BACKGROUND:Buprenorphine is currently under investigation as a pharmacotherapy to treat pregnant women for opioid dependence. This research evaluates buprenorphine (BUP), norbuprenophine (NBUP), buprenorphine-glucuronide (BUP-Gluc), and norbuprenorphine-glucuronide (NBUP-Gluc) pharmacokinetics after high-dose (14-20 mg) BUP sublingual tablet administration in three opioid-dependent pregnant women.METHODS:Oral fluid and sweat specimens were collected in addition to plasma specimens for 24 hours during gestation weeks 28 or 29 and 34, and 2 months after delivery. Time to maximum concentration was not affected by pregnancy; however, BUP and NBUP maximum concentration and area under the curve at 0 to 24 hours tended to be lower during pregnancy compared with postpartum levels.RESULTS:Statistically significant but weak positive correlations were found for BUP plasma and OF concentrations and BUP/NBUP ratios in plasma and oral fluid. Statistically significant negative correlations were observed for times of specimen collection and BUP and NBUP oral fluid/plasma ratios. BUP-Gluc and NBUP-Gluc were detected in only 5% of oral fluid specimens. In sweat, BUP and NBUP were detected in only four of 25 (12 or 24 hours) specimens in low concentrations (less than 2.4 ng/patch).CONCLUSION:These preliminary data describe BUP and metabolite pharmacokinetics in pregnant women and suggest that, like methadone, upward dose adjustments may be needed with advancing gestation.

Introduction: Oral fluid testing is widely used for detecting drug exposure, but data describing methadone and metabolites in oral fluid during pharmacotherapy for opioid-dependence are relatively limited.Methods: 414 oral fluid specimens from 16 opioid-dependent pregnant women receiving daily methadone were analyzed for methadone, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), and methadol by liquid chromatography-mass spectrometry.Results: All oral fluid specimens contained methadone greater than 1 ng/mL; 88% were positive for EDDP and 12% for methadol. Over 95% of oral fluid specimens exceeded the 20 ng/mL methadone cutoff set by the European Driving Under the Influence of Drugs, Alcohol and Medicines (DRUID) study. Methadone and EDDP oral fluid concentrations were highly variable within and between participants, did not predict methadone dose, but were negatively correlated with pH.Conclusion: Methadone was readily identified in oral fluid at concentrations greater than 20 ng/mL following daily 30-110 mg/day methadone pharmacotherapy. As no specimens contained only EDDP or methadol, there was no advantage to including these analytes for identification of methadone exposure. As nearly all oral fluid specimens from methadone-maintained patients exceeded the DRUID guideline, the 20 ng/mL cutoff appears to be sensitive enough to detect daily methadone exposure; however, additional indicators of behavioral and/or motor impairment would be necessary to provide evidence of driving impairment. Published by Elsevier Ireland Ltd.

AIMSMethadone is standard pharmacotherapy for opioid-dependent pregnant women, yet the relationship between maternal methadone dose and neonatal abstinence syndrome (NAS) severity is still unclear. This research evaluated whether quantification of fetal methadone and drug exposure via meconium would reflect maternal dose and predict neonatal outcomes.DESIGNProspective clinical study.SETTINGAn urban drug treatment facility treating pregnant and post-partum women and their children.PARTICIPANTSForty-nine opioid-dependent pregnant women received 30-110 mg methadone daily.MEASUREMENTSMaternal methadone dose, infant birth parameters and NAS assessments were extracted from medical records. Thrice-weekly urine specimens were screened for opioids and cocaine. Newborn meconium specimens were quantified for methadone, opioid, cocaine and tobacco biomarkers.FINDINGSThere was no relationship between meconium methadone concentrations, presence of opioids, cocaine and/or tobacco in meconium, maternal methadone dose or NAS severity. Opioid and cocaine were also found in 36.7 and 38.8 of meconium specimens, respectively, and were associated with positive urine specimens in the third trimester. The presence of opioids other than methadone in meconium correlated with increased rates of preterm birth, longer infant hospital stays and decreased maternal time in drug treatment.CONCLUSIONSMethadone and its metabolite 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) concentrations in meconium did not predict infant birth parameters or NAS severity. Prospective urine testing defined meconium drug detection windows for opiates and cocaine as 3 months, rather than the currently accepted 6 months. The presence of opioids in meconium could be used as a biomarker for infants at elevated risk in the newborn period.

Sweat patches (n = 350) were collected throughout gestation from 29 opioid-dependent pregnant women participating in an outpatient methadone-assisted therapy program. Volunteers provided informed consent to participate in institutional review board-approved protocols. Methadone was eluted from sweat patches with sodium acetate buffer, followed by solid-phase extraction and quantification by gas chromatography mass spectrometry (limit of quantification >= 10 ng/patch). Methadone was present in all weekly patches (n = 311) in concentrations ranging from 10.2 to 12,129.7 nanograms per patch and in 92.3% of short-term patches (n = 39, worn for 12 or 24 hours) in concentrations up to 3303.9 nanograms per patch. Correlation between patch concentrations and total amount of drug administered (r = 0.224), and concentrations and duration of patch wear (r = 0.129) were both weak. Although there were large intra-and intersubject variations in sweat drug concentrations, sweat testing was an effective alternative technique to qualitatively monitor illicit drug use and simultaneously document methadone medication-assisted treatment.

Dependence on illicit drugs during pregnancy is a major public health concern as there may be associated adverse maternal, fetal, and neonatal consequences. Sweat patches (n = 389) were collected from 39 pregnant volunteers who provided written informed consent for this Institutional Review Board-approved protocol and wore patches, replaced approximately weekly, from study entry until delivery. Patches were analyzed for opiates (heroin, 6-acetylmorphine, 6-acetylcodeine, morphine and codeine) and cocaine (cocaine, benzoylecgonine, ecgonine methyl ester, anhydroecgonine methyl ester) by solid phase extraction and gas chromatography mass spectrometry. Seventy-one percent (276) of collected sweat patches were ≥5 ng per patch (limit of quantification) for one or more analytes. Cocaine was present in 254 (65.3%) patches in concentrations ranging from 5.2 to 11,835 ng per patch with 154 of these high enough to satisfy the proposed Substance Abuse and Mental Health Services Administration guidelines for a confirmatory drug test (25 ng per patch). Interestingly, 6-acetylmorphine was the most prominent opiate analyte documented in 134 patches (34.4%) with 11.3% exceeding the proposed opiate Substance Abuse and Mental Health Services Administration cut-off (25 ng per patch). Heroin was identified in fewer patches (77), but in a similar concentration range (5.3-345.4 ng per patch). Polydrug use was evident by the presence of both cocaine and opiate metabolites in 136 (35.0%) patches. Sweat testing is an effective method for monitoring abstinence or illicit drug use relapse in this high-risk population of pregnant opiate- and/or cocaine-dependent women.

Buprenorphine is under investigation in the U.S. as pharmacotherapy for opioid-dependent pregnant women. Buprenorphine and metabolites were quantified in umbilical cord specimens from women receiving daily buprenorphine doses. Correlations between maternal buprenorphine dose, buprenorphine and metabolite umbilical cord concentrations, and neonatal outcomes were investigated, as well as the ability to identify heroin and cocaine relapse during pregnancy. Umbilical cord concentrations were compared to those of matched umbilical cord plasma and meconium. Buprenorphine metabolites were detected in all cords, but buprenorphine itself was absent. Concentration ranges were 1.2-5.1 ng/g norbuprenorphine, 1.7-4.2 ng/g buprenorphine-glucuronide, and 8.3-23 ng/g norbuprenorphine-glucuronide. Cord concentrations were similar to those in plasma, and lower (16-210-fold), although statistically correlated, than those in meconium. Significant positive correlations were observed for buprenorphine-glucuronide concentrations in umbilical cord and mean maternal BUP daily dose throughout pregnancy and third trimester, but buprenorphine biomarker concentrations did not predict neonatal outcomes. Opiate concentrations were lower (200-fold) in umbilical cord than in meconium, and when cocaine was present in meconium, it was not identified in cord. Umbilical cord can serve as an alternative matrix for identifying prenatal drug-exposure, but is much less sensitive than meconium. Buprenorphine provided a controlled drug administration model for evaluating drug disposition in the maternal-fetal dyad.

Buprenorphine is approved as pharmacotherapy for opioid dependence in nonpregnant patients in multiple countries and is currently under investigation for pregnant women in the United States and Europe. This research evaluates the disposition of buprenorphine, opiates, cocaine, and metabolites in five term placentas from a US cohort. Placenta and matched meconium concentrations were compared, and relationships among maternal buprenorphine dose, placenta concentrations, and neonatal outcomes after controlled administration during gestation were investigated. Buprenorphine and/or metabolites were detected in all placenta specimens and were uniformly distributed across this tissue (coefficient of variation less than 27.5%, four locations), except for buprenorphine in three placentas. In two of these, buprenorphine was not detected in some locations and in the third placenta was totally absent. Median (range) concentrations were 1.6 ng/g buprenorphine (not detected to 3.2), 14.9 ng/g norbuprenorphine (6.2-24.2), 3 ng/g buprenorphine-glucuronide (1.3-5.0), and 14.7 ng/g norbuprenorphine-glucuronide (11.4-25.8). Placenta is a potential alternative matrix for detecting in utero buprenorphine exposure, but at lower concentrations (15- to 70-fold) than in meconium. Statistically significant correlations were observed for mean maternal daily dose from enrollment to delivery and placenta buprenorphine-glucuronide concentration and for norbuprenorphine-glucuronide concentrations and time to neonatal abstinence syndrome onset and duration, for norbuprenorphine/norbuprenorphine-glucuronide ratio and maximum neonatal abstinence syndrome score, and newborn length. Analysis of buprenorphine and metabolites in this alternative matrix, an abundant waste product available at the time of delivery, may be valuable for prediction of neonatal outcomes for clinicians treating newborns of buprenorphine-exposed women.

Opiates, cocaine, and metabolites were quantified by liquid chromatography-mass spectrometry (LC-MS) in 284 urine specimens, collected thrice weekly, to monitor possible drug relapse in 15 pregnant heroin-dependent women. Opiates were detected in 149 urine specimens (52%) with limits of quantification (LOQ) of 10-50 microg/L. Morphine, morphine-3-glucuronide, and/or morphine-6-glucuronide were positive in 121 specimens; 6-acetylmorphine, a biomarker of heroin ingestion, was quantifiable in only 7. No heroin, 6-acetylcodeine, papaverine, or noscapine were detected. One hundred and sixty-five urine specimens (58%) from all 15 participants were positive for one or more cocaine analytes (LOQ 10-100 microg/L). Ecgonine methylester (EME) and/or benzoylecgonine were the major cocaine biomarkers in 142. Anhydroecgonine methylester, a biomarker of smoked cocaine, was positive in six; cocaethylene and/or ecgonine ethylester, biomarkers of cocaine and ethanol co-ingestion, were found in 25. At the current Substance Abuse Mental Health Services Administration cutoffs for total morphine (2000 microg/L), codeine (2000 microg/L), 6-acetylmorphine (10 microg/L), and benzoylecgonine (100 microg/L), 16 opiate- and 29 cocaine-positive specimens were identified. Considering 100 microg/L EME as an additional urinary cocaine biomarker would identify 51 more positive cocaine specimens. Of interest is the differential pattern of opiate and cocaine biomarkers observed after LC-MS as compared to gas chromatography-mass spectrometry analysis.

BACKGROUND:Buprenorphine (BUP) is under investigation as a medication therapy for opioid-dependent pregnant women. We investigated BUP and metabolite disposition in urine from women maintained on BUP during the second and third trimesters of pregnancy and postpartum.METHODS:We measured BUP, norbuprenorphine (NBUP), buprenorphine glucuronide (BUP-Gluc), and NBUP-Gluc concentrations in 515 urine specimens collected thrice weekly from 9 women during pregnancy and postpartum. Specimens were analyzed using a fully validated liquid chromatography-mass spectrometry method with limits of quantification of 5 microg/L for BUP and BUP-Gluc and 25 microg/L for NBUP and its conjugated metabolite. We examined ratios of metabolites across trimesters and postpartum to identify possible changes in metabolism during pregnancy.RESULTS:NBUP-Gluc was the primary metabolite identified in urine and exceeded BUP-Gluc concentrations in 99% of specimens. Whereas BUP-Gluc was identified in more specimens than NBUP, NBUP exceeded BUP-Gluc concentrations in 77.9% of specimens that contained both analytes. Among all participants, the mean BUP-Gluc:NBUP-Gluc ratio was significantly higher in the second trimester compared to the third trimester, and there were significant intrasubject differences between trimesters in 71% of participants. In 3 women, the percent daily dose excreted was higher during pregnancy than postpregnancy, consistent with other data indicating increased renal elimination of drugs during pregnancy.CONCLUSIONS:These data are the first to evaluate urinary disposition of BUP and metabolites in a cohort of pregnant women. Variable BUP excretion during pregnancy may indicate metabolic changes requiring dose adjustment during later stages of gestation.

OBJECTIVE. In a sample of methadone-maintained breastfeeding women and a matched group of formula-feeding women, this study evaluated concentrations of methadone in breast milk among breastfeeding women and concentrations of methadone in maternal and infant plasma in both groups.METHODS. Eight methadone-maintained (dose: 50-105 mg/day), lactating women provided blood and breast milk specimens on days 1, 2, 3, 4, 14, and 30 after delivery, at the times of trough and peak maternal methadone levels. Paired specimens of foremilk and hindmilk were obtained at each sampling time. Eight matched formula-feeding subjects provided blood samples on the same days. Infant blood samples for both groups were obtained on day 14. Urine toxicological screening between 36 weeks of gestation and 30 days after the birth confirmed that subjects were not using illicit substances in the perinatal period.RESULTS. Concentrations of methadone in breast milk were low (range: 21.0-462.0 ng/mL) and not related to maternal dose. There was a significant increase in methadone concentrations in breast milk over time for all 4 sampling times. Concentrations of methadone in maternal plasma were not different between groups and were unrelated to maternal dose. Concentrations of methadone in infant plasma were low (range: 2.2-8.1 ng/mL) in all samples. Infants in both groups underwent neurobehavioral assessments on days 3, 14, and 30; there were no significant effects of breastfeeding on neurobehavioral outcomes. Fewer infants in the breastfed group required pharmacotherapy for neonatal abstinence syndrome, but this was not a statistically significant finding.CONCLUSION. Results contribute to the recommendation of breastfeeding for methadone-maintained women.

Breastfeeding among methadone-maintained women is frequently challenged because of unclear guidelines regarding this practice. Previous research has confirmed that concentrations of methadone in breastmilk in the neonatal period are low. Currently unknown are the concentrations of methadone in breastmilk among women who breastfeed for longer periods of time. The purpose of this research is to examine concentrations of methadone in the plasma and breastmilk of women who breastfeed their infants beyond the neonatal period. Four methadone-maintained women provided blood and breastmilk samples up to 6 months postpartum. The concentrations of methadone in blood and breastmilk were low, contributing to the recommendation of breastfeeding for some methadone-maintained women.

For the first time, relationships among maternal buprenorphine dose, meconium buprenorphine and metabolite concentrations, and neonatal outcomes are reported. Free and total buprenorphine and norbuprenorphine, nicotine, opiates, cocaine, benzodiazepines, and metabolites were quantified in meconium from 10 infants born to women who had received buprenorphine during pregnancy. Neither cumulative nor total third-trimester maternal buprenorphine dose predicted meconium concentrations or neonatal outcomes. Total buprenorphine meconium concentrations and buprenorphine/norbuprenorphine ratios were significantly related to neonatal abstinence syndrome (NAS) scores >4. As free buprenorphine concentration and percentage free buprenorphine increased, head circumference decreased. Thrice-weekly urine tests for opiates, cocaine, and benzodiazepines and self-reported smoking data from the mother were compared with data from analysis of the meconium to estimate in utero exposure. Time of last drug use and frequency of use during the third trimester were important factors associated with drug-positive meconium specimens. The results suggest that buprenorphine and metabolite concentrations in the meconium may predict the onset and frequency of NAS.

This study evaluates concentrations of methadone in breast milk and plasma among a sample of methadone-maintained women in the immediate perinatal period. Twelve methadone-maintained, lactating women provided blood and breast milk specimens 1, 2, 3, and 4 days after delivery. Specimens were collected at the time of trough (just before methadone dose) and peak (3 hours after dosing) maternal methadone levels. Paired specimens of foremilk (prefeed) and hindmilk (postfeed) were obtained at each sampling time. Although there was a significant increase in methadone concentration in breast milk over time for the peak postfeed sampling time, t (22) = 2.40, P = .0255, methadone concentrations in breast milk were small, ranging from 21 to 314 ng/mL, and were unrelated to maternal methadone dose. Results obtained from this study contribute to the recommendation of breastfeeding for methadone-maintained women regardless of methadone dose. J Hum Lact. 23(2):184-190.

Interest in oral fluid as an alternative matrix for monitoring drug use is due to its ease-of-collection and non-invasiveness; however, limited data are available on the disposition of drugs into oral fluid. The objective of this research was to provide data on the presence and concentrations of heroin, cocaine and multiple metabolites in oral fluid after illicit opioid and cocaine use. Thrice weekly oral fluid specimens (N = 403) from 16 pregnant opiate-dependent women were obtained with the Salivette® oral fluid collection device. Evidence of heroin (N = 62) and cocaine (N = 130) use was detected in oral fluid by LC–APCI-MS/MS. 6-Acetylmorphine (6-AM), heroin and morphine were the major opiates detected, with median concentrations of 5.2, 2.3, and 7.5 μg/L, respectively. Cocaine and benzoylecgonine (BE) had median concentrations of 6.4 and 3.4 μg/L. Application of the Substance Abuse Mental Health Services Administration (SAMHSA) recommended cutoffs for morphine and codeine (40 μg/L), 6-AM (4 μg/L) and cocaine and BE (8 μg/L), yielded 28 opiate- and 50 cocaine-positive specimens. Oral fluid is a promising alternative matrix to monitor opiate and cocaine use in drug testing programs. These data guide interpretation of oral fluid test results and evaluate currently proposed SAMHSA oral fluid testing cutoffs.

BACKGROUNDBuprenorphine is under investigation as a pharmacotherapeutic agent for treating opioid dependence in pregnant women. We hypothesized that there would be a relationship between the cumulative maternal dose of buprenorphine during pregnancy and the concentration of buprenorphine and norbuprenorphine in maternal and infant hair.METHODSThis study examined buprenorphine and norbuprenorphine concentrations in hair obtained from 9 buprenorphine-maintained pregnant women and 4 of their infants. Specimens were analyzed by liquid chromatography-tandem mass spectrometry with limits of quantification of 3.0 pg/mg. All maternal hair specimens were washed with methylene chloride before analysis, and when sufficient amounts of maternal hair were available, specimens also were analyzed without washing. Infant hair specimens were not washed.RESULTSBuprenorphine concentrations were significantly greater in unwashed hair than washed hair (P = 0.031). Norbuprenorphine concentrations were significantly greater than buprenorphine concentrations in both maternal (P = 0.0097) and infant hair (P = 0.0033). There were statistically significant associations between the cumulative maternal dose of buprenorphine administered and the concentrations of buprenorphine (washed, P <0.0001; unwashed, P = 0.0004), norbuprenorphine (washed, P <0.0001; unwashed, P = 0.0005), and buprenorphine plus norbuprenorphine (washed, P <0.0001; unwashed, P = 0.0005) for both washed and unwashed maternal hair specimens. There was a significant positive association between concentrations of buprenorphine and norbuprenorphine in maternal hair (washed, P <0.0001; unwashed, P = 0.0003), a trend for this association in infant hair (P = 0.08), and an association between buprenorphine concentrations in maternal unwashed hair and infant hair (P = 0.0002). The buprenorphine:norbuprenorphine ratio increased in distal segments.CONCLUSIONBuprenorphine treatment during gestation provides an opportunity for monitoring drug disposition in maternal and fetal tissues under controlled conditions.

This manuscript details a validated liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry (LC-APCI-MS-MS) method for the quantification of methadone and its metabolites 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) and 2-ethyl-5-methyl-3,3-diphenylpyroline (EMDP) in 0.5 mL human breast milk. Limits of detection were 5 ng/mL for methadone and EDDP, and 10 ng/mL for EMDP. Linearity ranged from 10 to 500 ng/mL for all analytes. Breast milk is a complex biological fluid, necessitating several specimen preparation steps to separate methadone and metabolites from the lipophilic matrix. Recoveries were 66-97% following protein precipitation and solid-phase extraction with minimal matrix effect. Acceptable accuracy (89-101%) and precision (15-20% RSD) were achieved for all analytes. This is the first LC-APCI-MS-MS method for the sensitive and specific detection of methadone, EDDP, and EMDP in human breast milk. The method proved suitable for quantification of methadone and metabolites in breast milk of methadone-maintained opiate-dependent women.

This paper details a validated liquid chromatography atmospheric pressure chemical ionization tandem mass spectrometry (LC–APCI–MS/MS) method for the quantification of methadone, and its metabolites 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), 2-ethyl-5-methyl-3,3-diphenylpyraline (EMDP) and methadol in human meconium. Limits of detection (LOD) were determined to be 1.0 ng/g for methadone, EDDP and EMDP and 2.5 ng/g for methadol. The limits of quantitation (LOQ) for methadone, EDDP, EMDP were 5 and 25 ng/g for methadol. Linearity ranged from 5.0 to 500 ng/g. Following solid-phase extraction, no matrix effect was observed. This method proved to be suitable for the quantification of methadone, EDDP and EMDP and the semi-quantitation of methadol in meconium. Literature review revealed no other published LC–APCI–MS/MS method for the detection of methadone and its three main metabolites in meconium specimens.

The social and economic impact of drug use on our global population continues to increase leaving no geographical, social or cultural group untouched. The National Institute on Drug Abuse (NIDA), in one of the few large surveys of maternal abuse, found that 5.5% of mothers reported taking an illicit substance during gestation. These figures certainly are underestimates due to the stigma of drug use during pregnancy and the accompanying legal, ethical and economic issues. Although drugs of choice and routes of administration vary by country, exposure of our most valuable resource, our children, to the developmental effects of drugs is an enormous problem. In utero drug exposure can have a severe impact not only on the development of the fetus, but also on the child during later stages of life. More than 75% of infants exposed to drugs have major medical problems as compared to 27% of unexposed infants. The cost of treating drug-affected infants was twice the cost of non-affected infants. Obstetrical complications including placental insufficiency, miscarriage, intrauterine death, and increased incidence of infectious and sexually-transmitted diseases are higher in the drug-abusing mother. Treatment for pregnant addicts should be a high priority for our governments. Increased awareness and improvement in our understanding of drug abuse in the medical, legal and social realms will enable us to reduce the barriers to treatment for this important population.Accurate identification of in utero drug exposure has important implications for the care of the mother and child, but can raise difficult legal issues. Society discourages prenatal care with the infliction of harsh criminal penalties. Maternal drug use during pregnancy can be monitored with urine, sweat, oral fluid and/or hair testing. Detection of in utero drug exposure has traditionally been accomplished through urine testing; however, the window of detection is short, reflecting drug use for only a few days before delivery. Monitoring exposure through testing of alternative matrices, such as neonatal meconium and hair, offers advantages including non-invasive collection and detection earlier in gestation. There are many unresolved issues in monitoring in utero drug exposure that urgently require research. These can be divided into research to definitively differentiate drug exposed and non-drug-exposed fetuses, determine the most efficient methods to routinely monitor women's drug use, and determine how these drug test results relate to neonatal and maternal outcomes. Research in this area is difficult and expensive to perform, but necessary to assess accurately drug effects on the fetus. By increasing our understanding of the physiological, biochemical and behavioral effects of gestational drug exposure, we may ultimately provide solutions for better drug prevention, treatment and a reduction in the number of drug-exposed children.

0.5 1.0 Y = -0.0033+0.0054*X 0.995 1 - 500 m-hydroxybenzoylecgonine benzoylecgonine-d8 0.5 1.0 Y = -0.0034+0.0066*X 0.992 1 - 500 Benzoylnorecgonine benzoylecgonine-d8 0.5 1.0 Y = 0.0103+0.0056*X 0.996 1 - 500 Benzoylecgonine benzoylecgonine-d8 0.25 0.5 Y = -0.0029+0.0099*X 0.993 0.5 - 500 p-hydroxycocaine norcocaine-d3 0.5 1.0 Y = -0.0023+0.0055*X 0.993 1 - 500 m-hydroxycocaine norcocaine-d3 0.5 1.0 Y = -0.0052+0.0091*X 0.991 1 - 500 Norcocaine norcocaine-d3 0.5 1.0 Y = -0.0063+0.0118*X 0.992 1 - 500 Cocaine cocaine-d8 0.5 1.0 Y = -0.0004+0.0106*X 0.995 1 - 500 Norcocaethylene cocaethylene-d8 0.5 1.0 Y = 0.0027+0.0123*X 0.996 1 - 500 Cocaethylene cocaethylene-d8 0.5 1.0 Y = -0.0010+0.0105*X 0.995 1 - 500 Limits of quantitation (LOQ, concentration with accuracy and precision < 20%) were established in the low ng/mL range, i.e. 0.5-10 ng/mL. Intra- and inter-day reproducibility, as well as accuracy, were investigated at 3 concentrations, i.e. low, medium, and high. An overview of the results is given in Table 3.