Novel High Affinity Sigma-1 Receptor Ligands from Minimal Ensemble Docking-Based Virtual Screening.

Sigma-1 receptor (S1R) is an intracellular, multi-functional, ligand operated protein that also acts as a chaperone. It is considered as a pluripotent drug target in several pathologies. The publication of agonist and antagonist bound receptor structures has paved the way for receptor-based in silico drug design. However, recent studies on this subject payed no attention to the structural differences of agonist and antagonist binding. In this work, we have developed a new ensemble docking-based virtual screening protocol utilizing both agonist and antagonist bound S1R structures. This protocol was used to screen our in-house compound library. The S1R binding affinities of the 40 highest ranked compounds were measured in competitive radioligand binding assays and the sigma-2 receptor (S2R) affinities of the best S1R binders were also determined. This way three novel high affinity S1R ligands were identified and one of them exhibited a notable S1R/S2R selectivity.

Hollow fiber-based liquid-liquid-liquid microextraction combined with electrospray ionization-ion mobility spectrometry for the determination of pentazocine in biological samples.

A new method based on liquid-liquid-liquid microextraction combined with electrospray ionization-ion mobility spectrometry (LLLME-ESI-IMS) was used for the determination of pentazocine in urine and plasma samples. Experimental parameters which control the performance of LLLME, such as selection of composition of donor and acceptor phase, type of organic solvent, ionic strength of the sample, extraction temperature and extraction time were studied. The limit of detection and relative standard deviation of the method were 2 ng/mL and 5.3%, respectively. The linear calibration ranged from 10 to 500 ng/mL with r(2)=0.998. Pentazocine was successfully determined in urine and plasma samples without any significant matrix effect.

Pentazocine transport by square-wave AC iontophoresis with an adjusted duty cycle.

So far, pentazocine iontophoresis has never been studied, although pentazocine is widely used in pain management. The purpose of this study was to determine whether pentazocine transportation through a cellophane membrane could be enhanced using square-wave alternating current (AC) iontophoresis with an adjusted duty cycle and dependence on the voltage and the duty cycle. Voltages of 10, 25 and 40 V with duty cycles of 50%, 51%, 52%, 53%, 54% and 55% were applied for 60 minutes at a high frequency of 1 MHz to diffusion cells on both sides of a cellophane membrane. The donor compartment was filled with a solution containing pentazocine. Square-wave AC iontophoresis with an adjusted duty cycle enhanced pentazocine transportation at higher voltages and duty cycles. These results suggested that the direct current (DC) component of the square-wave AC played an important role in enhancing pentazocine transportation despite changes in polarity at very high frequency of 1 MHz. The higher voltages and duty cycles induced a pH change. The practical electrical conditions that could be applied clinically were 25 V with a 54% duty cycle or 40 V with a 53% duty cycle.

Pentazocine monitoring in rat hair and plasma by HPLC-fluorescence detection with DIB-Cl as a labelling reagent.

Pentazocine (PZ) in rat hair and plasma was determined by HPLC-fluorescence detection with 4-(4,5-diphenyl-1H-imidazol-2-yl)benzoyl chloride (DIB-Cl) as a labelling reagent and cyclazocine (CZ) as an internal standard (IS). PZ and IS extracted from hair or plasma sample were derivatized with DIB-Cl and the resulted solution was cleaned up with solid phase extraction. The isocratic separation of DIB-PZ and -CZ within 20 min could be achieved by a Wakopak Handy-ODS column (250 x 4.6 mm i.d.) using a mobile phase composed of 0.1 mol/L acetate buffer (pH 6.2):acetonitrile (25:75, v/v). The detection limits of PZ at a signal-to-noise ratio of 3 for rat hair and plasma were 0.18 ng/mg and 0.57 ng/mL, respectively. Reproducible and precise results could be obtained by an IS method with RSD values less than 6.6% for within- and between-day measurements. The method was successfully applied for the monitoring of PZ levels in Zucker rat hair and plasma samples after a single administration of 25 mg/kg PZ. Moreover, incorporation rates of PZ into black and white hair of Zucker rat were evaluated.

[UV-densitometry determination of pentazocine hydrochloride in tablets after low temperature extraction]. / UV-denzitometrijsko odredjevanje pentozocin hidrohlorida u tabletama nakon termomikro extrakcije.

TLC-UV densitometric method has been used for quantitative analysis of pentazocine hydrochloride in the pure form, as well as in tablets after thermomicro extraction and direct application on the thin layer plate. This procedure was based on the conversion of the investigated substance into the gaseous state, quantitative application on the silica gel GF254 plate and the elution was made with 100:0.15 (v/v) methanol-conc. ammonium hydroxide. Gas chromatography-mass spectometric identification was performed by NIST 90 mass spectra library. Densitometric determination of pentazocine hydrochloride was performed by HPTLC Scanner on 278 nm and the results were compared to the officinal BP 93 method. The method was precise (RSD % 0.69-3.88), accurate (recovery 96.04-101.65) and reproducible (SD 0.03-0.23). The linearity level (correlation coefficient r = 0.9911) was very satisfactory.

Modified liquid junction interface for nonaqueous capillary electrophoresis-mass spectrometry.

Electrospray ionization (ESI) is the most widely used ionization method in on-line coupling of capillary electrophoresis-mass spectrometry (CE-MS). The conventional coaxial sheath flow electrospray interface is currently being replaced by the more sensitive nanoelectrospray technique. The usual limitation of nanoelectrospray CE-MS interface has been its short lifetime caused by deterioration of the metal coating on the CE capillary terminus. This article describes an easy way to construct a more durable and sensitive nanospray interface for nonaqueous CE-MS. In this approach a very thin glass spray capillary (ca. 30 microm outer diameter) is partly inserted inside the CE capillary, the junction being surrounded by the electrolyte medium, which is in contact with the platinum electrode. The interface was tested with five pharmaceuticals: methadone, pentazocine, levorphanol, dihydrocodeine, and morphine. Detection limits ranged from 12 to 540 fmol. Separation efficiency and reproducibility were also studied. The CE current was found to be stable and the migration times were highly reproducible. All the CE separations were carried out in a nonaqueous background electrolyte solution.

Liquid chromatography-fast atom bombardment mass spectrometry for detection and determination of pentazocine in human tissues.

A reliable and sensitive method was developed for the detection and determination of pentazocine in human solid tissues using liquid chromatography-dynamic fast atom bombardment (FAB) mass spectrometry, combined with a three-step liquid-liquid extraction procedure. Levallorphan tartrate served as an internal standard. The extract was evaporated to dryness and dissolved in the mobile phase, acetonitrile-10 mM ammonium acetate solution (20:80, pH 4.0) containing 0.5% glycerol as FAB matrix. The eluent was pumped at a flow rate of 25 microl/min and split before introduction to FAB mass spectrometer. Quantitative analysis was carried out by means of monitoring quasi-molecular ions with m/z 286 for pentazocine and m/z 284 for levallorphan. The lower limit of detection of pentazocine in each tissue tested was 1 ng/g with scan mode and 0.1 ng/g with SIM mode. Using this method, the concentrations of pentazocine were determined in the tissues of an autopsied individual to perform toxicological evaluation.

Screening for drugs of abuse (II): Cannabinoids, lysergic acid diethylamide, buprenorphine, methadone, barbiturates, benzodiazepines and other drugs.

Requirements for the provision of an efficient and reliable service for drugs of abuse screening in urine have been summarized in Part I of this review. The requirements included rapid turn-around times, good communications between requesting clinicians and the laboratory, and participation in quality assessment schemes. In addition, the need for checking/confirmation of positive results obtained for preliminary screening methods was stressed. This aspect of the service has assumed even greater importance with widespread use of dip-stick technology and the increasing number of reasons for which drug screening is performed. Many of these additional uses of drug screening have possible serious legal implications, for example, screening school pupils, professional footballers, parents involved in child custody cases, persons applying for renewal of a driving licence after disqualification for a drug-related offence, doctors seeking re-registration after removal for drug abuse, and checking for compliance with terms of probation orders; as well as pre-employment screening and work-place testing. In many cases these requests will be received from a general practitioner or drug clinic with no indication of the reason for which testing has been requested. This also raises the serious problems of a chain of custody, provision of two samples, stability of samples, and secure and lengthy storage of samples in the laboratory-samples may be requested by legal authorities several months after the initial testing. The need for confirmation of positive results is now widely accepted but it may be equally important to confirm unexpected negative results. Failure to detect the presence of maintenance drugs may lead to the patient being discharged from a drug treatment clinic and, if attendance at the clinic is one of the terms of continued employment, to dismissal. It seems likely that increasing abuse of drugs and the efforts of regulatory authorities to control this, will lead to the manufacture of more designer drugs. Production of substituted phenethylamines was facilitated by the drug makers' cook book, 'PIHKAL' (Phenethylamines I Have Known And Loved) by Dr Alexander Shulgin and Ann Shulgin, and production of substituted tryptamines is promised in their next book, TIHKAL. Looking to the future, laboratories will need to ensure that they can detect and quantitate an ever-increasing number of drugs and related substances. The question of confidence in results of drugs of abuse testing raised in 1993 by Watson has assumed even greater importance as a result of attention focused on the OJ Simpson trial in Los Angeles. Toxicological investigations are likely to be challenged more frequently in the future. Even if analyses have been performed by GC-MS, there is a need to establish the level of match between the spectrum of the unknown substance and a library spectrum which is considered acceptable for legal purposes. It will also be essential to ensure that computer libraries contain spectra for all substances likely to be encountered in drugs of abuse screening.

Analysis of zopiclone (Imovane) in postmortem specimens by GC-MS and HPLC with diode-array detection.

A 26-year-old black female was found dead at home. Her mouth was covered with a fluid containing chalky particles. Empty strips of Imovane (zopiclone) and an empty bottle of Fortal (pentazocine) were also found. No urine was available at autopsy. Screening of postmortem blood and stomach contents with enzyme-multiplied immunoassay technique (EMIT) detected only caffeine. Further screening using routine high-performance liquid chromatography (HPLC) with diode-array detection and gas chromatography (GC) with mass spectrometric detection revealed the presence of large amounts of pentazocine in the blood and stomach contents. In the HPLC chromatogram, a second peak that was only partially resolved from the solvent front was observed. Thin-layer chromatography demonstrated the presence of zopiclone, but optimized HPLC and GC conditions had to be used for proper identification and quantitation. This case illustrates the fact that zopiclone can be easily overlooked during routine forensic screening.

Interaction of enantiomeric pairs of opiates with phencyclidine binding sites in rat brain: identification of (+) pentazocine as a ligand potentially suitable for imaging sigma binding sites using positron emission tomography.

Some unnatural opiates, which do not interact with classical opiate receptors, interact with phencyclidine (PCP) receptors. Among their many pharmacological actions, drugs which bind to the PCP receptor antagonize the actions of glutamic acid mediated via the NMDA excitatory amino acid receptor, leading to their potential use as anti-ischemic and anticonvulsant agents. Despite an enormous effort, identification of a PCP receptor antagonist, which would be useful for research and therapeutics, has not yet been reported. Chemical modification of unnatural opiates as a means to produce a PCP antagonist, or PCP agonists with properties different than PCP, has not been fully explored. Towards this end, we determined the equilibrium dissociation constants of eight enantiomeric pairs of opiates for the rat brain PCP receptor.

The effect of antihistaminic drugs on pentazocine antinociception in the rat.

The antinociception produced by pentazocine, diphenhydramine, promethazine, chlorpheniramine, cyclizine and chlorcyclizine in the rat has been measured with a low-temperature (51.5 degrees C) hot-plate from 15 to 75 min following drug administration. The mean reaction times measured at 15 min and the area under the antinociception curves following administration of pentazocine (5 to 30 mg/kg) were linear. Diphenhydramine, chlorpheniramine, promethazine, cyclizine, and chlorcyclizine showed mild antinociceptive potency. The antinociception produced by SC pentazocine (5 and 10 mg/kg) was potentiated, rather than in a simple additive manner, by simultaneous IP administration of 20 mg/kg of diphenhydramine, promethazine, cyclizine, or chlorpheniramine, but not by chlorcyclizine. After concurrent administration of pentazocine and diphenhydramine, diphenhydramine did not alter pentazocine concentrations in the brain and plasma 15 to 75 min following drug administration, nor did pentazocine change diphenhydramine concentrations. Results of this study demonstrate that pentazocine antinociception can be potentiated by several antihistamines and that the potentiation was not due to a mutual effect on metabolism but rather through an as yet undefined mechanism.

Spontaneous live birth with a maternal history of intravenous use of pentazocine and tripelennamine (T's and blues).

A 24-year-old black female presented a live birth of six-months gestation. The 700-g neonate survived for 11 h. After toxicology revealed the presence of pentazocine and tripelennamine (T's and Blues), the mother admitted to using this combination intravenously 9 h previous to admission. Concentrations of pentazocine and tripelennamine were simultaneously determined by gas-liquid chromatography (GLC) combined with nitrogen selective detection. Analyses were performed on a 3% OV-101 column, with the added internal standard, dexbrompheniramine. Both pentazocine and tripelennamine were qualitatively confirmed by their electron impact mass spectra. Concentrations of pentazocine and tripelennamine in various fluids and tissues were determined.

[FAB (fast atom bombardment)-mass spectrometry. A new study method in the hands of the (forensic) toxicologist]. / FAB (Fast Atom Bombardment)-Massenspektrometrie. Eine neue Untersuchungsmethode in der Hand des (forensischen) Toxikologen.

The FAB (Fast Atom Bombardment)-mass spectrometric ionization technique, which has now been available for about 1 year, has been successfully employed in forensic toxicology. The mass spectral behaviour of some representative drug-glucuronides (Codeine, p-Nitrophenol and 2-Phenyl-1-propanol) were studied by positive- and negative-ion-FAB-MS. The presented promising results may be of some interest, not only for the analytical toxicologist.

Drugs of abuse radioimmunoassay directory.

This clinical brief is a survey of procedures, components and kits presently available for detecting and quantitating drugs of abuse in biologic fluids by radioimmunoassay (RIA).