Opiate concentrations following the ingestion of poppy seed products – evidence for `the poppy seed defence'

The universally accepted 300 ng/ml cut-off limit for opiate assays stated to be mandatory for all drug screening laboratories by the Substance Abuse and Mental Health Services Administration, has been questioned recently due to positive results being obtained following the ingestion of poppy seed containing food products. To establish the plausibility of the `the poppy seed defence' the concentrations of codeine, norcodeine, morphine, normorphine and thebaine (a potential marker for seed ingestion) in several varieties of poppy seeds from different countries were quantified by GC–MS. The country of origin of the seed specimen analysed and the preparation of the seeds prior to their culinary use was found to influence the alkaloid concentration determined. The maximum morphine and codeine concentrations determined in the seeds were found to be 33.2 and 13.7 μg/g seed respectively. In addition, thebaine concentrations were found to vary with each seed sample analysed. Following the consumption of bread rolls (mean 0.76 g seed covering per roll) by four subjects, all urine specimens analysed produced negative results (using the Dade Bebring EMIT II opiate screening assay) with the exception of one subject (body weight 63.0 kg) who consumed two poppy seed rolls. In this subject opiate positive screening results were obtained for up to 6 h post ingestion with maximum urinary morphine and codeine concentrations of 832.0 ng/ml (@ 2–4 h post ingestion) and 47.9 ng/ml (@ 0–2 h post ingestion) respectively being achieved. Following the ingestion of poppy seed cake containing an average of 4.69 g of seed per slice by four individuals, opiate positive screening results were obtained for up to 24 h. In one subject (dose equivalent to 0.07 g poppy seed/kg body weight) maximum urinary morphine and codeine concentrations of 302.1 ng/ml (@ 0–2 h) and 83.8 ng/ml (@ 2–4 h) respectively were recorded. The elimination of thebaine was found to vary widely between individuals, therefore suggesting that its absence from a specimen is not necessarily indicative of opiate abuse. These findings demonstrate that the poppy seed defence could be used as an argument in medico-legal and employment medical cases. Great care should therefore be taken when interpreting the data produced when screening for opiates.     

Poppy seed ingestion: the Oregon perspective.

Numerous articles have been published regarding the positive morphine and codeine urinalysis results from the ingestion of poppy seeds. Oregon's perspective towards ingestion of controlled substances focuses around driving under impaired conditions. To determine the influence of the residual opium on poppy seeds to impairment, seven volunteers each ingested 25 grams of poppy seeds baked into bundt cakes. Urine samples were screened by EMIT using 300 ng/ml cutoff levels. All of the urine specimens were found to be opiate positive shortly after consuming the cake; however, after administering a series of standardized drug recognition evaluation tests, no subjects were found to exhibit symptoms of opiate impairment.     

Gas chromatographic/mass spectrometric analysis of morphine and codeine in human urine of poppy seed eaters

In this study, poppy seeds were examined for a natural constituent that might serve as a maker for the seeds' ingestion as opposed to opiate abuse. Thebaine was selected as possible marker, since it was found to be a component of all poppy seeds examined and was not a natural component of different heroin samples. During the course of this investigation, a new extraction and cleanup procedure was developed for the gas chromatographic/nitrogen phosphorus detection (GC/NPD) and gas chromatographic/mass spectrometric (GC/MS) analysis of morphine and codeine in urine. A linear response, over a concentration range of 25 to 600 ng/mL, was obtained for codeine and morphine (r = 0.9982 and 0.9947, respectively). The minimum detectable level (LOD) and limit of quantitation (LOQ) for morphine were 10 and 30 ng/mL, respectively; whereas LOD and LOQ for codeine were 2 and 8 ng/mL, respectively. The coefficients of variance (CV, n = 6) for morphine and codeine analyses at the 100-ng/mL level were 13.3 and 4.6%, respectively. This procedure was used for the analysis of urine samples from five poppy seed eaters who each ingested 200 g of poppy seed cake. Results indicated that significant amounts of morphine and codeine are excreted in urine and that in all subjects, at least at one point in time, the apparent morphine concentration as determined by radioimmunoassay (RIA) analysis exceeded the cutoff value (300 ng/mL) established for screening. Thebaine was not detected in urine specimens collected following poppy seeds ingestion and thus could not be used as a marker.     

Poppy seed ingestion as a contributing factor to opiate-positive urinalysis results: the Pacific perspective.

The possible contribution of poppy seed foods to positive opiate urinalysis results, especially from foods available in the Pacific Rim area, has recently become an issue for the U.S. Army Forensic Toxicology Drug Testing Laboratory in Hawaii. To assess the likelihood of this possible contribution, seven different poppy seed food products were consumed by male and female volunteers, and urine specimens were collected at time increments up to either 24 or 72 h. Specimens were evaluated for opiates using Roche Abuscreen radioimmunoassay (RIA), and all RIA positive specimens were analyzed for morphine and codeine using gas chromatography/mass spectrometry (GC-MS). Poppy seed cake, bagels, muffins, and rolls did not contain sufficient quantities of poppy seeds to give rise to opiate positive specimens by U.S. Department of Defense (DOD) GC-MS cutoff levels (morphine = 4000 ng/mL, codeine = 2000 ng/mL), although a number of specimens were positive by National Institute on Drug Abuse (NIDA) cutoff levels (morphine and codeine = 300 ng/mL). However, ingestion of poppy seed streusel or Danish pastry led to confirmed morphine and codeine positive specimens, irrespective of the use of DOD or NIDA confirmation cutoff values. In addition, significant amounts of codeine were observed in a number of these specimens. These findings argue against the unqualified application of previously published quantitative guidelines for eliminating poppy seed ingestion as a possible cause for a positive opiate urinalysis result.     

Poppy seed consumption and toxicological analysis of blood and urine samples

Poppy seeds contain morphine in different amounts. Reported concentrations are up to 294 mg morphine/kg poppy seeds. Since penalties based on Street Traffic Law (parapgraph 24a StVG) in Germany (administrative offence) require definitive proof of morphine in blood samples, and the "Grenzwertkommission" in consultation with the Ministry of Transportation recommended a threshold of free morphine of 10 ng/mL, the question arose whether the consumption of poppy seeds can lead to a blood concentrations equal or higher than 10 ng/mL of free morphine. Therefore, five volunteers ate poppy seed products (50 mg morphine/kg poppy seeds). In urine, all on-site tests were enzyme immunologically positive for opiates and were positive to morphine by GC/MS. All the blood samples were negative to morphine by EIA and to free morphine by GC/MS. However, after hydrolysis, morphine was detected by GC/MS in all cases. Accordingly, in Germany, penalties based on parapgraph 24a StVG are not likely to cause road users any concerns should they have consumed poppy seeds. Driver Licensing Authorities, however, should be advised of this problem to avoid unjustified legal measures.     

The GC-MS detection and characterization of reticuline as a marker of opium use

Reticuline (a precursor of opium alkaloids) was detected and characterised as its trimethylsilyl ethers, acetyl esters and methyl ethers by GC-EIMS and GC-CIMS in opium and the urine of opium users after hydrolysis by acid or beta-glucuronidase as coextractive of morphine. Because this compound cannot be detected in heroin and poppy seeds, it is suggested as a differentiating marker between opium and heroin use, opium and poppy seeds use, or opium and "pharmaceutical" codeine use in cases when opiate use has been confirmed by detection of morphine and codeine in the urine. As well as being a constituent of opium, reticuline in the urine of opium users may also result from the metabolic demethylation of the three other benzyltetrahydroisoquinoline opium alkaloids: codamine, laudanosine and laudanine.     

Quantification of Morphine,Codeine, and Thebaine in Home‐Brewed Poppy Seed Tea by LC‐MS/MS

Recently, medical examiners reported two cases of a 21‐year‐old male and 24‐year‐old male with high amounts of morphine in their blood at autopsy. It was suspected that the decedents ingested lethal amounts of morphine from home‐brewed poppy seed tea. No studies to date have investigated opium alkaloid content extracted from poppy seeds by home‐brewing methods. Various poppy seed products were purchased from online sources and extracted with four home‐brewing methods representative of recipes found on drug user forums. Morphine, codeine, and thebaine were quantified in the tea extracts by liquid chromatography‐tandem mass spectrometry using a validated analytical method. Morphine, codeine, and thebaine concentrations from seeds were <1–2788 mg/kg, <1–247.6 mg/kg, and <1–124 mg/kg, respectively. Alkaloid yield varied between extractions, but regardless of extraction conditions, lethal amounts of morphine can be rinsed from poppy seed coats by home‐brewing methods.     

The GC–MS detection and characterization of reticuline as a marker of opium use

Reticuline (a precursor of opium alkaloids) was detected and characterised as its trimethylsilyl ethers, acetyl esters and methyl ethers by GC–EIMS and GC–CIMS in opium and the urine of opium users after hydrolysis by acid or β-glucuronidase as coextractive of morphine. Because this compound cannot be detected in heroin and poppy seeds, it is suggested as a differentiating marker between opium and heroin use, opium and poppy seeds use, or opium and “pharmaceutical” codeine use in cases when opiate use has been confirmed by detection of morphine and codeine in the urine. As well as being a constituent of opium, reticuline in the urine of opium users may also result from the metabolic demethylation of the three other benzyltetrahydroisoquinoline opium alkaloids: codamine, laudanosine and laudanine.     

Fatal intoxications in Denmark following intake of morphine from opium poppies

In Denmark it is legal to grow opium poppies for the production of poppy seeds and until 1986 for decoration purposes, too. Danish poppy capsules contain 0.3-5 mg morphine per capsule and the content of morphine in opium exuded from the capsules may amount to 24%. This has resulted in misuse as both fresh and dried poppy capsules have been used for the production of "opium tea". During the period 1982-1985, seven casualties occurred among drug addicts in Denmark which were solely or partly caused by these opium poppies.     

Fatal versus non-fatal heroin "overdose": blood morphine concentrations with fatal outcome in comparison to those of intoxicated drivers

The study was performed to distinguish fatal from non-fatal blood concentrations of morphine. For this purpose, blood levels of free morphine and total morphine (free morphine plus morphine conjugates) in 207 cases of heroin-related deaths were compared to those in 27 drivers surviving opiate intoxication. The majority of both survivors and non-survivors were found to show a concomitant use of depressants including alcohol or stimulants. Blood morphine levels in both groups varied widely, with a large area of overlap between survivors (free morphine: 0-128 ng/ml, total morphine: 10-2,110 ng/ml) and non-survivors (free morphine: 0-2,800 ng/ml, total morphine: 33-5,000 ng/ml). Five (18.5%) survivors and 87 (42.0%) non-survivors exhibit intoxication only by morphine. In these cases, too, both groups overlapped (survivors-free morphine: 28-93 ng/ml, total morphine: 230-1,451 ng/ml; non-survivors-free morphine: 0-2,800 ng/ml, total morphine: 119-4,660 ng/ml). Although the blood levels of free or total morphine do not allow a reliable prediction of survival versus non-survival, the ratio of free/total morphine may be a criterion to distinguish lethal versus survived intoxication. The mean of the ratio of free to total morphine for all lethal cases (N=207) was 0.293, for those that survived (N=27) 0.135, in cases of intoxication only by morphine 0.250 (N=87) and 0.080 (N=5), respectively. Applying a cut-off of 0.12 for free/total morphine and performing ROC analyses, fatal outcome can be predicted in 80% of the cases correctly, whereas 16% of the survivors were classified as dead. Nevertheless, in this study, all cases with a blood concentration of 200 ng/ml and more of free morphine displayed a fatal outcome.     

中毒致死者体内芬氟拉明的GC/NPD,GC/MS法测定

建立了生物检材中芬氟拉明的定性定量分析方法。体液及脏器组织经有机溶剂提取后 ,用GC/MS法进行药物筛选、定性 ,生物检材中的芬氟拉明浓度用4 -苯丁胺作内标、GC/NPD法测定。测得芬氟拉明中毒致死者的血液、尿液、肝等组织中浓度分别为7.8μg/ml、64.2μg/ml、31.3μg/g。并对尸体解剖所见及方法可行性进行讨论     

The detection of acetylcodeine and 6-acetylmorphine in opiate positive urines

Acetylcodeine (AC), an impurity of illicit heroin synthesis, was investigated as a urinary biomarker for detection of illicit heroin use. One hundred criminal justice urine specimens that had been confirmed positive by GC/MS for morphine at concentrations >5000 ng/ml were analyzed for AC, 6-acetylmorphine (6AM), codeine, norcodeine and morphine. The GC/MS analysis was performed by solid phase extraction and derivatization with propionic anhydride. Total codeine and morphine concentrations were determined by acid hydrolysis and liquid/liquid extraction. AC was detected in 37 samples at concentrations ranging from 2 to 290 ng/ml (median, 11 ng/ml). 6AM was also present in these samples at concentrations ranging from 49 to 12 600 ng/ml (median, 740 ng/ml). Of the 63 specimens negative for AC, 36 were positive for 6AM at concentrations ranging from 12 to 4600 ng/ml (median, 124 ng/ml). When detected, the AC concentrations were an average of 2.2% (0.25 to 10.2%) of the 6AM concentrations. There was a positive relationship between AC concentrations and 6AM concentrations (r=0.878). Due to its very low concentration in urine, AC was found to be a much less reliable biomarker for illicit heroin use than 6AM in workplace or criminal justice urine screening programs. However, AC detection could play an important role in determining if addicts in heroin maintenance programs are supplementing their supervised diacetylmorphine doses with illicit heroin.     

Mixed fatal poisoning caused by taking L-methadone and chloral hydrate

A 42-year-old female drug user who was enrolled in a methadone maintenance program was found dead in her apartment. Cause of death was an intoxication with chloral hydrate and L-methadone. Trichloroethanol (TCE), the primary metabolite of chloral hydrate, was quantified by solid phase microextraction (SPME) and GC/MS in heartblood (27 micrograms/ml) and urine (338 micrograms/ml). D- and L-methadone were differentiated by chiral HPLC, which showed that only L-methadone had been taken. The quantitation of L-methadone and its metabolite EDDP was carried out by GC/MS from heartblood (1300 ng/ml and 86 ng/ml, respectively), urine (5239 ng/ml and 4960 ng/ml, respectively) and gastric contents (159 ng/ml and 122 ng/ml, respectively). The concentrations of both--trichloroethanol and methadone--were in toxic ranges.     

The stability of tetramine, morphine and meperidine in formalin solution.

The stability of tetramine, morphine and meperidine in formalin solution is an important factor for drug analysis in forensic investigation. In this paper, the tissues (liver, kidney, lung and heart) from poisoned rabbits were immersed in 50 ml 10% formalin solutions for 4 months before examination. We compared the levels of tetramine, morphine, meperidine and the main metabolite normeperidine, measured by GC/NPD or GC-MS, in frozen rabbit tissues, formalin-fixed rabbit tissues, and formalin solution. There was a decrease in the levels of tetramine, morphine, meperidine in formalin-preserved tissues compared with the levels of these drugs in the frozen tissues. It is suggested that the formalin-fixed tissues and formalin solution should be analyzed at the same time to assure the accurate results.     

尿中氯喹定性定量分析方法的研究

建立了人尿中氯喹的定性定量分析方法,2ml尿样用2ml×2环己烷：乙酸乙酯（8：2）提取净化后,60℃水浴室气吹干,残留物定容溶解后,气相色谱分析,氯喹的保留时间为9．44min。方法最低检测限为200ng／ml,回收率为87．0％,RSD＝7．9％（n＝5）,在0～50μg／ml浓度范围内,有良好的线性关系：A＝1778．9＋13686C,r＝0．999。方法同时可用于血中氯喹的分析。附一例应用报告,测得尿中氯喹的含量为0．745mg／ml,血中氯喹的含量为3．68μg／ml。尿液中同时检出氯喹的N－去单已基代谢物。定性结果经质谱法验证。     

An Assessment of the Utility of Universal and Specific Genetic Markers for Opium Poppy Identification

Abstract: The proper identification of illicit plants such as Papaver somniferum L (opium poppy) is important for law enforcement agencies. The identification of opium poppy was presently tested using 10 genetic markers that are universal for all plants or specific to a few poppy plants. The genetic distances of universal markers such as nuclear internal transcribed spacer (ITS), 18S rRNA, plastid rbcL, and trnL‐trnF intergenic spacer (IGS) of 14 species included in the Papaveraceae and Fumariaceae family were acquired by sequence comparisons. Both the ITS region and trnL‐trnF IGS showed high levels of interspecific divergence. Six Papaver genera‐specific markers were developed from coding regions involved in morphine biosynthesis. Three markers (TYDC, NCS, and BBE) produced amplicons only in opium poppy, providing a presence/absence test for opium poppy, while three additional markers (CYP80B1, SAT, and COR) were genus specific. These 10 markers might be useful for the forensic DNA analysis of opium poppy.     

内标法固相提取-GC/MS检测海洛因吸食者尿液中代谢物

本文建立了以SKF-525A作内标、GDX－301作固相提取剂、用GC／MS方法测定海洛因吸食者尿液中代谢物浓度的方法。系统考查了吗啡、单乙酰吗啡的提取回收率、线性关系等指标。所建立的方法对吗啡和单乙酰吗啡的回收率均大于80％;最小检测浓度小于0.1μg／ml;用代谢物特征离子与内标物特征离子峰高比法测定吗啡与单乙酰吗啡的浓度,在每ml尿添加0．25μg～250μg范围内,二者均有良好的提取线性关系。并对几例海洛因滥用者的尿液进行了检测。     

Computer-assisted interpretation in forensic toxicology: morphine-involved deaths

Case data from 200 morphine-involved deaths (Spiehler, V. and Brown, R., Journal of Forensic Sciences, Vol. 32, No. 4, July 1987, pp. 906-916) were analyzed for patterns and relationships using artificial intelligence (AI) computer software. Case parameters were blood unconjugated morphine, blood, brain, and liver total morphine, sex, age, frequency of use, time of death after injection, cause of death, and presence of other drugs. The programs used were Expert 4 (Biosoft-Cambridge), BEAGLE (Warm Boot, Ltd.), and KnowledgeMaker (Knowledge Garden Inc.). Interpretation was defined as estimating the dose, response, and time after drug dosing. The AI programs were used to advise on time and response outcomes for cases, to calculate the probability of the estimate being true, to develop rules for interpretation of morphine-involved cases, and to diagram a decision tree. On known cases the AI programs were successful 70 to 90% of the time in classifying the cases as to response and time. No data on dose were available in this database. The success rate in individual cases was proportional to the program-estimated probability. All three programs found the case parameters of most value in predicting response to be blood unconjugated morphine, blood total morphine, and liver total morphine. The case data most useful in estimating time of death since drug injection were blood unconjugated morphine, percent unconjugated morphine in blood, and brain total morphine. The rule induction programs found that morphine overdoses were characterized by blood unconjugated morphine greater than 0.24 micrograms/mL, liver morphine greater than 0.50 to 0.75 micrograms/g, brain morphine greater than 0.08 micrograms/g or greater than blood unconjugated morphine, and percent blood unconjugated morphine greater than 37%. Rapid deaths were characterized by percent unconjugated morphine greater than 44 to 50%; blood unconjugated morphine, as a function of other drugs present, greater than 0.09 to 0.21 micrograms/mL; and brain total morphine greater than 0.16 to 0.22 micrograms/g. This work demonstrates that inexpensive AI programs commercially available for personal computers can be useful in interpretation in forensic toxicology.     

Time of drug elimination in chronic drug abusers. Case study of 52 patients in a "low-step" detoxification ward

The elimination time of illicit drugs and their metabolites is of both clinical and forensic interest. In order to determine the elimination time for various drugs and their metabolites we recruited 52 volunteers in a protected, low-step detoxification program. Blood samples were taken from each volunteer for the first 7 days, daily, urine sample for the first 3 weeks, daily. Urine was analyzed using a fluorescence-polarization immunoassay (FPIA) and gas chromatography/mass spectrometry (GC/MS), serum using GC/MS. The elimination times of the drugs and/or their metabolites in urine and serum as well as the tolerance intervals/confidence intervals were determined. Due to the sometimes extremely high initial concentrations and low cut-off values, a few of the volunteers had markedly longer elimination times than those described in the literature. The cut-off values were as follows: barbiturates II (200ng/ml), cannabinoids (20ng/ml), cocaine metabolites (300ng/ml), opiates (200ng/ml). GC/MS detected the following maximum elimination times: total morphine in urine up to 270.3h, total morphine and free morphine in serum up to 121.3h, monoacetylmorphine in urine up to 34.5h, 11-nor-9-carboxy-delta-9-tetrahydrocannabinol (THC-COOH) in urine up to 433.5h, THC-COOH in serum up to 74.3h, total codeine in urine up to 123h, free codeine in urine up to 97.5h, total codeine in serum up to 29h, free codeine in serum up to 6.3h, total dihydrocodeine (DHC) in urine up to 314.8h, free DHC in urine up to 273.3h, total and free DHC in serum up to 50.1h. Cocaine and its metabolites were largely undetectable in the present study.