution. Then make it up to 50 mL. If the absorbances you measure are not quite right, you can make final (careful) adjustments with single drops of concentrated hydrochloric acid or caustic soda. You can only use a few drops, or the total volume will change significantly, affecting the concentrations. (7) When you eventually have a solution whose absrobances are exactly halfway between those of the acidic and basic forms, measure its pH with a carefully calibrated pH meter. That pH will be exactly equal to the pKa of the dye. (8) Some of these manipulations are not trivial. Seek advice and help from a teacher or trained chemist who is familiar with these sorts of measurements.
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