Forced degradation studies of norepinephrine and epinephrine from dental anesthetics: Development of stability-indicating HPLC method and in silico toxicity evaluation.

Injectable solutions containing epinephrine (EPI) and norepinephrine (NE) are not stable, and their degradation is favored mainly by the oxidation of catechol moiety. As studies of these drugs under forced degradation conditions are scarce, herein, we report the identification of their degradation products (DP) in anesthetic formulations by the development of stability-indicating HPLC method. Finally, the risk assessment of the major degradation products was evaluated using in silico toxicity approach. HPLC method was developed to obtain a higher selectivity allowing adequate elution for both drugs and their DPs. The optimized conditions were developed using a C18 HPLC column, sodium 1-octanesulfonate, and methanol (80:20, v/v) as mobile phase, with a flow rate of 1.5 mL/min, UV detection at 199 nm. The analysis of standard solutions with these modifications resulted in greater retention time for EPI and NE, which allow the separation of these drugs from their respective DPs. Then, five DPs were identified and analyzed by in silico studies. Most of the DPs showed important alerts as hepatotoxicity and mutagenicity. To the best of our acknowledgment, this is the first report of a stability-indicating HPLC method that can be used with formulations containing catecholamines.

Nanoparticles Loaded with a New Thiourea Derivative: Development and In vitro Evaluation Against Leishmania amazonensis.

BACKGROUND: Leishmaniasis is a neglected tropical disease caused by protozoa of the genus Leishmania. Current treatments are restricted to a small number of drugs that display both severe side effects and a potential for parasites to develop resistance. A new N-(3,4-methylenedioxyphenyl)-N'- (2-phenethyl) thiourea compound (thiourea 1) has shown promising in vitro activity against Leishmania amazonensis with an IC50 of 54.14 µM for promastigotes and an IC50 of 70 µM for amastigotes. OBJECTIVE: To develop a formulation of thiourea 1 as an oral treatment for leishmaniasis, it was incorporated into Nanoparticles (NPs), a proven approach to provide long-acting drug delivery systems. METHODS: Poly (D,L-Lactic-co-Glycolic Acid) (PLGA) polymeric NPs containing thiourea 1 were obtained through a nanoprecipitation methodology associated with solvent evaporation. The NPs containing thiourea 1 were characterized for Encapsulation Efficiency (EE%), reaction yield (% w/w), surface charge, particle size and morphology by Transmission Electron Microscopy (TEM). RESULTS: NPs with thiourea 1 showed an improved in vitro leishmanicidal activity with a reduction in its cytotoxicity against macrophages (CC50>100 µg/mL) while preserving its IC50 against intracellular amastigotes (1.46 ± 0.09 µg/mL). This represents a parasite Selectivity Index (SI) of 68.49, which is a marked advancement from the reference drug pentamidine (SI = 30.14). CONCLUSION: The results suggest that the incorporation into NPs potentiated the therapeutic effect of thiourea 1, most likely by improving the selective delivery of the drug to the phagocytic cells that are targeted for infection by L. amazonensis. This work reinforces the importance of nanotechnology in the acquisition of new therapeutic alternatives for oral treatments.

Experimental composites containing quaternary ammonium methacrylates reduce demineralization at enamel-restoration margins after cariogenic challenge.

OBJECTIVE: This study evaluated the influence of experimental composites containing quaternary ammonium monomers (QAM) at different concentrations and alkyl chains on demineralization at enamel-composite margins after cariogenic challenge. METHODS: Standardized 4×4mm cavities were cut into 35 bovine enamel blocks, which were randomly divided into seven groups (n=5) and restored with the following experimental composites and commercial materials: (G12.5) - 5% dimethylaminododecyl methacrylate (DMADDM) with a 12-carbon alkyl chain (G12.10) - 10% DMADDM, (G16.5) - 5% dimethylaminohexadecyl methacrylate (DMAHDM) with a 16-carbon alkyl chain (G16.10) - 10% DMAHDM, (CG) - control group (without QAM), (GZ250) - commercial composite (Filtek Z250®), and (GIC) - glass ionomer cement (Maxxion R®). After restorative procedures, initial microhardness was measured and experimental composites were subjected to Streptococcus mutans biofilm formation for 48h. After cariogenic challenge, the samples were washed and microhardness was reassessed. A 3D non-contact profilometer was used to determine surface roughness and enamel demineralization was assessed by micro-CT. Microhardness results were analyzed by the Kruskal-Wallis and Mann-Whitney tests and micro-CT results were analyzed by Tukey's HSD test (95% confidence interval). RESULTS: None of the materials could prevent mineral loss at the enamel-restoration margins. The addition of 10% DMAHDM yielded the lowest, albeit statistically significant, mineral loss (p<0.05). 3D non-contact profilometry showed enamel surface roughness modification after biofilm exposure. The CG had the highest roughness values. Micro-CT analysis revealed mineral loss, except for GIC. SIGNIFICANCE: The addition of 10% QAM with a 16-carbon chain in experimental composites reduced mineral loss at the enamel-restoration margins after cariogenic challenge.

Physical and chemical properties of model composites containing quaternary ammonium methacrylates.

OBJECTIVE: Investigate physical and chemical properties of model composites formulated with quaternary ammonium salt monomers (QAS) at different concentrations and alkyl chains lengths METHODS: QAS with 12 dimethylaminododecyl methacrylate (DMADDM) and 16 dimethylaminohexadecyl methacrylate (DMAHDM) chains lengths were synthesized and incorporated at 5 and 10% in model composites, resulting in four groups: G12.5 (DMADDM 5%), G12.10 (DMADDM 10%), G16.5 (DMAHDM 5%), G16.10 (DMAHDM 10%). One group was used as control group (CG 0%). Degree of conversion (DC); water sorption (WS) and solubility (SL); hygroscopic expansion (HE); degradation temperature (DT); glass transition temperature (Tg) and polymerization shrinkage (PS) were determined. Knoop hardness (KNH), flexural strength (FS) and elastic modulus (EM) were measured before and after storage Data were submitted to ANOVA and Tukey's test (p≤0.05). RESULTS: DC ranged between 76.1 (G12.10) and 70.7 (G16.5) %; CG had the lowest WS, SL and HE. There was no statistical difference for PS and FS. KHN values ranged between 30.2 (GC) and 25 (G16.10) and after storage the performance was depended on QAS concentration and chain length. For EM, CG had the highest values before and after storage and no difference was observed in the QAS groups before storage. After storage, the results were dependent on QAS concentration (3.5-4.3GPa). SIGNIFICANCE: In general, the addition of QAS increased composite's degradation compared with the CG. In the tested QAS, the addition of DMADDM at 5% concentration resulted in a less degradable material.

Antibiofilm properties of model composites containing quaternary ammonium methacrylates after surface texture modification.

OBJECTIVE: Investigate antimicrobial properties and surface texture of model composites with different concentration and alkyl chain length of quaternary ammonium monomers (QAS). METHODS: Monomers derived from QAS salts with alkyl chain lengths of 12 carbons ((dimethylaminododecyl methacrylate) DMADDM) and 16 carbons (dimethylaminohexadecyl methacrylate-DMAHDM) were obtained from the reactions of their respective organo-halides with the tertiary amine 2-(dimethylamino)ethyl methacrylate (DMAEMA). DMADDM and DMAHDM were incorporated into model composite in concentrations of 5 or 10%, resulting the following groups: G12.5 (DMADDM 5%), G12.10 (DMADDM 10%), G16.5 (DMAHDM 5%), G16.10 (DMAHDM 10%) and GC (control). Biofilm viability, lactic acid production and surface roughness were analysed 24h after samples preparation (initial), repeated after toothbrush abrasion and after polishing simulation. Data were submitted to ANOVA and Tukey's test (p≤0.05). RESULTS: The longer the molecular chain size of QAS and the higher its concentration (G16.10), the lower was the viability and the production of lactic acid by the biofilm. No differences were detected in initial roughness' measurements among groups. However, after abrasion, there was an increase of biofilm viability and lactic acid production. Composites containing QAS presented rougher surfaces compared to the CG. After polishing, biofilm viability and surface roughness were statistically similar for all groups. Nevertheless, DMAHDM at 10% showed reduction in lactic acid production. SIGNIFICANCE: Chain length and concentration of QAS influenced biofilm development and production of lactic acid. Longer chains and higher concentrations of QAS promoted better antimicrobial properties. Changes in surface texture caused by abrasion, decreased antibiofilm properties.