**Table 2.**

 *Selective studies on bioactivities and potential health benefits of main virgin olive oil phenolic compounds.*

#### *Virgin Olive Oil Phenolic Compounds: Insights on Their Occurrence, Health-Promoting… DOI: http://dx.doi.org/10.5772/intechopen.98581*

which is similar to that consumed daily in a typical Mediterranean diet [110]. These simple phenols were largely identified in both urine and plasma mainly in glucuronides and sulphate conjugates, while their free forms were not detected in plasma samples. In fact, aglycones and glycosides forms of tyrosol and hydroxytyrosol undergo a prompt hydrolysis phenomenon under gastric conditions together with a substantial rise in the contents of tyrosol and hydroxytyrosol free forms penetrating the small intestine [108]. The intestinal transport of hydroxytyrosol occurs through a bidirectional passive diffusion mechanism as demonstrated in an *in vitro* study conducted by Manna and co-workers [111]. While crossing epithelial cells of the gastrointestinal tract, hydroxytyrosol is usually transformed through enzymatic reactions into homovanillyl alcohol and its glucuronide forms [108].

Regarding secoiridoids, they remain highly stable in the mouth but suffer significant losses in the gastric, duodenal, and colonic regions, with a recovery rate at the duodenal level ranging between 7% and 34%. Glycosylation and cleavage of glycosidic linkages take part in the secoiridoids absorption, and it is thought that some of them, such as oleacein, are absorbed in the small intestine by passive diffusion through the membrane of intestinal cells [89]. In the case of oleuropein, the mechanism of absorption is still confusing and remains unclear. However, several studies showed that it exerts its biological effects *via* its conversion into hydroxytyrosol [112]. This was mainly explained by the fact that the content of oleuropein was in the mass range of few nanograms in plasma while hydroxytyrosol was detected in high concentrations after the intake of great doses of oleuropein in both rats and humans trials [113, 114]. When considering other secoiridoids, Vissers et al. [114] could not analyse oleuropein glycoside, oleuropein- or ligstroside-aglycons in urinary excretions which supports the idea that they may be hydrolysed into hydroxytyrosol and tyrosol and extensively metabolised once absorbed from the small intestine.

#### **4. Concluding remarks**

In the Mediterranean area, the largest virgin olive oil producing region in the world, new agronomic practices and processing technologies are steadily developed and adopted over the past decade. Thereby, virgin olive oil production has achieved outstanding performance both in terms of increasing oil yield and quality. Nevertheless, the future of this sector in a context of globalisation, and the consequent changes in lifestyle and consumers adherence to the Mediterranean diet, has become an important subject of attention for the producers and governmental bodies alike. For instance, in the search for new opportunities to boost virgin olive oil consumption, growing importance is attributed to promote its health value and organoleptic features among consumers, paying special attention to those compounds responsible of these characteristics. In developing such strategies, a central role should clearly be reserved for the phenolic fraction of this product if considering its health-promoting proprieties and its contribution to the oxidative stability and sensory quality of virgin olive oil. Undoubtedly, the recent agro-technological advancements supported by the scientific data available up till now offer promising tools to produce virgin olive oils rich on phenolics; however, the bitter taste of such oils is often not appreciated by consumers. For this reason, what still remains a challenge in this sector is the development of holistic agro-technological approaches to produce virgin olive oils with phenolics content that comply not only with institutional regulations (EFSA health claim for example) but also with consumer preferences. Furthermore, the lack of an official method of determining these compounds in compliance with health claim requirements is an obstacle to be overcome in a not-toodistant future if we want to avoid a lack of credibility of these health claims.

*Virgin Olive Oil Phenolic Compounds: Insights on Their Occurrence, Health-Promoting… DOI: http://dx.doi.org/10.5772/intechopen.98581*

#### **Acknowledgements**

Authors are grateful to the Erasmus+ Programme of the European Union, FoSaMed project (618518-EPP-T-2020-1-PT-EPPKA2-CBHE-JP) for Salah Chaji's predoctoral fellowship; and to the National Centre for Scientific and Technical Research (CNRST Morocco) for Sanae Moussafir's predoctoral fellowship (Research Excellence Fellowships Program).

### **Author details**

El Amine Ajal1 , Salah Chaji2 , Sanae Moussafir2 , Rachid Nejjari1 , Abdelmajid Soulaymani3 and Aadil Bajoub2,3\*

1 UPR of Pharmacognosy, Faculty of Medicine and Pharmacy of Rabat, Mohammed V University, Rabat, Morocco

2 Laboratory of Food and Food By-Products Chemistry and Processing Technology, National School of Agriculture in Meknès, Meknès, Morocco

3 Laboratory of Health and Biology, Faculty of Sciences, University Ibn Tofail, Kénitra, Morocco

\*Address all correspondence to: aliam80@hotmail.com

© 2021 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

#### **References**

[1] Farràs M, Martinez-Gili L, Portune K, Arranz S, Frost G, Tondo M, et al. Modulation of the gut microbiota by olive oil phenolic compounds: Implications for lipid metabolism, immune system, and obesity. Nutrients. 2020;12(8):2200. DOI: 10.3390/ nu12082200

[2] Fitó M, Cladellas M, de la Torre R, Martí J, Muñoz D, Schröder H, et al. Anti-inflammatory effect of virgin olive oil in stable coronary disease patients: A randomized, crossover, controlled trial. Eur J Clin Nutr. 2008;62(4):570-574.

[3] Buckland G, Travier N, Barricarte A, Ardanaz E, Moreno-Iribas C, Sánchez MJ, et al. Olive oil intake and CHD in the European Prospective Investigation into Cancer and Nutrition Spanish cohort. Br J Nutr. 2012;108(11):2075-2082. DOI: 10.1017/ S000711451200298X

[4] Guasch-Ferré M, Hu FB, Martínez-González MA, Fitó M, Bulló M, Estruch R, et al. Olive oil intake and risk of cardiovascular disease and mortality in the PREDIMED Study. BMC Med. 2014;12(1):78. DOI: 10.1186/1741-7015-12-78

[5] Covas MI, Ruiz-Gutiérrez V, De La Torre R, Kafatos A, Lamuela-Raventós RM, Osada J, et al. Minor components of olive oil: Evidence to date of health benefits in humans. Nutr Rev. 2006;64(10):S20–S30.

[6] Preedy VR, Watson RR, editors. Olives and olive oil in health and disease prevention. 2nd edition. Elsevier; 2021.

[7] Boskou D, editor. Olive and olive oil bioactive constituents. 1st edition. American Oil Chemists' Society Press; 2015.

[8] Jimenez-Lopez C, Carpena M, Lourenço-Lopes C, Gallardo-Gomez M, Lorenzo JM, Barba FJ, et al. Bioactive compounds and quality of extra virgin olive oil. Foods. 2020;9(8):1014. DOI: 10.3390/foods9081014

[9] Rodríguez-López P, Lozano-Sanchez J, Borrás-Linares I, Emanuelli T, Menéndez JA, Segura-Carretero A. Structure – biological activity relationships of extra-virgin olive oil phenolic compounds: Health properties and bioavailability. Antioxidants. 2020;9(8):685. DOI: 10.3390/ antiox9080685

[10] Servili M, Sordini B, Esposto S, Urbani S, Veneziani G, Maio I Di, et al. Biological activities of phenolic compounds of extra virgin olive oil. Antioxidants. 2014;3(1):1-23. DOI: 10.3390/antiox3010001

[11] De La Torre R. Bioavailability of olive oil phenolic compounds in humans. Inflammopharmacology. 2008;16(5):245-247.

[12] De La Torre R, Fitó M, Covas M-I. The bioavailability of olive oil phenolic compounds and their bioactive effects in humans. In: Preedy VR, Watson RR, editors. Olives and Olive Oil in Health and Disease Prevention (second edition). Elsevier; 2021. p.193-203.

[13] Uceda M. Composición y calidad de los aceites de oliva vírgenes extra. Influencia de la variedad. In: Hernández J.V. et al., editors. Algunas contribuciones sobre olivicultura y Elaiotecnia desde la perspectiva de la experiencia. GEA Westfalia Separator Ibérica, S.A. 2009.

[14] Rodis PS, Karathanos VT, Mantzavinou A. Partitioning of olive oil antioxidants between oil and water phases. J Agric Food Chem. 2002;50:596-601.

*Virgin Olive Oil Phenolic Compounds: Insights on Their Occurrence, Health-Promoting… DOI: http://dx.doi.org/10.5772/intechopen.98581*

[15] Emma MR, Augello G, Di Stefano V, Azzolina A, Giannitrapani L, Montalto G, et al. Potential uses of olive oil secoiridoids for the prevention and treatment of cancer: A narrative review of preclinical studies. Int J Mol Sci. 2021;22(3):1234.

[16] Termentzi A, Halabalaki M, Skaltsounis LA. From drupes to olive oil: An exploration of olive key Metabolites. In: Boskou D, editor. Olives and olive oil Bioactive Constituents. American Oil Chemists' Society Press; 2015. p.147-170.

[17] Fregapane G, Salvador MD. Production of superior quality extra virgin olive oil modulating the content and profile of its minor components. Food Res Int. 2013;54(2):1907-1914. DOI: 10.1016/j.foodres.2013.04.022

[18] Jerman Klen T, Golc Wondra A, Vrhovšek U, Sivilotti P, Vodopivec BM. Olive fruit phenols transfer, transformation, and partition trail during laboratory-scale olive oil processing. J Agric Food Chem. 2015;63(18):4570-4579.

[19] Velázquez-palmero D, Romerosegura C, García-rodríguez R, Hernández ML, Vaistij FE, Graham IA, et al. An oleuropein β-Glucosidase from olive fruit is involved in determining the phenolic composition of virgin olive oil. Front Plant Sci. 2017;8(November): 1902. DOI: 10.3389/fpls.2017.01902

[20] Artajo LS, Romero MP, Suarez M, Motilva MJ. Partition of phenolic compounds during the virgin olive oil industrial extraction process. Eur Food Res Technol. 2007;225:617-625.

[21] Tsimidou M. Polyphenols and quality of virgin olive oil in retrospect [*Olea europaea* L.]. Ital J Food Sci. 1998;10.

[22] Bajoub A, Medina-Rodríguez S, Olmo-García L, Ajal EA, Monasterio RP, Hanine H, et al. In-depth two-year study of phenolic profile variability among olive oils from autochthonous and mediterranean varieties in Morocco, as revealed by a LC-MS chemometric profiling approach. Int J Mol Sci. 2017;18(1):52. DOI: 10.3390/ ijms18010052

[23] Miho H, Díez CM, Mena-bravo A, Medina VS De, Moral J, Melliou E, et al. Cultivar influence on variability in olive oil phenolic profiles determined through an extensive germplasm survey. Food Chem. 2018;266:192-199. DOI: 10.1016/j. foodchem.2018.06.002

[24] Di Lecce G, Piochi M, Pacetti D, Frega NG, Bartolucci E, Scortichini S, et al. Eleven monovarietal extra virgin olive oils from olives grown and processed under the same conditions: Effect of the cultivar on the chemical composition and sensory traits. Foods. 2020;9(7):904. DOI: 10.3390/ foods9070904

[25] Martín-Peláez S, Covas MI, Fitó M, Kušar A, Pravst I. Health effects of olive oil polyphenols : Recent advances and possibilities for the use of health claims. Mol Nutr Food Res. 2013;57:760-771. DOI: 10.1002/mnfr.201200421

[26] Lockyer S, Rowland I. Authorised EU health claims for polyphenols in olive oil. In: Sadler MJ, editor. Foods, Nutrients and Food Ingredients with Authorised EU Health Claims. Woodhead Publishing; 2014. p.212-228.

[27] Pinto J, Paiva-Martins F, Corona G, Debnam ES, Jose Oruna-Concha M, Vauzour D, et al. Absorption and metabolism of olive oil secoiridoids in the small intestine. Br J Nutr. 2011;105(11):1607-1618. DOI: 10.1017/ S000711451000526X

[28] Serreli G, Deiana M. Biological relevance of extra virgin olive oil polyphenols metabolites. Antioxidants. 2018;7(12):170. DOI: 10.3390/ antiox7120170

[29] Bendini A, Cerretani L, Carrasco-Pancorbo A, Gómez-Caravaca AM, Segura-Carretero A, et al. Phenolic molecules in virgin olive oils: A survey of their sensory properties, health effects, antioxidant activity and analytical methods. An overview of the last decade. Molecules. 2007;12(8): 1679-1719.

[30] Owen RW, Mier W, Giacosa A, Hull WE, Spiegelhalder B, Bartsch H. Phenolic compounds and squalene in olive oils: The concentration and antioxidant potential of total phenols, simple phenols, secoiridoids, lignansand squalene. Food Chem Toxicol. 2000;38:647-659.

[31] Bianco A, Coccioli F, Guiso M, Marra C. The occurrence in olive oil of a new class of phenolic compounds: Hydroxy-isochromans. Food Chem. 2002;77(4):405-411. DOI: 10.1016/ S0308-8146(01)00366-1

[32] Taticchi A, Esposto S, Servili M. The Basis of the Sensory Properties of Virgin Olive Oil. In: Monteleone E, Langstaff S, editors. Olive Oil Sensory Science. John Wiley & Sons; 2014. p.33-54. DOI: 10.1002/9781118332511.ch2

[33] Miho H, Moral J, Barranco D, Ledesma-Escobar CA, Priego-Capote F, Díez CM. Influence of genetic and interannual factors on the phenolic profiles of virgin olive oils. Food Chem. 2021;342:128357. DOI: 10.1016/j. foodchem.2020.128357

[34] Pedan V, Popp M, Rohn S, Nyfeler M, Bongartz A. Characterization of phenolic compounds and their contribution to sensory properties of olive oil. Molecules. 2019;24(11):2041. DOI: 10.3390/molecules24112041

[35] Beltrán G, Ruano MT, Jiménez A, Uceda M, Aguilera MP. Evaluation of virgin olive oil bitterness by total phenol content analysis. Eur J Lipid Sci Technol. 2007;109(3):193-197. DOI: 10.1002/ ejlt.200600231

[36] Borges TH, Cabrera-Vique C, Seiquer I. Antioxidant properties of chemical extracts and bioaccessible fractions obtained from six Spanish monovarietal extra virgin olive oils: Assays in Caco-2 cells. Food Funct. 2015;6(7):2375-2383.

[37] Franco MN, Galeano-díaz T, Sánchez J, Miguel C De, Martínvertedor D. Total phenolic compounds and tocopherols profiles of seven olive oil varieties grown in the south-west of Spain. J Oleo Sci. 2014;63(2):115-125.

[38] Borges TH, Serna A, López LC, Lara L, Nieto R, Seiquer I. Composition and antioxidant properties of Spanish extra virgin olive oil regarding cultivar, harvest year and crop stage. Antioxidants. 2019;8(7):217. DOI: 10.3390/antiox8070217

[39] Borges TH, Pereira JA, Cabrera-Vique C, Seiquer I. Study of the antioxidant potential of Arbequina extra virgin olive oils from Brazil and Spain applying combined models of simulated digestion and cell culture markers. J Funct Foods. 2017;37:209- 218. DOI: 10.1016/j.jff.2017.07.059

[40] Sicari V, Leporini M, Giuffré AM, Aiello F, Falco T, Pagliuso MT, et al. Quality parameters, chemical compositions and antioxidant activities of Calabrian (Italy) monovarietal extra virgin olive oils from autochthonous (Ottobratica) and allochthonous (Coratina, Leccino, and Nocellara Del Belice) varieties. J Food Meas Charact. 2021;15(1):363-375. DOI: 10.1007/ s11694-020-00640-9

[41] Bayram B, Esatbeyoglu T, Schulze N, Ozcelik B, Frank J, Rimbach G. Comprehensive analysis of polyphenols in 55 extra virgin olive oils by HPLC-ECD and their correlation

*Virgin Olive Oil Phenolic Compounds: Insights on Their Occurrence, Health-Promoting… DOI: http://dx.doi.org/10.5772/intechopen.98581*

with antioxidant activities. Plant Foods Hum Nutr. 2012;67(4):326-336. DOI: 10.1007/s11130-012-0315-z

[42] Loizzo MR, Di Lecce G, Boselli E, Menichini F, Frega NG. Radical scavenging, total antioxidant capacity, and antiproliferative activity of phenolic extracts from extra virgin olive oil by Cultivar "Frantoio." Int J Food Prop. 2012;15(6):1345-1357. DOI: 10.1080/10942912.2010.522291

[43] Leporini M, Loizzo MR, Tenuta MC, Falco T, Sicari V, Pellicanò TM, et al. Calabrian extra-virgin olive oil from Frantoio cultivar: Chemical composition and health properties. Emirates J Food Agric. 2018;30(7):631-637. DOI: 10.9755/ ejfa.2018.v30.i7.1743

[44] Kalogeropoulos N, Kaliora AC, Artemiou A, Giogios I. Composition, volatile profiles and functional properties of virgin olive oils produced by two-phase vs three-phase centrifugal decanters. LWT - Food Sci Technol. 2014;58(1):272-9. DOI: 10.1016/j. lwt.2014.02.052

[45] Bouaziz M, Jemai H, Khabou W, Sayadi S. Oil content, phenolic profiling and antioxidant potential of Tunisian olive drupes. J Sci Food Agric. 2010;90(10):1750-1758. DOI: 10.1002/ jsfa.4013

[46] Nakbi A, Issaoui M, Dabbou S, Koubaa N, Echbili A, Hammami M, et al. Evaluation of antioxidant activities of phenolic compounds from two extra virgin olive oils. J Food Compos Anal. 2010;23:711-5. DOI: 10.1016/j. jfca.2010.05.003

[47] Gouvinhas I, Machado J, Gomes S, Lopes J, Martins-Lopes P, Barros AIRNA. Phenolic composition and antioxidant activity of monovarietal and commercial portuguese olive oils. J Am Oil Chem Soc. 2014;91(7): 1197-1203.

[48] Kelebek H, Kesen S, Selli S. Comparative study of bioactive constituents in Turkish olive oils by LC-ESI/MS/MS. Int J Food Prop. 2015;18(10):2231-2245. DOI: 10.1080/10942912.2014.968788

[49] Köseoǧlu O, Sevim D, Kadiroǧlu P. Quality characteristics and antioxidant properties of Turkish monovarietal olive oils regarding stages of olive ripening. Food Chem. 2016;212:628-634. DOI: 10.1016/j.foodchem.2016.06.027

[50] Bajoub A, Carrasco-Pancorbo A, Ajal EA, Beltrán Maza G, Fernández-Gutiérrez A, Ouazzani N. Contribution to the establishment of a protected designation of origin for Meknès virgin olive oil: A 4-years study of its typicality. Food Res Int. 2014;66:332-343. DOI: 10.1016/j. foodres.2014.09.021

[51] El Qarnifa S, El Antari A, Hafidi A. Effect of maturity and environmental conditions on chemical composition of olive oils of introduced cultivars in Morocco. J Food Qual. 2019;2019. DOI: 10.1155/2019/1854539

[52] Olmo-García L, Polari JJ, Li X, Bajoub A, Fernández-Gutiérrez A, Wang SC, et al. Study of the minor fraction of virgin olive oil by a multiclass GC–MS approach: Comprehensive quantitative characterization and varietal discrimination potential. Food Res Int. 2019;125:108649. DOI: 10.1016/j. foodres.2019.108649

[53] Bajoub A, Pacchiarotta T, Hurtadofernández E, Olmo-garcía L, García-Villalba R, Fernández-Gutiérrez A, et al. Comparing two metabolic profiling approaches (liquid chromatography and gas chromatography coupled to mass spectrometry) for extra-virgin olive oil phenolic compounds analysis : A botanical classification perspective. J Chromatogr A. 2016;1428:267-279. DOI: 10.1016/j.chroma.2015.10.059

[54] Mohamed M Ben, Rocchetti G, Montesano D, Ali S Ben, Guasmi F, Grati-Kamoun N, et al. Discrimination of Tunisian and Italian extra-virgin olive oils according to their phenolic and sterolic fingerprints. Food Res Int. 2018;106:920-927. DOI: 10.1016/j. foodres.2018.02.010

[55] Nescatelli R, Bonanni RC, Bucci R, Magrì AL, Magrì AD, Marini F. Geographical traceability of extra virgin olive oils from Sabina PDO by chromatographic fingerprinting of the phenolic fraction coupled to chemometrics. Chemom Intell Lab Syst. 2014;139:175-180.

[56] Romero M, Motilva M. Effect of climatic conditions on quality of virgin olive oil. In: Preedy VR, Watson RR, editors. Olives and Olive Oil in Health and Disease Prevention. Elsevier Inc.; 2010. p.43-50.

[57] Ben Rouina Y, Zouari M, Zouari N, Ben Rouina B, Bouaziz M. Olive tree (*Olea europaea* L. cv. Zelmati) grown in hot desert climate : Physio- biochemical responses and olive oil quality. Sci Hortic. 2020;261:108915. DOI: 10.1016/j. scienta.2019.108915

[58] Caruso G, Gucci R, Urbani S, Esposto S, Taticchi A, Di Maio I, et al. Effect of different irrigation volumes during fruit development on quality of virgin olive oil of cv. Frantoio. Agric Water Manag. 2014;134:94-103. DOI: 10.1016/j.agwat.2013.12.003

[59] Issaoui M, Flamini G, Brahmi F, Dabbou S, Ben Hassine K, Taamali A, et al. Effect of the growing area conditions on differentiation between Chemlali and Chétoui olive oils. Food Chem. 2010;119(1):220-225.

[60] Morales-Sillero A, Jiménez R, Fernández JE, Troncoso A, Beltrán G. Influence of fertigation in "Manzanilla de Sevilla" olive oil quality. HortScience. 2007;42(5):1157-1162. DOI: 10.21273/ HORTSCI.42.5.1157

[61] Dag A, Erel R, Kerem Z, Ben-Gal A, Stern N, Bustan A, et al. Effect of nitrogen availability on olive oil quality. Acta Hortic. 2018;1199:465-469. DOI: 10.17660/ActaHortic.2018.1199.74

[62] Alagna F, Mariotti R, Panara F, Caporali S, Urbani S, Veneziani G, et al. Olive phenolic compounds: Metabolic and transcriptional profiling during fruit development. BMC Plant Biol. 2012;12:162. DOI: 10.1186/1471- 2229-12-162

[63] Malheiro R, Rodrigues N, Pereira JA. Olive oil phenolic composition as affected by geographic origin, olive cultivar, and cultivation systems. In: Boskou D, editor. Olive and Olive Oil Bioactive Constituents. American Oil Chemists' Society Press; 2015. p.93-121. DOI: 10.1016/B978-1-63067-041-2. 50010-0

[64] Romero N, Saavedra J, Tapia F, Sepúlveda B, Aparicio R. Influence of agroclimatic parameters on phenolic and volatile compounds of Chilean virgin olive oils and characterization based on geographical origin, cultivar and ripening stage. J Sci Food Agric. 2016;96(2):583-592. DOI: 10.1002/ jsfa.7127

[65] Kalogeropoulos N, Kaliora AC. Effect of fruit maturity on olive oil phenolic composition and antioxidant capacity. In: Boskou D, editor. Olive and Olive Oil Bioactive Constituents. American Oil Chemists' Society Press; 2015. p.123-145.

[66] Servili M, Taticchi A, Esposto S, Urbani S, Selvaggini R, Montedoro G. Influence of the decrease in oxygen during malaxation of olive paste on the composition of volatiles and phenolic compounds in virgin olive oil. J Agric Food Chem. 2008;56(21):10048-10055. DOI: 10.1021/jf800694h

[67] Frankel E, Bakhouche A, Lozano-Sánchez J, Segura-Carretero A, *Virgin Olive Oil Phenolic Compounds: Insights on Their Occurrence, Health-Promoting… DOI: http://dx.doi.org/10.5772/intechopen.98581*

Fernández-Gutiérrez A. Literature review on production process to obtain extra virgin olive oil enriched in bioactive compounds. Potential use of byproducts as alternative sources of polyphenols. J Agric Food Chem. 2013;61(22):5179-5188.

[68] Antonini E, Farina A, Scarpa ES, Frati A, Ninfali P. Quantity and quality of secoiridoids and lignans in extra virgin olive oils : The effect of two- and three-way decanters on Leccino and Raggiola olive cultivars. Int J Food Sci Technol. 2015;67(1):9-15. DOI: 10.3109/09637486.2015.1121473

[69] Guerrini L, Migliorini M, Giusti M, Parenti A. The influence of crusher speed on extra virgin olive oil characteristics. Eur J Lipid Sci Technol. 2017;119(1):1600156. DOI: 10.1002/ ejlt.201600156

[70] Ouni Y, Guerfel M, Chedly A, Salma Nayet M, Zarrouk M, Flamini G. Changes in volatile compounds and oil quality with malaxation time of Tunisian cultivars of *Olea europea*. Int J Food Sci Technol. 2013;48(1):74-81.

[71] Jolayemi OS, Tokatli F, Ozen B. Effects of malaxation temperature and harvest time on the chemical characteristics of olive oils. Food Chem. 2016;211:776-783. DOI: 10.1016/j. foodchem.2016.05.134

[72] Lolis A, Badeka A V., Kontominas MG. Quality retention of extra virgin olive oil , Koroneiki cv. packaged in bag-in- box containers under long term storage : A comparison to packaging in dark glass bottles. Food Packag Shelf Life. 2020;26:100549. DOI: 10.1016/j.fpsl.2020.100549

[73] Li X, Zhu H, Shoemaker CF, Wang SC. The effect of different cold storage conditions on the compositions of extra virgin olive oil. J Am Oil Chem Soc. 2014;91(9):1559-1570. DOI: 10.1007/s11746-014-2496-0

[74] Olmo-García L, Carrasco-Pancorbo A. Chromatography-MS based metabolomics applied to the study of virgin olive oil bioactive compounds: Characterization studies, agrotechnological investigations and assessment of healthy properties. TrAC - Trends Anal Chem. 2021;135:116153. DOI: 10.1016/j.trac.2020.116153

[75] Wang P, Sun J, Zhang T, Liu W. Vibrational spectroscopic approaches for the quality evaluation and authentication of virgin olive oil. Applied Spectroscopy Reviews. 2016;51(10): 763-790. DOI: 10.1080/05704928.2016.1176034

[76] Lioupi A, Nenadis N, Theodoridis G. Virgin olive oil metabolomics: A review. J Chromatogr B. 2020;1150:122161. DOI: 10.1016/j.jchromb.2020.122161

[77] Waterhouse AL. Determination of Total Phenolics. Current Protocols in Food Analytical Chemistry. 2003;6:I1.1.1-I1.1.8. DOI: 10.1002/0471142913.fai0101s06

[78] Din I, Seccia S, Senatore A, Coppola D, Morelli E. Development and validation of an analytical method for total polyphenols quantification in extra virgin olive oils. Food Anal Methods. 2020;13:457-464. DOI: 10.1007/ s12161-019-01657-7

[79] Dais P, Hatzakis E. Quality assessment and authentication of virgin olive oil by NMR spectroscopy: A critical review. Anal Chim Acta. 2013;765:1-27.

[80] Hatzakis E. Nuclear Magnetic Resonance (NMR) Spectroscopy in Food Science: A Comprehensive Review. Compr Rev Food Sci Food Saf. 2019;18(1):189-220. DOI: 10.1111/1541-4337.12408

[81] Ramautar R, Somsen GW, de Jong GJ. CE-MS for metabolomics: Developments and applications in the period 2012-2014. Electrophoresis. 2015;36(1):212-224. DOI: 10.1002/ elps.201400388

[82] Valdés A, Cifuentes A, León C. Foodomics evaluation of bioactive compounds in foods. TrAC - Trends Anal Chem. 2017;96:2-13. DOI: 10.1016/j.trac.2017.06.004

[83] Rigacci S. Olive oil phenols as promising multi-targeting agents against Alzheimer's disease. Adv Exp Med Biol. 2015;863:1-20. DOI: 10.1007/978-3-319-18365-7\_1

[84] Rigacci S, Guidotti V, Bucciantini M, Nichino D, Relini A, Berti A, et al. Aβ(1-42) aggregates into non-toxic amyloid assemblies in the presence of the natural polyphenol oleuropein aglycon. Curr Alzheimer Res. 2011;8(8):841-852. DOI: 10.2174/156720511798192682

[85] Estruch R, Martínez-González MA, Corella D, Salas-Salvadó J, Ruiz-Gutiérrez V, et al. Effects of a Mediterranean-style diet on cardiovascular risk factors: A randomized trial. Ann Intern Med. 2006;145(1):1-11.

[86] Marcelino G, Hiane PA, Freitas K de C, Santana LF, Pott A, Donadon JR, et al. Effects of olive oil and its minor components on cardiovascular diseases, inflammation, and gut microbiota. Nutrients. 2019;11(8):1826. DOI: 10.3390/nu11081826

[87] Priore P, Caruso D, Siculella L, Gnoni G V. Rapid down-regulation of hepatic lipid metabolism by phenolic fraction from extra virgin olive oil. Eur J Nutr. 2015;54(5):823-833. DOI: 10.1007/ s00394-014-0761-5

[88] Casaburi I, Puoci F, Chimento A, Sirianni R, Ruggiero C, Avena P, et al. Potential of olive oil phenols as chemopreventive and therapeutic agents against cancer : A review of *in vitro* studies. Mol Nutr Food Res. 2013;57:71-83.

[89] Lozano-Castellón J, López-Yerena A, De JFRA, Castillo-Alba JR del, Vallverdú-Queralt A, Elvira E-F, et al. Health-promoting properties of oleocanthal and oleacein: Two secoiridoids from extra-virgin olive oil. Crit Rev Food Sci Nutr. 2020;60(15):2532-2548. DOI: 10.1080/10408398.2019.1650715

[90] Tuck KL, Hayball PJ. Major phenolic compounds in olive oil: Metabolism and health effects. J Nutr Biochem. 2002;13:636-644.

[91] D'Archivio M, Filesi C, Varì R, Scazzocchio B, Masella R. Bioavailability of the polyphenols: Status and controversies. Int J Mol Sci. 2010;11(4):1321-1342.

[92] Fabiani R, Sepporta MV, Rosignoli P, De Bartolomeo A, Crescimanno M, Morozzi G. Antiproliferative and pro-apoptotic activities of hydroxytyrosol on different tumour cells: The role of extracellular production of hydrogen peroxide. Eur J Nutr. 2012;51(4):455-464. DOI: 10.1007/ s00394-011-0230-3

[93] St-Laurent-Thibault C, Arseneault M, Longpre F, Ramassamy C. Tyrosol and hydroxytyrosol, two main components of olive oil, protect N2a cells against amyloid-β-induced toxicity. Involvement of the NF-κB signaling. Curr Alzheimer Res. 2011;8(5):543-551. DOI: 10.2174/156720511796391845

[94] Cao K, Xu J, Zou X, Li Y, Chen C, Zheng A, et al. Hydroxytyrosol prevents diet-induced metabolic syndrome and attenuates mitochondrial abnormalities in obese mice. Free Radic Biol Med. 2014;67:396-407. DOI: 10.1016/j.freeradbiomed. 2013.11.029

*Virgin Olive Oil Phenolic Compounds: Insights on Their Occurrence, Health-Promoting… DOI: http://dx.doi.org/10.5772/intechopen.98581*

[95] Ozbek N, Bali EB, Karasu C. Quercetin and hydroxytyrosol attenuates xanthine/xanthine oxidaseinduced toxicity in H9c2 cardiomyocytes by regulation of oxidative stress and stress-sensitive signaling pathways. Gen Physiol Biophys. 2015;34(4):407-414. DOI: 10.4149/gpb\_2015021

[96] Muriana FJG, Montserrat-De La Paz S, Lucas R, Bermudez B, Jaramillo S, Morales JC, et al. Tyrosol and its metabolites as antioxidative and antiinflammatory molecules in human endothelial cells. Food Funct. 2017;8(8):2905-2914. DOI: 10.1039/ c7fo00641a

[97] Abe R, Beckett J, Abe R, Nixon A, Rochier A, Yamashita N, et al. Olive oil polyphenol oleuropein inhibits smooth muscle cell proliferation. Eur J Vasc Endovasc Surg. 2011;41(6):814-820. DOI: 10.1016/j.ejvs.2010.12.021

[98] Sepporta MV, Fuccelli R, Rosignoli P, Ricci G, Servili M, Morozzi G, et al. Oleuropein inhibits tumour growth and metastases dissemination in ovariectomised nude mice with MCF-7 human breast tumour xenografts. J Funct Foods. 2014;8(1):269-273. DOI: 10.1016/j. jff.2014.03.027

[99] Galanakis PA, Bazoti FN, Bergquist J, Markides K, Spyroulias GA, Tsarbopoulos A. Study of the interaction between the amyloid beta peptide (1-40) and antioxidant compounds by nuclear magnetic resonance spectroscopy. Biopolymers. 2011;96(3):316-327. DOI: 10.1002/ bip.21558

[100] Mehraein F, Sarbishegi M, Aslani A. Evaluation of effect of oleuropein on skin wound healing in aged male Balb/c Mice. Cell J. 2014;16(1):25-30.

[101] Polini B, Digiacomo M, Carpi S, Bertini S, Gado F, Saccomanni G, et al. Oleocanthal and oleacein contribute to the in vitro therapeutic potential of extra virgin oil-derived extracts in non-melanoma skin cancer. Toxicol Vitr. 2018;52:243-250. DOI: 10.1016/j. tiv.2018.06.021

[102] Rosignoli P, Fuccelli R, Fabiani R, Servili M, Morozzi G. Effect of olive oil phenols on the production of inflammatory mediators in freshly isolated human monocytes. J Nutr Biochem. 2013;24(8):1513-1519. DOI: 10.1016/j.jnutbio.2012.12.011

[103] Czerwińska ME, Kiss AK, Naruszewicz M. Inhibition of human neutrophils NEP activity, CD11b/CD18 expression and elastase release by 3,4-dihydroxyphenylethanol-elenolic acid dialdehyde, oleacein. Food Chem. 2014;153:1-8. DOI: 10.1016/j. foodchem.2013.12.019

[104] Monti MC, Margarucci L, Riccio R, Casapullo A. Modulation of tau protein fibrillization by oleocanthal. J Nat Prod. 2012;75(9):1584-1588.

[105] Pei T, Meng Q, Han J, Li HSL, Song R, Sun B, et al. (−)-Oleocanthal inhibits growth and metastasis by blocking activation of STAT3 in human hepatocellular carcinoma. Oncotarget. 2016;7(28):43475-43491. DOI: 10.18632/ oncotarget.9782

[106] Scotece M, Gómez R, Conde J, Lopez V, Gómez-Reino JJ, Lago F, et al. Further evidence for the antiinflammatory activity of oleocanthal: Inhibition of MIP-1α and IL-6 in J774 macrophages and in ATDC5 chondrocytes. Life Sci. 2012;91(23- 24):1229-1235. DOI: 10.1016/j. lfs.2012.09.012

[107] Cicerale S, Lucas L, Keast R. Biological activities of phenolic compounds present in virgin olive oil. Int J Mol Sci. 2010;11(2):458-479.

[108] Corona G, Tzounis X, DessÌ MA, Deiana M, Debnam ES, Visioli F, et al. The fate of olive oil polyphenols in the gastrointestinal tract : Implications of gastric and colonic microfloradependent biotransformation. Free Radic Res. 2006;40(6):647-658. DOI: 10.1080/10715760500373000

[109] Visioli F, Galli C, Grande S, Colonnelli K, Patelli C, Galli G, et al. Hydroxytyrosol excretion differs between rats and humans and depends on the vehicle of administration. J Nutr. 2003;133(8):2612-2615.

[110] Miró-Casas E, Covas M-I, Fitó M, Farré-Albadalejo M, Marrugat J, Torre R de la. Tyrosol and hydroxytyrosol are absorbed from moderate and sustained doses of virgin olive oil in humans. Eur J Clin Nutr. 2003;57:186-190.

[111] Manna C, Galletti P, Maisto G, Cucciolla V, D'Angelo S, Zappia V. Transport mechanism and metabolism of olive oil hydroxytyrosol in Caco-2 cells. FEBS Lett. 2000;470:341-344.

[112] Edgecombe SC, Stretch GL, Hayball PJ. Oleuropein, an antioxidant polyphenol from olive oil, is poorly absorbed from isolated perfused rat intestine. J Nutr. 2000;130(12):2996-3002.

[113] Boccio P Del, Deo A Di, Curtis A De, Celli N, Iacoviello L, Rotilio D. Liquid chromatography–tandem mass spectrometry analysis of oleuropein and its metabolite hydroxytyrosol in rat plasma and urine after oral administration. J Chromatogr B. 2003;785(1):47-56.

[114] Vissers MN, Zock PL, Roodenburg AJC, Leenen R, Katan MB. Olive oil phenols are absorbed in humans. J Nutr. 2002;132(3):409-417.

#### **Chapter 9**

## Experimental Carcinogenesis with 7,12-Dimethylbenz(a)Anthrazene (DMBA) and Its Inhibition with Extra Virgin Olive Oil and a Diet of Mature Olives (*Picual* Variety)

*Juan José Soto-Castillo and Isicio Ortega-Medina*

### **Abstract**

7,12-Dimethylbenz(a)anthrazene (DMBA) is a carcinogen that induces carcinomas within a few weeks of application. Forty-four male hamsters were divided into four groups: DMBA dissolved in paraffin oil (DMBA-PO), DMBA dissolved in olive oil (DMBA-OO), paraffin oil and olive oil. Their mouths were swabbed daily with paraffin oil or extra virgin olive oil alternatively for the first two weeks, during the biweekly application of DMBA at 0.5% diluted in paraffin oil or olive oil for five weeks and daily until the twentieth week. The animals in the DMBA-OO and olive oil groups received an additional diet of mature *Picual* olives. The DMBA-PO carcinogen effect (35 carcinomas) is 100% and the inhibitory effect 0. The use of olive oil as DMBA solvent and the *ad libitum* diet with *Picual* olive has an inhibitory effect of 80%, with only three intraepithelial carcinomas and four verrucous carcinomas occurring and no invasive carcinoma.

**Keywords:** DMBA, carcinogenesis, olive oil, chemoprevention, squamous cell carcinoma

### **1. Introduction**

One of the first models of experimental carcinogenesis in animals was carried out by Salley in 1954 [1]. After applying various carcinogens, including 9,10-dimethyl-1,2 benzanthracene on the oral epithelium of Syrian hamsters for 3 months, Salley was able to verify the existence of squamous cell carcinomas (SCC) and lymphatic metastases. Subsequently, several authors have standardized this model and repeated it in order to achieve new knowledge about DMBA and the process of experimentally induced carcinogenesis [2, 3].

7,12-Dimethylbenz[a]anthracene (DMBA) is a polycyclic aromatic hydrocarbon which may, on its own, induce premalignant lesions and carcinomas within a few weeks after it is administered in mucosae [4, 5]. Commonly, it has been used in combination with ethanol as a promoter. DMBA is released after the combustion of tobacco -especially with cigarettes- or from animal fat when meat is grilled, and is also found in smoked meat and fish. This substance is, therefore, strongly involved in the carcinogenesis of oral, bronchiopulmonary and digestive tract malignancies [6–9].

In order to discover new drugs with cancer preventive effects, some authors have obtained promising outcomes at basic research level, specifically with substances such as salvinolic acid B [10] -derived from *Salvia miltiorrhiza*, used in fluorescence-, isothiocyanates [11] -synthetic derivatives of cabbages, squash, turnips and turnip greens-, *Buddleja incana* leaves, a tree that grows in Peru and Bolivia, Toona sinensis leaves [12], and olive oil extracts [9, 13–15]. In relation to the latter, and especially regarding its phenolic compounds, its antioxidant and cardiovascular protective properties are well known. In this sense, we have data stating that olive oil may act as preventive or inhibitor of carcinogenesis, and could even modify the nature of premalignant lesions that have already arisen, providing them a more benign and indolent behavior [14, 16].

#### **2. Objective**

To experimentally test the inhibitory effect on the carcinogenesis process of *Picual* variety extra virgin olive oil.

#### **3. Methods**

Forty-four male hamsters (*Syrian Golden*), 4-6 weeks old and weighing 60-80 g, were divided into four groups (two control and two experimental):


The animals in each group were sacrificed after twenty weeks. Then, a macroscopic description and histological analysis of the induced tumors in the oropharynx, esophagus and stomach were performed.

*Experimental Carcinogenesis with 7,12-Dimethylbenz(a)Anthrazene (DMBA) and Its… DOI: http://dx.doi.org/10.5772/intechopen.96514*

A carcinogenic effect of 100% was assigned to the total number of induced tumors in hamsters who received DMBA-PO. The inhibitory effect in the DMBA-OO group was established by the percentage difference over 100. An inhibitory effect >50% was considered significant in the DMBA-OO group.

This research work was examined and approved by the Ethical Committee for Animal Experimentation of the University of Seville (November 7, 2005). It met the requirements for experimentation with animals and was in accordance with the regulations in force in Spain and elsewhere the European Union.

#### **4. Results**

#### **4.1 Macroscopically**

The groups exposed to DMBA showed tumors of different characteristics. Nonspecific lesions and others more suggestive of malignancy were found in the oral pouches of the DMBA-OO group, with a predominance of the former. These findings included leukoplakia, denudation of the mucosa, ulcerations or tumors with a benign appearance. However, tumors in DMBA-OO group were less common and smaller than in DMBA-PO (**Figure 2A** and **B**).

On the other hand, the DMBA-PO group mostly showed malignantlooking neoplastic formations in the oral mucosa, such as ulcerated nodules, necrosis areas, exophytic and verrucous tumors, and areas with abundant vascularization.

In addition to the oral pouches, both DMBA groups presented tumors in the esophagus and stomach. Maximum and minimum measures of all lesions are shown in **Table 1**.

No visible lesions were found in the control groups which only received paraffin oil or olive oil.

#### **Figure 2.**

*Macroscopic comparison of the digestive tract of two animals belonging to the DMBA-OO group (A) and DMBA-PO group (B). (A) The oral pouches, esophagus and stomach showed few small lesions and benign appearance. (B) The oral pouches, esophagus, and stomach showed tissue retractions, larger tumors and apparently more malignant lesions.*


#### **Table 1.**

*Number, size and type of tumors in DMBA groups at 20th week.*

#### **4.2 Microscopically**

The histological study at 20 weeks evidenced different types of lesions, demonstrating a complete carcinogenesis process in both DMBA groups: Squamous papillomas, intraepithelial carcinomas, verrucous carcinomas and invasive SCC. *Experimental Carcinogenesis with 7,12-Dimethylbenz(a)Anthrazene (DMBA) and Its… DOI: http://dx.doi.org/10.5772/intechopen.96514*

SQUAMOUS PAPILLOMAS: Papillary projections lined with squamous epithelium were noted, showing hyperkeratosis and epithelial thickening. No atypia or mitotic activity was observed (**Figure 3**). Twelve papillomas were found among the groups exposed to DMBA (one in the DMBA-PO group and eleven in the DMBA-OO group). The differences regarding incidence of this kind of lesion were statistically significant (p .004).

INTRAEPITHELIAL CARCINOMAS (**Figure 4**): Twenty four intraepithelial carcinomas were identified. Twenty one occurred in the DMBA-PO group, and three in the DMBA-OO group. The differences observed between both groups were statistically significant (p .003).

#### **Figure 3.**

*Squamous papilloma is an exophytic lesion which shows typically papillary growth and highly differentiated epithelium.*

#### **Figure 4.**

*Intraepithelial carcinoma is classically characterized by full-thickness with hyperkeratosis and parakeratosis, hypercellurality, nuclear atypia and mitotic figures. The epithelium-stroma interface is preserved.*

#### **Figure 5.**

*SCC, verrucous carcinoma. Verrucous carcinoma is warty-appearing, highly differentiated, and shows hyperkeratosis. There is minimal atypia, abundant eosinophilic cytoplasm and normal mitotic figures. No invasion of the stroma by isolated neoplastic cells was observed.*

#### **Figure 6.**

*Invasive SCC (A). The SCC is composed of infiltrating islands or nets of malignant cells, which form an irregular growth pattern (B).*


#### **Table 2.**

*Carcinogenic and inhibitory effects of DMBA-PO/DMBA-OO, according to lesion subtypes.*

*Experimental Carcinogenesis with 7,12-Dimethylbenz(a)Anthrazene (DMBA) and Its… DOI: http://dx.doi.org/10.5772/intechopen.96514*

VERRUCOUS CARCINOMAS: Several exophytic lesions with papillomatosis and infiltrative growth (**Figure 5**). Thirteen verrucous carcinomas were found, nine in the in the DMBA-PO group and four in the DMBA-OO group. This was not statistically significant (p .523).

INVASIVE CARCINOMAS (**Figure 6**): Light microscopy revealed epithelial proliferations that, like cords, invaded the adjacent stroma. In addition, the proliferating cells showed marked atypia and mitotic activity. Five invasive carcinomas were found in the DMBA-PO group.

The carcinogenic effect in the DMBA-PA group (35 carcinomas) corresponded to 100%, while in the DMBA-OO group (7 carcinomas), it was of 20%. According to the observed results, inhibitory effect seen in the DMBA-OO group was 86% for intraepithelial carcinoma, 56% for verrucous carcinoma, and 100% for SCC (**Table 2**).

No tumors were observed in the control animals.

#### **5. Discussion**

This research work about carcinogenesis is based on an experimental model of induced SCC after the administration of DMBA at 0.5% -dissolved in mineral oil- into the oral pouches of the hamster. We think, like Nagini and Kowshik [3], that the DMBA carcinogenesis model in hamster oral pouches is characteristic and highly representative of the "cancer induction". In addition, it is advantageous for the reproducibility of lesions, facilitates experimental research, and can be used as a test for chemotherapy and preventive agents. Also, in this work, the olive oil inhibitory effect on carcinogenesis has been studied alone -extra virgin olive oil applied before, during, and after DMBA, and *ad libitum* diet with ripe olives ripe of the *Picual* variety, from the olive harvest-, and combined -as a solvent for DMBA- [16].

The carcinomas produced in the upper gastrointestinal tract were SCC, similar to SCC of the oral mucosa in humans. These results coincide with those obtained in other experimental works [17, 18].

In oral carcinogenesis, using DMBA in hamsters, some authors have described the development of precancerous lesions and, subsequently, their progression towards intraepithelial carcinoma and invasive carcinoma after a few months. At 8 weeks, precancerous lesions usually appear. At 12 weeks, these evolve to intraepithelial carcinoma; eventually developing into invasive carcinomas at 18 weeks. This phenomenon, although slower, also occurs in humans [19]. The results obtained in our work resemble those of oral cancer progression described in the literature.

As in the field of experimental carcinogenesis, research on cancer prevention has continued to grow in recent decades, focusing on agents proposed for this purpose, although with few results yet. This is the case of the mediterranean diet, which is largely based on extra virgin olive oil, and that has been explored in the prevention of breast cancer [11, 13], and colorrectal cancer [9]. In the present work, the combination of olive oil as dissolvent, extra virgin olive oil applied before, during, and after DMBA application, and *ad libitum* diet with *Picual* variety olives, have been used as a preventive agent of DMBA carcinogenesis.

Menéndez et al. have shown that extra virgin olive oil polyphenols can inhibit erbB-2 malignant transformation of human breast cancer epithelial cells [14]. Owen et al. pointed out the importance of phenolic compounds isolated from olive oil as antioxidants and their anticancer potential [20].

In this sense, olive oil is composed of 99% different fatty acids, the most important being oleic acid, a monounsaturated fatty acid, with a richness of 60-80%,

and other fatty acids -palmitic, stearic, palmitoleic, linoleic, and linolenic-. The remaining 1% is made up of vitamin E and natural antioxidants. The most important antioxidants are phenolic compounds, present in the mesocarp of the olive and in extra virgin olive oil, which are mainly responsible for the antioxidant properties and which are not present in any other vegetable oil. For this reason, the diet added to the standard feed that the hamsters received was ripe olives from the tree, recently harvested and not spoiled. The variety of olive richest in phenolic compounds is the *Picual* variety.

Keys et al. demonstrated an inversely proportional relationship between the incidence of cardiovascular diseases and the adoption of eating habits established in seven countries in the Mediterranean area [21]. It seems that this "cardiovascular protection" resides in the creation of an anti-atherosclerotic plasma profile, which is defined by a decrease in total cholesterol and low-density lipoprotein (LDL) cholesterol levels, as well as by an increase in high-density lipoprotein (HDL) cholesterol. Some studies have attributed these properties to the high content of oleic acid -monounsaturated grade acid of the omega-9 series- of olive oil [22].

Analyzing our results, we can affirm that the combination of olive oil as a solvent for DMBA, extra virgin olive oil applied before, during, and after DMBA administration, and *ad libitum* diet with *Picual* olives has shown the capability to reduce the malignant progression of lesions already started, and modify the malignant phenotype of some neoplasms, making it less aggressive.

It is possible that in the DMBA-OO group, -COOH groups and unsaturated bonds of the vegetable oil could absorb or react with carcinogen, decreasing the effective concentration of the carcinogen. The antioxidant effect and anticancer properties of extra virgin olive oil expressed by some authors are reinforced [18, 19].

The study of the lesions at 20 weeks showed a total of 59 neoplasms in the DMBA-PO group and 58 in the DMBA-OO group, so there were no differences in the absolute incidence of tumors. However, clear differences were observed regarding the type of neoplasms and malignancy. Eighty-eight percent of the tumors in the DMBA-OO group corresponded to benign squamous papilloma-type tumors, compared to 41% that developed in the DMBA-PO group; the rest were carcinomas.

In addition, hamsters that did not eat ripe olives and did not receive extra virgin olive oil, developed 21 intraepithelial carcinomas, 9 verrucous carcinomas, and 5 invasive carcinomas; while animals that received the olive oil as a solvent for DMBA, extra virgin olive oil -before, during, and after DMBA-, and *ad libitum* diet with *Picual* olive developed 3 intraepithelial carcinomas, 4 verrucous carcinomas, and no invasive squamous carcinoma.

#### **6. Conclusions**

The inhibitory effect of extra virgin olive oil (*Picual* variety) on the experimental chemical carcinogenesis is higher than 50% for carcinomas, especially for intraepithelial carcinoma and invasive squamous carcinoma.

Furthermore, the tumors originated in animals who received DMBA mixed with olive oil were predominantly benign, specifically of the squamous papilloma subtype.

Therefore, these data suggest that the extra virgin olive oil and the diet with ripe olives extracted from the harvesting of the tree may modulate the experimental carcinogenesis with DMBA, originating very well differentiated and not very aggressive tumors.

*Experimental Carcinogenesis with 7,12-Dimethylbenz(a)Anthrazene (DMBA) and Its… DOI: http://dx.doi.org/10.5772/intechopen.96514*

#### **Conflict of interest**

The authors declare no conflict of interest.

#### **Author details**

Juan José Soto-Castillo\* and Isicio Ortega-Medina Faculty of Medicine, University of Seville, Seville, Spain

\*Address all correspondence to: jj27sc@gmail.com

© 2021 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

### **References**

[1] Salley JJ. Experimental carcinogenesis in the cheek pouch of the Syrian Hamster. J Dent Res. 1954;33: 253-62. DOI: 10.1177/00220345540330021201.

[2] Santis H, Shklar G, Chauncey HH. Histochemistry of experimentally induced leukoplakia and carcinoma of the hamster buccal pouch. Oral Surg Oral Med Oral Pathol. 1964;17:207-18. DOI: 10.1016/0030-4220(64)90144-6.

[3] Nagini S, Kowshik J. The hamster buccal pouch model of oral carcinogenesis. Methods Mol Biol. 2016;1422:341-50. DOI: 10.1007/978-1-4939-3603-8\_29.

[4] Hassan MM, Shklar G, Solt D, Szabo G. Acute effect of DMBA application on mitotic activity of hamster buccal pouch epithelium. Oral Surg Oral Med Oral Pathol. 1985;59:491- 8. DOI: 10.1016/0030-4220(85)90090-8.

[5] García FJG, Ortega VV, Sánchez NA, Jornet PL. Estudio comparativo de la aplicación del hidrocarburo aromático policíclico 7,12-dimetil-1,2 benzatraceno (DMBA) sobre la mucosa oral del hámster y del cobaya. Rev Española Patol. 2009;42: 287-95. DOI: 10.1016/S1699-8855(09)70196-5.

[6] Solt DB, Polverini PJ, Calderon L. Carcinogenic response of hamster buccal pouch epithelium to 4 polycyclic aromatic hydrocarbons. J Oral Pathol. 1987;16:294-302. DOI: 10.1111/j.1600- 0714.1987.tb00697.x.

[7] Nagabhushan M, Ng YK, Elias R, Polverini PJ, Solt DB. Acute inhibition of DNA synthesis in hamster buccal pouch epithelium exposed to indirect acting carcinogens. Cancer Lett. 1990;53:163-73. DOI: 10.1016/0304-3835(90)90210-o.

[8] Calderon-Solt L, Solt DB. Gamma-glutamyl transpeptidase in precancerous lesions and carcinomas of oral, pharyngeal, and laryngeal mucosa. Cancer. 1985;56:138-43. DOI: 10.1002/1097-0142(19850701) 56:1<138::aid-cncr2820560122>3.0. co;2-4.

[9] Bartolí R, Fernández-Bañares F, Navarro E, Castellà E, Mañé J, Alvarez M, et al. Effect of olive oil on early and late events of colon carcinogenesis in rats: Modulation of arachidonic acid metabolism and local prostaglandinE(2) synthesis. Gut. 2000;46:191-9. DOI: 10.1136/gut.46.2.191.

[10] Zhou ZT, Ge JP. The preventive effect of salvianolic acid B on malignant transformation of DMBA-induced oral premalignant lesion in hamsters. Carcinogenesis. 2006;27:826-32. DOI: 10.1093/carcin/bgi271.

[11] Warin R, Xiao D, Arlotti JA, Bommareddy A, Singh SV. Inhibition of human breast cancer xenograft growth by cruciferous vegetable constituent benzyl isothiocyanate. Mod Carcinog. 2010;49:500-7. DOI: 10.1002/mc.20600.

[12] Wang WC, Chen CY, Hsu HK, Lin LM, Chen YK. Chemopreventive effect of Toona sinensis leaf extracton 7,12-dimethylbenz(a)anthraceneinduced hamster buccal pouch squamous cell carcinogenesis. Arch Oral Biol. 2016;70:130-42. DOI: 10.1016/j. archoralbio.2016.06.015.

[13] Manzanares MA, Solanas M, Moral R, Escrich R, Vela E, Costa I, et al. Dietary extra-virgin olive oil and corn oil differentially modulate the mRNA expression of xenobioticmetabolizing enzimas in the liver and in the mammary gland in a rat chemical induced breast cancer model. Eur J Cancer Prev. 2015;24:215-22. DOI: 10.1097/CEJ.0000000000000032.

[14] Menéndez JA, Vázquez A, Oliveras C, García R, Carrasco A, *Experimental Carcinogenesis with 7,12-Dimethylbenz(a)Anthrazene (DMBA) and Its… DOI: http://dx.doi.org/10.5772/intechopen.96514*

Fernández A, et al. Extra-virgin olive oil polyphenols inhibit HER2 (erbB-2) induced malignant transformation in human breast epitelial cells: Relationship between the chemical structures of extra-virgin olive oil secoiridoids and lignans and their inhibitory activities on the tyrosine kinase activity of HER2. Int J Oncol. 2008;34:43-51.

[15] Solanas M, Hurtado A, Costa I, Moral R, Menéndez JA, Colomer R, et al. Effects of a high olive oil diet on the clinical behavior and histopathological features of rat DMBAinduced mammary tumors compared with a high corn oil diet. Int J Oncol. 2002;21:745-53.

[16] Soto-Castillo JJ, Ortega-Medina I. Carcinogénesis experimental con 7,12 dimetilbenzantraceno (DMBA) y su inhibición con aceite de oliva virgen extra y dieta con aceitunas maduras (variedad Picual). Rev Esp Patol. 2017:50(2):82-88. DOI: 10.1016/j. patol.2016.10.001.

[17] Chen YK, Lin LM. DMBA-induced hamster buccal pouch carcinoma and VX-induced rabbit cancer as a model for human oral carcinogenesis. Expert Rev Anticancer Ther. 2010;10:1485-96. DOI: 10.1586/era.10.108.

[18] Tang X-H, Knudsen B, Bemis D, Tickoo S, Gudas LJ. Oral cavity and esophageal carcinogenesis modeled in carcinogen-treated mice. Clin Cancer Res. 2004;10:301-13. DOI: 10.1158/1078- 0432.ccr-0999-3.

[19] Vairaktaris E, Spyridonidou S, Papakosta V, Vylliotis A, Laza- ris A, Perrea D, et al. The hamster model of sequential oral oncogenesis. Oral Oncol. 2008;44:315-24. DOI: 10.1016/j. oraloncology.2007.08.015.

[20] Owen RW, Giacosa A, Hull WE, Haubner R, Spiegelhalder B, Bartsch H. The antioxidant/anticancer potential of phenolic compounds isolated from olive oil. Eur J Cancer. 2000;36:1235-47. DOI: 10.1016/s0959-8049(00)00103-9.

[21] Keys A, Menotti A, Karvonen MJ, Aravanis C, Blackburn H, Buzina R, et al. The diet and 15-year death rate in the seven countries study. Am J Epidemiol. 1986;124(6):903-15. DOI: 10.1093/oxfordjournals.aje.a114480.

[22] Ruiz V, Muriana FJ, Villar J. El aceite de oliva virgen y las enfermedades cardiovasculares. Perfil lipídico en plasma y composición lipídica de la membrana de eritrocito humano. Grasas y Aceites. 1998;49(1):9-29. DOI: 10.3989/gya.1998.v49.i1.703.

## *Edited by Muhammad Akram*

*Olive Oil - New Perspectives and Applications* is a collection of reviewed and relevant research chapters, offering a comprehensive overview of recent developments in the field of agricultural, medical, and biological sciences. The book comprises single chapters authored by various researchers and edited by an expert active in the olive oil research area. All chapters are complete in themselves but united under a common research study topic. This publication aims at providing a thorough overview of the latest research efforts by international authors on olive oil and opening new possible research paths for further novel developments.

Published in London, UK © 2022 IntechOpen © kuppa\_rock / iStock

Olive Oil - New Perspectives and Applications

Olive Oil

New Perspectives and Applications

*Edited by Muhammad Akram*