Summer Research Program for Science Teachers

Tehilla Rieser

Bronx High School of Science

August 1997


How does a spectrophotometer work?


- To understand the relationship between absorption of light and color [9-12 Content Standard B- Interactions of matter and energy]

- To understand how a spectrophotometer works [9-12 Content Standard E- Understandings about science and technology]

1. Wave nature of light

• Wavelength: Distance between peaks or troughs of a wave

• Frequency: # of waves passing a point in space per second

• Visible light: Receptors in human eye can detect l= 400-750 nm

• Light appears "white" if it includes all the wavelengths of visible region.

• White light dispersed by a prism or grating produces the color spectrum.

2. Interaction of light with atoms and molecules

• Absorption: Transition of electrons from lower to higher energy levels.

• Emission: Electrons excited to high energy levels decay to lower levels by emitting light. [9-12 Content Standard B- Structure of atoms]

• Substance will appear a particular color because it absorbs visible light of every wavelength except that color.

3. Absorption of light

        • When white light is directed at a colored solution some wavelengths are absorbed. The transmitted light is now colored              because it is missing some wavelengths of white light.

Ex. A substance which absorbs blue and green light appears yellow-red.

4. Spectrophotometer:

Light bulb to produce white light

Diffraction grating to "break up" white light

Slit to allow only narrow wavelength bands to enter

Rotation of grating selects portion of spectrum illuminating sample

Sample chamber holds the cuvette

          • Meter has scale calibrated in transmittance (top) and absorbance (bottom)

Visible Absorption Spectroscopy

AIM: How is the Spectronic 20 used for qualitative analysis of colored solutions?

Equipment and material: Spectronic 20, cuvettes, a stick of white chalk, distilled water, cobalt chloride solution.

[Teaching Standard D- Make accessible science tools]

Before you begin the experiment, your teacher will review with you the proper techniques in handling the Spec. 20.

General Operating Procedure:

1. Turn on the machine; pilot lamp will light. Allow 5 minutes for warm up.

2. Select the desired wavelength.

3. With the sample chamber empty, set the meter to zero T.

(The power switch is also the zero control knob.)

4. Insert a cuvette containing the reference solvent (distilled water) and set the

meter to 100% T.

5. Insert the sample; record % T or Abs.

6. Repeat steps 2 through 5.

To ensure good readings

1. All solutions must be free of bubbles. (We are not analyzing air!)

2. Cuvette should be 1/2 to 3/4 filled.

3. Cuvette should be clean, and dry on the outside.

4. Index mark on the cuvette should be aligned with the mark on the instrument.










1. Adjust the power switch so the % transmittance reading is 0.

2. Insert a cuvette containing distilled water in the sample chamber. Close the cover and set the absorbance reading to 100% transmittance by adjusting the front right knob. Remove the cuvette.

Sample absorbance

1. Take a sample of CoCl2 solution and insert it into the sample chamber. Follow the operating procedure. Read and record the % T from 350 nm to 650 nm at increments of 25 nm.

Wavelength (nm)

CoCl2 % T




























1. Plot the data for CoCl2 solution. The wavelength should be on the x-axis and the % T on the y-axis.

[Content Standard Unifying Concepts- Change, constancy, and measurement]

2. How could this data be helpful in identifying an unknown substance? [Teaching Standard B- Orchestrate scientific discourse]

Note to teacher:

• In some models of Spec 20 the light shuts off when the sample chamber lid is open.

• Since this experiment is not quantitative, any tube that fits the sample chamber would be OK.

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