[Mechanism of the "acetylene effect" during interaction of organophosphorus compounds with cholinesterases]. / O mekhanizme "atsetilenovogo éffecta" pri vzaimodeistvii fosfororganicheskikh soedinenii s kholinésterazami.

Here we present data on the anticholinesterase activity of 58 synthesized ethers of phosphorus thioacids with an acetylene bond in the thioether group. Anticholinesterase activity of the compounds, with acetylene group in beta and especially alpha position, in the thioether radical is many times that of their saturated analogs. Reaction between the enzymes and acetylene organophosphorous inhibitors, as well as their saturated analogs, results in phosphorylated enzyme. The triple bond plays a significant role in the acceleration of cholinesterases phosphorylation. Antienzyme activity of acetylene organophosphorous inhibitors is discussed.

[Biochemical characteristics of aminostigmine--a new anticholinesterase agent]. / Biokhimicheskaia kharakteristika aminostigmina--novogo lekarstvennogo antikholinésteraznogo sredstva.

The properties of aminostigmine in comparison with those of other carbamate inhibitors of cholinesterases have been studied in vitro using potentiometric titration and Ellman methods. The bimolecular constants of the inhibition rate of acetyl-, butyryl- and propionylcholinesterase were found to be equal to (8.0-14.0).10(5) (3.8-7.7).10(5) and 11.0.10(5) M-1.min-1, respectively. In terms of inhibitory activity, aminostrigmine is comparable to neostigmine methylsulphate, being inferior to physostigmine and superior to pyridistigmine. The rate of decarbamylation of acetylcholinesterase inhibited by aminostigmine measured by the dilution method, by creating excessive acetylcholine and by dialysis is characterized by k2c constants equal to (1.1-1.6).10(-2), (2.5-2.8).10(-2) and 0.025.10(-2) min-1, respectively. On the whole, aminostigmine belongs to slowly reversible inhibitors. Being carbamylated by aminostigmine, the enzyme is resistant to reactivation by TMB-4 and HI-6. At (4-6).10(-7) M aminostigmine prevents by 50% the irreversible binding of cholinesterase by certain organophosphate inhibitors of cholinesterase when the latter are used at concentrations needed to inhibit the enzymatic activity by 85-90%.

[A comparative study of carboxylesterase in the white mouse and in the caterpillar of the cotton bollworm Heliothes armigera]. / Sravnitel'noe issledovanie karboksilésterazy beloi myshi i gusenitsy khlopkovoi sovki Heliothis armigera.

Studies have been made on enzymic hydrolysis of p-nitrophenylacetate (p-NPhAc), n-nitrophenylbutyrate (p-NPhBu) and indophenylacetate (IPhAc) by carboxylesterase (CE) from mouse blood plasma and liver as well as from caterpillar of the cotton worm haemolymph, intestine and fat body. Different KM and V max values were obtained for CE from these sources. The highest specific activity of CE from mouse liver and caterpillar intestine and fat body was observed with p-NPhBu. p-NPhAc is the best substrate for CE from mouse blood serum and caterpillar haemolymph. Carboxylesterases from mouse and caterpillar differed in their sensitivity to armine and paraoxone by 1-2 orders depending on the substrate used. Species and tissue differences in the kinetics of CE-catalyzed reactions with different substrates and inhibitors were revealed.

[The mechanism of anticholinesterase action of acetylene organophosphorus inhibitors]. / O mekhanizme antikholinésteraznogo deistviia atsetilenovykh fosforororganicheskikh ingibitorov.

Introduction of the triple bond in the leaving group of the organophosphorus inhibitor molecule gives a sharp raise of the inhibitor activity but does not change principal characteristics of the cholinesterase inhibition mechanism. The reactivation experiments suggest that inactivation of cholinesterases by these compounds occurs due to phosphorylating of the serine hydroxyl by the corresponding phosphoric acid. A close similarity was shown between acetylenic and saturated organophosphorus inhibitors in altering ka upon change of pH and tetraalkylammonium ions action. It is demonstrated that S-alkynyl esters of thioacetic acid are slowly hydrolyzed by acetylcholinesterase and cholinesterase without irreversible inhibition of the enzymes.

[The effect of pH and reversible inhibitors on decarbamylation of acetylcholinesterase]. / Vliianie pH i obratimykh ingibitorov na dekarbamilirovanie atsetilkholinésterazy.

Decarbamylation rate of membrane-bound methyl- and dimethyl-carbamylated acetylcholinesterase of human erythrocytes and bovine brain is reliably 1.1-1.6 times lower than that of the soluble enzyme. Such reversible inhibitors as tacrine (of non-competition action), ambenonium (mixed action) and galanthamine (competitive type of action) decelerate the decarbamylation rate of acetylcholinesterase. At pH 6 tacrine inhibits the reduction rate of soluble acetylcholinesterase activity of human erythrocytes more intensively than that of membrane-bound acetylcholinesterase. No differences in decarbamylation rate were found for the both forms of the enzyme at pH 8. Tacrine, a non-competitive inhibitor in concentrations below the inhibition constant (Ki = 1.4 x 10(-7) M) exerts the most intensive effect on the decarbamylation rate of methyl- and dimethylcarbamylated acetylcholinesterase of the mouse brain, while ambenonium and galanthamine in concentrations much (tens times) exceeding their Ki (3.1 x 10(-10) M and 4.4 x 10(-7) M, respectively) provide a decrease of the decarbamylation rate.

[The lymphocyte acetylcholinesterase activity in rats poisoned by pesticides]. / Aktivnost' atsetilkholinésterazy limfotsitov krys pri intoksikatsii pestitsidami.

The experiments carried out present the evidence of acetylcholinesterase activity of Wistar rat lymphocytes. It was shown that splenocytes and thymocytes had significantly different levels of the enzyme activity. Peroral administration of phosphor-organic pesticide antio (phormothion) 1/100 and 1/20 LD50 induced the dose-dependent inhibition of splenocyte acetylcholine-esterase activity after 2 months of treatment. It suggests the relation of the immunosuppressive action of pesticide with the interference into the neuromediator mechanisms regulating the lymphoid cell function.

[Acetylenic amines as inhibitors of microsomal monooxygenases]. / Atsetilenovye aminy kak ingibitory mikrosomal'nykh monooksigenaz.

A new class of compounds: acetylenic amines, possessing structural similarity with the known inhibitor SKF-525A, and their saturated analogues, has been studied for its effect on the microsomal cytochrome P-450-dependent monooxygenases (MM). Significant differences in sensitivity of different substrate oxidation reactions in experiments with the mouse liver MM were observed. It was shown that the acetylenic amines investigated 13-30 times exceeded their saturated analogues as to the ability to inhibit aminopyrine and benzo[a]pyrene oxidation, and differed but slightly from their analogues with respect to p-nitroanisole and paraoxon oxidation. Benzo[a]pyrene hydroxylase of the house-fly abdomens was less sensitive to the compound investigated than that of the mouse liver, however, in contrast to it exhibited selectively to the diphenyl derivatives and not to the monophenyl ones.

[A comparative study of the effect of vinyl phosphoric acid esters on cholinesterase and carboxylesterase activities in mammals and arthropods]. / Sravnitel'noe issledovanie vliianiia éfirov vinilfosfornoi kisloty na aktivnost' kholinésterazy i karboksilésterazy mlekopitaiushchikh i chlenistonogikh.

Studies have been made of the effect of organophosphorus inhibitors on cholinesterase and carboxylesterase from various mammals (human erythrocytes, mouse brain, blood serum of mouse and rat, blood serum of horse) and arthropods (Calliphora vicina, Schizaphis graminum, Myzus persicae, Sitophilus oryzae, Pseudococcus maritimus, Tetranychus urticae). Organophosphorus inhibitors were presented by esters of vynylphosphoric acid containing normal and branched alkyls in the phosphoryl part of the molecule. The increase of the radical up to a propyl one increased the effect of organophosphorus inhibitors with respect to cholinesterase from the majority of the arthropods investigated. Organophosphorus compound with an isopropyl radical was found to be weaker for all the enzymes studied. Extremely high sensitivity of carboxylesterase from all arthropods to all organophosphorus inhibitors was noted; in some of the cases, anticarboxylesterase activity of all drugs was 2-3 orders higher than anticholinesterase one (P. maritimus, T. urticae). Regularities established for cholinesterase practically completely were confirmed on carboxylesterase. This finding evidently reveals similar structure of catalytic surface at the vicinity of esterase center in both enzymes.

[Interaction of membrane-bound and solubilized erythrocyte acetylcholinesterase with carbamates]. / Vzaimodeistvie membranosviazannoi i soliubilizirovannoi atsetilkholinésterazy éritrotsitov s karbamatami.

Physostigmine and neostigmine methylsulphate are shown to be the most strong inhibitors of acetylcholine esterase of human erythrocytes. The action of baigon is less pronounced and pyrimor is characterized as the weakest inhibitor. No differences are found between the membrane-bound and solubilized acetylcholine esterase relative to their ability to be inhibited by these carbamates. The preliminary treatment of acetylcholine esterase with carbamates protects the enzyme from the subsequent inhibition by the organophosphoric inhibitor. A higher concentration (1.6-2.1 times) of physostigmine and pyrimor and lower (1.7-1.9 times) one of baigon and neostigmine methylsulphate are needed for protection of the soluble enzyme than of the membrane-bound enzyme.

[Interaction of membrane-bound and solubilized acetylcholinesterase from human and bovine erythrocytes with organophosphorus inhibitors]. / Vzaimodeistvie membranosviazannoi i soliubilizirovannoi atsetilkholinésterazy éritrotsitov cheloveka i byka s fosfororganicheskimi ingibitorami.

Differences are found between the membrane-bound and soluble acetylcholinesterases of human and bovine erythrocytes when the enzyme interacts with organophosphoric inhibitors in the presence of acetylc choline and galantamine, a reverse inhibitor of acetylcholinesterase. In most cases prevention of inhibition of the soluble enzyme activity necessitates a higher (2-3 times higher) concentration of the protecting agent than protection of the membrane-bound enzyme. Concentrations of acetylcholine and galantamine providing a 50% protection of the enzyme did not practically depend on the strength of the anticholinesterase action of organophosphoric inhibitors.

[Effect of organophosphorus inhibitors, lupinine and epilupinine derivatives, on mammalian and arthropod cholinesterases]. / Deistvie fosfororganicheskikh ingibitorov proizvodnykh lupinina i épilupinina, na kholinésterazy mlekopitaiushchikh i chlenistonogikh.

Studies have been made on the effect of organophosphorus inhibitors on cholinesterases (CHe) from mammals (human and rabbit erythrocytes, mice brain) and arthropods (fly, spring grain aphid, rice weevil and spider tick). Organophosphorus inhibitors were presented by dialkylthiophosphates which contained normal or branching alkyls in the phosphoryl part of their molecule and lupinine or epilupinine residue in the other part of the latter. Increasing the length of alkyl in lupinine derivatives increased their inhibitory effect with respect to mammalian CHe, but decreased it in relation to all the investigated arthropod CHe. For epilupinine derivatives, the same relationship was found with mammalian CHe, being absent in arthropod enzymes. These data indicate the existence of significant differences in spatial structure of both esterase and anionic centers of the enzyme from mammals and arthropods. In arthropod CHe, hydrophobic regions of the esterase center are less developed that in mammalian CHe. The distance between the anionic and esterase centers of CHe is presumably different in the enzymes from different species.

[Intracellular distribution of phosphoorganic cholinesterase inhibitors in rat brain]. / Vnutrikletochnoe raspredelenie fosfororganicheskikh ingibitorov kholinésterazy v mozge krys.

Content of organophosphorous inhibitors (with the structure RO/CH3/P/O/SC2H4SC2H5) of cholinesterase as well as their hydrophobic properties (distribution coefficient in hexan/water system) were studied in subcellular fractions of rat brain. Relative content of organophosphorous inhibitors was distinctly decreased in supermicrosomal fraction with increase of hydrophobic properties of the fraction. Nuclear and mitochondrial fractions contained the more hydrophobic substances in relatively higher amount. When homogenate of supermicrosomal fraction was incubated at 37 degrees, own brain cholinesterase was not depressed by organophosphorous inhibitors, containing in the fraction at low concentration. The phenomenon exhibits that content of free organophosphorous inhibitors is distinctly lower in the subcellular fractions studied than amount of the inhibitors, extracted with chloroform.

[Proton acceptor properties of the blood plasma of a series of vertebrates]. / Izuchenie protonoaktseptornykh svoistv plazmy krovi v riadu pozvonochnykh.

Comparative studies have been made on the kinetics of thermal denaturation of the blood plasma of various vertebrates (lampreys, teleosts, frogs, tortoises, pigeons, mice, rats, rabbits, cats, dogs, man) by measuring ionization equilibrium of protein solution at elevated temperature. It was demonstrated that during the initial stage of heat denaturation at 58 degrees the blood plasma of all the animals studied binds protons practically linearly. Among the animals studied, the highest protonoacceptor capacity exhibited the plasma of warm-blooded animals; it was lower in tortoises frogs and teleost fishes, being the lowest one in lampreys. Comparison of total electric charge of plasma proteins evaluated by potentiometric titration with data on thermal denturation of plasma revealed positive correlation between the charge and protonoacceptor properties of the plasma.

[Effect of environment on allosteric properties of acetylcholinesterase]. / Vliianie uslovii sredy na allostericheskie svoistva atsetilkholinèsterazy.

In the presence of organophosphorus inhibitors (OPI) AChE inhibition is initiated at a lower concentration of ACh; the plot reaction rate versus substrate concentration shows two maxima with a distinct minimum between them. It was shown that extremely mild conditions (short-term heating up to 50 degrees C; acidic or alkaline pH shift by 0.5 units; high concentrations of bivalent cations; erythrocyte storage) which do not affect substrate inhibition, remove this effect. The data obtained suggest that OPI effect is not directed to the site of AChE responsible for enzyme inhibition by ACh excess ("substrate inhibition site"), but to some other area. This results in a change in the conformation of the substrate inhibition site and a pronounced inhibition of the AChE activity takes place at lower substrate concentration.

[Effect of diazepam on the properties of an erythrocyte-plasma protein complex and the development of shock during the transfusion of heterogenic blood]. / Vliianie diazepama na svoistva kompleksa éritrotsity-belki plazmy i razvitie shoka pri perelivanii geterogennoi krovi.

Preliminary administration of 2 mg/kg of diazepam eliminated the pyrogenic reaction and increased survival of rabbits following intravenous injection of cat blood to them. A study of physico-chemical properties of the blood by measurement of the ionization balance in thermal denaturing demonstrated that injection of heterogenous blood to rabbits and suspension of rabbit erythrocytes in cat plasma in vitro led to a sharp reduction of the rate of proton binding with plasma proteins in the presence of foreign erythrocytes. Preliminary administration of diazepam to rabbits, and diazepam addition to the cat plasma in vitro prevented such changes and promoted retention of normal blood properties. Addition of diazepam in a concentration of about 10--4 M to the heterogenous plasma decreased erythrocyte agglutination considerably.

[Binding of phosphoorganic cholinesterase inhibitors in rat brain tissue]. / Osobennosti sviazyvaniia fosfororganicheskikh ingibitorov kholinésterazy v tkani mozga krys

Hydrophobity (coefficient in distribution in the hexane water system) and the content of cholinesterase organophosphorous inhibitors (OPI) of the structure Ro (CH3) P (O) SC2H4 SC2H5 were studied in the rat brain. When the O-alkyl radical is increased hydrophobity rises and the relative content of free OPI in the brain extracted by chloroform decreases. With an increase in R from the ethyl to butyl one the ability to the additional inhibition of the brain own cholinesterase lowers due to incubation of homogenate at 37 degrees C, that evidences for an essential drop in the studied series of the free OPI fraction relative to the free OPI extracted by chloroform.