Bioaccesibilidad de cianotoxinas en mejillones mediante un modelo de digestión in vitro

Las cianobacterias son un diverso grupo de microorganismos procariotas cuya proliferación en masa da lugar a floraciones en las que se liberan sus metabolitos secundarios, entre los que destacamos las cianotoxinas, como las Microcistinas (MCs) y la Cilindrospermopsina (CYN), afectando así a la población expuesta, generando diferentes efectos tóxicos, además de crear problemas de índole tanto medioambiental, como económico.Universidad de Sevilla. Grado en Farmaci

In vitro assessment of cyanotoxins bioaccessibility in raw and cooked mussels

The oral route by ingestion of water and food contaminated with cyanotoxins is the main route of exposure to these toxins. This study addresses for the first time the bioaccessibility of some of the most common Microcystins (MC-LR, MC-RR and MC-YR) and Cylindrospermopsin (CYN) simultaneously in raw and steamed mussels spiked at 250 ng/g fresh weight of each cyanotoxin, after an in vitro digestion, including the salivary (incubation with artificial saliva, 30s), gastric (with pepsin, 2h, pH 2), duodenal (with pancreatin and bile salts, 2h, pH 6.5) and colonic phases (with lactic-acid bacteria, 48h, pH 7.2). The results obtained suggest that the potential absorption of these cyanotoxins by consumption of contaminated mussels is lower than expected. After the total effect of cooking and digestion, the mean bioaccessibility levels recorded were 24.65% (CYN), 31.51% (MC-RR), 17.51% (MC-YR) and 13.20% (MC-LR). Moreover, toxins were transferred to the steaming waters at 3.77 ± 0.24 μg L−1 CYN, 2.29 ± 0.13 μg L−1 MC-LR, 6.60 ± 0.25 μg L−1 MC-RR and 3.83 ± 0.22 μg L−1 MC-YR. These bioaccessibility results should be considered for a more accurate risk assessment related to these cyanotoxins in mussels, including the fact that the steaming waters could also represent a risk after human consumption.Ministerio de Economía y Competitividad (AGL2015-64558-R, MINECO/FEDER, UE

Safety assessment of propyl-propane-thiosulfonate (PTSO): 90-days oral subchronic toxicity study in rats

Propyl-propane-thiosulfonate (PTSO) is one of the main organosulfur compounds present in Allium essentials oil. Different applications in the food sector have been proposed for PTSO, such as food and feed additive and as active packaging. However, the authorization of its use depends on its toxicity profile. Thus, as a part of its safety assessment, in this work a repeated dose 90-day oral toxicity study has been conducted for the first time in rats following the OECD guideline 408. PTSO was administered to groups of 10 male and 10 female rats at dose levels of 0, 14, 28, and 55 mg/kg/day. No clinical signs or mortality and no changes in body weight, food consumption and feed conversion efficiency were detected through the study. Moreover, no treatment-related changes in hematological and biochemical parameters were observed, for either sex or dose groups. The histopathology study performed revealed no differences in organ weights, and no morphological and histopathological changes were observed. Based on these results, the no-observed-adverse-effect level (NOAEL) of PTSO was judged to be ≥ 55 mg/kg/day for both sexes.Ministerio de Ciencia e Innovación (Project RTC-2017-6199-2)Junta de Andalucía (Project AT 2017-5323

Mutagenicity and genotoxicity assessment of a new biopreservative product rich in Enterocin AS-48

PostprintA biopreservative derived from the fermentation of a dairy byproduct by Enterococcus faecalis UGRA10 strains being developed. This product possesses a strong and wide antibacterial spectrum mainly due to the presence of Enterocin AS-48 in its composition. To assess its potential as food additive, the mutagenicicity and genotoxicity has been assayed by means of the bacterial reverse-mutation assay in Salmonella typhimurium TA97A, TA98, TA100, TA102, TA1535 strains (Ames test, OECD 471, 2020) and the micronucleus test (MN) (OECD 487, 2016) in L5178Y/Tk ± cells. The results in the Ames test after exposure to the byproduct (6.75–100 μg/plate) with absence and presence of the metabolic activation system from rat liver (S9 fraction), revealed not mutagenicity at the conditions tested. For the MN test, the exposition to five enterocin AS-48 concentrations (0.2–1 μg/μl) was tested in the absence and presence of S9 fraction, with no evidence of genotoxicity. Negative results in the mutagenicity and genotoxicity assays point out the good safety profile of the byproduct and support its use as additive. Further toxicological studies are required before its approval and commercial application.Ministerio de Economía y Competitividad RTC-2017-6199- 2Junta de Andalucía AT 2017–5323Postprin

Acute and subchronic 90-days toxicity assessment of propyl-propane-thiosulfinate (PTS) in rats

The organosulfur compounds (OSC) extracted from Allium spp. exhibit antibacterial, antifungal, and antioxidant properties. The agri-food industry is taking advantage of these properties by using them as natural feed and food additives. In the present work, an acute and a subchronic 90-days toxicity studies have been conducted for the first time to assess the safety of the OSC propyl-propane-thiosulfinate (PTS). Both studies were carried out following the Organization for Economic Co-operation and Development test guidelines (425 and 408, respectively). The acute study provided a maximum tolerated dose (MTD) of 175 mg/kg and the subchronic study established the Non Observed Adverse Effect Level (NOAEL) ≥ 55 mg/kg body weight (b.w.)/day in both sexes. In addition, the subchronic study performed on rats exposed to 14, 28 and 55 mg/kg b.w./day PTS, revealed no changes in any of the hematological parameters measured as well as no differences in body weight and water/food consumption. However, biochemical parameters were altered in some groups, although they were not biologically significant (Ca2+ in female rats, and the thyroids hormones T3 and T4 in rat males). Furthermore, the histopathological assessment evidenced no abnormality on the gastrointestinal, respiratory, lymphoid, urinary, circulatory, nervous, musculoskeletal, and reproductive systems.Ministerio de Ciencia e Innovación RTC-2017-6199-2Junta de Andalucía AT 2017–5323 and P18-TP-214

Genotoxicity Evaluation of Two Derived Products from Allium Extracts: s-propylmercaptocysteine and s-propyl Mercaptoglutathione

Propyl-propane-thiosulfonate (PTSO) is one of the main organosulfur compounds present in Allium essential oils with a widely documented biological activity. For this reason, it could be used as a food and feed additive in the agri-food industry. A genotoxicity evaluation of substances and their metabolites present in food is necessary to guarantee the consumer's health following the recommendations of the European Food Safety Authority (EFSA). To evaluate the toxicological profile of derivatives of PTSO, the cytotoxicity, an Ames test, a micronucleus test and the comet assay were performed. Results showed that non-cytotoxic effects were observed in Caco-2 exposed to s-propyl mercaptocysteine (CSSP) and s-propyl mercaptoglutathione (GSSP) (0–450 μM). The mutagenicity index remained in the range of 0.6–1.4 for both compounds, showing no mutagenic effects for the concentrations of 5000–312.5 μg GSSP/plate and 250–15.63 μg CSSP/plate. Moreover, the % binucleated cells with micronuclei were 1.3–2.2 and 1.6–2.7 for GSSP and GSSP, respectively. For comet assays there was no DNA-genotoxic or oxidative damage in a concentration range of 112.5–450 μM. Therefore, we can conclude that these compounds are not genotoxic at the conditions tested. These results support that the presence of CSSP and GSSP in the food/feed is not of concern, although further studies are needed to complete their safety profile.Junta de Andalucía AT17_5323_USE, P18-TP-2147Ministerio de Ciencia, Innovación y Universidades FPU2019-0124

Simultaneous determination of Allium compounds (Propyl propane thiosulfonate and thiosulfinate) in animal feed using UPLC-MS/MS

Propyl-propane-thiosulfonate (PTSO) and Propyl-propane-thiosulfinate (PTS) are organosulfur compounds used to supplement the diet of livestock because of their beneficial effects on feed palatability, their antibacterial, anti-inflammatory, and antimethanogenic activities. Besides, antibiotic residues in the environment can be reduced by using these natural bioactive compounds. The objective of this study was to optimize the extraction parameters for the analysis of PTSO and PTS in feed matrices by performing a solid-liquid extraction and quantification by Ultra performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS). Optimization was performed using the Response Surface Methodology on a Box–Behnken experimental design, optimizing the following parameters: solvent:sample ratios and evaporation temperature set for the rotary evaporator. The method was validated for 3 concentration levels for both PTSO (100, 500, 1000 ng g−1) and PTS (500, 1150, 2300 ng g−1). The highest recoveries of PTSO and PTS were obtained using 12.5 mL of 100% acetonitrile, stirring for 15 min, and an evaporation temperature of 20 °C. The validated method was further applied to detect and quantify these compounds in different feed matrices. In conclusion, this is the first study to simultaneously analyze PTSO and PTS at low concentrations, employing a sensitive technique such as UPLC-MS/MS.Ministerio de Ciencia, Innovación y Universidades (RTC-2017-6199-2)Junta de Andalucía (AT-17-5323

Risk Assessment and Environmental Consequences of the Use of the Allium-derived Compound Propyl-propane Thiosulfonate (PTSO) in Agrifood Applications

The organosulfur compound propyl-propane thiosulfonate (PTSO), mainly found in Allium cepa, has a promising use in the agrifood industry. To confirm its safety for livestock, consumers, and environment, toxicological assessment is needed. In this regard, endocrine-disrupting chemicals (EDCs) are in the spotlight of research. Therefore, as part of the risk assessment of PTSO, in the present work, an in vivo study was performed in mice exposed to PTSO to investigate its potential reproductive toxicity considering fertility, genetic and endocrine endpoints. Five-weeks-old CD1 mice (80 males, 80 females) were exposed for 11 or 16 weeks (males or females, respectively) to different doses of PTSO (0, 14, 28 and 55 mg PTSO/kg b.w./day; 20 animals per group and sex) through the food pellets. No clinical observations or mortality and no changes in absolute organ weights and relative organ weights/body weight or brain ratios occurred during the study. The estrous cycle did not undergo any significant toxicologically relevant change. Most of the sex hormones displayed normal values. Some alterations in the expression of some genes related to reproduction is only observed in females, but they do not appear to have consequences in the development of sex organs. Docking results showed the impossibility of stable binding to estrogen and androgen receptors. Considering all the results obtained, the safe profile of PTSO can be confirmed for different agrifood applications at the conditions assayed.Junta de Andalucía P18-TP-2147Ministerio de Ciencia, Innovación y Universidades FPU2019-0124

Scientific Dissemination Conference: Contributions of Toxicology in Substance Safety Research and for Society

Las “Jornadas de divulgación científica: Aportaciones de la Toxicología en la investigación de la seguridad de las sustancias y para la sociedad” se celebraron el 19 de mayo de 2023 en la Facultad de Farmacia de la Universidad de Sevilla, gracias a una Ayuda recibida por el VII Plan Propio de la Universidad de Sevilla (VII PPIT-US) para actividades de divulgación científica. El objetivo fue divulgar entre nuestros alumnos los conocimientos científicos por parte de personal docente e investigador altamente cualificado, y también dar a conocer a la sociedad en general el trabajo de investigación científica de alto nivel que se está realizando en la US. La actividad iba dirigida a los alumnos de las asignaturas de “Toxicología”, “Quimioinformática, Investigación e Historia de la Farmacia”, “Laboratorio de Farmacia” (del Grado en Farmacia, Grado en Óptica y Optometría, y Doble Grado en Farmacia y Óptica y Optometría), y las asignaturas de “Toxicología de las Drogas de Abuso” o “Introducción a las Ciencias Forenses” (Grado en Criminología). Durante las Jornadas, que duraron 5,5 horas, se impartieron un total de 6 comunicaciones orales con diferente temática: el grafeno, las cianotoxinas, los aditivos aliáceos en alimentación humana y animal, los estilbenos en los vinos, o los bioplaguicidas; todas ellas sintetizadas en unos trípticos divulgativos repartidos entre los asistentes para su divulgación en su entorno social. Asimismo, se realizaron 2 ponencias invitadas por parte de expertos en Toxicología Forense y Ambiental, respectivamente. La actividad contó con casi un total de 75 asistentes y los resultados de las encuestas mostraron en general un alto grado de satisfacción con las mismas, mostrando las Jornadas como una enriquecedora actividad de divulgación científica, por la variedad de temas tratados, los nuevos conocimientos adquiridos y el acercamiento a la investigación e interés suscitado entre los asistentes.The " Scientific dissemination conference: Contributions of Toxicology in substance safety research and for society" were held on May 19th, 2023 at the Faculty of Pharmacy of the University of Seville, thanks to a financial support received by the VII Plan Propio of the University of Seville (VII PPIT-US) for scientific dissemination activities. The objective was to disseminate scientific knowledge among our students by highly qualified teaching and scientific personnel, and also to inform society in general of the high-level scientific research work that is being carried out in the University of Seville. The activity was aimed at students of the subjects of "Toxicology", "Chemoinformatics, Research and History of Pharmacy", " Laboratory of Pharmacy" (of the Degree in Pharmacy, Degree in Optics and Optometry, and Double Degree in Pharmacy and Optics and Optometry), and the subjects of "Toxicology of the Drugs of Abuse" or "Introduction to Forensic Sciences" (Degree in Criminology). During the Conference, which lasted 5.5 hours, 6 oral communications were given on graphene, cyanotoxins, alliaceous additives in human and animal feed, stilbenes in wines, or biopesticides. All of them were synthesized in informative triptychs distributed among the attendees for their dissemination in their social environment. Likewise, 2 invited presentations were given by experts in Forensic and Environmental Toxicology, respectively. The activity had 75 attendees and the results of the surveys generally showed a high degree of satisfaction with them, showing the Conference as an enriching outreach activity, due to the variety of topics covered, the new knowledge acquired and the approach to research and interest aroused among attendees

In Vitro Toxicity Studies of Bioactive Organosulfur Compounds from Allium spp. with Potential Application in the Agri-Food Industry: A Review

Organosulfur compounds (OSCs) are secondary metabolites produced by different Allium species which present important biological activities such as antimicrobial, antioxidant, anti-inflammatory antidiabetic, anticarcinogenic, antispasmodic, etc. In recent years, their use has been promoted in the agri-food industry as a substitute for synthetic preservatives, increasing potential accumulative exposure to consumers. Before their application in the food industry, it is necessary to pass a safety assessment as specified by the European Food Safety Authority (EFSA). This work reviews the scientific literature on OSCs regarding their in vitro toxicity evaluation following PRISMA guidelines for systematic reviews. Four electronic research databases were searched (Web of Science, Scopus, Science Database and PubMed) and a total of 43 works were selected according to predeterminate inclusion and exclusion criteria. Different data items and the risk of bias for each study were included. Currently, there are very few in vitro studies focused on investigating the potential toxicity of OSCs. Most research studies aimed to evaluate the cytotoxicity of OSCs to elucidate their antiproliferative effects focusing on their therapeutic aspects using cancer cell lines as the main experimental model. The results showed that diallyl disulfide (DADS) is the compound most studied, followed by diallyl trisulfide (DATS), diallyl sulfide (DAS), Allicin and Ajoene. Only 4 studies have been performed specifically to explore the safety of OSCs for agri-food applications, and genotoxicity studies are limited. More toxicity studies of OSCs are necessary to ensure consumers safety and should mainly be focused on the evaluation of genotoxicity and long-term toxicity effects.Junta de Andalucía AT17_5323_USE and P18-TP-21