Molecular mechanism of action of chlorogenic acid on erythrocyte and lipid membranes

Figures & data

Figure 1. Percentages of different shapes of erythrocytes induced by CGA at 0.05 mg/ml (grey bar) and 0.01 mg/ml (black bar) concentration, control (white bar). Results are expressed as average ± SD (n = 5). Various erythrocyte shapes are given morphological indices as follows: Spherostomatocytes (−4), stomatocytes II (−3), stomatocytes I (−2), discostomatocytes (−1), discocytes (0), discoechinocytes (1), echinocytes (2), spheroechinocytes (3), spherocytes (4). Statistical analysis was conducted using the Dunnett test. Statistically significant results are denoted by **α = 0.01, *α = 0.05.

Figure 2. Shapes of unmodified erythrocytes (A) and modified with CGA (B) observed with electron microscope at 0.05 mg/ml concentration.

Table 1. Values of fluorescence anisotropy (A) of DPH and TMA-DPH probes for erythrocyte membranes and liposomes formed of erythrocyte lipids (RBCL) modified by CGA at 37 °C. Results are expressed as average ± SD (n = 5). The statistical significance of differences between average values of A and A_control was checked using the Dunnett test (<A_control).

	Anisotropy (A) ± SD
Membrane CGA [mg/ml]	Erythrocyte membranes	RBCL
DPH
Control	0.239 ± 0.002	0.219 ± 0.003
0.0050	0.239 ± 0.002	0.218 ± 0.003
0.0075	0.236 ± 0.011	0.212 ± 0.002
0.0100	0.240 ± 0.002	0.211 ± 0.002
0.0250	0.244 ± 0.010	0.212 ± 0.003
0.0500	0.248 ± 0.006	0.215 ± 0.003
TMA-DPH
Control	0.268 ± 0.001	0.244 ± 0.001
0.0050	0.264 ± 0.002	0.243 ± 0.002
0.0075	0.263 ± 0.002	0.246 ± 0.002
0.0100	0.263 ± 0.002	0.249 ± 0.001
0.0250	0.265 ± 0.003	0.256 ± 0.002*
0.0500	0.266 ± 0.003	0.266 ± 0.004*

DPH, 1,6-diphenyl-1,3,5-hexatriene; CGA, chlorogenic acid; RBCL, red blood cell lipids. Statistically significant results at α = 0.05 are denoted by *.

Figure 3. Values of generalized polarization (GP) of Laurdan probe for erythrocyte membrane (□) and RBCL (▪) modified with CGA at 37 °C. Results are expressed as average ± SD (n = 5). Statistical analysis was conducted using Dunnett’s test. Statistically significant results at: *α = 0.05, **α = 0.01.

Figure 4. Relative transmembrane capacity of BLM formed of erythrocyte lipids vs. time. Control symbol is (+) and in the presence of CGA at 0.001 mg/ml (•) and 0.01 mg/ml (♦). Statistical significance of differences between average values of capacitance for modified (C) and unmodified (C₀) BLM was checked using the Dunnett test. Statistically significant results at *α = 0.05 and **α = 0.01.

Table 2. Relative resistivity of black lipid membranes formed of erythrocyte membranes modified with CGA at different concentrations. Results are expressed as average ± SD (n = 5).

Relative transmembrane resistance
CGA [mg/ml]	R/R0
0	0.971 ± 0.049
0.001	0.898 ± 0.108
0.005	0.885 ± 0.069*
0.010	0.597 ± 0.048**

CGA, chlorogenic acid; Statistically significant results at *α = 0.05, **α = 0.01.