Lab #6

Stress Factors on Biological Membranes

Procedure

Within each of the three science sections, six stress factors were prepared for “beet bricks” which leaked a red color (from cellular tonoplasts) depending on the level of stress. Medium sized test tubes contained each set of stress factors. The stress factors were:

1. Methanol concentrations

Five test tubes were prepared with concentration mixtures of 10.0 ml ranging from 0% through 100% methanol.

2. Different pH’s

Five different pH’s were prepared using vinegar (3% acetic acid) or small tablets of solid NaOH. In this case, the control pH was between 6-7, very near neutral. Color strips were used to determine actual pH levels.

3. Sodium chloride (NaCl) concentrations

Osmotic pressure was applied to each sample by adding various amounts (0.5-2.0 g) of NaCl to each of 5 10.0 ml samples. These samples ranged by weigh from 0% - 17% NaCl.

4. Liquid detergent concentrations

Variable numbers of drops (0-10) of Joy liquid detergent were added to 10.0 ml of distilled water to create five different stress factor samples.

5. Hydrogen peroxide (H₂O₂) concentrations

Five different hydrogen peroxide samples (0-30%) were prepared.

6. Different temperature baths.

Five different water baths ranging from 2º-100ºC were prepared. Medium sized test tubes with 10.0 ml distilled water were placed in each bath.

In each stress factor treatment a control was prepared, 10.0 ml distilled water into which three beet bricks were placed. After stress factors were prepared, 3 “beet bricks” measuring approximately 0.8 cm/side were introduced into the test tubes and remained for 8-10 minutes before 4.0 ml of the liquid was decanted into cuvettes for light absorbance analysis. The light absorbance of each sample was measured on a colorimeter; blue light (470 nm) was used. LoggerPro data analysis software was used to plot the stress factor versus light absorbance.