Evaluation of the ROS Inhibiting Activity and Mitochondrial Targeting of Phenolic Compounds in Fibroblast Cells Model System and Enhancement of Efficiency by Natural Deep Eutectic Solvent (NADES) Formulation.

PURPOSE: Many phenolics have already been tested for their antioxidant activities using in vitro methods. However, such assays do not consider the complexity of real cellular systems, and most of the phenolics characterized with such assays shows disappointing results when evaluated in cells. Accordingly, there is a need to develop effective screening methods. METHODS: Antioxidants were first evaluated by CAT assay and then, evaluated for their ability (i) to reduce the level of ROS using fluorescent probe, (ii) to cross fibroblast cell membranes using confocal microscopy, and (iii) to target mitochondria. Antioxidants were also formulated in NADES. RESULTS: Correlation was obtained when comparing CAT results with short term inhibition (2 h) in the fibroblast cells. On the contrary, it was difficult to anticipate ROS inhibiting efficiency at long term (24 h) from both the CAT assay and the short term inhibition measurements. Indeed, some molecules displayed activity rapidly but lost it over time. In contrast, other molecules were better for long term. The comparable efficiency at long term of Bis-Ethylhexyl Hydroxydimethoxy Benzylmalonate (Bis-EHBm) and decyl rosmarinate, prompted us to further investigate the potential mitochondrial targeting of the former. Using mitochondrial probes, our results confirmed its mitochondrial location. Finally, the formulation of antioxidants in NADES could greatly improve their activity. CONCLUSIONS: Combinations of fast acting and slow acting molecules could be promising strategies to identify a performant antioxidant system. Bis-EHBm behaves as decyl rosmarinate with a confirmed mitochondrial location. Finally, the formulation of antioxidants in NADES could greatly improve their activity for ROS inhibition.

Adhesion of powders for inhalation: an evaluation of drug detachment from surfaces following deposition from aerosol streams.

PURPOSE: To evaluate micronized powder retention and detachment from inhaler surfaces following reproducible deposition by impaction, coupled with centrifugal particle detachment (CPD). METHODS: Micronized albuterol sulfate (AS) and beclomethasone dipropionate (BDP) were aerosolized as dry powders and deposited by cascade impaction onto different contact surfaces. Drug detachment from the surfaces was characterized using CPD, coupled with HPLC assay and scanning electron microscopy. RESULTS: Drugs which accumulated as aggregates on model surfaces detached with distinctive profiles for % remaining vs. applied centrifugal force; each profile showed reproducible values for the minimum force required to initiate drug detachment, Fyield. While differences occurred in the observed detachment profiles for different drugs and contact surfaces (polyacetal vs. aluminum), the deposited drug particle size had the most significant effect on these profiles, e.g., Fyield for AS (2.1-3.3 microm) was 383 +/- 12.7 microN compared with 18 +/- 13.8 microN for AS (4.7-5.8 microm). CONCLUSIONS: A technique was developed which enabled the experimental review, and subsequent data analysis, of the adhesive properties between different DPI construction materials and drug substances deposited from aerosol clouds. The technique appears to be of greater relevance to inhaler design decisions than earlier studies in the literature claiming to show differences in the adhesion of single drug particles to surfaces.