Administrative Office
St. Joseph's Hospital Site, L301-10
50 Charlton Avenue East
HAMILTON, Ontario, CANADA L8N 4A6
PHONE: (905) 521-6141
FAX: (905) 521-6142
http://www.fhs.mcmaster.ca/hrlmp/
Urine Drug Testing
Drug testing is performed for a variety of reasons. We can loosely divide the drug testing into two categories. One is performed in clinical laboratories as a part of their ‘special chemistry’ sample workup for diagnostic purposes (St. Joseph’s Healthcare toxicology laboratory), the other is ‘forensic’ drug testing which has to follow specific rules in order that the results would be defendable in the ‘courts of law’. In the clinical laboratory the primary reason for drug testing is to produce fast and reliable results for physicians who ordered the test to help them in their diagnostic work. The other major task for the biochemistry laboratory is therapeutic drug monitoring. In both cases the work has to be done fast, reliably, accurately and inexpensively. The biological detection of drug use is a two-step process. It involves a screening test which, if found to be presumptive positive, is followed by a confirmatory test. There are two primary methods of analyzing specimens for drugs: immunoassay and chromatography . Immunoassay is typically used to screen for drugs, as it is quick and comparatively inexpensive. The main limitations of immunoassay screens are low specificity and high cross-reactivity resulting in relatively high rates of false-positive test results. A confirmatory test is conducted to guard against this using a different analytical technique of equal or greater sensitivity. Chromatographic tests such as gas chromatography separation coupled with mass spectrometry (GC/MS) detection or quantitation are recommended. The confirmation test should be based on a physical or chemical principle different from the first test and should offer a higher degree of specificity for the analyte than the first test. It is imperative that the positives obtained by the immunoassay screening are confirmed. The only exemptions may be use of the point-of-care devices in the emergency departments, where the speed of the testing is important for diagnostic purposes.
There is also much information associated with drug use that cannot be determined by biological analysis. For example, conclusions regarding current intoxication, quantity of drug used, frequency of use, and physical or psychological dependency cannot be made. Drug use determination is undertaken through the analysis of a drug's metabolites as well as the drug itself depending on the sample being examined. This is important for two reasons. Firstly, metabolite(s) are most likely to be detected in some samples, primarily urine, as they often have a longer half-life than the parent compound (drug consumed). Secondly, identification of relative metabolite concentrations is often necessary to determine the drug that has been consumed. Different drugs can metabolize into the same compounds. Drug use is currently most often assessed in urine, hair, saliva and sweat. Urine is the most widely used matrix. Urine offers only an intermediate window of detection (in most cases 1-3 days) thus making test scheduling a significant issue for many applications. Its susceptibility to tampering and adulteration is also a problem.
In USA and Canada SAMHSA (Substance Abuse and Mental Health Services Administration, previously NIDA: National Institute on Drug Abuse) instituted guidelines for urine drug testing in the work place. The programme was started in late eighties in USA and is primarily aimed at detection of street drugs such as amphetamines (amphetamine, methamphetamine), cannabinoids (11-nor delta-9 THC carboxylic acid), phencyclidine (PCP), opiates (codeine, morphine, 6-monoacetyl morphine) and cocaine (benzoylecgonine). The programme has several flavours depending if the testing is done for the Department of Transport (for truck drivers and pilots), US Federal Employees or employees of nuclear energy departments. All Canadian truck drivers going to USA from Canada have to be randomly tested twice a year. The program (i) serves as a deterrent to continued use of an illicit substance, (ii) provides a means to detect and, thereby, identify employees or job applicants who are using illicit substances and (iii) assists employees in recognizing and admitting their abuse problems so that they may obtain necessary treatment.
There are currently around 80 SAMSHA certified laboratories, three of them in Canada. The laboratories are challenged 4 times a year with 30 proficiency test samples and are inspected twice a year for 2-3 days with a team of 2-4 inspectors (based on the laboratory size).
A urine sample is declared positive only after the quantitative result is equal or exceeds the cut-off concentration of a drug or metabolite in the tested urine sample. Reagents used for initial screening for several drugs of abuse classes are based on principles of immunology. Antibodies are prepared to recognize specific drugs/metabolites or drug classes. Immunoassay drug screening is performed semi-quantitatively; applying administratively defined cut-off values or concentrations for each drug/drug class screened. These immunoassay-screening techniques are fast and efficient, but may not always be specific for one drug or drug class.
SAMHSA Immunoassay Screening Cut-Off Concentrations
Cocaine (as benzoylecgonine) 150 ng/mL
Opiates (morphine equivalents) 2000 ng/mL (formerly 300 ng/mL)
Phencyclidine (PCP) 25 ng/mL
Amphetamines (D methamphetamine equivalents) 1000 ng/mL
Cannabinoids (THC-carboxylic acid) 50 ng/mL
Creatinine* (Used to monitor urine dilution) 1.768 mmol/L (20mg/dL)
* All creatinine concentrations greater than 1.768 mmol/L (20 mg/dL) are considered reliable markers of ‘normal’, undiluted urine. When a creatinine test result is less than 1.768mmol/L, it suggests that the donor may have intentionally diluted their urine. A second test (specific gravity) is measured. For urine sample to be tested the specimens must have a creatinine concentration 1.768mmol/L (>20mg/dL) and a specific gravity value >1.003 g/L
All “presumptive” positive results from the initial screening process are confirmed by GC-MS that eliminates the possibility of false positives that may be found by immunoassay screening tests. The mass spectrometer “breaks down” the drug molecules into fragments, which are unique for every drug/metabolite, analogous to a “fingerprint” being unique to one individual. The computer generated mass spectra are essentially fingerprints of different drugs/metabolites. The intensities of these mass fragments are directly proportional to the quantity of the analyzed drug in a particular specimen. Calibrating the GC-MS with varying concentrations of drug standards allows for a very sensitive, specific, precise and accurate quantitation of drugs/metabolites in urine specimens. As is the case with immunoassay screening, the confirmation cut-off concentrations are designed to separate positive and negative specimens. Urine specimens in which the drug concentration is less than the cut-off concentration are reported as negative, even though some drug may be present. The ability of these GC-MS methods to detect small amounts of drug/metabolites is much greater than the cut-off concentrations used.
SAMHSA GC-MS confirmation Cut-Off Concentrations
Amphetamines (amphetamine, methamphetamine*) 500 ng/mL
Cannabinoids (as 11-nor-D-9 THC COOH) 15 ng/mL
Cocaine (as benzoylecgonine) 150 ng/mL
Phencyclidine (PCP) 25 ng/mL
Opiates (morphine, codeine) 2000 ng/mL
6-monoacetylmorphine (heroin metabolite) 10 ng/mL
* Amphetamine (methamphetamine metabolite) must also be present at >200ng/mL
Laboratories are also mandated to extensively test for adulterants used by some individuals in hope ‘to beat the urine drug test’.
Jiri Zamecnik, PhD, PEng
Consultant, Discipline of Special Chemistry
Hamilton Regional Laboratory Medicine Program,
St. Joseph’s Healthcare