David A. Kidwell

Testing for drugs of abuse in saliva and sweat

The detection of marijuana, cocaine, opiates, amphetamines, benzodiazepines, barbiturates, PCP, alcohol and nicotine in saliva and sweat is reviewed, with emphasis on forensic applications. The short window of detection and lower levels of drugs present compared to levels found in urine limits the applications of sweat and saliva screening for drug use determination. However, these matrices may be applicable for use in driving while intoxicated and surveying populations for illicit drug use. Although not an illicit drug, the detection of ethanol is reviewed because of its importance in driving under the influence. Only with alcohol may saliva be used to estimate blood levels and the degree of impairment because of the problems with oral contamination and drug concentrations varying depending upon how the saliva is obtained. The detection of nicotine and cotinine (from smoking tobacco) is also covered because of its use in life insurance screening and surveying for passive exposure.

Decontamination procedures for drugs of abuse in hair: are they sufficient?

This paper reviews the methods for decontaminating hair exposed to external solutions of drugs of abuse. Exposure of hair to cocaine at 1 microgram/ml for 5 min is sufficient to contaminate hair, yet decontamination is a very slow process. Using externally contaminated hair, a number of decontamination procedures were attempted, and none removed all the contamination. The percentage of external contamination removed depended on the hair type, with thick black hair being the most resistant to decontamination. Hair treated by dying incorporated externally applied drugs differently, depending on the hair type. Thick black hair became more absorbent whereas thin brown hair became less absorbent. Kinetic wash criteria are evaluated for their ability/inability to determine if hair has been contaminated from external sources. A theoretical framework for the incorporation and removal of drugs from hair is discussed, and the hypothesis that inaccessible domains exist in hair which trap drugs is critically examined. The results presented in this paper strongly suggest that much more information on the decontamination of hair and the differentiation of exogenously and endogenously incorporated drugs is needed before hair analysis can be employed in most forensic applications. We propose that the radioactive tracer methods discussed herein are well suited for evaluating any new decontamination or extraction technique.

Chromogenic substrates for horseradish peroxidase

Two new detection systems for horseradish peroxidase (HRP) have been developed for the staining of membranes used in immunoassays. These systems use dimethyl or diethyl analogues of p-phenylenediamine with 4-chloro-1-naphthol to generate a blue product or 3-methyl-2-benzothiazolinone hydrazone with 4-chloro-1-naphthol to generate a red product. These reagents offer increased sensitivity and lower background staining than currently available chromogenic detection substrates. In addition, the incorporation of these substrates increases the sensitivity of HRP labels to be comparable to that of alkaline phosphatase with the 5-bromo-4-chloro-3-indolyl phosphate + nitro blue tetrazolium substrate.

External contamination of hair by cocaine: an issue in forensic interpretation

This paper explores the variables by which hair samples may become contaminated with cocaine and thereby generate false positives during analysis of hair samples. A novel method for following the incorporation and removal of cocaine from hair was developed. This method allowed a large number of specimens to be analysed under a variety of conditions with high precision. The quantity of cocaine was carefully followed in each step of a published procedure. Regardless of washing technique, a substantial amount of cocaine could still be found in the final hair digest. Very few of our externally contaminated samples could be identified as externally contaminated by previously published criteria and washing procedures. Attempts to further decontaminate this hair were without success. Our data strongly suggest that external contamination of hair by drugs of abuse may make the interpretation of forensic results problematical.

Cocaine in hair, saliva, skin swabs, and urine of cocaine users' children

The concentrations of cocaine and benzoylecgonine (BE) in the hair, saliva, skin secretions, and urine samples of cocaine-using mothers, their children, and other adults living in the same environment were compared. Subjects were screened from urban cocaine dependence treatment patients. Drug using adults had mean hair concentrations of 2.4 ng cocaine/mg hair (range = 0-12.2, sigma = 3.1, 15/16 positive) and 0.39 ng BE/mg hair (range = 0-1.9, sigma = 0.62), compared with childrens mean hair concentrations of 2.4 ng cocaine/mg of hair (range = 0-14.4, sigma = 3.8, 22/24 positive) and 0.74 ng benzoylecgonine/mg hair (range = 0-5.4 sigma = 1.3). None of the childrens urine specimens (0/22) were positive above 300 ng BE/ml. In contrast, 3/16 adult urine specimens were positive, even though they were enrolled in drug treatment. Saliva had detectable levels of BE for only one child (1/17) and one adult (1/17). Forehead swabs contained measurable quantities of cocaine for most children (19/26) and adults (15/17) and BE for children (7/26) and adults (7/17). Unlike urine results, overall hair cocaine concentrations for adults paralleled those of children and a clear cut-off concentration could not be established to differentiate these two groups.

Cocaine detection in a university population by hair analysis and skin swab testing.

The ability to detect cocaine use/exposure by either hair or sweat analysis was compared in a random population of adults at a major US university. Sweat was obtained by wiping the forehead with a cosmetic puff containing isopropanol. Using cut-off levels for sweat of 2.2 ng cocaine/wipe and of hair of 0.05 ng cocaine/mg hair, sweat detected two times more cocaine use/exposure than did hair. Sweat analysis detected a use rate of 12% compared to a 6% rate by hair analysis, both greater than the 2% that would be expected in this population. The high rate of detection was surprising and suggests that use of, if not exposure to, cocaine is underreported. Controlled experiments showed that cocaine could remain on the skin for about 3 days after external exposure. At the current state of knowledge, sweat appears to measure both use and exposure. Nevertheless, sweat testing could be used in several scenarios (such as roadside driving while intoxicated) where the case of collection and testing of sweat could outweigh the passive exposure considerations. Cocaine concentrations in skin swabs > 15 ng/swab would appear to indicate recent use/exposure.

Susceptibility of PharmChek™ drugs of abuse patch to environmental contamination

The key component of the PharmChek sweat patch, the membrane, has been tested for the passage of externally applied materials. Drugs in the uncharged state rapidly penetrated the membrane but charged species were greatly slowed. In basic media, detectable concentrations of cocaine, methamphetamine, and heroin were observed at the earliest collection time (ca. 30 s), after drugs were placed on the outside of the membrane. Drug concentrations increased over the 2 h time course, when amounts detected (1710 ng cocaine, 1060 ng methamphetamine, 550 ng heroin per pad at 2 h) represented 5-17% of the drug deposited on the surface of the sweat patch. Drugs externally applied to human skin were shown to bind readily. Drugs deposited on the skin of drug-free volunteers several days prior to application of the sweat patch were not completely removed by normal hygiene or the cleaning procedures recommended before application of the sweat patch. Even 6 days of normal hygiene did not remove all drugs from externally contaminated skin and positive sweat patches resulted. A mechanism for passage of drugs through the sweat patch membrane, a mechanism for retention of drugs on skin, and a redesign of the sweat patch and modification of its use to reduce external contamination are proposed. Appropriate care should be taken in the interpretation of positive results from a sweat patch test until more research is conducted.

Analysis of phencyclidine and cocaine in human hair by tandem mass spectrometry.

A confirmation procedure for the analysis of cocaine, benzoylecgonine, ecgonine, and phencyclidine (PCP) in human hair using tandem mass spectrometry has been developed. This procedure requires no solvent extraction and thus can examine the metabolites of drugs such as cocaine. Hairs from six cocaine users were examined and the amount of cocaine and its ratio to the metabolites was not correlated to the reported use pattern. Only PCP was detected in the hair from a PCP user, no metabolites were found. In passive exposure experiments, hair was found to tightly absorb PCP from aqueous solutions, which mimics the incorporation of PCP by the body. These results indicate that drugs may be absorbed onto the hair of nonusers and become tightly bound.

Evidence for bias in hair testing and procedures to correct bias

A number of in vitro experiments show that different hair samples incorporate differing amounts of drugs under identical conditions. Incorporation of cocaine and morphine tends to be correlated with race, in that the hair of African American females incorporates higher concentrations of cocaine than does the hair of Caucasian males or females. Extrapolation of these data into populations has been fraught with difficulties because the dosages of drugs and their use patterns are unknown. Cosmetic treatments and hygiene alter drug binding, which must be considered in comparing populations because cosmetic treatments are often group dependent. Four reasons are proposed that account for the uptake and retention of drugs by hair and that may differ among groups: (1) permeability and other characteristics of the hair due to genetic influences, (2) cosmetic hair treatments and hair care habits (which may be culturally influenced), (3) drug removal during personal hygiene, and (4) manner and route of drug administration which can affect passive exposure to residual drugs in the environment. The data supporting bias in hair testing are reviewed and methods are proposed that use either the uptake of dyes or the incorporation of drug homologs to reduce bias.

Bilayer lipid membrane (BLM) based ion selective electrodes at the meso-, micro-, and nano-scales

This paper presents a novel method for making micron-sized apertures with tapered sidewalls and nano-sized apertures. Their use in bilayer lipid membrane-based ion selective electrode design is demonstrated and compared to mesoscale bilayers and traditional PVC ion selective electrodes. Micron-sized apertures are fabricated in SU-8 photoresist films and vary in diameter from 10 to 40 microm. The tapered edges in SU-8 films are desired to enhance bilayer lipid membrane (BLM) formation and are fabricated by UV-light overexposure. Nano-apertures are made in boron diffused silicon film. The membranes are used as septa to separate two potassium chloride solutions of different concentrations. Lecithin BLMs are assembled on the apertures by ejecting lipid solution. Potassium ionophore, dibenzo-18-crown-6, is incorporated into BLMs by dissolving it in the lipid solution before membrane assembly. Voltage changes with increasing potassium ion concentrations are recorded with an A/D converter. Various ionophore concentrations in BLMs are investigated. At least a 1% concentration is needed for consistent slopes. Electrode response curves are linear over the 10(-6) to 0.1M range with a sub-Nernstian slope of 20mV per Log concentration change. This system shows high selectivity to potassium ions over potential interfering sodium ions. BLMs on the three different aperture sizes at the meso-, micro-, and nano-scales all show similar linear ranges and limits of detection (LODs) as PVC ion selective membranes.