Neuroprotective effects of C-phycocyanin and allophycocyanin from Spirulina purified using a newly developed method: in vitro barrier permeability, antioxidant and anti-inflammatory activities

Phycobiliproteins, particularly C-phycocyanin (C-PC) and allophycocyanin (APC), are major bioactive pigments in Arthrospira platensis (Spirulina) with well-documented antioxidant properties, yet their individual neuroprotective potential and bioavailability remain poorly characterized. Here, we developed an efficient, potentially scalable, two-step ultrasound-based extraction followed by membrane chromatography to obtain analytical-grade purity C-PC and APC from fresh A. platensis biomass. Both purified phycobiliproteins, together with the crude extract (CE), demonstrated relevant cell viability up to 35 µg/mL in SH-SY5Y neuronal and BV2 microglial cells. In vitro assays revealed that APC exhibited significantly higher ferric-reducing antioxidant power (FRAP) than C-PC, while both phycobiliproteins effectively reduced intracellular ROS levels and protected SH-SY5Y cells from tert-butyl hydroperoxide-induced oxidative stress, with C-PC and APC showing the strongest cytoprotective effects. All preparations also significantly decreased LPS-induced nitric oxide production in BV2 microglia, indicating anti-inflammatory activity. Using Caco-2 monolayers, we demonstrated for the first time that both purified C-PC and APC exhibit time-dependent, low but moderate transcellular permeability, while preserving monolayer integrity. Fluorescence microscopy confirmed substantial intracellular uptake in SH-SY5Y neurons (punctate cytoplasmic pattern) and lower, yet detectable, internalization in BV2 microglia. These findings highlight distinct yet complementary bioactivities of highly purified C-PC and APC and provide the first evidence of their ability to cross physiological barriers in vitro, offering new mechanistic insights into their potential as neuroprotective nutraceuticals targeting oxidative and neuroinflammatory processes in the central nervous system.