Evaluation of the Performance of Nanostructured Proteins from Micro-Nematodes Extracted from Dairy Silage Fermentation Waste on the Synthesis of Novel Bio-Polypeptides and Their Applications in PharmaFood

In recent years, there has been a growing interest in sustainable and bio-based resources for the production of food and pharmaceutical materials under the PharmaFood paradigm. This study investigates the performance of nanostructured proteins extracted from micro-nematodes (Microbacterium nematodis) isolated from dairy silage fermentation waste in the synthesis of novel bio-polypeptides. Dairy silage fermentation residues represent a rich source of organic matter and beneficial microorganisms, offering high potential for the extraction of innovative biomacromolecules. Micro-nematodes, as efficient biological models, are capable of degrading these wastes and producing nanostructured proteins with particle sizes ranging from ۱۰ to ۵۰ nm, exhibiting unique catalytic properties. The primary objective of this research was to evaluate the efficacy of these proteins in synthesizing bio-polypeptides with novel structures, such as peptide chains featuring enhanced disulfide bonds, and to explore their potential applications in enhancing the nutritional and therapeutic properties of dairy products and PharmaFood formulations. The research methodology encompassed extraction, purification, and characterization of nanostructured proteins from nematodes cultured in silage waste media. Nematodes were cultivated under controlled conditions (temperature ۲۵°C, pH ۶.۵), and proteins were isolated using centrifugation and chromatography techniques. The extracted proteins were characterized via dynamic light scattering (DLS), transmission electron microscopy (TEM), and Fourier-transform infrared spectroscopy (FTIR) to confirm nanostructural integrity and functional groups. For polypeptide synthesis, solid-phase peptide synthesis (SPPS) was adapted with protein-mediated catalysis, incorporating amino acid precursors such as L-cysteine, L-arginine, and L-glutamine at molar ratios of ۱:۲:۱. Reaction kinetics were monitored over ۴۸ hours at ۳۷°C in phosphate-buffered saline (PBS) at pH ۷.۴. Synthesized polypeptides were purified by high-performance liquid chromatography (HPLC) and analyzed for molecular weight distribution using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Results demonstrated that nanostructured proteins accelerated polypeptide synthesis by ۳.۲-fold compared to conventional enzymatic methods, yielding polypeptides with average chain lengths of ۲۵-۳۵ residues and molecular weights between ۲.۸-۴.۱ kDa. Structural analysis revealed α-helical dominance (۶۲%) with stable disulfide linkages, enhancing thermal stability up to ۸۵°C. In vitro bioactivity assays showed significant antioxidant capacity (DPPH radical scavenging: ۷۸.۴ ± ۳.۱%) and immunomodulatory effects (IL-۶ suppression: ۴۱.۲% in macrophage models). Application trials in model dairy matrices (yogurt and cheese analogs) indicated improved shelf-life extension by ۱۸ days and enhanced probiotic viability under simulated gastric conditions. The integration of these bio-polypeptides into PharmaFood systems exhibited dual functionality: nutritional enrichment with essential amino acid profiles and therapeutic modulation via anti-inflammatory pathways. Stability under pasteurization (۷۲°C, ۱۵ s) was retained at ۹۴%, supporting industrial scalability. Comparative analysis with synthetic peptides highlighted superior biocompatibility and lower immunogenicity. This work establishes a circular bioeconomy model wherein dairy waste is valorized into high-value biomolecular catalysts for next-generation functional foods. The findings underscore the transformative potential of nematode-derived nanostructures in sustainable biomanufacturing, paving the way for eco-efficient PharmaFood innovations with minimal environmental footprint. Future directions include in vivo efficacy studies and pilot-scale production optimization to bridge laboratory-scale synthesis with commercial viability.

کلیدواژه ها:

Nano-biosensor ، Zinc oxide nanoparticles ، Peroxidase enzyme ، Minced red meat ، Smart packaging ، Microbial control ، Lipid oxidation ، Colorimetric indicator ، Refrigerated storage ، Shelf life extension ، Food safety ، Sensory quality