Influence of Plantaginaceae species on E. coli K12 growth in vitro: Possible relation to phytochemical properties

Figures & data

Table 1. Phytochemical content in Plantago and Veronica leaves (mg/g).

Species	P. maxima	P. lanceolata	P. major	V. teucrium	V. spicata	V. incana
Flavonoids	3.25 ± 0.68	1.24 ± 0.48	1.13 ± 0.33	1.36 ± 0.25	2.19 ± 0.44	2.43 ± 0.41
Iridoids	41.54 ± 1.12	5.82 ± 0.19	4.47 ± 0.25	6.41 ± 0.24	8.22 ± 0.20	9.87 ± 0.30
Tannins	9.37 ± 0.29	8.57 ± 0.15	4.15 ± 0.18	3.16 ± 0.16	4.06 ± 0.14	3.16 ± 0.12

Table 2. Chromatographic characterization of flavonoids, phenylcarboxylic acids, and iridoids found in P. maxima leaves.

	Rf		Color
Parameters	BAAW 4:1:2	15% СН3СООН	UV-light	UV-light + NH3	AlCl3	Hepfner reagent	Compound
Flavonoids and  phenylcarboxylic acids	0.35	0.15	Lavender	Yellow	Daffodil	–	Cynaroside
	0.27	0.15	Lavender	Canary	Daffodil	–	n.d. (Flavonoid)
		0.13	0.26	Petunia	Yellow	Yellow	–	n.d. (Flavonoid)
	0.08	0.37	Petunia	Canary	Canary	–	n.d. (Flavonoid)
	0.13	0.49	Violet	Yellow	Yellow	–	n.d. (Flavonoid)
	0.21	0.47	Lavender	Yellow	Daffodil	–	n.d. (Flavonoid)
	0.50	0.32	Violet	Canary	Canary	–	Apigenin
	0.21	0.86	–	Blue	–	Orange-brown	n.d. (Flavonoid)
	0.36	0.84	–	Blue	–	Orange-brown	n.d. (Flavonoid)
	0.52	0.76	Blue	Yellow-green	–	Orange-brown	Neochlorogenic acid
	0.63	0.73	Blue	Bright yellow-green	–	Orange-brown	Chlorogenic acid
	0.49	0.02	–	Blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.77	0.88	Blue	Bright blue	–	Orange-brown	Caffeic acid
	Rf,		Trim–Hill reagent processing
Parameters	BAAW 4:1:2		Visible colors		Color in UV-light		Compound
Iridoids	0.15		Grey		Violet		Isocatalpol
	0.23		Dark grey		Violet		Catalpol
	0.35		Dark grey		Violet		Aucubin
	0.40		Grey		Violet		n.d. (Iridoid)
	0.45		Dark gray		Violet		Aucubin acetate
	0.56		Green-gray		Violet		n.d. (Iridoid)

n.d. – not defined.

Table 3. Chromatographic characterization of flavonoids, phenylcarboxylic acids, and iridoids found in P. lanceolata leaves.

	Rf		Color
Parameters	BAAW 4:1:2	15% СН3СООН	UV-light	UV-light + NH3	AlCl3	Hepfner reagent	Compound
Flavonoids and  phenylcarboxylic acids	0.24	0.16	–	–	Daffodil	–	n.d. (Flavonoid)
	0.39	0.11	–	Yellow	Daffodil	–	Cynaroside
		0.13	0.26	Violet	Yellow	Canary	–	n.d. (Flavonoid)
	0.40	0.31	Violet	Yellow	Canary	–	Luteolin
	0.40	0.46	–	Yellow	Daffodil	–	n.d. (Flavonoid)
	0.85	0.55	Violet	Yellow	Canary	–	n.d. (Flavonoid)
	0.5	0.03	Blue	Bright blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.63	0.03	Blue	Bright blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.21	0.86	Blue	Bright blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.36	0.84	Blue	Bright blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.47	0.82	Blue	Bright yellow-green	–	Orange-brown	Neochlorogenic acid
	0.64	0.78	Blue	Bright yellow-green	–	Orange-brown	Chlorogenic acid
	Rf,		Trim–Hill reagent processing
Parameters	BAAW 4:1:2		Visible colors		Color in UV-light		Compound
Iridoids	0.23		Dark grey		Violet		Catalpol
	0.35		Dark grey		Violet		Aucubin
	0.56		Green-grey		Violet		n.d. (Iridoid)

n.d. – not defined.

Table 4. Chromatographic characterization of flavonoids, phenylcarboxylic acids, and iridoids found in P. major leaves.

	Rf		Color
Parameters	BAAW 4:1:2	15% СН3СООН	UV-light	UV-light + NH3	AlCl3	Hepfner reagent	Compound
Flavonoids and  phenylcarboxylic acids	0.24	0.16	–	–	Daffodil	–	n.d. (Flavonoid)
	0.37	0.11	–	Yellow	Daffodil	–	Cynaroside
		0.13	0.26	Violet	Yellow	Canary	–	n.d. (Flavonoid)
	0.40	0.31	Violet	Yellow	Canary	–	Luteolin
	0.40	0.46	Violet	Yellow	Daffodil	–	n.d. (Flavonoid)
	0.51	0.55	Pale blue	Bright blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.53	0.73	Blue	Yellow-green	–	Orange-brown	Neochlorogenic acid
	0.63	0.78	Blue	Bright yellow-green	–	Orange-brown	Chlorogenic acid
	0.69	0.79	Blue	Bright yellow-green	–	Orange-brown	Ferulic acid
	Rf,		Trim–Hill reagent processing
Parameters	BAAW 4:1:2		Visible colors		Color in UV-light		Compound
Iridoids	0.23		Dark grey		Violet		Catalpol
	0.35		Dark grey		Violet		Aucubin
	0.56		Grey		Violet		n.d. (Iridoid)

n.d. – not defined.

Table 5. Chromatographic characterization of flavonoids, phenylcarboxylic acids, and iridoids found in V. teucrium leaves.

	Rf		Color
Parameters	BAAW 4:1:2	15% СН3СООН	UV-light	UV-light + NH3	AlCl3	Hepfner reagent	Compound
Flavonoids and  phenylcarboxylic acids	0.12	0.16	Umber	Yellow-green	Daffodil	–	n. d. (Flavonoid)
	0.12	0.25	–	Yellow-green	Daffodil	–	n.d. (Flavonoid)
		0.23	0.31	Umber	Dark brown	Yellow	–	n.d. (Flavonoid)
	0.40	0.43	Umber	Umber	Canary	–	Cynaroside
	0.80	0.25	–	Light green	Yellow	–	Luteolin
	0.18	0.45	Umber	Umber	Daffodil	–	n.d. (Flavonoid)
	0.21	0.54	–	Yellow-green	Yellow	–	n.d. (Flavonoid)
	0.31	0.64	–	Yellow-green	Yellow-green	–	n.d. (Flavonoid)
	0.48	0.60	–	Yellow-green	Yellow-green	–	n.d. (Flavonoid)
	0.24	0.59	Blue	Blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.38	0.61	Blue	Blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.80	0.51	Bright blue	Bright blue	–	Orange-brown	Caffeic acid
	0.61	0.74	Bright blue	Bright yellow-green	–	Foxy	Chlorogenic acid
	0.54	0.84	Dark blue	Bright blue	–	Orange-brown	Neochlorogenic acid
	0.42	0.81	Blue	Bright blue	–	Light orange-brown	n.d. (Phenylcarboxylic acid)
	0.27	0.81	Blue	Blue	–	Orange-brown	Quinic acid
	Rf,		Trim–Hill reagent processing
Parameters	BAAW 4:1:2		Visible colors		Color in UV-light		Compound
Iridoids	0.34		Dark grey		Violet		Aucubin
	0.16		Grey		Violet		n.d. (Iridoid)
	0.56		Grey green		Violet		n.d. (Iridoid)
	0.68		Dark grey		Violet		Catalposide

n.d. – not defined.

Table 6. Chromatographic characterization of flavonoids, phenylcarboxylic acids, and iridoids found in V. spicata leaves.

	Rf		Color
Parameters	BAAW 4:1:2	15% СН3СООН	UV-light	UV-light + NH3	AlCl3	Hepfner reagent	Compound
Flavonoids and phenylcarboxylic acids	0.16	0.17	Petunia	Yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.11	0.10	Petunia	Yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.13	0.18	–	Yellow-green	Yellow	–	n.d. (Flavonoid)
	0.12	0.25	Petunia	Yellow-green	Canary	–	n.d. (Flavonoid)
	0.25	0.20	Petunia	Yellow-green	Yellow	–	n.d. (Flavonoid)
	0.22	0.33	Petunia	Yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.42	0.29	Petunia	Yellow-green	Bright yellow-green	–	n.d. (Flavonoid)
	0.84	0.11	Violet	Yellow-green	Yellow-green	–	Luteolin
	0.92	0.06	–	Yellow-green	Yellow-green	–	Apigenin
	0.72	0.28	–	Yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.57	0.35	Violet	Yellow-green	Yellow-green	–	Apigenin 7-glucuronide
	0.44	0.47	Light yellow-green	Yellow-green	Daffodil	–	Cynaroside
	0.31	0.60	–	Chlorine	Daffodil	–	n.d. (Flavonoid)
	0.15	0.65	–	Chlorine	Daffodil	–	n.d. (Flavonoid)
	0.28	0.71	–	Chlorine	Daffodil	–	n.d. (Flavonoid)
	0.78	0.51	Bright blue	Bright blue	–	Orange-brown	Caffeic acid
	0.83	0.53	Bright blue	Bright blue	–	Light brown	Ferulic acid
	0.77	0.74	Blue	Blue	–	Cherry-brown	Isoferulic acid
	0.58	0.74	Bright blue	Bright yellow-green	–	Foxy	Chlorogenic acid
	0.52	0.85	Dark blue	Bright yellow-green	–	Orange-brown	Neochlorogenic acid
	0.28	0.78	Blue	Bright blue	–	Light orange-brown	Quinic acid
	0.18	0.86	Blue	Blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.14	0.88	Cyan	Cyan	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	Rf,		Trim–Hill reagent processing
Parameters	BAAW 4:1:2		Visible colors		Color in UV-light		Compound
Iridoids	0.16		Grey green		Violet		Isocatalpol
	0.35		Dark grey		Violet		Aucubin
	0.56		Grey green		Violet		n.d. (Iridoid)
	0.76		Dark grey		Violet		n.d. (Iridoid)
	0.69		Dark grey		Violet		Catalposide
	0.93		Grey		Violet		n.d. (Iridoid)

n.d. – not defined.

Table 7. Chromatographic characterization of flavonoids, phenylcarboxylic acids, and iridoids found in V. incana leaves.

	Rf		Color
Parameters	BAAW 4:1:2	15% СН3СООН	UV-light	UV-light + NH3	AlCl3	Hepfner reagent	Compound
Flavonoids and phenylcarboxylic acids	0.07	0.17	Petunia	Yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.11	0.09	Petunia	Yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.13	0.19	–	Yellow-green	Yellow	–	n.d. (Flavonoid)
	0.18	0.19	–	Yellow-green	Canary	–	n.d. (Flavonoid)
	0.35	0.20	Petunia	Yellow-green	Yellow	–	n.d. (Flavonoid)
	0.43	0.21	Petunia	Yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.47	0.42	Petunia	Light yellow-green	Yellow	–	Cynaroside
	0.56	0.31	Violet	Yellow-green	Yellow-green	–	Apigenin 7-glucuronide
	0.74	0.06	–	Yellow-green	Yellow-green	–	n.d. (Flavonoid)
	0.83	0.13	–	Yellow-green	Daffodil	–	Luteolin
	0.92	0.06	Violet	Bright yellow-green	Yellow-green	–	Apigenin
	0.73	0.31	Light yellow	Light yellow-green	Daffodil	–	n.d. (Flavonoid)
	0.31	0.42	–	Chlorine	Daffodil	–	n.d. (Flavonoid)
	0.14	0.44	–	Chlorine	Daffodil	–	n.d. (Flavonoid)
	0.11	0.69	–	Chlorine	Daffodil	–	n.d. (Flavonoid)
	0.35	0.68	–	Chlorine	Daffodil	–	n.d. (Flavonoid)
	0.07	0.17	Bright blue	Bright blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	0.86	0.56	Bright blue	Bright blue	–	Light brown	Caffeic acid
	0.81	0.79	Blue	Blue	–	Cherry-brown	n.d. (Phenylcarboxylic acid)
	0.62	0.73	Bright blue	Bright yellow-green	–	Foxy	Chlorogenic acid
	0.53	0.82	Dark blue	Bright yellow-green	–	Orange-brown	Neochlorogenic acid
	0.30	0.79	Blue	Bright blue	–	Light orange-brown	Quinic acid
	0.18	0.85	Blue	Blue	–	Orange-brown	n.d. (Phenylcarboxylic acid)
	Rf,		Trim–Hill reagent processing
Parameters	BAAW 4:1:2		Visible colors		Color in UV-light		Compound
Iridoids	0.23		Dark grey		Violet		Catalpol
	0.34		Dark grey		Violet		Aucubin
	0.56		Grey green		Violet		n.d. (Iridoid)
	0.68		Dark grey		Violet		Catalposide
	0.82		Grey green		Violet		n.d. (Iridoid)

n.d. – not defined.

Figure 1. Dose-dependence (A) and time-dependence (B) of iron chelating activity, total antioxidant, (C), and reducing (D) activities of plant extracts. □, P. maxima; Δ, P. lanceolata; ⋄,- P. major; ¤, V. teucrium; •, V. spicata; ○, V. incana.

Table 8. Influence of plant extracts on E. coli K12 growth (units of optical density at λ=492 nm).

Extract	t (h)	Control	P. maxima	P. lanceolata	P. major	V. teucurium	V. spicata	V. incana
1/20 plant extracts	0	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001
	2	0.074 ± 0.003	0.092 ± 0.006*	0.082 ± 0.007	0.102 ± 0.020*	0.097 ± 0.009*	0.094 ± 0.006*	0.087 ± 0.006
	4	0.138 ± 0.011	0.218 ± 0.017*	0.210 ± 0.025*	0.229 ± 0.040*	0.179 ± 0.019*	0.173 ± 0.005*	0.167 ± 0.003*
	6	0.195 ± 0.017	0.291 ± 0.009*	0.310 ± 0.010*	0.331 ± 0.025*	0.281 ± 0.041*	0.270 ± 0.021*	0.251 ± 0.010*
	24	0.451 ± 0.037	0.571 ± 0.018*	0.567 ± 0.051*	0.616 ± 0.061*	0.657 ± 0.097*	0.654 ± 0.113*	0.647 ± 0.138*
	1/40 plant extracts	0	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001	0.017 ± 0.001
2		0.074 ± 0.003	0.089 ± 0.005*	0.081 ± 0.001	0.093 ± 0.008*	0.086 ± 0.001*	0.089 ± 0.007*	0.087 ± 0.002*
4		0.138 ± 0.011	0.184 ± 0.010*	0.185 ± 0.012*	0.200 ± 0.022*	0.162 ± 0.005*	0.166 ± 0.012*	0.156 ± 0.006*
6		0.195 ± 0.017	0.237 ± 0.028*	0.255 ± 0.016*	0.275 ± 0.023*	0.250 ± 0.014*	0.257 ± 0.011*	0.248 ± 0.030*
24		0.451 ± 0.037	0.470 ± 0.036	0.467 ± 0.029	0.479 ± 0.041	0.539 ± 0.031*	0.521 ± 0.010*	0.553 ± 0.022*

*Significant difference in comparison to control.

Table 9. Influence of plant extracts on E. coli K12-specific growth rate.

Extract	t (h)	Control	P. maxima	P. lanceolata	P. major	V. teucurium	V. spicata	V. incana
1/20 water extracts	0–2	0.591 ± 0.060	0.717 ± 0.040*	0.652 ± 0.068	0.766 ± 0.124	0.742 ± 0.032*	0.727 ± 0.030*	0.683 ± 0.081
	2–4	0.345 ± 0.062	0.472 ± 0.029*	0.521 ± 0.073*	0.435 ± 0.046	0.346 ± 0.045	0.341 ± 0.027	0.358 ± 0.048
	4–6	0.195 ± 0.009	0.131 ± 0.019*	0.177 ± 0.015	0.165 ± 0.054	0.215 ± 0.020	0.224 ± 0.040	0.214 ± 0.017
	6–24	0.049 ± 0.002	0.038 ± 0.003*	0.032 ± 0.004*	0.034 ± 0.002*	0.046 ± 0.001	0.047 ± 0.004	0.049 ± 0.007
	1/40 water extracts	0–2	0.591 ± 0.060	0.693 ± 0.078	0.622 ± 0.063	0.717 ± 0.102	0.677 ± 0.067	0.681 ± 0.062	0.681 ± 0.052
2–4		0.345 ± 0.062	0.401 ± 0.039	0.456 ± 0.033*	0.412 ± 0.034	0.356 ± 0.027	0.350 ± 0.046	0.344 ± 0.035
4–6		0.195 ± 0.009	0.133 ± 0.065	0.168 ± 0.036	0.155 ± 0.032	0.225 ± 0.017	0.227 ± 0.039	0.240 ± 0.068
6–24		0.049 ± 0.002	0.039 ± 0.002*	0.031 ± 0.005*	0.031 ± 0.007*	0.044 ± 0.003	0.040 ± 0.003	0.046 ± 0.008

*Significant difference in comparison to control.

Table 10. Correlation between phytochemical content and specific bacterial growth rate.

	1/20 water extracts			1/40 water extracts
t1–t2	Flavonoids	Iridoids	Tannins	Flavonoids	Iridoids	Tannins
0–2	−0.136	−0.019	−0.447	0.232	0.216	−0.395
2–4	−0.077	0.315	0.871†	−0.348	0.064	0.737†
4–6	−0.269	−0.690†	−0.787†	−0.081	−0.542	−0.785†
6–24	0.325	−0.114	−0.689†	0.502	0.148	−0.480

†Significant correlation.

Table 11. Correlation between plant extract in vitro activity and specific bacterial growth rate.

	1/20 water extracts			1/40 water extracts
t1−t2	Iron-binding activity	Antioxidant activity	Reducing activity	Iron-binding activity	Antioxidant activity	Reducing activity
0–2	−0.610†	−0.333	−0.132	−0.818†	0.081	−0.224
2–4	0.654†	−0.361	−0.662†	0.803†	−0.640†	−0.699†
4–6	−0.236	0.290	0.740†	−0.454	0.535	0.844†
6–24	−0.503	0.589	0.721†	−0.534	0.576	0.620†

†Significant correlation.