Characterization of the pharmacokinetics of entrectinib and its active M5 metabolite in healthy volunteers and patients with solid tumors.

BACKGROUND: Entrectinib is an oral, CNS-active, potent inhibitor of tyrosine receptor kinases A/B/C, tyrosine kinase ROS proto-oncogene 1, and anaplastic lymphoma kinase approved for use in patients with solid tumors. We describe 3 clinical studies, including one investigating the single/multiple dose pharmacokinetics of entrectinib in patients and two studies in healthy volunteers investigating the absorption/distribution/metabolism/excretion (ADME) of entrectinib, its relative bioavailability, and effect of food on pharmacokinetics. METHODS: The patient study is open-label with dose-escalation and expansion phases. Volunteers received entrectinib (100-400 mg/m2, and 600-800 mg) once daily with food in continuous 28-day cycles. In the ADME study, volunteers received a single oral dose of [14C]entrectinib 600 mg. In the third study, volunteers received single doses of entrectinib 600 mg as the research and marketed formulations in the fasted state (Part 1), and the marketed formulation in the fed and fasted states (Part 2). Entrectinib and its major active metabolite M5 were assessed in all studies. RESULTS: Entrectinib was absorbed in a dose-dependent manner with maximum concentrations at ~4 h postdose and an elimination half-life of ~20 h. Entrectinib was cleared mainly through metabolism and both entrectinib and metabolites were eliminated mainly in feces (minimal renal excretion). At steady-state, the M5-to-entrectinib AUC ratio was 0.5 (with 600 mg entrectinib research formulation in patients). The research and marketed formulations were bioequivalent and food had no relevant effect on pharmacokinetics. CONCLUSIONS: Entrectinib is well absorbed, with linear PK that is suitable for once-daily dosing, and can be taken with or without food.

Review of bioanalytical assays for the quantitation of various HDAC inhibitors such as vorinostat, belinostat, panobinostat, romidepsin and chidamine.

Histone deacetylase inhibitors (HDAC inhibitors) are used to treat malignancies such as cutaneous T cell lymphoma and peripheral T cell lymphoma. Only four drugs are approved by the US Food and Drug Administration, namely vorinostat, romidepsin, panobinostat and belinostat, while chidamide has been approved in China. There are a number of bioanalytical methods reported for the measurement of HDAC inhibitors in clinical (human plasma and serum) and preclinical (mouse plasma, rat plasma, urine and tissue homogenates, etc.) studies. This review covers various HDAC inhibitors such as vorinostat, romidepsin, panobinostat, belinostat and chidamide. In addition to providing a comprehensive review of the available methods for the above mentioned HDAC inhibitors, it also provides case studies with perspectives for chosen drugs. Based on the review, it is concluded that the published methodologies using either HPLC or LC-MS/MS are well suited for the quantification of HDAC inhibitors in various biological fluids to delineate pharmacokinetic data.

Plasma concentration-dependent suppression of endogenous hydrocortisone in the horse after intramuscular administration of dexamethasone-21-isonicotinate.

Detection times and screening limits (SL) are methods used to ensure that the performance of horses in equestrian sports is not altered by drugs. Drug concentration-response relationship and knowledge of concentration-time profiles in both plasma and urine are required. In this study, dexamethasone plasma and urine concentration-time profiles were investigated. Endogenous hydrocortisone plasma concentrations and their relationship to dexamethasone plasma concentrations were also explored. A single dose of dexamethasone-21-isonicotinate suspension (0.03 mg/kg) was administered intramuscularly to six horses. Plasma was analysed for dexamethasone and hydrocortisone and urine for dexamethasone, using UPLC-MS/MS. Dexamethasone was quantifiable in plasma for 8.3 ± 2.9 days (LLOQ: 0.025 µg/L) and in urine for 9.8 ± 3.1 days (LLOQ: 0.15 µg/L). Maximum observed dexamethasone concentration in plasma was 0.61 ± 0.12 µg/L and in urine 4.2 ± 0.9 µg/L. Terminal plasma half-life was 38.7 ± 19 h. Hydrocortisone was significantly suppressed for 140 h. The plasma half-life of hydrocortisone was 2.7 ± 1.3 h. Dexamethasone potency, efficacy and sigmoidicity factor for hydrocortisone suppression were 0.06 ± 0.04 µg/L, 0.95 ± 0.04 and 6.2 ± 4.6, respectively. Hydrocortisone suppression relates to the plasma concentration of dexamethasone. Thus, determination of irrelevant plasma concentrations and SL is possible. Future research will determine whether hydrocortisone suppression can be used as a biomarker of the clinical effect of dexamethasone.

Structure identification and elucidation of mosapride metabolites in human urine, feces and plasma by ultra performance liquid chromatography-tandem mass spectrometry method.

1. Mosapride citrate (mosapride) is a potent gastroprokinetic agent. The only previous study on mosapride metabolism in human reported one phase I oxidative metabolite, des-p-fluorobenzyl mosapride, in human plasma and urine using HPLC method. Our aim was to identify mosapride phase I and phase II metabolites in human urine, feces and plasma using UPLC-ESI-MS/MS. 2. A total of 16 metabolites were detected. To the best of our knowledge, 15 metabolites have not been reported previously in human. 3. Two new metabolites, morpholine ring-opened mosapride (M15) and mosapride N-oxide (M16), alone with one known major metabolite, des-p-fluorobenzyl mosapride (M3), were identified by comparison with the reference standards prepared by our group. The chemical structures of seven phase I and six phase II metabolites of mosapride were elucidated based on UPLC-MS/MS analyses. 4. There were two major phase I reactions, dealkylation and morpholine ring cleavage. Phase II reactions included glucuronide, glucose and sulfate conjugation. The comprehensive metabolic pathway of mosapride in human was proposed for the first time. 5. The metabolites in humans were compared with those in rats reported previously. In addition to M10, the other 15 metabolites in humans were also found in rats. This result suggested that there was little qualitative species difference in the metabolism of mosapride between rats and humans. 6. In all, 16 mosapride metabolites including 15 new metabolites were reported. These results allow a better understanding of mosapride disposition in human.

Monitoring phosphodiesterase-4 inhibitors using liquid chromatography/(tandem) mass spectrometry in sports drug testing.

RATIONALE: The recent discovery of resveratrol's capability to inhibit cAMP-specific phosphodiesterases (PDEs) and, as a consequence, to enhance particularly the activity of Sirt1 in animal models has reinforced the interest of preventive doping research organizations, especially in PDE4 inhibitors. Among these, the archetypical PDE4-inhibitor rolipram significantly increased the number of mitochondria in laboratory rodents, which further demonstrated a performance increase in a treadmill-test (time-to-exhaustion) of approximately 40%. Besides rolipram, a variety of new PDE4-inhibiting substances including cilomilast, roflumilast, and numerous additional new drug entities were described, with roflumilast being the first-in-class having received clinical approval for the treatment of chronic obstructive pulmonary disease (COPD). Due to the availability of these substances, and the fact that a misuse of such compounds in sport cannot be excluded, it deems relevant to probe for the prevalence of these compounds in sports drug testing programs. METHODS: Known urinary phase-I metabolites of rolipram, roflumilast, and cilomilast were generated by in vitro incubations employing human liver microsomal preparations. The metabolites obtained were studied by liquid chromatography with high-resolution/high-accuracy tandem mass spectrometry (LC/MS/MS) and the reference product ion mass spectra of established and most relevant metabolites were utilized to provide the information necessary for comprehensive doping controls. The analytical procedure was based on conventional routine doping control assays employing enzymatic hydrolysis followed by liquid-liquid extraction and subsequent LC/MS/MS measurement. RESULTS: Structures of diagnostic product ions and dissociation pathways of target analytes were elucidated, providing the information required for implementation into an existing test method for routine sports drug testing. The established method allowed for detection limits for the intact drugs of 1-5 ng/mL, and further assay characteristics (intraday precision 1.5-13.7%, interday precision 7.3-18.6%, recovery 20-100%, ion suppression/enhancement, and specificity) were determined. In addition, proof-of-concept analyses concerning roflumilast were conducted with a urine sample obtained from a COPD patient under roflumilast treatment.

[The test of benzamide derivative neuroleptics using the technique of thin-layer chromatography].

The article presents the data concerning the application of technique of thin-layer chromatography in the conditions of laboratory diagnostics to identify benzamide derivative neuroleptics (amisupride, sulpiride, tiapride) in derivate from human biologic fluids and tissues with the possibility to separate the analyzed tissues from soextractive tissues.

Assessment of exposure of metabolites in preclinical species and humans at steady state from the single-dose radiolabeled absorption, distribution, metabolism, and excretion studies: a case study.

The exposure of a drug candidate and its metabolites in humans and preclinical species during drug development needs to be determined to ensure that the safety of drug-related components in humans is adequately assessed in the standard toxicology studies. The in vivo radiolabeled studies in preclinical species and human volunteers provide the total fate of the drug-derived radioactivity including the relative abundance of metabolites. Here, we describe how the single-dose radiolabeled human studies could provide the exposure of circulating metabolites at steady state using a case study of an extensively metabolized drug, lixivaptan. After an oral dose of [(14)C]lixivaptan to humans, a total of nine metabolites were detected in the systemic circulation; eight of them exceeded 10% of the parent exposure (2-41% of total radioactivity). The plasma samples were profiled for all subjects at each time point by high-performance liquid chromatography, and metabolites were quantified using a radioactive detector. On the basis of single-dose area under the concentration-time curve (AUC) values, exposure of six human metabolites was greater at least in one preclinical species used in toxicology evaluation. On the basis of the t(1/2) of lixivaptan and two major metabolites from a single dose in humans, their AUC and C(max) values were simulated at the steady state. The simulated exposure (C(max) and AUC) values of parent drug and the two most abundant metabolites were similar to those from a 7-day clinical study obtained using a validated liquid chromatography-mass spectrometry assay, suggesting that a well designed single-dose radiolabeled human study can help in addressing the metabolites in safety testing-related issues.

A nonclinical safety and pharmacokinetic evaluation of N6022: a first-in-class S-nitrosoglutathione reductase inhibitor for the treatment of asthma.

S-nitrosoglutathione reductase is the primary enzyme responsible for the metabolism of S-nitrosoglutathione (GSNO), the body's main source of bioavailable nitric oxide. Through its catabolic activity, GSNO reductase (GSNOR) plays a central role in regulating endogenous S-nitrosothiol levels and protein S-nitrosation-based signaling. By inhibiting GSNOR, we aim to increase pulmonary GSNO and induce bronchodilation while reducing inflammation in lung diseases such as asthma. To support the clinical development of N6022, a first-in-class GSNOR inhibitor, a 14-day toxicology study was conducted. Sprague-Dawley rats were given 2, 10 or 50 mg/kg/day N6022 via IV administration. N6022 was well tolerated at all doses and no biologically significant adverse findings were noted in the study up to 10 mg/kg/day. N6022-related study findings were limited to the high dose group. One male rat had mild hepatocellular necrosis with accompanying increases in ALT and AST and several male animals had histological lung assessments with a slight increase in foreign body granulomas. Systemic exposure was greater in males than females and saturation of plasma clearance was observed in both sexes in the high dose group. Liver was identified as the major organ of elimination. Mechanistic studies showed dose-dependent effects on the integrity of a rat hepatoma cell line.

Metabolism of flumatinib, a novel antineoplastic tyrosine kinase inhibitor, in chronic myelogenous leukemia patients.

4-(4-Methyl-piperazin-1-ylmethyl)-N-[6-methyl-5-(4-pyridin-3-yl-pyrimidin-2-ylamino)-pyridin-3-yl]-3-trifluoromethyl-benzamide (flumatinib, HH-GV678), an antineoplastic tyrosine kinase inhibitor, is currently in Phase I clinical trials in China for the treatment of chronic myelogenous leukemia (CML). The purpose of this study was to identify the metabolites of flumatinib in CML patients, with the aim of determining the main metabolic pathways of flumatinib in humans after oral administration. Ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry revealed 34 metabolites; 7 primary metabolites were confirmed by comparison with synthetic reference standards. The results show that the parent drug flumatinib was the main form recovered in human plasma, urine, and feces. The main metabolites of flumatinib in humans were the products of N-demethylation, N-oxidation, hydroxylation, and amide hydrolysis. In addition to these phase I metabolites, several phase II glucuronidation and acetylation products were detected in plasma, urine, and feces. The observed circulating metabolites included an N-demethylated metabolite (M1), two hydrolytic metabolites (M3, M4), oxidation metabolites (M2-1, M2-4, M2-7, M2-9, and M14), a glucuronide conjugate (M16-2), and several multiple metabolic products. Flumatinib was predominantly metabolized by amide bond cleavage to yield two corresponding hydrolytic products. By comparison with the related drug 4-(4-methyl-piperazin-1-ylmethyl)-N-[4-methyl-3-(4-pyridin-3-yl-pyrimidin-2-ylamino)-phenyl]-benzamide (imatinib), we concluded that the electron-withdrawing groups of trifluoromethyl and pyridine facilitated the amide bond cleavage and led to the in vivo formation of a carboxylic acid and an amine.

Identification of urine metabolites of TFAP, a cyclooxygenase-1 inhibitor.

Only a few COX-1-selective inhibitors are currently available, and the research on COX-1 selective inhibitors is not fully developed. The authors have produced several COX-1 selective inhibitors including N-(5-amino-2-pyridinyl)-4-trifluoromethylbenzamide: TFAP (3). Although 3 shows potent analgesic effect without gastric damage, the urine after administration of 3 becomes red-purple. Since the colored-urine should be avoided for clinical use, in this research we examined the cause of the colored-urine. UV-vis spectra and LC-MS/MS analyses of urine samples and metabolite candidates of 3 were performed to afford information that the main reason of the colored urine is a diaminopyridine (4), produced by metabolization of 3. This information is useful to design new COX-1 selective inhibitors without colored urine based on the chemical structure of 3.

Are pigs a suitable animal model for in vivo metabolism studies of new psychoactive substances? A comparison study using different in vitro/in vivo tools and U-47700 as model drug.

Being highly potent, New Synthetic Opioids (NSO) have become a public health concern. Little is known though about the metabolism and toxicokinetics (TK) of many of the non fentanyl NSO such as U-47700. Obtaining such data in humans is challenging and so we investigated if pigs were a suitable model species as TK model for U-47700. The metabolic fate of U-47700 was elucidated after intravenous administration to one pig in vivo and results were compared to metabolic patterns formed by different other in vitro systems (human and pig liver microsomes, human liver S9 fraction) and compared to rat and human in vivo data. Furthermore, monooxygenase isozymes responsible for the major metabolic steps were elucidated. In total, 12 phase I and 8 phase II metabolites of U-47700 could be identified. The predominant reactions were N-demethylation, hydroxylation, and combination of them followed by glucuronidation or sulfation. The most predominant monooxygenase catalyzed conversions were N-demethylation, and hydroxylation by CYP3A4 and 2B6, and FMO3 catalyzed N-oxidation. Similar main phase I metabolites were found in vitro as compared to in vivo (pig/human). The metabolic pattern elucidated in the pig was comparable to human in vivo data. Thus, pigs seem to be a suitable animal model for metabolism and further TK of U-47700.

Multiplex detection of 14 fentanyl analogues and U-47700 in biological samples: Application to a panel of French hospitalized patients.

Synthetic opioids (SO) associated with the recent alarming increase of deaths and intoxications in United States of America and Europe are not detected by the usual first-line opiates drug screening assays. We developed a liquid chromatography tandem mass spectrometry analytical method for the multiplex detection of 14 fentanyl analogues (2-furanylfentanyl, 4-ANPP, 4-methoxybutyrylfentanyl, acrylfentanyl, alfentanil, carfentanil, despropionyl-2-fluorofentanyl, fentanyl, methoxyacetylfentanyl, norfentanyl, ocfentanil, remifentanil, sufentanil and valerylfentanyl) and U-47700 in whole blood and urine samples. The method was validated according to the requirements of ISO 15189. A simple and fast liquid-liquid extraction (LLE) with De-Tox Tube-A was performed leading to better recovery of molecules in urine than in blood samples. Depending on the compound, the limits of detection (LODs) ranged from 0.01 to 0.10 ng/mL and from 0.02 to 0.05 ng/mL in whole blood and urine, respectively. Calibration curves were linear in the range 0.5-50.0 ng/mL and the limit of quantification (LOQ) ranged from 0.10 to 0.40 ng/mL in blood. Internal quality controls at 1 and 40 ng/mL showed intra-day and between-day precision and accuracy bias below 10% in urine and 15% in blood. The method was applied to the screening of 211 urine samples from patients admitted in emergency or addiction departments. The presence of legal fentanyl analogues in 5 urine samples was justified by their therapeutic use as analgesics. Only one patient was concerned by fentanyl misuse and addiction whereas no illegal SO was detected. This study is not in favor of a huge misuse of SO in the Lorraine region.

Identification of human metabolites of (-)-N-{2-[(R)-3-(6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline-2-carbonyl)piperidino]ethyl}-4-fluorobenzamide (YM758), a novel If channel inhibitor, and investigation of the transporter-mediated renal and hepatic excretion of these metabolites.

(-)-N-{2-[(R)-3-(6,7-Dimethoxy-1,2,3,4-tetrahydroisoquinoline-2-carbonyl)piperidino]ethyl}-4-fluorobenzamide (YM758) is a novel inhibitor of the "funny" If current channel (If channel) that is expressed in the sinus node of heart and is being developed as a treatment for stable angina and atrial fibrillation. Its metabolites were identified in human urine, plasma, and feces by radio-high-performance liquid chromatography and liquid chromatography-tandem mass spectrometry analyses after oral administration of [(14)C]YM758. 6,7-Dimethoxy-2-[(3R)-piperidin-3-ylcarbonyl]-1,2,3,4-tetrahydroisoquinoline (YM-252124), (5R)-5-[(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)carbonyl]piperidin-2-one (YM-385459), 2-{[(3R)-1-{2-[(4-fluorobenzoyl)amino]ethyl}piperidin-3-yl]carbonyl}-7-methoxy-1,2,3,4-tetrahydroisonolin-6-yl beta-D-glucopyranosiduronic acid (AS2036329), and the unchanged drug were detected as major constituents in both urine and plasma, whereas N-(4-fluorobenzoyl)glycine (YM-385461) was detected in plasma, but not in urine. The renal and hepatic uptake transporters for these metabolites were investigated by assessing their inhibitory effect on uptake activity in human (h) organic cation transporter (OCT) 1-3/rat (r) Oct1-3, human organic anion transporter (OAT) 1/rOat1, hOAT3/rOat3, and organic anion-transporting protein 1B1/1B3-expressing HEK293 cells. IC(50) values of YM-252124 for 1-methyl-4-phenylpyridinium uptake via hOCT2 and rOct2 were 93.9 and 1700 microM, respectively, suggesting that this metabolite is secreted into urine via hOCT2/rOct2 and that the large difference in the inhibitory potentials between hOCT2 and rOct2 explains the species difference in the urinary excretion ratio of the radioactivity. The renal secretion of YM-385461, one derivative of p-aminohippuric acid, via hOAT1/rOat1, and hepatic uptake of YM-252124 via hOCT1/rOct1 was also expected. This kind of study was useful in investigating the relationship between the urinary/hepatic elimination and the transport activity for metabolites.

Metabolism of novel opioid agonists U-47700 and U-49900 using human liver microsomes with confirmation in authentic urine specimens from drug users.

Recently, the number of adverse events, including death, involving novel opioids has continued to increase, providing additional and sustained challenges for forensic and medical communities. Identification of emerging novel opioids can be challenging, compounded by detection windows and unknown metabolic profiles. In this study, human liver microsomes were used for the generation of in vitro metabolic profiles of U-47700 and U-49900. Generated metabolites were analyzed via a SCIEX TripleTOF® 5600+ quadrupole time-of-flight mass spectrometer and resulting data files were processing using MetabolitePilot™. Characterized metabolites were verified in vivo by analysis of authentic human urine specimens collected after analytically confirmed cases of overdose involving U-47700 or U-49900. In total, four metabolites were identified and present in urine specimens for U-47700, and five metabolites for U-49900. N-Desmethyl-U-47700 was determined to be the primary metabolite of U-47700. Parent U-47700 was identified in all urine specimens. N-Desmethyl-U-47700 and N,N-didesmethyl-U-47700 were structurally confirmed for the first time during this study following acquisition of standard reference material. N-Desethyl-U-49900 was determined to be the primary metabolite of U-49900 following microsomal incubations, while N,N-didesethyl-N-desmethyl-U-49900 was the most abundant in a urine specimen. Similarities in metabolic transformation were identified between U-47700 and U-49900, resulting in a common metabolite and isomeric species. These phenomena should be considered in cases involving U-47700 or U-49900. This study is the first to map the metabolic profiles of U-47700 and U-49900 using human liver microsomes, as well as the first to report any literature involving U-49900 and analysis of case specimens.

Two cases of intoxication with new synthetic opioid, U-47700.

BACKGROUND: Novel substances often referred to as "designer drugs" have emerged as drugs of abuse, and recognition of these is difficult as routine blood and urine screening tests do not detect these agents. U-47700 is a synthetic selective µ-opioid agonist that can be bought online for as little as $40 per gram. We report two patients presenting after insufflation of U-47700, with subsequent confirmation of this substance in urine samples. CASE DETAILS: A 26-year-old man and 24-year-old woman insufflated a substance they believed to be "synthetic cocaine." The man was found down with cyanosis and agonal respirations. He was intubated and taken to hospital where he recovered well with supportive care. The woman presented with anxiety, tremors and drowsiness and was admitted for observation. Urine samples from both patients were analyzed using GC/MS/MS and LC/QToF, and U-47700 was isolated in both cases. No other opioids were detected. DISCUSSION: These cases are concerning because U-47700 is a relatively new agent that is easy to obtain over the internet and has the potential to cause significant morbidity and mortality.

Near death from a novel synthetic opioid labeled U-47700: emergence of a new opioid class.

BACKGROUND: In the last decade there has been a worldwide surge in the recreational abuse of novel psychoactive substances, particularly amphetamine derivatives and synthetic cannabinoids. Synthetic opioids such as AH-7921, MT-45, and U-47700, with structures distinct from those ever used therapeutically or described recreationally, have also recently emerged. CASE DETAILS: We report a patient who suffered respiratory failure and depressed level of consciousness after recreationally using a novel synthetic opioid labeled U-47700. A single dose of naloxone administered by paramedics completely reversed his opioid poisoning. Comprehensive laboratory analysis confirmed the presence of a novel synthetic opioid and excluded other drugs. The drug used appeared to have caused a false positive benzodiazepine result on the initial urine drugs of abuse panel. CONCLUSION: The case we describe of toxicity from the synthetic opioid labeled U-47700 highlights the emerging trend of novel synthetic opioid abuse.

A case of U-47700 overdose with laboratory confirmation and metabolite identification.

CONTEXT: The opioid epidemic has included use of traditional drugs and recently newer synthetics. It is critically important to recognize and identify these new drugs both clinically and through appropriately designed toxicology testing. There is little available information on a synthetic gaining popularity, U-47700. CASE DETAILS: A 23-year-old female presented after using "U4" by nasal insufflation and injection. She was cyanotic with respiratory depression and responded to naloxone in the field. She was found to have non cardiogenic pulmonary edema and hemoptysis which improved with BiPAP. Urine and serum samples were analyzed using mass spectrometry, confirming 3,4-dichloro-N-[(1R,2R)-2-(dimethylamino)cyclohexyl]-N-methylbenzamide or U-47700. The sample was further analyzed elucidating metabolism specifics. Drug and metabolite concentrations were subsequently measured in both serum and urine. The parent compound of U-47700 was detected at 394 ng/mL and 228 ng/mL in serum and urine, respectively. Metabolites detected in appreciable amounts included the desmethyl (1964 ng/mL in urine), bisdesmethyl (618 ng/mL), desmethyl hydroxy (447 ng/mL), and bisdesmethyl hydroxy forms (247 ng/mL) of U-47700. DISCUSSION: U-47700 is a potent µ-opioid receptor agonist and has recently been used recreationally, contributing to hospitalizations and likely deaths in the community. This is a case report describing an exposure to U-47700 with subsequent laboratory analysis. Based upon this one case, parent U-47700 appear to be an appropriate marker of use in a serum sample. However, demethylated metabolites appear dominant as urinary markers of U-47700 use.

Analysis of U-47700, a Novel Synthetic Opioid, in Human Urine by LC-MS-MS and LC-QToF.

The illicit drug market has rapidly evolved from synthetic cannabinoids to cathinone derivatives and now a new emerging threat of synthetic opioids. These compounds were mostly developed by pharmaceutical companies during drug discovery. The new psychoactive substances are not routinely covered in drug screening and may go undetected. Recently fentanyl analogous, AH-7921, MT-45 and now U-47700 have been encountered in clinical and forensic casework. U-47700 is gaining popularity on drug user forms as a legal alternative to heroin. It is a µ-receptor agonist that is part of the trans-1-2-diamine opioid analgesic drug class developed by The Upjohn Company in an attempt to develop a non-addicting analgesic. A LC-MS-MS method was developed and validated to detect and quantify U-47700. Additional analysis was conducted with an LC-QToF to identify the presence of the parent drug and metabolites. A total of four cases have been evaluated by the LC-MS-MS methodology which has an analytical range of 1-1,250 ng/mL and limit of detection of 1 ng/mL. The concentration of U-47700 in urine specimens ranged from below the limit of quantification to 224 ng/mL. The ToF analysis detected the presence of suspected phase I demethylated metabolites that may assist future analysis of this compound. The prevalence of designer opioids in casework highlights the importance of analysis for new psychoactive substances. Traditional opiates/opioids were not detected in the presented cases, but the available case histories revealed an opioid toxidrome. These findings suggest that U-47700 drug may cause significant morbidity and mortality within the United States as an emerging drug threat.

Molecularly imprinted polymer-based bulk optode for the determination of itopride hydrochloride in physiological fluids.

We report here for the first time on the use of Molecularly Imprinted Polymers as modifiers in bulk optodes, Miptode, for the determination of a pharmaceutical compound, itopride hydrochloride as an example in a concentration range of 1×10(-1)-1×10(-4)molL(-1). In comparison to the optode containing the ion exchanger only (Miptode 3), the optode containing the ion exchanger and the MIP particles (Miptode 2) showed improved selectivity over the most lipophilic species, Na(+) and K(+), by more than two orders of magnitude. For instance, the optical selectivity coefficients using Miptode 2, [Formula: see text] , were as follow: NH4(+)˂-6; Na(+)=-4.0, which were greatly enhanced in comparison with that obtained by Miptode 3. This work opens a new avenue for using miptodes for the determination of all the pharmaceutical preparations without the need for the development of new ionophores.

A case of acute intoxication due to combined use of fentanyl and 3,4-dichloro-N-[2-(dimethylamino)cyclohexyl]-N-methylbenzamide (U-47700).

A 30-year old man was found dead in his home after inhaling fumes of a powder burned on aluminum foil. Blood and urine were taken by the medical examiner during the external body examination and submitted to the laboratory for a comprehensive systematic toxicological analysis. A toxic fentanyl level of 10.9µg/L was measured in the subclavian blood. Police investigation revealed that the man searched the internet for information on new psychotropic substances, among others including U-47700. A powder found in the victims' home was transferred to the laboratory for analysis, in which trace amounts of fentanyl (0.0035%, m/m) and U-47700 (0.0012%, m/m) were identified by gas chromatography mass spectrometry. U-47700 is an opioid analgesic drug, considered to have a potency of approximately 7.5 times that of morphine. A target analysis on U-47700 was performed using liquid-liquid extraction and ultra performance liquid chromatography tandem mass spectrometry operating in multiple reaction monitoring mode. The method validation was based on the Scientific Working Group of Forensic Toxicology document 'Standard Practices of Method Validation in Forensic Toxicology'. In blood and urine the U-47700 concentration was 13.8 and 71.0µg/L, respectively. To the author's knowledge, this is the first case report of a fatal intoxication involving U-47700 abused as a new psychotropic substance.