QUANTUM CHEMICAL INTERPRETATION OF THE REACTION BETWEEN L-ASCORBIC ACID AND FORMALDEHYDE AND ITS BIOLOGICAL SIGNIFICANCE

We have recognized a new nucleophilic addition reaction between L-ascorbic acid and formaldehyde in biological circumstances (pH 7.4, T = 300 K). UV spectroscopic investi- gation showed that the C2=C3 double bond of ascorbic acid rapidly disappears during the reaction, verifying that the addition breaks the double bond. On the other hand, 13C NMR studies have proved that on atom C2 an additional C-C single bond is formed. Ex- act interpretation of the reaction mechanism is possible only by using quantum chemical argumentations. Ab initio calculations show that in ionized (anionic) form of L-ascorbic acid in water solution the Mulliken excess atomic charge is strongly negative on atom C2 while C1 and C3 exhibit significant positive net charges. Coulombic attraction arranges formaldehyde's C=O double bond over the C2=C3 double bond of ascorbic acid and the interaction of the two loose π-electron systems causes forming of a new σ C-C bond. The described reaction plays a protective role in biological systems, since in this way the toxic carcinogenic formaldehyde can be eliminated

Role of arginine and its methylated derivatives in cancer biology and treatment

Both L-arginine supplementation and deprivation influence cell proliferation. The effect of high doses on tumours is determined by the optical configuration: L-arginine is stimulatory, D-arginine inhibitory. Arginine-rich hexapeptides inhibited tumour growth. Deprivation of L-arginine from cell cultures enhanced apoptosis. The pro-apoptotic action of NO synthase inhibitors, like NG-monomethyl-L-arginine, is manifested through inhibition of the arginase pathway. NG-hydroxymethyl-L-arginines caused apoptosis in cell cultures and inhibited the growth of various transplantable mouse tumours. These diverse biological activities become manifest through formaldehyde (HCHO) because guanidine group of L-arginine in free and bound form can react rapidly with endogenous HCHO, forming NG-hydroxymethylated derivatives. L-arginine is a HCHO capturer, carrier and donor molecule in biological systems. The role of formaldehyde generated during metabolism of NG-methylated and hydroxymethylated arginines in cell proliferation and death can be shown. The supposedly anti-apoptotic homozygous Arg 72-p53 genotype may increase susceptibility of some cancers. The diverse biological effects of L-arginine and its methylated derivatives call for further careful studies on their possible application in chemoprevention and cancer therapy

FORMATION OF BURST CHEMILUMINESCENCE, EXCITED ALDEHYDES, AND SINGLET OXYGEN IN MODEL REACTIONS AND FROM CARCINOGENIC COMPOUNDS IN RAT LIVER S9 FRACTIONS

It is shown that in physiological circumstances (T=298 K, pH=7.4) various aldehydes (formaldehyde, acetaldehyde, glyoxal, methyl-glyoxal, etc.) can be activated by hydrogen peroxide (H202) in the presence of the ε-amino group of L-lysine, with simultaneous formation of singlet oxygen (1O2 ) and chemiluminescence. The activated aldehydes are in an excited state and have radical structure causing extremely high reactivity and a possible attack of cellular components like proteins, amino acids, RNA and DNA. In the reactions methylated, formylated, acetylated, etc. compounds are formed as well. Activated aldehydes can also be formed during the biological oxidation of different nitrosamines (DMN, DEN, N-nitroso-morpholine) and formaldehyde-hydrazine adduct. It is proved that in rat liver S9 fractions these activated (excited) aldehydes, especially formaldehyde, have been formed with a simultaneous burst chemiluminescence emission and appearance of singlet oxygen. In rat liver S9 fractions excited formaldehyde can be liberated only after 30 min preincubation time from dimethyl-nitrosamine (DMN) and monomethyl-hydrazine (MMH). On the other hand, excited formaldehyde is liberated immediately from hydrazine-formaldehyde-adduct (HZ/FORM) and isonicotinic acid-hydrazide-formaldehyde adduct (INH/FORM), i.e. no preincubation is necessary

PROTEIN AND AMINO ACID CHEMICAL STUDIES

A cross-linking procedure not deteriorating the physico-chemical characteristics of wool has been elaborated. The thermal destruction processes in wool have been clarified. Solutions have been found for accelerating and inhibiting reactions taking place between wool and formaldehyde. The reactions between L-Iysine and formaldehyde as well as L-arginine and formaldehyde have been identified. Procedures have been elaborated for the practical utilization of the reaction products

INVESTIGATION OF CYANO-METHYLATION REACTION BY CYANO-HYDRINE AND ITS DETERMINATION IN TOBACCO-SMOKE. ( STRECKER-REACTIONS )

The results of our research work [1-7] of several years aimed at the binding of the unhealthy components of tobacco smoke directed our attention to the analysis and identification of the unknown peak observed during the high sensitivity radiochromatographic analysis of tobacco smoke condensates.It was a fruitless effort to remove the formaldehyde, the ac- etaldehyde and hydrogen cyanide present in tobacco smoke, or even to try to eliminate them separately, during the analysis the new, unknown peak appeared again and again on the radiochromatogram. During our research work we stated, that we were facing the Strecker reaction [8-9], known since 1850, that is, with the formation of cyanohydrine of the aldehydes and hydrogen cyanide present in tobacco smoke, corresponding to the new peak. Cyanohydrines to react quickly and energetically with basic aminoacids, during which reaction cyanomethyl derivatives are being formed. This reaction is proceeding also under physiological circumstances with the basic amino groups of proteins, contributing to the development of respiratory and cardiovascular insufficiencies. By means of model reactions and using solutions of tobacco smoke gases the process of the reaction was proved in a primary way, also its circumstances having been cleared. By radiochromatographic measurements it was proved that the labelled L-lysine-6 H is being cyanomethylated by formaldehyde-cyanohydrin at an extraordinary speed and cyanomethylation is complete within a very short time (5 min). The tests carried out on formaldehyde- C, cyanide- C and inactive L-lysine led to similar results, with a yield 93.9 percent within 5 min in all these cases. It was proved also by 13C NMR, 13C NMR DEPT and 1H NMR spectroscopic measurements that inactive L-lysine is cyanomethylated by cyanohydrine in an extraordi- narily energetic and quick reaction, according to the reaction mechanism supposed by us. Our results obtained during the radiochromatographic analysis of tobacco smoke conden- sate agreed in all respects with the model test results. That is, the cyanohydrine formation is proceeding very quickly in the tobacco smoke too (by the reaction of aldehydes with hydrogen cyanide contained by tobacco smoke) which can be unequivocally traced back to the Strecker-reaction recognized 140 years ago. The mechanism of the reaction was described by LAPWORTH in 1903. [10)

ESSENTIAL DIFFERENCES IN SPONTANEOUS REACTION OF L-LYSINE AND L-ARGININE WITH FORMALDEHYDE AND ITS QUANTUM CHEMICAL INTERPRETATION

It has been recently reported by us that spontaneous reaction between formaldehyde and L-lysine yields N-methylated products at room temperature. In this paper we discussed this reaction extending to L-arginine. While the reaction between formaldehyde and L-lysine produces different N-methylated derivatives, no methylated only NG-hydroxymethylated derivatives can be observed in the reaction mixture between formaldehyde and L-arginine. The difference in the reaction between L-lysine and L-arginine with formaldehyde can be explained with the different nucleophilicity of the amino group in lysine and that of the imino group in arginine examined by the electrostatic potential of N atoms by the ELPO quantumchemical calculation method of Náray-Szabó