EVALUATION OF NOVEL PROTEIN-BASED INGREDIENTS DERVIED FROM INDUSTRIAL HEMPSEED (Cannabis sativa L.)

Industrial hempseed (Cannabis sativa L.) is gaining recognition as a sustainable plant-based protein source, yet its allergenic potential remains largely unexplored. Similarly, the impact of consuming hemp ingredients long term has not been assessed. Thus, this study aimed to investigate the physicochemical properties and immunogenicity of hemp protein isolate (HPI) extracted at pH 10 and 12, with and without 30-minute enzymatic hydrolysis using bromelain, Alcalase, and trypsin. Furthermore, HPI-10, HPI-12 and hemp meal derived at pH 10 (HM-10) were used as supplementary sources of protein (in place of casein in the AIN-93G diet) in a 90-day feeding murine model to investigate the impact of prolonged hemp-protein consumption on growth parameters, blood chemistry, hematological, and histopathological changes.

Results found that enzymatic hydrolysis significantly altered the protein structure of HPI-10, redistributing the molecular weight via an increase in low molecular weight peptides (< 10 kDa), significantly increasing the degree of hydrolysis for all 3 hydrolysates (p < 0.05), and disrupting native folded tertiary conformations, as assessed by SDS-PAGE, degree of hydrolysis via Orthophtaldialdehyde, and fluorescence spectroscopy, respectively. Immunogenicity was evaluated through Enzyme-linked Immunosorbent Assay (ELISA) and Western blot using human allergic IgE antibodies corresponding to the "Big 9" food allergens, as well as hempseed. Results demonstrated that hydrolysis for both HPI-10 and HPI-12 effectively minimized the qualitative immunogenic responses with hemp-allergic plasma while revealing potential immunogenic interactions with unhydrolyzed HPI-10 and wheat and soy allergic individuals. Results from the long-term feeding study outlined no significant differences between the control diet and the three supplementary proteins in histopathological, hematological, and growth parameters (p < 0.05), outlining the safety for hemp protein supplementation in diets. These findings suggest that both enzymatically hydrolyzed HPI and HM may serve as a hypoallergenic, nutritionally safe, high protein ingredients, warranting further investigation into their application in functional food formulations.