Oxidation Communications 33, No 1, 167–174 (2010)
Study on Phenolics and Antioxidant Activity of Selected Wines from Some Greek Aegean Sea Islands N. Gougoulias Department of Plant Production, Techological Educational Institute of Larissa, 41 110 Larissa, Greece E-mail:
[email protected] ABSTRACT In this work, 17 red, white and rose wines of different quality (W.A.O.H.Q., W.A.O., table wine and traditional), from the Rodos, Limnos, Paros and Santorini islands, produced on the basis of traditional local grape varieties Monemvasia, Mandilaria, Athiri, Assyrtiko, Aidani and Limnio, and selected imported grape varieties were assayed in relation to their total phenols (ТР) content, antiradical (DPPH) activity, ferric reducing antioxidant power (FRAP) and antioxidant capacity (ЕС50). It has been found that the content of TP is ranging from 120 to 2850 mg/l, the antiradical activity – from 0.34 to 6.34 μmol DPPH/ml, and the antioxidant activity – from 1.15 to 22.60 μmol FRAP/ml wine. Among the investigated wines, the wines from the Paros and Santorini islands have higher amount of TP, higher antiradical activity and higher efficiency (ЕС50) of phenol compounds. Keywords: wine, total phenols (ТР), DPPH activity, FRAP activity, ЕС50. Aims and background Phenol compounds are one of the most widely spread secondary compounds in plants, and are present in many plan-derived foods and beverages1,2. Recently, the interest to them has increased strongly due to the establishment of a number of physiological effects on the human organism, and first of all due to their antioxidant properties3–5. Grape, wine and other grape products are rich in phenol compounds, particularly in flavonoids and phenolic acids, which exhibit a strong antioxidant effect6–10. Many epidemiological studies have established an inverse correlation between the moderate wine consumption and mortality of coronary heart diseases in France, Italy and other countries although the high fat consumption. This phenomenon is known as the ‘French paradox’11,12. It has been found that polyphenols in wine, defending the human organism from the action of free oxygen radicals (ROS), exert a strong antioxidant action, and thus inhibit the risk of heart diseases as well as from other diseases such atherosclerosis, ageing-related changes of eyes, nervous and immune system, cancer, 167
etc.13,14 Polyphenols present in fruits, vegetables, tea and wine become a topic of increasing interest. A number of studies have been devoted to investigating the phenol contents and antioxidant effectiveness of Greek wines15–17. The aim of the present study is to provide information for the content of total phenols in wines of different quality from some Greek Aegean sea islands – Rodos, Limnos, Paros and Santorini. Experimental Wines. Seventeen wines, 7 of which are of quality according to the European standard as ‘Wine of Appellation Origin of High Quality (W.A.O.H.Q.) or in Greek ‘ОРАР wines’, 5 wines of quality ‘Wine of Appellation Origin(W.A.O.), in Greek ‘ТOPIKO’, 4 wines – ‘table wine’, in Greek ‘epitrapezio’ and 1 wine – ‘traditional’, in Greek –‘paradosiako’ were the object of the present study. Among the investigated wines 6 are red, 10 – white and 1 – rose (Table 1). Table 1. Investigated wines from the islands
No Designation 1 Chevalier de Rhodes 2 Rodos 2400 3 Archondiko 4 5 6 7 8
Dodekanisiakos Egeopelagitikos Grand Rose Moulin Limnos (retsina)
9 10 11 12
Limnos Aroma Limnou Egeopelagitikos Kalabaki
13 Paros 14 Paros 15 Egeopelagitikos 16 Santorini 17 Vedema
Colour Grape variety Rodos island red Mandilaria white Athiri red Cabernet Sauvignon + Grenache Rouge white Athiri white Athiri red Amorgiano white Athiri white Athiri Limnos island white Muscat Alechandrias white Muscat Ottonel white Assyrtiko + Muscat red Limnio Paros island red Monemvasia + Mandilaria white Monemvasia red Monemvasia + Mandilaria Santorini island white Aidani + Assyrtiko + Athiri white Aidani + Assyrtiko + Athiri
Appellation W.A.O.H.Q. W.A.O.H.Q. W.A.O. W.A.O. W.A.O. table wine table wine traditional W.A.O.H.Q. W.A.O.H.Q. W.A.O. table wine W.A.O.H.Q. W.A.O.H.Q. W.A.O. W.A.O.H.Q. table wine
Determination of total polyphenols content. Total polyphenols (TP) contents were determined with the Folin–Ciocalteu (F.–C.) reagent according to the method of Sin168
gleton and Rossi and its micro-variant18,19: to 1580 μl distilled water were added 20 μl appropriately diluted wine, 100 μl the F.–C. reagent and 360 μl solution of Na2CO3. After 2 h the extinction at 750 nm was measured on a UV-9200 spectrophotometer (England). The phenol contents were expressed as gallic acid equivalent (GAE) and catechin equivalent (CE). Determination of antiradical activity. The antiradical activity was determined according to the method of Brand-Williams et al.20, with the stable radical 2,2′-diphenyl-1picrylhydrazyl (DPPH). The activity was evaluated in µmol DPPH/ml wine as well as in mg Trolox equivalent (synthetic vitamin E) per 100 ml wine. Determination of the antioxidant activity (FRAP). The ferric reducing antioxidant power (FRAP) was evaluated according to the method of Benzie et al.21 and was expressed as µmol FRAP reagent/ml wine. The activity was also presented as mg ascorbic acid (AA, vitamin C) per 100 ml wine Determination of the antioxidant efficiency (EC50). The antioxidant efficiency (EC50) was evaluated employing a DPPH solution according to the method of Vinson et al.22 All results of the chemical analyses were the mean of three replications and the standard deviations (sd) were calculated. The correlation coefficients and the statistical significance were determined using MINITAB RELLASE 13-ANNOVA ANALYSIS program according to well-accepted methods. RESULTS AND DISCUSSION The results for TP content in the investigated wines are shown in Table 2. They vary in wide ranges depending on the wine type, grape cultivar, climatic conditions and wine-making technologies. The concentration of TP determined with the Folin–Ciocalteu reagent varies from 1600 to 2850 mg/l GAE (2200 mg/l on average) for the red wines, and from 120 to 480 mg/l GAE (299 mg/l on average) for the white wines. Our results are in accordance with the data of other authors who have established a higher TP content in the red wines than in the white ones, which is related to the differences in the enological treatments at wine-making process6,8,23,24. It has been found that the TP levels in the red wines do not depend on the quality type of the wine (W.A.O.H.Q., W.A.O. or table wine), but are in close dependence on the grape variety type and wine-making technology. The comparison of the results with other authors data reveals that the wines from the Aegean sea, produced from local grape varieties, are characterised by a moderate content of total phenols7,8,15. The content of TP in the white wines is closely related with the grape variety and the ecological conditions of vine breeding. The white wine Paros (No 14, W.A.O.H.Q.), which in fact is the richest in phenol substances (480 mg/l GAE), is produced on the Paros island from the Monemvasia grape variety. It should be noted the high level of total phenols in the white wines Santorini (No 16, W.A.O.H.Q.), and Vedema (No 17, table wine), 169
(415 mg/l GAE on average), the two being produced from one and the same white grape varieties – Assyrtiko, Aidani and Athiri, grown on the Santorini island. The content of TP in the white wines of the Rodos island varies from 120 to 345 mg/ml GAE, although they are produced from one and the same grape variety – Athiri. This indicates its dependence on wine-making technological processes and wine quality. Our findings about the relation between the TP contents in the white wines of high quality (W.A.O.H.Q.), and the grape variety type produced on the islands are in accordance with the results of other authors. However, for some other types of wines the results reveal a strong dependence on the wine-making technology. Table 2. Content of total phenols (ТР)
No Designation
1 2 3 4 5 6 7 8
Chevalier de Rhodes Rodos 2400 Archondiko Dodekanisiakos Egeopelagitikos Grand Rose Moulin Limnos (retsina)
9 10 11 12
Limnos Aroma Limnou Egeopelagitikos Kalabaki
13 14 15
Paros Paros Egeopelagitikos
16 17
Santorini Vedema
Total phenols (mg/l)
GAЕ Rodos island 1600 2464 230 354 2200 3388 310 447 345 531 1400 2156 120 185 320 493 Limnos island 240 370 200 309 210 323 2150 3311 Paros island 2500 3850 481 739 2850 4389 Santorini island 430 662 400 616
СЕ
29 12 88 18 17 43 6 23
73 18 103 37 24 59 14 41
14 16 9 94
23 26 29 112
34 56 25
137 68 196
18 14
39 43
The results for the antiradical activity determined after the discolouring of the DPPH radical by wine phenolics are given in Table 3. The antiradical activity expressed as μmol DPPH/ml wine varies in wide ranges – from 0.32 to 6.34 μmol DPPH, depending on the wine colour. The red wines are characterised by a higher antiradical activity than the white wines, on average by 5.32 and 0.64 μmol DPPH/100 mol wine. The red and white wines from the Paros island (Nos 13 and 14), both being of high quality (W.A.O.H.Q.) are distinguished by the highest antiradical activity – 6.34 and 1.96 μmol DPPH/ml wine, respectively. Although the two red wines from the Paros 170
island are produced from one and the same local grape varieties, i.e. Monemvasia and Mandilaria, the wine of the higher quality (No 13) has a higher activity than the wine Egeopelagitikos (No 15), which has a higher TP content. The comparison of the antiradical activity of the white wines from the Rodos island produced from the Athiri grape variety reveals that the activity depends not only on the TP level, but also on the technology for the production of different quality wines. Among the white wines produced on the Limnos island, the wine Aroma Limnou (No 10), which is on the basis of the European grape variety Muscat Ottonel, has a higher antiradical activity (1.07 μmol DPPH/ml) in comparison with the wines Limnos (No 9) from Muscat Alechandrias and Egeopelagitikos from Assyrtiko and Muscat, regardless of their similar TP content. Table 3. Antiradical activity
No Designation
1 2 3 4 5 6 7 8
Chevalier de Rhodes Rodos 2400 Archondiko Dodekanisiakos Egeopelagitikos Grand Rose Moulin Limnos (retsina)
9 10 11 12
Limnos Aroma Limnou Egeopelagitikos Kalabaki
13 14
Paros Paros
15
Egeopelagitikos
16 17
Santorini Vedema
DPPH activity mg Trolox/100 ml µmol DPPH/ml Rodos island 4.53 0.13 412 0.32 0.02 29 5.34 0.35 486 0.55 0.09 50 0.45 0.03 41 2.85 0.12 259 0.40 0.03 36 0.49 0.02 45 Limnos island 0.33 0.03 30 1.07 0.08 97 0.34 0.05 31 4.51 0.15 410 Paros island 6.34 0.24 577 1.46 0.05 178 5.79 Santorini island 0.57 0.54
0.30
527
0.06 0.04
52 49
The antioxidant assay as measured by the FRAP method and expressing the ferric reducing power of phenol compounds is presented in Table 4. The red wines exert a higher ferric reducing power that the white ones, by 20.53 and 2 µmol FRAP/ml wine on average. The highest antioxidant activity was manifested by the Egeopelagitikos wine (No 15, W.A.O.) produced from the Mandilaria and Monemvasia grape varieties grown on the Paros island, which has also the highest TP content. Among the white 171
wines, the high quality wines (W.A.O.H.Q.), Santorini (No 16 – 3.48 µmol FRAP) and Paros (No 14 – 3.18 µmol FRAP), produced on the basis of different local grape varieties, but containing similar TP content, exert the highest ferric reducing capacity. The rose wine, Grand Rose (No 6, table wine), although the high TP content, has lower antioxidant activity than the high-quality wine Chevalier de Rhodes (No 1), which has relatively low TP content (1600 mg/l), and similar variety composition – Monemvasia, called also Amorgiano. Table 4. Antioxidant activity
No Designation
1 2 3 4 5 6 7 8
Chevalier de Rhodes Rodos 2400 Archondiko Dodekanisiakos Egeopelagitikos Grand Rose Moulin Limnos (retsina)
9 10 11 12
Limnos Aroma Limnou Egeopelagitikos Kalabaki
13 14
Paros Paros
15
Egeopelagitikos
16 17
Santorini Vedema
FRAP activity mg Trolox/100 ml µmol DPPH/ml Rodos island 21.92 0.30 193 2.15 0.07 19 20.29 0.32 179 2.11 0.08 19 1.70 0.10 15 7.80 0.24 69 1.15 0.11 10 2.73 0.20 24 Limnos island 1.96 0.08 17 1.62 0.10 14 1.60 0.10 14 19.30 0.54 170 Paros island 18.70 0.72 165 3.18 0.09 27 22.60 Santorini island 3.43 2.80
1.12
199
0.08 0.56
30 25
In literature are advanced different opinions concerning the correlation between antioxidant activity and the individual and group composition of the wines. Some authors consider that the activity correlates with flavanols amount, others with that of anthocyanins and other groups or individual compounds. Most often is assumed that the antioxidant activity of the wines correlates with their TP content7,15,24,25. We have established a good correlation between the total phenols content in the investigated wines and the antiradical (DPPH) activity from the one hand (R2= 0.9525), and from the other, between the total phenols and ferric reducing power (FRAP) (R2 = 0.9133). This is not valid in the case of the red wine Paros (No 13) with lower TP amount, but higher antiradical activity than the Egeopelagitikos wine (No 150) with higher TP 172
content and lower antiradical activity. This is attributed to the higher efficiency (ЕС50 = 16 µmol) of the phenol constituents of the Paros wine in comparison with that of the TP efficiency of the Egeopelagitikos wine – ЕС50= 32 µmol (р< 0.01). In the different countries are recommended different amounts of polyphenols which has to be included in the daily food for humans. The antiradical activity (in vitro) of the investigated wines expressed as vitamin E (Trolox) equivalent is ranging from 29 to 576 mg/100 ml, and the antioxidant activity presented as vitamin C (EAA) equivalent, is in the range of 10–199 mg/100 ml wine. The results of the study demonstrate that the investigated red and wine wines are rich in TP with antioxidant activity and the amount of 50 to 250 ml covers the daily dietary intake of polyphenols5. CONCLUSIONS The investigated red, white and rose wines from the Rodos, Limnos, Paros and Santorini islands divided in 4 quality grade – W.A.O.H.Q., W,A.O., table wine and traditional, are produced from typical for the islands local grape varieties such as Monemvasia, Mandilaria, Athiri, Assyrtiko, Aidani and Limniо and a selected imported grape varieties. The wines are characterised by a moderate TP content in comparison with the other wines on the basis of imported grape varieties and normal in vitro antiradical (DPPH) and antioxidant (FRAP) activity. The red and white wines from the Paros island and the white wines from the Santorini island are characterised by a higher TP content and higher antiradical and antioxidant activity, which is due to the higher efficiency of the phenol compounds (ЕС50). Amounts of 50 to 300 ml of the investigated wines cover the need of the human organism of daily dietary intake of polyphenols with antioxidant action. REFERENCES 1. J. B. Harborne: The Flavonoids (Ed. J. B. Harborne). Chapman and Hall, London, 1988. 2. J. J. Machiex, A. Flurient, J. Billot: Fruit Phenolics. CRC Press, Boca Raton, FL,1990. 3. F. Shahidi: Natural Antioxidants: Chemistry, Health Effects and Applications (Ed. F. Shahidi). AOCS Press, Champaign Illinois, 1997, 1–11. 4. J. Pokorny, N. Yanashlieva, M. Gordon: Antioxidants in Food. CSC Press, England, 2001. 5. L. BRAVO: Polyphenols: Chemistry, Dietary Sources, Metabolism and Nutritional Significance. Nutrition Reviews, 56 (11), 317 (1998). 6. J. Kanner, E. Frankel, R. Granit, B. German, J. E. Kinsella: Natural Antioxidants in Grapes and Wines. J. Agric. Food Chem., 42, 64 (1994). 7. J. BURNS, P. GARDNER, S. O’NEIL, S. CLOFORD, I. MORECROFT, D. B. McPHAIL, C. LISTER, D. MATTHEWS, M. R. McLEAN, M. E. S. LEAN, G. G. DUTHIE, A. CLOSIER: Relationship among Antioxidans Activity Vasodilation Capacity and Phenolic Content of Red Wines. J. Agric. Food Chem., 48, 220 (2000). 8. P. Simonetti, P. Pietta, G. Testolin: Polyphenol Content and Total Antioxidant Potential of Selected Italian Wines. J. Agric. Food Chem., 45, 1152 (1997).
173
9. N. Pellegrini, P. Simoneti, Cl. Gardana, O. Brenna, F. Brighenti, P. Pietta: Polyphenol Content and Total Antioxidant Activity of Vini Novelli (Young Red Wines). J. Agric. Food Chem., 48, 732 (2000). 10. J. A. Larrauri, C. Sanchez-Moreno, P. Ruperez, F. Saura-Calixto: Free Radical Scavenging Capacity in the Aging of Selected Red Spanish Wines. J. Agric. Food Chem., 47, 1603 (1999). 11. S. Renaud, M. de Lorgeril: Wine, Alcohol, Platelets and the French Paradox for Coronary Heart Disease. Lancet, 339, 1523 (1992). 12. M. Bourzeix, D. Weyland, N. Heredia: A Study of Catechins and Procyanidins of Grape Clusters, the Wine and Other By-products of the Wine. Bulletin de L’OIV, 59, 1171 (1987). 13. E. N. Frankel, A. L. Waterhouse, P. L. Teissedre: Principal Phenolic Phytochemicals in Selected California Wines and Their Antioxidant Activity in Inhibiting Oxidation of Human Lowdensity Lipoproteins. J. Agric. Food Chem., 43, 890 (1995). 14. C. A. RICE-EVANCE, N. J. MILLER: Antioxidants: the Case for Fruit and Vegetables in the Diet. British Food J., 97 (9), 35 (1995). 15. A. ARNOUS, D. P. MAKRIS, P. KEFALAS: Effect of Principal Polyphenolic Components in Relations to Antioxidant Characteristics of Aged Red Wines. J. Agric. Food Chem., 49 (12), 5736 (2001). 16. D. MAKRIS, E. PSARRA, S. KALLITHRAKA, P. KEFALAS: The Effect of Polyphenolic Compo sition as Related to Antioxidant Capacity in White Wines. Food Research Intern., 36, 805 (2000). 17. N. GUGULIAS: Phenol Content and Antioxidant Activity of Some Bulgarian and Greek Wines. J. Envir. Prot. Ecology, 6 (2), 392 (2005). 18. V. L. Singleton, S. A. Rossi: Colorimetry of Total Phenolics with Phosphomolibdic-phosphotungestic Acid Reagents. J. Enol. Viticult., 16, 144 (1965). 19. B. Badenscheider, D. Luthiria, L. A. Wasterhouse, P. Winterhalter: Antioxidants in White Wine (cv. Riesling). 1. Comparison of Different Testing Methods for Antioxidant Activity. Vitis, 38 (3), 127 (1999). 20. W. Brand-Williams, M. E. Cuvellier, C. Berset: Use of Free Radical Method to Evaluate Antioxidant Activity. Lebensm. Wiss. Technol., 28, 25 (1995). 21. I. F. F. Benzie, J. J. Strain: Ferric Reducing Antioxidant Power Assay. Methods in Enzymology, 299, 15 (1999). 22. J. A. Vinson, B. Hontz: Phenol Antioxidant Index: Comparative Antioxidant Effectiveness of Red and White Wines. J. Agric. Food Chem., 43, 401 (1995). 23. V. L. Singleton: Wine Phenols. In: Wine Analysis (Eds H. F. Liskens, S. F. Jackson). Springer Verlag, Berlin, 1988. 24. R. Meral: Antioxidant Effects of Wine Polyphenols. Trakia J. Sci., 6 (1), 57 (2008). 25 N. Gougoulias, N. Mashev: Evaluation of Polyphenols Antioxidant Activity of Grapes Seeds (V. vinifera). Oxid. Commun., 31 (1), 88 (2008). Received 9 June 2009 Revised 28 July 2009
174
