Gianpiero Boatto

Improvement of thymol properties by complexation with cyclodextrins: in vitro and in vivo studies

Thymol, an effective agent for microbial diseases, has a low aqueous solubility and a strong bitter/irritating taste. These physicochemical characteristics need to be improved to develop pharmaceutical preparations. This study evaluates whether β-cyclodextrin and a copolymer based on dimethylaminoethyl methacrylate (DMAEMA) interact with thymol in order to control powderization, solubilization, and taste-masking properties. The thymol-β-cyclodextrin complex was prepared by co-precipitation and sealed-heating methods. The DMAEMA copolymer was mixed with the complex using a new approach, instead of spray coating, to decrease thymol volatility. In vivo studies were performed. Sealed-heating is a suitable method for including thymol in β-cyclodextrin with a good loading efficiency; thymol volatility control is achieved by mixing the complex with the DMAEMA copolymer. β-Cyclodextrin accelerates the in vivo thymol absorption rate compared with the free drug; the thymol half-life is still long. Therefore, a low number of administrations per day are required. Although bioavailability is unchanged with respect to free thymol, high doses could be administered of a selected formulation without compromising the compliance. Furthermore, thymol that is not absorbed is held along the intestine, where it can useful in the treatment and/or prevention of intestinal bacterial diseases.

Determination of amphetamines in human whole blood by capillary electrophoresis with photodiode array detection.

A capillary electrophoresis (CE) with photodiode array detection (DAD) method for the analysis of amphetamines in human whole blood samples is described. Amphetamines were applied to CE without any derivatization procedure and detected at 200 nm for a rapid and simple analysis. The UV-spectra are show in. Amphetamines were separated within 7 min through an uncoated fused-silica capillary (50 cm x 50 microm ID) using a 100 mM phosphate buffer (pH 2.5). A simple and fast extraction method of amphetamines from human whole blood was developed using acetonitrile. Very clean extracts were obtained in one step. Whole blood drugs free samples were spiked with amphetamine standard solution of known concentration. Linear calibration plots were obtained over a large concentration range, with correlation coefficients higher than 0.998. Recoveries between 81 and 99% were obtained. Limit of detection (LOD) was from 10 to 30 ng ml(-1) for most of amphetamines, except for MDMA, for which it was 80 ng ml(-1).

Synthesis, pharmacokinetics and anticonvulsant activity of 7-chlorokynurenic acid prodrugs

7-Chlorokynurenic acid 1 is a potent glycine-N-methyl-D-aspartate (NMDA) receptor antagonist, but it shows weak activity after systemic administration. In order to overcome the Blood-brain barrier (BBB), we synthetized three new esters 2-4 of 1 obtained by chemical conjugation with essential nutrients such as glucose and galactose, that are actively transported across the BBB. These compounds were assayed to evaluate their in vitro chemical and enzymatic hydrolysis. In addition the prodrugs 2-4 were tested for their ability to protect mice against NMDA-induced seizures after systemic administration. All the prodrugs 2-4 appeared moderately stable in pH 7.4 buffered solution and were susceptible to in vitro enzymatic hydrolysis. Intraperitoneal administration of either esters 2 or 4 was highly protective against seizures induced by NMDA in mice, with the latter prodrug showing the highest anticonvulsive activity. In addition, ester 4 undergoes a time-dependent extracellular hydrolysis into 1 when applied to mixed cultures of mouse cortical cells, a model that reproduces in vitro the cellular milieu encountered by the prodrugs once they penetrate the brain parenchyma.

l-Cysteine reduces oral ethanol self-administration and reinstatement of ethanol-drinking behavior in rats.

Our previous findings have shown that l-cysteine, a non essential amino acid, prevented ethanol (EtOH) induced conditioned place preference. The aim of the present study was to examine the effect of l-cysteine on the acquisition and maintenance of oral EtOH self-administration and on the reinstatement of EtOH-drinking behavior in Wistar rats. Rats were pretreated intraperitoneally with saline or l-cysteine (20 and 40 mg/kg) 30 min before each acquisition trial, in an operant nose-poking paradigm where they were given the opportunity to orally self-administer tap water or EtOH (5-10% v/v). Further, to evaluate if l-cysteine reduces the acquired oral EtOH self-administration, we carried out an independent experiment in which rats were trained to self-administer EtOH (10%); after all groups of rats developed similarly stable oral EtOH self-administration, the effect of l-cysteine (0, 40, 60, 80 and 100mg/kg) was tested. An additional group of rats was pretreated with saline or l-cysteine (80 mg/kg) and tested on reinstatement after EtOH extinction and, at the end of last reinstatement session, were utilized to measure blood and brain EtOH levels. The animals that had access to EtOH solution discriminated between the active and inactive nose-pokes and showed rates of active nose-pokes significantly higher than the tap water group. Furthermore, rats self-administering EtOH (10%) also demonstrated extinction behavior and gradually reinstated active nose-poke responding when EtOH was reintroduced. l-cysteine reduced both the acquisition and maintenance of oral EtOH self-administration. The reduced reinstatement of EtOH-drinking behavior was paralleled by a significant reduction of EtOH intake and correlated with blood and brain EtOH levels. The efficacy of l-cysteine on the various phases of alcohol drinking in rats, could represent an interesting pharmacological approach and could open a new line of research for the development of therapies to reduce EtOH intake in alcoholic patients.

Multi‐residue analysis of eight thioamphetamine designer drugs in human urine by liquid chromatography/tandem mass spectrometry

An analytical procedure for the simultaneous determination in human urine of several thioamphetamine designer drugs (2C-T and ALEPH series) is reported. The quantitative analysis was performed by liquid chromatography/tandem mass spectrometry and has been fully validated. The mass spectrometer was operated in positive-ion, selected reaction monitoring (SRM) mode. In order to minimize interferences with matrix components and to preconcentrate target analytes, solid-phase extraction was introduced in the method as a clean-up step. The entire method was validated for selectivity, linearity, precision and accuracy. The method turned out to be specific, sensitive, and reliable for the analysis of amphetamine derivatives in urine samples. The calibration curves were linear over the concentration range of 1 to 100 ng mL(-1) for all drugs with correlation coefficients that exceeded 0.996. The lower limits of detection (LODs) and quantification (LOQs) ranged from 1.2 to 4.9 ng mL(-1) and from 3.2 to 9.6 ng mL(-1), respectively.

LC-MS analysis of trimethoxyamphetamine designer drugs (TMA series)from urine samples

A sensitive liquid chromatography-mass spectrometric (LC-MS) method for quantification of an active psychedelic hallucinogenic drugs (trimethoxyamphetamines) in human urine after solid-phase extraction (SPE) with C(18) cartridge was developed and validated. Chromatographic separation was achieved on reversed-phase Phenomenex 3.0 microm Polar Plus column (150 mm x 2.1 mm) with acetonitrile -0.2% acetic acid as mobile-phase and the step gradient elution resulted in a total run time of about 20 min. The analytes were detected by using an electrospray positive ionization mass spectrometry in selected ion monitoring (SIM) mode. In the evaluated concentration range (10-200 ng/mL) (R(2) > or = 0.998) a good linear relationship was obtained. The lower limits of detection (LLODs) and quantification (LLOQs) ranged from 4.26 to 9.12 ng/mL and from 13.18 to 29.22 ng/mL, respectively. Average recoveries ranged from 68.52 to 97.90% in urine at the concentrations of 25, 50 and 100 ng/mL. Intra- and inter-day relative standard deviations were 3.70-10.77% and 7.63-12.94%, respectively. This LC-MS method proved to be robust and reliable, and suitable for the use as a confirmation method in clinical urine drug testing.

Prepuberal Stimulation of 5-HT7-R by LP-211 in a Rat Model of Hyper-Activity and Attention-Deficit: Permanent Effects on Attention, Brain Amino Acids and Synaptic Markers in the Fronto-Striatal Interface

The cross-talk at the prefronto-striatal interface involves excitatory amino acids, different receptors, transducers and modulators. We investigated long-term effects of a prepuberal, subchronic 5-HT7-R agonist (LP-211) on adult behaviour, amino acids and synaptic markers in a model for Attention-Deficit/Hyperactivity Disorder (ADHD). Naples High Excitability rats (NHE) and their Random Bred controls (NRB) were daily treated with LP-211 in the 5th and 6th postnatal week. One month after treatment, these rats were tested for indices of activity, non selective (NSA), selective spatial attention (SSA) and emotionality. The quantity of L-Glutamate (L-Glu), L-Aspartate (L-Asp) and L-Leucine (L-Leu), dopamine transporter (DAT), NMDAR1 subunit and CAMKIIα, were assessed in prefrontal cortex (PFC), dorsal (DS) and ventral striatum (VS), for their role in synaptic transmission, neural plasticity and information processing. Prepuberal LP-211 (at lower dose) reduced horizontal activity and (at higher dose) increased SSA, only for NHE but not in NRB rats. Prepuberal LP-211 increased, in NHE rats, L-Glu in the PFC and L-Asp in the VS (at 0.250 mg/kg dose), whereas (at 0.125 mg/kg dose) it decreased L-Glu and L-Asp in the DS. The L-Glu was decreased, at 0.125 mg/kg, only in the VS of NRB rats. The DAT levels were decreased with the 0.125 mg/kg dose (in the PFC), and increased with the 0.250 mg/kg dose (in the VS), significantly for NHE rats. The basal NMDAR1 level was higher in the PFC of NHE than NRB rats; LP-211 treatment (at 0.125 mg/kg dose) decreased NMDAR1 in the VS of NRB rats. This study represents a starting point about the impact of developmental 5-HT7-R activation on neuro-physiology of attentive processes, executive functions and their neural substrates.

Sensitization to methyl methacrylate in the plastic catheter of an insulin pump infusion set

Cases of allergic contact dermatitis due to acrylics in the adhesive used to attach the needle to the tubing have previously been reported in pump infusion set users, but never previously due to an acrylic in the plastic tubing itself, as in this case. The presence of methyl methacrylate demonstrated in the catheter of the patient’s own infusion set correlated with her positive patch test results. Since patch testing and chemical-physical analysis was negative for methyl methacrylate in the alternative Cliniset ® microinfusion set, this was supplied to the patient for her insulin therapy and no further adverse skin reactions were reported. Patch testing demonstrated probable cross-sensitivity between methyl methacrylate and other acrylics.

Galactosyl prodrug of ketorolac: synthesis, stability, and pharmacological and pharmacokinetic evaluations.

Although ketorolac is one of the most potent anti-inflammatory and analgesic drugs, its use has been strongly limited owing to the high incidence of adverse effects reported, particularly in the gastrointestinal tract. Using the prodrug approach, which allows the reduction of toxicological features of the parent drug without altering its pharmacological properties, we synthesized an orally administrable prodrug of ketorolac by means of its reversible conjugation to D-galactose (ketogal). In a single dose study, its pharmacokinetic profile was compared with that of ketorolac. Moreover, we found that this prodrug was able to maintain the anti-inflammatory and the analgesic activity of the drug without giving rise to gastric ulcer formation. Thus, these results indicate that ketogal is a highly effective and valid therapeutic alternative to ketorolac itself.

Essential Oil Composition of Hypericum perforatum L. var. angustifolium DC Growing Wild in Sardinia (Italy)

Abstract The oil from inforescences of Hypericum perforatum var. angustifolium growing wild and harvested in Sardinia (Italy) was analyzed by GC and GC/MS. The major compounds in the oil were 2-methyloctane (21.1%), germacrene D (17.6%) and α-pinene (15.8%).