Desenvolvimento e aplicação de procedimentos envolvendo reações quimiluminescentes em fluxo para a determinação de analitos de interesse alimentício, farmacêutico e bioquímico.
Abstract
In this work, analytical flow procedures were developed using multicommutation
concept and chemiluminescent detection for the determination of hydrogen
peroxide, glucose and cholesterol in food, biochemistry and pharmaceutical
samples. A personal computer Pentium 233 MHz equipped with PCL 711S
interface was employed to control the flow manifold and data acquisition. The
detection module consist of a flow-cell constructed with a coiled polyethylene
tubing (0.8 mm i.d.), a silicon photodiode (OSD-50E) and an electronic device
(model FAC 500) for signals conditioning and amplification, are devices were
placed in a dark box. The manifold and detection module were characterized by
hydrogen peroxide determination in pharmaceutical samples, monitoring the
radiation produced in the chemiluminescent reaction between luminol, H2O2 and
hexacyanoferrate(III). The analytical curve obtained was linear in the hydrogen
peroxide concentration ranging from 2.2 x 10-6 to 2.1 x 10-4 mol L-4 and the
detection limit obtained was 1.8 x 10-6 mol L-1. In the procedures for glucose
determination in food, pharmaceutical and biochemical samples, a solid-phase
reactor containing glucose oxidase immobilized in glass beads was employed. The
hydrogen peroxide generated in this enzymatic reaction was monitored by
chemiluminescent emission produced by the luminol/hexacyanoferrate(III)
reaction. The proposed procedure showed linear response from 5.0 to 160.0 mg
L-1 and a sampling rate of 160 samples per hour was obtained. The analytical
procedure was employed with success in the glucose determination in honey,
glucose pharmaceutical formulation, grape and tomato juices. For glucose
determination in blood serum, the manifold was reconfigured to minimize the
sample manipulation. The results obtained in the glucose determination in serum
using this proposed procedure are in close agreement with those obtained using
the comparative procedure at a confidence level of 95%. In the proposed
procedure for cholesterol determination in eggs a solid-phase reactor containing
immobilized cholesterol oxidase was used, the hydrogen peroxide produced was
monitored by the same chemiluminescent reaction. The linear range of analytical
curve obtained in optimized procedure ranged from 250 to 2500 mg L-1, and are
described by the equation: Intensity (mV)= 43.0 + 0.325 x [cholesterol], r= 0.999,
The luminol and hexacyanoferrate(III) consumption per determination were 356 µg
and 2.64 mg, respectively. The manifold was reconfigured for cholesterol
determination in serum. The results obtained were in agreement with those
obtained using the comparative procedure. Extraction and pre-purification of
enzyme peroxidase were carried out with the purpose of employing this enzyme
as a catalyst in the chemiluminescent reaction between the luminol and hydrogen
peroxide.