Determination of pholcodine in syrups and human plasma using the chemiluminescence system of tris(1,10 phenanthroline)ruthenium(II) and acidic Ce(IV)

Pholcodine is an opiate derivative drug which is widely used in pediatric medicine. In this study, a chemiluminescence (CL) method is described that determines pholcodine in human plasma and syrup samples. This method is based on the fact that pholcodine can greatly enhance the weak CL emission of reaction between tris(1,10 phenanthroline)ruthenium(II), Ru(phen)₃²⁺, and acidic Ce(IV). The CL mechanism is described in detail using UV–vis light, fluorescence and CL spectra. Effects of chemical variables were investigated and under optimum conditions, CL intensity was proportional to the pholcodine concentration over the range 4.0 × 10^?8 to 8.0 × 10^?6 mol  L^?1. The limit of detection (LOD) (S/N = 3) was 2.5 × 10^?8 mol  L^?1. Percent of relative standard deviations (%RSD) for 3.0 × 10^?7 and 3.0 × 10^?6 mol  L^?1 of pholcodine was 2.9 and 4.0%, respectively. Effects of common ingredients were investigated and the method was applied successfully to the determination of pholcodine in syrup samples and human plasma.     

Determination of thyroxine in pharmaceuticals using flow injection with luminol chemiluminescence inhibition detection.

A simple flow injection method is reported for the determination of thyroxine, based on its inhibition effect on luminol-iron(II) chemiluminescence in alkaline medium in the presence of molecular oxygen. The detection limits (2s) for d- and l-thyroxine are 0.08 and 0.1 mg/L, respectively, with a sample throughput of 100/h. The calibration data for d- and l-thyroxine over the range 0.2-1.0 mg/L gives correlation coefficients (r(2)) of 0.9915 and 0.984 with relative standard deviations (RSD; n = 4) in the range 1.2-2.8%. The effects of some organic compounds was studied on luminol-iron(II) CL system for thyroxine determination. The method was applied to pharmaceutical thyroxine tablets and the results obtained (in the range 50.5 +/- 2.0-51.6 +/- 1.2 microg l-thyroxine/tablet) were in reasonable agreement with the value quoted.     

A comparison of whole blood neutrophil chemiluminescence measured with cuvette and microtitre plate luminometers

Results obtained by measuring human whole blood neutrophil chemiluminescence (CL) using the BioOrbit 1251 cuvette luminometer and the Immunotech LM‐01T microtitre plate luminometer are compared in this study. Opsonized zymosan, phorbol myristate acetate, N‐formyl–Met–Leu–Phe and calcium ionophore A23187 were used as activators. The CL response of neutrophils to their stimulation with the individual types of activators tested was fully detectable using either type of the luminometers. The kinetic curves of CL activity obtained from both the cuvette and the microtitre plate luminometers had similar characteristics. The only insignificant difference observed when comparing the kinetic curves was in the rates of the CL reactions. The peak CL response of activated neutrophils was reached faster when using the luminometer BioOrbit 1251 than with the luminometer Immunotech LM‐01T. A likely reason for this difference is the mode of transporting samples during the measurement, inducing different degrees of agitation. However, although this fact needs to be considered when interpreting results, both types of luminometer can be fully utilized in both research and clinical laboratories. Copyright © 2002 John Wiley & Sons, Ltd.     

Destructive effect of anticancer oxali-palladium on heme degradation through the generation of endogenous hydrogen peroxide

The interaction between human hemoglobin (Hb) and oxali-palladium was studied using different spectroscopic methods of UV–vis, fluorescence, circular dichroism (CD), and chemiluminescence at two temperatures of 25 and 37°C. The experimental results showed that both dynamic and static quenching is occurred simultaneously when oxali-palladium quenches the fluorescence of Hb. According to the fluorescence quenching method, the binding site number, apparent binding constant, and corresponding thermodynamic parameters were measured at two temperatures. The values of ΔH°, ΔS°, and ΔG° indicate that process of the formation of oxali-palladium–Hb complex is a spontaneous interaction procedure in which electrostatic interaction plays a major role. In addition, UV–vis and CD results showed that the addition of oxali-palladium changes the conformation of Hb. To evaluate the functional changes of Hb via destruction of the heme structure, fluorescence studies were performed. The results demonstrated that two fluorescent heme degradation products are found during the interaction of oxali-palladium with Hb. Also, the amount of hydrogen peroxide produced in the solution of Hb due to the interaction of oxali-palladium with Hb using chemiluminescence method indicated heme degradation in the protein is occurred. Structural and functional changes induced in Hb via heme degradation are considered as side effects of this synthesized anticancer drug.     

Effect of sulphite on the oxidative metabolism of human neutrophils: Studies with lucigenin- and luminol-dependent chemiluminescence

To assess the effect of sulphite on the oxidative metabolism of human neutrophils, chemiluminescence (CL) measurements were performed using lucigenin and luminol as chemiluminigenic probes. Lucigenin-dependent CL was used for measuring superoxide anion (O) production, and luminol-dependent CL was used for determination of myeloperoxidase (MPO)-connected processes. With sulphite concentrations of 0.01 to 1 mmol/L, resting neutrophils showed an up to sixfold increase of lucigenin-dependent CL, but only a 1.9-fold increase of luminol-dependent CL. Subsequent stimulation of sulphite-treated neutrophils with phorbol myristate acetate (PMA) (soluble stimulant) or zymosan (particulate stimulant) resulted in an additional significant increase of lucigenin-dependent CL compared to stimulated control cells, whereas luminol-dependent CL increased slightly by 0.01 mmol/L sulphite and decreased then continuously. Sulphite concentrations above 1 mmol/L decreased both lucigenin- and luminol-dependent CL of resting and PMA- or zymosan-stimulated neutrophils. Lucigenin-dependent CL of sulphite-treated and subsequently stimulated neutrophils was strongly inhibited by extracellularly added superoxide dismutase, whereas luminol-dependent CL was markedly reduced by the MPO inhibitor azide. The intracellular activity of MPO in neutrophils stimulated with PMA in the presence of sulphite (2 mmol/L) was reduced by 55%. Sulphite (0.1 mmol/L) also inhibited strongly the activity of MPO in a cell-free system. These results indicate that micromolar concentrations of sulphite exert a stimulating effect on the O production of neutrophils extracellularly, but have an inhibitory effect on MPO-catalysed reactions intracellularly.     

Continuous Cultivation and Drug Susceptibility Testing of Plasmodium falciparum in a Malaria Endemic Area

ABSTRACT Isolates (UCH-23 and OM) and cloned strains of Plasmodium falciparum (Clones W-2 and D-6) were maintained in continuous culture for 28 to 150 days using culture media supplemented with 10% (v/v) heat inactivated semi-immune human plasma. Microscopic appearance and growth rates (R) of the parasites in media supplemented with semi-immune human plasma [R = 1.13 (W-2), 0.92 (D-6), 0.75 (OM) and 0.84 (UCH-23)] were comparable to those of parallel cultures maintained in media supplemented with 10% (v/v) heat inactivated non-immune human plasma [R = 1.42 (W-2), 0.83 (D-6), 0.66 (OM) and 0.89 (UCH-23)]. In addition, IC50 for chloroquine and mefloquine against the two cloned strains of P. falciparum maintained in culture media supplemented with either non-immune human plasma or semi-immune human plasma were identical. Although growth rates of new isolates (UCH-23 and OM) fluctuated over time, they stabilized between the 12th and 19th day of adaptation to culture. This fluctuation in growth rates of the new isolates underscores the influence of population dynamics during adaptation of P. falciparum to continuous culture. Sixty-eight percent of the primary isolates (170 of 250) obtained from patients in Ibadan were successfully adapted and maintained in continuous culture using semi-immune human plasma. The results of these studies indicate that semi-immune human plasma is a suitable supplement for continuous cultivation and drug susceptibility testing of P. falciparum. This finding will have practical implications in malaria endemic areas where difficulties in obtaining non-immune human plasma or serum limits establishment of continuous culture of P. falciparum and its application in studies on malaria.