Background: Oral fluid (OF) is an exciting alternative matrix for monitoring drugs of abuse in workplace, clinical toxicology, criminal justice, and driving under the influence of drugs (DUID) programs. During the last 5 years, scientific and technological advances in OF collection, point-of-collection testing devices, and screening and confirmation methods were achieved. Guidelines were proposed for workplace OF testing by the Substance Abuse and Mental Health Services Administration, DUID testing by the European Union’s Driving under the Influence of Drugs, Alcohol and Medicines (DRUID) program, and standardization of DUID research. Although OF testing is now commonplace in many monitoring programs, the greatest current limitation is the scarcity of controlled drug administration studies available to guide interpretation.

Content: This review outlines OF testing advantages and limitations, and the progress in OF that has occurred during the last 5 years in collection, screening, confirmation, and interpretation of cannabinoids, opioids, amphetamines, cocaine, and benzodiazepines. We examine controlled drug administration studies, immunoassay and chromatographic methods, collection devices, point-of-collection testing device performance, and recent applications of OF testing.

Summary: Substance Abuse and Mental Health Services Administration approval of OF testing was delayed because questions about drug OF disposition were not yet resolved, and collection device performance and testing assays required improvement. Here, we document the many advances achieved in the use of OF. Additional research is needed to identify new biomarkers, determine drug detection windows, characterize OF adulteration techniques, and evaluate analyte stability. Nevertheless, there is no doubt that OF offers multiple advantages as an alternative matrix for drug monitoring and has an important role in DUID, treatment, workplace, and criminal justice programs.

Oral Fluid Testing for Drugs of Abuse

Over the last decade there have been considerable developments in the use of oral fluid (saliva) for drug testing. Oral fluid can provide a quick and non-invasive specimen for drug testing. However, its collection may be thwarted by lack of available fluid due to a range of physiological factors, including drug use itself. Food and techniques designed to stimulate production of oral fluid can also affect the concentration of drugs. Current applications are mainly focused on drugs of abuse testing in employees at workplaces where drug use has safety implications, in drivers of vehicles at the roadside and in other situations where drug impairment is suspected. Testing has included alcohol (ethanol) and a range of clinical tests eg antibodies to HIV, therapeutic drugs and steroids. Its main application has been for testing for drugs of abuse such as the amphetamines, cocaine and metabolites, opioids such as morphine, methadone and heroin, and for cannabis. Oral fluid concentrations of basic drugs such as the amphetamines, cocaine and some opioids are similar or higher than those in plasma. Tetrahydrocannabinol (THC), the major species present from cannabis use, displays similar concentrations in oral fluid compared to blood in the elimination phase. However, there is significant local absorption of the drug in the oral cavity which increases the concentrations for a period after use of drug. Depot effects occur for other drugs introduced into the body that allow local absorption, such as smoking of tobacco (nicotine), cocaine, amphetamines, or use of sub-lingual buprenorphine. Screening techniques are usually an adaptation of those used in other specimens, with an emphasis on the parent drug since this is usually the dominant species present in oral fluid. Confirmatory techniques are largely based on mass spectrometry (MS) with an emphasis on Liquid Chromatography-Mass Spectrometry (LC-MS), due to low sample volumes and the low detection limits required. Drug testing outside laboratory environments has become widespread and provides presumptive results within minutes of collection of specimens. This review focuses on the developments, particularly over the last 10 years, and outlines the roles and applications of testing for drugs in oral fluid, describes the difficulties associated with this form of testing and illustrates applications of oral fluid testing for specific drugs.

Drug Testing in Oral Fluid

Nine different oral fluid (OF) collection devices were studied to evaluate their suitability for collecting samples for drug analysis. The devices were Greiner Bio-One, Orasure Intercept, Immunalysis Quantisal, StatSure Saliva.Sampler, Cozart, Sarstedt Salivette, Malvern Medical OraCol, Acro Biotech Salicule, and Varian OraTube. For comparison, OF was also collected into plastic tubes. The volume of collected OF was quantified for samples collected both in vitro and from volunteers. Drug recovery was studied by collecting OF fortified at 1000 ng/mL with amphetamine, 3,4-methylenedioxymethamphetamine, cocaine, Delta(9)-tetrahydrocannabinol, morphine, codeine, diazepam, and alprazolam with the devices in vitro and analyzing the samples with gas chromatography-mass spectrometry. Recovery of ethanol was measured from 0.2% in OF by headspace gas chromatography-flame-ionization detection. The stability of drugs in the samples was studied by analyzing the samples after 0, 14, and 28 days storage. The study shows that there are substantial differences between the OF collection devices on the market. Some are well suited for collecting samples for toxicological analysis, but some give quite poor results.

/pdf/drug-testing-in-oral-fluid-evaluation-of-sample-collection-437aztmie4.pdf

Drug testing in oral fluid-evaluation of sample collection devices

Several minor tobacco alkaloids have been found to exhibit properties pharmacologically relevant to the addictive profile of tobacco; however, little is known of their effects on a behavioural model of drug addiction. In this study we compared the locomotor and reinforcing effects of intravenous nicotine (30 microg/kg per infusion) vs. a cocktail of nicotine plus five minor alkaloids found in tobacco smoke (anabasine, nornicotine, anatabine, cotinine and myosmine). Rats were initially tested for their locomotor response to nicotine or nicotine plus the minor alkaloids with six intravenous injections over 1 h. We then assessed the spontaneous acquisition of intravenous self-administration with nicotine or nicotine plus the minor alkaloids, under a fixed-ratio 1 schedule followed by responding on a fixed-ratio 5 schedule, progressive-ratio schedule and a single within-session ascending dose-response test. The activity test was repeated following the progressive-ratio phase to assess locomotor sensitization. A second group of rats were then tested on the locomotor procedure to better clarify the role of each individual minor alkaloid in nicotine-induced locomotor activity. Compared to nicotine alone, addition of the minor tobacco alkaloids increased locomotor activity and increased locomotor sensitization following self-administration. During fixed-ratio 5, progressive ratio and the dose-response test, rats receiving nicotine plus the minor alkaloids responded significantly more than those receiving nicotine alone. Testing of each minor alkaloid in the second experiment indicated that anatabine, cotinine and myosmine individually increased nicotine-induced locomotor activity. These results suggest that the minor tobacco alkaloids, particularly anatabine, cotinine and myosmine, may increase the motivation for nicotine and thus facilitate smoking behaviour.

The addition of five minor tobacco alkaloids increases nicotine-induced hyperactivity, sensitization and intravenous self-administration in rats.

A simple and reliable procedure for the determination of psychoactive drugs in oral fluid by gas chromatography-mass spectrometry.

The recovery of illicit drugs from oral fluid sampling devices

The effect of total particulate matter (TPM) from cigarette smoke on the expression and binding properties of nicotinic acetylcholine receptors (nAChRs) was investigated using a human neuroblastoma cell line (SH-SY5Y). TPM but not nicotine on its own inhibited cell growth at nicotine concentrations above 5 卮 To examine effects on nAChR expression, intact cells were incubated with 3H-epibatidine, and a Bmax of 13 fmoles/105 cells (7.8 x 104 binding sites/cell) was measured in unexposed cells as well as in cells treated with 2占nicotine alone or with TPM containing 2占nicotine. Using Scatchard analysis, we measured a Kd of 0.3nM for 3H-epibatidine binding to nAChRs. This Kd was increased to 1.3 nM by addition of nicotine or TPM extract, both at 2占nicotine. Bmax, however, was unaffected, suggesting competitive binding of nicotine to its receptor. Short-term and prolonged 3-day exposures of SH-SY5Y cells to either TPM or nicotine at nicotine concentrations ranging from 0.2 占to 20 占increased specific binding, suggesting upregulation of nAChR expression. Most significant, binding was consistently greater in cells pretreated with TPM than in cells pretreated with nicotine. We conclude that TPM contains compounds that are toxic to cells at high concentrations (cell growth inhibition) but that do not compete with nicotine for binding to nAChRs (Scatchard analysis). These non-nicotinic compounds are capable of increasing the expression of one or more of the nAChR subunits. Furthermore, our cell culture assay provides a useful in vitro model for assessing the relative addictiveness of different tobacco products, including that of non-nicotine components.

Tobacco particulate matter is more potent than nicotine at upregulating nicotinic receptors on SH-SY5Y cells.

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The recovery of illicit drugs from oral fluid sampling devices

Tobacco particulate matter is more potent than nicotine at upregulating nicotinic receptors on SH-SY5Y cells.