Elimination of Cocaine and Metabolites in Saliva Following Repeated Oral Cocaine Administration
Rebecca A. Jufer*1, Sharon L. Walsh2, and Edward J. Cone1
1Addiction Research Centre, IRP, NIDA, Baltimore, MD, USA
2Department of Psychiatry, Johns Hopkins University School of Medicine, Baltimore, MD, USA
Saliva is an alternative matrix that is useful for detecting recent drug use. Saliva collection is noninvasive and concentrations show a strong correlation with plasma drug concentrations under some circumstances. The excretion profile of cocaine in saliva following acute cocaine administration has been examined; however, few studies have examined the elimination of cocaine in saliva following chronic cocaine use. As part of a controlled clinical study investigating the effects of repeated oral cocaine administration, we collected saliva specimens for cocaine analysis. In this study, healthy male and female volunteers with a history of cocaine abuse were administered multiple doses of oral cocaine in up to 16 daily sessions. In each session, volunteers received 5 equal doses of oral cocaine with 1 hour between doses. Across sessions, cocaine was administered in ascending doses with an initial dose of 100 mg (500 mg/day) up to 400 mg (2 g/day), increasing by 25 mg/dose/session (125 mg/session). Cocaine capsules were double encapsulated and polished to prevent contamination of the oral cavity during drug administration. Saliva specimens were collected during and up to 120 hours after the final dosing session. Specimens were collected by expectoration into a tube while stimulating saliva flow with sour candy. Specimens collected during and after the final dosing session were analyzed for cocaine and metabolites by solid phase extraction followed by gas chromatography-mass spectrometry in the SIM mode (1). The limit of detection of the assay was approximately 1 ng/ml for all analytes. Cocaine (COC) and ecgonine methyl ester (EME) were the primary analytes detected. Other analytes identified included benzoylecgonine (BZE), norcocaine (NCOC), and p-hydroxycocaine (p-HOCOC). Detection times for cocaine ranged from 48-120 hours. Cocaine elimination half-lives were calculated from saliva concentration vs. time plots constructed on a semi-log scale where kel= -2.303*slope and t1/2= 0.693/ kel. Peak cocaine and metabolite concentrations from the final dose and cocaine elimination half-life for each subject are tabulated below:
| |
|
|
Peak Saliva
Concentration (ng/ml) |
| Subject ID |
Final Dose |
Cocaine |
BZE |
EME |
NCOC |
p-HOCOC |
Cocaine
Half-Life (hr) |
| A |
2000 mg
(5x400 mg) |
7590 |
2717 |
10022 |
809 |
292 |
4.4 |
| B |
1100 mg
(4x275 mg) |
10922 |
5046 |
22197 |
945 |
362 |
2.2 |
| D |
2000 mg
(5x400 mg) |
13822 |
3412 |
14352 |
1784 |
540 |
4.3 |
| E |
1500 mg
(5x300 mg) |
8844 |
3727 |
14741 |
1737 |
609 |
2.2 |
| G |
375 mg
(1x375 mg) |
1978 |
926 |
1313 |
139 |
55 |
2.6 |
Considerably higher saliva concentrations of EME and NCOC were produced by chronic oral dosing compared to other routes of cocaine administration. In addition, cocaine elimination half-lives following repeated oral administration were slightly longer than those reported from previous studies. The detection of substantial amounts NCOC and p-HOCOC in saliva has not been previously reported. Both of these compounds are pharmacologically active and NCOC has been shown to be a precursor to hepatotoxic metabolites in animals. In summary, these data suggest that the elimination profile of cocaine in saliva following repeated oral administration is altered from that observed following acute parenteral cocaine administration.
1. E.J. Cone, M. Hillsgrove, and W.D. Darwin. Simultaneous measurement of cocaine, cocaethylene, their metabolites, and "Crack" pyrolysis products by gas chromatography-mass spectrometry. Clin. Chem., 40, 1299-1305 (1994).
Key words: Oral Cocaine, Chronic Cocaine, Saliva.
The Application of Opiates Determination in Saliva for Diagnostic and Prognostic Purposes during Detoxification of Addicted Persons
Ewa Janowska1, Wojciech Piekoszewski*1, Janusz Pach2, Lidia Winnik2
1Institute of Forensic Research, Cracow, Poland
2 Department of Clinical Toxicology, Collegium Medicum, Cracow, Poland
The abuse of "kompot" - home-made heroin - is very common in Poland. During detoxification and monitoring of abstinence, frequent blood sampling has to be done which can cause some problems. Saliva can be an alternative body fluid for determination of morphine and other chemical substances co-existing in "kompot".
The aim of the study was to compare serum and saliva concentrations of morphine and codeine in patients admitted to the Detoxification Unit and during their stay at the Toxicology Clinic.
The first step was to develop and validate the GC/MS methods of morphine and codeine determination. Sample preparation consisted of enzymatic hydrolysis (b-hydroxylase and arylsulphatase), deproteinisation (trichloroacetic acid), solid phase extraction (Bond Elut Certify) and derivatisation (bis(trimethylsilil) trifluoracetamide). D3-morphine and D3-codeine were used as an intrinsic standard. LOD for morphine and codeine was 4 and 5 ng/ml; LOQ was 14 and 15 ng/ml respectively. The range of quantification was 30-850 ng/ml and the recovery range 50-60%.
The study was carried out on 37 patients (29 male and 8 female) admitted to the Toxicology Clinic. In all patients the presence of opiates in urine was preliminarily confirmed by enzymoimmunoassay. The concentration of morphine (measured by GC/MS) in serum ranged from 0 to 671 ng/ml (mean 179 ng/ml) and codeine from 0 to 553 ng/ml (mean 59 ng/ml), in saliva 0-142 ng/ml (mean 44 ng/ml) and 0-428 ng/ml (mean 63.4 ng/ml) respectively. In the studied group a correlation between the concentration of morphine in serum and saliva was shown (r = 0.4854), but there was no correlation for the concentration of codeine in these body fluids. The results of this study indicate that the measuring of morphine in saliva could be used as a means of monitoring the taking of home-made heroin; however, the fact that the concentration of morphine in saliva is three times lower than in serum necessitates the use of more sensitive analytical methods.
Keywords: Alternative Materials, Saliva, Morphine, GC/MS.
Testing for the Anabolic Steroid DHEA in Human Hair
Pascal Kintz*1, Vincent Cirimele1, Marc Deveaux2 and Bertrand Ludes1
1 Institut de Medecine Legale, 11 rue Humann, F-67000 Strasbourg, France
2 Institut de Medecine Legale, Lille, France
In 1974, steroids were added to the list of doping agents banned by the International Olympic Committee because of their effects on the performance of the athletes.
Dehydroepiandrosterone (DHEA) is a steroid hormone naturally produced by the adrenal glands and by the ovaries. DHEA can be converted into other hormones, including estrogen and testosterone. In the United States, DHEA is classified as a nutritional supplement. This is not the case in France, where the drug is listed as a doping agent. As athletes can abuse DHEA to benefit from its conversion to testosterone, there is a need to establish the physiological range of DHEA concentrations in human hair.
DHEA was investigated in hair obtained from 27 control subjects, including 15 males and 12 female, aged 17-42 years. After decontamination of hair samples with dichloromethane, 100 mg of hair was incubated in 1 M NaOH in the presence of 1 ng of testosterone-d3. After neutralisation, the extract was purified using solid phase extraction with Isolute C18 columns, and subsequent liquid-liquid extraction with pentane. After silylation, DHEA was analyzed by GC/MS. Results were linear in the range 1 to 20 pg/mg. Relative extraction recovery was 91.6% with a limit of detection of 0.5 pg/mg. Concentrations measured were in the range 1.2 to 6.7 pg/mg, with a mean value of 4.3 pg/mg (males), and 0.5 to 10.6 pg/mg, with a mean value of 5.3 pg/mg (females). Extensive chromatographic procedures (2 purification steps by solid phase and liquid-liquid extraction, combined with injection of 4 µl through the column in pulsed mode) were analytical prerequisites for successful identification of DHEA in hair due to the low target concentrations.
After daily oral administration of 25 mg of DHEA for 1 month to 3 subjects (males), segmental hair analyses were: (12.6-76.3 pg/mg), (7.5-13.0 pg/mg) and (32.7-48.9 pg/mg).
This new technology may find useful applications in anabolic abuse control.
Comparative Efficiency of Hair and Urine Analysis when Assessing Drug Consumption
Carmen Jurado*, Teresa Soriano, Manuel Menendez and Manuel Repetto
1Instituto Nacional de Toxicologia, P.O.Box 863, 41080-Sevilla, Spain
The correct diagnosis of drug abuse is very important in dealing with drug addicts. The toxicological analysis for drugs of abuse can be performed in all biological fluids or tissues. Traditionally urine has been the sample of choice for analysis. More recently, however, hair analysis has been introduced as an alternative or complementary method of testing for drug consumption.
The aim of this presentation is to compare the effectiveness of hair analysis and urinalysis in identifying drug-using individuals.
This comparison was performed with 100 subjects. Urine and hair were collected from all of them and analyzed for opiates, cocaine and cannabis.
Urine and hair were in agreement on 29 positives and 51 negatives for opiates, 35 positives and 35 negatives for cocaine and 27 positives and 46 negatives for cannabis. Nevertheless, hair analysis identified 19, 28 and 21 heroin, cocaine and cannabis abusers, respectively, who were undetected by urinalysis; while urinalysis detected 1, 2 and 6 heroin, cocaine and cannabis consumers undetected by hair analysis.
There was a concordance between hair and urine analysis in the case of heavy and intermediate consumers, while the majority of the cases where urinalysis failed to confirm positive hair analysis were from light abusers.
The study demonstrates that both samples are useful in establishing drug addiction. However, hair analysis is better to assess chronic drug consumption and thus gives a more real picture of drug use, in chronological terms, than that provided by standard urinalysis. Not only by confirming the positive results, but also by avoiding the negative analytical results due to temporary abstention.
Keywords: Hair Analysis, Urinalysis, Comparison.
The Development of a Liquid Chromatography - Mass Spectrometry Method for the Detection of Allopurinol and its Metabolites in Human Hair
Tamsin Kelly*1, Costa Conn1, Ken Williams2, Harry Rose1, Jim Keegan1
1 Department of Chemistry, Materials and Forensic Sciences, University of Technology, Sydney, P0 Box 123, Broadway, N.S.W. Australia 2007
2 Clinical Pharmacology, St Vincent's Hospital, Darlinghurst, N.S.W. Australia 2010
The development of a liquid chromatography - mass spectrometry (LC-MS) method for the detection of allopurinol, and it's metabolite, oxypurinol in human hair is reported. The method combined a SymmetryShield RP8 column with atmospheric pressure chemical ionisation (APCI) and selective ion monitoring (SIM) of the molecular ions of allopurinol and oxypurinol. The experimental results indicate that allopurinol is not incorporated into hair, while oxypurinol is. Furthermore, the results indicate that the developed LC-MS method may provide a means by which clinical monitoring of oxypurinol can be achieved; an application of potential importance due to the possible adverse effects associated with the accumulation of oxypurinol in the body. The LC-MS method developed is the first method to detect these analytes in hair, and among the first methods to utilise the technique of LC-MS for the analysis of drugs in hair.
Keywords: Allopurinol, Hair, LC-MS.
