morphine-3-glucuronide and Opioid-Related-Disorders

Current hair testing methods that rely solely on quantification of parent drug compounds are unable to definitively distinguish between drug use and external contamination. One possible solution to this problem is to confirm the presence of unique drug metabolites that cannot be present through contamination, such as phase II glucuronide conjugates. This work demonstrates for the first time that codeine-6-glucuronide, hydromorphone-3-glucuronide, oxymorphone-3-glucuronide, morphine-3-glucuronide and morphine-6-glucuronide are present at sufficient concentrations to be quantifiable in hair of opioid users and that their concentrations generally increase as the concentrations of the corresponding parent compounds increase. Here, we present a validated liquid chromatography tandem mass spectrometry method to quantify codeine-6-glucuronide, dihydrocodeine-6-glucuronide, hydromorphone-3-glucuronide, morphine-3-glucuronide, morphine-6-glucuronide, oxymorphone-3-glucuronide, codeine, dihydrocodeine, dihydromorphine, hydrocodone, hydromorphone, morphine, oxycodone, oxymorphone and 6-acetylmorphine in human hair. The method was used to analyze 46 human hair samples from known drug users that were confirmed positive for opioids by an independent laboratory. Glucuronide concentrations in samples positive for parent analytes ranged from ~1 to 25 pg/mg, and most samples had glucuronide concentrations in the range of ~1 to 5 pg/mg. Relative to the parent concentrations, the average concentrations of the four detected glucuronides were as follows: codeine-6-glucuronide, 2.33%; hydromorphone-3-glucuronide, 0.94%; oxymorphone-3-glucuronide, 0.77%; morphine 3-glucuronide, 0.59%; and morphine-6-glucuronide, 0.93%.

Topics: Chromatography, Liquid; Codeine; Glucuronates; Hair; Humans; Hydromorphone; Limit of Detection; Morphine Derivatives; Opioid-Related Disorders; Reproducibility of Results; Specimen Handling; Substance Abuse Detection; Tandem Mass Spectrometry

The toxicodynamics and, to a lesser degree, toxicokinetics of the widely used opiate codeine remain a matter of controversy. To address this issue, analytical methods capable of providing reliable quantification of codeine metabolites alongside codeine concentrations are required. This article presents a validated method for simultaneous determination of codeine, codeine metabolites codeine-6-glucuronide (C6G), norcodeine and morphine, and morphine metabolites morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) in post-mortem whole blood, vitreous fluid, muscle, fat and brain tissue by high-performance liquid chromatography mass spectrometry. Samples were prepared by solid-phase extraction. The validated ranges were 1.5-300 ng/mL for codeine, norcodeine and morphine, and 23-4,600 ng/mL for C6G, M3G and M6G, with exceptions for norcodeine in muscle (3-300 ng/mL), morphine in muscle, fat and brain (3-300 ng/mL) and M6G in fat (46-4,600 ng/mL). Within-run and between-run accuracy (88.1-114.1%) and precision (CV 0.6-12.7%), matrix effects (CV 0.3-13.5%) and recovery (57.8-94.1%) were validated at two concentration levels; 3 and 150 ng/mL for codeine, norcodeine and morphine, and 46 and 2,300 ng/mL for C6G, M3G and M6G. Freeze-thaw and long-term stability (6 months at -80°C) was assessed, showing no significant changes in analyte concentrations (-12 to +8%). The method was applied in two authentic forensic autopsy cases implicating codeine in both therapeutic and presumably lethal concentration levels.

Topics: Adipose Tissue; Autopsy; Brain; Calibration; Cause of Death; Chromatography, High Pressure Liquid; Codeine; Forensic Toxicology; Humans; Limit of Detection; Mass Spectrometry; Morphine Derivatives; Muscle, Skeletal; Opioid-Related Disorders; Reference Standards; Reproducibility of Results; Solid Phase Extraction; Substance Abuse Detection; Vitreous Body

Morphine-3- and morphine-6-glucuronide are morphine's major metabolites. As morphine-6-glucuronide produces stronger analgesia than morphine, we investigated the effects of acute and chronic morphine glucuronides on adenylyl cyclase (AC) activity. Using COS-7 cells cotransfected with representatives of the nine cloned AC isozymes, we show that AC-I and V are inhibited by acute morphine and morphine-6-glucuronide, and undergo superactivation upon chronic exposure, while AC-II is stimulated by acute and inhibited by chronic treatment. Morphine-3-glucuronide had no effect. The weak opiate agonists codeine and dihydrocodeine are also addictive. These opiates, in contrast to their 3-O-demethylated metabolites morphine and dihydromorphine (formed by cytochrome P450 2D6), demonstrated neither acute inhibition nor chronic-induced superactivation. These results suggest that metabolites of morphine (morphine-6-glucuronide) and codeine/dihydrocodeine (morphine/dihydromorphine) may contribute to the development of opiate addiction.

Topics: Adenylyl Cyclase Inhibitors; Adenylyl Cyclases; Animals; CHO Cells; Codeine; Cricetinae; Dihydromorphine; Dose-Response Relationship, Drug; Enzyme Activation; Isoenzymes; Morphine; Morphine Derivatives; Opioid-Related Disorders; Receptors, Opioid, mu; Thyrotropin; Transfection

A solid-phase extraction and gas chromatographic-mass spectrometric method for the simultaneous determination of codeine, dihydrocodeine, morphine, and 6-monoacetylmorphine in serum, blood or postmortem blood is described. The extraction technique allows the determination of free or total morphine (morphine plus morphine glucuronide). Experiments with spiked blood samples resulted in recoveries of 96.4% +/- 4.2% for codeine, 95.8% +/- 5.1% for dihydrocodeine, 90.3% +/- 7.8% for 6-monoacetylmorphine and 92.5% +/- 8.1% for morphine. Excellent linearity was obtained over the range 1-1500 ng/mL. The detection limit for all analytes is less than 1 ng/mL.

Topics: Codeine; Gas Chromatography-Mass Spectrometry; Humans; Morphine; Morphine Derivatives; Opioid-Related Disorders; Postmortem Changes