Concluding the Relationship Between Concentration of Color and Light Absorption Properties

Solutions of different colors will absorb a certain amount of light at each wavelength. The absorbance levels for a solution depend on both the concentration of the color in the solution and the actual color itself. Each differently colored solution will have a certain wavelength of maximum absorbance, called λmax. This wavelength for each solution can be used to determine the proportionality constant for the solution, because the relationship between absorbance and concentration is linear, as defined by Beer’s Law.1 A spectrophotometer is the tool used to measure the absorbance of a solution at desired wavelengths.

Experimental

The spectrophotometer was turned on and warmed up for 15 minutes. A colored solution, either yellow or blue was selected, and the color along with the wavelength interval for that color was recorded. Two cuvettes were cleaned with distilled water. One cuvette was filled with distilled water, and was used as a blank to calibrate the spectrophotometer to 0 absorbance. The second cuvette was first rinsed with the colored solution, then filled with the colored solution. The spectrophotometer was set to the first wavelength with the blank was inserted, and the “0 Abs” button was pressed to set the absorbance to zero. The blank was removed to allow the cuvette with the colored solution to be inserted, and the absorbance reading was recorded. The wavelength was then incremented by 10 nm, and the the blank was used again to set the absorbance to zero, and the absorbance of the colored solution was recorded. The wavelength was incremented about 15 times by 10 nm, and each time the blank was used to calibrate the spectrophotometer along with a reading of the colored solution’s absorbance. After all the data was obtained, the wavelength that produced the highest absorbance was selected as λmax. The spectrophotometer was set to λmax calibrated to 0 Abs with the blank. The absorbance of the colored solution was measured again, and the concentration was recorded. A portion of the colored solution was diluted in half, and the absorbance was measured. A portion of the dilute solution was diluted in half again, and the absorbance was measured. A portion of the double dilute solution was diluted in half again, and the absorbance was measured. The concentration for each dilute solution was also recorded.

Results

Calculations

1) Beer’s Law

A = ε b c ; b = 1

ε = A/c = 0.562/(1.0*10-5) = 56200

ε = A/c = 0.255/(5.0*10-6) = 51000

ε = A/c = 0.126/(2.5*10-6) = 50400

ε = A/c = 0..061/(1.25*10-6) = 48000

(56200 + 51000 + 50400 + 48000) / 4 = 5.16*104

2) Concentration from A

A = ε b c ; b = 1

c = (0.375)/(5.16*104) = 7.26*10-6 M

3) Concentration from %T

A = 2 - log(%T)

A = 2 - log(55.0) = 0.26

c = (0.260)/(5.16*104) = 5.04*10-6 M

Discussion

Graphing absorbance vs concentration at λmax yields a graph with a linear trendline. The line does not pass through the origin even though it should if there were a direct relationship between absorbance and concentration. This may be due to experimental errors such as inaccurate dilutions, or possibly a miscalibrated spectrophotometer. Using the graphs and Beer’s Law, the concentration of a solution can be determined from absorbance, and also transmittance. With a 0.375 absorbance, the concentration is 7.26*10-6 M. At 55.0% transmittance, the concentration is 5.04*10-6 M. Both these points lie between the concentrations of the original solution and the first dilution.

Conclusion

The value of λmax is consistent with the color of the solution, but it is not the wavelength of the solution, rather, it is the opposite color of the solution. λmax for the blue solution is 630 nm and it is the wavelength of orange, and orange and blue are opposite colors. λmax for the yellow solution is 424 nm and it is the wavelength of violet, and violet and blue are opposite colors. The graphs were mostly consistent with Beer’s law, they all followed a positive linear trend that showed a direct relationship between concentration and absorbance.