RESUMEN
Omega-3 (ω3) fatty acids are a family of polyunsaturated fats. Two of the ω3 long-chain polyunsaturated fatty acids (LC-PUFA), eicosapentaenoic acid (EPA, ω3, 20:5Δ5,8,11,14,17) and docosahexaenoic acid (DHA, ω3, 22:6Δ4,7,10,13,16,19) are sourced primarily from fish. Higher consumption, limited fishing quotas and other environmental factors (e.g., heavy metals) have warranted a need for alternative sources. Nuseed offers a genetically engineered canola (Brassica napus) event,1 DHA canola (OECD Unique Identifier NS-B5ØØ27-4), which has been modified to introduce a pathway for production of the ω3 LC-PUFAs DHA and EPA from oleic acid (OA) in the seed oil. To accomplish this, genes were sourced from marine microalgae and common yeast then incorporated into canola to produce DHA canola, one of the first land-based production systems for ω 3 PUFAs. Safety was evaluated in part by conducting a repeated dose 28-day toxicity study and a dietary 13-week toxicity study using CD® IGS [Crl:CD(SD)] rats. In the 28-day study, conventional and DHA canola oil were administered orally (via gavage); no treatment-related adverse effects were observed. The 13-week toxicity study was subsequently conducted where DHA canola oil and meal were administered by dietary admixture. No adverse effects were noted in clinical observations, clinical pathology, or histopathology. These studies support the food and feed safety of DHA canola oil and meal.
Asunto(s)
Ácidos Docosahexaenoicos/administración & dosificación , Aceite de Brassica napus/administración & dosificación , Animales , Peso Corporal , Brassica napus , Ácidos Docosahexaenoicos/efectos adversos , Femenino , Masculino , Microalgas/genética , Distribución Aleatoria , Aceite de Brassica napus/efectos adversos , RatasRESUMEN
Soybean (Glycine max) seed contain some proteins that are allergenic to humans and animals. However, the concentration of these allergens and their expression variability among germplasms is presently unknown. To address this problem, 10 allergens were quantified from 20 nongenetically modified commercial soybean varieties using parallel, label-free mass spectrometry approaches. Relative quantitation was performed by spectral counting and absolute quantitation was performed using multiple reaction monitoring (MRM) with synthetic, isotope-labeled peptides as internal standards. During relative quantitation analysis, 10 target allergens were identified, and five of these allergens showed expression levels higher than technical variation observed for bovine serum albumin (BSA) internal standard (â¼11%), suggesting expression differences among the varieties. To confirm this observation, absolute quantitation of these allergens from each variety was performed using MRM. Eight of the 10 allergens were quantified for their concentration in seed and ranged from approximately 0.5 to 5.7 µg/mg of soy protein. MRM analysis reduced technical variance of BSA internal standards to approximately 7%, and confirmed differential expression for four allergens across the 20 varieties. This is the first quantitative assessment of all major soybean allergens. The results show the total quantity of allergens measured among the 20 soy varieties was mostly similar.
Asunto(s)
Alérgenos/análisis , Proteómica/métodos , Proteínas de Soja/análisis , Espectrometría de Masas en Tándem/métodos , Alérgenos/química , Alérgenos/metabolismo , Animales , Bovinos , Fragmentos de Péptidos/análisis , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Reproducibilidad de los Resultados , Albúmina Sérica Bovina , Proteínas de Soja/química , Proteínas de Soja/metabolismo , Glycine max/química , Tripsina/metabolismoRESUMEN
DHA canola, a genetically engineered Brassica napus (OECD Unique Identifier NS-B5ØØ27-4), has been developed as one of the first land-based production systems for omega-3 long-chain polyunsaturated fatty acids (LCPUFA), whose health benefits are well-established. Yet, the marine sources of these nutrients are under high pressures due to over-fishing and increasing demand. DHA canola is a plant-based source for these essential fatty acids that produces a high level of docosahexaenoic acid (DHA). This terrestrial system allows for sustainable, scalable and stable production of omega-3 LCPUFA that addresses not only the increasing market demand, but also the complex interplay of agriculture, aquaculture, and human nutrition. The vector used to produce the desired oil profile in DHA canola contains the expression cassettes of seven genes in the DHA biosynthesis pathway and was specifically designed to convert oleic acid to DHA in canola seed. The characterization and safety evaluation of food and feed produced from DHA canola are described and supported by a detailed nutritional analysis of the seed, meal, and oil. Aside from the intended changes of the fatty acid profile, none of the other compositional analytes showed biologically meaningful differences when compared to conventional canola varieties. In addition, the meal from DHA canola is compositionally equivalent to conventional canola meal. Further evidence of nutritional value and safety of DHA canola oil have been confirmed in fish feeding studies. Given that most human populations lack sufficient daily intakes of omega-3 LCPUFA, a dietary exposure assessment is also included. In conclusion, the results from these studies demonstrate it is safe to use products derived from DHA canola in human foods, nutraceuticals, or animal feeds.
RESUMEN
The measurement of protein digestibility is one of the key steps in determining the safety of a genetically modified crop that has been traditionally accomplished using antibodies. Membrane proteins are often extremely difficult to express with replicated authentic tertiary structure in heterologous systems. As a result raising antibodies for use in safety assessment may not be feasible. In this study, LC-MS based proteomics was used to characterise seven transmembrane enzymes from the docosahexaenoic acid biosynthetic pathway that had been introduced into canola. The application of a two-stage digestion strategy involving simulated gastric fluid followed by trypsin enabled the measurement of protein digestibility in vitro. Tryptic peptide markers that spanned the length of each desaturase protein were monitored and showed that these proteins were readily degraded (>95% within 5â¯min) and highlighted regions of the elongase enzymes that showed limited resistance to simulated gastric digestion. Traditional gel-based and Western blotting analysis of ω3-desaturase and Δ6-elongase revealed rapid hydrolysis of the intact proteins within seconds and no fragments (>3â¯kDa) remained after 60â¯min, complementing the novel approach described herein. The LC-MS approach was sensitive, selective and did not require the use of purified proteins.
Asunto(s)
Ácidos Docosahexaenoicos/biosíntesis , Enzimas/metabolismo , Proteómica/métodos , Secuencia de Aminoácidos , Cromatografía Liquida/métodos , Espectrometría de Masas , Reproducibilidad de los ResultadosRESUMEN
The International Life Science Institute's Health and Environmental Sciences Institute's Protein Allergenicity Technical Committee hosted an international workshop October 23-25, 2007, in Nice, France, to review and discuss existing and emerging methods and techniques for improving the current weight-of-evidence approach for evaluating the potential allergenicity of novel proteins. The workshop included over 40 international experts from government, industry, and academia. Their expertise represented a range of disciplines including immunology, chemistry, molecular biology, bioinformatics, and toxicology. Among participants, there was consensus that (1) current bioinformatic approaches are highly conservative; (2) advances in bioinformatics using structural comparisons of proteins may be helpful as the availability of structural data increases; (3) proteomics may prove useful for monitoring the natural variability in a plant's proteome and assessing the impact of biotechnology transformations on endogenous levels of allergens, but only when analytical techniques have been standardized and additional data are available on the natural variation of protein expression in non-transgenic bred plants; (4) basophil response assays are promising techniques, but need additional evaluation around specificity, sensitivity, and reproducibility; (5) additional research is required to develop and validate an animal model for the purpose of predicting protein allergenicity.
Asunto(s)
Alérgenos/toxicidad , Proteínas en la Dieta/toxicidad , Hipersensibilidad a los Alimentos/diagnóstico , Alérgenos/química , Animales , Basófilos/inmunología , Biotecnología , Biología Computacional , Modelos Animales de Enfermedad , HumanosRESUMEN
One component of the safety assessment of agricultural products produced through biotechnology is evaluation of the safety of newly expressed proteins. The ILSI International Food Biotechnology Committee has developed a scientifically based two-tiered, weight-of-evidence strategy to assess the safety of novel proteins used in the context of agricultural biotechnology. Recommendations draw upon knowledge of the biological and chemical characteristics of proteins and testing methods for evaluating potential intrinsic hazards of chemicals. Tier I (potential hazard identification) includes an assessment of the biological function or mode of action and intended application of the protein, history of safe use, comparison of the amino acid sequence of the protein to other proteins, as well as the biochemical and physico-chemical properties of the proteins. Studies outlined in Tier II (hazard characterization) are conducted when the results from Tier I are not sufficient to allow a determination of safety (reasonable certainty of no harm) on a case-by-case basis. These studies may include acute and repeated dose toxicology studies and hypothesis-based testing. The application of these guidelines is presented using examples of transgenic proteins applied for agricultural input and output traits in genetically modified crops along with recommendations for future research considerations related to protein safety assessment.
Asunto(s)
Seguridad de Productos para el Consumidor , Alimentos Modificados Genéticamente/efectos adversos , Proteínas de Plantas/efectos adversos , Plantas Modificadas Genéticamente/efectos adversos , Animales , Biotecnología , Productos Agrícolas/genética , Productos Agrícolas/normas , Hipersensibilidad a los Alimentos/prevención & control , Tecnología de Alimentos , Humanos , Modelos Animales , Nivel sin Efectos Adversos Observados , Valor Nutritivo , Proteínas de Plantas/química , Proteínas de Plantas/inmunología , Medición de Riesgo , Gestión de Riesgos , Pruebas de Toxicidad , Estados UnidosRESUMEN
BACKGROUND: Growers have enthusiastically embraced crops genetically modified to express Bacillus thuringiensis (Bt) proteins for insect control because they provide excellent protection from key damaging insect pests around the world. Bt crops also offer superior environmental and health benefits while increasing grower income. However, insect resistance development is an important concern for all stakeholders, including growers, technology providers and seed companies that develop these genetically modified crops. Given the marked benefits associated with Bt crops, insect resistance management (IRM) must be a consideration when cultivating these crops. RESULTS: The technical data and practical experience accumulated with Bt crops in many global regions can inform different aspects of resistance management leading to robust, science-based IRM plans. A range of elements should be considered in assembling any IRM strategy, including: pest biology/ecology, product deployment patterns, local cropping systems, insect susceptibility monitoring, stakeholder/grower communications, and a remedial action plan should resistance develop. Each of these elements is described in more detail, with specific examples of how these elements can be combined and tailored to the local/regional environments and grower practices. CONCLUSION: IRM plans need to be suitable for the given production situation. What works for large monoculture production systems in North America is unlikely to be appropriate for the small, more diverse agriculture of southeast Asia or Africa. Though it is clear that Bt crops impart considerable value to growers, it is also clear that it is in the best interest of all stakeholders to preserve Bt proteins for the long-term benefits they provide.