Neurosteroids and epileptogenesis.

Epileptogenesis is defined as the latent period at the end of which spontaneous recurrent seizures occur. This concept has been recently re-evaluated to include exacerbation of clinically-manifested epilepsy. Thus, in patients affected by pharmacoresistant seizures, the progression toward a worse condition may be viewed as the result of a durable epileptogenic process. However, the mechanism potentially responsible for this progression remains unclear. Neuroinflammation has been consistently detected both in the latent period and in the chronic phase of epilepsy, especially when brain damage is present. This phenomenon is accompanied by glial cell reaction, leading to gliosis. We have previously described rats presenting an increased expression of the cytochrome P450 cholesterol side-chain cleavage (P450scc) enzyme, during the latent period, in glial cells of the hippocampus. The P450scc enzyme is critically involved in the synthesis of neurosteroids and its up-regulation is associated with a delayed appearance of spontaneous recurrent seizures in rats that experienced status epilepticus induced by pilocarpine. Moreover, by decreasing the synthesis of neurosteroids able to promote inhibition, such as allopregnanolone, through the administration of the 5α-reductase blocker finasteride, it is possible to terminate the latent period in pilocarpine-treated rats. Finasteride was also found to promote seizures in the chronic period of epileptic rats, suggesting that neurosteroids are continuously produced to counteract seizures. In humans, exacerbation of epilepsy has been also described in patients occasionally exposed to finasteride. Overall, these findings suggest a major role of neurosteroids in the progression of epilepsy and a possible antiepileptogenic role of allopregnanolone and cognate molecules.

Proteomic analysis in the lichen Physcia adscendens exposed to cadmium stress.

This work was undertaken to explore the potential of proteomics to dissect parallel and consecutive events of cadmium stress response in the lichen Physcia adscendens (Fr.) H. Olivier. Thalli were exposed to 0 (control) and 36 microM Cd for 6, 18, 24 and 48 h. Two-dimensional electrophoresis and mass spectrometry analyses showed an 80-85% spot identity between 6 and 18 h vs. 24 and 48 h of Cd exposure. Putative heat-shock proteins and glutathione S-transferase generally increased their expression all over the Cd treatments. By contrast, ABC transporters were underexpressed after 6-18 h, but in some cases induced after 24-48 h of Cd exposure. The cytochrome P450 appeared to have a variable expression pattern over time. Overall these data suggest that a considerable importance in the response of P. adscendens thalli to Cd stress can be assumed by differential expression of various protein families.

Effects of alpha- and beta-cyclodextrin complexation on the physico-chemical properties and antioxidant activity of some 3-hydroxyflavones.

Inclusion complexes of some flavonols (3-hydroxyflavone, morin and quercetin) have been obtained with alpha- and beta-cyclodextrins, by the co-evaporation method. Different analytical techniques (DSC, XRPD, FT-IR, 1H-NMR, UV-Vis) have been employed for a throughout investigation of the structural characteristics of such supramolecular aggregates, which exhibited distinct spectroscopic features and properties from both "guest" and "host" molecules. The stoichiometric ratios and stability constants describing the extent of formation of the complexes have been determined by phase-solubility studies; in all cases type-AL diagrams have been obtained (soluble 1:1 complexes). The effect of molecular encapsulation on the flavonols antioxidant activity has been afterwards evaluated, by means of different biological assays (Bathophenanthroline test; Comet assay; Lipid peroxidation). Complexation with cyclodextrins further improved the antioxidant activity, increasing drugs solubility in the biological moiety.

Study of flavonoids/beta-cyclodextrins inclusion complexes by NMR, FT-IR, DSC, X-ray investigation.

Flavonoids are natural substances with a lot of biological activities, including the antioxidant one. Their use in pharmaceutical field is, however, limited by their aqueous insolubility. As the formation of the inclusion complexes can improve their solubility in water, the flavonoids hesperetin, hesperidin, naringenin and naringin have been complexed with beta-cyclodextrin (beta-CD) by the coprecipitation method and studied in solution and in solid state by NMR, FT-IR, differential scanning calorimetry and X-ray techniques. The effects of complexation on the chemical shifts of the internal and external protons of beta-CD in the presence of each flavonoid were observed.

New combined treatment of hypermelanosis: analytical studies on efficacy and stability improvement.

The aim of this work was to evaluate the synergy between different lightening agents when associated; depigmenting activity was tested in vitro by monitoring the appearance of dopachrome, an intermediate in the melanogenesis process. The results obtained were compared with the depigmenting activity of each single compound, keeping the same global concentration of inhibitor. Our studies showed that the combination of hydroquinone and kojic acid had a synergistic effect, and that the maximum inhibiting action was achieved with an equimolecular mixture. This result could serve in the cosmetics field to prepare skin-lightening formulations that are less irritant. We also investigated the feasibility of complexing hydroquinone with cyclodextrin and evaluated the effectiveness of the complex obtained in the treatment of hyperpigmentation.

Study of beta-blockers/beta-cyclodextrins inclusion complex by NMR, DSC, X-ray and SEM investigation.

The formation of inclusion complexes between beta-cyclodextrin with the two beta-blockers, atenolol and celiprolol, have been studied in the aqueous environment and in the solid state by nuclear magnetic resonance (NMR) spectroscopy, X-ray, differential scanning calorimetry (DSC) and scanning electron microscopy (SEM) techniques. The magnitude of the chemical shifts of the interior and exterior beta-cyclodextrin protons in the presence of each beta-blocker indicated that these are included within the beta-cyclodextrin cavity. In aqueous solution they form 1:1 complexes. In the solid state the formation of the beta-cyclodextrin/atenolol (celiprolol) complexes is confirmed by X-ray, DSC and SEM, also employed to characterize pure substances and their physical mixtures.

Solid-state study of polymorphic drugs: carbamazepine.

Polymorphs of a compound have solid crystalline phases with different internal crystal lattices; in pharmaceuticals, differences due to polymorphism and pseudopolymorphism can affect bioavailability and effective clinical use. The aim of this work was to obtain the different polymorphic modifications of the anticonvulsant drug, carbamazepine, and to characterise them by means of typical structure-sensitive analytical techniques, such as FT-IR spectroscopy, XRPD and DSC. Further investigations were also performed by Hot Stage FT-IR thermomicroscopy, which permitted the visible and spectroscopic characterisation of the polymorphic forms during heating. Our results confirm the existence of three different polymorphic forms for anhydrous carbamazepine: Form III, the commercial one, Form I, obtained by heating Form III and Form II, crystallised from ethanolic solution. Substantial differences were detected among the polymorphs with regard to solid-state properties. Moreover, Hot Stage FT-IR thermomicroscopy proved its analytical potential to characterise the drug's polymorphism.

Properties of the racemic species of verapamil hydrochloride and gallopamil hydrochloride.

It is well known that the stereoselective actions associated with the enantiomeric constituents of a racemic drug can differ markedly in their pharmacodynamic or pharmacokinetic properties. Nevertheless, molecular chirality manifests itself in the solid, that is, crystalline state. The aim of this work was to characterize the solid-state properties of verapamil HCl and gallopamil HCl, two well-known chiral calcium channel antagonists. The characterization of the solid state for the single enantiomers and equimolecular mixtures for both the calcium antagonists was performed by solid-state techniques such as Fourier transform infrared (FT-IR spectroscopy), X-ray powder diffractometry (XRD) and differential scanning calorimetry (DSC). The FT-IR spectra and XRD of the single enantiomers are different from those of the corresponding equimolecular mixture owing to their different crystalline structure. The thermal behavior of the racemates and pure enantiomers were examined by DSC, and the resultant experimental and theoretical binary phase diagrams are discussed. Spectroscopic solid-state techniques, such as FT-IR and XRD, are useful in combination with thermal analysis for characterizing the racemic species of chiral drugs. The data obtained prove that the equimolecular mixtures of both verapmil hydrochloride and gallopamil hydrochloride exist as racemic compounds. Determination of the enantiomeric purity of the enantiomers and racemic compounds of both the calcium antagonists analyzed was performed by DSC.

Determination of azelaic acid in pharmaceuticals and cosmetics by RP-HPLC after pre-column derivatization.

This paper reports a RP-HPLC method for the determination in topics of azelaic acid, a keratolytic and anti-comedogenic agent widely used in the treatment of all types of acne. A derivatization step was needed prior to chromatographic analysis because the analyte is lacking in chromophore. A sample clean-up procedure by solid-phase extraction was also developed to analyse azelaic acid in complex matrices, such as pharmaceutical and cosmetic formulations.