Use of headspace solid-phase microextraction (HS-SPME) in hair analysis for organic compounds

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Abstract

Headspace solid phase microextraction (HS-SPME) has advantages of high purity of the extract, avoidance of organic solvents and simple technical manipulation and can be used in combination with gas chromatography–mass spectrometry (GC-MS) in the hair analysis of a number of drugs. HS-SPME coupled with the hydrolysis of the hair matrix by 4% sodium hydroxide in the presence of excess sodium sulphate and of a suitable internal standard proved to be a convenient one-step method for the measurement of many lipophilic basic drugs such as nicotine, amphetamine derivatives, local anaesthetics, phencyclidine, ketamine, methadone, diphenhydramine, tramadol, tricyclic antidepressants and phenothiazines. Detection limits were between 0.05 and 1.0 ng/mg. From spiked 10-mg hair samples absolute recoveries between 0.04 and 5.7% were found. These recoveries decreased considerably if larger sample amounts were used, perhaps due to increased drug solubility in the aqueous phase or to elevated viscosity in the presence of dissolved hair proteins. Because of the phenolic hydroxyl group a change of pH after alkaline hair digestion (by adding excess orthophosphoric acid) was necessary for the detection of Δ9-tetrahydrocannabinol (Δ9-THC), cannabinol (CBN) and cannabidiol (CBD) by HS-SPME. Nevertheless, the detection limits were such that only CBN could be detected in hair of a consumer. Clomethiazole, a compound hydrolysed in alkali, was measured by HS-SPME after extraction with aqueous buffer. The detection limit was 0.5 ng/mg. Cocaine could not be detected by HS-SPME. The application of HS-SPME to hair samples from several forensic and clinical cases is described.

Introduction

Methods used in sample preparation for analysis of illicit or therapeutic drugs in hair generally involve extraction or digestion of the hair matrix and subsequent clean up by solid phase extraction [1]. Such methods involve several steps and a high degree of experience is needed to obtain reproducible results. On the other hand, in headspace gas chromatography the analyte is separated from the biological matrix in a simple way, but this method is limited to volatile compounds and there are problems in attempting to combine it with mass spectrometry because of the relatively large amount of air injected with the sample.

Headspace-solid phase microextraction (HS-SPME) invented by Pawliszyn and coworkers [2], [3] has proved to be a simple way of avoiding this problem and increasing sensitivity. The principle of HS-SPME is shown in Fig. 1. A fused silica fibre coated with a 7–100-μm layer of, for example, polyacrylate (PA), polydimethylsiloxan/polydivinylbenzene (PDMS/DVB) or polydivinylbenzene/polyethylenglycol (DVB/Carbowax) and protected in a stainless steel injection needle (SPME-device) is placed in the vapour phase of the headspace vial. The fibre is then exposed for a certain time at a certain temperature (adsorption time and adsorption temperature) and substances evaporated from the liquid or solid sample are absorbed into or adsorbed onto the layer. After that the fibre is retracted into the needle, which is injected into the injection port of the GC-MS. There the fiber is exposed again, and at the high temperature the substances are immediately desorbed for separation and identification.

In toxicological analysis HS-SPME has been used in several cases for the detection and quantitation of volatile compounds from blood or urine [4], [5], [6]. But, surprisingly, compounds with a low volatility (‘semivolatile compounds’ [7], [8]) such as amphetamines [9], [10], [11], phencyclidine [12], tri- and tetracyclic antidepressants [13], [14], local anaesthetics [15], [16], phenothiazines [17], diphenylmethane antihistamines [18] and some pesticides [19], [20] can be measured by HS-SPME in body fluids with high sensitivity.

SPME was used in hair analysis by Strano-Rossi and Chiarotti to detect cannabinoids, methadone and cocaine [21], not in the headspace mode but by direct immersion of the SPME fibre in the solution remaining after alkaline or enzymatic hair digestion. However, the analysis of amphetamine and methamphetamine from hair by HS-SPME was described by Koide et al. [22]. In a previous investigation [16] we found that the local anaesthetic lidocaine (lignocaine), which is frequently used as an adulterant of cocaine and cocaine–heroin mixtures, can be measured easily in 10-mg hair samples from drug consumers with a detection limit of less than 0.1 ng/mg using HS-SPME after alkaline hydrolysis of the matrix in the headspace vessel. In order to examine to what extent HS-SPME can be used as a general method in hair analysis for organic compounds, hair samples were investigated for a series of drugs, which were either naturally present from incorporation after intake or were added by spiking.

Section snippets

Reference substances and reagents

Therapeutic drugs used as reference substances were generously donated by the corresponding manufacturers. Samples of illicit drugs and deuterated standards were from Sigma or Promochem. All solvents and reagents were obtained from Merck/Darmstadt (Germany) in analytical purity.

Hair samples

For the investigations with spiked samples a pool was prepared of head hair from volunteers who had not taken illicit or therapeutic drugs. The hair was washed (5 min) with deionised water and with acetone in an

Results and discussion

Digestion of the hair matrix in alkaline solution, for example with 1 M sodium hydroxide, is one of the most efficient and convenient sample preparation methods if the analyte(s) are stable under these conditions. Examples are carbamazepine [23], amphetamines [24], and tricyclic antidepressants [25]. However, esters such as heroin or cocaine are hydrolysed under these conditions. An advantage of HS-SPME is that the dissolution of the hair matrix and the extraction can be combined in the same

Conclusions

HS-SPME can be applied with advantage to hair analysis in a convenient one-step method for a wide variety of basic drugs including frequently abused compounds such as phencyclidine, ketamine and methadone. Some of the results presented here are preliminary, and the methods are not fully evaluated. By adaptation of the conditions to the special properties of a particular substance an extension to some other drugs and a further decrease in detection limits are possible. Practical advantages of

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