Protein Bioaccessibility in Meat-Free Deli Ham Enriched with Electrospun Spirulina Biomass

The growing demand for meat analogues has stimulated the search for alternative protein sources and processing strategies capable of improving their nutritional and technological properties. Spirulina (Limnospira spp; SP) is a source of protein, while the residual biomass obtained after C-phycocyanin extraction (Spirupower®; SPW) also represents a potential protein ingredient for food applications within a circular production approach. However, protein enrichment does not necessarily ensure nutritional accessibility, as protein release during gastrointestinal digestion is strongly influenced by processing and food-matrix interactions. Therefore, this study aimed to evaluate the in vitro protein bioaccessibility of meat-free deli ham analogues containing electrospun zein/polyethylene oxide (PEO) composites (Bioinicia, Spain, 24 kV, 10 mm tip-to-collector distance) loaded with SP or SPW. The meat-free matrix was formulated with water/ice, sunflower oil, egg white, potato starch, psyllium, and seasonings. SP and SPW were incorporated at 5.77% through electrospun microfibers containing 3.46% zein and 0.28% PEO. A formulation without microfibers (C) and a zein-rich microfiber formulation (ZC) were used as controls. Protein content was determined by the Dumas method, and simulated gastrointestinal digestion was performed according to the INFOGEST protocol. Protein bioaccessibility was calculated from the initial protein content and the protein remaining after digestion. SP and SPW significantly increased (p < 0.05) protein content from 10.74 ± 0.36% in C to 16.12 ± 0.48% and 16.30 ± 0.43%, respectively, corresponding to approximately 50% protein enrichment. All formulations exhibited high protein bioaccessibility (>89%); however, SPW reached 95.01 ± 0.34%, significantly exceeding C (90.16 ± 0.93%), ZC (89.78 ± 1.06%), and SP (89.24 ± 0.72%). Interestingly, ZC exhibited the highest protein content (19.93 ± 0.12%) without a corresponding increase in bioaccessibility. Since SP and SPW were incorporated at the same concentration within identical zein/PEO systems, the superior performance of SPW suggests that compositional and/or structural modifications associated with C-phycocyanin extraction may have favored protein release. Overall, electrospun SPW-containing microfibers enabled substantial protein enrichment while maintaining high protein bioaccessibility, supporting the valorization of residual cyanobacterial biomass in structured meat analogues.

Acknowledgments: This study was financed, in part, by the São Paulo Research Foundation (FAPESP), Brazil, process Number #2024/19218-0; 2023/00857-0. This study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES) - Finance Code 001, process number 88887.917438/2023-00; and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) grant number 405238/2025-8, and A.R.C.B (PQ 305518/2024-0) was a recipient of a research productivity fellowship from CNPq. Agenda VIIAFOOD PRR - Plataforma de Valorização, Industrialização e Inovação comercial para o AgroAlimentar (C644929456-00000040) – project AltProtein (SONAE); FCT (strategic project UID/AGR/04129/2020 (LEAF)).