Münchausen Syndrome as an Unusual Cause of Pseudo-resistant Hypertension: A Case Report.

Münchausen syndrome can be characterized by simulated illness, pathological lying and wandering from place to place (the patient typically presents to numerous hospitals). Individuals with elevated blood pressure due to non-adherence to medication have the so-called pseudo-resistant hypertension. A 45-year-old woman was admitted to hospital on an emergency basis because of a hypertensive crisis. Despite combination antihypertensive treatment, normalization of blood pressure was not achieved and a device to produce a therapeutic arteriovenous fi stula was implanted. Aft er the procedure, a signifi cant increase in pulmonary artery pressure was observed and closure of the fistula was performed by implantation of the stent graft . The suspicion was raised that the patient had not been taking her prescribed medications. Therefore, blood samples were taken and the serum was analyzed for presence of the prescribed drugs (atorvastatin, bisoprolol, chlorthalidone, clonidine, doxazosin, furosemide, nitrendipine, oxazepam and valsartan). The results confirmed suspected failure of the patient to take the prescribed medications. Münchausen syndrome is usually first suspected when inexplicable laboratory test results are noted. To our knowledge, this is the first reported case of Münchausen syndrome with pseudo-resistant hypertension leading to the implantation of a device to produce a therapeutic arteriovenous fi stula.

Detection of legal highs in the urine of methadone-treated patient by LC-MS.

Urine tests are the commonly accepted methods to control abstinence and adherence to treatment of patients who undergo methadone maintenance treatment (MMT). Depending on various national guidelines and accessibility of techniques, only selected psychoactive substances are routinely tested in urine of MMT patients. In general, they belong to the few groups of compounds: THC, cocaine, amphetamines, opiates, PCP and benzodiazepines. It is, however, well known that patients enrolled in such replacement programmes take psychoactive substances that are not routinely detected by the toxicology laboratories, to escape unexpected tests. Here, we report semiquantitative detection of legal highs taken by the MMT patient, using high-pressure liquid chromatography coupled to the flowing atmospheric pressure afterglow ion source (LC-FAPA-MS). To demonstrate effectivity of this technique, the data were confirmed by quantitative analysis using LC-ESI-MS/MS. In the analysed sample of MMT patient, a mixture of psychoactive compounds was found, namely 3-MMC (3-methylmethcathinone), pentedrone and methcathinone and determined at the concentrations of 670; 50 and 0.2 µg/mL, respectively. Such fast analytical technique may be useful for the efficient control of substances taken intentionally by MMT patients.

Severe clinical toxicity caused by 25I-NBOMe confirmed analytically using LC-MS-MS method.

Rhabdomyolysis is a relatively rare, but potentially serious complication of various diseases. Muscular injury and resultant release of electrolytes, myoglobin and other enzymatic proteins e.g. creatine kinase (CK) into circulation may result in multi-organ failure requiring an extensive treatment. Non-traumatic causes of rhabdomyolysis, like poisonings, appear to be much more frequent than traumatic causes. We present the case of a patient who developed exceptionally massive rhabdomyolysis, with CK up to 516 455 U/l, after ingestion of a relatively small dose of a novel psychoactive substance known as "Alice in Wonderland". Laboratory quantification was performed using a validated LC-MS/MS method in a whole blood sample.

Acidity of substituted cathinones studied by capillary electrophoresis using the standard and fast alternative approaches.

Cathinone derivatives are notorious but still weakly characterized molecules, known mainly as components of the designer and illicit drugs. The knowledge on their acidity is scarce and incomplete, therefore, we decided to determine the pKa values for six of them: 2-methylmethcathinone, 3-methylmethcathinone, 4-methylmethcathinone, α-pyrrolidinovalerophenone, methylenedioxypyrovalerone and ephedrone. For that purpose we employed capillary electrophoresis, which is known for its accurateness in comparison to other analytical techniques. We used and compared two methodologically different approaches. The standard method relied on measuring electrophoretic mobility across the broad pH range and fitting the sigmoidal function. The obtained pKa values were in the range 8.59-9.10, thus these molecules remain mostly protonated and positively ionized in the physiological conditions. The alternative two-values (TVM) and one-value methods (OVM), proposed by us previously, have been used herein for the first time to the cationic molecules. TVM enables estimation of pKa using only two electrophoretic mobility values, referring to the total and partial ionization. OVM requires only a single measurement because mobility of ion is predicted theoretically. Both TVM and OVM yielded only a small deviation of pKa from the standard approach, averagely 0.07-0.09pH unit. Two important issues have also been addressed: the potential of a maximally fast calculation method using no repetition at the given pH, and the accuracy of method with regard to pH attributed to partial ionization. As a whole, the analytical potential of the TVM/OVM approach seems to be huge and invaluable for fast pKa screening/estimation.

Assessment of adherence to treatment in patients with resistant hypertension using toxicological serum analysis. A subgroup evaluation of the RESIST-POL study.

INTRODUCTION: Nonadherence to antihypertensive therapy is one of the main causes of resistant hypertension. OBJECTIVES: The aim of our study was to evaluate adherence to therapy in patients with resistant hypertension by determining serum antihypertensive drug levels with the use of liquid chromatography-tandem mass spectrometry (LC-MS/MS). PATIENTS AND METHODS: The study included 36 patients with primary resistant hypertension selected from the RESIST-POL study (23 men and 13 women; mean age, 52.5 ±9.1 years; range, 22-67 years; mean number of antihypertensive drugs, 5.3 ±1.4), who met all 3 inclusion criteria: use of ≥4 antihypertensive drugs; average daytime ambulatory systolic blood pressure ≥140 mmHg; one of the clinical features suggesting nonadherence. All patients had their serum drug levels assessed using LC-MS/MS. Patients in whom the serum level of at least 1 drug was below the limit of quantification for the method used were regarded as nonadherent. RESULTS: Of all study patients, nonadherence was observed in 31 patients (86.1%), and none of the prescribed drugs was detected (complete nonadherence) in 5 patients (13.9%). In 26 patients (72.2%), at least 1 of the prescribed drugs could not be detected (partial nonadherence). CONCLUSIONS: In our study, we documented a surprisingly low adherence to antihypertensive treatment in patients with resistant hypertension. Our results suggest that, particularly in those patients, the analysis of serum antihypertensive drug levels using LC-MS/MS might allow to avoid a comprehensive and costly diagnostic work-up including biochemical and imaging studies.

Acute intoxication due to tert-amyl alcohol--a case report.

We presented a case of 28 year-old male, who was found in a deep coma complicated with acute respiratory failure because of recreational intoxication with tert-amyl alcohol (TAA). The TAA blood level at the admission was 83 µg/mL determined by gas chromatography-mass spectrometry (GC-MS). In the last few months popularity of TAA among alcohol and drug addicted people in Europe is still growing. The main reasons of these are: self-healing of addiction, low price of this xenobiotic compare to alcohol, and problem to detect this xenobiotic in generally used screening tests.

[Resurgence in paramethoxamphetamine use. Review of poisonings and two recent cases study]. / Powrót do uzywania parametoksyamfetaminy. Przeglad zatruc i studium dwóch nowych przypadków.

Two recent cases of death due to p-methoxyamphetamine (PAM) intake, a methoxylated phenethylamine derivative, were described and compared with previous PMA related deaths that occurred in many countries. Following a review of the available literature from 1974 to 2011 three periods of resurgence of an unsuspected increase of lethal PMA causation were considered. Signs of intoxications, concentrations of PMA found in biological materials were discussed. Based on the case reports can be concluded the great number of victims were unconscious of taking PMA as substitute of MDMA. Two new methods for screening, identification and quantification of amphetamine derivatives in biosamples (blood and urine) using LC-MS/MS techniques were developed. The methods have proven to be appropriate for clinical and forensic toxicology purposes.

Simultaneous screening for and determination of 128 date-rape drugs in urine by gas chromatography-electron ionization-mass spectrometry.

Date-rape drugs (DRDs) are used for the purpose of "drugging" unsuspected victims and raping or robbing them while under the influence of the drug. The wide variety of substances used for criminal purposes, their low concentrations in body fluids and, often, a long time delay between the event and clinical examination make comprehensive screening analysis of biological materials collected from crime victims for the presence of these drugs very difficult. Detection of a drug used to facilitate sexual assault in biological fluids can be very important evidence of a committed crime. The purpose of this study was to develop a simple GC-EI-MS screening procedure for date-rape drugs in urine. Target analytes were isolated by solid-phase extraction. 2-mL urine samples were extracted and then derivatized by using BSTFA+1%TMCS reagent. Detection of all compounds was based on full-scan mass spectra and for each compound one ion was chosen for further quantification. The method allowed the simultaneous screening, detection and quantification of 128 compounds from different groups (number of compounds): opioids (20), amphetamines (11), GHB and related products (3), hallucinogens (9), benzodiazepines (18), antihistamines (9), antidepressants (14), selective serotonin-reuptake inhibitors (4), antipsychotics (7), barbiturates (7), other sedatives (5), muscle relaxants (2) and other drugs (19). The procedure can easily be expanded to encompass more substances. The developed method appeared to be suitable for screening for the target DRDs. The procedure was successfully applied to the analysis of authentic urine samples collected from victims of rapes and other crimes in routine casework.

[Drugs scene in Poland from a forensic toxicologist's view]. / Scena narkotykowa w Polsce z punktu widzenia toksykologa sadowego.

This paper reviews the current use agents influencing the central nervous system. Among them are new synthetic psychoactive drugs, "legal highs" (herbal and synthetic), anabolic steroids, untypical adulterants to classic drugs of abuse, unusual mixtures, dietary supplements, energising drinks and substances similarly acting to alcohol. All these agents were characterized regarding to their acting on human organism, impairing on driving ability and legal regulations that means the Drug Addiction Counteraction Act of 29 July, 2005, the Road Traffic Act of 20 June, 1997 and the Ministry of Health Decree concerning the list of substances acting similarly to alcohol, and the conditions and the methods for carrying out examinations for the presence of these substances in the human body of 11 June 2003. The variety of abused, used and misused agents leads to the application of very sensitive analytical methods of their detection (screening), identification (confirmation) and quantification. Attention has been drawn to some specific difficulties caused currently by the identification of active components of "legal highs" in biological fluids, which influences the diagnosis of poisoning by these substances. The inconsistence of legal regulations allowing for arbitrary interpretation of the results of toxicological analyses concerning even forensic purposes has been critically discussed. It has been proposed that the manner of legal controlling of new psychoactive agents should be changed.

Stress-opioid interactions: a comparison of morphine and methadone.

The utility of methadone and morphine for analgesia and of methadone for substitution therapy for heroin addiction is a consequence of these drugs acting as opioid receptor agonists.We compared the cataleptogenic and antinociceptive effects of single subcutaneous doses of methadone hydrochloride (1-4 mg/kg) and morphine sulfate (2.5-10 mg/kg) using catalepsy and hot-plate tests, and examined the effects of the highest doses of the drugs on Fos protein expression in selected brain regions in male Sprague-Dawley rats. Methadone had greater cataleptogenic and analgesic potency than morphine. Fos immunohistochemistry revealed substantial effects on the Fos response of both the stress induced by the experimental procedures and of the drug exposure itself. There were three response patterns identified: 1) drug exposure, but not stress, significantly elevated Fos-positive cell counts in the caudate-putamen; 2) stress alone and stress combined with drug exposure similarly elevated Fos-positive cell counts in the nucleus accumbens and cingulate cortex; and 3) methadone and morphine (to a lesser extent) counteracted the stimulatory effect of nonpharmacological stressors on Fos protein expression in the somatosensory cortex barrel field, and Fos-positive cell counts in this region correlated negatively with both the duration of catalepsy and the latency time in the hot-plate test. The overlap between brain regions reacting to nonpharmacological stressors and those responding to exogenous opioids suggests that stress contributes to opioid-induced neuronal activation.

[Determination of fentanyl, atropine and scopolamine in biological material using LC-MS/APCI methods]. / Oznaczanie fentanylu, atropiny i skopolaminy w materiale biologicznym metodami LC-MS/APCI.

In this paper methods for determination of fentanyl (FL) and its three analogues: alfentanyl (AL), sufentanyl (SL) and remifentanyl (RL), atropine (AT) and scopolamine (SK) in biological material (whole blood and urine) using liquid chromatography atmospheric pressure chemical ionisation mass spectrometry technique (LC-MS/ APCI) are presented. Separation of analytes was performed in gradient conditions, using a LiChroCART LiChrospher 60 RP-select B column. The mobile phase consisted of 0.1% (v/v) formic acid in water and in acetonitrile. Target analytes were isolated from biological matrices using liquid-liquid extraction technique with n-butyl chloride or diethyl ether as extraction solvents. The validation data of the methods were: limit of detection (LOD, S/N = 3) and limit of quantification (LOQ, S/N = 10) - 0.05 and 0.25 ng/ml for FL, and 0.7 and 0.9 ng/ml for AT, both in blood, whereas 1.9 and 2.1 ng/ ml for FL, and 0.6 and 0.9 ng/ml for AT in urine. Calibration curves showed linearity in concentration ranges from LOQ to 25 ng/ml in blood and from LOQ to 50 ng/ml in urine. Determination coefficients (R2) of linear regression equation were higher then 0.98. Extraction recovery, intra-day precision (CV(w.g.)) and inter-day precision (CV(m.g.)) were determined at analytes and internal standard (I.S.) concentration of 5 ng/ml for blood, and at analytes and I.S. concentrations of 20 and 5 ng/ml, respectively for urine. Extraction recovery ranged from 76 to 100% for blood and 53--72% for urine. CV (n=5) and CV(m.g.) (n=15) equal from 4.8 to 7.5% and from 6.8 to 16.2% respectively for blood, and from 4.3 to 5.4% and from 5.8 to 9.5% for urine. The application of elaborated methods of the determination of FL, AT and SK in blood and urine for 8 expert opinions elaborated at the Institute of Forensic Research in Krakow is described. FL was detected and quantified in 3 cases, whereas AT and SK in 7.

Date-rape drugs scene in Poland.

Since the beginnings of twenty-first century in Poland increasing number of reports about the drug-facilitated sexual assaults have been observed. Many drugs have been identified as so-called "date-rape drugs", because of their pharmacological properties, especially inducing amnesia. These drugs are used for the purpose of "drugging" unsuspected victims and than raping them. In a typical scenario, the perpetrator surreptitiously adds "date-rape drug" to the alcoholic or non-alcoholic beverage of an unsuspecting person, who is subsequently sexually assaulted while under the influence of this substance. Many victims do not report the incident until several days after the event or even do not report it at all. They report the incident so late after the events because they often have problems with remember the course of incident. It causes that victim is not reliable witness for justice. Detection of "date-rape drugs" in biological fluids is unequivocal evidence of perpetration. Analysis of biological fluids collected from victims of rapes for presence of drugs was rare in Poland up to now. The aim of this study is to show the use of "date-rape drugs" in Poland. Materials for this study were from the routine casework elaborated at the Institute of Forensic Research in Kraków. APCI-LC-MS methods were applied for screening of biological fluids (blood and/or urine) for amphetamine and its 6 analogues, for 12 substances from benzodiazepine group and for quantification of the detected drugs. HPLC-DAD was used as a screening method for wide range of medicinal drugs, and NCI-GC-MS methods for determination of ketamine and tetrahydrocannabinols (delta9-tetrahydrocannabinol, 9THC) and its metabolite (11-nor-carboxy-delta9-tetrahydrocannabinol, THCCOOH). In 2000-2004, the biological fluids taken from 33 persons, both sexually assaulted or perpetrators were analysed. In 2000 and 2002 not any case of this type was registered, in 2001 only two cases were recorded. After 2003 significant increase in the number of these cases was observed. Eleven and twenty cases involving "date-rape drugs" were submitted to the Institute in 2003 and 2004, respectively. The most common substances detected in analysed materials were amphetamine (in concentrations ranged from 10 to 85 ng/ml) and 9THC (0.36-1.4 ng/ml). Alcohol (0.27-2.3% per hundred), MDMA (8-201 ng/ml), benzodiazepines (oxazepam, nordazepam, estazolam), propranolol and lidocaine were also found in blood and urine specimens.

Tetrahydrocannabinols in clinical and forensic toxicology.

Cannabinoids are the natural constituents of marihuana (cannabis). The main of them are delta9-tetrahydrocannabinol (9THC)--psychoactive agent, cannabinol (CBN) and cannabidiol (CBD). Cannabis is administered either by smoking or orally. 9THC potency and duration of action as well as its and two of its major metabolites concentrations in organism highly depend on the route of administration. A single active dose of 9THC is estimated on 520 mg. 9THC is rapidly metabolised. It is hydroxylated to an active metabolite, I1 -hydroxy-delta9-tetrahydro-cannabinol (11-OH-THC), then oxidised to an inactive 11-nor-9-carboxy-delta9-tetrahydrocannabinol (THCCOOH), which is conjugated with glucuronic acid and predominantly excreted in the urine. The maximum psychological effect persists for 4-6 h after administration despite of very low 9THC blood concentrations. 9THC plasma concentration declined to values of 2-3 ng/ml during 3-4 h after smoking. Such a low concentration of the active compound in human organism create a demand for use of sensitive analytical methods for detection and determination of 9THC and its metabolites. The most effective techniques for 9THC and related compounds determination in biological material are chromatographic ones (gas and liquid) with mass spectrometric detection and different ionization modes. 9THC and its two metabolites (11-OH-THC and THCCOOH) are present in blood and hair, 9THC in saliva, and THCCOOH in urine. 9THC and related compounds are determined in autopsy material, although deaths by overdose of cannabis are exceptionally rare. Fatalities happen most often after intravenous injection of hashish oil. 9THC and its metabolites determination in different biological materials gives the basis for a wide interpretation of analytical results for clinical and forensic toxicology purposes.

Fentanyl and its analogues in clinical and forensic toxicology.

Fentanyl and its analogues are commonly named synthetic opiates. Some compounds from fentanyl group (e.g. fentanyl, alfentanil, sufentanil, remifentanil) are used as anaesthetic and analgesic of high potency and short duration of action. Effects of acting fentanyls are indistinguishable from those produced by nasal inhalation of street heroin. In view of this, fentanyls are very "attractive" for the narcotic market. In least years an increase in number of reports about illegal production and non-medical use of fentanyl and its analogues have appeared. Numerous fentanyl analogues are sold under one name synthetic heroine or China white. Fentanyl and its analogues also can be used as gas warfare agents. One of fentanyl-related compound was used during action of easing hostages from Dubrovka theatre. Because of respiratory depression, which was caused by used compound, died over 100 hostages. Because of high potency fentanyl-related compounds are used in very low doses. So fentanyls appear in biological material in very low concentrations, not higher than several nanograms per millilitre or per gram. The drugs in these concentrations cannot be detected by means of routinely used screening procedures. Therefore direct methods for determination of this group of compounds are needed. Screening method for determination of fentanyl and its analogues was elaborated and validated. Liquid chromatography-mass spectrometry (LC/MS) technique was applied. The method is characterised by the limit of quantification (LOQ) and the limit of detection (LOD) ranged from 0.6 to 2 ng/ml and from 0.2 to 0.6 ng/ml, respectively for four above-mentioned compounds. The method was used for determination of fentanyl in three forensic cases. The blood, bile and blood from lung samples were taken during autopsy of persons who died shortly after surgical procedure in which fentanyl was used as an adjunct to general anaesthesia.

Urinary excretion rates of ketamine and norketamine following therapeutic ketamine administration: method and detection window considerations.

Ketamine is widely used in veterinary medicine. Its medical application in humans is limited to children because in adults it induces severe psychedelic episodes. In recent years, teenagers have abused ketamine as a recreational and "club drug" because of its hallucinogenic and stimulant effects. Ketamine is also misused as a "date-rape" drug (to induce amnesia in unsuspecting victims). Sensitive gas chromatography-mass spectrometry-negative chemical ionization (GC-MS-NCI) and liquid chromatography-mass spectrometry-atmospheric pressure chemical ionization (LC-MS-APCI) methods were applied for the simultaneous quantification of ketamine and its major metabolite, norketamine, in urine. Urine samples were collected from hospitalized children who had received ketamine as an anesthetic. Individual urine samples were collected up to 16 days after drug administration. Using the GC-MS-NCI method, ketamine was detected in the urine of the children from only the day of drug administration up to 2 days after drug administration. Its concentrations ranged from 29 to 1410 ng/mL. Norketamine (measured in concentrations of 0.1-1442 ng/mL) was detected up to 14 days. Using the LC-MS-APCI method, norketamine was detected up to 6 days after drug administration, ranging in concentrations of 2-1559 ng/mL, while ketamine was detected up to 11 days (2-1204 ng/mL). In the urine taken from one child, ketamine was not detected through the entire 16-day period using both methods. The detection window for the analytes is highly dependent on the method used for determination and varies between individuals.

Instability of pancuronium in postmortem blood and liver taken after a fatal intramuscular Pavulon injection.

The present study was designed to determine the stability of pancuronium in postmortem blood and liver during storage. Results were obtained using the method by Kerskes et al. [C.H.M. Kerskes, K.J. Lusthof, P.G.M. Zweipfenning, J.P. Franke, The detection and identification of quaternary nitrogen muscle relaxants in biological fluids and tissues by ion-trap LC-ESI-MS, J. Anal. Toxicol. 26 (2002) 29-34.], modified and validated in our laboratory. Target analytes were isolated after enzymatic hydrolysis followed by solid phase extraction (BondElut C18 column). Internal standardisation was carried out using laudanosine and the target ions were monitored by LC-ESI-MS (monitoring ions m/z 358 for IS and 286 for pancuronium). Materials were taken from a 46-year-old woman, who had been found dead. A syringe (2 ml) and an empty ampoule of Pavulon (4 mg/2 mL) were found in her hand. The residual volume of fluid in the syringe was 0.7 ml. An autopsy was performed six days after death. It revealed a needle mark on the left thigh. Postmortem materials (muscle from the injection site, blood and liver) and the syringe with fluid were stored for four months in a freezer at -20 degrees C. The initial pancuronium concentrations were 81 ng/mL in blood and 532 ng/g in liver. The analyte was stable when stored at -20 degrees C in blood even up to seven months. In liver samples its concentrations were variable. Pancuronium in blood stored at 20 degrees C underwent degradation very rapidly. After three months of storage these blood samples had concentrations not greater about 10% of the initial value. The degradation patterns of pancuronium depended on temperature and the biological matrix.

HPLC/DAD Screening Method for Selected Psychotropic Drugs in Blood.

The influence of experimental conditions on gradient high-performance liquid chromatography/diode array detector separation and identification of 13 psychotropic drugs belonging to two groups--phenothiazines and tricyclic antidepressants--was examined. The main interaction effects of three experimental factors were determined according to a 2 3 factorial design, and the optimum conditions of analysis were searched for. The degree to which the chromatographic peaks overlap is taken into account in a proposed criterion for separation quality. The screening analysis proposed was tested on whole blood samples spiked with mixtures of the examined drugs. The method was characterized by such validation parameters as relative retention times, absorbance ratios at two wavelengths, detection limits, linearity ranges, and extraction recoveries. The method proved to be a suitable tool for the identification of psychotropic drugs tested for forensic purposes.