Maria Addolorata Saracino

HPLC-F analysis of melatonin and resveratrol isomers in wine using an SPE procedure.

An original analytical method has been developed for the determination of the antioxidants trans-resveratrol (t-RSV) and cis-resveratrol (c-RSV) and of melatonin (MLT) in red and white wine. The method is based on HPLC coupled to fluorescence detection. Separation was obtained by using a RP column (C8, 150 mm x 4.6 mm id, 5 mum) and a mobile phase composed of 79% aqueous phosphate buffer at pH 3.0 and 21% ACN. Fluorescence intensity was monitored at lambda = 386 nm while exciting at lambda = 298 nm, mirtazapine was used as the internal standard. A careful pretreatment of wine samples was developed, using SPE with C18 cartridges (100 mg, 1 mL). The calibration curves were linear over the following concentration ranges: 0.03-5.00 ng/mL for MLT, 3-500 ng/mL for t-RSV and 1-150 ng/mL for c-RSV. The LOD values were 0.01 ng/mL for MLT, 1 ng/mL for t-RSV and 0.3 ng/mL for c-RSV. Precision data, as well as extraction yield and sample purification results, were satisfactory. Thus, the method seems to be suitable for the analysis of MLT and resveratrol isomers in wine samples. Moreover, wine total polyphenol content and antioxidant activity were evaluated.

Liquid chromatographic analysis of oxcarbazepine and its metabolites in plasma and saliva after a novel microextraction by packed sorbent procedure.

A rapid and reliable analytical method suitable for the simultaneous determination of the antiepileptic drug, oxcarbazepine and its metabolites in human plasma and saliva by means of liquid chromatography with diode array detection (DAD) has been developed. Oxcarbazepine and its metabolites (10,11-dihydro-10-hydroxycarbamazepine, trans-10,11-dihydro-10,11-dihydroxycarbamazepine and 3-hydroxycarbamazepine) were baseline separated within 6.5 min on a reversed-phase C18 column with a phosphate buffer-acetonitrile-triethylamine mixture as the mobile phase. The DAD detector was set at 240 nm. A sample preparation method for biological samples using a microextraction by packed sorbent technique has been implemented, employing a C18 sorbent inserted into a microvolume syringe and using only a small volume (25 microL) of plasma or saliva. The extraction yield values were satisfactory for all analytes (>86.5%) as well as the precision data, which were always in the low percentage of relative standard deviation values (<4.6%). The method was successfully applied to both plasma and saliva samples drawn from psychiatric and neurological patients undergoing treatment with oxcarbazepine (Tolep) tablets.

Rapid assays of clozapine and its metabolites in dried blood spots by liquid chromatography and microextraction by packed sorbent procedure

A novel analytical approach has been developed for the determination of clozapine and its metabolites in dried blood spots on filter paper, using a chromatographic method coupled with a microextraction by packed sorbent procedure. The analytes were separated on a RP-C18 column using a mobile phase composed of 20% methanol, 16% acetonitrile and 64% aqueous phosphate buffer. Coulometric detection was used, setting the guard cell at +0.050 V, the first analytical cell at -0.200 V and the second analytical cell at +0.500 V. Clozapine and its metabolites were extracted from dried blood spots with phosphate buffer and, then, a microextraction by packed sorbent procedure for the sample clean-up was implemented obtaining good extraction yields. The calibration curve was linear over the 2.5-1000 ng mL(-1) blood concentration ranges for all the analytes. The method validation gave satisfactory results in terms of sensitivity, precision, selectivity and accuracy. The analytical method was successfully applied to dried blood spots from several psychiatric patients for therapeutic drug monitoring purpose.

Separation and analysis of glycyrrhizin, 18β-glycyrrhetic acid and 18α-glycyrrhetic acid in liquorice roots by means of capillary zone electrophoresis

Abstract Glycyrrhizin is the main active compound of Glycyrrhiza glabra root extracts; according to recent studies, glycyrrhizin and its aglycon, glycyrrhetic acid, have interesting therapeutic properties. A new capillary electrophoretic method has been developed for the separation and quantification of glycyrrhizin, β-glycyrrhetic acid and its isomer α-glycyrrhetic acid. Separation of the analytes was achieved in less than 3min on a fused silica capillary, by injecting the samples at the short end of the capillary (effective length: 8.5cm). The background electrolyte was composed of pH 10.0 carbonate buffer, methanol and ethylene glycol (80/10/10) and contained 0.4% β-cyclodextrin; indomethacin was used as the internal standard. Diode array detection was used, with quantitative assays carried out at 254nm. Linearity was found over the 5–200 and 2.5–100μgmL−1 concentration ranges for glycyrrhizin and glycyrrhetic acid, respectively. This method has been applied to the determination of the analytes in different matrices (liquorice roots and commercial confectionery products), and to the purity control of β-glycyrrhetic acid obtained from the hydrolysis of glycyrrhizin. When analysing β-glycyrrhetic acid and its epimer in roots, the samples were purified by means of a suitable solid-phase extraction (SPE) procedure with Oasis HLB cartridges, which granted good selectivity, eliminating matrix interference.

Analysis of risperidone and its metabolite in plasma and saliva by LC with coulometric detection and a novel MEPS procedure

A new analytical method, based on the use of liquid chromatography with coulometric detection, has been developed and applied to quantify risperidone and its main active metabolite 9-hydroxyrisperidone in human plasma and saliva. The analytes were separated on a reversed phase C18 column, using a mobile phase composed of acetonitrile (26%) and a pH 6.5 phosphate buffer (74%). Pipamperone was used as the internal standard. A high sensitivity coulometric detection analytical cell containing two flow-through working electrodes was used: electrode 1 was set at +0.500V and electrode 2 at +0.700V. The detector response was linear over a plasma and saliva concentration range of 0.5-50.0ngmL(-1) for risperidone and 0.5-100.0ngmL(-1) for 9-hydroxyrisperidone. The limit of quantitation and the limit of detection for risperidone and 9-hydroxyrisperidone were 0.5ngmL(-1) and 0.17ngmL(-1), respectively. A novel clean-up procedure of biological samples was developed using the microextraction by packed sorbent technique, which gave good extraction yield for both the analytes, with absolute recovery values higher than 90.1%. The intra-day and the inter-day precision results, expressed by relative standard deviation values, were lower than 5.8%. Accuracy and selectivity assays were also satisfactory. The validated method has been successfully applied to the analysis of risperidone and 9-hydroxyrisperidone in plasma and saliva of psychiatric patients undergoing therapy with risperidone.

Antipsychotic and antiepileptic drugs in bipolar disorder: the importance of therapeutic drug monitoring.

Bipolar disorder (BD) is a long-term illness with mood swings which are characterized by recurrent episodes of mania/hypomania and depression, with variable interpolations of relatively asymptomatic periods, called euthymic, in which, however, some psychopathological symptoms may persist. Although mood stabilizers, such as lithium, are the first-line treatment for the prevention of new BD episodes, combination therapy has become the standard of care for BD patients. Besides lithium, the use of a mood stabilizer along with an atypical antipsychotic is recommended in many patients. Recently, atypical antipsychotics (quetiapine, olanzapine, risperidone and aripiprazole) and antiepileptic agents (valproate, lamotrigine and oxcarbazepine) are increasingly used as mood stabilizers. To reduce side effects and optimize treatment it is important to perform accurate monitoring of drug blood levels in these patients, who are often treated with multiple drugs. Therapeutic drug monitoring (TDM) is in fact a powerful tool that, starting from clinical-chemical correlation data, allows to tailor-cut treatment to the specific needs of individual patients; hence the need to have reliable analytical methods available for the determination of plasma levels of drugs and their metabolites. Analyses of biological samples are mainly carried out using high-performance liquid chromatography (HPLC) coupled with different detectors, capillary electrophoresis and gas-chromatography. Various procedures are employed to remove biological interferences before analyzing the samples. This review focuses on currently available analytical TDM methods for atypical antipsychotics and antiepileptic agents used in the treatment of patients with bipolar disorder. Advantages and limitations of the various analytical methods will be reviewed and discussed, together with an evaluation of the role of TDM.

Simultaneous high-performance liquid chromatographic determination of olanzapine and lamotrigine in plasma of bipolar patients.

An original method based on the use of high-performance liquid chromatography with both coulometric and diode array detection has been developed for the therapeutic drug monitoring of patients with bipolar disorders being treated with olanzapine and lamotrigine. Chromatographic separation was achieved on a reversed-phase C8 column (150 × 4.6 mm internal diameter, 5 μm) using a mobile phase composed of methanol (27%) and a 50.0 mmol/L, pH 3.5 phosphate buffer (73%). For the analysis of olanzapine and its main metabolite, N-desmethylolanzapine, a coulometric detector was used, with electrode 1 set at -200 mV and electrode 2 at +500 mV. Lamotrigine was determined using a diode array detection at 220 nm. The two detectors were connected in series. For the analysis of biological samples, a clean-up procedure was implemented by means of solid-phase extraction using phenyl cartridges and eluting the analytes with methanol; only a small volume of plasma (150 μL) was needed to analyze both olanzapine and lamotrigine. Linear responses were obtained between 0.1 and 50.0 ng mL−1 for olanzapine, 0.1 and 25.0 ng mL−1 for N-desmethylolanzapine, and between 0.25 and 10.0 μg mL−1 for lamotrigine. The extraction yield values were always higher than 90% for all the analytes, with precision (expressed as relative standard deviation values) lower than 3.4%. The method was applied successfully to some human plasma samples drawn from bipolar patients undergoing combined therapy with the two drugs. Satisfactory values for accuracy were obtained, with mean recovery higher than 91%. Thus, the method appears suitable for the investigation of the chemical-clinical correlations in patients receiving therapy with olanzapine and lamotrigine.

Determination of selected phenothiazines in human plasma by solid-phase extraction and liquid chromatography with coulometric detection.

A new analytical method, based on liquid chromatography with coulometric detection, has been developed and applied to the determination of selected phenothiazines (chlorpromazine, promazine, fluphenazine and levomepromazine) in human plasma. The drugs were separated on a Discovery pentafluorophenylpropyl column, using a mobile phase composed of acetonitrile (32%) and a pH 1.9 phosphate buffer (68%). Promethazine was used as the internal standard. Detection was carried out at an oxidation potential of +0.500 V. A novel clean-up procedure was developed by means of solid-phase extraction, using cyanopropyl cartridges, which gave good extraction yield for all the analytes, with absolute recovery values higher than 91.0%. The detector response was linear over a plasma concentration range of 0.5-250.0 ng mL(-1) for chlorpromazine, promazine and levomepromazine and of 0.2-4.0 ng mL(-1) for fluphenazine. Precision results, expressed by the intra-day and the inter-day relative standard deviation values, were good, being lower than 3.9%. Accuracy data were satisfactory as well. The method has been successfully applied to the analysis of drug plasma levels of psychiatric patients undergoing therapy with selected phenothiazines.

Combined liquid chromatography–coulometric detection and microextraction by packed sorbent for the plasma analysis of long acting opioids in heroin addicted patients

The sublingual combination of buprenorphine and naloxone (Suboxone(®)) and Methadone Maintenance Therapy have been found effective in treating heroin addiction. A new analytical method suitable for the simultaneous determination of buprenorphine, norbuprenorphine, methadone and naloxone in human plasma by means of liquid chromatography with coulometric detection has been developed. The chromatographic separation was achieved with a phosphate buffer-acetonitrile mixture as the mobile phase on a cyano column. The monitoring cell of the coulometric detector was set at an oxidation potential of +0.600 V. A rapid clean-up procedure of the biological samples using a microextraction by packed sorbent technique has been implemented, employing a C8 sorbent inserted into a syringe needle. The extraction yield values were satisfactory for all analytes (>85%). The calibration curves were linear over a range of 0.25-20.0 ng mL(-1) for buprenorphine and norbuprenorphine, 3.0-1000.0 ng mL(-1) for methadone and 0.13-10.0 ng mL(-1) for naloxone. The sensitivity was also high with limits of detection of 0.08 ng mL(-1) for both buprenorphine and norbuprenorphine, 0.9 ng mL(-1) for methadone and 0.04 ng mL(-1) for naloxone. The intraday and interday precision data were always satisfactory. The method was successfully applied to plasma samples obtained from former heroin addicts treated with opioid replacement therapy.

Promising Medications for Cocaine Dependence Treatment

Cocaine dependence is characterized by compulsive drug seeking and high vulnerability to relapse. Overall, cocaine remains one of the most used illicit drugs in the world. Given the difficulty of achieving sustained recovery, pharmacotherapy of cocaine addiction remains one of the most important clinical challenges. Recent advances in neurobiology, brain imaging and clinical trials suggest that certain medications show promise in the treatment of cocaine addiction. The pharmacotherapeutic approaches for cocaine dependence include medications able to target specific subtypes of dopamine receptors, affect different neurotransmitter systems (i.e. noradrenergic, serotonergic, cholinergic, glutamatergic, GABAergic and opioidergic pathways), and modulate neurological processes. The systematic reviews concerning the pharmacological treatment of cocaine dependence appear to indicate controversial findings and inconclusive results. The aim of future studies should be to identify the effective medications matching the specific needs of patients with specific characteristics, abandoning the strategies extended to the entire population of cocaine dependent patients. In the present review we summarize the current pharmacotherapeutic approaches to the treatment of cocaine dependence with a focus on the new patents.