Simple extraction of phencyclidine from human body fluids by headspace solid-phase microextraction (SPME)

Summary Phencyclidine (PCP) was found to be extractable by headspace solid-phase microextraction (SPME) from human whole blood and urine. Sample solutions were heated at 90°C in the presence of NaOH and K₂CO₃, and an SPME fiber was exposed in the headspace of a vial for 30 min. Immediately after withdrawal of the fiber, it was analyzed by gas chromatography with surface ionization detection (GC-SID). Recoveries of PCP were approximately 9.3–10.8% and 39.8–47.8% for whole blood and urine samples, respectively. The calibration curve for PCP showed good linearity in the range 2.5–100 ng mL^–1 whole blood and 0.5–100 ng mL^–1 urine. The detection limits were approximately 1.0 ng mL^–1 for whole blood and 0.25 ng mL^–1 for urine.     

Detection of ethanol in human body fluids by headspace solid-phase micro extraction (SPME)/capillary gas chromatography

Summary Ethanol has been found extractable from human whole blood and urine samples by headspace solid-phase micro extraction (SPME) with a Carbowax/divinylbenzene-coated fiber. After heating a vial containing the body fluid sample with ethanol, and isobutanol as internal standard (IS) at 70°C in the presence of (NH₄)₂SO₄, a Carbowax/divinylbenzene-coated SPME fiber was exposed in the headspace of the vial to allow adsorption of the compounds. The fiber needle was then injected into a middle-bore capillary gas chromatography (GC) port. The headspace SPME-GC gave intense peaks for both compounds; a small amount of background noises appeared, but did not interfere with the detection of the compounds. Recoveries of ethanol and IS were 0.049 and 0.026% for whole blood, respectively, and 0.054 and 0.085% for urine, respectively. The calibration curves for ethanol showed excellent linearity in the range of 80–5000 mg L^–1 for whole blood and 40–5000 mg L^–1 for urine; the detection limits for both samples were 20 and 10 mg L^–1, respectively. The data on actual determination of ethanol after the drinking of beer are also presented for two subjects.     

Automated headspace solid-phase microextraction and in-matrix derivatization for the determination of amphetamine-related drugs in human urine by gas chromatography-mass spectrometry

An automated extraction and determination method for the gas chromatography (GC)-mass spectrometry (MS) analysis of amphetamine-related drugs in human urine is developed using headspace solid-phase microextraction (SPME) and in-matrix derivatization. A urine sample (0.5 mL, potassium carbonate (5 M, 1.0 mL), sodium chloride (0.5 g), and ethylchloroformate (20 microL) are put in a sample vial. Amphetamine-related drugs are converted to ethylformate derivatives (carbamates) in the vial because amphetamine-related drugs in urine are quickly reacted with ethylchloroformate. An SPME fiber is then exposed at 80 degrees C for 15 min in the headspace of the vial. The extracted derivatives to the fiber are desorbed by exposing the fiber in the injection port of a GC-MS. The calibration curves show linearity in the range of 1.0 to 1000 ng/mL for methamphetamine, fenfluramine, and methylenedioxymethamphetamine; 2.0 to 1000 ng/mL for amphetamine and phentermine; 5.0 to 1000 ng/mL for methylenedioxyamphetamine; 10 to 1000 ng/mL for phenethylamine; and 50 to 1000 ng/mL for 4-bromo-2,5-dimethoxyphenethylamine in urine. No interferences are found, and the time for analysis is 30 min for one sample. Furthermore, this proposed method is applied to some clinical and medico-legal cases by taking methamphetamine. Methamphetamine and its metabolite amphetamine are detected in the urine samples collected from the patients involved in the clinical cases. Methamphetamine, amphetamine, and phenethylamine are detected in the urine sample collected from the victim of a medico-legal case.     

Determination of triazine herbicides in human body fluids by solid-phase microextraction and capillary gas chromatography

Summary Eight triazine herbicides, prometon, propazine, atrazine, simazine, prometryn, ametryn, metribuzin, and cyanazine, have been extracted from human whole blood and urine samples by headspace solid-phase microextraction (SPME) with a polydimethylsiloxane-coated fiber and quantified by capillary gas chromatography with nitrogen-phosphorus detection. Extraction efficiencies for all compounds were 0.21–0.99% for whole blood, except for cyanazine (0.06%). For urine, the extraction efficiencies for prometon, propazine, atrazine, prometryn and ametryn were 13.6–38.1%, and those of simazine, metribuzin and cyanazine were 1.35–8.73%. The regression equations for the compounds extracted from whole blood were linear within the concentration ranged 0.01–1 μg (0.5 mL)⁻¹ for prometon, propazine, atrazine, prometryn, and ametryn, and 0.02–1 μg (0.5 mL)⁻¹ for simazine, metribuzin, and cyanazine. For urine, regression equations for all compounds were linear within the concentration range 0.005–0.25 μg mL⁻¹. Compound detection limits were 2.8–9.0 ng (0.5 mL)⁻¹ and 0.4–2.0 ng mL⁻¹ for whole blood and urine, respectively. The coefficients of within-day and day-to-day variation were satisfactory for all the compounds, and not greater than 10.3 and 14.2%, respectively. Data obtained from determination of atrazine in rat whole blood after oral administration of the compound are also presented.     

Simple analysis of naphthalene, fluorene, and anthracene in whole blood by gas chromatography/mass spectrometry after headspace-solid phase microextraction

A simple and sensitive method for the simultaneous analysis of naphthalene, fluorene, and anthracene in whole blood was developed using headspace-solid-phase microextraction (SPME) and GC/MS. A 0.5 g whole blood sample, 5 microL naphthalene, fluorene, and anthracene (50 microg/mL) as spiked standards, and 0.5 mL sodium hydroxide were placed into a 12 mL vial and sealed rapidly. The vial was immediately heated to 70 degrees C in an aluminium block heater, the needle of the SPME device was inserted through the septum of the vial, and the extraction fiber was exposed to the headspace for 15 min. Afterwards, the compounds extracted by the fiber were desorbed simultaneously by exposing the fiber in the gas chromatograph injection port. No interferences were found, and the time for analysis was about 30 min for one sample. This method was applied to a suicide case in which the victim ingested naphthalene, fluorene, and anthracene.     

Sensitive determination of n-hexane and cyclohexane in human body fluids by capillary gas chromatography with cryogenic oven trapping

A sensitive method was developed for determination of n-hexane and cyclohexane in human body fluids by headspace capillary gas chromatography (GC) with cryogenic oven trapping. Whole blood and urine samples containing n-hexane and cyclohexane were heated in a 7.5 mL vial at 70 degrees C for 15 min, and 5 mL of the headspace vapor was drawn into a glass syringe. All vapor was introduced through an injection port of a GC instrument in the splitless mode into an Rtx-Volatiles middle-bore capillary column at an oven temperature of -40 degrees C for trapping volatile compounds. The oven temperature was programmed to 180 degrees C for GC with flame ionization detection. These conditions gave sharp peaks for both n-hexane and cyclohexane, a good separation of each peak, and low background impurities for whole blood and urine. The extraction efficiencies of n-hexane and cyclohexane were 13.2-30.3% for whole blood and 12.7-20.7% for urine. The coefficients of within-day variation in terms of extraction efficiency of both compounds were 5.0-9.5% for whole blood and 3.8-10.8% for urine; those of day-to-day variation for the compounds were not greater than 16.6%. The regression equations for n-hexane and cyclohexane showed good linearity in the range of 5-500 ng/0.5 mL for whole blood and urine. The detection limits (signal-to-noise ratio = 3) for both compounds were 1.2 and 0.5 ng/0.5 mL for whole blood and urine, respectively. The data on n-hexane or cyclohexane in rat blood after inhalation of each compound are also presented.     

Radiolytic degradation of malathion and lindane in aqueous solutions

Degradation of malathion and lindane pesticides present in an aqueous solution was investigated on a laboratory scale upon gamma-irradiation from a ⁶⁰Co source. The effects of pesticide group, presence of various additives and absorbed dose on efficiency of pesticide degradation were investigated. Gamma-irradiation was carried out in distilled water solutions (malathion and lindane) and in combination with humic solution (HS), nitrous oxide (N₂O) and HS/N₂O (lindane) over the range 0.1–2 kGy (malathion) and 5–30 kGy (lindane). Malathion was easily degraded at low absorbed doses compared to lindane in distilled water solutions. Absorbed doses required to remove 50% and 90% of initial malathion and lindane concentrations in distilled water solutions were 0.53 and 1.77 kGy (malathion) and 17.97 and 28.79 kGy (lindane), respectively. The presence of HS, N₂O and HS/N₂O additives in aqueous solutions, significantly improved the effectiveness of radiolytic degradation of lindane. Chemical analysis of the pesticides and the by-products resulted from the radiolytic degradation were made using a gas chromatography associated with mass spectrometry (GC–MS). Additionally, the final degradation products of irradiation as detected by ion chromatography (IC) were acetic acid and traces of some anions (phosphate and chloride).     

Simple Analysis of Amphetamines in Human Whole Blood and Urine by Headspace Capillary Gas Chromatography with Large Volume Injection

Summary A very simple method for the analysis of methamphetamine and amphetamine in human whole blood and urine by headspace gas chromatography (GC) has been presented. It neither needs solid-phase microextraction nor cryogenic trapping devices, but only a conventional capillary GC instrument with flame ionization detection (FID). The two special points to be mentioned in this method are the in-matrix derivatization of amphetamines for vaporization and the capability of injection of as large as 5 mL of the headspace vapor into a GC instrument in the splitless mode for sensitive detection. After heating a whole blood or urine sample containing amphetamines, -methylbenzylamine (internal standard, IS) and heptafluoro-n-butyryl chloride under alkaline conditions in a 7.0-mL vial at 90 °C for 20 min, 5 mL of the headspace vapor was drawn with a glass syringe and injected into the gas chromatograph. During injection the column was at 40 °C to trap the analytes, and then the oven temperature was programmed up to 320 °C. Sharp peaks were obtained for each analyte and IS, and only a relatively small number of background impurity peaks for the whole blood and urine samples. The detection limits for each amphetamine were estimated to be 0.1 g mL^–1 for whole blood and 0.03 g mL^–1 for urine. Precision and linearity were also tested to confirm the reliability. Methamphetamine and amphetamine could be determined from whole blood and urine obtained at autopsy in three methamphetamine poisoning cases. The identity of each peak appearing in the gas chromatograms was confirmed by GC/mass spectrometry.     

Extraction of local anaesthetics from human blood by direct immersion-solid phase micro extraction (SPME)

Summary Local anaesthetics have been shown to be extractable from human whole blood samples by direct immersion (DI)-solid phase micro extraction (SPME). After deproteinization with perchloric acid, the pH of the clear supernatants of human whole blood samples containing the drugs were adjusted to about 7 with 10 M NaOH in the presence of NaCl; a polydimethylsiloxanecoated SPME fiber was then immersed directly into the sample solution to allow adsorption of the drugs before capillary gas chromatography (GC) with flame ionization detection. The DI-SPME for 1-mL whole blood gave peaks for all the drugs; only a small amount of background noise appeared and this gave no problems in the detection of the drugs. Recoveries of the ten drugs from human whole blood was 0.74–19.7 %. The calibration curves for seven drugs showed linearity in the range of 0.25–12 g mL^–1 whole blood, with detection limits of 54–158 ng mL^–1.     

Pipette tip solid-phase extraction and gas chromatography – mass spectrometry for the determination of methamphetamine and amphetamine in human whole blood

Methamphetamine and amphetamine were extracted from human whole blood samples using pipette tip solid-phase extraction (SPE) with MonoTip C₁₈ tips, on which C₁₈-bonded monolithic silica gel was fixed. Human whole blood (0.1 mL) containing methamphetamine and amphetamine, with N-methylbenzylamine as an internal standard, was mixed with 0.4 mL of distilled water and 50 μL of 5 M sodium hydroxide solution. After centrifugation, the supernatant was extracted to the C₁₈ phase of the tip (pipette tip volume, 200 μL) by 25 repeated aspirating/dispensing cycles using a manual micropipettor. Analytes retained in the C₁₈ phase were eluted with methanol by five repeated aspirating/dispensing cycles. After derivatization with trifluoroacetic anhydride, analytes were measured by gas chromatography – mass spectrometry with selected ion monitoring in the positive-ion electron impact mode. Recoveries of methamphetamine and amphetamine spiked into whole blood were more than 87.6 and 81.7%, respectively. Regression equations for methamphetamine and amphetamine showed excellent linearity in the range of 0.5–100 ng/0.1 mL. The limits of detection for methamphetamine and amphetamine were 0.15 and 0.11 ng/0.1 mL, respectively. Intra- and interday coefficients of variation for both stimulants were not greater than 9.6 and 13.8%, respectively. The determination of methamphetamine and amphetamine in autopsy whole blood samples is presented, and was shown to validate the present methodology.     

Detection of 2,4,6-trichloroanisole in chlorinated water at nanogram per litre levels by SPME–GC–ECD

A method involving solid-phase micro extraction (SPME) and gas chromatography with electron capture detection (SPME–GC–ECD) has been optimised for identification and quantification of 2,4,6-trichloroanisole (TCA) at ng L^–1 concentrations in disinfected (chlorinated) water samples. A central composite design was used for factorial analysis of four factors, three factors related to the SPME (PDMS fibre) procedure (adsorption time, temperature of the sample during headspace sampling, and desorption time) and one related to the GC operation (the rate of increase of the temperature of the GC oven). Good linearity (linear correlation coefficient greater than 0.999) was observed for TCA concentrations up to 50 ng L^–1, limits of detection and quantification of 0.7 and 2.3 ng L^–1, respectively, and good precision (relative standard deviation 2.8% and 3.4% for 5 and 30 ng L^–1 of TCA, respectively). Besides TCA, this system also enables the detection and quantification of the four trihalomethanes in the g L^–1 concentration range with limits of detection and quantification of approximately 0.3 g L^–1 and 1 g L^–1, respectively.     

Sensitive analysis of alkyl alcohols as decomposition products of alkyl nitrites in human whole blood and urine by headspace capillary GC with cryogenic oven trapping

The abuse of alkyl nitrites is becoming a serious social problem worldwide. In this report, a simple and sensitive method is presented for the determination of n-butyl alcohol, isobutyl alcohol, and isoamyl alcohol as decomposition products of alkyl nitrites in human whole blood and urine samples using capillary gas chromatography (GC) with cryogenic oven trapping. After heating a whole blood or urine sample containing each alkyl alcohol and t-butyl alcohol [the internal standard (IS)] in a 7-mL vial at 55 degrees C for 15 min, 5 mL of the headspace vapor is drawn into a gas-tight syringe and injected into a GC inlet port. The vapor is introduced into an Rtx-BAC2 medium-bore capillary column in the splitless mode at 0 degrees C oven temperature in order to trap the entire analytes, and then the oven temperature is programmed up to 240 degrees C for the GC measurements by flame ionization detection. These conditions give sharp peaks for each compound and the IS and low background noise for whole blood or urine samples. The detection limits of the analytes are 10 ng/mL for whole blood and 5 ng/mL for urine. Linearity and precision are also tested to confirm the reliability of this method. Isobutyl alcohol and methemoglobin could be determined from the whole blood samples of three male volunteers who had sniffed isobutyl nitrite.     

A high-throughput approach for the determination of pesticide residues in cucumber samples using solid-phase microextraction on 96-well plate

A high-throughput solid-phase microextraction (SPME) on 96-well plate together with gas chromatography–mass spectrometry (GC–MS) was developed for the determination of some selected pesticides in cucumber samples. Pieces with the length of 1.0 cm of silicon tubing were precisely prepared and then coated on the end part of stainless steel wires. The prepared fibers were positioned in a home-made polytetrafluoroethylene (PTFE)-based constructed ninety-six holes block to have the possibility of simultaneous immersion of the SPME fibers into the center of individual wells. Pesticides such as diazinon, penconazol, tebuconazol, bitertanol, malathion, phosalone and chlorpyrifos-methyl were selected for their highly application in cucumber field. The performances of the SPME fibers, such as intra and inter-fibers reproducibility, were evaluated and the results showed a good similarity in extraction yields. A volume of 1 mL of the aquatic supernatant of the cucumber samples was transferred into the 96-well plate and the array of SPME fibers was applied for the extraction of the selected pesticides. The important parameters influencing the whole extraction process including, organic solvent percent, salt addition, dilution factor, stirring rate and extraction time were optimized. The inter- and intra-day RSD% were found to be less than 15.4%. Limits of detection (LOD) and limits of quantification (LOQ) were below 60 and 180 μg kg⁻¹, respectively. The coefficient of determination was satisfactory (r² > 0.99) for all the studied analytes. The developed method was successfully applied to the monitoring of several samples gathered from local markets.     

Determination of parathion in biological fluids by means of direct solid-phase microextraction

A new and simple procedure for the determination of parathion in human whole blood and urine using direct immersion (DI) solid-phase microextraction (SPME) and gas chromatography/mass spectrometry (GC/MS) is presented. This technique was developed using only 100 μL of sample, and ethion was used as internal standard (IS). A 65-μm Carbowax/divinylbenzene (CW/DVB) SPME fibre was selected for sampling, and the main parameters affecting the SPME process such as extraction temperature, adsorption and desorption time, salt addition, agitation and pH effect were optimized to enhance the sensitivity of the method. This optimization was also performed to allow the qualitative determination of parathion’s main metabolite, paraoxon, in blood. The limits of detection and quantitation for parathion were 3 and 10 ng/mL for urine and 25 and 50 ng/mL for blood, respectively. For paraoxon, the limit of detection was 50 ng/mL in blood. The method showed linearity between the LOQ and 50 μg/mL for both matrices, with correlation coefficients ranging from 0.9954 to 0.9999. Precision and accuracy were in conformity with the criteria normally accepted in bioanalytical method validation. The mean absolute recoveries were 35.1% for urine and 6.7% for blood. Other parameters such as dilution of sample and stability were also validated. Its simplicity and the fact that only 100 μL of sample is required to accomplish the analysis make this method useful in forensic toxicology laboratories to determine this compound in intoxications, and it can be considered an alternative to other methods normally used for the determination of this compound in biological media.     

Adsorptive voltammetric procedure for the determination of platinum baseline levels in human body fluids

Summary An extremly sensitive procedure for the determination of platinum in human body fluids is presented. A high pressure decomposition of the samples is followed by adsorptive voltammetric measurement. A detection limit down to 0.2 ng Pt/l sample allowed baseline levels of platinum in body fluids (urine: 0.5–15 ng/l, blood and blood plasma: 0.8–6.9 ng/l) to be evaluated. The concentration ranges in body fluids of occupationally exposed people were determined to 21–2900 ng/l (urine), 32–180 ng/l (blood) and 95–280 ng/l (blood plasma).     

Solid-phase microextraction for the investigation of sex pheromone of Eucosma notanthes Meyrick

A simple and efficient technique that does not require solvent and uses less operating time for the investigation of sex pheromones of the carambola fruit borer (Eucosma notanthes Meyrick) by utilizing headspace solid-phase microextraction (SPME) followed by GC-MS analysis has been developed. Variables such as types of SPME fiber, number of pests, temperature and extraction time have been studied. Whole sex glands of Eucosma notanthes Meyrick were dissected from 5 virgin insects, placed in a 2 mL vial, equilibrated at 170 °C for 10 min, and then extracted by headspace SPME at room temperature for 5 min. The results of the GC-MS analyses of headspace SPME of these sex glandular solid samples were much better than those obtained with hexane extraction of sex glandular from 117 insects followed by either headspace SPME or direct injection due to higher absorption efficiency. The simplicity of this technique renders it a very suitable method for research on the biological control of pests.     

Multiresidue analysis of 83 pesticides and 12 dioxin-like polychlorinated biphenyls in wine by gas chromatography–time-of-flight mass spectrometry

A multiresidue method is described for simultaneous estimation of 83 pesticides and 12 dioxin-like polychlorinated biphenyls (PCBs) in red and white wines. The samples (20 mL wine, acidified with 20 mL 1% HCl) were extracted with 10 mL ethyl acetate (+20 g sodium sulphate) and cleaned by dispersive solid-phase extraction (DSPE) with anhydrous calcium chloride and Florisil successively. The final extract (5 mL) was solvent exchanged to 1 mL of cyclohexane:ethyl acetate (9:1), further cleaned by DSPE with 25 mg primary secondary amine sorbent and analyzed by gas chromatography–time-of-flight mass spectrometry (GC–TOF-MS) within 31 min run time. The limits of quantification of most analytes were ≤10–20 μg/L. Acidification of wine prior to extraction prevented hydrolysis of organophosphorous pesticides as well as dicofol, whereas treatment with CaCl₂ minimized the fatty acid co-extractives significantly. Solvent exchange to cyclohexane:ethyl acetate (9:1) further minimized the co-extractives. Recoveries at 5, 10 and 20 ng/mL were >80% for most analytes except cyprodinil, buprofezin and iprodione. The expanded uncertainties at 10 ng/mL were <20% for most analytes. Intra-laboratory precision in terms of Horwitz ratio of all the analytes was below 0.5, suggesting ruggedness of the method. Effectively, the method detection limit for most analytes was as low as up to 1 ng/mL in both red and white wine, except for cyfluthrin and cypermethrin.     

Application of solid-phase microextraction in analytical toxicology

Solid-phase microextraction (SPME) is a miniaturized and solvent-free sample preparation technique for chromatographic–spectrometric analysis by which the analytes are extracted from a gaseous or liquid sample by absorption in, or adsorption on, a thin polymer coating fixed to the solid surface of a fiber, inside an injection needle or inside a capillary. In this paper, the present state of practical performance and of applications of SPME to the analysis of blood, urine, oral fluid and hair in clinical and forensic toxicology is reviewed. The commercial coatings for fibers or needles have not essentially changed for many years, but there are interesting laboratory developments, such as conductive polypyrrole coatings for electrochemically controlled SPME of anions or cations and coatings with restricted-access properties for direct extraction from whole blood or immunoaffinity SPME. In-tube SPME uses segments of commercial gas chromatography (GC) capillaries for highly efficient extraction by repeated aspiration–ejection cycles of the liquid sample. It can be easily automated in combination with liquid chromatography but, as it is very sensitive to capillary plugging, it requires completely homogeneous liquid samples. In contrast, fiber-based SPME has not yet been performed automatically in combination with high-performance liquid chromatography. The headspace extractions on fibers or needles (solid-phase dynamic extraction) combined with GC methods are the most advantageous versions of SPME because of very pure extracts and the availability of automatic samplers. Surprisingly, substances with quite high boiling points, such as tricyclic antidepressants or phenothiazines, can be measured by headspace SPME from aqueous samples. The applicability and sensitivity of SPME was essentially extended by in-sample or on-fiber derivatization. The different modes of SPME were applied to analysis of solvents and inhalation narcotics, amphetamines, cocaine and metabolites, cannabinoids, methadone and other opioids, fatty acid ethyl esters as alcohol markers, γ-hydroxybutyric acid, benzodiazepines, various other therapeutic drugs, pesticides, chemical warfare agents, cyanide, sulfide and metal ions. In general, SPME is routinely used in optimized methods for specific analytes. However, it was shown that it also has some capacity for a general screening by direct immersion into urine samples and for pesticides and other semivolatile substance in the headspace mode.     

Simple and sensitive analysis of nereistoxin and its metabolites in human serum using headspace solid-phase microextraction and gas chromatography-mass spectrometry

A simple method for the analysis of nereistoxin and its metabolites in human serum using headspace solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS) is developed. A vial containing a serum sample, 5M sodium hydroxide, and benzylacetone (internal standard) is heated to 70 degrees C, and an SPME fiber is exposed for 30 min in the headspace of the vial. The compounds extracted by the fiber are desorbed by exposing the fiber in the injection port of the GC-MS. The calibration curves show linearity in the range of 0.05-5.0 micrograms/mL for nereistoxin and N-methyl-N-(2-methylthio-1-methylthiomethyl)ethylamine, 0.01-5.0 micrograms/mL for S,S'-dimethyl dihydronereistoxin, and 0.5-10 micrograms/mL for 2-methylthio-1-methylthiomethylethylamine in serum. No interferences are found, and the analysis time is 50 min for one sample. In addition, this proposed method is applied to a patient who attempted suicide by ingesting Padan 4R, a herbicide. Padan 4R contains 4% cartap hydrochloride, which is an analogue of nereistoxin. Nereistoxin and its metabolites are detected in the serum samples collected from the patient during hospitalization. The concentration ranges of nereistoxin in the serum are 0.09-2.69 micrograms/mL.     

Evaluation of solid-phase micro-extraction coupled to gas chromatography-mass spectrometry for the headspace analysis of volatile compounds in cocoa products

The aroma profile of cocoa products was investigated by headspace solid-phase micro-extraction (HS-SPME) combined with gas chromatography–mass spectrometry (GC–MS). SPME fibers coated with 100 μm polydimethylsiloxane coating (PDMS), 65 μm polydimethylsiloxane/divinylbenzene coating (PDMS-DVB), 75 μm carboxen/polydimethylsiloxane coating (CAR-PDMS) and 50/30 μm divinylbenzene/carboxen on polydimethylsiloxane on a StableFlex fiber (DVB/CAR-PDMS) were evaluated. Several extraction times and temperature conditions were also tested to achieve optimum recovery. Suspensions of the samples in distilled water or in brine (25% NaCl in distilled water) were investigated to examine their effect on the composition of the headspace. The SPME fiber coated with 50/30 μm DVB/CAR-PDMS afforded the highest extraction efficiency, particularly when the samples were extracted at 60 °C for 15 min under dry conditions with toluene as an internal standard. Forty-five compounds were extracted and tentatively identified, most of which have previously been reported as odor-active compounds. The method developed allows sensitive and representative analysis of cocoa products with high reproducibility. Further research is ongoing to study chocolate making processes using this method for the quantitative analysis of volatile compounds contributing to the flavor/odor profile.