[Development of allergenic potential's assessment approaches of protein hydrolysates obtained from alternative food...

The investigation of the perspectives for the application of alternative protein sources in food production has allowed us to identify two main directions requiring comprehensive scientific support: firstly, the development of production and processing of food protein technologies to obtain food hydrolysates with a given composition and properties, and secondly, the assessment of the such product safety, including evaluation of their allergenic potential. Currently, within the second direction, research is being conducted to improve approaches to assessing the allergenic potential of innovative food, search for new in vitro and in vivo models and biomarkers to guarantee the safety of these products for consumers. The purpose of the study was to research the allergenic potential of the methanotrophic bacteria protein hydrolysates on the model of systemic anaphylaxis in rats and to formulate approaches to assessing the novel foods' safety. Material and methods. The allergenic potential of acid and enzymatic hydrolysates of Methylococcus capsulatus methanotrophic bacteria was investigated on the systemic anaphylaxis' model in two 30-day experiments on Wistar rats' male fed diet containing acid hydrolysate in the 1st experiment and enzymatic hydrolysate in the 2nd experiment. Animals of the corresponding control groups were kept on semi-synthetic casein diet. The ingredients of the experimental groups' diets were substituted taking into account the content of protein, fat, carbohydrates and dietary fibers in the hydrolysates in compliance with the principle of isocaloric content and identity of chemical composition. Allergenic potential of hydrolysates was assessed using an expanded pool of indicators: severity of active anaphylactic shock (mortality, number of severe anaphylaxis reactions); cytokine profile (content of pro-inflammatory and anti-inflammatory cytokines, as well as regulators of cellular and humoral immune response), ovalbumin specific IgG immunoglobulin level before administration of ovalbumi permissive dose. Results. In the sensitised rats the inclusion of methanotrophic bacteria hydrolysates in the diet caused an increase in allergic reactivity. The systemic anaphylaxis severity was 20% (p<0.05) higher in rats of the 1st experimental group receiving acid hydrolysate with the diet than in animals of the 1st control group; in rats of the 2nd experimental group it was 33% (p<0.05) lower than the corresponding control. The assessment of the humoral immune response intensity shows a slight statistically unreliable increase by 11% (p>0.05) in the sum of IgG1 + IgG4 antibodies in rats of the 1st experimental group compared to the corresponding control group. This indicator in rats of the 2nd experimental group was 81% (p<0.05) higher than in rats of the 2nd control group. The study of serum cytokine profiles revealed an increase in Th1/Th2-regulators of the immune response in rats of both experimental groups compared to the corresponding controls: in rats of the 1st experimental group 8 out of 12 indices exceeded control values by 4-69% (p>0.05), in rats of the 2nd experimental group 7 out of 12 indices exceeded control values by 5-19% (p>0.05), and 2 out of 12 - by 88-191% (p<0.05). The experimental data processed by nonparametric ranked two-factor dispersion analysis confirmed the influence of the methanotrophic bacteria protein hydrolysis' technology on the hydrolysates allergenicity and the influence of the diet protein source. Conclusion. The results of the allergenic potential' investigation of Methylococcus capsulatus protein hydrolysates in the systemic anaphylaxis rats' model indicate the promising use of enzymatic hydrolysis. However, further optimisation of the technology is required to create a product with low residual antigenicity and immunogenicity. Approaches to allergenic potential assessment of innovative food obtained by microbial synthesis, including a number of in vitro and in vivo studies, were formed on the basis of the obtained data analysis.