Phenylpiperazine/dithioates as novel H2S donors: design, synthesis, H2S release, formulation as PEG-niosomes and antifungal activity against Candida albicans.

A series of novel phenylpiperazine-based carbodithioate derivatives (3a-j) were designed, synthesized, formulated as PEG-Niosomes and evaluated for their potential hydrogen sulphide (H2S) donating properties and their antifungal activities. Structural modifications were inspired by the antifungal drug tolnaftate, incorporating a dithiocarbamate moiety to enhance biological activity and target engagement. H2S-releasing capacity was quantified via the methylene blue assay. Compound 3j (S-phenacyl p-chloro derivative) exhibiting the highest release (89.71%), followed by compounds 3a and 3i. Antifungal efficacy against Candida albicans was determined using the microbroth dilution method, where compounds 3a, 3b, 3f, 3i and 3j demonstrated potent inhibitory activity (MIC = 4.68 μg/mL), exceeding that of fluconazole. Further investigations revealed that these compounds also effectively suppressed hyphal formation and biofilm development, two major virulence factors of C. albicans. Notably, compound 3j exhibited superior performance across all assays. In a preliminary safety evaluation, compound 3j displayed low cytotoxicity toward mammalian fibroblasts (WS-1) (IC₅₀ = 65.73 ± 4.20 μg/mL) compared to doxorubicin (IC₅₀ = 26.72 ± 2.20 μg/mL), providing an 14-fold selectivity index relative to its antifungal MIC (4.68 μg/mL). Molecular docking studies corroborated these findings by showing favorable binding of compound 3j to the lanosterol 14α-demethylase enzyme (CYP51), highlighting π-sulfur and hydrophobic interactions critical for binding affinity. Additionally, compound 3j directly inhibited CYP51 in vitro (IC₅₀ = 0.331 ± 0.025 μM), showing stronger inhibition than the reference azole fluconazole (IC₅₀ = 1.388 ± 0.099 μM. H₂S release was also verified by scavenger reversal. In the presence of MgO (1.5 mg/mL), the antifungal activity of compound 3j decreased, with the MIC rising to 32 μg/mL. These results underscore the potential of phenylpiperazine dithioates, particularly compound 3j, as promising antifungal agent with multi-target activity. Additionally, the PEG-Niosomes of compound 3j represent a promising delivery system for enhancing the dissolution and potential bioavailability of compound 3j.