H. Kleszczyńska

Erythrocyte Hemolysis by Organic Tin and Lead Compounds

The effect of trialkyllead and trialkyltin on pig erythrocyte hemolysis has been studied and compared. The results of experiments showed that the hemolytic activity of organoleads increases with their hydrophobicity and follows the sequence: triethyllead chloride < tri-npropyllead chloride < tributyllead chloride. And similarly in the case of organotins: triethyltin chloride < tri-n-propyltin chloride < tributyltin chloride. Comparison of the hemolytic activity of organoleads and organotins indicates that the lead compounds exhibit higher hemolytic activity. The methods of quantum chemistry allowed to determine the maximum electric potential of the ions R3Pb+ and R3Sn+, and suggest a relationship between the potential and toxicity.

Protective Effect of Quaternary Piperidinium Salts on Lipid Oxidation in the Erythrocyte Membrane

Abstract A new series of amphiphilic compounds with incorporated antioxidant functional group has been investigated. Piperidinium bromides, differing in the alkyl chain length (8, 10, 12, 14 and 16 carbon atoms in the chain) were synthesised to protect biological and/or model membranes against peroxidation and following negative consequences. Their antioxidant activity was studied with erythrocytes subjected to UV radiation. The salts used inhibited lipid oxidation in the erythrocyte membrane. The degree of this inhibition depended on the alkyl chain length of the bromide used and increased with increasing alkyl chain length. A comparison of the results obtained for piperidinium bromides with those obtained for the widely used antioxidant 3,5-di-t-butyl-4-hydroxytoluene (BHT) revealed that only two shortest alkyl chain salts were less efficient than BHT in protecting erythrocyte membranes. A similar comparison with antioxidant efficiency of flavonoids extracted from Rosa rugosa showed that they protected the membranes studied more weakly than the least effective eight-carbon alkyl chain piperidinium bromide. The three compounds of longest alkyl chains were the most active antioxidants. Their activities did not differ significantly.

Inhibition of Lipid Peroxidation in the Erythrocyte Membrane by Quaternary Morpholinium Salts with Antioxidant Function

Abstract The antioxidant activity of five new compounds from a group of quaternary ammonium salts with antioxidant function has been investigated. The effect of the compounds on the degree of lipid oxidation in the erythrocyte membrane subjected to UV radiation was studied. It was found that all the salts used decreased oxidation of the lipids in the erythrocyte membrane. The antioxidant activity of the ammonium salts studied increased with their alkyl chain length. Three compounds with the longest alkyl chains were the most active antioxidants, and their antioxidant properties were comparable to those of a flavonoid extracted from haw thorn. The antioxidant effectiveness of the ten-carbon alkyl chain compound was comparable with that of the known antioxidant 3,5-di-t-butyl-4-hydroxytoluene (BHT). The least effective antioxidant studied proved to be the eight-carbon alkyl chain compound. The effect of these compounds on fluidity of the erythrocyte membrane has been studied, and for all an increase of fluidity of the membrane was observed. The changes in erythrocyte ghosts fluidity depended both on concentration and type of compound. A fluorimetric study also indicated that rigidity of the erythrocyte membrane increased with degree of its oxidation, but with antioxidants present membrane rigidity increased less.

Aminoethyl esters of fatty acids as model lysosomotropic substances.

Conclusions1.The activity of the DM-n compounds is chain length and pH dependent; the highest activity being observed for 11–13 carbon atoms chain.2.The highest biological activity was observed at pH above 7, which is characteristic for lysosomotropic agents.3.Hemolytic experiments conformed the lysosomotropic character of DM-n. The free amine (DMF-11) can pass through membranes and at lower pH can interact with the vacuolar membrane.4.In yeast, DM-n mobilized respiration of endogenous substrates.5.In yeast protoplasts treated with DM-n, temporary increase in Ca2+ uptake (characteristic for apoptosis) takes place. This indicates that the compounds have a killing effect.

N-substituted aminoethylesters of fatty acid as lysosomotropic substances

[1] SUBIK J., KOLAROV J., KOVA(~ L.: BiochemBiophys.Res.Commun. 49, 192-198 (1972). [2] HAPALA 1.: Biochem.Biophys.Res.Commun. 159, 612-617 (1989). [3] HAPALA I., HUNAKOVA A., GERZANIt~OVA G., BUTKO P.: Biochim.Biophys.Acta 1190, 40-42 (1994). [4] DRGO~q T., ~ABOVA L., NELSON N., KOLAROV J.: FEBS Lett. 289, 159-162 (1991). [5l HAPALA 1,, HUNAKOVA A.: Folia Microbiol. 41, 95-96 (1996). [6] ZINSER E., PALTAUF F., DAUM G.: J.Bacteriol. 175, 2853-2858 (1993).

Antioxidative activity of new N-oxides of tertiary amines: membrane model and chromogen studies.

Potential antioxidative activities of three series of newly synthesized N-oxides were studied. Individual components in each of the series differed in the lipophilicities and number of free radical scavenging groups. Various methods were used to determine their antioxidative efficiencies: Prevention of erythrocyte membrane lipid oxidation induced by UV irradiation and chromogen experiments in which antioxidative efficiencies of compounds were compared to that of the standard antioxidant Trolox (a water-soluble vitamin E analogue). Additionally, some hemolytic (pig erythrocytes) and differential scanning calorimetry (DSC) measurements were performed to determine a mechanism of the interaction between membranes and N-oxides. It was found that N-oxides, especially those of long alkyl chains (> C12H25), readily interacted with both, erythrocyte and liposomal membranes. No marked differences were found in their protection of erythrocytes against oxidation. In most cases inhibition of oxidation changed between 15% and 25%. Still, it was far better than in chromogen experiments where suppression of free radicals reached 20% in the best case. It may be concluded that antioxidative capabilities of N-oxides are moderate. Studies on the interaction mechanism showed that incorporation of particular compounds into model membranes varied. Hemolysing activities of compounds increased with the elongation of the alkyl chain but differed for corresponding compounds of particular series indicating that lipophilicity of compounds is not the only factor determing their interaction with erythrocyte membranes. DSC experiments showed that N-oxides, upon incorporation into 1,2-dipalmitoyl-3-snphosphatidylcholine liposomes, shifted the subtransition (Tp) and the main transition (Tm). The shifts observed depended on the alkyl chain length. The effects differed for each series. It seems that in the case of long alkyl chain compounds the domain formation may take place. Generally, the decrease of Tm was greatest for the same compounds that exhibited the best hemolytic efficacy. The same conclusion concerns the decrease of cooperativity of the main transition and the observed changes suggest an increase in membrane fluidity. Both, erythrocyte and DSC experiments seem to indicate that compounds of particular series incorporate in a somewhat different way into membranes.

Biological activity of new N-oxides of tertiary amines.

Abstract Potential biological properties of newly synthesized single and double alkyl chain N-oxides of tertiary amines (NTA) were studied. Individual compounds in each of the series had alkyl chains of different length. Various experiments were performed to determine a mechanism of the interaction between NTA and model and biological membranes. These were measurements of hemolytic efficiencies of NTA (pig erythrocytes), their influence on the transition temperatures (DPPC liposomes), on potassium leakage from cucumber, its growth and chlorophyll content (Cucumis sativus cv. Krak F1), and on the resting membrane potential in alga cells (Nitellopsis obtusa). Also, prevention of erythrocyte membrane lipid oxidation induced by UV irradiation was studied. Potential antioxidative properties of NTA were additionally tested in radical chromogen (ABTS●+) experiments in which antioxidative efficiencies of NTA were compared to that of the standard antioxidant Trolox. It was found that NTA readily interacted with erythrocyte membranes. Their hemolyzing efficiency increased with the alkyl chain length. Slightly more intensive interaction was found for double alkyl chain compounds. Similar results were obtained in DSC experiments, where incorporation of NTA into liposomal membranes shifted the main transition temperatures and caused a broadening of the main transition peaks depending on the alkyl chain length. Double alkyl chain compounds were also found more efficiently influencing the growth of cucumber. Influence of NTA on the resting membrane potential of algae cells was not quite following the alkyl chain length rule found in erythrocyte and liposome experiments. Also potassium leakage and chlorophyll content determined in physiological experiments were not following the increase of lipophilicity of compounds. Most efficiently influencing those parameters were NTA having shorter alkyl chains, and efficiencies of single alkyl chain compounds were evidently stronger. Both methods used to test the antioxidative properties of NTA showed that they depended on the alkyl chain lengths of compounds within each series, but double alkyl chain ones exhibited markedly greater efficiency.

Antioxidative Properties of Pyrrolidinium and Piperidinium Salts

Two series of pyrrolidinium (PYA-n) and piperidinium (PPPA-n) bromides with incorporated antioxidant function were synthesized. Both have hydrocarbon chains with odd number of the carbon atoms (n) ranging between 7 and 15. Pig erythrocytes (RBC) were used to study antioxidant activity of these compounds. They were incorporated into RBC membranes in sublytic (micromolar) concentrations and RBC were then subjected to UV radiation. It was found that all the salts used protected erythrocyte membranes against oxidation of membrane lipids. This protection increased with hydrocarbon chain length. Such effect may be the result of an incorporation of particular compounds to different depths into the lipid phase of RBC membrane depending on their chain length. Such possibility was checked by studies on fluidity changes induced by the compounds studied in ghost membranes by fluorimetric measurements. The measurements showed that pyrrolidinium bromides were slightly more effective in a protection of erythrocytes than the corresponding piperidinium ones. The possible reason of such behaviour may be the difference in lipophilicity between piperidine and pyrrolidine rings

Hemolytic activity of aminoethyl-dodecanoates.

Abstract The effect of new lysosomotropic compounds on red blood cell hemolysis and erythrocyte membrane fluidity has been investigated. In earlier studies it was shown that the compounds inhibit the growth of yeast and plasma membrane H+-ATPase activity. The study was per formed with eight aminoethyl esters of lauric acid variously substituted at nitrogen atom. Esters of dodecanoic acid were chosen for study because at that chain length dimethylaminoethyl esters showed maximum activity. The hemolytic activity of the substances studied exhibits diversified activity in their interaction with the erythrocyte membrane: they differ in hemolytic activity and affect membrane fluidity differently. Erythrocyte membrane fluidity changes under the effect of those compounds which possess highest hemolytic activity. The hemolytic activity of the aminoesters investigated was found to follow a sequence that depended on basicity (i.e. ability of the protonated form formation) of the compound and its polar head group size. The most active are the compounds that possess not more than four carbon atoms substituted at nitrogen and highest pKa value.

Functionalized Permeatoxins: Derivatives of Dihydrocinnamic Acid with Antioxidant Function

[1] OBLAtK E., LACHOWlCZ T.M., WITEK S.: Folia Microbiol. 41, 116-118 (1996). [2] LACHOW1CZ T.M., WITEK S., LUCZY?;/SKI J., WITKOWSKA R., KLESZCZYlqSKA H., PRZESTALSK1 S.: Folia Microbiol. 42, 231-232 (1997). [3] GOFFEAU A., AMORY A., DUFOUR J.P.: Methods in Enzymology 157, 513-528 (1988). [4] GOFFEAU A., SLAYMAN C.W.: Biochem.Biophys.Acta 639, 197-223 (1981). [5] GALZY P., SLONIMSKI P.P.: C.R.Acad.Sci. 245, 2556-2559 (1957).