morphine-3-glucuronide and normorphine

To compare the time course of morphine and metabolite concentrations in serum and cerebrospinal fluid (CSF) after intravenous and intramuscular administration after surgery.. This was a randomized double-blind, double-dummy study in patients who had undergone hip replacement surgery. Morphine (M, 10 mg) was administered intravenously (IV) or intramuscularly (IM). Arterial blood and CSF samples (from a spinal catheter) were drawn simultaneously at 10, 30, 60, and 120 min after administration. Morphine and metabolites [morphine-3-glucuronide (M-3-G), morphine-6-glucuronide (M-6-G), and normorphine (NM)] were determined by a validated liquid chromatography-tandem mass spectrometry method.. Thirty-eight patients were included: 13 men and 25 women, 20 in the IV, 18 in the IM group. Serum concentrations of M after 10 min were consistently higher after IM than IV, concentrations of M-3-G and M-6-G after IM surpassed those of IV after 45 min. NM was not found. None of the metabolites was found in CSF. CSF morphine concentrations and CSF/serum concentration ratios were consistently higher after IV compared to IM. The mean AUC(CSF)/AUC(serum) (0-120 min) concentration ratios were 0.18 and 0.09 after IV and IM, respectively.. The uptake of morphine to the CSF was consistently higher after IV administration than after IM already after 10 min. The higher CSF concentration may be caused by an initially higher morphine blood/CSF gradient following IV morphine injection. The pharmacokinetic findings are compatible with a more rapid and extensive initial effect of IV morphine compared with IM.

Topics: Aged; Analgesics, Opioid; Area Under Curve; Arthroplasty, Replacement, Hip; Bupivacaine; Double-Blind Method; Female; Humans; Injections, Intramuscular; Injections, Intravenous; Male; Midazolam; Middle Aged; Morphine; Morphine Derivatives; Pain Measurement; Pain, Postoperative; Postoperative Care

Subtle changes in molecular structure often lead to significant differences in host-guest interactions, which result in different host-guest recognition capabilities and dynamics behaviours in complex formation. Herein, we reveal the influence of the guest substituents on host-guest molecular recognition by molecular dynamics (MD) simulation and density functional theory (DFT) approaches. The results suggest that the binding energy barrier of acyclic cucurbit[4]uril (ACB[4]) with opiate metabolites gradually decreases. The methyl group in morphine (MOR) and morphine-3-glucuronide (M3G) strengthens the hydrophobicity of the guest, while depressing the energy loss of the desolvation of polar groups (e.g. hydroxyl) inside the ACB[4] cavity. However, in M3G, the 3-glucuronide group located outside the ACB[4] host cavity effectively alleviates the unfavourable desolvation effect of the hydroxyl and increases the binding constant by two orders of magnitude (compared with normorphine (NMOR)). Our findings stressed the essentiality of the binding mode and intermolecular noncovalent interactions in the host-guest selective binding ability.

Topics: Bridged-Ring Compounds; Density Functional Theory; Hydrogen Bonding; Imidazoles; Models, Chemical; Molecular Dynamics Simulation; Morphine; Morphine Derivatives

The USA and numerous other countries worldwide are currently experiencing a public health crisis due to the abuse of heroin and illicitly manufactured fentanyl. We have developed a liquid chromatography tandem mass spectrometry (LC-MS-MS)-based method for the detection of morphine, fentanyl and their metabolites, including morphine-3-glucuronide (M3G), morphine-6-glucuronide (M6G), normorphine, norfentanyl and deuterated internal standards in limited sample volumes with the limit of detection of 5.0/0.5 ng/mL (morphine, M3G, M6G, normorphine/fentanyl, norfentanyl). The inter-assay precision (%CV) was less than 12% for all assays, and the inter-assay bias (%) was less than 5%. The ruggedness of the method, dilution effect and carryover were also investigated as part of the study. The simultaneous quantification of morphine, fentanyl and its metabolites via this simple and time- and cost-efficient method could be successfully applied to samples taken for pharmacokinetic evaluation (antemortem and postmortem) after a single dose of morphine or co-administration of morphine with other drugs (e.g., fentanyl) in rats.

Topics: Animals; Chromatography, Liquid; Fentanyl; Heroin; Morphine; Morphine Derivatives; Rats; Reproducibility of Results; Substance Abuse Detection; Tandem Mass Spectrometry

Development and validation of a selective, robust high-performance liquid chromatography-tandem mass spectrometric (HPLC/MS-MS) method for the quantification of morphine, morphine-3-β-glucuronide, morphine-6-β-glucuronide, hydromorphone, and normorphine in human serum.. Drug-free human serum samples spiked with morphine, morphine-3-β-glucuronide, morphine-6-β-glucuronide, hydromorphone, and normorphine were prepared by protein precipitation using methanol containing the internal standards. Samples were injected onto a Thermo Scientific AccuCore PFP column for chromatographic separation. Detection was achieved using a Thermo Scientific TSQ Vantage mass spectrometer. Assay validation followed the new Clinical and Laboratory Standards Institute (CLSI) C62-A guidelines.. The analytical measuring range for all analytes was determined to be 5 to 1000 ng/mL. Intra- and inter-assay precision for three quality control levels were ≤ 7.0% and ≤ 13.5%, respectively. Carryover, stability, linearity, matrix effects, extraction and processing efficiency and method comparison characteristics were acceptable relative to the CLSI C62 guidelines.. The validation of this HPLC-MS/MS method demonstrated a robust and rapid assay for the quantification of morphine, morphine-3-β-glucuronide, morphine-6-β-glucuronide, hydromorphone, and normorphine.

Topics: Analgesics, Opioid; Chromatography, High Pressure Liquid; Guidelines as Topic; Humans; Hydromorphone; Linear Models; Morphine; Morphine Derivatives; Observer Variation; Reference Standards; Tandem Mass Spectrometry

This work presents two cases of codeine intoxication in 3-year-old monozygotic twin brothers while treated with a codeine slow-release formulation. One child had to be admitted to the hospital, whereas the other one died at home after aspiration of gastric content. The concentrations of codeine and major metabolites including morphine and corresponding glucuronide conjugates were measured by liquid chromatography-tandem mass spectrometry in serum, urine, cerebrospinal fluid, and brain tissue, respectively. A genetic polymorphism study was carried out in order to determine the ability of the children to metabolize codeine by O-demethylation. A pharmacokinetic calculation was also performed to estimate the administered dose of codeine in question. High concentrations of all substances were found in samples of both children. The pharmacokinetic estimate suggests an overdose of codeine, and the possible reasons for the high opiate concentrations are discussed. Furthermore, the postmortem distribution--during and after resuscitation--might play a major role in the interpretation of postmortem concentration levels.

Topics: Antitussive Agents; Brain Chemistry; Brain Edema; Child, Preschool; Chromatography, Liquid; Codeine; Cytochrome P-450 CYP2D6; Delayed-Action Preparations; Drug Overdose; Fatal Outcome; Forensic Toxicology; Genotype; Glucuronides; Humans; Medication Errors; Morphine; Morphine Derivatives; Polymorphism, Genetic; Respiratory Aspiration; Tandem Mass Spectrometry; Tissue Distribution; Twins, Monozygotic

A cation-selective exhaustive injection and sweeping micellar EKC (CSEI-Sweep-MEKC) was established to analyze morphine and its four metabolites, including codeine, normorphine (NM), morphine-3-glucuronide (M3G), and morphine-6-glucuronide (M6G). After SPE, the urine samples were analyzed by this CE method. The phosphate buffer (75 mM, pH 2.5) containing 30% methanol was first filled into an uncoated fused-silica capillary (40 cm, 50 microm id), then a high-conductivity buffer (120 mM phosphate, 10.3 kPa for 99.9 s) followed. The pretreated urine sample was loaded by electrokinetic injection (10 kV, 600 s). The stacking and separation were performed by using phosphate buffer (25 mM, pH 2.5) containing 22% methanol and 100 mM SDS at -20 kV, and detected at 200 nm. During method validation, calibration plots were linear (r > or = 0.998) over a range of 30-3000 ng/mL for morphine, NM, and codeine, 100-2000 ng/mL for M6G, and 80-3200 ng/mL for M3G. The LODs (S/N = 5, sampling 600 s at 10 kV) were 10 ng/mL for morphine, NM, and codeine, 35 ng/mL for M6G, and 25 ng/mL for M3G. This stacking CE method could increase 2500-fold sensitivity of codeine, when comparing with CZE. Five addicts' urine specimens were analyzed. Their results were compared with those of LC-MS-MS, and showed good coincidence. This method could be feasible for monitoring morphine and its metabolites in forensic interest and pharmacokinetic investigations.

Topics: Chromatography, Liquid; Chromatography, Micellar Electrokinetic Capillary; Codeine; Humans; Morphine; Morphine Dependence; Morphine Derivatives; Reproducibility of Results; Solid Phase Extraction; Tandem Mass Spectrometry

A high-performance liquid chromatography tandem mass spectrometry-mass spectrometry (LC-MS-MS) assay was developed for the analyses of morphine, morphine glucuronides and normorphine in plasma samples from rats. The analytes were extracted by using C2 solid-phase extraction cartridges. The extraction recoveries were 100% for morphine, 84% for morphine-3-glucuronide, 64% for morphine-6-glucuronide and 88% for normorphine. Both intra- and inter-assay variabilities were below 11%. Using a plasma sample size of 100 microliters, the limits of detection were 13 nmol l-1 (3.8 ng ml-1) for morphine, 12 nmol l-1 (5.5 ng ml-1) for morphine-3-glucuronide, 26 nmol l-1 (12 ng ml-1) for morphine-6-glucuronide and 18 nmol l-1 (5.0 ng ml-1) for normorphine, at a signal-to-noise ratio of 3. The present assay was applied to a pharmacokinetic study in rats after intraperitoneal administration of morphine.

Topics: Analgesics, Opioid; Animals; Area Under Curve; Biotransformation; Calibration; Chromatography, High Pressure Liquid; Half-Life; Male; Mass Spectrometry; Morphine; Morphine Derivatives; Rats; Rats, Sprague-Dawley; Solutions

A rapid and selective reversed-phase high-performance liquid chromatographic assay with gradient elution and diode-array detection for diacetylmorphine, morphine, codeine, and their free and glucuronidated metabolites in plasma, was developed. After addition of ethylmorphine as internal standard the plasma samples were extracted using C18 ODS-2 solid-phase columns with a recovery better than 80%. The limit of quantitation using an injection volume of 2 microl was 25 ng/ml for each compound. The intra- and inter-day precision was better than 5%. The described method cannot only be used for pharmacokinetic studies but also for intoxication cases to monitor a wide range of opiates.

Topics: Chromatography, High Pressure Liquid; Heroin; Humans; Injections, Intravenous; Morphine; Morphine Derivatives; Narcotics; Spectrophotometry, Ultraviolet

It has previously been shown that guinea pig hepatocytes metabolise morphine in a fashion similar to humans. The metabolism of morphine (5 muM) and the formation of metabolites morphine-3-glucuronide, morphine-6-glucuronide and normorphine was studied in the absence and presence of ethanol (5, 10, 25, 60 and 100 mM) in freshly isolated guinea pig hepatocytes. In order to gain more detailed information, a mathematical model was estimated on experimental data and used to analyse the effects of ethanol on the reaction rates of the different morphine metabolites. Ethanol inhibited the rate of morphine elimination in a dose-related manner, at the high ethanol concentrations the elimination rate was 40 per cent of the control rate. The formation of morphine-glucuronides was influenced in a biphasic manner. Five and 10 mM ethanol increased both the morphine-3-glucuronide and morphine-6-glucuronide levels after 60 min incubation compared to the control, whereas at the higher ethanol concentrations (25-100 mM) the levels of morphine-glucuronides were reduced. Data from the mathematical model, however, demonstrated that the reaction rates for morphine-glucuronide formation were decreased at all ethanol concentrations and in a dose-dependent manner, the interpretation of this being that at the lower (5 and 10 mM) ethanol concentrations employed in this study, other metabolic pathways of morphine are more heavily inhibited than the glucuronidations, resulting in a shunting towards morphine-3-glucuronide and morphine-6-glucuronide. The pharmacodynamic consequences of these pharmacokinetic effects are thus somewhat difficult to predict since morphine-6-glucuronide has a higher agonist potency than morphine. At high concentrations ethanol inhibition of morphine metabolism will increase the concentration of morphine and subsequently the euphoric and the toxic effects. The lower quantities of morphine-6-glucuronide formed in the presence of high ethanol concentrations on the other hand most probably imply reduction of such effects and the net pharmacodynamic effect would be uncertain. At low ethanol concentrations, however, morphine-6-glucuronide concentrations increased and morphine metabolism was less inhibited leading to a possible potentiation of the effects of morphine. Thus, a low ethanol concentration might exert a more pronounced ethanol-drug effect interaction than a higher ethanol concentration.

Topics: Animals; Cell Separation; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Drug Interactions; Ethanol; Guinea Pigs; Liver; Male; Models, Theoretical; Morphine; Morphine Derivatives; Narcotics; Regression Analysis; Solvents

A sensitive method for the analysis of unconjugated morphine, codeine, normorphine and total morphine after hydrolysis of glucuronide conjugates is described. The method was applicable to 50-microliters volumes of plasma. The analytes were converted to heptafluorobutyryl (HFB) derivatives before analysis by gas chromatography-negative ion chemical ionization mass spectrometry. Morphine and codeine were quantified against their [2H3]-isotopomers. Linearity, precision and accuracy were quite acceptable (in the 10(-10)-10(-9) g range), and the absolute limits of detection were < 1 pg.

Topics: Child; Codeine; Gas Chromatography-Mass Spectrometry; Humans; Morphine; Morphine Derivatives; Sensitivity and Specificity; Tritium

A specific HPLC method with UV detection was used to investigate the disposition of morphine and its metabolites in the in-situ rat isolated perfused liver preparation. Livers of male Sprague-Dawley rats (n = 4) were perfused under single pass conditions with protein- and erythrocyte-free perfusate, containing 2.66 microM morphine, for up to 90 min. The concentration of morphine, normorphine and morphine-3-glucuronide (M3G) in outflow perfusate, and the biliary excretion of M3G and normorphine glucuronide, all reached steady-state levels within 15-20 min after commencing perfusion. At steady-state, the mean (+/- s.d.) extraction ratio of morphine was 0.87 +/- 0.06 and clearance (26.0 +/- 1.7 mL min-1) approached perfusate flow rate (30 mL min-1). Although M3G was the main metabolite, accounting for 72.8 +/- 12.7% of eliminated morphine, a significant proportion (21.6 +/- 13.5%) was N-demethylated to normorphine and was recovered as unchanged normorphine in outflow perfusate and normorphine glucuronide in bile. The biliary extraction ratio of hepatically-formed M3G was 0.61 +/- 0.31. Results from an additional six experiments, in which livers were perfused with 1.33 and 2.66 microM of morphine for 30 min each in a balanced cross-over manner, indicated that the disposition of morphine and its metabolites was approximately linear within this concentration range.

Topics: Animals; Chromatography, High Pressure Liquid; Liver; Male; Morphine; Morphine Derivatives; Perfusion; Rats; Rats, Sprague-Dawley

An isocratic high-performance liquid chromatographic method has been developed for the determination of morphine, codeine, normorphine, morphine 3-glucuronide and morphine 6-glucuronide in plasma using a diol column and diode-array detection. Samples were extracted using solid-phase extraction with recoveries in excess of 90%. The limit of determination was 1 ng/ml for morphine, codeine and morphine 3-glucuronide, and 10 ng/ml for normorphine and morphine 6-glucuronide. Inter- and intra-day precision were better than 10%.

Topics: Chromatography, High Pressure Liquid; Codeine; Humans; Hydrogen-Ion Concentration; Morphine; Morphine Derivatives; Solvents

Morphine, morphine-6-glucuronide (M6G), morphine-3-glucuronide (M3G) and normorphine were analysed with high performance liquid chromatography in plasma and urine, collected over 72 h after administration of single intravenous 5 mg and oral 20 mg doses of morphine to 7 healthy volunteers. Systemic plasma clearance of morphine was on average 21.1 +/- 3.4 ml/min/kg (1.27 +/- 0.20 L/h/kg), volume of distribution was 2.9 +/- 0.8 L/kg and oral bioavailability was 29.2 +/- 7.2%. Clearance of morphine to form M3G and M6G comprised 57.3% and 10.4%, respectively, and renal clearance comprised 10.9% of total systemic plasma clearance; hence, more than one-fifth of a dose (20.8%) remained as unidentified residual clearance. On the basis of the area under the plasma concentration-time curves determined after oral and intravenous administration, the ratios of M6G:morphine were 3.6 +/- 1.2 and 0.7 +/- 0.3, respectively. The corresponding figures for M3G:morphine were 29.9 +/- 6.8 and 7.7 +/- 1.4. Differences in metabolic ratios between the parenteral and oral routes could be attributed solely to differences in morphine concentrations as evidenced both by plasma concentrations and amounts excreted in urine. An oral:parenteral potency ratio of 1:3 may, thus, be due to differences in circulating amounts of morphine since the proportions of an administered dose found as M6G and M3G after administration by both routes were equal. A major finding was a slowly declining terminal phase of morphine and metabolites that was evident both in plasma and in urinary excretion versus time curves, where the half-lives of morphine, M3G and M6G were 15.1 +/- 6.5 h, 11.2 +/- 2.7 h and 12.9 +/- 4.5 h, respectively. The terminal half-life of normorphine was 23.9 +/- 10.1 h after oral administration. Comparison of oral with intravenous excretion curves showed that a greater part of morphine and metabolites were excreted during the slowly declining phase after the oral dose than the intravenous dose, which is highly suggestive of enterohepatic cycling. The renal clearance of M6G and morphine was seen to exceed creatinine clearance, possibly due to an active secretion process.

Topics: Administration, Oral; Adult; Enterohepatic Circulation; Female; Humans; Injections, Intravenous; Male; Middle Aged; Morphine; Morphine Derivatives

This article describes a high-performance liquid chromatography (HPLC) method for the simultaneous determination of morphine (M) and its principal metabolites morphine-3-glucuronide (M3G), morphine-6-glucuronide (M6G), and normorphine (NM) in plasma. All four compounds are extracted from plasma using a C8 solid-phase extraction column, separated by reverse-phase HPLC on a C18 analytical column, and detected by spectrofluorometry at 210 nm excitation wavelength. The method takes advantage of the compounds' native fluorescence, so that derivitization is not required. Samples have been quantified over a concentration range of 25-100 ng/ml M and NM, 50-200 ng/ml M3G, and 100-300 ng/ml M6G, using nalorphine (500 ng/ml) as internal standard. Within-run and between-run errors were less than 10% for morphine and less than 13% for all the metabolites. The lower limit of quantitation for morphine is 10 ng/ml. The accuracy of the method was confirmed by including quality controls fitted to the standard curves of each compound. The assay described in this article represents a simplification of previous versions of the method, which included cumbersome extraction procedures and multiple detectors. For the first time, an internal standard has been employed. The assay is reliable and easy to use and can be performed in any therapeutic drug monitoring laboratory.

Topics: Chromatography, High Pressure Liquid; Fluorometry; Humans; Morphine; Morphine Derivatives

A method for the fast analysis of morphine (M), normorphine (NM), morphine-3- and -6-glucuronides (M3G and M6G) and codeine (C) is described which has the advantages of sensitivity, speed and specificity. Dihydrocodeine and heroin can also be assayed. The method is based on extraction of the opiates from serum, plasma and cerebrospinal fluid using reversed-phase solid-phase extraction columns, followed by reversed-phase high-performance liquid chromatography with native fluorescence detection. The extraction step provides greater than 95% recovery, and the response of the detection system is linear from 0.5 to beyond 750 ng. The method allows analysis of M, NM, M3G, M6G and C. No other drugs have been found to interfere with the assay. The assay offers a quick, cheap and reliable method of specifically determining morphine and its metabolites, including the potent M6G, from a small sample volume; this will be of advantage to both clinician and basic scientist.

Topics: Chromatography, High Pressure Liquid; Codeine; Fluorescence; Humans; Microchemistry; Morphine; Morphine Derivatives; Quality Control