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Antioxidant Activity of Echinophora Platyloba DC Essential

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Antioxidant Activity of Echinophora Platyloba DC Essential Original Paper DOI: 10.22067/veterinary.v8i1.50620 Received: 14 October, 2015 Accepted after revision: 29 November, 2016 Published online: 10 April, 2017 Antioxidant activity of Echinophora platyloba DC essential oil: a comparative study on four different methods a b b Ali Ehsani, Mohammad Hashemi, Asma Afshari a Department of Food Science and Technology, Faculty of Nutrition, Tabriz University of Medical Sciences, Tabriz, Iran b Department of Nutrition, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran Keywords Echinophora platyloba, ABTS, DPPH, FRAP, β-caro- exhibited strong antioxidant activity in FRAP and BCBT tene bleaching test assays. It is concluded that FRAP and BCBT assays are more suitable spectrometric assays for antioxidant capacity Abstract evaluation of Echinophora platyloba. The present study is conducted in order to investigate Abbreviations the antioxidant capacity of Echinophora platyloba DC (Duncan) essential oil (EO), growing wild in west E. platyloba: Echinophora platyloba Azerbaijan, Iran, with four different assays. The aerial parts EO: Essential Oil BHT: Butylated Hydroxytoluene of Echinophora platyloba DC were provided from Maraghe DPPH: 2,2-Diphenyl-1-Picrylhydrazyl city district, northwest of Iran and its phytochemicals ABTS: 2,2’-Azino-Bis 3-ethylbenzothiazoline-6-Sul were determined using GC-MS analysis. In order to phonic acid evaluate and compare the antioxidant activity of the EO, FRAP: Ferric Reducing Antioxidant Power various concentrations (1, 2.5, 5 and 10 mg.ml-1) of the BCBT: β-Carotene Bleaching Test oil and reference antioxidants (ascorbic acid and BHT) SD: Standard Deviation were analyzed by four different assays such as 2,2’-azino- DC: Duncan bis 3-ethylbenzothiazoline-6-sulphonic acid (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH), Ferric reducing antioxidant power (FRAP) and β-carotene bleaching test (BCBT). The oil exhibited moderate antioxidant activity with a dose-response in DPPH and ABTS assays and it IRANIAN JOURNAL OF VETERINARY SCIENCE AND TECHNOLOGY Corresponding author: Mohammad Hashemi Department of Nutrition, Faculty of Medicine, Tel: +98 51 3880 3742 Fax: +98 51 3876 3852 Mashhad University of Medical Science, Mashhad, Iran. Web: ijvst.um.ac.ir Email: [email protected] Email: [email protected] Tel: +98 51 38002423 Echinophora platyloba DC essential oil as antioxidant IJVST 2016; VOLUME 8, NUMBER 1 47 Introduction Materials and Methods The genus Echinophora (family Umbelliferae, subfam- ily Apioideae, tribe Echinophoreae), is represented in the Plant material flora of Iran by four species two of which are endemics The aerial parts of E. platyloba DC were harvested at including Echinophora platyloba DC and Echinophora ci- the flowering stage (10th June to 15th August 2010) from nerea (Boiss). E. platyloba DC (with the Persian name wild grown plants in the Maragheh city district of East “Khosharizeh”, “Khosharouzeh” or “Tigh Toragh”) that is a Azerbaijan province, Iran. A voucher specimen (no. 6502) midsummer plant, i.e. one of the most fascinating species was deposited in the Herbarium of West Azerbaijan Agri- that wildly grows and is well known for its favorable aro- cultural and Natural Resource Center, Urmia, Iran. matic properties. Fresh and dried aerial parts of this spe- Extraction and analysis of E.platyloba were provided cies are used as a seasoning and anti-mold agent in pickled according to the method recommended by the European cauliflower, gherkin, and native dairy products like cheese, Pharmacopeia (Ahmad et al., 1999). The obtained chem- yoghurt and doogh (a savory yogurt-based beverage pop- ical composition of E.platyloba EO components were de- ular in Iran) in the west and northwest of Iran (Saei‐Deh- scribed by Hashemi et al., (2016). kordi et al., 2012). Different assays, including a wide range of spectro- DPPH radical-scavenging assay photometric assays have been employed to measure the This assay was performed as previously described with antioxidant capacity of EOs and their extracts. There is a minor modification (Erkan et al., 2008). 50 μL of various certain principle in most of these assays. This principle is concentrations (1, 2.5, 5 and 10 mg.ml-1) of the E. platy- that a colored radical or redox-active compound is gener- loba EO and reference antioxidants, ascorbic acid and ated; then the ability of a sample for scavenging the syn- BHT (Merck , India), in methanol were mixed with 2 ml of thetic radical or reducing the redox-active compound is methanolic DPPH solution (24 μg.mL-1). After an incuba- monitored using a spectrophotometer and an appropriate tion period of 60 min at room temperature in a dark place, standard such as ascorbic acid is applied to quantify anti- the absorbance at 517 nm, the wavelength of maximum oxidant capacity (Floegel et al., 2011). The most frequent- absorbance of DPPH, were measured as ASample, using ly used assays are 1,1-diphenyl-2-picrylhydrazyl (DPPH) a spectrophotometer (LKB Novaspec II; Pharmacia, Swe- and 2,2-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid den). A solution without the test material was also carried (ABTS) assays as well as β-carotene bleaching test (BCBT) out applying the same procedure and the absorbance was and ferric reducing antioxidant power (FRAP) (Ou et al., recorded as ABlank. The free radical-scavenging activity 2002; Thaipong et al., 2006). was calculated according to the following equation: Application of chemical antioxidants in food could Radical scavenging activity (%) = 100 × (ABlank – result in different negative health effects such as hepatic ASample )/ABlank damages, toxicities, and cancer. Therefore, a global interest is emerging in antioxidant characteristics of EOs (Rohani β-carotene bleaching test (BCBT) et al., 2011; Saei-Dehkordi et al., 2010; Saei‐Dehkordi et al., β-carotene bleaching test was carried out as previous- 2012). Antioxidants that have originated from plants have ly described with minor modifications (Miraliakbari and biological effects by scavenging free radicals and inhibiting Shahidi, 2008). A solution of 5 mg β-carotene in 10 ml oxidation. Application of these natural compounds with chloroform was prepared and 1 mL of this solution was antioxidant properties can prevent oxidation and extend added into a 100 ml round bottom flask. Chloroform was the shelf life of foods (Ani et al., 2006). On the other hand, removed under vacuum using a rotary evaporator (Hei- consumption of foods containing natural antioxidant may dolph laborta 4003, SchwaBach, Germany) at 40ºC.Then transmit these bioactive compounds to the body to play a 25 μL linoleic acid, 400 mg Tween 40 emulsifier and 100 role in the inhibition of oxidative damage due to free radi- ml of aerated distilled water were added to the flask and cals formed in the human cells (Sharafati-chaleshtori et al., shaken vigorously. Aliquots of 2.5 ml of this emulsion were 2012). To our best knowledge, there is no comprehensive transferred into a series of test tubes containing 350 μL of work focusing on antioxidant capacity of E. platyloba DC. various concentrations (1, 2.5, 5 and 10 mg.ml-1) of the E. EO. Moreover, no data have been published on the antioxi- platyloba EO and reference antioxidants (ascorbic acid and dant capacity of the oil of this plant grown in the northwest BHT). The absorbance of each tube was measured at 470 of Iran as one of the most geographically important regions nm immediately at time zero and subsequently over a two- of this plant’s growth due to its commonly conventional hour period at 20 min intervals while the tubes were kept consumption in various foods. Therefore, the present study in a water bath at 50ºC. The capacity of the E. platyloba EO is conducted to assess and compare the antioxidant prop- to protect against oxidation of β -carotene was determined erties of E. platyloba EO grown in the northwest of Iran as shown in the following equation: using four routine assays including DPPH, ABTS, BCBT I % = (Aβ-carotene after 2h assay / AInitial β-carotene) and FRAP. ×100 48 IJVST 2016; VOLUME 8, NUMBER 1 Ehsani/Hashemi/Afshari Table 1 DPPH radical scavenging activity of Echinophora platyloba DC essential oil Sample C 1 (mg/ml) C 2.5 (mg/ml) C 5 (mg/ml) C 10 (mg/ml) Essential oil 16.70 ± 3.17 aA 29.70 ± 1.40 bB 36.36 ± 1.25 cC 41.43 ± 2.81 dD Ascorbic acid 99.99 ± 0.00 aA 99.99 ± 0.00 aA 99.99 ± 0.00 aA 99.99 ± 0.00 aA BHT 99.98 ± 0.00 aA 99.98 ± 0.00 aA 99.99 ± 0.00 aA 99.99 ± 0.00 aA Same uppercase and lowercase letters indicate no significant differences within a row and column, respectively (p > 0.05). Ferric reducing antioxidant power (FRAP) assay wise comparison of the means which is indicated by sub- The FRAP assay was performed followed by the meth- script letters (for different concentrations c1-c10) or stars od previously described with some modifications (Benzie, (for different samples: EO, AA, and BHT). 1996). The working FRAP reagent was prepared by mix- ing 300 mM acetate buffer, pH 3.6, (3.1 g sodium acetate Results and 16 ml glacial acetic acid made up to 1 L with distilled The major compounds of the EO were p-cis-Ocimene water); 10mM 2,4,6-tri-2-pyridyl- 1,3,5-triazin (TPTZ) in (26.51%), 2,3 Dimethyl-1,3-cyclohexadiene (9.87%), Al- 40 mM HCL; 20 mM ferric chloride at 10:1:1. Aliquots of phapinene (7.96%), gamma-dodecalactone (5.84%) and 4.5 ml of the mixture were added into the tubes contain- Nerolidol (5.66%), representing 93.29% of the total content ing 150 μL of various concentrations (1, 2.5, 5 and 10 mg/ of the oil (Hashemi et al., 2016).
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