A Perspective Study on Antioxidant activity of Nerium oleander (flower)

 

Mrs. S. Dhanalakshmi*, N. Harikrishnan

Department of Pharmacognosy, Faculty of Pharmacy,

Dr. M. G. R. Educational and Research Institute University, Velappanchavadi, Cennai-600117.

 

ABSTRACT:

Herb has been derived from the Latin word, “herba” and an old French word “herbe. Antioxidants play a vital role in the defense system against reactive oxygen species. As oxidative stress is a reason for many human diseases. Based on the literature review the Nerium plant has a potent antioxidant activity when it should be taken in minor concentration. The selected Nerium oleander flowers were extracted with different solvent system by using Aqueous, methanol, Acetone and Hydro alcohol. For the collection of volatile Oil Hydro distillation techniques  were carried out. The extracted phytoconstituents were analyzed by the preliminary phytoconstituent studies. To support this phytoconstituent analysis, thin layer chromatography was carried out. From this Phytoconstituent study we revealed that the Maximum number of phytoconstituent were present in Methanolic extract of Nerium oleander which also supported by Thin layer Chromatography.

 

 

 

INTRODUCTION:

Plants have been used for medicinal purposes long before prehistoric period. Ancient Unani manuscripts Egyptian papyrus and Chinese writings described the use of herbs. Herbal products are innocuous or possess minimal toxicity, some contain toxic ingredients that may not be identified on the label¹. These unidentified ingredients may be unintentionally included in the product (eg, misidentification of a toxic plant as a desired nontoxic plant or contamination with pesticide residues or heavy metals) or adulterants introduced for increased effect (eg, addition of a pharmaceutical agent to an herbal preparation)².

 

METHODOLOGY:

Preparation of Nerium oleander flower extract:

The flowers of Nerium oleander was washed properly with water and dried at room temperature. About 200g of fresh flower were extracted separately by maceration procedure successively with solvents such as Water, Methanol, Acetone and Hydro alcohol^{3, 4}. The solvents was filtered and refluxed. The yield was noted and its percentage was calculated.

 

QUALITATIVE:

Phytochemical evaluation:

The preliminary phytochemical screening of aqueous, Methanol, Acetone, Hydro alcohol extract of flowers were carried out for the identification of the various phytoconstituents. The results were tabulated in Table No.1.

 

THIN LAYER CHROMATOGRAPHY:

The principle of separation is adsorption and partition. When the stationary phase is solid in nature the principle of separation is adsorption and when it is liquid in nature the principle of separation is partition^{5, 6, 7}. The results were tabulated in Table No.2.

 

 

Table No.1

SOLVENT SYSTEM	RATIO
Hexane : Ethyl acetate	9: 1	8: 2	7: 3	6: 4	5: 5	4: 6	3: 7	2: 8	1: 9
Chloroform: Methanol	9: 1	8: 2	7: 3	6: 4	5: 5	4: 6	3: 7	2: 8	1: 9

 

Table No.2

S.NO	STATIONARY PHASE	MOBILE PHASE	RATIO	DETECTING AGENT
1	Silica gel-G	Hexane: Ethyl acetate	6: 4	Vanillin in H2SO4
2	Silica gel-G	Chloroform: Methanol	5: 5, 6: 4	Vanillin in H2SO4

 

Antioxidant study:

Antioxidant plays a vital role in the defense system against reactive oxygen species. As oxidative stress is a reason for many human diseases^{8, 9}. Research suggests that consumption of antioxidants rich food reduces damage of cell from free radicals. Dietary antioxidants interact in a dynamic fashion, including recycling, sparing one another and to decrease oxidative stress¹⁰. Natural antioxidants are Vitamin C, Vitamin E, Citrus fruits and juices, green peppers, cabbage^{11, 12}.

 

In-Vitro antioxidant activity of Nerium oleander Extracts:

Nitric oxide was generated from sodium nitroprusside and measured by Griess reaction. Sodium nitroprusside (5mM) in standard phosphate buffer saline solution (0.025 M, pH: 7.4) was incubated with different concentrations of EESC (25-1000 ug/ml), ascorbic acid as reference standard (5-1000 ug/ml) and FGTR (5-100ug/ml) dissolved in phosphate buffer saline (0.025M, PH: 7.4) and the tubes were incubated at 25*c for 5hrs. Control experiment without the test compounds but equivalent amounts of buffer were conducted in an identical manner^{13, 14}. After 5hrs, 0.5ml of incubation solution was removed and diluted with 0.5ml of Griess reagent (1% sulphanilamide, 2% O-phosphoric acid and o.1% naphthyl ethylene diamine dihydrochloride). The absorbance of the chromophore formed during diazotization of nitrite with sulphanilamide and its subsequent coupling with naphthyl ethylene diamine was read at 546 nm¹⁵. All the determinations were performed in 6 replicates¹⁶. (Annie Shirwaikar, 2006) percentage inhibition of nitric oxide was calculated by using the formula;

    

 

 

                                           Abs _control-Abs _Test × 100

Percentage Inhibition (%)= -----------------------------------

                                                   Abs _control

 

 

 

RESULT: The results were tabulated given below.

 

Table No. 3 : Qualitative phytochemical evaluation

S. No	CHEMICAL TEST	Nerium oleander
		Aqu.	Meth	Act.	Hydro alcohol
1.	Alkaloid a) Mayer’s test	-	-	-	-
	b) Dragendorff’s test	-	-	-	-
	c) Hager’s test	-	-	-	-
	d) Wagner’s test	-	-	-	-
2.	Glycosides Anthrone test	-	+	-	+
	Cardiac glycoside Keller killani test	-	-	-	-
	Anthroquinone test a) Borntrager’s test	-	-	-	-
	b) Legal test	-	-	-	-
	c) Baljet test	-	-	-	-
3.	Carbohydrate a) Molish’s test	-	+	-	+
	b) Barfoed’s test	-	+	-	+
	c) Fehling’s	-	+	-	+
	d) Benedict’s test	-	+	-	+
4.	Protein a)Millon’s test	+	-	-	-
	b)Biuret test	+	-	-	-
	c)Xanthoprotein test	-	-	-	-
5.	Amino acid Ninhydrin test	+-	-	-	-
6.	Steroids a)Libermann’s test b)salkowski test	- -	+ +	- -	+ +
7.	Flavonoids a)Shinoda test	-	+	-	-
8.	Tannins a)ferric chloride b)lead acetate c)gelation solution	- - -	+ + +	- - -	- - -
9.	Quinones	-	-	-	-
10.	Coumarin	-	-	-	-
11.	Anthocyanins a)sodium hydroxide b)con.sulphuric acid	- -	- -	- -	- -
12.	Triterpenoids Nollar’s test	-	-	-	-
13.	Saponin Foam test	-	-	+	-
14.	Fixed oils and fats a)spot test b)saponification test	- -	- -	- -	- -
15.	Gums and mucilage	-	-	-	-
	Resins a)with distilled water b)with acetone	- -	- -	- +	- -

+ indicates presence of phytoconstituent and – indicates their absence

 

Table No. 4: TLC analysis of Nerium olender (flower part )

S.No	Mobile Phase	Detecting agent	Ratio	Colour of spot	RfValue
1. 2.	Hexane: Ethyl acetate Chloroform: Methanol	Vanillin/Sulphuric acid Vanillin/Sulphuric acid	6: 4 5: 5 6: 4	Red orange Red orange Red orange	0.64 0.75 0.6

 

Thin Layer Chromatography of Nerium oleander flower

Fig No : 1

 

Table No. 5: Nitric Oxide Scavenging Activity of Methanolic Extract of Nerium oleander

S. NO	CONCENTRA-TION (µg/ml)	ABSORBANCE (nm)	% OF INHIBITION
			Nerium oleander (flower )	QUERCETIN
1.	Control	0.03	03.±09.01	03.±09.01
2.	25	0.16	20±2.67***	5±1.16
3.	50	0.18	15± 1.53	10±1.16
4.	100	0.17	10±0.81**	10±1.16
5.	200	0.19	05±1.44	15±1.16**
6.	500	0.19	05±1.11	20±1.16
7.	1000	0.17	05±1.16	20±1.16***

Values are mean ± SEM(n=6). All the values are ** P< 0.01, ***P< 0.05 when compared against control

 

 

 

SUMMARY AND CONCLUSION:

The selected Nerium oleander flower was authentified and extracted with different solvent system by using Aqueous, methanol, Acetone and Hydro alcohol. For the collection of volatile Oil Hydro distillation techninc were carried out¹⁷. The extracted phytoconstituent were analysed by the preliminary phytoconstituent studies¹⁸. To support this phytoconstituent Thin layer chromatography were carried out. From this Phytoconstituent study we revealed that the Maximum number of phytoconstituent were present in Methanolic extract of Nerium olender which also supported by Thin layer Chromatography¹⁹. Methanol was proved to be the most efficient solvents for the extraction of antioxidants from Nerium oleander flower. Along with that report we were also finding the Maximum effect should only present at 50 µg/ml, (***P< 0.05) were the concentration of extract increased the antioxidant activity was decreased due to the presence of toxic principle present in the flower part^{20, 21}

 

ACKNOWLEDGEMENT:

The authors are highly thankful to the Department of Pharmacognosy, Faculty of Pharmacy, Dr. M. G. R. Educational and Research Institute University providing the facilities during research works.

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Accepted on 26.07.2017        © RJPT All right reserved

Research J. Pharm. and Tech 2017; 10(10):3449-3452.