Podophyllum hexandrum aqueous extract as a potential free radical scavenger

Figures & data

Figure 1. Effect of aqueous extract of P. hexandrum (P.H) and known antioxidant vitamin E (V.E) on DPPH radical scavenging activity. The results represent mean ± SD of three separate experiments.

Figure 2. Effect of aqueous extract of P. hexandrum (P.H) and catechin (an antioxidant) on hydroxyl radical scavenging activity. ***P < 0.001 when compared with catechin. The results represent mean ± SD of three separate experiments.

Figure 3. Reducing power of aqueous extract of P. hexandrum. The absorbance at 700 nm was measured in triplicate. BHT was used as a positive control. The results represent mean ± SD of three separate experiments.

Table 1. Effects of P. hexandrum aqueous extract on biochemical parameters in CCl₄-induced hepatotoxicity in albino rats

Treatment	Dose (mg/kg)	ALT (U/l)	AST (U/l)	LDH (U/l)
Control group (olive oil only)	5 ml/kg	20.06 ± 6.8	25.97 ± 5.05	35.51 ± 10.2
CCl4-treated group	1 ml/kg	120.60 ± 12.9	130.25 ± 9.3	235.70 ± 12.9
CCl4-treated + V.E	50 mg/kg	50.57 ± 5.1^$$$	62.48 ± 6.8^$$$	140.21 ± 5.7^$$$
CCl4-treated + PH extract	20 mg/kg	80.20 ± 4.2^$#	98.50 ± 6.1^$$###	215.85 ± 2.3^$###
CCl4-treated + PH extract	30 mg/kg	74.08 ± 8.6^$$###	75.02 ± 6.8^$#	190.06 ± 1.7^$$#
CCl4-treated + PH extract	50 mg/kg	68.27 ± 5.1^$$$#	69.70 ± 5.0^$$$ns	172.42 ± 5.3^$$$###

Each value represents the mean ± SD of seven animals. V.E; vitamin E, P.H; Podophyllum hexandrum.

^$$$P < 0.001, ^$$P < 0.01, ^$P < 0.05 compared to CCl₄ group, ^###P < 0.001 as compared with CCl₄ + V.E, ^#P < 0.05 as compared to CCl₄ + V.E, NS, non-significant when compared with CCl₄ + V.E. The data were presented as means ± SD of six parallel measures and evaluated by one way ANOVA followed by the Dunnett's test to detect inter group differences. Differences were considered to be statistically significant if P < 0.05.

Figure 4. Effect of aqueous extract of P. hexandrum on lipid peroxidation in CCl₄-treated rats. V.E stands for vitamin E. Values represent mean ± SD of seven animals in each group. ***P < 0.001 when compared with olive oil only, ^$$$P < 0.001 when compared with the CCl₄ group, NS, non-significant when compared with CCl₄ + V.E.

Table 2. Effects of treatment of aqueous extract of P. hexandrum on antioxidant enzymes in CCl₄-challenged rats

Parameters	Group I (olive oil only)	Group II CCl4 treated	Group III CCl4 + V.E	Group IV CCl4 + 20 mg/kg extract	Group V CCl4 + 30 mg/kg extract	Group VI CCl4 + 50 mg/kg extract
Glutathione reductase (μg GSSG utilized/minute/mg protein)	29.90 ± 1.3	3.27 ± 0.8	26.87 ± 3.5^$$$ns	9.52 ± 1.1^$$$***^###	14.98 ± 0.8^$$$***^###	18.22 ± 0.8^$$$***^##
Glutathione peroxidase (μg GSH utilized/minute/mg protein)	36.31 ± 3.2	3.30 ± 0.8	32.47 ± 2.8^$$$*	11.57 ± 0.7^NS***^###	17.54 ± 1.0^$$$**^###	26.31 ± 1.6^$$$ns##
Superoxide dismutase (units/mg protein)	54.34 ± 2.6	20.97 ± 1.2	41.425 ± 10.2^$$$*	31.06 ± 2.3^$***^#	40.63 ± 2.0^$$$*^NS	42.95 ± 5.5^$$$ns NS
Catalase activity (nm of H2O2 decomposed /minute/mg protein)	2138.57 ± 84.2	148.97 ± 7.0	1683.2 ± 41.5^$$$*	711.45 ± 21.0^$$$***^###	1127.76 ± 39.1^$$$***^###	1374.07 ± 52.9^$$$**^###
Glutathione-S-transferase (nmoles of CDNB conjugated/minute/mg protein)	11.55 ± 0.4	6.48 ± 0.5	9.89 ± 0.4^$$$*	6.51 ± 0.4***^#	8.66 ± 0.4^$$$**^##	9.78 ± 0.4^$$$ns NS

***P < 0.001, **P < 0.01, *P < 0.05, and ns, non-significant as compared with the normal group. ^$$$P < 0.001, ^$P < 0.05 as compared with the CCl₄-induced group. ^###P < 0.001, ^##P < 0.01, ^#P < 0.05, and NS, non-significant as compared to CCl₄ + V.E. The data were presented as means ± SD for seven animals in each observation and evaluated by one-way ANOVA followed by Dunnett's test to detect inter-group differences. Differences were considered to be statistically significant if P < 0.05.

Figure 5. Aqueous extract of P. hexandrum induced DNA fragmentation in HL-60 cells. Cells 2 × 10⁶/ml/well were treated with indicated conc. of plant extract for 24 hours. Genomic DNA was isolated and put to electrophoresis as described in the Materials and methods to assess the fragmentation at different concentrations. Lane 1, 1 kb ladder; Lane 2, –ve control (untreated cells); Lane 3, +ve control (camptothecin at 5 µM for 6 hours); Lanes 4–7, extract at 100, 70, 50, 30 µg/ml.

Figure 6. Inhibitory effects of plant extract on DNA nicking caused by hydroxyl radicals. Lane 1, native pBR322 plasmid DNA; Lane 2, Fenton's reagent + DNA; Lanes 3–6, Fenton's reagent + DNA + P. hexandrum aqueous extract (20–80 µg/ml); Lane 7, Fenton's reagent + DNA + 5 U catalase.