ABSTRACT
Prototype technologies of a bioluminescent signal system (BSS) based on the luminous bacterium Photobacterium phosphoreum and three enzymatic bioluminescence systems have been proposed for detecting and signalling the presence of toxicants in water systems. A number of pesticides, mostly known as poisonous substances, similar in their structures and physicochemical properties, have been taken as model compounds of chemical agents. The effect of toxicants (organophosphates, derivatives of dithiocarbamide acid, and pyrethroid preparations) on the bioluminescence of the four systems has been analysed. EC(50) and EC(80) have been determined and compared to the maximum permissible concentration for each of the analysed substances. The triple-enzyme systems with ADH and trypsin have been shown to be more sensitive to organophosphorous compounds (0.13-11 mg/L), while the triple-enzyme system with trypsin is highly sensitive to lipotropic poison, a derivative of dithiocarbamine acid (0.03 mg/L). Sensitivities of the triple-enzyme systems to pyrethroid preparations are similar to those of luminous bacteria (0.9-5 mg/L). The results can be used to construct an alarm-test bioluminescence system for detecting chemical toxicants, based on intact bacteria or enzyme systems.
Subject(s)
Chemical Warfare Agents/analysis , Water Pollutants, Chemical/analysis , Biological Assay , Chemical Warfare Agents/toxicity , FMN Reductase/metabolism , Luciferases/metabolism , Luminescence , Water Pollutants, Chemical/toxicityABSTRACT
The Ca(2+)-regulated photoprotein obelin has been examined as a label for bioluminescence immunoassay of infective agents. The hepatitis B virus (HbsAg) and the bacteria Escherichia coli and Shigella sonnei lipopolysaccharide (LPS) were chosen as model antigens. Chemically synthesized obelin-corresponding antibody conjugates were used in a solid-phase microplate immunoassay. The sensitivities achieved by the assay were 0.25 ng/mL for S. sonnei LPS and 0.375 ng/mL for HbsAg. A novel, filter-based immunoassay to determine bacterial admixtures in the environment was proposed. The NanoCeram filters were effectively applied to 'trap' and pre-concentrate pathogens from samples under study for the purposes of further detection and measurement of the absorbed material by bioluminescence immunoassay.
Subject(s)
Escherichia coli/isolation & purification , Hepatitis B virus/isolation & purification , Immunoassay/methods , Luminescence , Shigella sonnei/isolation & purification , Signal Transduction , Antigens, Bacterial/isolation & purification , Escherichia coli/immunology , Escherichia coli/pathogenicity , Hepatitis B Surface Antigens/isolation & purification , Hepatitis B virus/immunology , Hepatitis B virus/pathogenicity , Lipopolysaccharides/isolation & purification , Shigella sonnei/immunology , Shigella sonnei/pathogenicityABSTRACT
The integral bioluminescent biotest with lyophilized fluorescent bacteria was used for monitoring of LPO processes in tissue extracts and serum of rats exposed to stress. A relationship between the content of MDA (LPO indicator) and fluorescence of bacteria was observed in all biological samples.
Subject(s)
Biological Assay/methods , Luminescent Measurements , Oxidative Stress , Animals , Bacteria/metabolism , Lipid Peroxidation , Malondialdehyde/metabolism , Photobacterium/chemistry , Rats , Tissue Extracts/chemistry , Tissue Extracts/metabolismABSTRACT
The study addressed the effects of redox-active compounds on trypsin activity. Series of organic oxidizers (quinones) and reducers (phenols) were chosen as model redox-active compounds. Trypsin activity was quantified by bioluminescent technique. Interactions of these compounds with trypsin were studied by fluorescent and light absorption methods. Luminescence intensity decay constants in the reduced nicotinamidadeninedinucleotide (NADH): flavinmononucleotide (FMN)-oxidoreductase (R)-luciferase (L)-trypsin (T) (R + L + T) triple-enzyme system were calculated and compared in the presence of different concentrations of quinones and phenols. The triple-enzyme system was shown to be sensitive to quinones and not sensitive to phenols. It has been found that the effects produced by quinones on the coupled enzyme system (R + L) and on the trypsin molecule (T) are not related. The conclusions were extrapolated to the properties of other proteases and antiproteases.
Subject(s)
Environmental Monitoring/methods , Phenols/analysis , Quinones/analysis , Endopeptidases/metabolism , FMN Reductase/metabolism , Fluorescence Polarization , Luciferases/metabolism , Luminescence , Oxidation-Reduction , Trypsin/metabolismABSTRACT
We studied the possibility of evaluation of the intensity of pathological oxidative processes in rat liver using an integral bioluminescent test with controlled perfusion of the isolated organ. The test revealed a significant correlation between the level of TBA-reactive products and bioluminescence intensity.
