Squillante, Jonathan (2025) Biosynthesis and functional assessment of bioactive compounds derived from agri-food by-products fermentation. [Tesi di dottorato]

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Abstract

The agri-food industry generates a significant amount of by-products, which require appropriate disposal, resulting in both higher costs for industry and environmental impact. However, several by-products have shown great functional and technological properties, motivating researchers and industries to reuse or valorize them. Nevertheless, by-products widely differ for chemo-physical composition, determining their “valorization power” as well as the potential output. They can be employed for the industrial production of (bio)gases, (bio)ethanol, (bio)diesel, compost and fertilizers, (bio)absorbents, organic acids, enzymes, (bio)materials, and bioactive compounds. For this purpose, several singular or combinate biological, chemical, and physical approaches are used based on the final new product. Among them, fermentation has gained great interest over the years for its relative practicality on an industrial scale, low cost, and variety of applications. It is one of the oldest technologies that uses the metabolic activity of microorganisms for the transformation of complex substrates. The modern fermentation distinguishes two techniques: submerged fermentation (SmF) and solid-state fermentation (SsF). They stand out mainly for the amount of water that is high in the former and low in the latter. SmF takes place in a liquid medium, favoring a higher homogeneity of the nutrients dissolved and easier parameter control. On the other hand, SsF requires a low amount of water, reducing the production cost and mimicking the natural environment of most microorganisms. They also differ in output yields and post-fermentation processes that depend on the substrate and microorganism. Among the most industrially valuable outputs through both techniques, bioactive compounds have attracted growing attention, especially for their human health-promoting effects. They refer to a wide class of compounds with biological activity and can be released by substrate or synthesized ex novo as secondary metabolites through microbial metabolism. Bioactive compounds are highly sought after in pharmaceutical, nutraceutical, cosmetic, and food industries, depending on their key functionality. Specifically, compounds that support overall well-being arouse great interest due to growing awareness and, consequently, increasing market demand. These compounds contribute to maintaining physiological balance and preventing chronic conditions through a wide range of biological effects, including antioxidant, anti-inflammatory, antimicrobial, antihypertensive, immunomodulatory, hypocholesterolemic, hypoglycemic, neuroprotective, and prebiotic activity. Delving deeper, bioactive compounds with cardioprotective effects are of great relevance in preventive medicine and public health given the heavy and growing burden of cardiovascular diseases (CVDs). Aims and outline The overall aim of the present research was to explore the application of agri-food by-products to produce bioactive compounds through fermentation and to evaluate the resulting biological activity. Therefore, this investigation sought to accomplish of three main goals: i) screening of fermentable by-products and microorganisms producing target compounds; ii) optimization of the bioactive compound biosynthesis via fermentation and analysis of the phenolic profiling of the final product; iii) evaluation of bioactivity of bioactive compound-enriched products. These three key points are addressed in this thesis, following the outline: Chapter 1: A state-of-the-art review of the production of bioactive compounds through by-products fermentation is provided with special focus on monacolin K, peptides, gamma-aminobutyric acid (GABA), phenolic compounds, coenzyme Q10, nattokinase, and exopolysaccharides involved in CVDs prevention. Chapter 2: Technological properties of by-products from various food supply chains were assessed, and microbial screening for monacolin K and GABA-producer microorganism in SsF was performed. Chapter 3: Optimization of monacolins K from SsF of tomato pomace with Monascus ruber, modulating different environmental parameters Under optimal conditions, phenolic profiling of the fermented tomato pomace was characterized and compared with the unfermented tomato pomace Chapter 4: The bioactivity and cytotoxicity of Monascus-fermented tomato pomace extract were assessed through chemical, enzyme, and cellular-based assays to reveal the effects of fermentation on the functional properties of the by-product Chapter 5: GABA synthesis from SsF of tomato pomace using Levilactobacillus brevis was investigated and then optimized by a Response Surface Methodology (RSM). Under the optimal conditions for GABA production, phenolic profiling of the fermented tomato pomace was characterized and compared with the unfermented tomato pomace Chapter 6: The bioactivity and cytotoxicity of Levilactobacillus-fermented tomato pomace extract were assessed through chemical, enzyme, and cellular-based assays, to reveal the effects of fermentation on the functional properties of the by-product Chapter 7: A comprehensive conclusion summarizing the key findings and implications of this research is provided.

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