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The relevance of considering the problems and prospects of biological control is associated with the need to form scientific, methodological and organizational foundations for the inclusion of biological control of industrial exposure to harmful chemicals in the system of social and hygienic monitoring to ensure the chemical safety of the Russian Federation. The purpose of the work is to substantiate the need to develop and implement methodological recommendations for the development and validation of chromatographic methods for biological control of industrial exposure to harmful substances with the system of sanitary and epidemiological regulation of the Russian Federation. The published results of works in the department of toxicology of the "Research Institute of Hygiene, Occupational Pathology and Human Ecology", carried out under the guidance of the authors, were used, and a number of new additions to the toxicokinetic equations were made, in particular, a method for calculating the material cumulation coefficient (Km.cum) was proposed for the first time.

Introduction. Currently, there are no standard methodological approaches that allow assessing the danger to human and animal reproduction of poorly soluble chemicals (industrial waste, products of destruction of organic substances, polymer materials), including substances that form complex poorly soluble systems with environmental objects. It may be impossible or ineffective to use standard routes of administration of such substances and complexes to assess their reproductive hazard. An attempt to find an experimental model for conducting toxicological and embryotoxicological studies that allow assessing the danger of prolonged interaction of poorly soluble compounds directly with tissue fluids was the main aim of the work. The study aims to investigate the possibility of using diffusion chambers to assess the reproductive hazard of poorly soluble chemicals. Material and methods. In experiments with chemicals, the researchers placed samples of dry preparations in the inner cavity of one of the rings. Experts sealed the ring on both sides with millipore membranes. The diffusion chambers obtained in this way were implanted into the abdominal cavity or under the skin in the cervical region of female rats during the most sensitive days of pregnancy to pathogenic effects. Results. When implanting diffusion chambers to animals, the possibility of assessing the embryotoxic properties of poorly soluble chemicals, such as bitumen-salt masses, polymer materials and coatings, etc., as well as soil contamination with substances potentially dangerous to the reproductive function of humans and animals is shown. With the help of diffusion chambers, experts assessed the embryotoxic activity of known dysmorphogens, cyclophosphamide, cadmium sulfate, as well as a number of polymer materials (di-paraxilylene film, acrylic acid copolymer, silloxane rubber), Scientists assessed soil contamination with a known insecticide and acaricide - endosulfan. Limitations. In the work, the experts used diffusion chambers with a volume of 400, 900 or 3000 m3. Conclusion. As a result of the experiments conducted, scientists determined that the presented method can be used to assess the reproductive hazard of poorly soluble chemicals (products of destruction of organic matter, polymer and endoprosthesis materials), when assessing the risk of prolonged presence of foreign bodies in tissues, when assessing soil contamination in places of chemical accidents, rocket crashes, storage and destruction of chemical weapons. Ethics. The study was carried out in compliance with the rules of bioethics approved by the European Convention for the Protection of Vertebrates Used for Experimental and Other Purposes (Protocol No. 4 of 8.11. 2022).

An approach for the preparation of a polymer delivery system consisting of sodium alginate (SA) and carbopol was developed. The efficiency of the inclusion of medicinal compounds in these systems was investigated using the polyphenols curcumin and resveratrol as examples. Curcumin and resveratrol possess anti-inflammatory, antitumor, and antibacterial effects, which makes them promising compounds for use in clinical practice. The main disadvantage of these polyphenols is their low bioavailability. The formation of microcapsules occurs due to the precipitation of a mixture of SA and carbopol (carbopol 940 or ETD 2020), containing a model object, in a calcium chloride solution. The dependence of the inclusion of drugs in microcapsules and their release into gastrointestinal environment imitators on the viscosity characteristics of carbopol, which is part of the microcapsule shell, was investigated.

The paper presents the results of an experimental study of the dose-dependent nature of functional changes in the body systems under chronic administration of uranyl acetate dihydrate in doses of 0.5 and 5.0 mg/kg per element for 18 weeks. The study was performed on 45 male outbred rats. It has been shown that uranyl acetate dihydrate in a dose of 0.5 mg/kg had no significant effect on hematological parameters. At the same time, activation of bactericidal activity of neutrophils, a decrease in the immunoregulatory index, and an increase in the blood concentration of tumor necrosis factor (TNF-α) have been revealed. The toxicant administered to rats in a dose of 5 mg/kg led to a decrease in the absolute number of erythrocytes, hemoglobin, hematocrit, platelets, the release of myelocytes into the blood, basophilia, monocytosis, the appearance of leukolysis cells and plasmatization of lymphocytes. On the part of the immune system, an increase in the biocidal capacity of neutrophilic granulocytes, TNF-α production, an increase in the number of CD8+ cells, and a reduction in the CD4+/CD8+ ratio have been found. Uranyl acetate dihydrate had a dose-dependent effect only on the number of cytotoxic T-lymphocytes, T-cells with the CD4+CD8+ phenotype, on the immunoregulatory index, and on the level of TNF-α. Hyperglycemia and glucosuria were also dose-dependent. An increase in glucose in the blood and urine indicated a violation of carbohydrate metabolism and kidney function. There was a decrease in the concentration of thyroxine, testosterone and an increase in the level of insulin. Uranyl acetate dihydrate led to the development of insulin resistance. The level of hormones did not depend on the dose of the toxicant administered to the animals.

The results of our own and foreign studies have shown the absence of sensitizing properties of terephthalic acid, and therefore there was a need to review the current hygienic standard – MAC – in the air of the working area and change the hazard class of the substance. The materials of the German Research Community (DFG) were used for substantiation of MAC. Recommended MAC in the air of working area is set at 5 mg/m3 (maximum single), a mixture of vapors and aerosol, hazard class 3.

A procedure for the determination of phosphonylated tyrosine, a marker of exposure to organophosphorus nerve agents, in dried blood plasma spot (DBS) samples by means of liquid chromatography/tandem high-resolution mass spectrometry was developed. The DBS samples on Protein Saver 903 Card substrates were shown to be stable for two months on storage at 22 and 40°C. It was found that the temperature of storage has a high impact on the stability of the DBS samples: the analyte concentration in the samples stored at 40°C was about three times lower than that in the samples stored at room temperature. It was shown that the special Protein Saver 903 Card substrates could be replaced by an ordinary blue ribbon filter paper. In the latter case, the tyrosine adducts of nerve agents were reliably determined in the DBS samples stored for a month at room temperature (22°C). However, for storage at elevated temperatures (40°C), the use of filter paper substrates is not recommended. The proposed approach to storage of blood samples allows one to take and easily transport large series of samples and automate their analysis.

Several hybrid systems for oral delivery of a therapeutic peptide have been prepared and investigated. Poly(l-glutamic acidco-d-phenylalanine) and silica sub-microparticles, porous CaCO3 microparticles and Lycopodium clavatum spore capsules were employed as the first level carriers included in sodium alginate granules as the second level carriers. Efficiency of the peptide encapsulation and release rate strongly depended on the nature and structure of the carriers.

Introduction. Russia annually produces about 400 thousand tons of Hazard Class I (extremely hazardous) and Hazard Class II (highly hazardous) wastes. Material and methods. The Program includes the study of the hazardous chemical pollution produced by the waste disposed at the Krasny Bor Landfill within a radius of 1-4 km over the period from 2009 to 2013. Samples of water, sediments, soil, and atmospheric air were analyzed by gas and liquid chromatography with highly sensitive detectors, as well as IR spectroscopy. Results. At all sampling sites, the air concentrations of priority pollutants characteristic of the waste disposed at the Landfill did not exceed the MAC level. Analysis of water and sediments from channels at the Landfill showed that the concentrations of a number of chemical pollutants at the border of the 1-km zone exceed the corresponding regulatory standards. Heavy pollution of wastewater and bottom sediments at the Landfill with some waste components, including organochlorine compounds, oil products, and heavy metals, as well as uneven pollution of soil in the 1-km zone from the Landfill in the northern and western directions, i.e. toward the potential residential development, with lead, copper, zinc, petroleum products, polychlorinated biphenyls, benzo[a]pyrene, and xylene isomers was revealed. Conclusion. The resulting data provides evidence for the adverse environmental impact of the Krasny Bor Landfill. The territory of the Landfill has been stabilized to allow further trouble-free development of a complex of measures on a scheduled basis.

Organophosphates’ toxic effect causes covalent binding to serine-198 in the active site of human plasma butyrylcholinesterase (BChE) with loss of enzymatic function (covalent inhibition). Mass spectrometric detection of modified FGESAGAAS peptide at the active site is a powerful exposure biomarker tool. The aim of this study was to develop mass spectrometry-based method for BChE adduct formation screening, avoiding the use of standard peptides. Immunomagnetic separation of proteins from plasma was optimized. Commercially available anti-butyrylcholinesterase monoclonal antibodies, immobilized on magnetic beads, resulted in stable and reusable affinity sorbent. The method was tested on horse serum BChE and real human plasma from healthy donors, treated with Russian VX (VR). The BChE isolated from blood plasma was digested with pepsin and analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). The method was evaluated by using synthetic peptides and by comparison to the enzymatic activity Ellman's assay. The minimum concentration of VR exposure, resulting in detectable VR-adduct, was 0.2 ng/mL, which corresponded to the relative BChE inhibition of less than 2%. Adduct formation assessment was performed via monitoring of decrease in non-modified peptide LC-MS/MS signal and increase in VR-modified peptide signal. The designed approach was tested in a pilot study with 5 blood samples from healthy volunteers. Mass spectrometry-based method for BChE adduct formation was found to be in agreement with Ellman's inhibition assay, so the method is applicable for direct BChE inhibition assessment.

Features of the electron and chemical ionization mass spectra of derivatives of N-[2-[(hydroxyethyl)thio]ethyl]-DL-valine (НЕТЕ-Val)—a biomarker of sulfur mustard—recorded by GC–MS and GC–MS/MS are studied. The retention indices of the studied derivatives were measured. An optimized procedure of the GC–MS/MS (MRM mode) identification of НЕТЕ-Val was tested in the analysis of donor blood samples in vitro exposed to sulfur mustard. The linear range of the detector was from 10 to 100 ng mL–1, and the limit of detection for sulfur mustard was 10 ng mL–1.

A unified procedure is proposed for the determination of thiodiglycol (TDG) and its oxide (TDGO) as markers of exposure to sulfur mustard in biomedical samples (urine, blood plasma, hairs) by gas chromatography–electron ionization mass spectrometry. The limit of detection of the procedure is comparable with the lowest level of endogenous TDG in biosamples (1 ng mL–1). Solid-phase microextraction of the derivatized analytes from the reaction mixture decreases the matrix effect. The procedure was tested in the analysis of rat biosamples obtained after in vivo exposure to sulfur mustard. The procedure was found to offer a higher potential in the retrospective determination of the total TDG and TDGO level in rat urine (within 7 days after exposure to 2 mg kg–1 of sulfur mustard) compared to the determination of the same markers in blood plasma. Blood plasma can be used for confirmatory analysis, provided the samples are taken within 2 days after exposure. Hair was found to be the least suitable biomatrix in view of the variability of the TDG background level in these samples and the possibility of contamination with TGD from other sources.

COVID-19 is a disease characterized by damage to the lower respiratory tract, development of the acute respiratory distress syndrome, in severe cases — multiple organ failure, including acute heart failure and cardiomyopathy. This study aimed to evaluate the effectiveness of the developed COVID-19 pathogenetic therapy and complications prevention agents using the ex vivo isolated lung and heart models. Isolated organs of white rats were used for the research; the dynamics of functional indicators were analyzed. An amino acid-peptide complex (APC) from a thermally treated milk protein hydrolyzate was used as the experimental COVID-19 pathogenetic therapy and complications prevention agent. Introduction of the APC to the isolated cardiopulmonary complex perfusate slowed down development of pulmonary edema in the experimental group; the organ's weight was 1.5 times less than in the control group (p = 0.0158). We have also registered an airway resistance downtrend. APC supported contractile activity of the isolated myocardium suffering ischemia-reperfusion: the growth of the left ventricular end diastolic pressure was 34% smaller than that registered in the control group (p < 0.05). The APC's cardioprotective effect relies on the endothelium-dependent mechanisms. The ex vivo method is highly informative. It allows assessing reactivity of the isolated organs exposed to biologically active substances and determining the possibilities of compensating for functional changes.

Two-level systems for oral delivery of therapeutic peptides were developed; the carriers consist of CaCO3 cores included into alginate granules. Such systems were first used for the delivery of low molecular weight drugs. It was shown that efficiency of encapsulation of peptides depends on their pI value, hydrophobicity, characteristics of the compounds used for doping CaCO3 cores, their surface potential and the techniques employed for loading peptides into the first-level carriers. Doping CaCO3 cores with dextran sulphate save their viability compared to the pristine CaCO3 cores, but ensures delivery of the desired quantity of peptide when using a smaller amount of delivery systems. Introducing the inhibitor of peptidases leads to an increase in the concentration of peptide in rat blood after intragastric administration of the developed delivery systems. Scanning electron microscopy and energy-dispersive X-ray spectroscopy demonstrated the presence of fragments of destructed first-level carriers in blood and plasma of experimental animals.

The title compound was synthesized as an internal standard for the quantitative determination of Arimistane in biological samples. [2H7] Arimistane was obtained from Arimistane-d0 by alkaline H/2H exchange of the С2,4,6,8,16 protons. The location of the labels was confirmed by NMR spectroscopy. The spatial structures were studied by X-ray diffraction.

Toxicity and hazard assessment of the mixture of saturated hydrocarbons C6-C10 (hexane, heptane, octane, nonane, decane)) at a single and chronic exposure was PERFORMED in animal experiments. It was shown that in terms of acute toxicity, the mixture is low hazard, exhibits moderate irritant effect on skin and eye mucous membrane and shows a slight dermal resorptive effect. The threshold limit concentration for a single inhalation exposure (Limac) as estimate4d by the resorptive effect is 5250±mg/m3 At a long 90 day continuous exposure, neuro-, hepato- and embryo toxicity was revealed at a concentration of 160mg/m3. The threshold limit concentration for chronic exposure (Limch) was estimated as 31.4 mg/ m3 and no effect concentration as 5.2 mg/m3. The threshold limit concentration of the mixtu re based on reflex action was estimated on a level of 280 mg/m3. Based on the investigations outcome, the mixture of saturated hydrocarbons C6-C10 is referred to hazard class 3. Maximum allowable concentrations (MACs) in the atmospheric air of residential areas were substantiated: MAC average daily based on resorptive effect equals to 5 mg/m3; MAC maximum single based on reflex action equals to 50 mg/m3.

Experimental studies of toxicity and hazard assessment of a mixture of С1-С5 saturated hydrocarbons as pollutants of atmospheric air were performed. It was shown that the mixture exhibits a low toxicity at a single exposure and does not irritate skin and eye mucous membranes. The Limac of the С1-С5 mixture was set at 25300 and 3100 mg/ m3 by the resorptive and reflex effects respectively. After 90-day chronic continuous exposure to the mixture, symptoms of chronic intoxication, specifically functional disorders in the nervous, cardiovascular, and respiratory systems and in biochemical processes, were revealed. The threshold concentration of the mixture was estimated at 255.7 mg/m3 . The experimental results were used to substantiate hygiene regulatory standards (MAC average day dose, and MAC maximum one-time dose) and hazard class for the mixture of С1-С5 saturated hydrocarbons in the atmospheric air of residential areas.

The effect of the Pelixim antidote on the possibility to detect markers of G type nerve agents sarin and soman was studies in bio probes obtained in an acute in vivo experiment after exposure of rats to organophosphorus nerve agents in doses of 0.5LD50. It was found out that the intake of equitoxic doses of soman and sarin leads to a decrease of acetylcholinesterase erythrocyte membrane (AChE) activity for up to 7 days after exposure. The effect of Pelixim on the recovery of erythrocyte AChE activity is mostly pronounced a day after sarin poisoning. The fluoride regeneration of nerve agents from blood plasma protein adducts was possible for up to 7 days after soman poisoning without antidotal therapy and for 3 days after with antidotal therapy; in case of sarin intoxication, fluoride regeneration was possible for 3 days regardless of the use of antidotal therapy . The antidote strongly affected the excretion of hydrolytic metabolite of sarin О-isopropyl methylphosphonic acid (IMPA) and had no effect on the excretion of hydrolytic metabolite of soman О-pinacolyl methylphosphonic acid (PMPA). A day after poisoning and Pelixim injection, IMPA was detected in urine at a level of 15.3 ng/ml, whereas its level in the urine samples of animals in the absence of antidotal therapy was 55.0 ng/ml; 3 days after poisoning, IMPA was detected at a level of 4.9 ng/ml exclusively in the urine of animals subjected to antidotal therapy. The urine levels of PMPA in animals subjected and not subjected to antidotal therapy were respectively 44 and 53 ng/ml a day after poisoning and 12 and 14 ng/ml respectively 3 days after poisoning. Thus, the antidote impact on the excretion profile of hydrolytic metabolites is more significant for sarin than that of soman.

The metabolic profile of plasma of white non-linear rats was investigated under normal conditions and after chronic inhalation exposure to low doses of aliphatic hydrocarbons (HC) with the number of carbon atoms from 6 to 10. The metabolic profile was determined by a combination of gas chromatography/mass spectrometry and high-performance liquid chromatography/high-resolution mass spectrometry with the subsequent use of chemometrical methods for data treatment and presentation. It was shown that continuous 90-day exposure to a mixture of C6–C10 saturated hydrocarbons at a concentration of 160 ± 20.5 mg/m3 leads to various metabolic impairments in the liver and kidneys. Exposure to hydrocarbon mixtures at doses of 31.4 ± 5.6 and 5.2 ± 1.8 mg/m3 evoked significantly smaller changes. Novel metabolic markers of the toxic effect of low concentrations of C6–C10 aliphatic hydrocarbons were revealed. The concentration ratio of pyrophosphoric and oxalic acids in rat blood plasma was found to be the most sensitive marker called a pyrophosphate index. A hypothesis was proposed about the disturbance of the redox balance as a leading pathogenetic mechanism of neuropathies and concomitant pathologies associated with chronic HC intoxication.

The potentials of different chromatography–mass spectrometry methods for the determination of alkyl methylphosphonic acids (AMPAs)—the chemical markers of nerve agents in urine—are compared. The gas chromatography–mass spectrometry (GC–MS) characteristics of various volatile AMPA derivatives are studied. The preference of perfluorobenzyl derivatives over methyl, trimethylsilyl and tert-butyldimethylsilyl esters for the GC–MS determination of AMPAs in urine is demonstrated. An optimal technique for the determination of AMPAs in urine is HPLC combined with high-resolution MS2 mass spectrometry with the isotope–labeled forms of target compounds as internal standards. The detection limits of AMPAs in the proposed analytical procedures vary from 0.1 to 1.0 ng/mL.

The metabolic profile of plasma of white non-linear rats was investigated under normal conditions and after chronic inhalational exposure to low doses of aliphatic hydrocarbons with the number of carbon atoms from 6 to 10. Metabolic profile was determined with combination of gas chromatography-mass spectrometry and high performance, high resolution liquid chromatography-mass spectrometry with subsequent use of chemometrical methods for data treatment and presentation. It was shown that continuous 90-day exposure to a mixture of C₆-C₁₀ saturated hydrocarbons at concentration of 160 ± 20.5 mg/m³ results in various impairments of metabolic processes in liver and kidneys. Exposure to hydrocarbons at doses of 31.4 ± 5.6 mg/m³ and 5.2 ± 1.8 mg/m³ evoked significantly smaller changes. Novel metabolic markers of the toxic effect of low concentrations of C₆-C₁₀ aliphatic hydrocarbons were revealed. The ratio of concentrations of pyrophosphoric and oxalic acids in rat blood plasma was found to be the most sensitive marker calledpyrophosphate index. A hypothesis is put forward about the redox balance violation as the leading pathogenetic mechanism of neuropathies and concomitant pathologies associated with hydrocarbon chronic intoxication.

A comparative study of the efficiency of procedures for the determination of the biomarkers of organophosphorus agents (OPAs) in blood plasma was performed. It was found that the gas chromatography–mass spectrometry determination of OPAs reactivated from the composition of protein adducts is a rapid method for the detection of exposure to OPAs. The liquid chromatography–mass spectrometry determination of phosphonylated butyrylcholinesterase and albumin fragments modified with OPA residues provides an opportunity to perform more sensitive and retrospective analysis. The tyrosine adducts of OPAs with serum albumin and other blood plasma proteins are not prone to dealkylation in the course of aging; in the series of the test markers, they possess the greatest diagnostic value because they make it possible to determine the precise structure a toxic agent after the longest time interval after exposure. The tentative limit of detection of OPA markers varies from 0.1 to 1.0 ng/mL.

The His-Phe-Arg-Trp-Pro-Gly-Pro [ACTH-(6–9)-PGP] peptide was synthesized. Proteolysis of ACTH-(6–9)-PGP and semax (Met-Glu-His-Phe-Pro-Gly-Pro) in the presence of leucine aminopeptidase and carboxypeptidase Y was studied. If the proteolysis of ACTH-(6–9)-PGP is mainly defined by aminopeptidases, the basic metabolite is Trp-Pro-Gly-Pro. Identification of major metabolites of ACTH-(6–9)PGP when incubated with the enzymes in vitro made it possible to evaluate proteolysis pathways of the peptide and prepare necessary amounts of its metabolites for using them as standards. Kinetics of ACTH-(6–9)PGP degradation in the presence of enzyme systems of nasal mucus, blood, and plasma were also explored. It was found that proteolysis of ACTH-(6–9)-PGP in rat blood and plasma occurs mainly under the effect of enzymes whose action is similar to leucine aminopeptidase.

Molecular aspects of the biological mechanism of action of the synthetic peptide HFRWPGP, including an assessment of the stability to membrane proteolytic enzymes and the determination of several intermolecular interaction parameters of the peptide and rat-brain cytoplasmic membranes. It was hypothesized based on the results that the peptide apparently acts as an allosteric activity modulator for several receptor systems and can after the nature of signal transfer to cells. It was established that heptapeptide HFRWPGP in the presence of cerebellum and hippocampus cytoplasmic membranes was degraded in a manner similar to that of Semax proteolysis (used as a reference peptide) under the same incubation conditions.

We have studied the effect of Р-endorphin (an endogenous opioid peptide) on the functional characteristics of isolated rat heart. In physiological concentrations (6.3 X 10-10 M) Р-endorphin induces a negative inotropic effect, accompanied by a slight decrease in the heart rate, and has no effect on the perfusion pressure of the coronary vessels. It is shown that the effect of Р-endorphin is mediated by 5-opioid receptors. ATP-dependent K+-channels contribute the greatest contribution to the implementation of the effect of Р-endorphin on the isolated heart of a rat. The role of voltage-dependent K+-channels and NO-mediated mechanisms in the implementation of the effects of Р-endorphin is minor.

When metal catalysts of the same group are used, the nature of the inorganic support with the surface-deposited peptide becomes the crucial factor determining the yield and hydrogen isotope content of 5-oxo-Pro-His-Pro-NH2. The label distribution in the amino acid residues of 5-oxo-Pro-His-Pro-NH2 was found. The label was shown to be mainly incorporated in His. The synthesis in the presence of deuterium or tritium afforded labeled specimens containing about 3 deuterium atoms or 2.5 tritium atoms (about 70 Ci/mmol) per peptide molecule.

The purpose of research is to learn the influence of β-endorphin on the functional activity indicators of the isolated heart of rat and to evaluate the role of opioid receptors. Materials and methods. The research was performed on isolated hearts of males nonlinear white rats. Perfusion of the heart was performed on the Langendorff System at constant flow. Register indicators are volume rate of coronary perfusion, perfusion pressure, the pressure in the left ventricle, pulse pressure, heart rate, speed of changing the pressure developed by the left ventricle. Additionally was calculated integral indicator minute performance of the left ventricle corresponding to the area under the curve of change of pressure in the left ventricle per a minute, index of contractile function and the integral indicator of the average performance of the left ventricle per one contraction. Results and discussion. It was identified depressant effect of β-endorphin in slightly declining trend to decrease of myocardial contractility indicators in a concentration range from 6.3x10 -10 to 6.3x10 -18 M. In the presence of a blocker of δ-opioid receptor naltrindole inhibitory effect of β-endorphin identified in isolated use, practically was not manifested. Thus, the peripheral δ-opioid receptors of cardiomyocytes mediate the decrease of myocardial contractility caused by β-endorphin using. Conclusions. β-endorphin in physiological concentrations has a depressing effect on functional parameters of the isolated heart of rat. Stimulation of δ-opioid receptors is one of the mechanisms leading to formation of the negative inotropic effect of β-endorphin as a component of local cardiac activity regulation.

Deuterium- and tritium-labeled His–Phe–Arg–Trp–Pro–Gly–Pro [ACTH(6–9)PGP] samples were prepared. The formation of isotopomers depending on the reaction conditions was monitored by mass spectrometry. The isotope exchange efficiency strongly depends not only on the ratio of the substance amount to the amount of hydrogen isotopes, on the nature of the catalyst and support, and on temperature, but also on the procedure for preparing the same support. Tritium-labeled ACTH(6–9)PGP with the radiochemical purity of no less than 97–98% was prepared at 200°С on 5% Pd/Al2O3 in 30–40% yield with the molar radioactivity of 26 Ci mmol–1. Deuterium-labeled ACTH(6–9)PGP with 97–98% content of the main substance was prepared in 67% yield and contained, on the average, 2.9 deuterium atoms per peptide molecule.

A method is proposed for the extractive alkylation of free fatty acids (FFA) in blood plasma and urine for their subsequent chromatographic–mass spectrometric determination as methyl esters. The list of target compounds includes 19 saturated acids (C6–C24) and 20 unsaturated acids (C16–C24), including ω-3, ω-6, ω-9, and polyunsaturated fatty acids. For the simultaneous extraction of FFA from a matrix and derivatization, extractive alkylation with iodomethane was used. The peculiarity and advantage of the method is integration of the stages of extraction and preparation of volatile derivatives under mild conditions. The recovery and conversion are from 60 to 90%. The analytical range was from 0.02 to 20 µg/mL of a sample. The effect of samples storage conditions on the results of measurements is evaluated. The method was tested in the determination of FFA in blood plasma and urine of volunteers and laboratory animals.

A GC-MS method was developed for quantification of 32 free and esterified fatty acids: 12 saturated fatty acids (C 11 -C 24 ) and 20 unsaturated and polyunsaturated fatty acids (C 16 -C 24 ). GC-MS quantification was performed in two stages. At first, esterified fatty acids were converted to methyl esters by transesterification with KOH in MeOH. At the second stage free fatty acids were converted to methyl esters by extractive alkylation with methyl iodide in CH 2 Cl 2 and tetrabutylammonium hydroxide. Relative calibration was performed with deuterated palmitic acid and methyl palmitate as internal standards. Range of measured concentrations was 0.2 to 800 mkg/ml. Relative standard deviation was below 15%. The soft conditions for preparing the samples tested lead to the absence of or minor transesterification side reactions, oxidation and hydrolysis. It is proved that during the extraction procedure esterified fatty acid does not increase the concentration of free fatty acids, indicating the absence of conversion of esterified to free fatty acids. The technique was tested in the study of fatty acids of blood plasma of human volunteers. Key words : blood plasma, free fatty acids, esterified fatty acids, extractive alkylation, GC-MS (Russian) DOI: http://dx.doi.org/10.15826/analitika.2015.19.2.002 T.I. Orlova, A.I. Ukolov, E.I. Savel’eva and A.S. Radilov Research Institute of Hygiene, Occupational Pathology and Human Ecology Federal State Unitary Enterprise, Federal Medical Biological Agency of Russia Kapitolovo Station, g/p Kuz’molovsky, Vsevolozhsky District, 188663 Leningrad Region, Russian Federation

Review substantiates the key role of gas chromatography mass spectrometry in determining organophosphorus chemical warfare agents and products of their transformations in the environment. Modern methods used in sample preparation for analysis, solid-phase and liquid extraction techniques, are presented. The problem of identification of organophosphorus chemical agents and their decomposition products and precursors in complex matrices in the untargeted analysis mode is considered.

Paraoxonase-1 (PON1) and butyrylcholinesterase (BCHE) are natural bioscavengers of organophosphate acetylcholinesterase inhibitors in the human body, which can determine individual sensitivity to organophosphate toxicity. Interindividual differences in activity of PON1 (catalytic bioscavenger) and substrate specificity are strongly associated with the substitution of two amino acids: Leu/Met (L/M) at position 55 (rs854560) and Gln/Arg (Q/R) at position 192 (rs662). In the case of BCHE (stoichiometric bioscavenger) substitution, Ala/Thr (A/T) at position 539 produces the so-called “K-variant” of the enzyme (rs1803274). Threonine allele is often co-inherited with an atypical BCHE allele (rs1799807). The atypical variant of BCHE displays a lower affinity for cholinesterase inhibitors. Genotyping rs662 and rs1803274 single-nucleotide polymorphisms (SNP) by high-resolution melting (HRM) is facilitated by the nucleotide substitution A>G (G>A), which resulted in a changed number of hydrogen bonds in the PCR product and, consequently, shifted T m. In the case of rs854560, genotyping is complicated by the nucleotide substitution T>A, which has no significant effect on the T m of the PCR product. An addition of a small quantity of LL homozygote DNA into the reaction mixture before PCR discriminates the three genotypes by the melt curves due to different amounts of heteroduplexes formed in the LM and MM samples. HRM analysis can be applied for genotyping human rs854560, rs662, and rs1803274 SNPs.

Mitochondrial aconitase has been shown to be inactivated under the effects of many compounds and critical states. Fluoroacetate (FA) is the best-known aconitase-inhibiting toxic agent. The biochemistry of the toxic action of FA has been rather well studied; however, no effective therapy has been developed over the past six decades. To search for new approaches to the development of possible antidotes, experiments were carried out in vitro with rat liver mitochondria, Ehrlich ascite tumor (EAT) cells, and cardiomyocytes exposed to FA or fluorocitrate (FC). FA produced its effects at much higher concentrations as compared with FC; in experiments with mitochondria these effects depended on respiratory substrates: with pyruvate, FA induced a slow oxidation and/or a leak of pyridine nucleotides and inhibition of respiration. Oxidation of pyridine nucleotides (PN) was prevented by the incubation of mitochondria with cyclosporin A. Studies of the PN level and dynamics of Ca2+ in EAT cells during activation by ATP also revealed the PN leak from mitochondria, which led to a shift in the balance of mitochondrial and cytosolic NAD(P)H under action of FA. Moreover, an increase of cytosolic Ca2+ was revealed in the cells exposed to FA, which could be explained by the activation of plasma membrane calcium channels. This mechanism could affect the amplitude and rate of calcium waves in cardiomyocytes under the effects of FA. We emphasize the reciprocal relationship between intracellular PN dynamics and calcium balance and discuss possible pathways of metabolic modulation in the context of development of effective therapy of poisoning with FA and other aconitase inhibitors.

The maximum amounts of the thyroliberin in the blood and brain of rats at intranasal and intravenous administration were determined. It is found that rat hippocampal, cortical, and cerebellar membranes contain two types of specific binding sites (high- and low-affinity) for the labeled ligand. It was shown that, at intranasal and intravenous administration, maximum amounts of the thyroliberin were detected in the cerebellum and then in the cortex and hippocampus. The degradation of the thyroliberin in the rat brain and its regions at intranasal and intravenous administration was studied. It is shown that the degree of degradation and the formation of proteolytic products of the thyroliberin is different in different regions of the rat brain.

Presented in this chapter are data on the principal hydrolysis products of Russian VX (RVX) with excess and equimolar quantities of water, as well as a generalized list of compounds extracted from samples of construction materials of a former RVX production facility. Toxicokinetic experiments have been conducted to determine the decrease of O-alkyl methylphosphonates in blood plasma of rats after intramuscular (i.m.) administration of RVX. Tandem mass spectrometry (MS/MS) peptide spectra obtained with commercially available human albumin incubated with RVX revealed covalent binding of RVX to Tyr411 and Tyr150. A new covalent adduct of N,N-diethyl-2-aminoethanthiol (an RVX hydrolysis product) with blood plasma proteins has been detected by high-performance liquid chromatography with mass spectrometry (HPLC-MS/MS), and this can be another marker of exposure to RVX. New data on effects developed under chronic exposure of rats to RVX are presented, including kinetic parameters of blood platelet aggregation and electrophysiology of Nervus tibialis. According to these data, morphofunctional changes at the level of the microcirculatory bed may prove to be a significant factor in the etiology of delayed effects.

Fluoroacetate (FA; CH2FCOOR) is highly toxic towards humans and other mammals through inhibition of the enzyme aconitase in the tricarboxylic acid cycle, caused by 'lethal synthesis' of an isomer of fluorocitrate (FC). FA is found in a range of plant species and their ingestion can cause the death of ruminant animals. Some fluorinated compounds -- used as anticancer agents, narcotic analgesics, pesticides or industrial chemicals -- metabolize to FA as intermediate products. The chemical characteristics of FA and the clinical signs of intoxication warrant the re-evaluation of the toxic danger of FA and renewed efforts in the search for effective therapeutic means. Antidotal therapy for FA intoxication has been aimed at preventing fluorocitrate synthesis and aconitase blockade in mitochondria, and at providing citrate outflow from this organelle. Despite a greatly improved understanding of the biochemical mechanism of FA toxicity, ethanol, if taken immediately after the poisoning, has been the most acceptable antidote for the past six decades. This review deals with the clinical signs and physiological and biochemical mechanisms of FA intoxication to provide an explanation of why, even after decades of investigation, has no effective therapy to FA intoxication been elaborated. An apparent lack of integrated toxicological studies is viewed as a limiter of progress in this regard. Two principal ways of developing effective therapies for FA intoxication are considered. Firstly, competitive inhibition of FA interaction with CoA and of FC interaction with aconitase. Secondly, channeling the alternative metabolic pathways by orienting the fate of citrate via cytosolic aconitase, and by maintaining the flux of reducing equivalents into the TCA cycle via glutamate dehydrogenase.

Products of the chemical neutralization of O-isobutyl S-(2-diethylaminoethyl) methylthiophosphonate, a VX Group warfare agent, in bitumen–salt matrices were studied by gas chromatography–mass spectrometry. Forty-four compounds were characterized by their electron impact ionization mass spectra and gas chromatographic retention indices on standard nonpolar polydimethylsiloxane phases; seven of these compounds were determined as derivatives. The structures of 26 compounds were identified, including structures of 24 O-isobutyl S-(2-diethylaminoethyl) methylthiophosphonate neutralization products. To solve this problem, chromatographic retention indices of unknown compounds were evaluated using reference data for their structural analogues. Partition coefficients in the binary heterophase hexane–octanol system were determined for the major components.

The feasibility of isolation and quantitative determination by gas chromatography of methylphosphonic acid and its mono- and dialkyl etsers, which originate from destruction of organophosphorus chemical warfare agents and have been detected in bitumen-salt compounds is examined.

Basic provisions of the medical and sanitary passport of chemically dangerous object and the territory adjoining to it are presented in article. Need of development of the medical and sanitary passport for systematization of sanitary and epidemiologic data with the purpose of a complex assessment of health of the population and the personnel working at chemically dangerous objects, harmful factors production and environment taking into account emissions of polluting substances of chemically dangerous objects is shown.

The main provisions concerning the need for a creation of medical and sanitary passport for territories in the protective measures zones (PMZ) of storage facilities and facilities for the destruction of chemical weapons (FDCW) are presented in the article. The assessment of results of certification of the territory of PMZ and FDCW in "Maradykovsky" in the Kirov region has been done.

The analysis of the data available in the literature has shown that nanoparticles (NP) have a high toxicity than usual microparticles, can penetrate unchanged across the cell barriers, the blood-brain barrier into the central nervous system, circulate and accumulate in the organs and tissues, by inducing more significant pathomorphological lesions in the visceral organs, and have a long elimination half-life. The toxicity of HPs is determined by their shape and sizes; moreover, minute fusiform particles cause more destructive effects than similar spherical particles; upon exposure, there is a clear dose-effect relationship. According to exposure to NPs, the classical target organs for the latter are the lung, liver, kidney, brain, gastrointestinal tract.

Mass spectrum analysis revealed differences in general contents of low-molecular peptides spectrums in chemical weapons extermination object staffers, in comparison with the reference group. Findings are that serum paraoxonase activity in chemical weapons extermination object staffers in significantly increased.

Publisher Summary This chapter describes the chemistry, analysis, toxicity, monitoring, and regulatory hygiene, and therapy of Russian VX nerve agent. One of the most abundant and most toxic chemical warfare agents in the chemical arsenals of the USA and Russia is VX and Russian VX respectively, whose development in the middle of the 20th century signified the peak of warfare chemistry. The arbitrary name VX relates to a group of O, S-diesters of methylphosphonic acid ROPO(CH3)S(CH2)2N(R1)2. V-gases are low-volatile liquids with high boiling points and, therefore, they are much more persistent than higher volatility organophosphorus (OP) agents of the G-series, such as sarin, soman, or tabun. V-series compounds are more toxic than OP nerve agents of the G-series. Poisoning occurs irrespective of route of exposure; specifically inhalation, ingestion of vaporous and liquid agents through intact or injured skin or eye mucosa, and on contact with contaminated surfaces. The most effective antidote complex for treating acute intoxications with RVX consists of an antagonist of M-cholinoceptors, a reversible inhibitor of cholinesterase, and a reactivator of cholinesterase. In addition, anticonvulsants can be used in cases where convulsions occur.