Felice Liberatore

Dimers of nicotinamide adenine dinucleotide: New evidence for the structure and the involvement in an enzymatic redox process

Abstract The composition and the structure of the product from the known electrochemical dimerization of the NAD+ have been conclusively demonstrated. A detailed analysis of the 1H and 13C nmr spectra has in fact led to the conclusion that the product contains three diastereoisomeric dimers of the 4,4′-tetrahydrobipyridyl type. Furthermore, the cytoplasmic fraction obtained from a standard mitochondrial preparation of rat liver has been shown to catalyze the oxygen uptake by the dimers. A 1 : 1 molar ratio of the reagents in the redox process is indicated by manometric data on oxygen uptake complemented by spectrophotometric analysis of the oxidized substrates, suggesting that H2O2 is the reduction product. NAD+ was identified as the oxidation product by an enzymatic method.

Choline pivaloyl esters improve in rats cognitive and memory performances impaired by scopolamine treatment or lesions of the nucleus basalis of Meynert.

The effects of two choline pivaloyl esters, [2-(2,2-dimethylpropionyloxy)ethyl]trimethylammonium iodide (1) and [2-(2,2-dimethylpropionyloxy)ethyl]trimethylammonium 2,2-dimethylpropionate (2), on learning and memory impairments induced in rats by scopolamine or lesions of nucleus basalis magnocellularis (NBM) have been evaluated by object recognition and Morris water maze tests in comparison with Tacrine (THA). Both 1 and 2 restored discrimination in object recognition test for assessing working-episodic memory and improved spatial memory in scopolamine or NBM-lesioned rats as well. The positive effects produced by 1 and 2 on cognitive and memory deficits were well comparable with those evoked by THA, used as reference compound.

New systems for the specific delivery and sustained release of dopamine to the brain

Abstract The present paper reports the synthesis of the chemical delivery system 5 and dopamine (DA) prodrug 6 as well as their application for the specific delivery and sustained release of DA to the rat brain. The ability of 5 and 6 to penetrate the blood-brain barrier (BBB) and release DA into the central nervous system (CNS) was assessed by comparing, on a molar basis, the behavioural effects produced by DA itself and the above compounds, when centrally or peripherally administered in conscious rats. When intravenously injected, both derivatives 5 and 6 elicited vacuous chewing behaviours comparable with those induced by intracerebroventricular (icv) injection of the parent drug. These results suggest that 5 and 6 are able to cross the BBB and enter the CNS, releasing DA. Furthermore, a long lasting effect was observed for the tripivaloyl-derivative 6, likely due to a slower release of DA following from an increased resistance of the sterically hindered pivaloylamide group to enzymatic hydrolysis. It must be pointed out that the α-adrenergic effect (piloerection) observed after DA was peripherally injected was not observed after systemic administration of the compounds 5 and 6. This finding may indicate that neither the chemical delivery system 5 nor the prodrug 6 release free DA at bioactive concentrations at a peripheral level.

On the structure of intermediate adducts arising from dithionite reduction of pyridinium salts: a novel class of derivatives of the parent sulfinic acid

Abstract 13C and 17O NMR spectroscopy show that adducts arising from dithionite reduction of 3- or 3,5-cyano- or carbamoyl-substituted pyridinium salts to the corresponding 1,4-dihydropyridines, are S-anions of esters of the simplest parent sulfinic acid. A pathway for formation of the 1,4-dihydropyridines, involving an intramolecular hydride transfer, is suggested.

Photocatalyzed oxidation of NAD dimers

Abstract A mixture of dimers of nicotinamide adenine dinucleotide, largely 4,4−-linked, obtained by electrochemical reduction of NAD + , can be photooxidized back to NAD + in the presence of oxygen. Oxygen is consumed during the photooxidation process with the production of hydrogen peroxide. The oxidation is almost pH independent and is stimulated by light whose wavelength exceeds 300 nm. Lactate dehydrogenase and alcohol dehydrogenase added to the solutions under irradiation increased the oxygen uptake by the NAD dimers in a concentration-dependent way. These observations suggest that light induces the homolytic cleavage of NAD dimers to NAD radicals which in turn are oxidized to NAD + by oxygen.

Evidence for binding of NAD dimers to NAD-dependent dehydrogenases.

The binding of dimers of nicotinamide adenine dinucleotide, (NAD)2, to lactate, malate and alcohol dehydrogenase has been studied by the fluorescence quenching technique. While the alcohol dehydrogenase shows a low binding ability, malate and lactate dehydrogenases have been found to bind (NAD)2 in a specific way with high affinity. Malate dehydrogenase binds (NAD)2 more than NADH. All three dehydrogenases are inhibited by (NAD)2, which behaves as a competitive inhibitor with respect to both NAD+ and NADH. The results show that (NAD)2 is bound to the nucleotide-specific binding site of the dehydrogenases. (NAD)2 was found to stoichiometrically react with ferricyanide at variance with NADH. The specific interactions with the NAD-dependent dehydrogenases and the ability to enter in monoelectronic redox cycles suggest possible physiological roles for (NAD)2.

On the regio- and stereoselectivity of pyridinyl radical dimerization

The present work was carried out in order to elucidate the combined effects of the electron spin density on the ring carbons and the steric hindrance of the ring substituents upon the regio- and stereoselectivity of the dimerization of 3-carbamoyl- and 3-cyanopyridinyl radicals. To this purpose the composition of mixtures of diastereoisomeric dimers arising from the one-electron electrochemical reduction of several 3-carbamoyl and 3-cyano substituted pyridinium salts has been studied by 1H NMR spectroscopy. In some cases, single diastereoisomers have been isolated from mixtures by preparative HPLC.The results show that: (a) hindering steric effect of substituents at coupling sites prevails over electron spin density on coupling carbons in governing regioselectivity of 3-carbamoylpyridinyl radical dimerization; (b) large bulky N-ring substituents produce a significant shielding effect on the adjacent dimerization site; (c) the relative amounts of diastereoisomers in the mixtures of 4,4′- and 6,6′-linked dimers indicate that the dimerization process is largely stereoselective; (d) otherwise, nearly equal amounts of 4,4′- and 4,6′-linked dimers, and relative diastereoisomers as well, arise from the reduction of 3-cyano substituted pyridinium salts. This finding indicates that the presence of the carbamoyl substituent at the 3 position is a primary factor in inducing the regio- and stereoselectivity of pyridinyl radical dimerization.

Reactivity of pseudobases from pyridinium salts competition between hydrogen transfer and ring-opening reactions

Abstract The reactions of 3-cyano-1-methylpyridinium iodide (1a) and l-benzyl-3-cyanopyridinium chloride (1b) in aqueous NaOH have been studied over a range of concentrations from N 0.5 to N 2.5 with respect to the hydroxide ion, and with the concentrations of the reactants at different ratios. It has been found that the intermediate, not isolable pseudobases can undergo, depending on the experimental conditions, two competitive reaction channels, namely, a redox process leading to the dihydropyridines 3–5 and pyridones 2,6, and a ring-opening reaction leading to the pentadienenitrile 7, 2-imino-3-pyridinecarbaldehyde 8, and 2-(aryl)alkylamino-3-pyridinecarbaldehyde 9.

Choline pivaloyl ester strengthened the benefit effects of Tacrine and Galantamine on electroencephalographic and cognitive performances in nucleus basalis magnocellularis-lesioned and aged rats.

The aim of the present work was the assessment of the effects produced on the electroencephalographic (EEG) activity and the cognitive and memory performances of nucleus basalis magnocellularis (NBM)-lesioned or aged rats by the combined treatment with [2-(2,2-dimethylpropionyloxy)ethyl]trimethylammonium 2,2-dimethylpropionate (choline pivaloyl ester) (CPE) and the Cholinesterase inhibitors (ChEIs) Tacrine (THA) and Galantamine (GAL). Intraperitoneal administration of CPE combined with THA or GAL to both NBM-lesioned or aged rats, produced EEG desynchronisation, and a significant decrease in the energy of the total EEG spectrum and the lower frequency bands (delta 0.25-3 and theta 4-7 Hz) lasting many minutes. Furthermore, drug associations reversed in aged rats the scopolamine (0.2 mg/kg, i.p.)-induced increase in EEG power, slow waves and high-voltage spindle (HVS). Furthermore, the combined administration of CPE and Cholinesterase inhibitors in both NBM-lesioned or aged animals, improved performances in all behavioural tasks, enhancing object discrimination, increasing locomotory activity and alternation choice in T-maze, ameliorating retention in passive avoidance and decreasing escape latency in Morris water maze. In all test, AChEIs and CPE combinations proved to be more effective than CPE, THA or GAL given alone. In conclusion, the present work shows the ability of choline pivaloyl ester in strengthening the positive cerebral activity of THA and GAL.

Reactivity of 3-cyano-1-methylpyridinium iodide in aqueous ammonia or amine solutions

Reaction of 3-cyano-1-methylpyridinium iodide (1) with aqueous methylamine or ethylamine yields 1-alkyl-3-alkyliminomethyl-2-imino-1,2-dihydropyridines 8. The related aldehydes 9 on alkaline treatment give 2-alkylamino-3-pyridinecarbaldehydes 10. Experimental evidence, including the related reactions of 1 with aqueous ammonia and diethylamine as well, suggest a general reaction sequence initiated by the attack of the nucleophile, followed by ring-opening, transamination and ring-closing steps.