The use of purified enzymes for the early assessment of toxicity

TR Number



Journal Title

Journal ISSN

Volume Title


Virginia Polytechnic Institute and State University


The increasing level and dispersion of toxic chemicals in the environment has stimulated a need for accurate methods capable of detecting and quantifying the activity of toxicants. The primary objective of this study was to determine, through in vitro tests, the potential of three purified enzymes: α-chymotrypsin, acid phosphatase, and carbonic anhydrase for use in the early assessment of toxicants at environmentally significant levels. Activities of α-chymotrypsin and acid phosphatase were measured spectrophotometrically, while carbonic anhydrase activities were determined through monitoring a pH change. The chemical agents investigated included several heavy metals, common herbicides and pesticides, and various environmentally significant anions. In addition, several techniques were explored to amplify enzyme response to chemical agents.

The results of the study demonstrated that α-chymotrypsin did not significantly respond to cadmium, nickel, or 2,4-D, and consequently, does not appear to be useful in indicating potential toxicity problems associated with these agents.

The acid phosphatase enzyme system appears to be useful in signaling the presence of low levels of certain anions (fluoride at 0.45 mg/ℓ and nitrate at 7.3 mg/ℓ), but does not appear to have potential for detecting toxic activity due to cadmium, nickel, or 2,4-D. Enzyme inhibition induced by fluoride at 37°C was not altered by changing the assay temperature to 50°C.

The results of the experiments with carbonic anhydrase show that the enzyme does not appear to be affected by cadmium or nickel ions. However, enzyme activity was inhibited by fluoride (4.5 mg/ℓ), sulfide (0.5 mg/ℓ), and nitrate (73 mg/ℓ). Enzyme inhibition was also induced by 10 mg/ℓ of atrazine, malathion, or carbaryl, and 150 to 500 mg/ℓ of 2,4-D. Inhibitory effects induced by sulfanilamide appeared to be slightly enhanced by the addition of Cd²⁺, Ni²⁺, or Zn²⁺ cations. These findings, although preliminary, suggest that carbonic anhydrase demonstrates potential for signaling the presence of anions, and appears to be useful in indicating potential toxicity problems due to pesticides and herbicides.