Fractionation of hexane extracts from Achyrocline satureioides and their biological activities against Paenibacillus larvae.

Previous studies carried out in our laboratory described the antimicrobial activity of the whole hexanic extract (HE) of Achyrocline satureioides (Lam.) DC against Paenibacillus larvae, the causal agent of American Foulbrood (AFB) a disease of the honey bee larvae. In this study, the HE was partitioned into five main fractions by chromatographic techniques leading to the isolation of four known compounds: two prenylated phloroglucinol α-pyrones (1 and 3), 5,7-dihydroxy-3,8-dimethoxyflavone (gnaphaliin A) (2), and 23-methyl-6-O-demethylauricepyrone (4). Isolated compounds were further analyzed towards structural elucidation using 1H RMN and 13C RMN spectroscopic techniques. For the first time, the antimicrobial activity of the isolated compounds was evaluated against P. larvae strains by broth microdilution method and compared with that of the whole HE. Compounds 1-4 displayed minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values ranging between 0.07 and 62.5 µg/mL and 0.26 and 12.5 µg/mL, respectively. The lowest MIC and MBC values were obtained with compounds 3 and 4, respectively. The antimicrobial activity of each single compound and the combination of them showed that the presence of all compounds is needed for the antimicrobial efficacy of whole HE.

Extraction-free colorimetric determination of thymol and carvacrol isomers in essential oils by pH-dependent formation of gold nanoparticles.

An extraction-free method is described for the colorimetric determination of thymol (TY) and carvacrol (CA) isomers in essential oils by making use of the pH-dependent formation of gold nanoparticles (AuNPs). In solutions of pH 12, TY and CA form gold nanoparticles, while at pH ≤ 11 only CA does so. By taking advantage of this finding, two different approaches based on colorimetric assay (absorption at 550 nm) were developed: one at pH 12 for the determination of total CA and TY, and other at pH 9 and pH 12 for differential quantification of TY and CA. The former agrees with the well-established Folin-Ciocalteu method, and the latter provides a simple way for calculation of TY/CA ratio. The linear ranges are from 100 to 1000 µM at pH 9, and from 50 to 200 µM at pH 12. The limits of detection are 0.09 µM at pH 9, and 0.02 µM at pH 12. These features make this method simple, fast and reliable. Conceivably, it can be used to assess the quality of essential oils and may become a valuable alternative to more sophisticated, laborious and high time-consuming methods. Graphical abstract Schematic of the assay: At pH 12 (blue color), thymol and carvacrol form gold nanoparticles (Au), while at pH 9 (red color) only carvacrol does so. This finding resulted in a colorimetric method for the differential quantification of both compounds in essential oils.

Square wave voltammetry with multivariate calibration tools for determination of eugenol, carvacrol and thymol in honey.

The simultaneous determination of eugenol (EU), thymol (Ty) and carvacrol (CA) in honey samples, employing square wave voltammetry (SWV) and chemometrics tools, is informed for the first time. For this purpose, a glassy carbon electrode (GCE) was used as working electrode. The operating conditions and influencing parameters (involving several chemical and instrumental parameters) were first optimized by cyclic voltammetry (CV). Thus, the effects of the scan rate, pH and analyte concentration on the electrochemical response of the above mentioned molecules were studied. The results show that the electrochemical responses of the three compounds are very similar and that the voltammetric traces present a high degree of overlap under all the experimental conditions used in this study. Therefore, two chemometric tools were tested to obtain the multivariate calibration model. One method was the partial least squares regression (PLS-1), which assumes a linear behaviour. The other nonlinear method was an artificial neural network (ANN). In this last case we used a supervised, feed-forward network with Levenberg-Marquardt back propagation training. From the accuracies and precisions analysis between nominal and estimated concentrations calculated by using both methods, it was inferred that the ANN method was a good model to quantify EU, Ty and CA in honey samples. Recovery percentages were between 87% and 104%, except for two samples whose values were 136% and 72%. The analytical methodology was simple, fast and accurate.

Optimization of the electrodeposition of copper on poly-1-naphthylamine for the amperometric detection of carbohydrates in HPLC.

A modified electrode consisting of copper dispersed in a poly-1-naphthylamine (p-1-NAP/Cu) film on a glassy carbon electrode was used as an amperometric detector for the on-line analysis of various carbohydrates separated by high performance liquid chromatography. The results obtained with this new sensor were compared to those obtained with a modified electrode based on the same polymer but with copper ions incorporated at open circuit, as described in a previous paper. In this new modified electrode the copper microparticles were electrochemically deposited into the polymeric matrix by single potential step chronoamperometry. A nucleation and growth mechanism was proposed to explain the current transients of copper electrodeposition. The experimental results were fitted to the proposed mechanism by using a mathematical equation that considers three-dimensional growth and progressive nucleation, assuming a no overlap and no diffusion mechanism. Cyclic voltammetric experiments showed that the electrodeposited copper microparticles provided a catalytic surface suited for the oxidation of glucose and several carbohydrates. The sensitivity of the electrode was influenced by the amount of copper electrodeposited, which in turn depended on the applied overpotential used for the deposition of copper. Liquid chromatographic experiments were carried out to test the analytical performance of these electrodes for the determination of various carbohydrates.