Búsqueda de sustancias nutraceuticas contenidas en la dieta mediterránea: análisis antigenotoxicológico, tumoricida, de antienvejecimiento y de marcas epigenéticas

La producción del sector hortofrutícola se ve incrementada cada año y, paralelamente, la sociedad actual demanda productos agrícolas de calidad. Esta calidad debe ser medida no sólo en un mejor sabor sino en sus potenciales efectos saludables. La dieta mediterránea es una dieta altamente valorada a nivel mundial debido a su alto contenido en antioxidantes naturales presentes en las matrices alimenticias que la constituyen como son las frutas y verduras entre otras (Visioli and Galli 2001; Oh et al. 2005; Saura-Calixto and Goñi 2006). Esta dieta es muy similar a la dieta de nuestros antepasados paleolíticos (Konner and Eaton 2010) y numerosos estudios multidisciplinares han mostrado los efectos positivos de esta dieta frente a enfermedades degenerativas como el cáncer, obesidad, diabetes y enfermedades cardiovasculares (Agarwal and Rao 2000; La Vecchia 2004; Salas-Salvadó et al. 2011; Estruch et al. 2013); además se le ha atribuido una asociación directa con la longevidad (Shahar and Itamar Grotto 2006)

Safety and Protective Activities of Manufactured Alcohol-Free Beers

Nowadays, a general interest in improving health in order to achieve better conditions of life is increasing. Diet is a complex factor affecting health conditions. We analysed the biological activities of three types of alcohol-free lager beer (a blond, a pale-blond and a stout beer) as well as epicatechin gallate (ECG) as one of their most abundant phenols with the aim of revealing them as nutraceuticals. For that purpose, we carried out safety and protective assays of the tested substances in the well-known Drosophila melanogaster animal model. Moreover, chemoprevention studies on human leukaemia cells (HL-60) in an in vitro model were carried out to evaluate the viability and genomic damage potential of the studied compounds on the tumour cell line. Results suggest the safety properties of all compounds, although pale-blond and stout beer only showed genotoxic activity at the lowest concentrations assayed. Moreover, alcohol-free beers and phenols were able to protect against H2O2 oxidative damage as well as to induce an increase in longevity with an improvement of the quality of life in the in vivo animal model assayed. Promising results were obtained with the alcohol-free beers and ECG in the in vitro assays with human leukaemia cells as they inhibited the tumour cells’ growth, induced DNA damage and modified the methylation status of such a cancer cell line. To sum up, alcohol-free beers should be of interest not only because of their reduced calories and isotonic properties but because they can be recognised as nutraceutical substances

Biological Effects of Food Coloring in In Vivo and In Vitro Model Systems

(1) Background: The suitability of certain food colorings is nowadays in discussion because of the effects of these compounds on human health. For this reason, in the present work, the biological effects of six worldwide used food colorings (Riboflavin, Tartrazine, Carminic Acid, Erythrosine, Indigotine, and Brilliant Blue FCF) were analyzed using two model systems. (2) Methods: In vivo toxicity, antitoxicity, and longevity assays using the model organism Drosophila melanogaster and in vitro cytotoxicity, DNA fragmentation, and methylation status assays using HL-60 tumor human cell line were carried out. (3) Results: Our in vivo results showed safe effects in Drosophila for all the food coloring treatments, non-significant protective potential against an oxidative toxin, and different effects on the lifespan of flies. The in vitro results in HL-60 cells, showed that the tested food colorings increased tumor cell growth but did not induce any DNA damage or modifications in the DNA methylation status at their acceptable daily intake (ADI) concentrations. (4) Conclusions: From the in vivo and in vitro studies, these results would support the idea that a high chronic intake of food colorings throughout the entire life is not advisable

Toxicological and Epigenetic Studies of Two Types of Ale Beer, Tyrosol and Iso-Alpha Humulone

Although many benefits drawn from beer consumption are claimed, the epidemiological records are contradictory with respect to cancer prevention. The purpose of this study was to investigate the possible health-related activities involving genome safety and the ageing processes of two types of lyophilised ale beers (blond and stout), as well as two of their bioactive compounds (tyrosol and iso-alpha humulone). A multipurpose trial set of in vivo toxicity, antitoxicity, mutagenicity, antimutagenicity, lifespan and healthspan assays using Drosophila melanogaster were used. In parallel, several in vitro assays were designed using the cancer cell line HL-60 in order to establish the possible chemopreventive activity of the selected substances, where epigenetic modulation of DNA methylation changes, clastogenic activity and tumour cell inhibition growth were evaluated. The safety of the four substances was confirmed: lyophilised blond ale beer (LBAB), lyophilised stout ale beer (LSAB), tyrosol and iso-alpha humulone were neither toxic nor genotoxic. Moreover, all substances, except tyrosol, revealed the ability to protect individual genomes against oxidative radicals and to exert antimutagenic activity against the genotoxin hydrogen peroxide. With respect to the degenerative process indicators of lifespan and healthspan, tyrosol was the only compound that did not exert any influence on the life extension of Drosophila; LBAB induced a significant lifespan extension in D. melanogaster; LSAB and its distinctive compound iso-alpha humulone induced a reduction in longevity. The in vitro assays showed the cytotoxic activity of LBAB, LSAB and tyrosol against HL-60 cells. Moreover, proapoptotic DNA fragmentation or DNA strand breakage was observed for both types of beers and iso-alpha humulone at different concentrations. Furthermore, the lyophilised ale beers and tyrosol exhibited an increasing genome-wide methylation status, while iso-alpha humulone exhibited a demethylation status in repetitive cancer cell sequences. Although the biological activities assigned to beer consumption cannot be linked to any specific molecule/element due to the complexity of the phenolic profile, as well as the multifactor brewing process, the results obtained let us propose lyophilised ale beers as safe potential nutraceutical beverages when consumed in moderate amounts. The prevention of toxicity and genetic oxidative damage, as well as the induction of tumor cell death and modulation of the methylation status, are the key activities of beer that were shown in the present research

In Vivo and In Vitro Genotoxic and Epigenetic Effects of Two Types of Cola Beverages and Caffeine: A Multiassay Approach

The aim of this work was to assess the biological and food safety of two different beverages: Classic Coca Cola™ (CCC) and Caffeine-Free Coca Cola (CFCC). To this end, we determined the genotoxicological and biological effects of different doses of lyophilised CCC and CFCC and Caffeine (CAF), the main distinctive constituent. Their toxic/antitoxic, genotoxic/antigenotoxic, and chronic toxicity (lifespan assay) effects were determined in vivo using the Drosophila model. Their cytotoxic activities were determined using the HL-60 in vitro cancer model. In addition, clastogenic DNA toxicity was measured using internucleosomal fragmentation and SCGE assays. Their epigenetic effects were assessed on the HL-60 methylation status using some repetitive elements. The experimental results showed a slight chemopreventive effect of the two cola beverages against HL-60 leukaemia cells, probably mediated by nonapoptotic mechanisms. Finally, CCC and CAF induced a global genome hypomethylation evaluated in LINE-1 and Alu M1 repetitive elements. Overall, we demonstrated for the first time the safety of this famous beverage in in vivo and in vitro models

In Vivo and In Vitro Assays Evaluating the Biological Activity of Taurine, Glucose and Energetic Beverages

Taurine is one of the main ingredients used in energy drinks which are highly consumed in adolescents for their sugary taste and stimulating effect. With energy drinks becoming a worldwide phenomenon, the biological effects of these beverages must be evaluated in order to fully comprehend the potential impact of these products on the health due to the fact nutrition is closely related to science since the population consumes food to prevent certain diseases. Therefore, the aim of this study was to evaluate the biological effects of taurine, glucose, classic Red Bull® and sugar-free Red Bull® in order to check the food safety and the nutraceutical potential of these compounds, characterising different endpoints: (i) Toxicology, antitoxicology, genotoxicology and life expectancy assays were performed in the Drosophila melanogaster model organism; (ii) The in vitro chemopreventive activity of testing compounds was determined by assessing their cytotoxicity, the proapoptotic DNA-damage capability to induce internucleosomal fragmentation, the strand breaks activity and the modulator role on the methylation status of genomic repetitive sequences of HL-60 promyelocytic cells. Whereas none tested compounds showed toxic or genotoxic effect, all tested compounds exerted antitoxic and antigenotoxic activity in Drosophila. Glucose, classic Red Bull® and sugar-free Red Bull® were cytotoxic in HL-60 cell line. Classic Red Bull® induced DNA internucleosomal fragmentation although none of them exhibited DNA damage on human leukaemia cells. In conclusion, the tested compounds are safe on Drosophila melanogaster and classic Red Bull® could overall possess nutraceutical potential in the in vivo and in vitro model used in this study. Besides, taurine could holistically be one of the bioactive compounds responsible for the biological activity of classic Red Bull®

Food Safety and Nutraceutical Potential of Caramel Colour Class IV Using In Vivo and In Vitro Assays

Nutraceutical activity of food is analysed to promote the healthy characteristics of diet where additives are highly used. Caramel is one of the most worldwide consumed additives and it is produced by heating natural carbohydrates. The aim of this study was to evaluate the food safety and the possible nutraceutical potential of caramel colour class IV (CAR). For this purpose, in vivo toxicity/antitoxicity, genotoxicity/antigenotoxicity and longevity assays were performed using the Drosophila melanogaster model. In addition, cytotoxicity, internucleosomal DNA fragmentation, single cell gel electrophoresis and methylation status assays were conducted in the in vitro HL-60 human leukaemia cell line. Our results reported that CAR was neither toxic nor genotoxic and showed antigenotoxic effects in Drosophila. Furthermore, CAR induced cytotoxicity and hipomethylated sat-α repetitive element using HL-60 cell line. In conclusion, the food safety of CAR was demonstrated, since Lethal Dose 50 (LD50) was not reached in toxicity assay and any of the tested concentrations induced mutation rates higher than that of the concurrent control in D. melanogaster. On the other hand, CAR protected DNA from oxidative stress provided by hydrogen peroxide in Drosophila. Moreover, CAR showed chemopreventive activity and modified the methylation status of HL-60 cell line. Nevertheless, much more information about the mechanisms of gene therapies related to epigenetic modulation by food is necessary

Antigenotoxicity and Tumor Growing Inhibition by Leafy Brassica carinata and Sinigrin

Cruciferous vegetables are well known and worldwide consumed due to their health benefits and cancer prevention properties. As a desirable cruciferous plant, Ethiopian mustard (Brassica carinata A. Braun) and its glucosinolate sinigrin were tested in the in vivo Drosophila melanogaster (SMART) and the in vitro HL60 (human promyelocytic leukaemia cell line) systems. High performance liquid chromatography (HPLC) analysis of plant samples confirmed the presence of sinigrin as principal B. carinata glucosinolate. SMART was performed by feeding D. melanogaster larvae either with different concentrations of plant/compound samples or combining them with hydrogen peroxide (a potent oxidative mutagen) being both antimutagenics. HL60 assays showed the tumoricidal activity of plant samples (IC50 = 0.28 mg·mL−1) and the breakdown products of sinigrin hydrolysis (IC50 = 2.71 μM). Our results enhance the potential of B. carinata as health promoter and chemopreventive in both systems and the leading role of sinigrin in these effects

Toxicological and Nutraceutical Screening Assays of Some Artificial Sweeteners

Artificial sweeteners are food additives worldwide used instead of fructose or glucose in many diet beverages. Furthermore, diet beverages intake has been increasing every year. Thus, some food agencies should regulate it based on toxicological studies. Debates and controversial results are demonstrated, and authority can revise its decision on the basis of new data reporting toxicological effects since cyclamate has been forbidden in some countries. Therefore, the aim of this study was to report new data about the toxicity of acesulfame-k, aspartame, and cyclamate, which are useful for authority agencies, determining the toxic potential and nutraceutical capabilities of these compounds. The toxicity, antitoxicity, genotoxicity, antigenotoxicity, and life expectancy assays were carried out in Drosophila as an in vivo model. In addition, in vitro HL-60 line cell was used to evaluate the chemopreventive activity determining the cytotoxic effect and the capability of producing DNA damage due to internucleosomal fragmentation or DNA strand breaks. Furthermore, the methylated status of these cancer cells treated with the tested compounds was assayed as a cancer therapy. Our results demonstrated that all tested compounds were neither toxic nor genotoxic, whereas these compounds resulted in antigenotoxic and cytotoxic substances, except for cyclamate. Aspartame showed antitoxic effects in Drosophila. All tested compounds decreased the quality of life of this in vivo organism model. Acesulfame-k, aspartame, and cyclamate induced DNA damage in the HL-60 cell line in the comet assay, and acesulfame-k generally increased the methylation status. In conclusion, all tested artificial sweeteners were safe compounds at assayed concentrations since toxicity and genotoxicity were not significantly induced in flies. Moreover, Aspartame and Cyclamate showed protective activity against a genotoxin in Drosophila Regarding nutraceutical potential, acesulfame-k and aspartame could be demonstrated to be chemopreventive due to the cytotoxicity activity shown by these compounds. According to DNA fragmentation and comet assays, a necrotic way could be the main mechanism of death cells induced by acesulfame-k and aspartame. Finally, Acesulfame-K hypermethylated repetitive elements, which are hypomethylated in cancer cells resulting in a benefit to humans

In Vivo and in Vitro Genotoxic and Epigenetic Effects of Two Types of Cola Beverages and Caffeine: A Multiassay Approach

The aim of this work was to assess the biological and food safety of two different beverages: Classic Coca Cola™ (CCC) and Caffeine-Free Coca Cola (CFCC). To this end, we determined the genotoxicological and biological effects of different doses of lyophilised CCC and CFCC and Caffeine (CAF), the main distinctive constituent. Their toxic/antitoxic, genotoxic/antigenotoxic, and chronic toxicity (lifespan assay) effects were determined in vivo using the Drosophila model. Their cytotoxic activities were determined using the HL-60 in vitro cancer model. In addition, clastogenic DNA toxicity was measured using internucleosomal fragmentation and SCGE assays. Their epigenetic effects were assessed on the HL-60 methylation status using some repetitive elements. The experimental results showed a slight chemopreventive effect of the two cola beverages against HL-60 leukaemia cells, probably mediated by nonapoptotic mechanisms. Finally, CCC and CAF induced a global genome hypomethylation evaluated in LINE-1 and Alu M1 repetitive elements. Overall, we demonstrated for the first time the safety of this famous beverage in in vivo and in vitro models.Instituto de Genética Veterinari