LAB 3 Work sheet

Lab 3 : Determination of pK_a for an indicator (Bromothymol Blue)

Add 0.1 ml of 0.05% bromothymol blue solution to each of the 10-ml tubes (from last week experiment) and vortex them.

Assuming that bromothymol blue is entirely in the dissociated form (In-)( blue colour) in 100% K₂HPO₄(tube 10), determine the maximum absorption by measuring absorbance (A) of solution in tube 10 at 10 nm intervals from 400-700 nm ( you can perform an automated spectrum instead of doing this manually).

Note: the blank you use for tube 10 is the same content of tube 10 but without the indicator.

Which wavelength gave maximum absorption for In-? __615___________

Measure the absorbance of tubes 2-9 at the wavelength of maximum absorption and calculate the % of In^- in each tube based on the assumption that tube 1 contains 0% In^- and tube 10 is 100% In^-.

Note: the blank you use for tubes 2-9is tube 1 with the indicator.

Calculations

% In-= assuming that tube 10 has 100% In- , so

Absorbance 10= 100%

Absorbance tube x= ?%

HIn= 100% - In%

5= at % 50 of In- = pH=pKa

When log is zero = pH=pKa

Test tube #	2	3	4	5	6	7	8	9	10(C1)
Absorbance	0.009	0.049	0.093	0.232	0.314	0.538	0.649	0.743
% of In- A1A2 =C1C2
% of HIn (100 - In-%)
Log In-HIn