Postepy Hig Med Dosw. (online), 2013; 67: 958-963
Original Article
Full Text PDF  

Antioxidant activity of selected phenols estimated by ABTS and FRAP methods
Aktywność przeciwutleniająca wybranych fenoli oznaczona testem ABTS i FRAP
Izabela Biskup1  ABCDEF, Iwona Golonka2  BCDEFG, Andrzej Gamian3,4  D, Zbigniew Sroka1  ADG
1Department of Pharmacognosy, Silesian Piasts Wrocław Medical University, Wrocław, Poland
2Department of Physical Chemistry, Silesian Piasts Wrocław Medical University, Wrocław, Poland
3Department of Immunology of Infectious Diseases, Institute of Immunology and Experimental Therapy, Wrocław, Poland
4Department of Medical Biochemistry, Silesian Piasts Wrocław Medical University, Wrocław, Poland
Corresponding author
Dr Iwona Golonka, Department of Physical Chemistry, Silesian Piasts Wrocław Medical University, pl. Nankiera 1, 50-140 Wrocław, Poland; e-mail: iwona.golonka@am.wroc.pl

Authors' Contribution:
A - Study Design, B - Data Collection, C - Statistical Analysis, D - Data Interpretation, E - Manuscript Preparation, F - Literature Search, G - Funds Collection

Source of support
The study was supported partly by internal grant ST-527 and partly by a research fellowship within the "Develop­ment program of Wroclaw Medical University" funded from the European Social Fund. Human Capital. National Cohesion Strategy (contract no. 184 UDA-POKL.04.01.01-00-010/08-01)"

Received:  2012.09.18
Accepted:  2013.05.27
Published:  2013.09.10

Summary
Introduction: Phenols are the most abundant compounds in nature. They are strong antioxi­dants. Too high level of free radicals leads to cell and tissue damage, which may cause asthma, Alzheimer disease, cancers, etc. Taking phenolics with the diet as supplements or natural medicines is important for homeostasis of the organism.
Materials and methods: The ten most popular water soluble phenols were chosen for the ex­periment to investigate their antioxidant properties using ABTS radical scavenging capacity assay and ferric reducing antioxidant potential (FRAP) assay.
Results and discussion: Antioxidant properties of selected phenols in the ABTS test expressed as IC50 ranged from 4.332 µM to 852.713 µM (for gallic acid and 4- hydroxy­phenylacetic acid respectively). Antioxidant properties in the FRAP test are expressed as µmol Fe2+/ml. All examined phenols reduced ferric ions at concentration 1.00 x 10-3 mg/ml. Both methods are very useful for determination of antioxidant capacity of water soluble phenols.
Key words: antioxidants • phenols • ABTS • FRAP




Introduction
Phenols (and other antioxidant compounds) are very significant for human health. They are widely present in fruits, vegetables, and beverages: coffee, tea, fruit juices, wines and other products [12,18,21,22,31,36]. In plants, phenolic acids occur mainly in bound forms, as components of complex structures such as lignins and hydrolysable tannins or derivatives of sugars and organic acids [14,17]. Two main classes of phenolic ac­ids exist: hydroxybenzoic acids (e.g. gallic, p-hydroxy­benzoic, vanillic, syringic, protocatechuic acids) and hydroxycinnamic acid (e.g. coumaric, caffeic, ferulic, sinapic acid). The antioxidant activity of phenolic ac­ids is related to the quantity, number and position of hydroxyl groups in the molecule [4,27]. Electrochemi­cal reduction potentials determined by cyclic voltam­metry give information about the capacity of reducing compounds and their results indicate that reactions involving phenols are not always reversible [8]. Phe­nolic compounds can act in many different ways [6]:
• chelating metals such as iron and copper, which can prevent their involvement in Fenton reactions that can generate high concentrations of hydroxyl radicals,
• breaking the chain of reactions triggered by free radi­cals,
• slowing down or accelerating enzyme activity.
Phenolic acids are involved in repair and adaptive sys­tems, and can act preventively and sometimes therapeu­tically in various diseases: cardiovascular, respiratory, dermatological and digestive system diseases [16,25,32]. The relationship between total phenolic and flavonoid contents and antioxidant, anti-inflammatory or antibac­terial activities is strong in many plants [10,11,13,23]. The main antioxidants of olive oil reduce the activity of xan­thine oxidase, an enzyme potentially involved in carcino­genesis [35]. Oleuropein and p-hydroxy benzoic, vanillic and p-coumaric acids exhibit antimicrobial activity. They completely inhibit the growth of Escherichia coli, Klebsiella pneumoniae and Bacillus cereus [1]. UV radiation damages the skin by increasing levels of free radicals. For their elimination antioxidants contained in creams and diet are helpful [37]. Pollution, cigarette smoke, drugs, illnesses, stress etc. can increase levels of free radicals in the human body and disrupt homeostasis. The consequent damage may contribute to the development of Alzheimer's dis­ease, Parkinson's disease, dementia, atherosclerosis and asthma [3,9,15,20,34,38]. Several methods are available to evaluate antioxidant activities of natural compounds.
The aim of this research was to estimate the antioxi­dant activity of selected water-soluble phenols, sup­plied in food of vegetable origin. These compounds dif­fer in structure, especially the number and location of substitution of hydroxyl groups, which has a strong in­fluence on their antioxidant properties. Two methods with different mechanisms were used: ABTS assay with 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) based on free radical scavenging [26] and FRAP (ferric reducing antioxidant power) assay based on reduction of ferric ion by phenol [2]. In addition, comparison of the efficiency of these two assays was done.
Materials and methods
Phenols shown in Fig. 1 were obtained from Extrasyn­thesis (4-hydroxyphenylacetic acid (10) and 3,4-dihy­droxyphenylacetic acid (3)), Fluka (3-caffeoylquinic acid= chlorogenic acid (6); 3,4-dihydroxybenzoic acid= protocatechuic acid (7); 3,5-dihydroxybenzoic acid= α- resorcylic acid (8); 2,3-dihydroxybenzoic acid= o-pyrocatechuic acid (4); 2,4- dihydroxybenzoic acid= β-resorcylic acid (9); 2,5-dihydroxybenzoic acid= gen­tisic acid (5)), Mallinckrodt GmbH (1,2,3-trihydroxy­benzene= pyrogallol (1)), Polskie Odczynniki Chemic­zne (3,4,5-trihydroxybenzoic acid= gallic acid (2)). Stock solutions of phenols were prepared by dissolution in distilled water. All measurements were done on the Ce­cil CE spectrophotometer model 3021.
Figure 1. Structures of selected phenols

ABTS ASSAY
Chemicals: Trolox (Hoffman-La Roche) (6-hydroxy- 2,5,7,8-tetramethylchroman-2- carboxylic acid; Sigma Aldrich) was used an antioxidant standard. Trolox (2.5 mM) was prepared in methanol for use as a stock stan­dard. Working standards were prepared daily on dilution with methanol. ABTS, 2,2'-azinobis(3-ethylbenzothia­zoline-6-sulfonic acid) diammonium salt, and potassi­um persulfate (di-potassium peroxdisulfate) were ob­tained from Sigma-Aldrich and HPLC grade methanol from POCh.
Assay: Experiments were performed according to [26] with small modifications. ABTS and potassium persul­fate were dissolved in distilled water to a final concen­tration of 7 mM and 2.45 mM respectively. These two solutions were mixed and the mixture allowed to stand in the dark at room temperature for 16 h before use in order to produce ABTS radical (ABTS•+). For the study of phenolic compounds the ABTS radical solution was diluted with distilled water to an absorbance of 1.00 at 734 nm.
Phenols (final concentrations 0.0001-0.01 mg/ml) or Trolox standards (final concentration 0-20 mM) were added to diluted ABTS•+ solution and the absorbance reading was taken 6 min after mixing using the spec­trophotometer. Results are presented as the ability of phenols to scavenge 50% of free radical ABTS • + (IC50) and TEAC (Trolox equivalent antioxidant capacity). Pa­rameters IC50 (µM) and TEAC (µM) were determined with a relative uncertainty of less than five percent.
All determinations were carried out in triplicate.
FRAP ASSAY
Chemicals: TPTZ (= 2,4,6-tripyridyl-s-triazine) (Fluka Analytical) was dissolved in HCl (POCh) for a few hours. FeSO4* 7H2O (= iron (II) sulfate heptahydrate) (Acros Or­ganics) dissolved in deionized water was used as standard solution. FeCl3*6H2O (= iron (III) chloride hexahydrate) was dissolved in deionized water for a few minutes prior to analysis.
Assay: Experiments were done according to [2] with modi­fications. FRAP working solution was prepared freshly each time: 0.3 M acetate buffer (pH=3.6), 0.01 M TPTZ (2,4,6-tripyridyl-s-triazine) in 0.04 M HCl and 0.02 M FeCl3*6H2O were mixed in 10:1:1 (v/v/v) and kept away from light. 0.075 ml of phenols (final concentration 0.0001-0.01 mg/ml) or FeSO4* 7H2O (final concentration 0-1.8 µmol Fe2+ /ml) solution were added to 2.25 ml FRAP working solution and 0.225 ml of deionized water. The mixture was vortexed and incubated at 37°C for 30 min away from light. Absorbance was measured at 593 nm us­ing the spectrophotometer. FRAP working solution with deionized water instead of a sample was used as a blank. All determinations were carried out in triplicate.
Results
Structurally different water-soluble phenols were com­pared: phenylacetic acids, cinnamic acids, hydroxyben­zoic acid derivatives and ester of caffeic acid with quinic acid. At least three different concentrations were chosen for each tested compound in both methods in order to avoid speculation of how sample concentration influences the antioxidant capacity [7].
Table 1 shows the antioxidant activity of phenols ex­pressed as the ability to scavenge 50% of free radical ABTS•+ (IC50) and TEAC (Trolox equivalent antioxidant ca­pacity). Phenols with a higher number of hydroxyl groups showed higher antioxidant capacity, which is expressed as the lowest IC50 value, 4.332 µM and 5.228 µM respectively for gallic acid and pyrogallol. In contrast, the highest IC50 value was obtained for 4-hydroxyphenylacetic acid (852.713 µM), almost 200 times greater, which confirms the stronger antioxidant activity of phenols with a higher number of hydroxyls [30].
Table 1. Antioxidant properties of selected phenolics expressed as the ability to scavenge 50% of free radical ABTS•+ (IC50) and TEAC (Trolox equivalent antioxidant capacity)

Antioxidant activity of selected phenols determined by FRAP is depicted in Fig. 2a,b.
Figure 2. Antioxidant activity (FRAP) of the various phenols; a - For strong antioxidants, b - For weak antioxidants

All phenols reduce Fe3+ ions to Fe2+ ions which are com­plexed by TPTZ at a concentration of 1.00 x10-3 mg/ ml. Phenol antioxidant activity at this concentration decreased in the order o-pyrocatechuic acid>3,4-dihy­droxyphenylacetic acid>pyrogallol>gallic acid>gentisic acid>protocatechuic acid>chlorogenic acid>α-resorcylic acid>4- hydroxyphenylacetic acid> β-resorcylic acid. In­hibitory effects of radicals have been identified for all compounds and their strength depends on the concentra­tion of phenol. No reaction occurred for the concentra­tion of 1.00x10-4 mg/ml of chlorogenic acid and gentisic acid. The remaining eight phenols with the same con­centration have antioxidant properties in the following decreasing order: pyrogallol> o-pyrocatechuic acid>3,4- dihydroxyphenylacetic acid> protocatechuic acid>gallic acid> 4-hydroxyphenylacetic acid>β-resorcylic acid>α- resorcylic acid. In the case of chlorogenic acid at a con­centration of 5x10-4 mg/ml it also does not absorb at 593 nm but 3,4-dihydroxyphenylacetic acid, o-pyrocatechuic acid and pyrogallol are still in the top three most pow­erful antioxidants. Phenols at concentrations 2.50 x10-3, 5.00 x10-3, 1.00 x10-2, 5.00 x10-2, 1.00 x10-1 (mg/ml) in most cases showed a very significant reduction of Fe3+ ions to Fe2+ ions and thus exceeded the scope of the absorbance.
The largest correlation was observed between ABTS method and FRAP method for phenols at a concentra­tion 1 x 10-3 (1), 5 x 10-4 (2), 1 x 10-4 (3) mg/ml as shown in Table 2. The results obtained using linear regression analysis were used to fit a line to a set of experimental results using the "least squares". The correlation coef­ficient in these cases is 0.746, 0.783, 0.786 and all depen­dencies can be considered as statistical type I error not more than p=0.0023.
Table 2. Correlation between ABTS method and FRAP method for phenols at concentration 1 x 10-3 (1), 5 x 10-4 (2), 1 x 10-4 (3) mg/ml

The correlation coefficient between ABTS method and FRAP method for three phenols at a concentration 2.5 x 10-3 (1) mg/ml is 0.997 and type I error p=0.0259. For the other, dependence on type I error is greater than 0.05; this means that no correlation exists between the two methods.
Discussion
Antioxidant activity depends on the number and posi­tion of the hydroxyl groups of the aromatic ring binding site and the type of substituent. The strongest antioxi­dant activity in the ABTS method was achieved by phe­nols with 3 hydroxyl groups from all phenols tested in this experiment. The carboxylic group slightly increases this activity (lower IC50 for gallic acid compared to IC50 for pyrogallol). The next antioxidants in line are two hy­droxylated phenolic acids with the strongest,3,4-dihy­droxyphenylacetic acid, among them. It was proved that the type of spacer between the carboxylic acid and ben­zene ring influences their activity, which increases for the methylenic group in phenolic acids [27] Our results confirmed this thesis. What is more, phenols with two hydroxyl groups linked to the aromatic ring at the ortho position more strongly quench ABTS•+ radical than com­pounds with hydroxyl groups substituted at the meta position [30]. Also presence of a carbonyl group like es­ter (chlorogenic acid) enhances antioxidant activity [5]. So far, the great majority of antioxidant assays have been based on methanol as a solvent. The present results are in agreement with this, which makes the ABTS method equal for methanolic and watery solutions. Pyrogallol, o-pyrocatechuic acid and 3,4 dihydroxyphenylacetic acid have the strongest antioxidant properties identified by the FRAP method. The amount of Fe3+ ions reduced to Fe2+ ions varies for different concentrations of phe­nols. Phenols with two hydroxyl groups bonded to the aromatic ring in the ortho position, such as o-pyrocate­chuic acid and 3,4 dihydroxyphenylacetic acid, showed strong antioxidant anti-radical activity at a concentra­tion of 1 x 10-3 and 5 x 10-4 mg/ml but pyrogallol with three hydroxyl groups bonded to the aromatic ring at a concentration of 1 x 10-4 mg/ml. 3,5-Dihydroxybenzoic (α- resorcylic) and 2,4-dihydroxybenzoic (β-resorcylic) acids with two hydroxyls bonded in the meta position in relation to each other are weak antioxidants. The results confirm earlier studies [30]. 4-Hydroxyphenylacetic acid behaves like the previous two acids but is involved in scavenging reactive oxygen and nitrogen species both in vitro and in vivo [19,33].
To sum up, these two methods complement one another and give full information of antioxidative capacity of wa­ter-soluble phenols. The correlation between ABTS and FRAP methods was indicated by Nilsson et al. [24]. The ABTS method for investigating hydroxybenzoic and hy­droxycinnamic acids takes advantage of the fact that they probably act as radical scavengers owing to their hydro­gen and electron donating capacity and their ability to delocalize/stabilize the resulting phenoxyl radical within the structure [28,29], while the FRAP method is based on Fe3+ being reduced to Fe2+ by the examined substance.
REFERENCES
[1] Aziz N.H., Farag S.E., Mousa L.A., Abo-Zaid M.A.: Comparative antibacterial and antifungal effects of some phenolic compounds. Microbios, 1998; 93: 43-54
[PubMed]  
[2] Benzie I.F., Strain J.J.: The ferric reducing ability of plasma (FRAP) as a measure of "antioxidant power": the FRAP assay. Anal. Biochem., 1996; 239: 70-76
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[3] Dworski R.: Oxidant stress in asthma. Thorax, 2000; 55 (Suppl. 2): S51-S53
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[4] Fukumoto L.R., Mazza G.: Assessing antioxidant and prooxidant activities of phenolic compounds. J. Agric. Food Chem., 2000; 48: 3597-3604
[PubMed]  
[5] Göcer H., Gülcin I.: Caffeic acid phenethyl ester (CAPE): correlation of structure and antioxidant properties. Int. J. Food Sci. Nutr., 2011; 62: 821-825
[PubMed]  
[6] Halliwell B., Gutteridge J.M.: Role of free radicals and catalytic metal ions in human disease: an overview. Methods Enzymol., 1990; 186: 1-85
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[7] Hengst C.,Werner S., Müller L., Fröhlich K., Böhm V.: Determination of the antioxidant capacity: influence of the sample concentration on the measured values. Eur. Food Res. Technol., 2009; 230: 249-254
[Abstract]  
[8] Joergensen L.V., Madsen H.L., Thomsen M.K., Dragsted L.O., Skibsted L.H.: Regeneration of phenolic antioxidants from phenoxyl radicals: an ESR and electrochemical study of antioxidant hierarchy. Free Radic. Res., 1999; 30: 207-220
[PubMed]  
[9] Kanazawa H., Kurihara N., Hirata K., Takeda T.: The role of free radicals in airway obstruction in asthmatic patients. Chest, 1991; 100: 1319-1322
[PubMed]  
[10] Kang J., Xie C., Li Z., Nagarajan S., Schauss A.G., Wu T., Wu X.: Flavonoids from acai (Euterpe oleracea Mart.) pulp and their antioxidant and anti-inflammatory activities. Food Chem., 2011; 128: 152-157
[Full Text HTML]  [Full Text PDF]  
[11] Karou D., Dicko M.H., Simpore J., Traore A.S.: Antioxidant and antibacterial activities of polyphenols from ethnomedicinal plants of Burkina Faso. Afr. J. Biotechnol., 2005; 4: 823-828
[Abstract]  [Full Text PDF]  
[12] Kasprzyk A., Żbikowska B., Sroka Z., Gamian A.: The antiradical activity of some plant raw materials and extracts obtained from these raw materials. Postępy Hig. Med. Dośw., 2012; 66: 146-152
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[13] Kiessoun K., Souza A., Meda N.T., Coulibaly A.Y., Kiendrebeogo M., Lamien-Meda A., Lamidi M., Millogo-Rasolodimby J., Nacoulma O.G.: Polyphenol contents, antioxidant and anti-inflammatory activities of six Malvaceae species traditionally used to treat hepatitis B in Burkina Faso. Eur. J. Scientific Res., 2010; 44: 570-580
[14] Klick S.; Herrmann K.: Glucosides and glucose esters of hydroxybenzoic acids in plants. Phytochem., 1988; 27: 2177-2180
[Abstract]  
[15] Koutsilieri E., Scheller C., Grünblatt E., Nara K., Li J., Riederer P.: Free radicals in Parkinson's disease. J. Neurol., 2002; 249 (Suppl. 2): ii01-ii05
[PubMed]  
[16] Kowalska K.: Natural compounds involved in adipose tissue mass control in in vitro studies. Postępy Hig. Med. Dośw., 2011; 65: 515-523
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[17] Lam T.B., Kadoya K., Iiyama K.: Bonding of hydroxycinnamic acids to lignin: ferulic and p-coumaric acids are predominantly linked at the benzyl position of lignin, not the β-position, in grass cell walls. Phytochemistry, 2001; 57: 987-992
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[18] López M., Martínez F., Del Valle C., Orte C., Miró M.: Analysis of phenolic constituents of biological interest in red wines by high-performance liquid chromatography. J. Chromatogr. A, 2001; 922: 359-363
[PubMed]  
[19] Mani A.R., Pannala A.S., Orie N.N., Ollosson R., Harry D., Rice-Evans C.A., Moore K.P.: Nitration of endogenous para-hydroxyphenylacetic acid and the metabolism of nitrotyrosine. Biochem. J., 2003; 374: 521-527
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[20] Mao P.: Oxidative stress and its clinical applications in dementia. J. Neurodegenerative Dis., 2013; 2013: 319898
[Full Text HTML]  [Full Text PDF]  
[21] Matilla P., Hellström J.: Phenolic acids in potatoes, vegetables, and some of their products. J. Food Comp. Anal., 2007; 20: 152-160
[Abstract]  [Full Text HTML]  [Full Text PDF]  
[22] Minussi R.C., Rossi M., Bologna L., Cordi L., Rotilio D., Pastore G.M., Durán N.: Phenolic compounds and total antioxidant potential of commercial wines. Food Chem., 2003; 82: 409-416
[Full Text HTML]  [Full Text PDF]  
[23] Mothana R.A., Al-Said M.S., Al-Rehaily A.J., Thabet T.M., Awad N.A., Lalk M., Lindequist U.: Anti-inflammatory, antinociceptive, antipyretic and antioxidant activities and phenolic constituents from Loranthus regularis Steud. ex Sprague. Food Chem., 2012; 130: 344-349
[Full Text HTML]  [Full Text PDF]  
[24] Nilsson J., Pillai D., Önning G., Persson C., Nilsson A., Akesson B.: Comparison of the 2,2'-azinobis-3-ethylbenzotiazo-line-6-sulfonic acid (ABTS) and ferric reducing anti-oxidant power (FRAP) methods to assess the total antioxidant capacity in extracts of fruit and vegetables. Mol. Nutr. Food Res., 2005; 49: 239-246
[PubMed]  [Full Text HTML]  
[25] Piasek A., Bartoszek A., Namieśnik J.: Phytochemicals that counteract the cardiotoxic side effects of cancer chemotherapy. Postępy Hig. Med. Dośw., 2009; 63: 142-158
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[26] Re R., Pellegrini N., Proteggente A., Pannala A., Yang M., Rice-Evans C.: Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic. Biol. Med., 1999; 26: 1231-1237
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[27] Rice-Evans C.A., Miller N.J., Paganga G.: Structure-antioxidant activity relationship of flavonoids and phenolic acids. Free Radic. Biol. Med., 1996; 20: 933-956
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[28] Silva A.M., Santos C.M., Cavaleiro J.A., Tavares H.R., Borges F., Silva F.A.: NMR studies on the antiradical mechanism of phenolic compounds towards 2.2-diphenyl-1-picrylhydrazyl radical. In: Webb G.A., Belton P.S., Gil A.M., Delgadillo I. editors. Magnetic resonance in food science - a view to the future. 8th ed. London: Royal Society Chemistry, 2001; 110-116
http://pubs.rsc.org/en/content/chapter/bk9780854048700-00110/978-0-85404-870-0
[29] Silva F.A., Borges F., Ferreira M.A.: Effects of phenolic propyl esters on the oxidative stability of refined sunflower oil. J. Agric. Food Chem., 2001; 49: 3936-3941
[PubMed]  
[30] Sroka Z., Cisowski W.: Hydrogen peroxide scavenging, antioxidant and anti-radical activity of some phenolic acids. Food Chem. Toxicol., 2003; 41: 753-758
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[31] Sroka Z., Gamian A., Cisowski W.: Low-molecular antioxidant compounds of natural origin. Postępy Hig. Med. Dośw., 2005; 59: 34-41
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[32] Szumiło J.: Protocatechuic acid in cancer prevention. Postępy Hig. Med. Dośw. 2005; 59: 608-615
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[33] Takahama U., Oniki T., Murata H.: The presence of 4-hydroxyphenylacetic acid in human saliva and the possibility of its nitration by salivary nitrite in the stomach. FEBS Lett., 2002; 518: 116-118
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[34] Tęgowska E., Wosińska A.: The role of biological sciences in understanding the genesis and a new therapeutic approach to Alzheimer's disease. Postępy Hig. Med. Dośw., 2011; 65: 73-92
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[35] Tripoli E., Giammanco M., Tabacchi G., Di Majo D., Giammanco S., La Guardia M.: The phenolic compounds of olive oil: structure, biological activity and beneficial effects on human health. Nutr. Res. Rev., 2005; 18: 98-112
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[36] Vinson J.A., Dabbagh Y.A.: Tea phenols: Antioxidant effectiveness of teas, tea components, tea fractions and their binding with lipoproteins. Nutr. Res., 1998; 18: 1067-1075
[Full Text HTML]  [Full Text PDF]  
[37] Viola P., Viola M.: Virgin olive oil as a fundamental nutritional component and skin protector. Clin. Dermatol., 2009; 27: 159-165
[PubMed]  [Full Text HTML]  [Full Text PDF]  
[38] Vogiatzi G., Tousoulis D., Stefanadis C.: The role of oxidative stress in atherosclerosis. Hellenic J. Cardiol., 2009; 50: 402-409
[PubMed]  [Full Text PDF]  
The authors have no potential conflicts of interest to declare.