Biomedical Engineering Reference
In-Depth Information
Product
Substrate
Reaction time
Figure 7.2
Changes in the substrate concentration
vs.
reaction time.
colorimetric or isotopic labelling procedures. h e choice of the proce-
dure is determined, identii ed and depicted by the nature of the substrate
or the product. For example, spectrophotometric assays are used when
the substrate or product was absorbing light of some specii c wavelength.
Fluoresecence or radioactive measurements determines the highly sensitive
assayy procedure.
In many instances substates and casese modii ed substrateby introduc-
ing specii c groups for the estimationed of their products. Such substrate
modii cation should not, however, result in the decrease in the rate of its
transformation to product by the enzyme. For instance the introduction of
p-nitrophenyl phosphate compound in substrate adds for making the sub-
strate useful for the assay of phosphatases. h e product of enzyme action
is an intence, ionised form having p-nitrophenylate ion which is of yellow
colour and the intensity of color indicates enormously the activity of the
enzyme assay.
Enzyme concentrations is generally expressed as activity per unit vol-
ume and must specify the temperature at which activity is measured. h e
simple enzyme-catalyzed reactions consist of at least three stages, which
are temperature dependent i.e the formation of the enzyme-substrate com-
plex, the conversion of this complex to the enzyme-product complex, and
the dissociation of the enzyme-product complex.
A combination of the ef ects produced at each stage is the overall ef ect
of temperature on reaction rates is. In general,the increase in T with 10
C
will leads to double the rate of an enzymatic reaction.To ensure the repro-
ducible measurement of reaction rates the temperature control at 0.1
°
°
C is
