Rey Vincent P.

Antonio Experiment 10: Spectrophotometric Determination of the Acid Dissociation Constant of Methyl Red Results and Discussion Spectrophotometry is the measurement of the amount of radiant energy absorbed by a chemical system to determine the concentration of the absorbing species. This absorptive capacity is based on the wavelength of the radiation and absorption measurements at a predetermined wavelength. It is detected using an UV-VIS spectrophotometer. The logarithm of the reciprocal of the transmittance of the solution defines the absorbance of the solution. The transmittance of the solution depends on the ratio of the intensities of the radiation entering the solution and the radiation unabsorbed and transmitted by the solution. Aside from the concentration of the analyte, the absorbance can be expressed as a function of the molar absorptivity constant of the solution at a specific wavelength. This relationship is represented by the Beers law:

A = bc
where A is the solutions absorbance, is the molar absorptivity constant, b is the path length (usually 1.00cm), and c is the concentration of the sample. In the experiment, the acidic (HMR) and basic (MR-) forms of the weak acid methyl red is analyzed spectrophotometrically.

HMR MR + H +
From the equation, the acid dissociation constant, Ka, is derived as:

Ka =

[ MR ][ H + ] = 5.0 [ HMR]

Using the Henderson-Hasselbach equation for this system, the following equation is obtained:

[ MR ] pH = pK a + log [ HMR]
A stock methyl red solution was prepared from 0.05000g of solid methyl red, diluted with 95% ethyl alcohol and distilled water to create a 50mL stock solution. Fifty milliliters of methyl red standard solution was prepared by adding 2.50mL of the stock solution to 95% ethyl alcohol and distilled water. An acidic (HMR) and a basic (MR-) solution were prepared from 5mL of the standard methyl red solution. A low pH of 2 was maintained for the HMR solution to force the equilibrium to shift towards the production of acidic methyl red. This process also minimizes the amount of MR- molecules which can affect the absorbance reading for HMR. Similarly, the basic solution was maintained to be at high pH to shift the reaction to produce MR- and minimize HMR. Three acidic and three basic solutions with different concentrations were produced by adding hydrochloric acid and sodium acetate, respectively, to the prepared solutions. This was done to produce a calibration curve of concentration against the absorbance to determine the molar absorptivity constant of the solutions. In measuring the absorbances of the solutions using a spectrophotometer, a blank reference sample was used throughout the experiment to minimize errors. Two reference absorbance wavelengths were used. The first one is based from the reference acidic solution and the other based from the reference basic solution. Assuming minimal interference from the other species, strong absorbances are expected at these absorbance wavelengths. The curves for the several measurements at different wavelengths are shown below.

Molarity (HMR) vs Absorbance (at HMR)

0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0.00E+ 2.00E- 4.00E- 6.00E- 8.00E- 1.00E- 1.20E- 1.40E- 1.60E00 06 06 06 06 05 05 05 05 Molarity y = 47924x - 0.0065

Figure1. Molarity of HMR solution against its absorbance at HMR

Absorbance

Molarity (HMR) vs Absorbance (at MR-)

0.12 0.1 Absorbance 0.08 0.06 0.04 0.02 0 0.00E+ 2.00E- 4.00E- 6.00E- 8.00E- 1.00E- 1.20E- 1.40E- 1.60E00 06 06 06 06 05 05 05 05 Molarity y = 7668.1x - 0.011

Figure2. Molarity of HMR solution against its absorbance at MR-

Molarity (MR-) vs Absorbance (at HMR)

0.025 Absorbance 0.02 0.015 0.01 0.005 0 0.00E+ 2.00E- 4.00E- 6.00E- 8.00E- 1.00E- 1.20E- 1.40E- 1.60E00 06 06 06 06 05 05 05 05 Molarity y = 1597.1x - 0.0019

Figure3. Molarity of MR- solution against its absorbance at HMR

Molarity (MR-) vs Absorbance (at MR-)

0.3 0.25 Absorbance 0.2 0.15 0.1 0.05 0 0.00E+ 2.00E- 4.00E- 6.00E- 8.00E- 1.00E- 1.20E- 1.40E- 1.60E00 06 06 06 06 05 05 05 05 Molarity y = 18956x - 0.0037

Figure4. Molarity of MR- solution against its absorbance at MR-

Based on Beers Law and looking at the linear equations of the calibration curves, the values for the molar absorptivity constants of the solutions on different wavelengths were obtained. The slopes of the lines were approximately equal to the molar absorptivity constant, . On the next part of the experiment, four solutions containing 6mL each of methyl red standard solutions were prepared. Acetic acid and sodium acetate of varying amounts were added to vary the acidity of the solutions. The absorbances of these solutions are due to both absorbances of both species. The absorbances of the solutions were measured at both HMR and MR-. Still based from Beers Law, the equations for the total absorbance on both wavelengths as a function of concentration is given by:

A1 = HMR ,1bc HMR + MR ,1bc MR A 2 = HMR , 2 bc HMR + MR , 2 bc MR

From the obtained total absorbances, the concentrations of the species per solution were determined using the equations above. The plot of the logarithm of the ratio of the concentrations of MR and HMR versus the measured pH of the solutions is given by the figure below.

log [MR-]/[HMR] vs pH
7 6 5 pH 4 3 2 1 0 -4.00E- -2.00E- 0.00E+0 2.00E- 4.00E- 6.00E01 01 0 01 01 01 log [MR-]/[HMR]

y = 0.991x + 5.0437

8.00E- 1.00E+0 01 0

Figure5. The plot of

log

[ MR] versus pH. [ HMR]

From the Henderson-Hasselbach equation, the y-intercept of the curve represents the pKa of the reaction. The experimental pKa was found out to be 5.04, while the Ka was calculated to be 9.04 x 10 -6. The calculated Ka is 9.57% deviated from the theoretical value of 1.00 x 10-5. This error can be caused by several factors throughout the experiment. One of these could be improper preparation of solutions. The concentrations of the solutions may have been misstated, therefore affecting the calibration curves as well as the value for the molar absorptivity constant. Contaminations on the solutions may have also occurred, affecting the actual concentrations of the solutions. Improper handling of the cuvette may have also caused contamination and affecting the absorbance reading of the sample. Machine error could also be the cause of the discrepancy. The spectrophotometer used in the experiment may not be in a good condition. Also, the assumption that the total absorbance was only caused by the two methyl red species in the solution may not be true, hence, causing overstated values for the concentrations of these species.