Visible Spectrophotometer
Visible Spectrophotometer
% 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
Visible Spectrophotometer Zero Adjust I0 I white light detector/recorder zero adjust (barrier inserted) prism I = 0 % transmittance = 0 absorbance = maximum
% 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
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
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
“Spec 20” Visible Spectrophotometer digital analog
1. Adjust wavelength
2. Zero adjust with sample compartment empty : 0 % T
3. 100 % adjust with distilled water (blank) in sample compartment : 100% transmittance
4. Record sample absorbance (Be certain to switch MODE to absorbance)
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
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
Sample Conc.(ppm) Absorbance 1 2 3 4 5 0.0 5.0 .0023 10.0 .0045 15.0 2 5.0 .0023 3 10.0 .0045 4 15.0 .0056 5 20.0 .0076 Unknown ? .0051
13 ppm
Recall Beer’s Law: A = b c + 0 y = mx + b = 13.0 .0051 = 0.00037 x C + 0.00030 C = = 13.0