Properties of acid phosphatase–tannic acid complexes formed in the presence of Fe and Mn
M.A. Rao, L. Gianfreda
Dipartimento di Scienze Chimico-Agrarie, Universita` di Napoli “Federico II”, via Universita` 100, 80055 Portici, Naples, Italy Accepted 15 May 2000
Abstract
The preparation and characterisation of synthetic enzyme complexes simulating those usually encountered in soil were investigated. Complexes were prepared at 30
8C by the interaction of acid phosphatase and tannic acid in the presence and absence of Fe and Mn ions as chlorides and oxides, with and without montmorillonite. In comparison with phosphatase–tannic acid complexes, the enzymatic complexes
formed in the presence of Fe oxide showed higher activity levels .70. Non significant increases were measured in the presence of Fe and
Mn ions and Mn oxide. In the presence of montmorillonite, a further increase in the activity of the resulting complexes was measured. For example, the activity of complexes formed with Fe
3 1
or Mn
2 1
increased by 25 and 28, respectively. In all cases, the kinetics of the immobilised enzyme conformed to the Michaelis–Menten equation and the V
max
values were significantly
P 0 :
001 lower and the K
m
values significantly higher than those of the free enzyme. For example, a two-fold reduction in the V
max
and four-fold increase in the K
m
parameters were measured for phosphatase complexed with tannic acid. In the presence of montmorillonite, no significant changes of V
max
values were measured whereas a detectable reduction of K
m
values was observed. Immobilised phosphatase showed a greater sensitivity to increases in temperature, but higher stability to proteolytic attack. Complexes with Fe ions and MnO
2
were the most stable. q 2000 Elsevier Science Ltd. All rights reserved.
Keywords: Soil-enzymes; Organo-mineral complexes; Inorganic catalysts; Fe and Mn oxides and ions
1. Introduction
In soil, the Fe and Mn compounds are involved in numer- ous processes, including adsorption andor transformation
Boyd and Mortland, 1990; Huang, 1990. An extensive literature is available concerning the role that Fe and Mn
compounds may play in soil as catalysts of processes invol- ving simple or complex aromatic compounds e.g. mono
and polyphenols, humic-like precursors Burns, 1986; Huang, 1990. Biotic catalysts, such as oxidoreductive
enzymes, may also promote similar processes Bollag, 1992. Depending on the nature, number and type of
aromatic compounds andor catalysts involved, the ultimate result of such processes is the formation of polymers andor
copolymers of various sizes and complexity, that contribute to the pool of humic materials Shindo and Huang, 1992;
Naidja et al., 1998.
Recently, Naidja et al. 1998 have demonstrated that the catechol –melanin complexes were formed whether the
polymerisation of a phenol such as catechol was catalysed by birnessite a d-manganese oxide or an oxidative enzyme
such as tyrosinase. However, the authors observed that the reaction products with a lower degree of aromatic ring
condensation and lower molecular mass were produced by the abiotic catalysis as compared with those generated in the
presence of tyrosinase.
Other molecules, such as proteins, may be involved in the polymerisation of phenolic compounds and polymeric
organic aggregates entrapping protein molecules may form Ladd and Butler, 1975. Adsorption of proteins on the exter-
nal surfaces of already formed organic aggregates may also occur Ladd and Butler, 1975. If the involved protein is an
enzyme, the enzymatic-copolymer complexes showing a cata- lytic activity may result Sarkar and Burns, 1983, 1984.
In the previous studies, we and others have demonstrated that active enzymatic complexes were generated by the
interaction of urease, invertase b-glucosidase and acid phosphatase with tannic acid, a humic-like precursor
Sarkar and Burns, 1984; Gianfreda et al., 1993, 1995a; Rao et al., 1996. The presence of montmorillo-
nite during the process usually produced organo-mineral complexes with higher enzymatic activity as compared
with those complexes obtained without clay.
Soil Biology Biochemistry 32 2000 1921–1926
0038-071700 - see front matter q 2000 Elsevier Science Ltd. All rights reserved. PII: S 0 0 3 8 - 0 7 1 7 0 0 0 0 1 6 7 - X
www.elsevier.comlocatesoilbio
Corresponding author. Tel.: 139-81-788-5225; fax: 139-81-775-5130.
E-mail address: mariaraounina.it M.A. Rao.
Further investigations showed that the addition of Fe and or Mn, as ions or oxides influenced significantly the
formation and the resulting enzymic properties of urease– tannic acid complexes Gianfreda et al., 1995a,b. The
purpose of the work reported here was to compare the ability of Fe and Mn as ions and oxides, to promote the formation
of complexes of tannic acid and acid phosphatase and to affect their catalytic properties. Acid phosphatase was
chosen as it is one of the most studied enzymes in soil because of its essential role in the phosphorus cycle Speir
and Ross, 1978. The influence of the clay montmorillonite on the enzymic properties of the various complexes was also
investigated.
2. Materials and methods
