A case report on fatal intoxication by tapentadol: Study of distribution and metabolism.

We report a case in which a tapentadol acute intoxication was suspected as the cause of death of a 39-year-old man: approximately two days after death, cardiac and femoral blood, as well as urine, bile, gastric content and chest hair, were collected during the autopsy. Tapentadol was detected before and after hydrolysis in femoral (530 ng/mL unconjugated and 1570 ng/mL conjugated) and cardiac (680 ng/mL unconjugated and 3440 ng/mL conjugated) blood, and additionally in bile (3200 ng/mL), urine (9300 ng/mL), chest hair (2850 pg/mg) and gastric content. LC-QTOF screening analysis confirmed the presence of five different tapentadol metabolites (tapentadol-O-glucuronide, tapentadol-O-sulfate, N-desmethyltapentadol, N-desmethyltapentadol-glucuronide and N-desmethyltapentadol-O-sulfate), in urine, bile, cardiac and femoral blood. Positivity of body hairs allowed us to conclude that the man had used tapentadol in the last weeks/months. Autopsy and toxicological results (also positive for clotiapine, diazepam and chlordesmethyldiazepam) suggested that tapentadol could have caused, even at low concentrations, a severe respiratory depression, which contributed to the death of the subject. This is one of the few cases in literature where tapentadol was detected in blood, together with its metabolites, and the only one in which the parent drug was identified in hairs.

Determination of 105 antibiotic, anti-inflammatory, antiparasitic agents and tranquilizers by LC-MS/MS based on an acidic QuEChERS-like extraction.

A procedure for screening 105 veterinary drugs in foods by liquid chromatography tandem mass-spectrometry (LC-MS/MS) is presented. Its scope encompasses raw materials of animal origin (milk, meat, fish, egg and fat) but also related processed ingredients and finished products commonly used and manufactured by food business operators. Due to the complexity of the matrices considered and to efficiently deal with losses during extraction and matrix effects during MS source ionisation, each sample was analysed twice, that is 'unspiked' and 'spiked at the screening target concentration' using a QuEChERS-like extraction. The entire procedure was validated according to the European Community Reference Laboratories Residues Guidelines. False-negative and false-positive rates were below 5% for all veterinary drugs whatever the food matrix. Effectiveness of the procedure was further demonstrated through participation to five proficiency tests and its ruggedness demonstrated in quality control operations by a second laboratory.

[Determination of three tranquillizer residues in animal foods by QuEChERS-high performance liquid chromatography-tandem mass spectrometry].

A method for simultaneous determination of chlorpromazine, diazepam and metolazone residues in porcine muscle, fish, liver and kidney was developed using QuEChERS and HPLC-MS/MS technique. The samples were extracted with ethyl acetate and cleaned up with C18, N-propylethylendiamine (PSA) and NH2 sorbents after using Na2SO4 as dehydrating agent. The analytes were separated by a special C18 column, Atlantis T3, and gradiently eluted with a mixed solution of 5 mmol/L formic acid and acetonitrile at a flow rate of 0.35 mL/min. The mass spectrometric analysis that quantified using isotope internal standard, was carried out with electrospray positive ion source (ESI+) and multiple reaction monitoring mode (MRM). The linearity of the calibration curves was good in the range of 0.2-5.0 µ g/L. The recoveries at three different spiked levels (0.5, 1 and 5 µ g/kg) in four matrices were in the range of 92.5%-117.8%. The repeatability expressed as relative standard deviations (RSDs) ranged from 0.7% to 11.6% (n=6). The method, with wide matrix range of application, is highly effective and sensitive and suitable for the rapid analysis of large quantities of samples.

Subdose de acepromazina no acuponto yin tang para tranquilização de cães / Acepromazine subdose in yin tang acupoint for dog tranquilization / Subdosis de acepromacina en el punto yin tang para tranquilizar perros

Foi realizado um estudo para investigar a tranquilização de cães com subdose de acepromazina no acuponto yin tang, localizado no ponto médio de uma linha traçada entre os cantos laterais dos olhos. O estudo foi delineado em quatro protocolos, utilizando-se oito cães. No primeiro protocolo (P1) foi administrada acepromazina no yin tang em subdose (0,01mg/ kg). No segundo protocolo (P2) foi administrada a mesma dose utilizada em P1 por via intramuscular (IM). No terceiro protocolo (P3) foi administrada dose terapêutica (0,1mg/kg) IM. No quarto protocolo (P4) foi colocada uma agulha de acupuntura no yin tang. Durante as etapas foram aferidos: frequência cardíaca (FC), frequência respiratória (FR) e temperatura retal (TR). Tais aferições foram realizadas antes dos tratamentos, quinze minutos após a administração do fármaco e de quinze em quinze minutos até duas horas. Nos mesmos momentos foi pesquisada a presença ou ausência de decúbito, sonolência, ptose palpebral e outros sinais de tranquilização. Os dados foram analisados estatisticamente. No tratamento experimental (P1) dois cães apresentaram tranquilização satisfatória e um moderada, sem diferença significativa com o grupo no qual se administrou acepromazina em dose terapêutica IM. Concluiu-se que a administração de acepromazina em subdose no acuponto Yin Tang pode ser usada com segurança na rotina clínica nas mais diversas manipulações nas quais haja necessidade de tranquilização de cães...

A study was conducted to investigate the tranquilization of dogs using a subdose of acepromazine in the yin tang acupoint located in the medium point of a line drawn between the lateral corners of the eyes. The study was designed in four protocols using eight dogs. In the first protocol (P1), acepromazine was administered in yin tang in subdose (0.01mg/kg). In the second protocol (P2), the same dose as in P1 was administered by intramuscular route (IM). In the third protocol (P3), a therapeutic dose (0.1 mg/kg) was administered IM. In the fourth protocol (P4), an acupuncture needle was placed in the yin tang point. During the steps, heart rate (HR), respiratory rate (RR) and rectal temperature (RT) were measured. These evaluations were performed before treatment, fifteen minutes after drug administration, and every fifteen minutes for two hours. At the same moments, the presence or absence of decubitus, drowsiness, ptosis and other signs of tranquilization were observed. Data were statistically analyzed. In the experimental treatment (P1), one dog showed moderate tranquilization, and two dogs showed satisfactory tranquilization, without statistical difference from the group in which acepromazine was administered IM in the therapeutic dose. It can be concluded that the administration of acepromazine in subdose in the yin tang acupoint can be safely used in clinical practice, in various manipulations in which the need of tranquilization of dogs is necessary...

Se ha realizado un estudio para investigar como tranquilizar perros con acepromacina en subdosis en el punto de acupuntura yin tang, que se encuentra en el punto medio de una línea trazada entre las bordas laterales de los ojos. El estudio ha sido diseñado en cuatro protocolos utilizando ocho perros. En el primer protocolo (P1) se administró acepromacina en el punto yin tang en subdosis (0,01 mg / kg). En el segundo protocolo (P2), se administró la misma dosis usada en P1 por vía intramuscular (IM). En el tercer protocolo (P3), se administró dosis terapéutica (0,1 mg / kg) IM. En el cuarto protocolo (P4), se ha puesto una aguja de acupuntura en el punto yin tang. Durante las etapas se midieron: frecuencia cardíaca (FC), frecuencia respiratoria (FR) y la temperatura rectal (TR). Estas evaluaciones se realizaron antes del tratamiento, quince minutos después de la administración del fármaco, y a cada quince minutos durante dos horas. En los mismos momentos se ha investigado la presencia o ausencia de decúbito, somnolencia, ptosis de pálpebra y otras señales de tranquilidad. Los datos han sido analizados estadísticamente. En el tratamiento experimental (P1), dos perros mostraron tranquilidad satisfactoria y un perro moderada, lo que no fue significativamente diferente del grupo donde se administró dosis terapéutica de acepromacina por vía intramuscular. Se concluyó que la administración de acepromacina en subdosis en el punto yin tang puede ser utilizado con seguridad en la práctica clínica en las más diversas manipulaciones en las que hay la necesidad de tranquilizar perros...

Development and validation of a confirmatory method for the determination of tranquilisers and a ß-blocker in porcine and bovine kidney by LC-MS/MS.

A method for the confirmation of the tranquilisers azaperone and its metabolite azaperol, of chlorpromazine, propionylpromazine and acepromazine, haloperidol, xylazine and the ß-blocker carazolol in porcine and bovine kidney with LC-MS/MS was developed. Various solvents and extraction techniques and different ways of clean-up were tested and verified with incurred pig kidney from an animal study. The best extraction efficiency was reached by extraction with a mixture of ammonia in ethyl acetate and overhead shaking without SPE clean-up. The optimised method with a twofold sample extraction was validated according to Commission Decision 2002/657/EC in a matrix-comprehensive in-house validation supported by the InterVal software, taking into account the defined MRLs and recommended concentrations. The validated method can be used for residue control of the above-mentioned analytes in bovine and porcine kidney of food-producing animals in accordance with Council Directive 96/23/EC.

Chemiluminescence and electrochemiluminescence detection of controlled drugs.

We review the determination of various controlled drugs (opioids, tranquilizers, stimulants, and hallucinogens) using flow-analysis methodologies (flow injection analysis, high performance liquid chromatography, capillary electrophoresis, and microfluidic devices) with chemiluminescence and electrochemiluminescence reagents such as luminol, diaryloxalates, tris(2,2'-bipyridine)ruthenium(II), permanganate, manganese(IV), and sulfite, for industrial, clinical, pharmaceutical, and forensic science applications.

Rapid method for the determination of tranquilizers and a beta-blocker in porcine and bovine kidney by liquid chromatography with tandem mass spectrometry.

A fast and simple liquid chromatography with tandem mass spectrometry method for detection and confirmation of tranquilizers (chlorpromazine, propionylpromazine, acepromazine, triflupromazine, promazine, azaperone and its metabolite, azaperol) and beta-blocker (carazolol) in porcine and bovine kidney has been presented. The method relies on the extraction with acetonitrile followed by centrifugation. After evaporation of acetonitrile, the residue was reconstituted in a mobile phase and filtrated. The separation of analytes was performed on a C18 column using a mobile phase of acetonitrile and ammonium formate buffer (0.05 M, pH 4.5) with gradient elution. The electrospray ionization was used to obtain the protonated molecules [M+H](+) and two product ions were monitored for each compound. For quantification deutered internal standards were used. The whole method has been validated according to the European Union requirements. Specificity, decision limit (CCalpha), detection capability (CCbeta), trueness and precision were determined. The results showed good trueness ranged from 73.2% to 110.6% with a good R.S.D., less than 13.0% under within-laboratory reproducibility conditions. The calculated critical concentrations of CCalpha for phenothiazines were between 5.8 and 6.6 microgkg(-1) while for azaperone CCalpha was 105.5 microgkg(-1) and for azaperol was 121.4 microgkg(-1). CCalpha for carazolol was 16.7 microgkg(-1) in bovine and 21.9 microgkg(-1) in porcine kidney. CCbeta for phenothiazines were between 6.3 and 7.6 microgkg(-1), for azaperone was 119.0 microgkg(-1) and for azaperol was 140.0 microgkg(-1). For carazolol in bovine kidney CCbeta was 18.6 microgkg(-1) whereas in porcine kidney was 24.4 microgkg(-1).

Fate of beta blockers and psycho-active drugs in conventional wastewater treatment.

The removal of beta blockers and psycho-active drugs was investigated in a representative conventional German WWTP by long-term measurement campaigns along different biological treatment processes. The activated sludge treatment with an elevated SRT of 18 d was the only process which led to a significant removal of certain beta blockers and psycho-active drugs. The removal efficiency was below 60% for all compounds except for the natural opium alkaloids codeine and morphine being removed by more than 80%. Primary biological transformation and sorption onto sludge as the main removal mechanisms were examined in lab-scale batch experiments. Sorption onto activated sludge was found to be negligible (<3%). The biological transformation could be described by pseudo-first order kinetics and the transformation constants k(biol) were used to predict the removal of beta blockers and psycho-active drugs in an activated sludge unit with a model. For most compounds the removal efficiencies measured on the full-scale WWTP were within the 95% confidence intervals predicted by the model. The results from full-scale measurements and modeling indicate that biological transformation in the nitrification tank together with parameters such as the sludge retention time and the temperature is crucial regarding the biological transformation of beta blockers and psycho-active drugs in conventional WWTPs.

Benzodiazepines: sample preparation and HPLC methods for their determination in biological samples.

Benzodiazepines (BDZs) belong to a group of substances known for their sedative, antidepressive, muscle relaxant, tranquilizer, hypnotic and anticonvulsant properties. Their determination in biological fluids is essential in clinical assays as well as in forensics and toxicological studies. Researchers focus on the development of rapid, accurate, precise and sensitive methods for the determination of BDZs and their metabolites. A large number of analytical methods using different techniques have been reported, but none can be considered as the method of choice. BDZs are usually present at trace levels (microgram or nanogram per milliliter) in a complex biological matrix and the potentially interfering compounds must be isolated by various extraction techniques before analysis. An extended and comprehensive review is presented herein, focusing on sample preparation (pretreatment and extraction) and HPLC conditions applied by different authors. These methods enable bioanalysts to achieve detection limits down to 1-2 ng/ml using UV/diode array detection, readily available in most laboratories, and better than 1 ng/ml using electron capture detection, which is lower than that obtained using a nitrogen phosphorus detector. MS interfaced with electrospray ionization offered a similar sensitivity, while negative chemical ionization MS or sonic spray ionization MS provided sensitivity down to 0.1 ng/ml.

Determination of residues of azaperone in the kidneys by liquid chromatography with fluorescence detection.

An analytical method has been developed for the quantitative determination of residues of the tranquillizer azaperone (AZN) in the kidneys of slaughtered animals. Samples were extracted with acetonitrile, extracts were acidified and further purified with solid-phase extraction (SPE) on a polymeric mixed-mode cation-exchange sorbent, Oasis. AZN and its main metabolite azaperol (AZL) were eluted by alkaline methanol (MeOH), the eluate was evaporated, re-dissolved and analysed by gradient high performance liquid chromatography (LC) on reversed and deactivated phase LiChrospher 60-RP select B at excitation and emission wavelengths of 245 and 345 nm, respectively. The method was validated according to the requirements of European Commission Decision 2002/657/EC, using fortified porcine kidneys. The method proved to be selective, specific against carazolol (CAR) and linear over a concentration range 10-150 microg kg(-1) (r2>0.99). Over a concentration range 50-150 microg kg(-1), mean recovery of AZN and AZL was 88.2 and 91.2%, respectively, with intra-laboratory reproducibility of <11.0 and <9.0%, respectively. The decision limit (CCalpha) of AZN and AZL was 112 and 111 microg kg(-1), respectively, and the limit of quantification (LOQ) was 10 and 5 microg kg(-1), respectively. The procedure was also applied to bovine, poultry and horse kidneys, giving similar results, and has been successfully implemented in statutory residue monitoring control in food of animal origin in Slovenia.

Multiresidue method for the detection of tranquillisers, xylazine, and a beta-blocker in animal production by liquid chromatography-tandem mass spectrometry.

Tranquillisers are often used in animal production, especially in pigs to calm them before transport to the slaughterhouse. The use of certain substances (derived from phenothiazine) is totally prohibited, whilst other compounds (butyrophenone and beta-blockers) are regulated through the establishment of MRLs. A physico-chemical detection method based on liquid chromatography-tandem mass spectrometry is described. Validation was carried out according to the criteria laid down in Directive 2002/657/EC. The method was also used to detect and quantify these substances in treated animals.

Analysis of certain tranquilizers in biological fluids.

A review with 282 references is presented that deals with the reported methods of analysis of phenothiazines, thioxanthenes, and benzodiazepine derivatives of pharmaceutical interest. The review includes the methods adapted in biological fluids.

Multiresidue analysis of tranquilizers and the beta-blocker Carazolol in meat by liquid chromatography/tandem mass spectrometry.

A fast and simple method for the quantification of a number of tranquilizers and the beta-blocker Carazolol in pork and bovine kidney is described. Extracts are purified/concentrated by a solid phase extraction step and separated on a reversed phase column with an alkaline (ammonia) acetonitrile gradient. The electrospray tandem mass spectrometer is operated in positive ion multireaction monitoring mode. Resulting chromatograms are free of interfering peaks. The recovery is >75% for all analytes and the limit of detection <1 ppb, which is well below the current maximum residue limit for the various compounds.

Evaluation of certain veterinary drug residues in food.

This report presents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of residues of certain veterinary drugs and to recommend maximum levels for such residues. The first part of the report considers an approach to assessing the safety of antimicrobial drug residues and their effects on the human intestinal microflora and general issues relating to the recommendation of Maximum Residue Levels (MRLs) for veterinary drug residues in food. A summary follows of the Committee's evaluations of toxicological and residue data on a variety of veterinary drugs: one beta-adrenoceptor-blocking agent (carazolol), one anthelminthic agent (doramectin), four antimicrobial agents (dihydrostreptomycin, streptomycin, neomycin and thiamphenicol), two insecticides (deltamethrin and phoxim), four production aids (estradiol-17 beta, progesterone, testosterone and porcine somatotropins) and one tranquillizing agent (azaperone). Annexed to the report are a summary of the Committee's recommendations on these drugs, including Acceptable Daily Intakes and MRLs, and other information required.

Determination of propionylpromazine hydrochloride in formulation matrixes using reversed-phase ion-pair small bore liquid chromatography.

Propionylpromazine hydrochloride (PPZHCl) has been investigated for use with leghold traps to reduce the amount of self-inflicted trauma experienced by animals restrained by these traps. Three types of PPZHCl formulations made with Karo dark syrup, K-Y Jelly, and Vaseline were used in 2 types of tranquilizer trap devices (TTDs). A reversed-phase ion-pair liquid chromatography (LC) method using a small bore C18 column was used to: (1) determine the purity of the PPZHCl material used in these formulations, and (2) to determine the resulting PPZHCl content of each formulation. Analyte quantitation was done using UV absorption at 280 nm. Regression analysis of calibration standard solutions indicated a linear and directly proportional relationship between analyte response and PPZHCl concentration over the range evaluated. Recovery data from: (1) Vaseline formulations containing 38.8, 16.2, and 8.78% PPZHCl were 104, 92.9, and 90.2%, respectively, (2) Karo dark syrup formulations containing 26.5, 18.1, and 10.3% PPZHCl were 97.7, 99.3, and 106%, respectively, and (3) K-Y Jelly formulations containing 33.0, 23.5, and 13.4% PPZHCl were 100, 99.4, and 88.7%, respectively. The relative standard deviation (RSD) values from triplicate analysis of these formulations ranged from 0.7 to 6.7%. The PPZHCl content from 9 manufactured TTDs, 3 for each formulation type, were analyzed in triplicate and produced RSD values ranging from 0.7-6.8%. These results indicate that the formulation extraction presented could be used to evaluate the PPZHCl content in TTDs prior to field use. The use of a small bore LC column reduced the amount of solvents consumed and hazardous waste generated, compared to sample analysis that uses a more conventional analytical LC column.

Multi-residue analysis of tranquillizers in meat: confirmatory assays using mass spectrometry.

A rapid and sensitive multi-residue method was developed to attempt to confirm the presence of the beta-blocker carazolol and the tranquillizers acepromazine, azaperone, chlorpromazine, propionylpromazine and xylazine in pig muscle tissues. The procedure involves determination by liquid chromatography coupled with tandem mass spectrometry. The liquid chromatographic separation was performed on a Symmetry C18 column with gradient elution. A mixture of aqueous buffer, containing 0.01% m/v trifluoroacetic acid (pH 3.5), and acetonitrile at a flow rate of 0.4 ml min-1 was used as the mobile phase. The abundant parent ions [M+ H+] produced by positive electrospray ionisation were selected for collisional dissociation with argon. Fragment ions were recorded with daughter ion scan and multiple reaction monitoring. The analytes were identified unambiguously by assessing retention times and diagnostic ions in meat samples spiked from 50 micrograms kg-1 [maximum residue limit (MRL) for azaperone and azaperol] to 5 micrograms kg-1 (MRL for carazolol).

Problems associated with drug residues in beef from feeds and therapy.

Drug residues in beef have been reported internationally. These include antimicrobials, anti-inflammatories, growth promotants, parasiticides and insecticides. The main factors associated with residues are animal age and use, and failure to observe withdrawal time for regular or extra-label use. Public health concerns include toxic and anaphylactic reactions, and development of drug-resistant strains of bacteria. The maximum residue level (MRL) is the current standard for residues in food adopted by the Codex Committees of the Food and Agriculture Organisation and World Health Organisation, but is not universally accepted or standardised. Detection of residues at slaughter is a critical point in residue control. Several live animal tests are available, but these vary in reliability and usage. After slaughter, tissues sampled and tests used are more uniform. To prevent international trade barriers associated with drug residues in beef, the following conditions should be implemented: standardisation of testing methods used to detect drug residues; standardisation of methods for determining MRLs; establishment of active surveillance programmes to monitor residues.

Analysis of toxic drugs by capillary electrophoresis using polyacrylamide-coated columns.

Toxic drugs, including barbiturates and benzodiazepines, were analyzed using polyacrylamide-coated columns in capillary electrophoresis (CE). The separations were carried out in absence of electroosmotic flow. Seven kinds of barbiturates were successfully separated with the coated columns in free solution without further additives. Benzodiazepines, the electrically neutral solutes were introduced onto the coated column, and separated in presence of sodium dodecyl sulfate above its critical micelle concentration in the running buffer. This CE method offered fast and efficient separation of more hydrophobic solutes, such as benzodiazepines. The separation of seven barbiturates was studied in linear (noncross-linked) polyacrylamide solutions and in acrylamide/N-isopropylacrylamide (AA/IPAA) copolymer solutions to explore the effect of isopropyl groups in the AA/IPAA copolymer chain.

Potential interference of cyclobenzaprine and norcyclobenzaprine with HPLC measurement of amitriptyline and nortriptyline: resolution by GC-MS analysis.

Cyclobenzaprine and its major metabolite, norcyclobenzaprine, differ from amitriptyline and nortriptyline only by the presence of a double bond in the cycloheptane ring. Three patients developed sufficient levels of cyclobenzaprine and norcyclobenzaprine because of either rapid or long-term ingestion of cyclobenzaprine to cause positive interferences in both a Syva EMIT assay and a high-performance liquid chromatographic (HPLC) assay for identification and quantitation of tricyclic antidepressants in serum. Cyclobenzaprine coeluted with amitriptyline, and norcyclobenzaprine eluted slightly earlier than, but was poorly resolved from, nortriptyline in this HPLC assay. We found that cyclobenzaprine could be distinguished from amitriptyline and that norcyclobenzaprine could be distinguished from nortriptyline on the basis of gas chromatographic retention times upon gas chromatographic-mass spectrometric analyses after derivatization with trifluoroacetic anhydride. The compounds were also distinguishable by mass spectrometric criteria.

[Determination of psoralen and isopsoralen in tincture of fructus Psoraleae by HPLC].

A reversed-phase HPLC method was described for the determination of psoralen and isopsoralen in tincture of Fructus Psoraleae. After adding the internal standard and diluting the sample to a definite volume, the sample was injected and analysed. The RSDs were within 2% and the minimum working concentrations below 1 microgram/ml; the average recoveries of psoralen and isopsoralen 102.8% and 99.5% respectively and the linear ranges of psoralen and isopsoralen 12-200 micrograms/ml and 13-210 micrograms/ml (r = 0.999) respectively.