Interaction of 4'-methylflavonoids with biological membranes, liposomes, and human albumin

Abstract

The aim of the study was to compare the impact of three synthesized chemical compounds from a group of methylated flavonoids, i.e. 2'-hydroxy-4-methylchalcone (3), 4'-methylflavanone (4), and 4'-methylflavone (5), on a red blood cell membranes (RBCMs), phosphatidylcholine model membranes (PC), and human serum albumin (HSA) in order to investigate their structure-activity relationships. In the first stage of the study, it was proved that all of the compounds tested do not cause hemolysis of red blood cells and, therefore, do not have a toxic effect. In biophysical studies, it was shown that flavonoids have an impact on the hydrophilic and hydrophobic regions of membranes (both RBCMs and PC) causing an increase in packing order of lipid heads and a decrease in fluidity, respectively. Whereas, on the one hand, the magnitude of these changes depends on the type of the compound tested, on the other hand, it also depends on the type of membrane. 4'-Methylflavanone and 4'-methylflavone are located mainly in the hydrophilic part of lipid membranes, while 2'-hydroxy-4-methylchalcone has a greater impact on the hydrophobic area. A fluorescence quenching study proved that compounds (3), (4) and (5) bind with HSA in a process of static quenching. The binding process is spontaneous whereas hydrogen bonding interactions and van der Waals forces play a major role in the interaction between the compounds and HSA.

Scheme 1

Synthesis of 2′-hydroxy-4-methylchalcone (3), 4′-methylflavanone (4), and 4′-methylflavone (5).

Figure 1

Percentage of hemolysis of RBCs at different concentrations of the compounds (3), (4) and (5) after: (A) 1 h, (B) 24 h and (C) 48 h of incubation.

Figure 2

GP of Laurdan probe for (A) PC and (B) RBCMs membranes in the presence of compounds (3), (4) and (5). Anisotropy fluorescence of DPH probe for (C) PC and (D) RBCMs membranes in the presence of compounds (3), and (5). Means labeled with asterisk (*) are significantly (p < 0.05) different from control. PC- phosphatidylcholine liposomes, RBCMs-red blood cells membranes. C—control, (3) 2′-hydroxy-4-methylchalcone(4) 4′-methylflavanone, (5) 4′-methylflavone.

Figure 3

FTIR spectra of RBCMs and of RBCMs modified with flavonoids, concentration of compounds 50 µM; (A) C–H stretching regions, (B) carbonyl band, (C) phosphate and choline band of lipids, (D) amide bands.

Figure 4

FTIR spectra of PC liposome’s and of PC modified with flavonoids, concentration of compounds 50 µM; (A) C–H stretching regions, (B) carbonyl band, (C) phosphate and choline bands of lipids.

Figure 5

Emission spectra of HSA in the presence of various concentrations of (A) 2′-hydroxy-4 methylchalcone, (B) 4′-methylflavanone, (C) 4′-methylflavone and Stern–Volmer plots of F₀/F against concentration of (D) 2′-hydroxy-4 methylchalcone, (E) 4′-methylflavanone and (F) 4′-methylflavone. Control is marked black and consecutive spectra of the studied compounds (marked color) are in the following concentrations 1, 2, 4, 6, 8, 10, 12 and 14 µM, λ_ex = 280 nm, T = 310 K.

Figure 6

The plots of log(F₀ − F)/F versus log[c] of (A) 2′-hydroxy-4 methyl chalcone, (B) 4′-methylflavanone, (C) 4′-methylflavone at 310 K and van't Hoff plot for temperature dependence of K_b obtained from HSA fluorescence quenching by (D) 2′-hydroxy-4 methyl chalcone, (E) 4′-methylflavanone and (F) 4′-methylflavone.