Spray-dried microencapsulation as a strategy for stabilizing chlorophyll-rich extracts: pigment profiling, digestive fate, and in vitro biological activity with and without Cu2+ incorporation.

The limited stability and digestive functionality of chlorophyll-rich systems remain major challenges in the development of functional green powders. This study aimed to investigate the stabilization, transformation, and biological functionality of chlorophyll-rich extracts microencapsulated using spray drying and freeze drying, with particular emphasis on carrier composition and copper-mediated pigment modification. Native and Cu2+-treated chlorophyll-rich extracts were encapsulated using inulin, soy protein isolate (SPI), and their binary system. Unlike previous studies focused primarily on storage stability, the present work simultaneously evaluated pigment transformations, digestive behavior, antioxidant capacity, and dipeptidyl peptidase-4 (DPP-4) inhibitory activity in native and Cu-modified chlorophyll systems. Carrier composition exerted a stronger influence than drying technique on pigment stability and functional properties. The highest total chlorophyll content was observed for powders obtained from the chlorophyll-rich extract by spray drying with SPI (247.39 mg 100 g-1 dry weight (dw)), whereas the formulation based on copper-modified chlorophyll-rich extract (Cu-ChRE) produced by freeze drying with inulin contained more than eight-fold lower chlorophyll levels (29.70 mg 100 g-1 dw). SPI-based systems generally promoted higher chlorophyll retention, antioxidant activity, and DPP-4 inhibitory activity, while inulin-based matrices more effectively limited secondary photooxidative transformations during storage. In the initial powders, ChRE-based formulations exhibited lower IC₅₀ values than Cu-ChRE systems (0.84-1.12 vs. 1.45-1.96 mg mL-1), indicating stronger enzyme inhibition by native pigment systems. Copper treatment improved pigment stability and reduced pheophytin formation; however, Cu-modified systems exhibited lower antioxidant and antidiabetic activity than native chlorophyll-rich extracts. Simulated gastrointestinal digestion substantially altered pigment composition and biological responses, partially restoring antioxidant capacity and enhancing DPP-4 inhibitory activity in selected formulations. The results demonstrate that stabilization strategies effective during storage do not necessarily ensure enhanced post-digestive functionality. Overall, spray-drying-based microencapsulation represents a promising alternative to Cu2+-based stabilization for the development of functional chlorophyll-rich powders with potential application in functional food systems.