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Oral fluid could be used in monitoring of antimicrobials with D9-THC and benzodiazepines. Sensitivity for amphet-
isoniazid, ciprofloxacin, and gentamicin as well.7 amines, opiates, and cocaine (S/P ratios 1) were generally better
Commercially available screening devices allow on-site than for THC and benzodiazepines.
testing. Immunologic methods such as the enzyme-linked In the ROSITA projects, the importance of size,
immunosorbent assay are commonly used for oral fluid practicability, rapidity, and portability of the testing device
analysis. Gas chromatography–mass spectrometry (GC-MS) is was emphasized from the operational point of view. In addition
also commonly used, but many laboratories have recently to thin lines in the test display and sometimes difficult
developed liquid chromatography–mass spectrometry (LC- interpretation of the test result, practical problems had been
MS) or liquid chromatography–tandem mass spectrometry noted, eg, with using cold water for testing and/or test devices
(LC-MS-MS) methods for analysis of drugs in oral fluid. in winter. Furthermore, the education of police officers was
important. Most of the police officers who performed the tests
European Union Roadside Testing Assessment saw the OF on-site test devices as valuable tools for helping in
ÔROSITA 1 and 2Õ: Projects and European Union the identification and confirming of initial suspicion of drug
Driving Under Influence of Drugs use. The police officers with more experience of using on-site
(DRUID) Project devices were generally more satisfied with the device.
There have been two European Union–(USA) projects In a few countries, legislation already allows the use of
(ROSITA) concerning on-site testing in road traffic control OF for screening and/or confirmation in suspected driving
since 1998.6,21 The purpose of these projects was to evaluate under influence of drugs cases. Although at the end of the
on-site testing devices. In ROSITA-1, both the available urine ROSITA-2 project, no device was considered reliable enough
and OF testing devices were evaluated.21,23 The survey was to be recommended for roadside screening of drivers, on-site
conducted in collaboration with the various national police tests are used routinely in some countries by police officers. In
officers on the road. Laboratories confirmed the drug findings Australia, random roadside tests for drugs are allowed and
in OF by using the same, most often GC-MS, methods as for drivers are tested by Drugwipe (Securetec) and RapiScan
blood analysis. Development of LC-MS methods in the (Cozart Bioscience Ltd.). If both devices give a positive drug
consortium started. A clear demand for OF on-site testing result, an OF sample is sent to the laboratory for confirmation
arose from the police side. Therefore, the ROSITA-2 project analysis. The police in Finland started using Drugwipe in 2003
(2003–2005) concentrated solely on the evaluation of usability as an on-site test in cases in which police officers already
and analytical reliability of on-site OF (saliva) drug testing suspect drug use. Amphetamine is the most commonly found
devices. illicit drug among Finnish drivers. The police are satisfied with
the ability of the device to detect amphetamine users. Since
On-Site Tests then, the number of amphetamine findings has more than
The aim of the ROSITA-2 project was to evaluate the doubled among driving under the influence suspected
available on-site devices for the detection of drugs in OF at the drivers.25
roadside or in a police station. For confirmation analysis, an
additional oral fluid sample was taken with the Intercept device
(OraSure Technologies, Inc., Bethlehem, PA, USA). Also, a Oral Fluid Sample Collection, Storage, and
blood sample was taken. Nine on-site devices were evaluated: Pretreatment for Confirmatory Analysis
Drugwipe (Securetec, Brunnthal, Germany), Lifepoint Impact Oral fluid collection is a critical step in the validity of
(San Francisco, CA, USA), OraLab (Varian, Lake Forest, CA, the OF drug testing process. Oral fluid can be collected by
USA), OraLine (Sun Biomedical Laboratories, Inc., Black- adsorbent swabs, by spitting, draining, or suction. The flow
wood, NJ, USA), Oral Stat device (American Bio Medica can be stimulated mechanically (chewing gum, parafilm,
Corporation, New York, NY, USA), Oratect II (Branan Teflon, rubber band) or chemically (citric acid). Although
Medical Corporation, Irvine, CA, USA), RapiScan (Cozart stimulation allows collection in a shorter time, changing the
Bioscience Ltd., Oxfordshire, UK), Saliva Screen 5 (Ultimed pH of OF can affect the concentrations of drugs and meta-
Products GmbH, Ahrensburg, Germany), and Uplink/Drug bolites and affect the S/P ratio of drugs. Citric acid stimulation
Test (Orasure Technologies Inc., Bethlehem, PA, USA exclu- of OF, for example, produced a fivefold decrease of cocaine,
sively for Dräger Safety AG & Co, Lübeck, Germany).6 benzoylecgonine (BE), and ecgonine methyl ester (EME) in
In the ROSITA-1 project, the criteria of sensitivity and OF. It can also cause interference in some immunoassays.
specificity (greater than 90%) and accuracy (greater than 95%) Absorption of highly lipophilic substances may occur when
was proposed. None of the devices tested in ROSITA-2 met using parafilm or other stimulation devices, leading to
these criteria. However, some of the devices detected a few of decreases in the amount of measured drugs.7
the substance classes well. Sensitivity, specificity, and accu- Oral fluid specimen collectors are valuable tools for
racy for amphetamine screening with Drugwipe calculated collecting OF. Collecting a valid and representative OF is
based on OF confirmation results were 95.5%, 92.9%, and a challenge. If drugs are administered orally or smoked, OF
95.3%, respectively (128 true-positives from 148 analyzed by may be contaminated and elevated concentrations of drugs
OF), whereas the sensitivity for cannabis was low (52.5% ).22,24 may be found. These elevated concentrations affect adversely
A significant variation existed between drug classes. Theoret- S/P and S/B ratios, which make reliable interpretation of OF
ically, both cocaine and opiates should be more easily detected results difficult.26 The preservation buffer solution of devices
in OF because of generally higher concentrations compared may cause long-term problems in GC-MS analysis and poor
precision and accuracy in the form of ion suppression (see ng/mL in OF), and ethanol (0.2% in OF). Analytes were quan-
subsequently). tified from OF or OF buffer solution by GC-MS (all except for
Oral fluid samples should be stored at +4°C and be ethanol) and gas-chromatography flame ionization detector
analyzed as soon as possible. If longer storage is needed, (GC-FID) (ethanol). Collected OF volume was tested with test
samples should be stored at –20°C. Freezing lowers the persons (n = 6). Collection time of different collectors was
viscosity and after thawing, samples can be easily centrifuged. recorded simultaneously by the test persons and also user
For pretreatment, both liquid–liquid and solid-phase extraction comments were requested. The summary of the results are
is used. presented in Table 1.
The variability of the recoveries of different substances The ordinary plastic tube did well in the experiments,
from the different collection devices was not well known at but it was considered somewhat messy to collect OF by
the time of the ROSITA-2 project. All the ROSITA partners spitting. Other collection devices with no buffer were not good
agreed to use the Intercept collector, although the drug enough in terms of recovery or stability. The Greiner was
stability in the buffer and absorption of drugs in the cotton roll considered to be too complicated to use at the roadside. The
of the specimen collector was not known. The average amount Statsure collector gave the best results for recovery (greater
collected with the device was low, only 224 mL (minimum, 0; than 80% for all substances) and good results for stability.
maximum, 795 mL) in roadside conditions. In laboratory It was quick to use (less than 2-minute collection time) and
conditions, however, much higher volumes were collected received good user feedback. The buffer solutions seem to
(500 to 700 mL).24 There was a wide variability in the volumes contain macromolecules that coextract with the analytes in the
of OF collected. Collection of OF was much more difficult, extraction procedure and cause long-term problems in the GC-
for example, from amphetamine abusers, whose oral fluid MS systems.1 Therefore, the buffer solutions from the devices,
secretion is lowered and who experienced dry mouth.5 including Statsure, contaminated the GC liner significantly.
Oral fluid was diluted with buffer in the Intercept device, The only exception was the Greiner that provided a clean
which complicated the quantitative determination, especially background in the chromatograms. None of the collection
because the amount of buffer solution was not constant. devices was thus perfect, but the Statsure was chosen as the OF
Furthermore, some nonvolatile components in the Intercept collection device of the DRUID project.27 Buffer macro-
buffer coextracted with the analytes, increasing the back- molecules seemed to coextract with target drugs and caused
ground interference in GC-MS analysis and decreasing the interfering background in chromatograms both after solid-
column life.1 phase extraction as well as after liquid–liquid extraction.1,27
A study27 was done find out the best choice of OF
collection device for the European Union project, DRUID. The Screening Tests Used by Laboratories
criteria were that the device should be suitable for roadside The analytical approaches for OF testing of laboratories
collection (fast, easy to use), provide enough sample for in ROSITA project were immunologic, GC-MS, and LC-MS
toxicologic analysis, be able to measure sample volume, and (-MS) methods. Some laboratories used immunologic screen-
enable good recovery and stability of drugs and medicinal ing like the enzyme-linked immunosorbent assay before
drugs. The tested analytes were amphetamine, MDMA, THC, confirmation, but most laboratories did screening and con-
cocaine, morphine, codeine, diazepam, alprazolam (all 1000 firmation simultaneously by chromatographic methods. Table 2
TABLE 4. Confirmatory Tests’ Cutoff Values for Drugs in Undiluted Oral Fluid Proposed by European Union
(EU) DRUID Project and Substance Abuse Mental Health Services Administration (SAMSHA)47
Whole Blood EU/DRUID Oral Fluid EU/DRUID Oral Fluid Cutoffs
Substance Cutoff (ng/mL) Cutoff (ng/mL) (ng/mL) Proposed by SAMSHA
Ethanol 0.1 g/L 0.1 g/L
Morphine 10 20 40
Amphetamine 20 25 50
MDMA 20 25 50
MDA 20 25 50
Cocaine 10 10 8
THC 1 1 2
THCCOOH 5
Diazepam 20 5
Alprazolam 10 1
Clonazepam 10 1
Benzoylecgonine 50 10
Codeine 10 20 40
6-acetylmorphine 10 5 4
Methamphetamine 20 25 50
Methadone 10 20
Oxazepam 50 5
Nordiazepam 20 1
Zopiclone 10 10
MDEA 20 25
Lorazepam 10 1
Flunitrazepam 2 1
Zolpidem 20 10
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