ANALYTICAL EVALUATION OF FIVE ORAL FLUID DRUG TESTING DEVICES

INTRODUCTION

The correlation between drug presence in blood and oral fluid and the ease of sample collection make oral fluid an ideal matrix for roadside drug tests aimed at detecting “driving under the influence” [1, 2].

The possibility of using the result of oral fluid tests to detect impaired drivers was introduced in the Belgian Traffic Legislation in July 2009 [3].

A reliable rapid on-site oral fluid testing device, that can be used routinely in police controls, is the basis for implementing the law.

CONCLUSIONS

In July 2009 the possibility of using oral fluid drug testing devices to target impaired drivers was introduced in the Belgian legislation.

The aim of this study was to evaluate the reliability of 5 oral fluid testing devices to determine whether any of them may be suitable to detect drug-impaired drivers during roadside police controls. The evaluation was carried out taking the Belgian law confirmation cut-offs for cocaine, opiates, cannabinoids and amphetamine in consideration.

All tested devices showed good specificity for all drug classes. Sensitivity and accuracy were very different among devices and drug classes. In particular, when applying Belgian low cut-offs, all devices showed unsatisfactory sensitivity for cocaine; the highest being equal to 50% for both Dräger DrugTest ® 5000 and Mavand RapidSTAT devices.

Considering that cannabis, followed by amphetamines and ecstasy, is the most prevalent drug among impaired drivers in Belgium [5], only Dräger DrugTest ® 5000 appeared to be sensitive enough to be used during roadside police controls.

Cristina Isalberti, Sylvie Van Stechelman, Sara-Ann Legrand, Trudy Van der Linden, Alain Verstraete

Department of Clinical Chemistry, Microbiology and Immunology, Ghent University, De Pintelaan 185, 9000 Ghent, Belgium

UPLC-MS/MS CONFIRMATION ANALYSIS

The oral fluid samples for confirmation analysis were collected using StatSureTM Saliva Sampler TM device. This consists of an absorptive pad with a volume indicator that turns blue when 1 mL of oral fluid has been collected, and a plastic tube containing 1 mL of buffer solution. To account for any possible variability in the volume of oral fluid collected, drug concentrations obtained after UPLC-MS/MS analysis were corrected on the basis of the average weight of an empty StatSureTM device and the weight of the StatSureTM device after sample collection.

For the liquid-liquid extraction 20 L of 200 ng/mL isotope-labelled internal standards solution and 200 L of NH4HCO3 (0.2 M, pH 9.3) were added to 400 L of sample from the StatSureTM Saliva Sampler. Samples were then extracted with 1.25 mL of heptane/ethyl acetate (1:4). After 15 minutes on a shaker and centrifugation, the organic phase was transferred into a clean test tube and evaporated to dryness at room temperature. Samples were reconstituted in 100 L methanol/water (50:50) and transferred into a vial for UPLC-MS/MS analysis. Calibration curves (range 0.5-200 ng/mL) and quality controls, made by spiking target compounds in drug free oral fluid samples collected using StatSure TM Saliva Sampler TM device, were extracted alongside each batch of samples.

UPLC-MS/MS analysis were performed on an AcquityTM ultra performance liquid chromatograph (Waters) equipped with an Acquity UPLC BEH C18 column (1.7µm, 2.1 x 100mm) and a Vanguard BEH C18 pre-column (1.7µm, 2.1 x 5mm). Detection was performed using a Waters Quattro Premier XE tandem mass spectrometer operating in MRM mode. UPLC and MS conditions are reported in the tables below.

AIM

The purpose of this study was to evaluate the performance of five oral fluid testing devices (Varian OraLab ® 6, Dräger DrugTest® 5000, Cozart® DDS 806, Mavand RapidSTAT®, Innovacon OrAlert) that may be used in roadside drug tests and assess their suitability for detecting cocaine, opiates, cannabinoids and amphetamine at the cut-offs set by the Belgian legislation.

Short descriptions of the tested devices are reported in the box to the right. Detailed descriptions, including classes of substances detected, cut-offs and operating procedures, can be found in Deliverable 3.2.2 on the DRUID project official website [4].

METHODS

More than 760 oral fluid samples were collected from volunteers either at drug addiction treatment centres or during roadside sessions.

At the time of collection volunteers provided two oral fluid samples. One was tested on-site with one of the selected devices, while the second sample was used for confirmation analysis by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS/MS) after liquid-liquid extraction (conditions described below).

Sensitivity, specificity, accuracy and prevalence for cocaine, opiates, cannabinoids and amphetamine were calculated for each device applying Belgian legal confirmation cut-offs (THC 10 mg/mL; amphetamine 25 ng/mL; MDMA 25 ng/mL; free morphine or 6-AM 5 ng/mL; cocaine or benzoylecgonine 10 ng/mL).

RESULTS

Specificity, sensitivity, accuracy and drug prevalence for each device are shown in the table below.

REFERENCES

[1] Aps JK, Martens LV (2005) Forensic Sci Int 150(2-3): 119-131 - [2] Verstraete AG (2005) Forensic Sci Int 150: 143-150 - [3] Belgisch Staatsblad/Moniteur Belge (15.09.2009 – Ed. 2): 62185-62189 - [4] www.druid-project.eu - [5] Raes E, Verstraete AG (2005) J Anal Toxicology 29: 632-636

DISCLAIMER - This abstract has been produced under the project “Driving Under the Influence of Drugs, Alcohol and Medicines” (DRUID) financed by the European Community within the framework of the EU 6 th Framework Program. This abstract reflects only the authors’ view. The European Community is not liable for any use that may be made of the information contained therein.

Target Q1 Q2 RT

(min) Cone (V)

Collision energy

(eV) Target Q1 Q2

RT (min)

Cone (V)

Collision energy

(eV) 6-acetylmorphine 328.1 152.1 3.61 47 61 MDA-D5 185.0 110.0 3.23 17 21 6-acetylmorphine-D3 331.1 164.9 3.58 45 37 MDEA 208.1 163.0 3.79 23 13 Amphetamine 136.1 119.1 3.43 15 9 MDEA-D5 213.1 162.9 3.69 21 13 Amphetamine-D5 141.0 92.9 3.37 17 27 MDMA 194.1 163.0 3.43 21 13 Benzoylecgonine 290.1 168.0 2.64 33 19 MDMA-D5 199.1 135.2 3.34 21 21 Benzoylecgonine-D3 293.1 171.0 2.65 33 19 Metamphetamine 150.0 91.0 3.61 21 17 Cocaine 304.1 182.1 4.66 31 19 Methamphetamine-D5 155.0 120.9 3.55 19 11 Cocaine-D3 307.1 185.0 4.64 31 19 Morphine 286.1 152.1 3.13 45 53 Codeine 300.1 165.0 3.85 41 43 Morphine-D3 289.1 164.9 3.10 43 37 Codeine-D3 303.1 215.0 3.83 45 25 THC 315.2 193.1 5.44 31 21 MDA 180.0 105.0 3.28 15 21 THC-D3 318.1 196.0 5.43 31 25

UPLC gradient elution Time (min)

Flow (mL/min)

%A %B

0.00 0.40 90.0 10.0 1.00 0.40 90.0 10.0 1.50 0.40 60.0 40.0 2.99 0.40 33.8 66.2 3.00 0.25 33.8 66.2 3.80 0.25 25.0 75.0 4.30 0.40 5.0 95.0 5.30 0.40 5.0 95.0 5.50 0.40 90.0 10.0 7.00 0.40 90.0 10.0

A = NH4HCO3 (2mM, pH 9.3); B = MeOH Column temperature = 60°C

Volume injected = 25 L

General parameters for mass spectrometry ES + Source Capillary Voltage 0.8 kV

Extractor Voltage 4 V RF Lens 0.0 V Source Temperature 140°C Desolvation Temperature 450°C Desolvation Gas Flow 1,000 L/h Cone Gas Flow 50 L/h

Analyser Collision Gas Flow 0,15 mL/min

Varian OraLab ® 6 - Consists of a collection stick and an expresser vial. Testing starts with oral fluid collection and results can be read in 10 - 15 minutes.

TESTED DEVICES

Dräger DrugTest ® 5000 - Comprises the Dräger DrugTest 5000 Analyzer and a test kit. The test kit consists of a test cassette with an oral fluid collector. Once the test cassette and the cartridge are placed into the analyzer results are shown within a few minutes.

Cozart ® DDS 806 - Comprises a collector swab, a buffer bottle, a disposable test cartridge and a handheld instrument for result interpretation. Two drugs can be tested in 90 seconds, and 5/6 drug classes in 5 minutes.

Mavand RapidSTAT ® - Consists of a collector stick, a buffer bottle and the test panel. The test results can be read either straight from the test device or by inserting the device into a mobile reader. The total time needed for testing is 7-12 minutes.

Innovacon OrAlert - Consists of a collector stick and of a test device. An oral fluid sample is collected during the testing and can be sent to laboratory for confirmation. The total test time is approximately 10 minutes.

DEVICE TARGET substance

True pos

False pos

True neg

False neg

N. of tests

Sens. (%)

Spec. (%)

Acc. (%)

Prev. (%)

Varian OraLab ®

6

Cocaine 19 0 195 35 249 35.2 100 85.9 21.7 Opiates 85 2 120 42 249 66.9 98.4 82.3 51.0

THC 18 2 159 70 249 20.5 98.8 71.1 35.3 Amph 19 0 216 14 249 57.6 100 94.4 13.3

Dräger DrugTest ®

5000

Cocaine 6 1 124 6 137 50.0 99.2 94.9 8.8 Opiates 75 3 45 14 137 84.3 93.8 87.6 65.0

THC 20 5 107 5 137 80.0 95.5 92.7 18.2 Amph 6 0 129 2 137 75.0 100 98.5 5.8

Cozart ® DDS 806

Cocaine 1 1 129 7 138 12.5 99.2 94.2 5.8 Opiates 49 0 66 23 138 68.1 100 83.3 52.2

THC 11 0 99 28 138 28.2 100 79.7 28.3 Amph 4 1 131 2 138 66.7 99.2 97.8 4.3

Mavand RapidSTAT

®

Cocaine 3 3 120 7 133 30.0 97.6 92.5 7.5 Opiates 62 2 52 17 133 78.5 96.3 85.7 59.4

THC 13 12 91 17 133 43.3 88.3 78.2 22.6 Amph 1 4 123 5 133 16.7 96.9 93.2 4.5

Innovacon OrAlert

Cocaine 7 0 96 7 110 50.0 100 93.6 12.7 Opiates 64 2 20 24 110 72.7 90.9 76.4 80.0

THC 3 0 97 10 110 23.1 100 90.9 11.8 Amph 1 10 97 2 110 - 90.7 89.1 2.7