β-Cyclodextrin Inclusion Complexes of 20-Hydroxyecdysone Derivatives: Synthesis, NMR Characterization, and In...

Background/Objectives: For the first time, water-soluble encapsulated forms of 20-hydroxyecdysone were synthesized as clathrate complexes with β-cyclodextrin. The derivatives included a ketoxime form (20-NOH), obtained by selective oximation of the C-6 carbonyl, and a triacetate form (3Ac-20E), obtained by regioselective acetylation of the C-2, C-3, and C-22 hydroxyl groups. Methods: The chemical structures of the obtained compounds were confirmed using spectroscopic methods, including ¹H and ¹³C NMR. The Z-configuration of the oxime in 20-NOH was established by the diagnostic upfield shift in the C-5 resonance (δ 36.0 ppm) relative to the parent compound (δ 51.4 ppm). Results: The antioxidant activity was evaluated using FRAP and DPPH assays. The β-cyclodextrin complexes were shown to exhibit significantly higher radical scavenging activity, with IC₅₀ values of <0.05 mg/mL compared to 0.24 mg/mL for the parent 20-hydroxyecdysone (20E). In the FRAP assay, the complexes demonstrated enhanced reducing capacity, with values of 3.452 ± 0.520 AAE/mL for the ketoxime form (20-NOH·β-CD) at a concentration of 0.75 mg/mL and 3.810 ± 0.279 AAE/mL for the acetate form (3Ac-20E·β-CD) at the same concentration, whereas the parent compound did not exceed 0.46 AAE/mL. In both in vitro and in vivo experiments, it was established that the investigated complexes demonstrated hepatoprotective properties, as evidenced by the normalization of biochemical parameters as well as the restoration of liver structure according to ultrasound and histological studies. The ketoxime-derived complex (20-NOH·β-CD) at a dose of 25 mg/kg demonstrates the greatest reduction in transaminase activity and MDA levels, indicating its high efficacy and the presence of an optimal therapeutic dose. In addition, the encapsulated forms demonstrated moderate antifungal activity against Candida albicans and bactericidal activity against four bacterial strains (MIC >10 mg/mL). Conclusions: The obtained results indicate that β-cyclodextrin complexes significantly enhance the biological activity of 20E and highlight the potential of these compounds for pharmaceutical applications.