Parallel Synthesis and Screening of Supramolecular Chemosensors That Achieve Fluorescent Turn-on Detection of Drugs in Saliva.

Programming and controlling molecular recognition in aqueous solutions is increasingly common, but creating supramolecular sensors that detect analytes in biologically relevant solutions remains a nontrivial task. We report here a parallel synthesis-driven approach to create a family of self-assembling dimeric sensors that we call DimerDyes and its use for the rapid identification of salt-tolerant sensors for illicit drugs. We developed an efficient method that involves parallel synthesis and screening in crude form without the need to purify each potential sensor. Structurally diverse "hit" DimerDyes were resynthesized and purified and were each shown to assemble into homodimers in water in the programmed way. DimerDyes provided a "turn-on" fluorescence detection of multiple illicit drugs at low micromolar concentrations in water and in saliva. The combination of multiple agents into a sensor array was successfully able to detect and discriminate between closely related drugs and metabolites in multiple important drug families.

"Lab-on-Click" Detection of Illicit Drugs in Oral Fluids by MicroNIR-Chemometrics.

A novel, entirely automated MicroNIR-chemometric platform was developed for the "lab-on-click" detection of illicit drugs in nonpretreated oral fluids, and a novel tool for the first-level test is proposed. Calibration of the method was achieved by collecting oral-fluid specimens from volunteers, and chemometric analysis was considered for the development of models for prediction for cocaine, amphetamine, and Δ9-tetrahydrocannabinol. In addition, a comprehensive model was optimized for the simultaneous prediction of positive-negative samples and the specific illicit drug used by abusers in a single "click". The detection ability of the method was checked for true-positive and false-positive outcomes, and results were validated by a GC-MS reference official method. The MicroNIR-chemometric platform provided the simultaneous prediction of the three most frequently abused addictive drugs with the sensitivity and accuracy of the confirmatory analyses, offering the advantages of rapidity and simplicity and demonstrating that it is a promising tool for supporting public-health surveillance.

Cannabinoids in oral fluid by on-site immunoassay and by GC-MS using two different oral fluid collection devices.

Oral fluid (OF) enables non-invasive sample collection for on-site drug testing, but performance of on-site tests with occasional and frequent smokers' OF to identify cannabinoid intake requires further evaluation. Furthermore, as far as we are aware, no studies have evaluated differences between cannabinoid disposition among OF collection devices with authentic OF samples after controlled cannabis administration. Fourteen frequent (≥4 times per week) and 10 occasional (less than twice a week) adult cannabis smokers smoked one 6.8% ∆(9)-tetrahydrocannabinol (THC) cigarette ad libitum over 10 min. OF was collected with the StatSure Saliva Sampler, Oral-Eze, and Draeger DrugTest 5000 test cassette before and up to 30 h after cannabis smoking. Test cassettes were analyzed within 15 min and gas chromatography-mass spectrometry cannabinoid results were obtained within 24 h. Cannabinoid concentrations with the StatSure and Oral-Eze devices were compared and times of last cannabinoid detection (t(last)) and DrugTest 5000 test performance were assessed for different cannabinoid cutoffs. 11-nor-9-Carboxy-THC (THCCOOH) and cannabinol concentrations were significantly higher in Oral-Eze samples than in Stat-Sure samples. DrugTest 5000 t(last) for a positive cannabinoid test were median (range) 12 h (4-24 h) and 21 h (1- ≥ 30 h) for occasional and frequent smokers, respectively. Detection windows in screening and confirmatory tests were usually shorter for occasional than for frequent smokers, especially when including THCCOOH ≥20 ng L(-1) in confirmation criteria. No differences in t(last) were observed between collection devices, except for THC ≥2 µg L(-1). We thus report significantly different THCCOOH and cannabinol, but not THC, concentrations between OF collection devices, which may affect OF data interpretation. The DrugTest 5000 on-site device had high diagnostic sensitivity, specificity, and efficiency for cannabinoids.

A systematic approach to the analysis of illicit drugs for DNA with an overview of the problems encountered.

Due to the restricted nature of illicit drugs, it is difficult to conduct research surrounding the analysis of this drug material for any potential DNA in sufficient quantities acceptable for high numbers of replicates. Therefore, the current research available in peer reviewed journals thus far regarding analysing illicit drugs for DNA has been performed under varying experimental conditions, often using surrogate chemicals in place of illicit drugs. The data presented within this study originated from the analysis of genuine illicit drugs prepared both in controlled environments and those seized at the Australian border (and therefore from an uncontrolled environment) to determine if DNA can be obtained from this type of material. This study has been separated into three main parts (total n=114 samples): firstly, methamphetamine synthesised within a controlled environment was spiked with both saliva and trace DNA to determine the yield following DNA extraction; secondly, methamphetamine also synthesised in a controlled environment but on a larger scale was tested for the amount of DNA added incidentally throughout the synthesis, including the additional steps of recrystallising, homogenising and "cutting" the drug material to simulate preparation for distribution; and thirdly, the detection of human DNA within samples of cocaine and heroin seized at the Australian border. The DNA Fast Flow Microcon Device was utilised to concentrate all replicates from the same source into one combined extract to improve the DNA profiles for the samples where no DNA spiking occurred. Full STR profiles were successfully obtained from drug samples spiked with both saliva and trace DNA. Methamphetamine was present in the final DNA extracts and caused incompatibilities with the quantification of DNA using Qubit. The yields of DNA from drugs not spiked with DNA sources were much lower, resulting in 36 % of samples yielding alleles where all others did not. These results were not unexpected given these were realistic drug samples where the history of the drug material was unknown. This is the first study to obtain DNA profiles from genuine illicit drug material in both controlled and uncontrolled environments and indicates that the analysis of illicit drugs for DNA is an avenue worth pursuing to provide information which can in turn assist with disrupting the supply of these drugs. Given that DNA profiling is carried out worldwide using essentially the same systems as described within this study, the potential for impact is on a national and international scale.

Identification of the polymer-bounded drugs on the fabric surface: A challenge to the forensic drug analysts.

Drug trafficking through concealment has always been a method of choice for drug traffickers all around the world. This case shares a new trend in the smuggling of illicit drugs by applying a coating of drug and polymer mixture on fabric. A white fabric sample was submitted by a law enforcement agency to detect the presence of any explosive material on its surface. Later on it was also tested for illicit drugs. Stereomicroscope and Scanning Electron Microscope/Energy Dispersive X-ray Detector (SEM/EDX) were applied for microscopic examination. Acetone extract of the sample was analyzed for explosives by explosive detection kit, Gas Chromatography Mass Spectrometry (GCMS), and Fourier Transform Infrared Spectroscopy (FTIR). The routine method involving methanol as solvent was used to check heroin presence. Methanol extract of the sample was analyzed by Mecke test and GCMS. Stereomicroscope and SEM/EDX revealed the presence of some unusual coating on one side of fabric. No explosive material was detected; instead GCMS (method 1) confirmed the presence of heroin (mass fragments 268, 310, 327, and 369 m/z) and FTIR spectrum revealed presence of a polymeric material (dyneema). No drug was identified by GCMS (method 2). Method 2 was modified by replacing methanol with acetone and including an additional step of sonication for 30 min. Acetone extract showed green color with Mecke reagent and a strong signal of heroin on GCMS. This modified extraction method acted well to unbind the coated material from the fabric and to disentangle the drug from the polymer.

Determination of eight illegal drugs in human urine by combination of magnetic solid-phase extraction with capillary zone electrophoresis.

Using magnetite/silica/poly(methacrylic acid-co-ethylene glycol dimethacrylate) (Fe(3)O(4)/SiO(2)/poly(MAA-co-EDMA)) magnetic microspheres, a rapid and high-throughput magnetic solid-phase extraction coupled with capillary zone electrophoresis (MSPE-CZE) method was developed for the determination of illegal drugs (ketamine, amphetamines, opiates, and metabolites). The MSPE of target analytes could be completed within 2 min, and the eight target analytes could be baseline separated within 15 min by CZE with 30 mM phosphate buffer solution (PBS, pH 2.0) containing 15% v/v ACN as background electrolyte. Furthermore, hydrodynamic injection with field-amplified sample stacking (FASS) was employed to enhance the sensitivity of this MSPE-CZE method. Under such optimal conditions, the limits of detection for the eight target analytes ranged from 0.015 to 0.105 µg/mL. The application feasibility of MSPE-CZE in illegal drugs monitoring was demonstrated by analyzing urine samples, and the recoveries of target drugs for the spiked sample ranging from 85.4 to 110.1%. The method reproducibility was tested by evaluating the intra- and interday precisions, and relative standard deviations of <10.3 and 12.4%, respectively, were obtained. To increase throughput of the analysis, a home-made MSPE array that has potential application to the treatment of 96 samples simultaneously was used.

Liquid chromatography/tandem mass spectrometry for the qualitative and quantitative analysis of illicit drugs and medicines in preserved oral fluid.

A qualitative and quantitative analytical method was developed for the simultaneous determination of 24 illicit drugs and medicines, in preserved oral fluid samples collected with the StatSure Saliva Sampler collection device. The samples were prepared by liquid-liquid extraction followed by liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis. The chromatographic separation was performed with an Atlantis T3 (100 x 2.1 mm i.d., 3 microm) reversed-phase column using an acetonitrile/2 mM ammonium formate buffer pH 3.4 gradient and the MS/MS detection was achieved with two precursor-product ion transitions per substance. The method was fully validated, including specificity and capacity of identification, limit of detection (0.2-2.1 microg/L), limit of quantitation (0.8-6.4 microg/L), recovery (34-98%), carryover, linearity (the method was linear in the range 1-200 microg/L), intra-assay precision (coefficient of variance (CV) <20% for 20 microg/L and CV <10% for 100 microg/L) and inter-assay accuracy (mean relative error <15%) and precision (CV <20%). The method showed to be specific and sensitive. It has already been successfully used in four proficiency tests and subsequently applied to oral fluid samples collected from road traffic volunteers in the driving population of Portugal (districts of Lisbon, Coimbra and Porto), within the DRUID project.

Dual enantioselective LC-MS/MS method to analyse chiral drugs in surface water: Monitoring in Douro River estuary.

This work presents the development of an enantioselective method to quantify chiral drugs (CDs) in surface water and its application in the Douro River estuary monitoring. Different classes of CDs were targeted, including 23 compounds, namely beta-blockers, antidepressants, one beta2-adrenergic agonist, non-steroidal anti-inflammatory drugs, stimulants, and some illicit drugs as cocaine (COC) and its metabolites, and amphetamines. The analytical method was based on an innovative application of solid phase extraction (SPE), followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) using a triple quadrupole analyzer. The ground-breaking approach of SPE consists in the use of Oasis® MCX cartridges to pre-concentrate 500 mL of water samples, allowing the simultaneous extraction of acidic, basic and neutral analytes, rather than the conventional recovery of basic compounds only. Two chiral columns were used for enantiomeric separation in reverse elution mode, a Chirobiotic™V and a Pirkle type Whelk-O®1, for basic and acidic compounds, respectively. The method validation demonstrated good linearity (r2 > 0.99), selectivity and sensitivity, with method detection limits between 0.01 and 2.66 ng L-1 and method quantification limits between 0.02 and 5.71 ng L-1. The developed method was successfully applied to monitor daily variations along one week in surface waters collected in 5 locations of the Douro River estuary. Tramadol (TRM) and its metabolite N-desmethyltramadol (NDT), presented high concentrations near the affluent of a tributary river, while the second eluted enantiomer of O-desmethyltramadol (ODT) was found at high concentrations at the mouth of the Douro River. The metabolite NDT was quantified at higher concentrations than TRM. Venlafaxine (VNF) was found at high concentrations near the affluent of the same tributary river, but its metabolite, O-desmethylvenlafaxine (ODV), was found at concentrations 3 times higher. COC was found every day at all sampling points along the estuary, with slight variations.

Cannabis resin in the region of Madrid: Adulteration and contamination.

The aim of this study is to analyze the adulteration and contamination of cannabis resin obtained on the streets of Madrid, in order to establish whether it is suitable for human consumption. A total of 90 samples obtained through street vending in the Region of Madrid (CAM) were analyzed. Our results showed a direct relationship between the shape of the samples (acorn or ingot) and the presence of foreign elements, adulterants and microbiological contamination. Foreign elements were found in 64.7% of the ingot-shaped samples and in 30.2% of the acorn-shaped samples (p < 0.01); 25% of the samples were deliberately adulterated, 66.7% of which had an ingot shape. With regard to microbiological contamination, 93% of acorns were contaminated by E. coli, compared to 29.4% of ingots (p < 0.0001). In addition, all samples with fecal odor were acorns and were contaminated by E. coli. Ten per cent of the samples were contaminated by Aspergillus; of these, 66.7% had the shape of an acorn. Overall, our results showed that most (88.3%) of the hashish samples were not suitable for consumption. This percentage was significantly higher (p < 0.0001) in acorn than in ingot samples (100% vs. 58.8%). Hence, illegal street vending of hashish constitutes a public health issue.

"Krokodil" drug - Related osteonecrosis of midface: A case series.

Krokodil is a cheap and effective home-made substitute for heroin. It is widely used over the territory of the former USSR (Russia, Ukraine, Armenia and others). Krokodil drug-related midface ON often occurs as a complication of maxillary ON. Treatment of Krokodil drug-related ON of the midface is challenging. It is difficult to determine the ON zone preoperatively and intraoperatively, due to the complex anatomy of the midface and the different periods of the disease onset in different areas. The aim of this study is to show variations of the clinical course and treatment options of Krokodil drug-related ON of the midface. In this study, 3 cases of Krokodil drug-related midface ON are reported. The main clinical feature of midface ON is extraoral fistula in the midfacial zone with purulent discharge or extraoral exposure of zygomatic bone. Surgery is the main treatment method for Krokodil drug-related midface osteonecrosis. Surgery includes necrotic bone removal and defect closure. Usually an extraoral approach is used to expose necrotic bone. Intraoral maxillary sinus floor defect is closed with the use of a buccal fat pad to prevent formation of oroantral communication. Drug withdrawal, radical necrectomy, and proper closure of formed defects are the main factors that lead to successful treatment of Krokodil drug-related midface ON patients.

Interpretation of oral fluid tests for drugs of abuse.

Oral fluid testing for drugs of abuse offers significant advantages over urine as a test matrix. Collection can be performed under direct observation with reduced risk of adulteration and substitution. Drugs generally appear in oral fluid by passive diffusion from blood, but also may be deposited in the oral cavity during oral, smoked, and intranasal administration. Drug metabolites also can be detected in oral fluid. Unlike urine testing, there may be a close correspondence between drug and metabolite concentrations in oral fluid and in blood. Interpretation of oral fluid results for drugs of abuse should be an iterative process whereby one considers the test results in the context of program requirements and a broad scientific knowledge of the many factors involved in determining test outcome. This review delineates many of the chemical and metabolic processes involved in the disposition of drugs and metabolites in oral fluid that are important to the appropriate interpretation of oral fluid tests. Chemical, metabolic, kinetic, and analytic parameters are summarized for selected drugs of abuse, and general guidelines are offered for understanding the significance of oral fluid tests.

Highly precise detection, discrimination, and removal of anionic surfactants over the full pH range via cationic conjugated polymer: an efficient strategy to facilitate illicit-drug analysis.

A water-soluble cationic conjugated polyelectrolyte (CPE), poly(1,4-bis(6-(1-methylimidazolium)-hexyloxy)-benzene bromide) (PMI) displays extraordinary stability over the full pH range of 1-14 as well as in seawater, brine, urine, and other solutions and carries out efficient detection, discrimination, and removal of moderately dissimilar anionic surfactants (viz., sodium dodecyl benzenesulfonate (SDBS) and sodium dodecyl sulfate (SDS)) at very low levels (31.7 and 17.3 parts per billion (ppb), respectively). PMI formed stable hydrogels in the presence of SDS that remained unaffected by strong acids/bases, heating, ultrasonication, or exposure to light, whereas SDBS formed precipitate with PMI as a result of its different interpolymer cofacial arrangement via Columbic attraction. The complex-forming ability of PMI with SDS and SDBS facilitated their elimination from water or drug-doped urine samples without the use of any organic solvent, chromatographic technique, or solid support. This protocol, the first of its kind for the removal of anionic surfactants at very low concentrations from any type of solution and competitive environments, demonstrates an original application using a CPE. The surfactant-free sample solutions could be precisely analyzed for the presence of illicit drugs by any standard methods. Using PMI, a newly developed CPE, a rapid and practical method for the efficient detection, discrimination, and removal of SDS and SDBS at ppb levels from water and urine, under harsh conditions, and in natural chemical environments is demonstrated.

[Multislice computed tomography in the diagnosis of toxic phosphorus necrosis of the jaw].

Objective: To estimate the possibilities of using and systematizing computed tomographic findings in patients with toxic phosphorus necrosis of the jaw. Material and Methods: The investigation enrolled 87 patients diagnosed as having toxic phosphorus osteonecrosis. Radiation examination consisted of two stages: primary and repeated radiologic examinations in the postoperative period (final examination before hospital discharge). All the patients underwent skull X-ray and multislice computed tomography (MSCT). Results: Clinical and radiation examination revealed toxic phosphorus osteonecrosis of the maxilla and mandible in 29 (33%) cases. Osteonecrosis affected only the mandible in 40 (46%) cases and only the maxilla in 18 (21%) cases. In all the patients, computed tomography showed main trends in the X-ray semiotics of toxic phosphorus necrosis of the facial skeleton, such as periostitis; osteosclerosis; development a lesion having a "soap-bubble" appearance; nonspecific and inflammatory bone destruction. The bone, being destroyed, was replaced by pus; inflammatory granulations were absent; osteonecrosis occurred. These processes were characterized by the absence of an obvious demarcation zone along the edges of the process. Sequestration commonly occurred to form sinus tracts. The process involved the adjacent bones; there were reactive changes in the accessory sinuses. Conclusion: MSCT data are of highly informative value in evaluating the status of bone tissue and teeth and in detecting a concomitant abnormality in patients with osteonecrosis of the facial skeleton and may be used to plan surgical treatment for this category of patients.

Rapid detection of drugs of abuse in saliva using surface enhanced Raman spectroscopy and microfluidics.

We present a microfluidic device that detects trace concentrations of drugs of abuse in saliva within minutes using surface-enhanced Raman spectroscopy (SERS). Its operation is demonstrated using methamphetamine. The detection scheme exploits concentration gradients of chemicals, fostered by the laminar flow in the device, to control the interactions between the analyte, silver nanoparticles (Ag-NPs), and a salt. Also, since all species interact while advecting downstream, the relevant reaction coordinates occur with respect to the position in the channel. The system was designed to allow the analyte first to diffuse into the side stream containing the Ag-NPs, on which it is allowed to adsorb, before salt ions are introduced, causing the Ag-NPs to aggregate, and so creating species with strong SERS signal. The device allows partial separation via diffusion of the analyte from the complex mixture. Also, the reproducible salt-induced NP aggregation decouples the aggregation reaction (necessary for strong SERS) from the analyte concentration or charge. This method enables the creation of a region where detection of the analyte of interest via SERS is optimal, and dramatically extends the classes of molecules and quality of signals that can be measured using SERS, compared to bulk solution methods. The spatial distribution of the SERS signals was used to map the degree of nanoparticle aggregation and species diffusion in the channel, which, together with numerical simulations, was used to describe the kinetics of the colloid aggregation reaction, and to determine the optimal location in the channel for SERS interrogation.

Drugs of abuse in oral fluid collected by two different sample kits--stability testing and validation using ultra performance tandem mass spectrometry analysis.

Oral fluid (OF) is an alternative matrix for monitoring drugs of abuse in workplace, clinical toxicology, criminal justice, and driving under the influence of drugs (DUID). OF is suitable for detection of drugs that have been taken recently. It is unproblematic to observe the collection and hence avoid the possibility of the samples being tampered. OF often contains compounds in low concentrations, and small volumes are often collected. It is therefore necessary to have a sensitive, multi component method for drug detection. In this study an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) method has been developed. The samples were prepared by liquid-liquid extraction (LLE) with ethyl acetate/heptane (4:1) and the separation was achieved by an Acquity HSS T3-column (2.1 mm × 100 mm, 1.8 µm particles). Mass detection was performed by positive ion mode electrospray MS-MS. 32 drugs of abuse were determined with a cycle time of 9 min. Stability of drugs in oral fluid before analysis is an important factor that must be evaluated for each sampling device. The collection devices Intercept(®) and StatSure Saliva Sampler™ were tested using pools of real samples containing various drugs. The testing showed that 6-MAM (6-acetylmorphine), cocaine and zopiclone were the least stable compounds. In the testing for short term stability, StatSure Saliva Sampler™ showed better results. The testing of 1 year of storage at -20°C showed that most of the compounds were stable for both sampling devices, except for 6-MAM, cocaine and zopiclone. Samples of OF should be analysed as soon as possible after collection, and they should be kept frozen if immediate analysis is not possible.

Endogenous γ-hydroxybutyric acid concentrations in saliva determined by gas chromatography-mass spectrometry.

Endogenous γ-hydroxybutyric acid (GHB) concentrations in blood and urine are well documented, but there are very little data on natural levels in saliva, a biological matrix increasingly used for drug testing. We measured endogenous GHB concentrations in 120 unpaid volunteers who also provided anonymous epidemiological data. Samples were analyzed using a rapid and reliable method, utilizing liquid-liquid extraction, silyl-derivatization, and gas chromatographic-mass spectrometric analysis. One sample, between the lower limit of quantitation (0.2 mg/L) and limit of detection (0.1 mg/L), was split to 0.15 mg/L for statistical purposes. Salivary GHB concentrations ranged from 0.15 to 3.33 mg/L (mean = 1.29; median = 1.13). Statistical analysis using the Mann-Whitney test indicated that endogenous GHB concentrations in saliva were not significantly affected by age, gender, medical conditions, use of medications, and recent food/drink consumption. Interpreting GHB concentrations in biological samples poses difficulties because of its endogenous presence and rapid elimination, and this is true for saliva as well as blood and urine. However, saliva has the merit of being easy to collect by law enforcement personnel.

An evaluation of on-site oral fluid drug screening devices DrugWipe 5+ and Rapid STAT using oral fluid for confirmation analysis.

In this study, the performance of two on-site oral fluid drug-testing devices, DrugWipe 5(+) (Securetec) and Rapid STAT (Mavand), was assessed. The results obtained by the devices were compared with gas chromatography-mass spectrometry confirmation analysis results in oral fluid. Sensitivity, specificity, and accuracy of the tests, as well as positive and negative predictive values, were calculated based on the classified results of the comparison. Both of the devices were evaluated for their ability to meet toxicological cutoffs as set in the DRUID project (www.druid-project.eu) as well as those reported by the manufacturers. The evaluation was performed for relevant drug groups of both devices: amphetamines, cannabis, cocaine, and opiates. Additionally, Rapid STAT has a test for benzodiazepines included in the same device. Both tests seemed to perform quite well for amphetamines although they also gave negative results for cases with high concentrations. Also, the benzodiazepine test of Rapid STAT was at a relatively good level although only half of the positive test results were true positives using the test cutoffs. The same phenomenon was detected for the cannabis tests of both devices. The proper evaluation of cocaine and opiates tests was not applicable because of the very low number of positive cases.

In vitro capturing of various lipophilic illicit drugs by lipid dispersions. An electrokinetic capillary chromatography and fluorescence polarization study.

Fatal drug overdoses are a cause for concern all over the world. We present here a lipid-based formulation which has a strong affinity for some common illicit street drugs and can be used in vivo as a lipid 'sink'. In this study, the in vitro interactions of nine lipophilic drugs and three lipid dispersions were determined by electrokinetic capillary chromatography and fluorescence polarization. Two lipid dispersions, zwitterionic 1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine (POPC) and an anionic mixture of POPC and 1-palmitoyl-2-oleyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG) were tested and compared with a commercial lipid dispersion Intralipid(®), which has been successfully used for resuscitation of patients in cases of anesthetic overdoses. The interactions between dispersions and the drugs were quantified by means of retention factors and distribution constants, which makes the results highly comparable to those obtained from any other formulation of lipids. The results demonstrate a stronger interaction between the drugs and an artificial liposome dispersion than with the commercial Intralipid dispersion. The liposome dispersion composed of POPC and POPG functions as a lipid 'sink' for efficient entrapment of various lipophilic drugs.

Design of molecular imprinted polymers compatible with aqueous environment.

The main problem of poor water compatibility of molecularly imprinted polymers (MIPs) was addressed in examples describing design of synthetic receptors with high affinity for drugs of abuse. An extensive potentiometric titration of 10 popular functional monomers and corresponding imprinted and blank polymers was conducted in order to evaluate the subtleties of functional groups ionisation under aqueous conditions. It was found that polymers prepared using 2-trifluoromethacrylic acid (TFMAA) in combination with toluene as porogen possess superior properties which make them suitable for effective template recognition in water. The potential impact of phase separation during polymerisation on formation of high quality imprints has been discussed. Three drugs of abuse such as cocaine, deoxyephedrine and methadone were used as template models in polymer preparation for the practical validation of obtained results. The polymer testing showed that synthesized molecularly imprinted polymers have high affinity and selectivity for corresponding templates in aqueous environment, with imprinting factors of 2.6 for cocaine and 1.4 for methadone and deoxyephedrine. Corresponding blank polymers were unable to differentiate between analytes, suggesting that imprinting phenomenon was responsible for the recognition properties.

Comparison of two extraction procedures for determination of drugs of abuse in human saliva by high-performance liquid chromatography.

High performance liquid chromatography in combination with diode array detection (HPLC-DAD) was used to determine morphine, 6-acetylmorphine, cocaine, benzoylecgonine, cocaethylene, methadone and 2-ethylene-1,5-dimethyl-3,3,-diphenylpyrrolidine in human saliva. For comparison, samples were prepared by either liquid-liquid extraction in Toxitubes A or microwave-assisted extraction (MAE), by mixing 1 ml of saliva with 10 ml of chloroform and operating at 100 degrees C for 10 min. Acetonitrile and 0.02 m phosphate buffer at pH 6.5 were used as mobile phase in HPLC in gradient mode. The detector response was linear over the drug concentration range of 0.05-2.0 microg ml(-1) in human saliva. The analytical method was validated by determining its precision and accuracy (n = 5), which were lower than 5% as relative standard deviation and 6% as relative error. Limits of detection ranged from 10 to 35 ng ml(-1); mean recoveries of drugs were from 53 to 95% with Toxitubes A and from 83 to 100% with MAE at two different concentrations (0.1 and 1.0 microg ml(-1)). The proposed method was applied to 24 saliva samples from individuals poisoned with opiates and/or cocaine.