Date of Award
Doctor of Philosophy
Amidase (acylamide amidohydrolase, EC 18.104.22.168) was studied in soils becase its substrates, aliphatic amides, have potentials as synthetic N fertilizers. This enzyme catalyzes the hydrolysis of amides producing NH(,3) and their corresponding carboxylic acids. Amidase activity was detected in soils and a simple, sensitive, and precise method was developed for its assay. This method involves steam distillation of the NH(,4)('+) produced by amidase activity when soil is incubated with buffered (0.1 M THAM, pH 8.5) amide solution and toluene at 37(DEGREES)C;Studies of the kinetic parameters of the reaction catalyzed by amidase showed that, by using the Lineweaver-Burk plot, the K(,m) values of amidase in 8 soils, with formamide, acetamide, and propionamide as substrates, averaged 12.3, 4.6, and 14.5 mM, respectively. The temperature dependence of the rate constants conformed to the Arrhenius equation up to the point of enzyme inactivation (65(DEGREES)C). The activation energy values for soil amidase averaged 46.9, 50.0, and 26.5 kJ/mole using formamide, acetamide, and propionamide as substrates, respectively;Amidase activity in soils was found to be influenced by the methods of handling, storing, and pretreating the sample before the enzyme assay. The distribution of amidase activity in soil profiles decreased with sample depth. Statistical analyses showed that amidase activity was significantly correlated with urease activity (r = 0.73***), organic C (r = 0.74***), total N (r = 0.74***), and percentage clay (r = 0.69***) in 21 Iowa surface soils samples;Tests indicated that of the 21 trace elements studied, Ag(I), Hg(II), As(III), and Se(IV) were the most effective inhibitors of amidase ((GREATERTHEQ) 20% inhibition at 5 (mu)mole/g soil). The average inhibition of soil amidase by each of 16 pesticides added at the rate of 10 (mu)g/g of soil ranged from 2% with Dinitramine, Eradicane, and Merpan to 10% with Sutan;The transformations of N in 25 amides and their derivatives were studied and compared with those of N in (NH(,4))(,2)SO(,4) and urea added to field-moist soil samples and incubated under aerobic conditions at 30(DEGREES)C for 14 days. With the majority of the amides studied, the inorganic N produced accumulated as No(,3)('-). More than 40% of the amide-N was recovered as NH(,4)-N when thioacetamide, fluoroacetamide, and 2-chloroacetamide were added to soils. With one sandy soil, treatment of urea, formamide, N-benzylformamide, and p-nitrobenzamide showed accumulation of NO(,2)-N;Studies of the properties of amidase extracted from bacteria isolated from soils showed that it has lower values than soil amidase in the following properties: optimal pH (7.0 vs 8.5), optimal temperature (50 vs 60(DEGREES)C), K(,m) value (5.6 vs 12.3 mM), activation energy (18.9 vs 46.9 kJ/mole) and temperature coefficients (1.28 vs 1.75).
Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/
William Thomas Frankenberger, Jr.
Frankenberger, William Thomas Jr., "Amidase activity in soils " (1980). Retrospective Theses and Dissertations. 6722.