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The paper describes the use of liposomes and erythrocytes membrane as a real membrane models to evaluate the potential benefits of several plants extracts and two flavones in relation to lipid peroxidation. The antioxidant behaviour of the plant extracts from pine (Pinus silvestris L), hawthorn (Crataegus oxyacantha L, two extracts: from hawthorn’s leaves-l and bark-b), evening primrose (Oenothena paradoxa Hudziok – three extracts differ in procyanidins content P1, P2 and P3) and rosemary (Rosmarinus officinalis, as a standard for extracts) and flavones of baicalin and rutin have been studied. The results obtained showed that the studied extracts exhibited differentiated, dose-dependent antioxidant activity against phospatidylcholine liposomes (rosemary>pine≈hawthorn-l>hawthorn-b≈P1≈P2≈P3; statistically significant differences were observed between the extracts at p≤0.05) and erythrocyte membranes (rosemary≥hawthorn-b≈hawthorn-l>P1≈pine>P2≈P3) when the oxidation was induced by UV-C radiation. They also reduce the oxidation of liposomes and erythrocyte membrane when its oxidation was induced by 2,2’-azobis(2-amidinopropane) dihydrochloride (pine≥P1≈P2≈rosemary≈P3 in the case of liposomes and rosemary>> pine≥P1≈P2≥P3 in a case of erythrocyte). Moreover, the results of the study show that baicalin is characterised by high inhibition ability towards liposome PC peroxidation, as well as towards erythrocyte ghosts, when oxidation was initiated by UV radiation. However, at the same experimental conditions, the inhibitory capacity of rutin was about 7-8 times weaker. The presence of cholesterol in liposome membrane decreased the level of membrane peroxidation but do not influenced on the antioxidant activity of hawthorn extract.
Celem pracy było określenie właściwości przeciwutleniających wybranych ekstraktów roślinnych (dziurawca, gryki, głogu i kocanki) oraz ich ważnych składników polifenolowych w stosunku do błon liposomów fosfatydylocholinowych (PC) utlenianych promieniowaniem UVC. Określono także właściwości przeciwrodnikowe tych substancji w stosunku do wolnego rodnika 2,2-difenylo-1-pikrylohydrazylu (DPPH•) oraz stałe asocjacji z błoną z zastosowaniem sondy DPH (l,6-difenylo-l,3,5-heksatrienu). Wykazano następującą relację aktywności przeciwutleniającej ekstraktów wyrażoną parametrem IC50PC: głóg – kora (21,69 mg/l) > dziurawiec (25,45 mg/l) > gryka – łuski (30,33 mg/l) > głóg – liście (32,52 mg/l) > gryka – łęciny (37,47 mg/l) > kocanka (115,06 mg/l). Aktywność przeciwutleniająca wybranych składników polifenolowych zmieniała się natomiast w następujący sposób: kwercetyna (0,53 mg/l) > epikatechina (29,19 mg/l) > kwas chlorogenowy (62,59 mg/l) > rutyna (78,97 mg/l). Dodatnia korelacja aktywności przeciwutleniającej badanych ekstraktów oraz związków fenolowych z aktywnością przeciwrodnikową (r ≥ 0,94) wskazuje, że mechanizm ich przeciwutleniającego działania polega na neutralizowaniu wolnych rodników. Kwercetyna spośród badanych flawonoidów wykazywała najwyższą aktywność przeciwutleniającą, wysoką aktywność przeciwrodnikową (IC50 DPPH• = 2,51 mg/l) oraz wysoką stałą asocjacji z błoną liposomów (Ka = 11,23·10³ l/mg). Możliwość głębokiego zakotwiczenia się molekuł kwercetyny w błonie liposomów fosfolipidowych ogranicza wnikanie wolnych rodników do wnętrza błony co wzmacnia jej skuteczność przeciwutleniającą.
The degree of dependence of a lipid bilayer’s surface properties on its conformational state is still an unresolved question. Surface properties are functions of molecular organization in the complex interfacial region. In the past, they were frequently measured using fluorescence spectroscopy. Since a fluorescent probe provides information on its local environment, there is a need to estimate the effect caused by the probe itself. In this paper, we address this question by calculating how lipid head-group orientation effects the fluorescence intensity of Fluorescein-PE (a probe that is sensitive to surface potential). In the theoretical model assumed the lipid bilayer state and the interactions between the charged fluorescent probe and the surrounding lipid molecules was evaluated. The results of this theoretical analysis were compared with experimentally obtained data. A lipid bilayer formed from DPPC was chosen as the experimental system, since it exhibits all the major conformational states within a narrow temperature range of 30°C - 45°C. Fluorescein-PE fluorescence intensity depends on local pH, which in turn is sensitive to local electrostatic potential in the probe’s vicinity. This local electrostatic potential is generated by lipid head-group dipole orientation. We have shown that the effect of the probe on lipid bilayer properties is limited when the lipid bilayer is in the gel phase, whereas it is more pronounced when the membrane is liquid-crystalline. This implies that Fluorescein-PE is a good reporter of local electrostatic fields when the lipid bilayer is in the gel phase, and is a poor reporter when the membrane is in the liquid-crystalline state.
We studied the effects of UV radiation on the degree of phosphatidylcholine (PC) liposome membrane oxidation in the presence of such toxic organometallic compounds (TOC) as diphenyltin dichloride (DPhT), triphenyltin chloride (TPhT), dibutyltin dichloride (DBT), tributyltin chloride (TBT), triphenyllead chloride (TPhL) and tributyllead chloride (TBL). PC liposome oxidation was also investigated in the presence of quercetin (Que) and equimolar mixtures of Que and TOC in order to determine the protective properties of this natural antioxidant with respect to liposome membrane oxidation induced by UV light. Concentration of the compounds studied was 10 µM. The degree of liposome oxidation was measured using the TBARS (Thiobarbitutric Reactive Substances) test. The sequence thus obtained of relative induction of PC oxidation due to TOC was as follows: TBT > TPhT = DBT > DPhT = TPhL > TBL. The results of studies with Que indicate its high efficacy in the antioxidative action of equimolar mixtures of Que and TOC. From our study it follows that quercetin forms complexes both with phenyl- and butyl- tin and lead compounds. A high antiradical (towards DPPH radical) activity of the associates with respect to the activity of quercetin alone was also found. This result can partly explain the antioxidative properties of Que in the studied solution, connected with their antiradical action and chelating posibility towards the organometallic compounds studied. A factor that differentiates the antioxidant activity of the complexes Que-TOC is probably their differentiated location in the liposome membrane bilayer. A factor that differentiates the antioxidant activity of the complexes Que-TOC is probably their differ­entiated location in the liposome membrane bilayer.
The nature of the interaction between some strong antifungal compounds and model membranes was studied. It was found that these compounds (N-dodecyl-N,N-dimethyl-N-/3- (β-methyl-β-nitrovinyl 1-6-methoxybenzyl/ammonium chloride and its derivatives) significantly change the properties of liposomes, planar lipid bilayers (BLM) and erythrocytes. The changes observed (i.e., release of praseodymium ions from liposomes and gradually diminished stability of BLM) in the presence of the compounds studied, as well as hemolysis of red blood cells, were time-dependent. It was also found that they depended on the polar head structure of the compounds.
The modyfication of model lipid membranes by amphiphilic ammonium salts of the general formulae |(CH3)3NCH2COOC12H25|+ • Cl (V-12), |(CH3)2C12H25NCH2COOC2H4Cl|+ • Cl- (V-A) and |(CH3)3NCH2COOC2H4Cl|+ • Cl- (V-B) were studied. It was found that the efficiency of liposomal and planar lipid membranes (BLM) modified by these salts can be sequenced as follows: V-12, VA, and VB. It was also found that sulpate-ion permeation through liposomal membranes modified by the salts studied was temperature-dependent and changed significantly in the case of membranes modified with V-12 and V-A, while the rate constant was only slightly dependent on temperature for liposomes modified by V-8 salt.
The effect of some biologically active amphilic quaternary ammonium chlorides (QA) on properties of bimolecular planar membranes (BLM) formed of lipids extracted from bovine erythrocytes was studied. Ammonium salts differed in the alkyl chain length or in the non-methylene group incorporated between the hydrocarbon chain and the polar head of a molecule. It was found that in the case of a series of QA of general structure (CH3)3N+CH2OCnH2n+1 (n=4, 10, 12, 14, 16 and 18) there exists a maximal interaction between QA and BLM for QA with n=16. It was also found that the introduction of different functional groups, between the nitrogen atom and the alkyl chain of the molecule significantly changed the interaction between ammonium salts and lipid membranes. The observed changes are explained in terms of changed amphiphilic properties of the molecule.
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