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Antioxidants, also known as protecting agents, are substances which are added to solutions used in biology, biochemistry or microbiology to prevent chemical changes caused by exposure to oxygen. Cultures of anaerobic bacteria may require the presence of substances buffering the redox potential at negative, reducing values. Breaking cells, especially with ultrasonic waves, introduces oxygen into the cell extract, inducing oxidation of cellular compounds such as lipids or the sulfhydryl groups in proteins. As a result, enzymes possessing functional sulfhydryl groups become inactivated. Such negative effects can be minimized by the addition of protecting agents, such as ascorbate, dithiothreitol, mercaptoethanol or cysteine. These chemicals are usually used in concentrations of 10-3 to 10-2 M. Their redox potential lies in the range of 0 to -350 mV at pH 7. Sulfhydryl agents maintain the free sulfhydryl groups of proteins in their reduced state and also reduce previously formed disulfide bridges. Additionally, these agents often display chelating properties, masking heavy metal ions in solution. Good protecting agents should be stable in air and under physiological conditions, and should not be oxidized during experimentation.
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Redox Properties of Antioxidants
| Antioxidant |
Redox System |
E0' |
Ref. |
|
| Ascorbic acid |
Dehydroascorbic acid/Asorbic acid |
58 mV |
[1] |
| Cysteine |
Cystine/Cysteine |
-340 mV |
[1] |
| Dithioerythritol |
C4H8O2S2/C4H10
O2S2
|
-330 mV |
[2] |
| Dithionite |
SO32-/S2O42- |
-527 mV |
[1] |
| Dithiothreitol |
C4H8O2S2/C4H10O2
S2 |
-330 mV |
[2] |
| Pyrosulfite |
|
* |
[3] |
|
E0': Midpoint potential at pH 7.
*: Pyrosulfite in aqueous solution, gives a mixture of pyrosulfite, bisulfite, sulfite, sulfurous acid, and SO2.
The proportions depend on temperature, concentration, and pH.
[1] CRC Handbook of Biochemistry and Molecular Biology, 3rd ed., Physical Chemical Data, vol. I, p. 122-130, G.D. Fasman, ed. CRC Press, Cleveland, Ohio 1976
[2] W.W Cleland, Biochemistry 3, 480 (1964)
[3] W.D. Loomis, Methods Enzymol. 31, 528 (1974)
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