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ABSTRACT
The marine macro algal species have great antioxidant potentials. They produce various numbers of bioactive compounds.
The aim of present investigation was to evaluate the phytochemical screening and antioxidant activity of green alga
Cladophora glomerata Linn belonging to family- Glomerataceae. This alga collected along the coasts of Raigad of Konkan
region of Maharashtra. Qualitative phytochemical investigations indicate that the extract of Cladophora glomerata
contained phytoconstituents like flavonoids, glycosides, phenolic compounds, saponins, tannin and proteins. The
antioxidant potential investigated by using four methods like DPPH radical scavenging(%), FRAP Assay, Reducing power
assay and determination of total phenolic contents. Obtained results from these methods clearly indicates that the
Cladophora glomerata have a great antioxidant potential.
KEY WORDS: Raigad coast, Cladophora glomerata Linn, Phyto -constituents, Antioxidant potential.
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Study of phytochemical screening & antioxidant activities of Cladophora glomerata
Preparation of organic extracts of sample for 593 nm with using single concentration of extract i.e. 10
Antioxidant activities mg/ml and standard curve being linear between 100 and
The dried sample of seaweeds ground to coarse powder, 1000 L FeSO4. Then results were expressed in L Fe
weighed and wrapped in Whatman No.1 filter paper and (II)/g dry mass and compared with that of BHT and BHA
successively extracted with 200 ml of different solvents (Kasote et al., 2011).
such as benzene, chloroform, ethanol, ethyl acetate, Reducing power assay
methanol and petroleum ether with their increasing order The reducing power of marine algal extracts was assessed
of polarity by soxhlation for 12-24 hours. The extract by using various concentrations i.e. 50-250 L/ml of
analyzed for the presence of antioxidant activities by marine algal extracts in methanol (10 mg/ml), by using
referring standard procedure (Thoudam et al., 2011). reference of standard BHT and BHA (1mg/ml), mixed
DPPH radical scavenging activity with 2.5 ml phosphate buffer (0.2 M, pH 6.6) and 1%
DPPH is a stable inorganic radical. To determine the potassium ferric cyanate (2.5 ml). The mixture was
radical scavenging effect of marine algal extracts, DPPH incubated at 500C for 20 minutes and (10%, 2.5 ml)
(1, 1-diphenyl-2-picryl hydrazyl) method is used. This trichloroacetic acid was added. This solution is centrifuged
method is based on estimating the reduction of alcoholic at 3000 rpm for 10 minutes. Then upper layer of the
DPPH solution in the presence of a hydrogen donor. reaction mixture (2.5 ml) is mixed with 2.5 ml distilled
A solution of 0.135 mM DPPH in methanol was prepared water and 0.5 ml FeCl3 (0.1%), and absorbance measured
and 1.0 ml of this solution was mixed with 1.0 ml of at 700 nm. Increased absorbance of the reaction mixture
marine algal extracts in methanol with different indicates the increased reducing power (Kasote et al.,
concentrations (0.5-2.5 mg/ml). Then the reaction mixture 2011).
mixed thoroughly and kept in the dark at room Determination of total phenolic content
temperature for 30 minutes and absorbance of the reaction Total phenolic content in marine algal extracts were
mixture measured spectrophotometrically at 690 nm. BHT determined by using modified Folin-Ciocalteau reagent
and BHA were used as references (Liyana-Pathiranan et method. According to (Zahin et al., 2009), gallic acid is a
al., 2005). standard phenolic compound. The reaction mixture
Total antioxidant activity (FRAP Method) contained single concentration of the extracts i.e. 10
Total antioxidant activity of marine algal extracts was mg/ml, and Folin- Ciocalteau reagent. To 500 L (10
determined by FRAP (Ferric reducing antioxidant mg/ml) of marine algal extracts in methanol, 2.5 ml of
potential) method. The stock solution included 300 ml 1:10 dilution of Folin-Ciocalteaus reagent and 2 ml of
acetate buffer (3.1g C2H3NaO2.3H2O and 16 ml C2H4O2), Na2CO3 (7.5% w/v) were added and mixed thoroughly and
pH 3.6, 10 ml TPTZ (2, 4, 6-tripyridyl-S-triazine) solution incubated at 45oC for 15 minutes. Same procedure is
in 40 ml HCL, and 20 ml FeCl3.6H2O solution. The fresh followed for other marine algal extracts with benzene,
working solution prepared by mixing 30 ml acetate buffer, chloroform, ethanol, ethyl acetate and petroleum ether
3 ml TPTZ and 3 ml FeCl3.6H2O solution. The respectively. The absorbance measured at 765 nm. The
temperature of the solution was maintained to 370C before concentration of total phenolic content in the marine algal
analysis. Then marine algal extracts (150L) allowed to extracts was determined as milligrams of Gallic acid
react with 2850 L of the FRAP solution for 30 minutes in equivalent per gram of dry weight (mg GAE/g dw) (Zahin
the dark condition. The coloured product (Ferrous et al., 2009).
tripyridyl triazine complex) read spectrophotometrically at
RESULTS
TABLE 1: Preliminary phytochemical study of Cladophora glomerata Linn.
Sr. Name of the Algal Solvent used a b c d e f g h i j k
No Species
water - + + + + - + + + - +
HCL - + - - + - - + + - +
Ethanol - + + + + - + + + - +
1 Cladophora Ethyl Acetate - + + + + - + + + - +
glomerata Linn. Methanol - + + + + - - + + - +
Chloroform - - - + + - + + - - -
Benzene - + + - + - - + - - -
Petroleum ether - - - - - - - - - - -
Where, a: Alkaloids, b: Flavonoids, c: Glycosides, d: Phenolic compounds, e: Saponins, f: Steroids, g: Tannins, h:
Carbohydrates, i: Proteins, j: Fats, k: Sugar and (+): Present, (-): Absent.
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I.J.S.N., VOL.7 (3) 2016: 659-663 ISSN 2229 6441
TABLE 2: DPPH-Radical scavenging activity of Cladophora glomerata Linn, at 517 nm is given in table,
Sr. Concn DPPH-Radical Scavenging Activity (%) of Cladophora glomerata Linn at
No (mg/ml) 517 nm.
Benzene Chloroform Ethanol Ethyl Methanol Petroleum
acetate Ether
1 0.5 3.744 6.349 4.342 2.371 27.464 14.836
2 1.0 8.183 13.227 7.196 5.039 40.845 18.442
3 1.5 13.453 24.338 10.669 7.509 52.112 21.065
4 2.0 19.278 34.126 15.012 10.671 71.830 23.196
5 2.5 27.045 42.857 18.858 13.438 86.619 26.672
6
Cladophora glomerata
5 Standard
at
4 Benzene
Absorbance
3 Chloroform
700 nm
Ethanol
2
Ethylacetate
1 Methanol
0 Pet. Ether
50 100 150 200 250
Concentration
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Study of phytochemical screening & antioxidant activities of Cladophora glomerata
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I.J.S.N., VOL.7 (3) 2016: 659-663 ISSN 2229 6441
of various extracts of Saragassum muticam, Intl. J. amylase inhibitory activity of methanol extract of
Pharmaceutical research and Development, 3(10):25-30. Colocasia esculenta corm. Pharmacologyonline. 2:715-721
Liyana-Pathiranan, C.M. and Shahidi, F. (2005) Zahin, M, Farukh, A. and Iqbal, A. (2009) The in vitro
Antioxidant activity of commercial soft and hard wheat antioxidant activity and total phenolic content of four
(Triticum aestivum L) as affected by gastric pH conditions. Indian medicinal plants. Int. J. of Pharm. and Pharm, Sci.
J. Agric. Food Chem. 53:2433-2440. 1: 88-95.
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