Title : Organic synthesis and NMR characterization of bisquaternary pyridinium salts associated with rivastigmine as a potential preventive antidotal system against neurotoxic organophosphorus compounds
Abstract:
Background: Organophosphorus (OP) compounds occupy an important position in both agricultural -related contexts due to their widespread use as pesticides. They exert their toxic effect through irreversible inhibition of acetylcholinesterase (AChE) due to the phosphorylation of hydroxyl-bearing residues including serine at the catalytic site of the enzyme. This modification inhibits acetylcholine hydrolysis and causes its accumulation at synaptic junctions, leading to a marked cholinergic overstimulation. Based on these risks, the design of efficient AChE reactivators and prophylactic agents has long been a core research and biomedicine area in its own right. Bisquaternary pyridinium oximes, such as the aforementioned traditional remedies of the class such as obidoxime or HI-6, are known for their potential of reactivation with enzymes, and the synthesis of novel structural analogues is therefore worthy of further study.
Objective: This work reports the synthesis of new symmetric bisquaternary pyridinium salts derived from the precursor 3-pyridylisonitrosoacetanilide and the potential association with a carbamamate such as rivastigmine in the preventive treatment of exposure to organophosphorus pesticides. The precursors were formed from 3-aminopyridine, chloral hydrate, and hydroxylamine hydrochloride in saturated sodium sulfate. Based on the isolation and purification, the precursor was characterized by ^1H and ^13C NMR spectroscopy and the expected oxime, carbonyl, and heteroaromatic structural elements were verified (research was done at Military Medical Research Center).
Methods: The target bisquaternary salts are generated by two quaternizing agents: symmetric dichlorodimethyl ether (DCMS) and 1,3-dibromopropane. Quaternization reactions were carried out in anhydrous dimethylformamide (DMF) at 30–55 °C, with a 2:1 precursor to dihalogenated reagent ratio and symmetric bisquaternary formation. Purification and analyses of the compounds were carried out by ^1H and ^13C NMR spectroscopy to confirm their molecular structures.
Results: NMR characterization of the precursor 3-pyridylisonitrosoacetanilide displayed spectral signatures associated with the suggested structure, such as typical oxime proton bands and distinct aromatic resonances during ^1H NMR, as well as unique carbonyl, heteroaromatic, and aromatic carbon signatures during ^13C NMR. The bisquaternary salt manufactured by DCMS exhibited duplicated pyridinium signals in both ^1H and ^13C NMR, indicating symmetric quaternization and oxapropane linker presence. Similarly, the compound formed using 1,3-dibromopropane generated downfield pyridinium proton resonances and methylene signals related to a propylene bridge of the compound. ^13C NMR spectra verified the purity of the bisquaternary formation and the preservation of oxime functional group critical for reactivation of AChE. Furthermore, a carbamate-type compound structurally related to rivastigmine was detected and confirmed by high-performance liquid chromatography (HPLC). The chromatographic retention time of the carbamate derivative in the analytical profile was 3.66 ± 0.25 min, which confirmed its presence and purity.
Conclusion: This work presents the successful synthesis, purification, and structural characterization of two new symmetric bisquaternary pyridinium salts derived from 3-pyridylisonitrosoacetanilide Their structural characteristics are similar to those of established AChE reactivators, making them an attractive area for biological evaluation. These new compounds will be chosen for introduction into preventive treatment strategies together with carbamate agents such as rivastigmine against neurotoxic organophosphorus compounds.
Key words: Bisquaternary pyridinium oximes, NMR spectroscopy, rivastigmine, organophosphates
Acknowledgement: This work was carried out through the Core Program within the National Research, Development and Innovation Plan 2022-2027, with the support of National Research Authority (ANC), project no. 23 44
