N. E. Basova

The effect of ionic strength on the reversible inhibition of acetylcholinesterase under the influence of thionephosphonates of different hydrophobicity.

280 A study of the anticholinesterase efficacy of a series of esters of thionephosphonic acids was a premise for obtaining a new group of reversible organophosphorous inhibitors (OPIs) of cholinesterases characterized by a competitive type of action [1‐3]. To this end, it was of interest to study the characteristics of the mechanism of action of these OPIs, such as the possibility of contribution of the uncompetitive component to the overall pattern of reversible inhibition, the effect of the structural characteristics of a reversible OPI molecule on the esterase catalysis upon change in ionic strength, and the role of hydrophobic‐hydrophilic interactions in the sorption of the ligand on the catalytic surface of the enzyme. Clarification of these issues will help to determine the advantages of one or another structure of the inhibitor’s molecule taking into account the ion‐ion and hydrophobic interactions taking place during their reaction with cholinesterase. The presence of the trimethylammonium group in the molecule of acetylcholine, the natural substrate of cholinesterase, determines the substantial effect of both ionic strength and the nature of ions on the enzymatic hydrolysis as such, which was studied in sufficient detail earlier [1, 3]. In the line with these studies, the effect of chlorides of alkali metals and alkali-earth metals on various kinetic characteristics, including the parameters of nonproductive substrate binding [4, 5], of enzymatic hydrolysis by serum cholinesterase of acetylcholine and its analogue, N -acetoxyethylene- N -ethylpiperidinium iodide [6]; the molecule of this analogue contains the N -ethylpiperidinium group instead of the trimethylammonium group. That study showed that the introduction of salts to the reaction mixture differentially influenced the catalytic site activity ( a c ) and the Michaelis constant ( K M ) depending on the nature of cation and the structure of substrate. However, the a c / K M ratio, which to a certain extent characterizes the affinity of the substrate for the enzyme [3], calculated for the two substrates of different structure, was similar irrespective of the presence of cations and their nature [6].

A comparative study of sulfonium reversible inhibitors of cholinesterases of various animals.

The majority of cholinergically active compounds in general and reversible inhibitors of cholinesterases in particular are the onium derivatives (i.e., contain a peralkylated heteroatom of a higher valence) [1, 2]. Such a hydrophobic organic cation ensures a fairly rapid penetration of the effector into the active center of the cholinesterase [3]. In addition to ammonium derivatives, which occupy the leading position among the reversible inhibitors of cholinesterases, the phosphonium and sulfonium compounds have been intensely studied in the past years [4–9]. In this work, we performed a comparative study of specially synthesized sulfonium analogues of the ammonium effectors, which were studied earlier. In addition, the erythrocytic acetylcholinesterase and serum butyrylcholinesterase, as well as the preparations of cholinesterases from the optical ganglia of the Comandor squid Berryteuthis magister from various areas of the northwestern area of the Pacific Ocean were studied as reference enzymes.

Element-Onium Silatran Derivatives As Reversible Inhibitors of Cholinesterases of Various Origin

The vast majority of cholinergically active compounds in general and reversible inhibitors of cholinesterases in particular are onium derivatives (i.e., contain a peralkylated heteroatom of a higher valence) [1, 2]. Such a hydrophobic organic cation ensures sufficiently rapid penetration of the effector into the cavity of the active center of cholinesterase [3]. Besides the ammonium derivatives, which take the top place among the reversible inhibitors of cholinesterases, phosphonium and sulfonium compounds have been actively studied in the past years [4–7]. In most cases, alkan derivates were used as carriers of onium groups [2, 6, 7]. We were the first to study the anticholinesterase effect of organosilicon compounds by the example of bisammonium oligomethylsiloxanes, among which high-specificity inhibitors of horse serum butyrylcholinesterase were found [8]. A new stage in these studies includes the synthesis and study of properties of silatran derivatives [9–13]. The selection of this tricyclic group, exhibiting the superelectrondonor inductive effect [10],

Differences in substrate and inhibitor specificity of cholinesterase activity of optical ganglia of the squid Ommastrephes bartrami (Les) as a characteristic of isolation of populations from different areas of a disjunctive home range

The squid Ommastrephes bartrami (LeSueur, 1821), belonging to the family Ommastrephidae (subfamily Ommastrephinae, genus Ommastrephes , d’Orbigny, 1839) is a common oceanic species inhabiting surface and subsurface water layers. Its subtropical (antiequatorial) disjunctive home range consists of three isolated parts: Northern Atlantics, the northern part of the Pacific Ocean, and the southern zone including the Southern Atlantics, the Indian Ocean, and the southwestern part of the Pacific Ocean (the region of the Great Gulf of Australia) [1–8]. The populations of these home-range parts have been completely isolated since the end of the Glacial Period (i.e., approximately ten thousand years). Squids inhabiting these three home-range parts are similar in general morphology, nutrition, mode of breeding, sizes in maturation, parasitic fauna, and other biological traits. They differ in relative sizes of large suckers of the palpi, spermatophore structure, and the set of metacercaries (flukes of the family Didymozoidae) parasitizing on them. These differences are less important than intraspecific differences [1–5]. It was of interest to distinguish squids from the three home-range parts using the biochemical taxonomic method [9], which is based on the comparison of enzymological properties of squid nervous tissue cholinesterase—the key enzyme involved in the cholinergic nerve transmission. It is known that the parameters of the substrate–inhibitory characteristics of cholinesterase are characterized by a pronounced species specificity [10, 11].

Conformational characteristics of the structure of acetylcholine amide derivatives and the specificity of their action as reversible inhibitors of cholinesterases of different animals.

In 1976, Chromov-Borisov et al. [1] performed an elegant study of the role of conformational characteristics of molecules of derivatives of acetylcholine, the mediator of cholinergic nerve transmission, occurring upon the interaction with nicotinic cholinergic receptors. For this purpose, they synthesized a series of derivatives of acetylcholine amide, 2-acetylamidoethylene trimethylammonium iodide (compound I ):

The Use of Reversible Inhibitors of Cholinesterases for Identification of Intraspecific Groups of the Comandor Squid Berryteuthis magister from Different Zones of the Northwestern Area of the Pacific Ocean

The classification of the great variety of squid species in the World Ocean is still based on the morphological traits and biological characteristics; however, precise determination of taxonomy of cephalopods is complicated and, sometimes, impossible [1, 2]. In view of this, we proposed enzymological ways of species determination, which are based on the determination of different catalytic parameters of cholinesterases of squid nervous tissue [3–5]. Recently, we have formulated the list of such parameters [6]. The latter include (1) the degree of homogeneity of cholinesterase activity of squid nervous tissue [6, 7]; (2) the substrate specificity of cholinesterases of different squid species [2, 8]; (3) inhibitory specificity, which is determined using irreversible organophosphorous inhibitors (OPI) of cholinesterases [2, 8]; and (4) reversible onium inhibitors of various structure, which may be used for revealing finer taxonomic (intraspecific) differences.

Elementorganic bisonium reversible inhibitors of different cholinesterases.

Bisammonium compounds have been long known as cholinergically active compounds—blockers of nicotinic cholinergic receptor and reversible inhibitors of cholinesterases [1–3]. In the past years, using the method of molecular mechanics [4–7], we studied the conformation–function relationship for a series of bisamonium derivatives upon their interaction with cholinesterases of various origin [7]. The variation of the structures of effectors consisted in changing the rigidity and composition of the interonium chain, using hexamethylene(trimethylammonium) (hexamethonium) [8] as a reference compound, as well as in increasing the sizes and configuration of the ammonium group [7].

Inhibition of Cholinesterases of Various Origin by Anabasine Derivatives

Nitrogen-containing alkaloids are widely used as various effectors of the cholinergic nervous system [1–3]. Analogs of anabasine, which exhibits a broad spectrum of biological activity, occupied a special place among alkaloid effectors [1]. For example, with respect to ganglion-stimulating activity, anabasine is only four times less effective than nicotine [1]. A pronounced ganglionstimulating activity was also observed in bisanabasides of carboxylic acids [1]. These findings stimulated us to test the effect of anabasine derivatives on the activity of different preparations of cholinesterases, taking into account the fact that the anabasine group itself is of certain structural interest as an anticholinesterase effector. Because anabasine [neonicotine, or 2-(3-pyridyl)piperidine] has two nitrogen-containing heterocycles, its alkylation may yield an asymmetric dicationoid structure, which may affect its sorptive interaction with the catioin-binding sites on the catalytic surface of cholinesterase [2]. In recent years, a series of alkaloid (including anabasine) derivatives exhibiting certain specificity of antianabasine effect has been synthesized [4–6]. It is known that cholinesterases are characterized by a pronounced species and tissue specificity; i.e., enzymes of different animals exhibit different reactivity with respect to the same effectors—substrates and inhibitors (both reversible and irreversible) [2, 5–7].

Alkylammonium derivatives of chlorobenzoic acids: a new group of reversible ester bond-containing inhibitors of cholinesterases of different animals.

Among the vast variety of substrates and inhibitors of cholinesterases of various origin, reversible inhibitors of cholinesterases fill a special and very important place [1–3]. It can be stated that, despite the great diversity of members of the cholinesterase family, determined by pronounced species and tissue specificity of cholinesterases, specific reversible inhibitors exist (or may exist) for nearly every enzyme of this family. For example, specific effectors of the cholinesterase of optical ganglia of the Comandor squid Berryteuthis magister have been revealed for squids captured in different habitats in the northwestern area of the Pacific Ocean [4]. A high selectivity of the effect of reversible inhibitors is probably related to different binding sites of these effectors on the catalytic surface of cholinesterase [5]. It is known that a reversible inhibitor (especially if it displays a noncompetitive or incompetitive inhibition type) may be adsorbed on the periphery of the catalytic surface of cholinesterase. This feature differs it from substrates and quasisubstrates (organophosphorous inhibitors), whose adsorption site should be located in the immediate vicinity of the serine hydroxyl of the triad of the “catalytic machinery” of the active center of cholinesterase [6, 7]. As a result, reversible inhibitors can detect even slight differences in the structure of the catalytic surface of cholinesterase. For this reason, studying every new group of anticholinesterase agents synthesized is promising in terms of searching for specific effectors.

Anticholinesterase efficiency of some tropolone alkaloids and their lumiderivatives.

Alkaloids of the tropolone group, which contain an aromatic seven-mer cyclic oxyketone structure of nonbenzoid type, occur predominantly in plants belonging to some genera of the family Liliacea and exhibit a broad spectrum of physiological activity [1–5]. This mostly applies to such Tropolonic alkaloids as colchicine (compound I ) and colcemid (compound IV ), isolated from bulbotubers of Colchicum speciosum Stev, a representative of the family Liliacea [2, 3]. Both preparations exhibit a pronounced antimitotic activity, blocking mitosis at the stage of metaphase, as well as have a karyoclastic effect and suppress leukoand lymphopoiesis [2, 3]. However, the physiological activity of colchicine and colcemid apaprenlty is not limited by these characteristics. It is known that polycyclic quinolysidine alkaloids, whose structure is similar to the structure of tropolone alkaloids, are blockers of cholinergic transmission and, hence, may serve as reliable tolls for studying the mechanism of functioning of this type of nervous system [1]. The cholinergic (in particular, anticholinesterase) effectiveness of some alkaloids of this group, such as lupinine (compound XI ), anabasine (compound XII ), and cytisine (compound XIII ) and their derivatives have been studied in most detail [1, 6–9]. The presence in the structure of tropolone alkaloids of a planar polycyclic core and nitrogen-containing groups allowed us to assume that thee compounds will also have an anticholinesterase effect. To test the possibility to use the group of colchicine alkaloids and the products of their modification as effectors of the cholinergic nervous system, we performed a comparative study of the effect of these compounds on the activity of references cholinesterases— mammalian erythrocytic acetylcholinesterase and serum butyrylcholinesterase. We studied the following series of tropolone alkaloids and some products of their modification (the formulae are shown below): colchicine (compound ( I ), 3 demethylcolchicine (compound II ), N -acetyl-3-demethylcolchicine (compound III ), colcemid (compound IV ), speciosin (compound V ), aminocolchicine (compound VI ), aminocolcemid (compound ( VII ), β -lumocolchicine (compound ( VIII ), 2 -dimeethylβ -lumocolchicine (compound IX ), and colchicinic acid (compound X ). Compounds I – X were synthesized at the Tashkent Institute of National Economy [10].