Natural Alkaloid Coptisine, Isolated from Coptis Chinensis, Inhibits Fungal Growth by Disrupting Membranes and Triggering Apoptosis

Objective: To evaluate the antifungal activity of coptisine, isolated from Coptis chinensis, against Cryptococcus neoformans both in vitro and in vivo, along with investigating its underlying mechanism. Methods: Microbroth dilution assay and disk diffusion assay were used to evaluate the antifungal activity of coptisine in vitro. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) was performed to observe the changes induced by coptisine in the morphology and structural integrity of C. neoformans cells. Flow cytometry was utilized for the analysis of cell apoptosis. To examine if apoptosis induced by coptisine includes initial mitochondrial impairment, changes in Δψm were assessed using flow cytometry with Rhodamine-123. Additionally, reactive oxygen species levels were identified through both confocal microscopy and flow cytometry. Furthermore, the therapeutic efficacy of coptisine in vivo was evaluated employing a murine model of C. neoformans infection induced by cyclophosphamide. Results: Coptisine has demonstrated significant antifungal efficacy against C. neoformans, exhibiting a half minimum inhibitory concentration (MIC50) of 61.64 µg/mL. SEM and TEM has unveiled that coptisine provokes significant alterations in morphology and structural integrity of C. neoformans cells, such as the formation of extensive cellular debris, compromised cell walls, and membrane distortions. Experimental data further elucidate that coptisine disrupts the cell membrane, encompassing the cytoplasmic membrane, culminating in the accumulation of reactive oxygen species, ultimately triggering cell apoptosis. Further, coptisine significantly improved the survival of infected mice, and reduced the damage caused by C. neoformans to the visceral tissues of infected mice. Conclusions: This study underscores the promising potential of coptisine, a natural alkaloid, as a prospective candidate for the development of an antifungal agent.