Agriculture Reference
In-Depth Information
Dehydrogenase (DHA) activity typically occurs in all viable microbial cells (Gianfreda et
al., 2005). Thus, its measurement is usually related to the presence of viable microorganisms
and their oxidative ability has been often used as a functional indicator of soil health. DHA is
related to a group of intracellular enzymes that are present in active soil microorganisms. This
enzyme is found in all living organisms and takes part in many metabolic reactions involved
in oxidative energy transfer in microbes. As dehydrogenases are not active as extracellular
enzymes in the soil, the management of DHA has been used as a good overall indicator of
microbial activity and of the capacity of microbes to oxidize soil organic matter (Bolton et al.,
1985; Dick, 1997). Several studies have demonstrated that dehydrogenase enzyme activity of
microorganisms is among the most sensitive parameters for the evaluation of toxicity (Shen et
al., 2005).
In this chapter, dehydrogenase activity was determined using a method developed by
Casida et al. (1964) using as subtract triphenyltetrazolium chloride (TTC) and glucose. After
24 hours of incubation triphenylformazan formed (TPF) from TTC was extracted with
methanol. According to Obbard et al. (2001) substrate-induced DHA was found to be a
sensitive assay for determining metal effects on the physiologically active soil microbial
biomass. The precision of the assay allows evaluating, separately, effects due to pH,
contaminant concentration and other soil properties.
DHA activity varied significantly (P< 0.05) between control soil and BA and BD soils
(Figure 4), being the DHA activity in BC soil 2 times higher. This finding is in agreement
with (Trasar-Cepeda et al. 2000) who reported less dehydrogenase activity in metal-
contaminated soil than in a similar uncontaminated soil, and with Marzadori et al. (1996) who
stated that dehydrogenase activity was inhibited by the toxic effects of heavy metals,
particularly Pb. Dar (1996) also reported a decrease in dehydrogenase activity at 50 mg Cd
kg
-1
in a laboratory study.
DHA activity appears to depend on the type of pollutant; for example, it is high in soils
polluted with pulp and paper mill effluents and low in soils polluted with fly ash (Trasar-
Cepeda et al., 2000). Nevertheless, several studies have demonstrated that dehydrogenase
enzymatic activity of microorganisms is among the most sensitive parameters for evaluation
of toxicity (12), and that may be the reason why dehydrogenases activity are a frequently used
test for determining the influence of various pollutants (heavy metals, pesticide, crude oil,
etc.) on the microbiological quality of soil (Margesin et al., 2002).
The fluorescein diacetate hydrolysis (FDAH) assay measures the hydrolytic enzyme
activities of microbial populations and can provide an estimate of overall microbial activity in
an environmental sample (Adam and Duncan, 2001; Bandick et al., 1999; Burns, 1982). The
assay is considered non-specific because it is sensitive to the activity of several enzyme
classes including urease, lipases, phosphatases, glucosidase, esterases and proteases (Bandick
et al, 1999; Hayano and Tubakai, 1985). The enzymes responsible for Fluorescein diacetate
(FDA) hydrolysis are plentiful in the soil environment, and are involved in the decomposition
of many types of tissues (Adam and Duncan, 2001). The ability to hydrolyse FDA thus seems
widespread, especially among the major decomposers, bacteria and fungi. Generally more tan
90% of the energy flow in a soil system passes through microbial decomposers, therefore an
assay which measures microbial decomposing activity will provide a good estimate of total
microbial activity (Adam and Duncan, 2001).
