In vivo and in vitro antiviral activity of five Tibetan medicinal plant extracts against herpes simplex virus type 2 infection

Figures & data

Table 1.  EC₅₀ and CC₅₀ determinations for the medicinal plant extracts.

Plant extract	Family	Used part	CC50^a	EC50^b	S(CC50/EC50)I^c
D. heterophyllum	Lamiaceae	aerial parts	8.79 ± 0.5	0.99 ± 0.12	8.91
D. tanguticum	Lamiaceae	aerial parts	9.09 ± 1.49	1.43 ± 0.13	6.36
A. tanguticum	Ranunculaceae	aerial parts	5.25 ± 0.59	1.83 ± 0.21	2.81
S. mussotii	Gentianaceae	aerial parts	12.77 ± 0.61	3.60 ± 0.13	3.55
L. brevituba	Scrophulariaceae	aerial parts	1.40 ± 0.23	0.35 ± 0.11	4.04
ACV			10.85 ± 0.92	1.42 ± 1.17	7.64

^aDetermined as the concentration of the extracts that reduced the cell viability to 50% of control (cells without addition of extracts). Each represents the mean value from three independent experiments. The unit for CC₅₀ values shown in the table is mg/mL.

^bDetermined as the concentration of the extracts needed to inhibit 50% of virus-induced PFU to of control value. Each represents the mean value from three independent experiments. The unit for EC₅₀ values shown in the table is mg/mL.

^cSelectivity index (CC₅₀/EC₅₀).

Figure 1.  Virucidal activity of extracts against HSV-2. 100 PFU HSV-2 was studied on each test extract at various concentration (0.25, 0.5, 1, 2, 4 mg/mL except 0.06, 0.13, 0.25, 0.5, 1 mg/mL for L. brevituba, respectively) for 1 h at 37°C. Each point represents the mean ± SD of three independent experiments.

Figure 4.  Effect of ACV on HSV-2 infectivity, attachment and biosynthesis to Vero cells. The tested concentration of ACV was 0.03, 0.06, 0.13, 0.25, 0.5 mg/mL. Each point represents the mean ± SD of three independent experiments. A, HSV-2 was mixed with various concentrations of ACV for 1 h at 37°C; B, cells treated with ACV before virus infection; C, cells treated with ACV after virus infection.

Figure 2.  Effect of extracts on HSV-2 attachment to Vero cells. Vero cell monolayer in 24-well plates were treated with extracts before virus infection at various concentration (0.25, 0.5, 1, 2, 4 mg/mL except 0.06, 0.13, 0.25, 0.5, 1 mg/mL for L. brevituba, respectively). Each point represents the mean ± SD of three independent experiments.

Figure 3.  Effect of extracts on HSV-2 biosynthesis to Vero cells. Vero cell monolayer in 24-well plates were treated with extracts after virus infection at various concentration (0.25, 0.5, 1, 2, 4 mg/ml except 0.06, 0.13, 0.25, 0.5, 1 mg/mL for L. brevituba, respectively). Each point represents the mean ± SD of three independent experiments.

Table 2.  The effect of extracts against HSV DNA synthesis quantitated using a quantitative real-time PCR.

	Dose (mg/mL)	Copies /mL	Reduction folds
Control-1 (only cell)		NA
Control-2 (HSV-2 and cell)		(7.93 ± 0.84) × 10^8
D. heterophyllum	2	(3.62 ± 0.28) × 10^4*	10^4
	4	(2.39 ± 0.38) × 10^2*	10^6
D. tanguticum	2	(2.12 ± 0.52) × 10^3*	10^5
	4	(3.87 ± 1.2) × 10^3*	10^5
A. tanguticum	2	(2.05 ± 0.47) × 10^6*	10^2
	4	(1.36 ± 0.42) × 10^6*	10^2
S. mussotii	2	(3.14 ± 0.58) × 10^5*	10^3
	4	(5.26 ± 1.23) × 10^5*	10^3
L. brevituba	2	(3.14 ± 0.58) × 10^5*	10^3
	4	(5.26 ± 1.23) × 10^5*	10^3

Each number represents the mean ± SD of three independent experiments.

*Significant difference between test extract and control (p < 0.05).

Table 3.  The effect of extracts against HSV-2-induced encephalitis in mice.

Extracts	LD50 (g/kg)	Dose (g/kg)	Mean days^a	Mortality within 7 days (%)	Protection rate (%)
D. hetero phyllum	9.82	0.5	7.1 ± 0.8*	60	40
		1.00	5.9 ± 0.8	70	30
D. tang uticum	5.66	0.50	8.2 ± 0.5*	60	40
		1.00	5.5 ± 0.4	80	20
A. tang uticum	0.85	0.15	5.7 ± 0.3	100	0
		0.30	4.4 ± 0.3	100	0
S. mussotii	12.43	1.00	6.1 ± 1.0*	90	10
		2.00	7.1 ± 1.1*	100	0
L. brevituba	1.31	0.125	4.3 ± 0.2	100	0
		0.25	4.8 ± 0.3	100	0
ACV		0.01	8.5 ± 0.2*	50	50
Control-1			30.0 ± 0.0	0	100
Control-2			4.5 ± 1.1	100	0
Control-3		5% DMSO	4.3 ± 0.8	100	0

^aEach number represents the mean ± SD of three independent experiments.

*Significant difference between test extract and solvent control (p < 0.05).