Krzysztof Bielecki

Toxicity and model membrane modifying properties of organolead compounds

Department of Botany and Plant Physiology, Agricultural University, Cybulskiego 32, 50–205 Wroclaw,PolandThe influence of trialkylleads on haemolysis ofred blood cells (RBCs), growth of Spirodelaoligorrhiza and stability of planar lipid mem-branes (PLMs) at different pH of solution hasbeen studied. The results obtained show that theefficiency of trialkylleads (methyl-, ethyl-, pro-pyl- and butyl-lead chlorides) in modifying thephysiological and mechanical properties of theobjects studied depended both on pH of solutionand hydrophobicity of the compounds. Namely,it was found that this efficiency increased withpH of solution. The most significant increase wasobserved in PLM experiments. Also, the hydro-phobicity of trialkylleads influenced the proper-ties mentioned. The more hydrophobic a com-pound the greater was its haemolytic toxicity.The same applies to the physiological toxicityof the compounds, whose measure was 50%inhibition of plant growth. Generally, thesequence of modifying possibilities of the com-pounds studied at any pH of the solution was thefollowing:tributyllead >tripropyllead >triethyllead >trimethylleadA possible mechanism of the interaction oforganolead species with model and biologicalmembranes is discussed. Copyright # 2001John Wiley & Sons, Ltd.Keywords: triorganolead chlorides; erythrocytehaemolysis; physiological toxicity; planar lipidmembranes destabilization; interaction mechan-ism.

Antioxidant Activity of Extracts from Apple, Chokeberry and Strawberry

The aim of the study was to determine the protective action of extracts from apple, strawberry and chokeberry with respect to linoleic acid and the biological membrane exposed to oxidation induced by physicochemical factors. The activity of the extracts was determined by measuring inhibition of lipid oxidation in red blood cell membrane, induced with UVC radiation and the AAPH radical. The protective effect of the extracts was essayed fl uorimetrically and spectrophotometrically. These results together with the ones obtained earlier explain the mechanism of the interaction between the extracts and the red blood cell membrane. The mechanism consists in the incorporation into the membrane and screening the cell against oxidation. The results indicate that the extracts possess very good antioxidant properties, since at the highest concentrations used (0.1 mg/mL) they protect the biological membranes almost entirely against oxidation. Among the extracts studied the best antioxidant properties were exhibited by the apple fruit, which gave 80% or 100% protection of the membrane at 0.05 mg/mL concentration of dry matter, for UVC and AAPH inductors, respectively.

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.

Binary Mixtures of (N-phosphonomethyl)-glycine with New Aminophosphonates

Abstract The potential biological activity of binary mixtures of some new organophosphorous compounds, aminoalkane- and aminofluorenephosphonates, with (N-phosphonomethyl)-glycine (glyphosate, PMG) was studied. The inhibition of growth of wheat (Triticum aestivum) induced by individual compounds and their equimolar mixtures with PMG was a measure of that activity. The experiments were expected to show if the new compounds exhibited good biological activity to be used for agrochemical applications and if this activity can be improved when they are used in mixtures with glyphosate which is the active component of the well-known herbicide Roundup. The results obtained show that aminofluorenephosphonates inhibited wheat growth when used in micromolar concentrations. Thus, their efficiency can be compared to that of PMG. The efficiency of aminoalkanephosphonates was one order of magnitude weaker. The measure of the efficiency was the effective concentration inhibiting wheat growth by 50% (EC50). The most demanded interaction, i.e., a synergistic was observed for only one of binary mixtures of the compounds studied with PMG. Mostly they showed antagonistic or strong antagonistic interactions. Some of them were of the additive type. Such results exclude the possibility of potential use of all the compounds studied in binary mixtures with phosphonomethylglycine, especially as the mentioned synergistic interaction found was rather weak. The influence of structural features of aminophosphonates on the results obtained is discussed.

Aminophosphonate-Induced Changes of Betacyanine and Ionic Efflux

Betacyanine and ionic leakage from red beet (Beta vulgaris ssp. L. rapacea) roots and lilac (Syringa vulgaris L.) leaves under the influence of new aminophosphonates were studied by spectroscopic and conductometric methods. It was found that the leakage of dye or electrolytes depended both on the concentration of the compounds used and their structural features. The results compared to those obtained for the well known herbicide Buminafos® (dibutyl 1-butylamino-1-cyclohexanephosphonate) enabled to conclude that some of the compounds studied exhibited comparable or better activity than this herbicide. That makes them potentially good herbicides. It is possible that the effects observed are the result of action on cell membranes of the tissues used. The possible role of the structural features of aminophosphonates in this action is discussed.

Influence of Triphenyllead Chloride on Biological and Model Membranes

Abstract The physiological and hemolytic toxicities of triphenyllead chloride (TPhL) as well as its modyfying influence on model lipid membranes were studied. The experiments allowed the determination of TPhL concentrations causing 50% inhibition of growth of Spirodela oligorrhiza, Lemna minor and Solvinia natans (EC50), 100% hemolysis of pig erythrocytes (C100) and destabilization of planar lipid membranes (CC). Also, fluidity of erythrocyte ghosts was measured by fluorescence technique and osmotic sensitivity of erythrocytes to the presence of TPhL. All parameters studied were found to be dependent on pH, of experimental solutions and the concentration of TPhL. Acidic conditions increased EC50, C100 and CC concentrations of TPhL. Fluorescence and osmotic measurements showed that osmotic stability and fluidity decreased with increasing trimethyllead concentration. A possible mechanism of TPhL toxicity is discussed. It is assumed that TPhL is interacting with the lipid phase of the models used. It is also assumed that there may exist various, ionic and nonionic, forms of TPhL as a result of its speciation under different experimental conditions. These species, due to their differentiated lipophilicity, may exert different effects on the model membranes studied.

Assessment of Potential Application of Binary Mixtures of 2,4-D with Novel Aminophosphonates

A series of new aminoalkane- and aminofluorenephosphonates was synthesized for agrochemical application. The particular compounds had different alkyl substituents at the carbon, nitrogen and phosphorus atoms. Their pesticidal activity was checked by applying various experimental methods. These included the measurements of compounds’ potency: to inhibit growth of cucumber and germination of white mustard seeds, to influence on the membrane potential of algae and to damage human erythrocyte membranes resulting in hemolysis. All the aminophosphonates were also used in equimolar binary mixtures with the well-known herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), to check, if using such mixtures, the biological efficiencies found for particular compounds could be enhanced due to interactions between aminophosphonates and 2,4-D. The results demonstrated, that depending on the structural features of the compounds, the final effects differed from antagonistic, through additive to the most promising synergistic ones. However, the type of interaction between 2,4-D and the compounds studied found in different experiments was somewhat different. In order to estimate those effects various statistical methods were used (toxic unit method, isobole method).