New quaternary pyridine aldoximes as casual antidotes against nerve agents intoxications

In this work, the ability of four newly synthesized oximes--K005 (1,3-bis(2-hydroxyiminomethylpyridinium) propane dibromide), K027 (1-(4-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium) propane dibromide), K033 (1,4-bis(2-hydroxyiminomethylpyridinium) butane dibromide) and K048 (1-(4-hydroxyiminomethylpyridinium)-4-(4-carbamoylpyridinium) butane dibromide) to reactivate acetylcholinesterase (AChE, EC 3.1.1.7) inhibited by nerve agents is summarized. Reactivation potency of these compounds was tested using standard in vitro reactivation test. Tabun, sarin, cyclosarin and VX agent were used as appropriate testing nerve agents. Rat brain AChE was used as a source of the enzyme. Efficacies of new reactivators to reactivate tabun-, sarin-, cyclosarin- and VX-inhibited AChE were compared with the currently used AChE reactivators (pralidoxime, obidoxime and HI-6). Oxime K048 seems to be promising reactivator of tabun-inhibited AChE. Its reactivation potency is significantly higher than that of HI-6 and pralidoxime and comparable with the potency of obidoxime. The best reactivator of sarin-inhibited AChE seems to be oxime HI-6. None of the new AChE reactivators reached comparable reactivation potency. The same results were obtained for cyclosarin-inhibited AChE. However, oxime K033 is also potent reactivator of AChE inhibited by this nerve agent. In the case of VX inhibition, obidoxime and new oximes K027 and K048 seem to be the best AChE reactivators. None from the currently tested AChE reactivators is able to reactivate AChE inhibited by all nerve agents used and, therefore, the search for new potential broad spectrum AChE reactivators is needed.     

Phosphoryl oxime inhibition of acetylcholinesterase during oxime reactivation is prevented by edrophonium.

Reactivation of organophosphate (OP)-inhibited acetylcholinesterase (AChE) is a key objective in the treatment of OP poisoning. This study with native, wild-type, and mutant recombinant DNA-expressed AChEs, each inhibited by representative OP compounds, establishes a relationship between edrophonium acceleration of oxime-induced reactivation of OP-AChE conjugates and phosphoryl oxime inhibition of the reactivated enzyme that occurs during reactivation by pyridinium oximes LüH6 and TMB4. No such recurring inhibition could be observed with HI-6 as the reactivator due to the extreme lability of the phosphoryl oximes formed by this oxime. Phosphoryl oximes formed during reactivation of the ethoxy methylphosphonyl-AChE conjugate by LüH6 and TMB4 were isolated for the first time and their structures confirmed by (31)P NMR. However, phosphoryl oximes formed during the reactivation of the diethylphosphoryl-AChE conjugate were not sufficiently stable to be detected by (31)P NMR. The purified ethoxy methylphosphonyl oximes formed during the reactivation of ethoxy methylphosphonyl-AChE conjugate with LüH6 and TMB4 are 10- to 22-fold more potent than MEPQ as inhibitors of AChE and stable for several hours at pH 7.2 in HEPES buffer. Reactivation of both ethoxy methylphosphonyl- and diethylphosphoryl-AChE by these two oximes was accelerated in the presence of rabbit serum paraoxonase, suggesting that organophosphorus hydrolase can hydrolyze phosphoryl oxime formed during the reactivation. Our results emphasize that certain oximes, such as LüH6 and TMB4, if used in the treatment of OP pesticide poisoning may cause prolonged inhibition of AChE due to formation of phosphoryl oximes.     

Changes of rat plasma total low molecular weight antioxidant level after tabun exposure and consequent treatment by acetylcholinesterase reactivators

These experiments were performed on a rat model. The rats were divided into eight groups and consequently exposed to either a saline solution (control), atropine or a combination of atropine and tabun. The reactivation efficacy of the oximes was estimated on the rats exposed to tabun, atropine and a reactivator of AChE. The oximes HI-6, obidoxime, trimedoxime, K203 and KR-22836 were used as representative compounds of commonly available and new AChE reactivators. Besides the positive effect of the administered reactivators on blood AChE activity, the sizable modulation of low molecular weight antioxidant (LMWA) levels was also determined. The LMWA levels in the the animals treated with the oxime reactivators were decreased in comparison with the animals treated by atropine alone. It was found that the levels of LMWA returned to the level found in the control animals when either trimedoxime, K203 or KR-22836 were administered. The principle of oxime reactivator function and a novel insight into AChE activity regulation and oxidative stress is discussed.     

Oxime-assisted acetylcholinesterase catalytic scavengers of organophosphates that resist aging

The cholinesterases, acetylcholinesterase (AChE) and butyrylcholinesterase, are primary targets of organophosphates (OPs). Exposure to OPs can lead to serious cardiovascular complications, respiratory compromise, and death. Current therapy to combat OP poisoning involves an oxime reactivator (2-PAM, obidoxime, TMB4, or HI-6) combined with atropine and on occasion an anticonvulsant. Butyrylcholinesterase, administered in the plasma compartment as a bio-scavenger, has also shown efficacy but is limited by its strict stoichiometric scavenging, slow reactivation, and a propensity for aging. Here, we characterize 10 human (h) AChE mutants that, when coupled with an oxime, give rise to catalytic reactivation and aging resistance of the soman conjugate. With the most efficient human AChE mutant Y337A/F338A, we show enhanced reactivation rates for several OP-hAChE conjugates compared with wild-type hAChE when reactivated with HI-6 (1-(2'-hydroxyiminomethyl-1'-pyridinium)-3-(4'-carbamoyl-1-pyridinium)). In addition, we interrogated an 840-member novel oxime library for reactivation of Y337A/F338A hAChE-OP conjugates to delineate the most efficient oxime-mutant enzyme pairs for catalytic bio-scavenging. Combining the increased accessibility of the Y337A mutation to oximes within the space-impacted active center gorge with the aging resistance of the F338A mutation provides increased substrate diversity in scavenging potential for aging-prone alkyl phosphate inhibitors.     

Design,synthesis and evaluation of new classes of nonquaternary reactivators for acetylcholinesterase inhibited by organophosphates

A new series of nonquaternary conjugates for reactivation of both nerve agents and pesticides inhibited hAChE were described in this paper. It was found that substituted salicylaldehydes conjugated to aminobenzamide through piperidine would produce efficient reactivators for sarin, VX and tabun inhibited hAChE, such as L6M1R3, L6M1R5 to L6M1R7, L4M1R3 and L4M1R5 to L4M1R7. The in vitro reactivation experiment for pesticides inhibited hAChE of these new synthesized oximes were conducted for the first time. Despite they were less efficient than obidoxime, some of them were highlighted as equal or more efficient reactivators in comparison to 2-PAM. It was found that introduction of peripheral site ligands could increase oximes’ binding affinity for inhibited hAChE in most cases, which resulted in greater reactivation ability.     

Preclinical Studies of Noncharged Oxime Reactivators for Organophosphate Exposure

A countermeasure that protects the brain from organophosphate toxicity is an unmet need. Few small molecule reactivators that can cross the blood brain barrier and reactivate brain acetyl cholinesterases have been reported. Herein, we describe preclinical investigations of a new class of amidine–oxime reactivator of cholinesterases with improved potency and blood brain barrier permeability. (Z)‐N‐((E)‐1‐(Dimethylamino)‐2‐(hydroxyimino)ethylidene)butan‐1‐aminium chloride, 1 , is zwitterionic at physiological pH but possesses increased oxime nucleophilicity because of the adjacent amidine functionality. The amidine–oximes reported herein were observed to be nontoxic (up to 200 mg/kg in vivo) and are chemically and metabolically stable. The results presented herein show that uncharged amidine–oxime reactivators such as 1 can penetrate the blood brain barrier in animals and protect from the toxicity of nerve agent model compounds.     

Refinement of structural leads for centrally acting oxime reactivators of phosphylated cholinesterases

We present a systematic structural optimization of uncharged but ionizable N-substituted 2-hydroxyiminoacetamido alkylamine reactivators of phosphylated human acetylcholinesterase (hAChE) intended to catalyze the hydrolysis of organophosphate (OP)-inhibited hAChE in the CNS. Starting with the initial lead oxime RS41A identified in our earlier study and extending to the azepine analog RS194B, reactivation rates for OP-hAChE conjugates formed by sarin, cyclosarin, VX, paraoxon, and tabun are enhanced severalfold in vitro. To analyze the mechanism of intrinsic reactivation of the OP-AChE conjugate and penetration of the blood-brain barrier, the pH dependence of the oxime and amine ionizing groups of the compounds and their nucleophilic potential were examined by UV-visible spectroscopy, (1)H NMR, and oximolysis rates for acetylthiocholine and phosphoester hydrolysis. Oximolysis rates were compared in solution and on AChE conjugates and analyzed in terms of the ionization states for reactivation of the OP-conjugated AChE. In addition, toxicity and pharmacokinetic studies in mice show significantly improved CNS penetration and retention for RS194B when compared with RS41A. The enhanced intrinsic reactivity against the OP-AChE target combined with favorable pharmacokinetic properties resulted in great improvement of antidotal properties of RS194B compared with RS41A and the standard peripherally active oxime, 2-pyridinealdoxime methiodide. Improvement was particularly noticeable when pretreatment of mice with RS194B before OP exposure was combined with RS194B reactivation therapy after the OP insult.     

Synthesis and in vitro reactivation study of isonicotinamide derivatives of 2-(hydroxyimino)-N-(pyridin-3-yl)acetamide as reactivators of Sarin and VX inhibited human acetylcholinesterase (hAChE)

Previously (Karade et al., 2014), we have reported the synthesis and in vitro evaluation of bis-pyridinium derivatives of pyridine-3-yl-(2-hydroxyimino acetamide), as reactivators of sarin and VX inhibited hAChE. Few of the molecules showed superior in vivo protection efficacy (mice model) (Kumar et al., 2014; Swami et al., 2016) in comparison to 2-PAM against DFP and sarin poisoning. Encouraged by these results, herein we report the synthesis and in vitro evaluation of isonicotinamide derivatives of pyridine-3-yl-(2-hydroxyimino acetamide) (4a–4d) against sarin and VX inhibited erythrocyte ghost hAChE. Reactivation kinetics of these compounds was studied and the determined kinetic parameters were compared with that of commercial reactivators viz. 2-PAM and obidoxime. In comparison to 2-PAM and obidoxime, oxime 4a and 4b exhibited enhanced reactivation efficacy toward sarin inhibited hAChE while oxime 4c showed far greater reactivation efficacy toward VX inhibited hAChE. The acid dissociation constant and IC₅₀ values of these oximes were determined and correlated with the observed reactivation potential.     

Novel tacrine-pyridinium hybrid reactivators of organophosphorus-inhibited acetylcholinesterase: Synthesis,molecular docking,and in vitro reactivation study

First-line medical treatment against nerve agents consists of co-administration of anticholinergic agents and oxime reactivators, which reactivate inhibited AChE. Pralidoxime, a commonly used oxime reactivator, is effective against some nerve agents but not against others; thus, new oxime reactivators are needed. Novel tacrine-pyridinium hybrid reactivators in which 4-pyridinealdoxime derivatives are connected to tacrine moieties by linear carbon chains of different lengths (C2–C7) were prepared (Scheme 1, 5a–f). Their binding affinities to electric eel AChE were tested because oximes can inhibit free AChE, and the highest AChE activity (95%, 92%, and 90%) was observed at 1?μM concentrations of the oximes (5a, 5b, and 5c, respectively). Based on their inhibitory affinities towards free AChE, 1?μM concentrations of the oxime derivatives (5) were used to examine reactivation of paraoxon-inhibited AChE. Reactivation ability increased as the carbon linker chains lengthened (n?=?2–5), and 5c and 5d showed remarkable reactivation ability (41%) compared to that of 2-PAM (16%) and HI-6 (4%) against paraoxon-inhibited electric eel AChE at 1?μM concentrations. Molecular docking simulation showed that the most stable binding free energy was observed in 5c at 73.79?kcal?mol^?1, and the binding mode of 5c is acceptable for the oxygen atom of oximate to attack the phosphorus atom of paraoxon and reactivate paraoxon-inhibited eel AChE model structure.     

Mono-oxime bisquaternary acetylcholinesterase reactivators with prop-1,3-diyl linkage-Preparation, in vitro screening and molecular docking

The treatment of organophosphorus (OP) poisoning consists of the administration of a parasympatholytic agent (e.g., atropine), an anticonvulsant (e.g., diazepam) and an acetylcholinesterase (AChE) reactivator (e.g., obidoxime). The AChE reactivator is the causal treatment of OP exposure, because it cleaves the OP moiety covalently bound to the AChE active site. In this paper, fourteen novel AChE reactivators are described. Their design originated from a former promising compound K027. These compounds were synthesized, evaluated in vitro on human AChE (hAChE) inhibited by tabun, paraoxon, methylparaoxon and DFP and then compared to commercial hAChE reactivators (pralidoxime, HI-6, trimedoxime, obidoxime, methoxime) or previously prepared compounds (K027, K203). Three of these novel compounds showed a promising ability to reactivate hAChE comparable or better than the used standards. Consequently, a molecular docking study was performed for three of these promising novel compounds. The docking results confirmed the apparent influence of π-π or cation-π interactions and hydrogen bonding for reactivator binding within the hAChE active site cleft. The SAR features concerning the non-oxime part of the reactivator molecule are also discussed.     

New group of xylene linker-containing acetylcholinesterase reactivators as antidotes against the nerve agent cyclosarin

Nerve agents such as sarin, cyclosarin and tabun are organophosphorus substances able to inhibit the enzyme acetylcholinesterase (AChE; EC 3.1.1.7). AChE reactivators and anticholinergics are generally used as antidotes in the case of intoxication with these agents. None of the known AChE reactivators is able to reactivate AChE inhibited by all nerve agents used. In this work, reactivation potency of nine newly developed AChE reactivators with an incorporated xylene ring in their structure was measured in vitro. Cyclosarin was chosen as an appropriate member of the nerve agent family. Reactivation potency of the tested AChE reactivators was compared with the gold standard of AChE reactivators--pralidoxime. Two oximes (K107 and K108) surpassed the reactivation potency of pralidoxime. Moreover, from the obtained results it could be deduced that AChE reactivators with a functional oxime group in position-2 are the most potent AChE reactivators in the case of cyclosarin intoxications.     

New group of xylene linker-containing acetylcholinesterase reactivators as antidotes against the nerve agent cyclosarin

Nerve agents such as sarin, cyclosarin and tabun are organophosphorus substances able to inhibit the enzyme acetylcholinesterase (AChE; EC 3.1.1.7). AChE reactivators and anticholinergics are generally used as antidotes in the case of intoxication with these agents. None of the known AChE reactivators is able to reactivate AChE inhibited by all nerve agents used. In this work, reactivation potency of nine newly developed AChE reactivators with an incorporated xylene ring in their structure was measured in vitro. Cyclosarin was chosen as an appropriate member of the nerve agent family. Reactivation potency of the tested AChE reactivators was compared with the gold standard of AChE reactivators – pralidoxime. Two oximes (K107 and K108) surpassed the reactivation potency of pralidoxime. Moreover, from the obtained results it could be deduced that AChE reactivators with a functional oxime group in position-2 are the most potent AChE reactivators in the case of cyclosarin intoxications.     

Potency of new structurally different oximes to reactivate cyclosarin-inhibited human brain acetylcholinesterases

Antidotes currently used for organophosphorus pesticide and nerve agent intoxications consist of anticholinergics (atropine mainly) and acetylcholinesterase (AChE, EC 3.1.1.7) reactivators called oximes. Owing to the wide-spread of these toxic compounds worldwide, development of antidotes in the case of first aid is needed. To select the most promising AChE reactivators is a very time consuming process, which is necessary before approval of these compounds to be used as human antidotes. Because of ethical reasons, many developing experiments have been conducted on laboratory animals. However, these results often could not be transferred directly to human. Here, we have tested five newly developed AChE reactivators--K027, K033, K048, K074 and K075, which showed promising reactivation activity on rodents, as reactivators of inhibited human brain cholinesterases. For this purpose, cyclosarin was used as member of the nerve agent family. Oxime HI-6 and pralidoxime were used as AChE reactivator standards. Two AChE reactivators, K027 and K033, achieved comparable reactivation potency as HI-6. Moreover, oxime K033 reached its maximal reactivation potency at the lowest concentration which could be attained in humans.     

Potency of new structurally different oximes to reactivate cyclosarin-inhibited human brain acetylcholinesterases

Antidotes currently used for organophosphorus pesticide and nerve agent intoxications consist of anticholinergics (atropine mainly) and acetylcholinesterase (AChE, EC 3.1.1.7) reactivators called oximes. Owing to the wide-spread of these toxic compounds worldwide, development of antidotes in the case of first aid is needed. To select the most promising AChE reactivators is a very time consuming process, which is necessary before approval of these compounds to be used as human antidotes. Because of ethical reasons, many developing experiments have been conducted on laboratory animals. However, these results often could not be transferred directly to human. Here, we have tested five newly developed AChE reactivators – K027, K033, K048, K074 and K075, which showed promising reactivation activity on rodents, as reactivators of inhibited human brain cholinesterases. For this purpose, cyclosarin was used as member of the nerve agent family. Oxime HI-6 and pralidoxime were used as AChE reactivator standards. Two AChE reactivators, K027 and K033, achieved comparable reactivation potency as HI-6. Moreover, oxime K033 reached its maximal reactivation potency at the lowest concentration which could be attained in humans.     

A comparison of reactivating efficacy of newly developed oximes (K074, K075) and currently available oximes (obidoxime, HI-6) in soman, cyclosarin and tabun-poisoned rats

The potency of newly developed oximes (K074, K075) and commonly used oximes (obidoxime, HI-6) to reactivate nerve agent-inhibited acetylcholinesterase was evaluated in rats poisoned with soman, tabun or cyclosarin at a lethal dose corresponding to their LD(50) value. In vivo determined percentage of reactivation of soman-inhibited blood and brain acetylcholinesterase in poisoned rats showed that only the oxime HI-6 was able to reactivate soman-inhibited acetylcholinesterase in the peripheral (blood) as well as central (brain) compartment. In vivo determined percentage of reactivation of tabun-inhibited blood and brain acetylcholinesterase in poisoned rats showed that obidoxime is the most efficacious reactivator of tabun-inhibited acetylcholinesterase among studied oximes in the peripheral compartment (blood) while K074 seems to be the most efficacious reactivator of tabun-inhibited acetylcholinesterase among studied oximes in the central compartment (brain). In vivo determined percentage of reactivation of cyclosarin-inhibited blood and brain acetylcholinesterase in poisoned rats showed that HI-6 is the most efficacious reactivator of cyclosarin-inhibited acetylcholinesterase among studied oximes. Due to their reactivating effects, both newly developed K oximes can be considered to be promising oximes for the antidotal treatment of acute tabun poisonings while the oxime HI-6 is still the most promising oxime for the treatment of acute soman and cyclosarin poisonings.     

Pyridinium-2-carbaldoximes with quinolinium carboxamide moiety are simultaneous reactivators of acetylcholinesterase and butyrylcholinesterase inhibited by nerve agent surrogates

The pyridinium-2-carbaldoximes with quinolinium carboxamide moiety were designed and synthesised as cholinesterase reactivators. The prepared compounds showed intermediate-to-high inhibition of both cholinesterases when compared to standard oximes. Their reactivation ability was evaluated in vitro on human recombinant acetylcholinesterase (hrAChE) and human recombinant butyrylcholinesterase (hrBChE) inhibited by nerve agent surrogates (NIMP, NEMP, and NEDPA) or paraoxon. In the reactivation screening, one compound was able to reactivate hrAChE inhibited by all used organophosphates and two novel compounds were able to reactivate NIMP/NEMP-hrBChE. The reactivation kinetics revealed compound 11 that proved to be excellent reactivator of paraoxon-hrAChE better to obidoxime and showed increased reactivation of NIMP/NEMP-hrBChE, although worse to obidoxime. The molecular interactions of studied reactivators were further identified by in silico calculations. Molecular modelling results revealed the importance of creation of the pre-reactivation complex that could lead to better reactivation of both cholinesterases together with reducing particular interactions for lower intrinsic inhibition by the oxime.     

A comparison of reactivating efficacy of newly developed oximes (K074, K075) and currently available oximes (obidoxime, HI-6) in cyclosarin-and tabun-poisoned rats

The potency of newly developed oximes (K074, K075) and commonly used oximes (obidoxime, HI-6) to reactivate nerve agent-inhibited acetylcholinesterase was evaluated in rats poisoned with tabun or cyclosarin at a lethal dose corresponding to the LD50 value. In vivo determined percentage of reactivation of tabun-inhibited blood and brain acetylcholinesterase showed that obidoxime is the most efficacious reactivator of tabun-inhibited acetylcholinesterase among studied oximes in the peripheral compartment (blood) although the differences between obidoxime and newly developed oximes were not significant. On the other hand, one of the newly developed oximes (K074) seems to be a significantly more efficacious reactivator of tabun-inhibited acetylcholinesterase in the central compartment (brain) than the other studied oximes. In addition, the oxime HI-6 is unable to sufficiently reactivate tabun-inhibited acetylcholinesterase in rats. In vivo determined percentage of reactivation of cyclosarin-inhibited blood and brain acetylcholinesterase in poisoned rats showed that HI-6 is the most efficacious reactivator of cyclosarin-inhibited acetylcholinesterase among the studied oximes in the peripheral (blood) as well as central (brain) compartment although the differences between the oxime HI-6 and other tested oximes in the brain were not significant. Due to their reactivating effects, both newly developed K-oximes can be considered to be promising oximes for the antidotal treatment of acute tabun poisoning while the oximes HI-6 is still the most promising oxime for the treatment of acute cyclosarin poisonings due to its high potency in reactivating cyclosarin-inhibited acetylcholinesterase in the peripheral as well as central compartment.     

Reactivation of organophosphate-inhibited human acetylcholinesterase by isonitrosoacetone (MINA): a kinetic analysis

Treatment of poisoning by highly toxic organophosphorus compounds (OP) with atropine and an acetylcholinesterase (AChE) reactivator (oxime) is of limited effectiveness in case of different nerve agents and pesticides. One challenge is the reactivation of OP-inhibited brain AChE which shows inadequate success with charged pyridinium oximes. Recent studies with high doses of the tertiary oxime isonitrosoacetone (MINA) indicated a beneficial effect on central and peripheral AChE and on survival in nerve agent poisoned guinea pigs. Now, an in vitro study was performed to determine the reactivation kinetics of MINA with tabun-, sarin-, cyclosarin-, VX- and paraoxon-inhibited human AChE. MINA showed an exceptionally low affinity to inhibited AChE but, with the exception of tabun-inhibited AChE, a moderate to high reactivity. In comparison to the pyridinium oximes obidoxime, 2-PAM and HI-6 the affinity and reactivity of MINA was in most cases lower and in relation to the most effective reactivators, the second order reactivation constant of MINA was 500 to 3400-fold lower. Hence, high in vivo MINA concentrations would be necessary to achieve at least partial reactivation. This assumption corresponds to in vivo data showing a dose-dependent effect on reactivation and survival in animals. In view, of the toxic potential of MINA in animals human studies would be necessary to determine the tolerability and pharmacokinetics of MINA in order to enable a proper assessment of the value of this oxime as an antidote in OP poisoning.     

Structures of paraoxon‐inhibited human acetylcholinesterase reveal perturbations of the acyl loop and the dimer interface

Irreversible inhibition of the essential nervous system enzyme acetylcholinesterase by organophosphate nerve agents and pesticides may quickly lead to death. Oxime reactivators currently used as antidotes are generally less effective against pesticide exposure than nerve agent exposure, and pesticide exposure constitutes the majority of cases of organophosphate poisoning in the world. The current lack of published structural data specific to human acetylcholinesterase organophosphate‐inhibited and oxime‐bound states hinders development of effective medical treatments. We have solved structures of human acetylcholinesterase in different states in complex with the organophosphate insecticide, paraoxon, and oximes. Reaction with paraoxon results in a highly perturbed acyl loop that causes a narrowing of the gorge in the peripheral site that may impede entry of reactivators. This appears characteristic of acetylcholinesterase inhibition by organophosphate insecticides but not nerve agents. Additional changes seen at the dimer interface are novel and provide further examples of the disruptive effect of paraoxon. Ternary structures of paraoxon‐inhibited human acetylcholinesterase in complex with the oximes HI6 and 2‐PAM reveals relatively poor positioning for reactivation. This study provides a structural foundation for improved reactivator design for the treatment of organophosphate intoxication. Proteins 2016; 84:1246–1256. © 2016 Wiley Periodicals, Inc.     

Assessment of antidotal efficacy of cholinesterase reactivators against paraoxon: In vitro reactivation kinetics and physicochemical properties

The search of proficient oximes as reactivators of irreversibly inhibited-AChE by organophosphate poisoning necessitates an appropriate assessment of their physicochemical properties and reactivation kinetics. Therefore, herein acid dissociation constant; pK_a, lipophilicity; log P, polar surface area, hydrogen bond donor and acceptor counts of structurally different oximes (two tertiary oximes and thirteen pyridinium aldoxime derivatives) have been evaluated. The experimentally obtained data for pK_a has been comparatively analyzed by using non-linear regression. Further the tested oximes were screened through in vitro reactivation kinetics against paraoxon-inhibited AChE. The pK_a values of all the examined oximes were within the range of 7.50–9.53. pK_a values of uncharged and mono-pyridinium oximes were in good correlation with their reactivation potency. The high negative log P values of pyridinium oxime reactivators indicate their high hydrophilic character; hence oximes with improved lipophilicity should be designed for the development of novel and more potent antidotes. Propane and butane linked oximes were superior reactivators than xylene linked bis-oxime reactivators. It is concluded from the present study that pK_a value is not only ruled by the position of oximino functionality in the pyridinium ring, but also by the position of linker. Although, pyridinium oximes are proved to be better reactivators but their lipophilicity has to be improved.