Research Articles

2018  |  Vol: 4(2)  |  Issue: 4(March-April)  |  https://doi.org/10.31024/ajpp.2018.4.2.10
In-vitro antioxidant activity and free radical scavenging potential of three different extract of leaves of Chinese Ornamental plant

Sailesh Narayan*

Department of Pharmacology, Radharaman College of Pharmacy, Bhopal, Madhya Pradesh, India

*Address for Corresponding Author

Prof. Sailesh Narayan

Department of Pharmacology, Radharaman College of Pharmacy, Bhopal, Madhya Pradesh, India


Abstract

Objective: The objective of the present work is to study for an in-vitro anti-oxidant activity of petroleum ether, ethyl acetate and methanolic extracts of leaves of Salvia splendensMaterial and methods: The extracts were studied using 1,1-diphenyl-2-picryl hydrazyl, hydrogen peroxide (H2O2), total phenolic content (TPC) and total flavonoid content (TFC). The TPC and TFC were estimated taking gallic acid and rutin calibration curve respectably. Results: All the extracts possess in-vitro anti-oxidant activity. However, the order of possessing activity was methanolic > ethyl acetate > petroleum ether extracts of S. splendens leaves. The TPC and TFC were highest in methanolic extract. Conclusion: It can be concluded that the extract of the leaves of S. splendens, possess anti-oxidant activity. The methanolic extract of the leaves of S. splendens, possess highest anti-oxidant activity in-vitro.

KeywordsSalvia splendens, anti-oxidant, 1,1-diphenyl-2-picryl hydrazyl, phenolic content, flavonoid content


Introduction

Oxidative damage to cellular biomolecules such as lipids, proteins and DNA is thought to play a crucial role in the incidence of several chronic diseases (Simonian and Coyle, 1996; Dhalla et al., 2000; Van’t Veer  et al., 2000 ; Bokov, 2004 ; Madamanchi  et al., 2005). Flavonoids are a group of polyphenolic compounds found abundantly in the plant kingdom. Interest in the ;possible health benefits of flavonoids and other polyphenolic compounds has increased in recent years owing to their potent antioxidant and free-radical scavenging activities (Bravo et al., 1998 ; Heim et al., 2002 ; Pier-Giorgio, 2000; Rice-Evans et al., 1997; Sealbert et al., 2005; Ross and Coyle, 2002; Rice-Evans et al., 1995).

The effects of free radicals on human beings are closely related to toxicity, disease and aging Most living species have an efficient defense system to protect themselves against the oxidative stress induced by reactive oxygen species (Dhalla et al., 2000). Recent investigations have shown that the antioxidant properties of plants could be correlated with oxidative stress defense and different human diseases including cancer, atherosclerosis and the aging process (Van’t Veer  et al., 2000 ; Bokov et al., 2004 ; Madamanchi  et al., 2005). The antioxidants can interfere with the oxidation process by reacting with free radicals, chelating free catalytic metals and also by acting as oxygen scavengers.

Salvia splendens of family Lamiaceae/Labiatae (Mint family) is commonly known as Scarlet sage (Shanker and Unnikrishnan, 2001; Gharpure, 2005; Bobbi, 2003). It also reported the activity like analgesic and anti-inflammatory of roots, anti-ulcerative activity, antimicrobial activity, laxative activity, anti-oxidant, hepatoprotective and anti-hyperlipidemic activity have also been reported (Shanker and Unnikrishnan, 2001; Gharpure, 2005; Bobbi, 2003; Nadkarni,1991; The Wealth of India, 1998; Kaplan, 2001; Erv, 2014) . Hence, the present investigation was conducted to study in-vitro antioxidant activities of various roots extracts so as to make researcher to route for other pharmacological activities.

Material and methods

Plant material, authentication and extraction procedures

S. splendens plant was collected from Bhopal (Madhya Pradesh) and Hazaribag, (Jharkhand) and was authenticated by Dr. V.P. Prasad, Scientist-C, Botanical Survey of India, Government of India, Howrah, (West Bengal). The specimen No. PY/JVD 1026/2011 had been submitted. The air-dried roots were made into coarse powder and extracted with methanol, ethyl acetate and petroleum ether and percentage yield were calculated. The dried roots were extracted with Hot continuous soxhlet apparatus for 72 hrs with three different solvents i.e. methanol, ethyl acetate and petroleum ether and concentrated to dryness under reduced temperature.

Preliminary phytochemical analysis

The various extracts of S. splendens were tested for different phytoconsituents such as alkaloids, glycosides, saponinins, tannins, terpinoids, phenolic compounds, protein, carbohydrates using standard procedures (Kokate, 1993).

In-vitro anti-oxidant activity

1,1-diphenyl-2-picryhydrazyl (DPPH) radical scavenging activity

The DPPH assay of S. splendens extract was determined by the method as reported by Patil et al. (2009). The procedure involved UV-spectrophotometric determination. Three solutions i.e. standard, test and control were prepared.

Preparation of standard ascorbic acid solutions

Different solutions (1-10 μg/ml) of the ascorbic acid were prepared in methanol. 1.5 ml of each solution of ascorbic acid were mixed with 1.5 ml of 200 μM DPPH solution and incubated for 30 minutes at room temperature in dark. Absorbance of each solution was taken after 30 minutes against methanol (as blank) at 517 nm (Premanath and Lakshmidevi, 2010; Pandey and Barve, 2011).

Preparation of test solutions

Different solutions of the S. splendens extract were prepared in methanol to give concentrations (10-100 μg/ml). 1.5 ml of each solution of S. splendens extract was mixed with 200 μM DPPH solution and incubated for 30 minutes at room temperature in dark. Absorbance of each solution of S. splendens extract was taken after 30 minutes against methanol (as blank) at 517 nm (Premanath and Lakshmidevi, 2010; Pandey and Barve, 2011).

Preparation of control solution

For control, 1.5 ml of methanol was mixed with 200 μM DPPH solution and incubated for 30 minutes at room temperature in dark. Absorbance of the control was taken after 30 minutes against methanol (as blank) at 517 nm.

Percentage inhibition was calculated using equation (1), while IC50 values were estimated from the % inhibition versus concentration plot, using a non-linear regression algorithm. The data were presented as mean values ± standard deviation (n=3) (Premanath and Lakshmidevi, 2010; Pandey and Barve, 2011).     

Where Equation (1)

I% = Percentage inhibition

Ac = Absorbance of control (methanol and 200 μM DPPH solution)

At = Absorbance of ascorbic acid/plant extract with 200 μM DPPH solution after 30 minutes.

Ab = Absorbance of ascorbic acid/plant extract without 200 μM DPPH solution.

Ferric reducing power activity

This method is based on the principle of increase in the absorbance of the reaction mixture. Increase in the absorbance indicates increase in the antioxidant activity. Substances, which have reduction potential, react with potassium ferricyanide (Fe3+) to form potassium Fe2+, which then react with ferric chloride to form ferric ferrous complex that has an absorption maximum at 700 nm. Increase in absorbance of the reaction mixture indicates the reducing power of the Samples.

Antioxidant= Potassium ferricyanide + Ferric chloride→Potassium ferrocyanide + Ferrous chloride

Preparation of standard ascorbic acid solutions

Different concentrations of the ascorbic acid were prepared in distilled water to give solutions of concentration (20-100 μg/ml). 1ml of each concentration of ascorbic acid solutions were mixed with 2.5 ml of phosphate buffer (0.2 M, pH 6.6) and 2.5 ml of 1% potassium ferricyanide solution. The reaction mixtures were incubated for 20 minutes at 50°C. Afterwards 2.5 ml of 10% trichloroacetic acid solution was added and centrifuged at 560 × g for 10 minutes. After separation, 2.5 ml of upper layer of each solution were mixed with 2.5 ml of distilled water and 1 ml of 0.1% ferric chloride (freshly prepared solution). Absorbance was recorded for each solution of ascorbic acid against (0.2 M, pH 6.6) phosphate buffer (used as blank) at 700 nm (Jayanthi and Lalitha, 2011).

Preparation of test solutions

Different solutions of extract were prepared in distilled water to give various concentrations (20-100 μg/ml). 1 ml of each solution of plant part extract was mixed with 2.5 ml of phosphate buffer (0.2 M, pH 6.6) and 2.5 ml of 1% potassium ferricyanide solution. The reaction mixtures were incubated for 20 min at 50°C. Afterwards 2.5 ml of 10% trichloroacetic acid solution was added and centrifuged at 560 × g for 10 minutes. After separation, 2.5 ml of upper layer of each solution were mixed with 2.5 ml of distilled water and 1 ml of 0.1% ferric chloride (freshly prepared solution). Absorbance was recorded for each solution of plant part extract against (0.2 M, pH 6.6) phosphate buffer (used as blank) at 700 nm.

The absorbance versus concentration curve for ascorbic acid and extract was plotted. The‘Y’ and ‘R2’ values obtained in both curve and the cases were comparatively studied to determine the reducing power of extract (Jayanthi and Lalitha, 2011).

Hydrogen peroxide (H2O2) scavenging activity

H2O2 is a biologically important oxidant because of its ability to generate the hydroxyl radical (•OH) which is extremely potent. The ability of the •OH to remove or add hydrogen molecules to unsaturated hydrogen bonds of organic lipids makes it potentially one of the most reactive oxidants in biological systems. It’s very short half-life (1 × 10−9 at 37°C), however, restricts its diffusion capability and potency.

The ability of the S. splendens extract to scavenge H2O2 was determined according to the method reported by Neha Panday et al. (2011).

The procedure involved UV-spectrophotometric determination of H2O2 radical scavenging. Three solutions i.e. standard, test and control were prepared.

Preparation of standard ascorbic acid solutions

Different concentrations (10-100 μg/ml) of the ascorbic acid were prepared in distilled water. 1ml of each solution of ascorbic acid was mixed with 2 ml of 0.1 M phosphate buffer solution and 600 μl of 100 mM H2O2 solution. After 10 minutes absorbance of different concentration of ascorbic acid solutions was taken at 230nm (Deore et al., 2009).

Preparation of test solutions

Various concentrations (10-100 μg/ml) of the S. splendens aq. extract were prepared in distilled water. 1 ml of each solution of aq. extract was mixed with 2 ml of 0.1 M phosphate buffer solution and 600 μl of 100 mM H2O2 solution. After 10 minutes (approximately) absorbance of different concentration of S. splendens extract solutions were taken at 230 nm (Deore et al., 2009).

Preparation of control solution

For control 2 ml of 0.1 M phosphate buffer solution was mixed with 600 μl of 100 mM H2O2 solution. After 10 minutes absorbance of control was taken at 230 nm. The percentage of inhibition was calculated by comparing the absorbance values of the control and test samples using Equation 1. IC50 values were estimated from the % inhibition versus concentration plot, using a non-linear regression algorithm (Deore et al., 2009).

Where,

I% = Percentage inhibition.

Ac = Absorbance of control (0.1 M phosphate buffer solution and H2O2).

At = Absorbance of ascorbic acid/plant extract with H2O2 after 10 minutes.

Ab = Absorbance of ascorbic acid/plant extract without H2O2.

Estimation of total phenolic content (TPR)

The amount of total TPC in extracts was determined with the Folin– Ciocalteu reagent. Gallic acid (GA) was used as a standard and the total phenolic were expressed as mg/g GA equivalent (GAE). Concentration of 0.01, 0.02, 0.03, 0.04 and 0.05 mg/ml of GA were prepared in methanol. Concentration of 0.1 and 1 mg/ml of plant extract were also prepared in methanol and 0.5 ml of each sample were introduced in to test and mixed with 2.5 ml of a 10 fold dilute Folin–Ciocalteu reagent and 2 ml of 7.5% sodium carbonate. The tubes were covered with parafilm and allowed to stand for 30 minutes at room temperature before the absorbance was at read at 760 nm spectrometrically. All determination was performed in triplicate. The Folin–Ciocalteu reagent is sensitive to reducing compounds including polyphenols. They produce a blue color upon reaction. This blue color was measured spectrophotometrically.

Line of regression from GA was used for estimation of unknown phenol content. From standard curve of GA line of regression was found to be

y = 0.005x + 0.065 and R2 = 0.976

Thus the goodness of fit was found to be good for selected standard curve. By putting the absorbance of test sample (y = absorbance) in line of regression of above mentioned GA (Maurya and Singh, 2010).

Total flavonoids determination

Total flavonoids were measured by a colorimetric assay according to Dewanto et al. An aliquot of diluted sample or standard solution of quercetin  was added to a 75 μl of NaNO2 solution, and mixed for 6 minutes, before adding 0.15 mL AlCl3 (100 g/L). After 5 minutes, 0.5 mL of NaOH was added. The final volume was adjusted to 2.5 ml with distilled water and thoroughly mixed. Absorbance of the mixture was determined at 510 nm against the same mixture, without the sample, as a blank. Total flavonoid content was expressed as mg quercetin/g dry weight (mg quercetin/g DW), through the calibration curve of quercetin. All samples were analyzed in three replications.

Line of regression from quercetin was used for estimation of unknown flavonoid content. From standard curve of quercetin line of regression was found to be

y = 0.001x − 0.118 and R2 = 0.985

Thus the goodness of fit was found to be good for selected standard curve. By putting the absorbance of test sample (y = absorbance) in line of regression of above mentioned quercetin (Zou et al., 2004; Zengin et al., 2011; Rahman et al., 2013).

Results

The successive solvent extraction was done using petroleum ether, ethyl acetate and methanol using standard procedure. The behavior of various extracts like texture and colour and extractive yield were calculated.

DPPH free radical scavenging activity

The DPPH radical scavenging activity of petroleum ether extract of S. splendens (PEESS), ethyl acetate extract of S. splendens (EAESS) and methanol extract of S. splendens (MESS) leaves were detected and compared with ascorbic acid. The percentage inhibition (% inhibition) at various concentration (10-100 μg/ml) of PEESS, EAESS and MESS as well as standard ascorbic acid (1-10 μg/ml) were calculated and plotted (Figure 1,2,3,4). The IC50 values of ascorbic acid were 25.07 μg/ml), PEESS (187.74 μg/ml), EAESS (112.22 μg/ml) and MESS 95.30 μg/ml).

Table 1. % Inhibition of DPPH by Ascorbic acid

Concentration (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

0.291

0.562

 

48.22

25.07

40

0.253

54.98

60

0.217

61.39

80

0.172

69.40

100

0.144

74.38

Figure 1. Regression curve of ascorbic acid by DPPH assay method

 

Table 2.  % Inhibition of DPPH by pet.ether leaves extract of Salvia splendens (S1)

Conc. (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

0.407

0.562

27.58

187.74

40

0.396

29.54

60

0.384

31.67

80

0.364

35.23

100

0.343

38.97

Figure 2. Regression curve of pet ether leaves extract of Salvia splendens by DPPH assay method (S1)

 

Table 3. % Inhibition of DPPH by ethyl acetate leaves extract of Salvia splendens (S1)

Conc. (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

0.381

0.562

 

32.21

112.22

40

0.368

34.52

60

0.343

38.97

80

0.314

44.13

100

0.294

47.69

Figure 3. Regression curve of ethyl acetate leaves extract of Salvia splendens by DPPH assay method (S1)

 

Table 4. % Inhibition of DPPH by methanolic leaves extract of Salvia splendens (S1)

Conc. (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

0.347

0.562

38.26

95.30

40

0.323

42.53

60

0.308

45.20

80

0.299

46.80

100

0.275

51.07

Figure 4. Regression curve of methanolic leaves extract of Salvia splendens by DPPH assay method (S1)
 

Ferric reducing power activity

The reductive capabilities of PEESS, EAESS and MESS leaves were detected and compared with ascorbic acid. The mean absorbance at various concentration (20-100 μg/ml) of PEESS, EAESS and MESS as well as standard Ascorbic acid (20-100 μg/ml) were calculated. The reductive capabilities were found to increase with increasing of concentration in various extract as well as standard ascorbic acid.

Table 5. Reducing power assay of leaves extract of Salvia splendens (S1)

S. No.

Conc. (µg/ml)

Pet. ether leaves extract

Ethyl acetate leaves extract

Methanolic leaves extract

1

10

0.029

0.035

0.049

2

20

0.036

0.046

0.063

3

40

0.057

0.069

0.088

4

60

0.078

0.094

0.112

5

80

0.105

0.119

0.137

6

100

0.113

0.126

0.144

H2O2 scavenging activity

The H2O2 scavenging activity of PEESS, EAESS and MESS leaves were detected and compared with ascorbic acid. The percentage inhibition (% inhibition) at various concentration (10-100 μg/ml) of PEESS, EAESS and MESS as well as standard Ascorbic acid (10-100 μg/ml) were calculated and plotted in figure 5. The IC50 values are calculated from graph and were found ascorbic acid (36.27 μg/ml), PEESS (170.85 μg/ml), EAESS (122.06 μg/ml) and MESS (68.54 μg/ml).

Table 6. % Inhibition of H2O2 by Ascorbic acid

Conc. (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

0.708

1.302

 

45.62

36.27

40

0.619

52.46

60

0.592

54.53

80

0.546

58.06

100

0.513

60.60

Figure 5. Regression curve of ascorbic acid by H2O2 assay method

 

Table 7. % Inhibition of H2O2 by pet ether leaves extract of Salvia splendens (S1)

Conc. (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

1.069

1.302

 

17.90

170.85

40

1.032

20.74

60

0.982

24.58

80

0.922

29.19

100

0.837

35.71

Figure 6. Regression curve of pet. ether leaves extract of Salvia splendens by H2O2 assay method (S1)

 

Table 8. % Inhibition of H2O2 by ethyl acetate leaves extract of Salvia splendens (S1)

Conc. (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

0.905

1.302

 

30.49

122.06

40

0.817

37.25

60

0.793

39.09

80

0.744

42.86

100

0.714

45.16

Figure 7. Regression curve of ethyl acetate extract of Salvia splendens by H2O2 assay method (S1)

 

 

Table 9. % Inhibition of H2O2 by methanolic leaves extract of Salvia splendens (S1)

Conc. (µg/ml)

Absorbance of Sample

Absorbance of Control

% Inhibition

IC50 (µg/ml)

20

0.834

1.302

 

35.94

68.54

40

0.774

40.55

60

0.673

48.31

80

0.598

54.07

100

0.537

58.76

Figure 8. Regression curve of methanolic extract of leaves of Salvia splendens by H2O2 assay method (S1)

 

Total phenolic contents (TPC)

The TPC in PEESS, EAESS and MESS roots were estimated using standard GAE of phenols. The various concentration of GA (10-50 μg/ml) calibration curve was plotted (figure 9). The TPC for PEESS, EAESS and MESS were obtained for 1 mg/ml of extracts from TPC calibration of GA. The phenolic compounds are absent in the petroleum ether. The TPC for EAESS and MESS were calculated using standard calibration curve (y = 0.007x + 0.056, R2 = 0.995) and found to have 202.06±0.611and 213.0±0.721 mg/g equivalent of GA respectably.

Figure 9. Standard curve of Gallic acid

 

Total flavonoid content (TFC)

The TFC in PEESS, EAESS and MESS roots were estimated using standard quercetin equivalent of phenols. The various concentration of quercetin (25-100 μg/ml) calibration curve was plotted and the results were given in figure 10. The TFC for PEESS, EAESS and MESS were obtained for 1000 μg/ml of extracts from TFC calibration of quercetin. The phenolic compounds are absent in the petroleum ether. The TFC for EAESS and MESS were calculated using standard calibration curve (y = 0.001x+ 0.120, R2 = 0.998) and found to have 92.33±3.055, and 115.33±1.154 mg/g equivalent of quercetin respectably.

Figure 10. Standard curve of quercetin

 

Discussion

The S. splendens roots were made coarse powder and extracted with using petroleum ether, ethyl acetate and methanol as solvent using standard procedure.

The various extracts of S. splendens roots were tested for different phytoconsituents like alkaloids, glycosides, saponinins, tannins, terpinoids, reducing sugars, phenolic compounds, flavonoids, protein, carbohydrates and volatile oils. The phenolic and flavonoids are widely distributed secondary metabolites in plants having anti-oxidant activity and have wide range of biological activities as anti-apoptosis, anti-aging, anti-carcinogen, anti-inflammation, anti-atherosclerosis, cardiovascular protection and improvement of endothelial function, as well as inhibition of angiogenesis and cell proliferation activities (Rice-Evans et al., 1996; Bors et al., 1987). Recent studies have shown that many dietary polyphenolic constituents derived from plants are more effective antioxidants in-vitro than vitamins E or C, and thus might contribute significantly to the protective effects in vivo (Rice-Evans et al., 1997)

In-vitro antioxidant studies are widely carried to screen various plant containing phenolic and flavonoids constituents. Plant derived antioxidant compounds, flavonoids and phenolics have received considerable attention because of their physiological effect like antioxidant, anti-inflammatory, antitumor activities and low toxicity compared with those of synthetic phenolicantioxidant such as butylated hydroxyanisole, butylated hydroxytoluene and Propyl gallate (Kumar et al., 2008; Sharififar et al., 2009).

DPPH is a purple colored stable free radical; when reduced it becomes the yellow-colored diphenyl-picryl hydrazine. DPPH radicals react with suitable reducing agents and then electrons become paired-off and the solution loses colour stoichimetrically with the number of electrons taken up (Chidambaraet al., 2003).. Such reactivity has been widely used to test the ability of compounds/plant extracts to act as free radical scavengers (Park et al., 2012). In this present study, the DPPH radical scavenging activity of MESS, PEESS and EAESS roots were detected and compared with ascorbic acid. The IC50 values for DPPH assay of for methanolic extract was found maximum, followed by ethyl acetate extract and for petroleum ether extract was minimum. Though the extracts showed good DPPH scavenging activity, but it was less effective than standard ascorbic acid. The difference of activity is due to presence of phenolic components in different extracts. Thus, choosing the appropriate solvent is one of the most important factors for obtaining extracts with a high content of bioactive compounds and antioxidant activity (Park et al., 2012).

In ferric reducing antioxidant power assay, a yellow color of the test solution changes to various shades of green and blue is depending on the reducing power of each compound. The presence of radicals (i.e. antioxidant) causes the conversion of the Fe3+/ferricyanide complex used in this method to the ferrous form. Therefore by measuring the formation of pearls prussian blue spectroscopically, the Fe2+ concentration can be monitored; a higher absorbance indicates a higher reducing power. The reductive capabilities of PEESS, EAESS and MESS roots were detected and compared with ascorbic acid. The methanolic extract showed highest reducing power, followed by ethyl acetate extracts and petroleum ether extracts respectively. The increased reducing power in the extracts indicated that some components in the extract were electron donors that could react with the free radicals to convert them into more stable products to terminate radical chain reaction. Antioxidants are strong reducing agents and this is principally based on the redox properties of their hydroxyl groups and the structural relationships between different parts of their chemical structure (Oboh et al., 2012; Mustafa et al., 2010).

H2O2, a biologically relevant, non-radical oxidizing species, may be formed in tissues through oxidative processes. H2O2 which in turn generate OH resulting in initiation and propagation of lipid peroxidation (Mustafa et al., 2010). The H2O2 scavenging activity of PEESS, EAESS and MESS roots were detected and compared with ascorbic acid. The IC50 values for H2O2 scavenging activity of for methanolic extract was found maximum followed by ethyl acetate extract and for petroleum ether extract was minimum. Though the extracts showed good H2O2 scavenging activity but it was less effective than standard ascorbic acid. The ability of the extracts to quench OH− seems to be directly related to the prevention of the lipid peroxidation and appears to be moderate scavenger of active oxygen species, thus reducing rate of chain reaction (Maity et al., 2013).

The TPC in PEESS, EAESS and MESS roots were estimated using standard GAE of phenols. The phenolic compounds are absent in the petroleum ether. The TPC for EAESS and MESS were found to have 202.06 and 213.00 mg/g equivalent of GA respectably. The methanolic extract was found to have maximum phenolic components and which may be one the reason of its to possess maximum antioxidant activity then other two extracts (Habbu et al., 2010). But in TFC, it was found methanolic extract to possess maximum 148.66 mg/g equivalent of rutin then other ethyl acetate (121.66 mg/g Eq). Flavonoids play some important pharmacological roles against diseases, such as cardiovascular disease, cancer, inflammation and allergy and other oxidative stress related diseases (Habbu et al., 2010). From, above discussion, it was clear that the most powerful anti-oxidant extract is MESS roots.

Conclusion

It can be concluded that S. splendens roots extracts possess anti-oxidant activities and the potency of anti-oxidant activities depends on the type of extract. The MESS roots possess highest anti-oxidant activity in-vitro. This anti-oxidant power depends on total phenolic and flavonoid contents on particular extract.

Conflicts of interest

Authors not have any conflicts

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