1. Visible Spectrophotometer

2. Visible Spectrophotometer

3. % transmittance = (I / I0) x 100 = 100
Visible Spectrophotometer
100 % Adjust I0 I
white light
detector/recorder
100 % adjust
blank
distilled water
prism
I0 = I
% transmittance = (I / I0) x 100 = 100
absorbance = 0

4. Visible Spectrophotometer
Zero Adjust I0 I
white light
detector/recorder
zero adjust
(barrier inserted)
prism
I = 0
% transmittance = 0
absorbance = maximum

5. % transmittance = (I / I0) x 100 = 50%
Visible Spectrophotometer
Sample I0 I
white light
detector/recorder
sample
solution
prism
I < I0
% transmittance = (I / I0) x 100 = 50%
absorbance = 1.7

6. Visible Spectrophotometer
white light
detector/recorder
sample
solution
prism
I0 = I
% transmittance = (I0 / I) x 100 = 100
absorbance = 0
In order for blue solution to absorb light,
wavelength must be changed
to blue’s complement

7. Wavelength is changed by either rotating prism
Visible Spectrophotometer
white light I0 I
detector/recorder
sample
solution
prism
Wavelength is changed by either rotating prism
or position of slit

8. “Spec 20” Visible Spectrophotometer
digital
analog

9. 1. Adjust wavelength

10. 2. Zero adjust with sample compartment
empty : 0 % T

11. 3. 100 % adjust with distilled water
(blank) in sample compartment :
100% transmittance

12. 4. Record sample absorbance
(Be certain to switch MODE to
absorbance)

13. Beer’s Law
As the intensity of the color of the sample solution increases…
the absorbance increases.
A =  b c
absorbance
constant
path length
concentration

14. Determination of the Concentration of an Unknown Solution
Prepare a set of standard solutions of known concentrations
Select the appropriate wavelength
Determine the absorbance of each
Determine the absorbance of the unknown solution

15. Sample Conc.(ppm) Absorbance 1 2 3 4 5 0.0 5.0 .0023 10.0 .0045 15.02
5.0
.0023 3
10.0
.0045 4
15.0
.0056 5
20.0
.0076
Unknown ?
.0051

16. 13 ppm

17. Recall Beer’s Law: A =  b c + 0 y = mx + b = 13.0
.0051 = x C
C =
= 13.0