Noroxycodone Research Articles

Rapid, Multianalyte Detection of Opioid Metabolites in Wastewater.

By monitoring opioid metabolites, wastewater-based epidemiology (WBE) could be an excellent tool for real-time information on the consumption of illicit drugs. A key limitation of WBE is the reliance on costly laboratory-based techniques that require substantial infrastructure and trained personnel, resulting in long turnaround times. Here, we present an aptamer-based graphene field effect transistor (AptG-FET) platform for simultaneous detection of three different opioid metabolites. This platform provides a reliable, rapid, and inexpensive method for quantitative analysis of opioid metabolites in wastewater. The platform delivers a limit of detection 2-3 orders of magnitude lower than previous reports, but in line with the concentration range (pg/mL to ng/mL) of these opioid metabolites present in real samples. To enable multianalyte detection, we developed a facile, reproducible, and high-yield fabrication process producing 20 G-FETs with integrated side gate platinum (Pt) electrodes on a single chip. Our devices achieved the selective multianalyte detection of three different metabolites: noroxycodone (NX), 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), and norfentanyl (NF) in wastewater diluted 20× in buffer.

The Quantification of Oxycodone and Its Phase I and II Metabolites in Urine.

The purpose of this research was to develop and validate an analytical method for the detection and quantification of noroxymorphone-3β-D-glucuronide (NOMG), oxymorphone-3β-D-glucuronide (NOMG), noroxymorphone (NOM), oxymorphone (OM), 6α-oxycodol (αOCL), 6β-oxycodol (βOCL), noroxycodone (NOC) and oxycodone (OC) in urine by liquid chromatography tandem mass spectrometry to be used in a human study. The method was validated according to the Academy Standards Board Standard Practices for Method Development in Forensic Toxicology. The method was then applied to a single-dose pilot study of a subject. Urine samples were collected from the subject after ingesting 10-mg OC as an immediate-release tablet. Additionally, urine specimens (n = 15) that had previously been confirmed positive for OC were analyzed using the validated method. The calibration range for NOMG and OMG was 0.05–10 μg/mL; for all other analytes, it was 0.015–10 μg/mL. Validation parameters such as bias, precision, carryover and dilution integrity, all met the validation criteria. After the method was validated, urine samples from the first subject in the controlled dose study were analyzed. It was observed that OC, NOC and OMG contained the highest concentrations and were present in either the 0.5 or 1 h void. NOC and OMG were detected until the 48 h collection, while OC was detectable till the 24 h collection. Time to reach maximum concentration (Tmax) in the urine was achieved within 1.5 h for OC and within 3 h for NOC and OMG. Maximum concentration (Cmax) in the urine for OC, NOC and OMG was 3.15, 2.0 and 1.56 μg/mg, respectively. OC concentrations in authentic urines ranged from 0.015 to 12 μg/mL. Ranges for NOMG and OMG were 0.054–9.7 μg/mL and 0.14–67 μg/mL, respectively. A comprehensive method for the quantification of NOMG, OMG, NOM, OM, αOCL, βOCL, NOC and OC in urine was optimized and met the validation criteria. The concentrations of NOMG and OMG presented in this study provide the details needed in the forensic community to better comprehend OC pharmacokinetics.

Oxycodone Concentrations and Metabolic Ratios in Femoral Blood from Fatal Intoxications and Other Causes of Death using LC-MS-MS.

Oxycodone (OC) is an opioid with strong analgesic effects widely used to treat acute and chronic pain. Interpretation of OC concentrations in postmortem cases is complicated due to tolerance and overlapping concentrations for fatal and non-fatal levels. In this study, our aim was to develop and validate a method for OC and its three metabolites: noroxycodone (NOC), oxymorphone (OM) and noroxymorphone (NOM) in postmortem femoral blood. Our goal was to define reference concentrations for intoxications and non-intoxications and investigate metabolic ratios in different causes of death. A rapid LC-MS-MS method using protein-precipitated postmortem blood was developed. Lower limit of quantitation was 0.005μg/g blood for all analytes; upper limit of quantitation was 1.0μg/g for OC and NOC and 0.25μg/g for OM and NOM. The method displayed high precision (3.3-7.7%) and low bias (-0.3 to 12%). In total, 192 cases were analyzed and concentrations ranged from 0.005 to 13μg/g for OC, 0.005 to 2.0μg/g for NOC, 0.005 to 0.24μg/g for OM, and 0.005 to 0.075μg/g for NOM. We found a significant difference in OC concentration between the cases where OC contributed and those where it did not. In spite of that, we do not recommend the use of a specific blood concentration to distinguish fatal intoxications. Instead, the percentiles from our data set suggest that concentrations >0.2μg/g are likely to have contributed to toxicity, but that concentrations as high as 0.3 might be tolerated without toxic effects. In addition, we also found that a low NOC/OC ratio could point toward an acute fatal intoxication. In conclusion, the OC concentration alone may not be sufficient to diagnose a fatal intoxication.

Prescription Opioids. I. Metabolism and Excretion Patterns of Oxycodone in Urine Following Controlled Single Dose Administration

The ongoing epidemic of prescription opioid abuse in the United States has prompted interest in semi-synthetic opioids in the federal workplace drug testing program. This study characterized the metabolism and disposition of oxycodone (OC) in human urine. Twelve healthy adults were administered a single oral 20 mg dose of OC in a controlled clinical setting. Urine specimens were collected at timed intervals up to 52 h and analyzed by liquid chromatography-tandem mass spectrometry (limit of quantitation: 50 ng/mL) for OC, oxymorphone (OM), noroxycodone (NOC) and noroxymorphone (NOM) with and without enzymatic hydrolysis. OC and NOC appeared in urine within 2 h, followed by OM and NOM. Peak concentrations of OC and metabolites occurred between 3 and 19 h. Mean peak concentrations in hydrolyzed urine were in the following order: NOC > OC > OM > NOM. Only OM appeared to be excreted extensively as a conjugated metabolite. OC concentrations declined more quickly than NOC and OM. At a cutoff concentration of 50 ng/mL, detection times were approximately 30 h for OC and 40 h for NOC and OM. Some specimens did not contain OC, but most contained NOC, thereby facilitating interpretation that OC was the administered drug; however, five specimens contained only OM. These data provide information that should facilitate the selection of appropriate test parameters for OC in urine and assist in the interpretation of test results.

Analysis of oxycodol and noroxycodol stereoisomers in biological samples by capillary electrophoresis

A capillary electrophoresis (CE) method for the separation of the diastereoisomers of 6-oxycodol (6OCOL) and nor-6-oxycodol (N6OCOL), the 6-keto-reduced metabolites of oxycodone (OCOD) and noroxycodone (NOCOD), respectively, is reported and employed to assess the stereoselectivity of these metabolic steps in vivo, in vitro, and in chemical synthesis. CE in an untreated fused-silica capillary with acidic buffers containing 2-hydroxypropyl-beta-cyclodextrin, randomly sulfated beta-cyclodextrin, or single isomer heptakis(2,3-diacetyl-6-sulfato)-beta-cyclodextrin (HDAS-beta-CD) is shown to permit the simultaneous separation of the stereoisomers of 6OCOL and N6OCOL. A 100 mM phosphate buffer of pH 2.0 containing 2.05% w/v HDAS-beta-CD provides a medium for rapid analysis and unambiguous identification of these stereoisomers in solid-phase extracts of (i) urines stemming from patients under pharmacotherapy with OCOD, (ii) incubations of OCOD and NOCOD with human liver cytosol and the human liver S9 fraction, and (iii) after chemical synthesis from OCOD and NOCOD using NaBH(4). In all cases, alpha-N6OCOL is shown to be the predominant stereoisomer of N6OCOL. For 6OCOL, the same is true for in vitro formation and for chemical synthesis. In urine, however, beta-6OCOL is observed to be excreted in a higher amount than alpha-6OCOL. For the urinary alpha-/beta-isomer ratio of 6OCOL and N6OCOL, there are no differences between the data obtained for nonhydrolyzed and enzymatically hydrolyzed urines. The data document the stereoselectivity of the 6-keto-reduction of OCOD and NOCOD in man.

Determination of Oxycodone and Its Metabolite, Noroxycodone, in Human Plasma by HPLC with Post‐column Chemiluminescence Detection Using Electrogenerated Tris(2,2′‐Bipyridyl)ruthenium(III)

A sensitive high performance liquid chromatography (HPLC) method with post‐column chemiluminescence detection was developed for the simultaneous determination of oxycodone (OXC) and its metabolite, noroxycodone (NOC), in human plasma. Sample preparation for human plasma was performed by means of solid extraction with Sep‐Pak Vac C18 (100 mg/mL). Separation was performed by reversed‐phase HPLC using a 5‐µm Develosil ODS‐HG‐5 column (150 mm × 4.6 mm) and a mobile phase consisting of a mixture of 0.1 mol/L phosphate buffer (pH 2.6) containing 5 mmol/L sodium dodecylsulfate and methanol (53:47, v/v). Quantitation was performed by the measurement of chemiluminescence using electrogenerated tris(2,2′‐bipyridyl)ruthenium(III). The lower and upper limits of quantitation for OXC and NOC were 0.5–250 ng/mL and 1–500 ng/mL, respectively. The calibration curve for this assay showed good linearity for the concentration range of 0.5–50 ng/mL for OXC and 1–100 ng/mL for NOC. Recoveries of the studied compounds were excellent at the individual assay ranges (102.9–108.7%), and validation of the assay gave results that were satisfactory in terms of within‐run or between‐run precision (1.9–11.9% as RSD) and accuracy (−13.5% to +9.2% as bias). Stability studies showed that OXC and NOC were stable in assay solutions for at least 6 days at 4°C storage in the autosampler. OXC and NOC were also stable in human plasma for at least 48 hr at room temperature, for at least 8 months at −20°C and after five repeated freeze–thaw cycles. Furthermore, OXC and NOC were stable in human whole blood for up to 5 days at 4°C and for up to 5 hr at room temperature. This system was applied to the clinical study of cancer patients after single oral administration doses of controlled‐released tablets of OXC hydrochloride. Consequently, this method should serve as a useful tool for pharmacokinetic studies of OXC and NOC.

Simultaneous Determination of Oxycodone and Noroxycodone in Dog Plasma and Urine by HPLC, with Post‐column Chemiluminescence Detection Using Electrogenerated tris(2,2′‐Bipyridyl) Ruthenium(III)

A sensitive and specific method was developed for the determination of oxycodone (OXC) and its metabolite, noroxycodone (NOC), in dog plasma and urine by high performance liquid chromatography (HPLC) with chemiluminescence (CL) detection. Plasma samples were treated with a disposable C18‐bonded column to extract oxycodone and noroxycodone. Urine samples were treated by dilution with water. The analytes were separated on the reversed phase HPLC column and detected by CL with electrochemically oxidized tris(2,2′‐bipyridyl) ruthenium(III). The assay concentration ranges for OXC and NOC were 0.5–250 ng/mL and 2.5–1250 ng/mL in dog plasma, and 0.1–40 µg/mL and 0.5–200 µg/mL in dog urine, respectively. Recoveries of the compounds from plasma and urine were more than 87%. In the within‐run and between‐run assay, the precision values were not more than 13.3%, and the accuracy values were between −3.6% and +15.0%, respectively. The methods were applied to the pharmacokinetic study of oxycodone in dogs.

Capillary electrophoresis and capillary electrophoresis–ion trap multiple-stage mass spectrometry for the differentiation and identification of oxycodone and its major metabolites in human urine

Oxycodone (OCOD) and its metabolites, including oxymorphone (OMOR), noroxycodone (NOCOD) and noroxymorphone (NOMOR), are opioids that carry an OH group at position 14. Using capillary electrophoresis (CE) with a binary phosphate buffer containing 60% ethylene glycol (pH 7.9), the migration order of OCOD and OMOR with respect to their N-demethylated analogs was found to be reversed compared to that observed for codeine, dihydrocodeine, morphine and dihydromorphine, compounds that do not have an OH group at position 14. OCOD and structurally related compounds can also be distinguished from these opioids by their absorbance spectra at low wavelengths and via a characteristic neutral H 2O loss at the MS 2 level. Using the binary phosphate buffer, CE with UV detection is shown to be capable of monitoring OCOD, NOCOD, OMOR (after hydrolysis only) and NOMOR (after hydrolysis and in patient urine only) in alkaline liquid–liquid extracts of urines that were collected after ingestion of 10 mg OCOD hydrochloride and in a patient urine collected at steady state (80 mg OCOD hydrochloride daily). Using an aqueous pH 9 ammonium acetate buffer, these results were confirmed by CE–MS 3. Based on CE–MS, MS 2 and MS 3 data, the absorbance spectra measured across the CE peaks and the relative position within the electropherogram, two peaks monitored in the UV absorbance electropherograms could be assigned to the two keto-reduced metabolites 6oxycodol (6OCOL) and nor6oxycodol, for which no standards were available. Comparison of data obtained with urines pretreated with two different enzyme products (β-glucuronidase and β-glucuronidase/arylsulfatase) suggest that OCOD, NOCOD and 6OCOL are mainly glucuronidated, whereas OMOR mainly forms other conjugates. Furthermore, in a first attempt to directly measure conjugates of the compounds of interest, solid-phase extracts were analyzed by CE–MS 4, which revealed the presence of the acyl glucuronides of 6OCOL and OMOR and an unidentified OMOR conjugate. The quantitation of free OCOD and NOCOD by CE–MS using deuterated internal standards is also discussed briefly.

The antinociceptive potencies of oxycodone, noroxycodone and morphine after intracerebroventricular administration to rats

The antinociceptive activity of noroxycodone (NOR) was determined and its potency compared with that of oxycodone (OXY) and morphine (MOR) after intracerebroventricular (icv) administration to Sprague-Dawley rats. The antinociceptive potencies of OXY and NOR relative to MOR were 0.44 and 0.17, respectively. Administration of naloxone (55 nmol icv) abolished the antinociceptive response produced by the subsequent administration of OXY (227 nmol icv), indicating that the antinociceptive effects of OXY are mediated by opioid receptors. In addition, the same dose of naloxone (55 nmol icv) markedly reduced the antinociceptive effects of NOR (332 nmol icv) and MOR (93 nmol icv) indicating the involvement of opioid receptors in the antinociceptive activities of NOR and MOR. NOR, administered by the icv route, also produced excitatory effects throughout the antinociceptive dose range, the severity of which was reduced but not abolished by prior administration of naloxone (55 nmol icv). As excitatory effects have not been observed in patients receiving OXY, it is unlikely that NOR contributes to the analgesic activity of OXY administered systematically to humans.