MIHAELA BERINDE 1,2 , MAURIZIO RUZZI 2 , ROSARIO MULEO 2 , MARIA TOFANA 1* , SONIA SOCACI 1
1
MIHAELA BERINDE 1,2 , MAURIZIO RUZZI 2 , ROSARIO MULEO 2 , MARIA TOFANA 1* , SONIA SOCACI 1 1-Universitaty of Agricultural Science and Veterinary Medicine, Cluj-Napoca 2 - University of Tuscia, Viterbo, Italy * Corresponding author: tofanam @ yahoo.com Purpose: To determine the polyphenols composition of apple (fruit pulp and leaf) from two varieties: Italian Red Passion and Annurca; Preliminary evaluation of the antimicrobial activity of this apple varieties POLYPHENOLS CONTENT IN APPLE ITALIAN RED PASSION AND ANNURCA Introduction Abstract • Apples of seedlings, M3, M4 and M5 from the experimental field of the University of Tuscia Italy, belonging to the Italian Red Passion group are investigated about the polyphenols content. These fruits are characterized by red coloration of the pulp. In this work we analyzed comparatively the polyphenols content of apples M3 and M4 and leaves of apples M4 and M5. As control are used Annurca apples and Annurca leaves. Materials and Methods Results Literature • The assortment of polyphenols present in apple fruit is wide, but differs between flesh and peel both in its range and in the total content (Wolfe et al., 2003). Typical apple fruit phenolic acids are hydroxy-cinnamic acids, dihydro-chalcones, flavonols, anthocyanin, flavanols and procyanidin (Tsao et al., 2003). The content changes of the main phenols in apples and their contributions to the total antioxidant activity are not very clear. •Numerous epidemiological studies, published toward the end of the twentieth century, have provided evidence that increased consumption of fruit and vegetables provides protection against cardiovascular diseases, various kinds of cancer, and other chronic diseases. The risk of a heart attack has been shown to decrease by 49% for apple consumers in diet at a daily amount of >110 g compared to those at <18 g. •In fruits such as apple, red coloration of the skin is a much valued trait. In the last 20 years the interest on the polyphenolic content of the fruit is continuously growing up, due to their health-related characteristics (Boyer and Liu, 2004); from this point of view, red fleshed apple could be a varietal innovation with favorable health-related characteristics and a strong appeal to consumers. •The accessions were obtained from cross pollinated by a plant of red apple from the wild germplasm Lazio in the experimental field at the farm of the University of Tuscia. •In this work we used only apples M3 and M4 and leaves of apples M4 and M5. As control was used Annurca apples and Annurca leaves. Extract preparation: Extract preparation: Lyophilized material Solvent extraction Methanol/Water/Acetic acid (70:29:1) Acetone/Water/Acetic acid (70:29:1) Methanol/Acetone/Water/Acetic acid (60:20:19:1) Dried samples were resuspended in pure MetOH Wolfe K, Liu RH. (2003), “ Apple peels as a value-added food ingredient” . J Agric Food Chem. 2003, 51:1676–1683. Tsao Rong, Yang Raymond, Young J. Christopher and Zhu Honghui, (2003), ”Polyphenolic Profiles in Eight Apple Cultivars Using High Performance Liquid Chromatography (HPLC)” , J. Agric. Food Chem., 51: 6347-6353. Alberto María Rosa, Matías Andrés, CanavosioRinsdahl, Manca de NadraMaría Cristina,(2006), “Antimicrobial effect of polyphenols from apple skins on human bacterial pathogens”, Electronic Journal of Biotechnology ISSN: 0717-3458 Vol.9 No.3, Special Issue, by Pontificia Universidad Católica de Valparaíso – Chile. Boyer J.; Liu, R.H,(2004), “Apple phytochemicals and their health benefits”, Nutrition Journal, 3: 5. Kondo Satoru, Tsuda Kazuhiko, Muto Norio, Ueda Jin-etsu, (2002) “Antioxidative activity of apples skin or flesh extracts associated whit fruit development on selected apple cultivars”, ScientiaHorticulturae, 96: 177-185. Mari Angela, Tedesco Idolo, Nappo Annunziata, Russo Gian Luigi, Malorni Antonio, Carbone Virginia, (2010) “Phenolic compound characterization and anti-proliferative activity of ‘‘Annurca” apple, a southern Italian cultivar”, Food Chemistry, 123: 157–164. D’AngeloStefania, Cimmino Amelia, Raimo Marianna, Salvatore Anna, Zappia Vincenzo , and GallettiPatrizia, (2007) “Effect of Reddening–Ripening on the Antioxidant Activity of Polyphenol Extracts from Cv. ‘Annurca’ Apple Fruits”,J. Agric. Food Chem, 55:9977–9985. HPLC Analysis was performed using a HPLC-DAD apparatus at 5 wavelengths (280, 307, 316, 365 and 520 nm), following the method used by Jeffrey et al. (1999) Bacterial antimicrobial susceptibility plate test: modified Kirby- Bauer Method Bacterial antimicrobial susceptibility liquid test: Broth Dilution Method Growth inhibition obtained using acetone extracts from fruit pulp of different Italian Red Passion genotypes (M3, M4) and Annurca SOLVENT SAMPLE INHIBITION ZONE DIAMETER (cm) 24h 48h E. coli DH 5α M3 0.325±0.00125 0.300±0.0 M4 0.600±0.0 0.600±0.0 0.575±0.00125 0.575±0.00125 Annurca 0.450±0.005 0.450±0.0 B.subtilis PB19 M3 0.275±0.00125 0.250±0.0 M4 0.550±0.005 0.550±0.005 0.525±0.00125 0.525±0.00125 Annurca 0.375±0.0 0.350±0.00125 P.aeruginosa M3 0.225±0.00125 0.225±0.00125 M4 0.400±0.0 0.400±0.0 Annurca 0.400±0.0 0.400±0.0 Control(methanol) 0.0 0.0 Higher antimicrobial activity was obtained using fruit pulp extracts of M4 genotype, independently from the solvent system SOLVENT SAMPLE GROWTH INHIBITION (%) 25 µl 50 µl Acetone M3 none 25.20±0.0017 2 M4 19.52±0.0053 67.87±0.0004 Annurca none 68.40 Methanol M3 none 25.02±0.0153 3 M4 35.00±0.0156 53.32±0.0013 Annurca 25.65±0.01346 59.18±0.0001 9 Mix M3 none 21.66±0.0027 4 M4 35.55±0.0008 60.25±0.0016 61.84±0.0004 Functional compounds in apple: organic acids, polyphenols, flavonoids 6 th International Conference on Polyphenols Applications June 7 & 8, 2012 – Paris, France •The total polyphenols content from all this samples were extracted with 3 types of solvents: Methanol/Water/Acetic Acid:70:29:1, Acetone/Water/Acetic Acid:70:29:1 and Methanol/ Acetone/Water /Acetic Acid: 60:20:19:1. The obtained extract was evaporated, lyophilized and resuspended in methanol and then analyzed by HPLC. The HPLC analysis was performed, simultaneously, at 5 wavelengths (280, 307, 316, 365 and 520 nm). •Comparative analysis of HPLC data confirmed that Italian Red Passion extracts from both fruit and leaf samples contained a larger array of compounds compared to Annurca samples. This compounds, belong to different chemical groups, such as anthocyanin, hydroxycinnamic acids, flavonols, procyanidins and dihydrochalcones. The better solvent systems for the chemical analysis of fruit pulp and leaf samples are acetone and methanol respectively. ANTIMICROBIAL ACTIVITY OF EXTRACTS ANTIMICROBIAL ACTIVITY OF EXTRACTS FROM APPLE ITALIAN RED PASSION AND ANNURCA FROM APPLE ITALIAN RED PASSION AND ANNURCA HPLC chromatogram of anthocyanins (A520) Compounds identified Graphical representation of HPLC Graphical representation of HPLC results results from methanol extracts of from methanol extracts of M4 fruit pulp M4 fruit pulp
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6 th International Conference on Polyphenols Applications June 7 & 8, 2012 – Paris, France. POLYPHENOLS CONTENT IN APPLE ITALIAN RED PASSION AND ANNURCA. MIHAELA BERINDE 1,2 , MAURIZIO RUZZI 2 , ROSARIO MULEO 2 , MARIA TOFANA 1* , SONIA SOCACI 1. - PowerPoint PPT Presentation
Transcript of MIHAELA BERINDE 1,2 , MAURIZIO RUZZI 2 , ROSARIO MULEO 2 , MARIA TOFANA 1* , SONIA SOCACI 1
MIHAELA BERINDE1,2, MAURIZIO RUZZI2, ROSARIO MULEO2, MARIA TOFANA1*, SONIA SOCACI1
1-Universitaty of Agricultural Science and Veterinary Medicine, Cluj-Napoca2 - University of Tuscia, Viterbo, Italy
* Corresponding author: [email protected]
Purpose: To determine the polyphenols composition of apple (fruit pulp and leaf) from two varieties: Italian Red Passion and Annurca; Preliminary evaluation of the antimicrobial activity of this apple varieties
POLYPHENOLS CONTENT IN APPLE ITALIAN RED PASSION AND ANNURCA
Introduction
Abstract
• Apples of seedlings, M3, M4 and M5 from the experimental field of the University of Tuscia Italy, belonging to the Italian Red Passion group are investigated about the polyphenols content. These fruits are characterized by red coloration of the pulp. In this work we analyzed comparatively the polyphenols content of apples M3 and M4 and leaves of apples M4 and M5. As control are used Annurca apples and Annurca leaves.
Materials and Methods
Results
Literature
• The assortment of polyphenols present in apple fruit is wide, but differs between flesh and peel both in its range and in the total content (Wolfe et al., 2003). Typical apple fruit phenolic acids are hydroxy-cinnamic acids, dihydro-chalcones, flavonols, anthocyanin, flavanols and procyanidin (Tsao et al., 2003). The content changes of the main phenols in apples and their contributions to the total antioxidant activity are not very clear. •Numerous epidemiological studies, published toward the end of the twentieth century, have provided evidence that increased consumption of fruit and vegetables provides protection against cardiovascular diseases, various kinds of cancer, and other chronic diseases. The risk of a heart attack has been shown to decrease by 49% for apple consumers in diet at a daily amount of >110 g compared to those at <18 g. •In fruits such as apple, red coloration of the skin is a much valued trait. In the last 20 years the interest on the polyphenolic content of the fruit is continuously growing up, due to their health-related characteristics (Boyer and Liu, 2004); from this point of view, red fleshed apple could be a varietal innovation with favorable health-related characteristics and a strong appeal to consumers.
•The accessions were obtained from cross pollinated by a plant of red apple from the wild germplasm Lazio in the experimental field at the farm of the University of Tuscia.
•In this work we used only apples M3 and M4 and leaves of apples M4 and M5. As control was used Annurca apples and Annurca leaves.
Extract preparation:Extract preparation: Lyophilized material
Solvent extraction Methanol/Water/Acetic acid (70:29:1)Acetone/Water/Acetic acid (70:29:1)Methanol/Acetone/Water/Acetic acid (60:20:19:1) Dried samples were resuspended in pure MetOH
Wolfe K, Liu RH. (2003), “ Apple peels as a value-added food ingredient”. J Agric Food Chem. 2003, 51:1676–1683. Tsao Rong, Yang Raymond, Young J. Christopher and Zhu Honghui, (2003), ”Polyphenolic Profiles in Eight Apple Cultivars Using High
Performance Liquid Chromatography (HPLC)”, J. Agric. Food Chem., 51: 6347-6353. Alberto María Rosa, Matías Andrés, CanavosioRinsdahl, Manca de NadraMaría Cristina,(2006), “Antimicrobial effect of polyphenols from apple
skins on human bacterial pathogens”, Electronic Journal of Biotechnology ISSN: 0717-3458 Vol.9 No.3, Special Issue, by Pontificia Universidad Católica de Valparaíso – Chile.
Boyer J.; Liu, R.H,(2004), “Apple phytochemicals and their health benefits”, Nutrition Journal, 3: 5.Kondo Satoru, Tsuda Kazuhiko, Muto Norio, Ueda Jin-etsu, (2002) “Antioxidative activity of apples skin or flesh extracts associated whit fruit
development on selected apple cultivars”,ScientiaHorticulturae, 96: 177-185.Mari Angela, Tedesco Idolo, Nappo Annunziata, Russo Gian Luigi, Malorni Antonio, Carbone Virginia, (2010) “Phenolic compound
characterization and anti-proliferative activity of ‘‘Annurca” apple, a southern Italian cultivar”, Food Chemistry, 123: 157–164.D’AngeloStefania, Cimmino Amelia, Raimo Marianna, Salvatore Anna, Zappia Vincenzo , and GallettiPatrizia, (2007) “Effect of Reddening–
Ripening on the Antioxidant Activity of Polyphenol Extracts from Cv. ‘Annurca’ Apple Fruits”,J. Agric. Food Chem, 55:9977–9985.
HPLC Analysis was performed using a HPLC-DAD apparatus at 5 wavelengths (280, 307, 316, 365 and 520 nm), following the method used by Jeffrey et al. (1999)
Bacterial antimicrobial susceptibility plate test: modified Kirby-Bauer Method
Bacterial antimicrobial susceptibility liquid test: Broth Dilution Method
Growth inhibition obtained using acetone extracts from fruit pulp of different Italian Red Passion genotypes (M3, M4) and Annurca
SOLVENT SAMPLE
INHIBITION ZONE DIAMETER
(cm)
24h 48h
E. coli DH 5α
M3 0.325±0.00125 0.300±0.0
M4 0.600±0.0 0.600±0.0 0.575±0.00125 0.575±0.00125
Annurca 0.450±0.005 0.450±0.0
B.subtilis PB19
M3 0.275±0.00125 0.250±0.0
M4 0.550±0.005 0.550±0.005 0.525±0.00125 0.525±0.00125
Annurca 0.375±0.0 0.350±0.00125
P.aeruginosa
M3 0.225±0.00125 0.225±0.00125
M4 0.400±0.0 0.400±0.0
Annurca 0.400±0.0 0.400±0.0
Control(methanol) 0.0 0.0
Higher antimicrobial activity was obtained using fruit pulp extracts of M4 genotype, independently from the solvent system
SOLVENT SAMPLE
GROWTH INHIBITION
(%)
25 µl 50 µl
Acetone
M3 none 25.20±0.00172
M4 19.52±0.0053 67.87±0.0004
Annurca none 68.40
Methanol
M3 none 25.02±0.01533
M4 35.00±0.0156 53.32±0.0013
Annurca 25.65±0.01346 59.18±0.00019
Mix
M3 none 21.66±0.00274
M4 35.55±0.0008 60.25±0.0016
Annurca 33.90±0.00273 61.84±0.00047
Functional compounds in apple: organic acids, polyphenols, flavonoids
6th International Conference on Polyphenols Applications June 7 & 8, 2012 – Paris, France
•The total polyphenols content from all this samples were extracted with 3 types of solvents: Methanol/Water/Acetic Acid:70:29:1, Acetone/Water/Acetic Acid:70:29:1 and Methanol/ Acetone/Water /Acetic Acid: 60:20:19:1. The obtained extract was evaporated, lyophilized and resuspended in methanol and then analyzed by HPLC. The HPLC analysis was performed, simultaneously, at 5 wavelengths (280, 307, 316, 365 and 520 nm). •Comparative analysis of HPLC data confirmed that Italian Red Passion extracts from both fruit and leaf samples contained a larger array of compounds compared to Annurca samples. This compounds, belong to different chemical groups, such as anthocyanin, hydroxycinnamic acids, flavonols, procyanidins and dihydrochalcones. The better solvent systems for the chemical analysis of fruit pulp and leaf samples are acetone and methanol respectively.
ANTIMICROBIAL ACTIVITY OF EXTRACTS ANTIMICROBIAL ACTIVITY OF EXTRACTS FROM APPLE ITALIAN RED PASSION AND FROM APPLE ITALIAN RED PASSION AND ANNURCAANNURCA
HPLC chromatogram of anthocyanins (A520)
Compounds identifiedGraphical representation of HPLC Graphical representation of HPLC results results from methanol extracts of from methanol extracts of M4 fruit pulpM4 fruit pulp
