Analysis of anaesthetics and analgesics in human urine by headspace SPME and GC.

The determination of widely used anaesthetic and analgesic drugs in biological fluids is of major clinical importance. Typical methods used for sample preparation employ liquid-liquid extraction protocols which are complex, costly, not handy and not amenable to automation. In the present communication, we report the development of a methodology that employs headspace solid-phase microextraction (HS-SPME) for the determination of four anaesthetic (lidocaine, midazolam, diazepam and ketamine) and three analgesic drugs (fentanyl, remifentanyl and codeine) in human urine. Important parameters controlling SPME were studied: selection of SPME fibre, type and amount of salt added, preheating and extraction time, extraction temperature, sample volume and desorption time. GC with nitrogen phosphorus detection (GC-NPD) facilitates sensitive and selective detection of the anaesthetics. The developed method renders an efficient tool for the precise and sensitive determination of the anaesthetics and analgesics in human urine (RSDs ranged from 7.7 to 12.6%, whereas LODs ranged from 0.01 to 1.5 ng/mL). The method was applied to the determination of the anaesthetics and analgesics in human urine from patients that had undergone coronary by-pass surgery operations. The proposed protocol can function as an attractive alternative for clinical acute intoxications and medico-legal cases.

Tolerance to propofol's sedative effect in mechanically ventilated Rabbits.

Propofol is commonly used for the sedation of critically ill patients undergoing mechanical ventilation. These patients may develop tolerance during long-term administration. Here, we describe the development of tolerance to propofol's sedative effect in rabbits during prolonged mechanical ventilation. Six healthy male New Zealand White rabbits were endotracheally intubated and received propofol by continuous IV infusion to maintain sedation for 48 h. The propofol infusion rate (IR) was adjusted to maintain the desired level of sedation. Assessments of the sedation level were made every 30 min or earlier if there were signs of awakening. Propofol concentrations were measured in arterial plasma after every other IR adjustment, provided there was an adequate level of sedation, using high performance liquid chromatography, and calculations of systemic clearance rates were made. The mortality rate was 100% with a survival period of 30.8 +/- 6.0 h (mean +/- sd). The course of IR adjustments followed a 5-phase pattern: 1) steady IR (mean +/- sd duration; 1.2 +/- 0.6 h), 2) increasing IR (9.4 +/- 5.5 h), 3) steady high-IR (2.3 +/- 1.2 h), 4) decreasing IR (13.7 +/- 1.9 h), and 5) steady low-IR (5.0 +/- 2.7 h). The course of propofol concentrations during the experiment in relation to propofol IR followed a 3-phase pattern: 1) steady concentration with increasing IRs (6.0 +/- 2.7 h), 2) increasing concentrations with increasing IR (5.8 +/- 2.5 h), and 3) increasing concentrations with decreasing IR (18.8 +/- 3.3 h). Propofol systemic clearance rates were progressively increased for 6.0 +/- 2.7 h and then gradually decreased for 24.6 +/- 4.7 h. In conclusion, all rabbits developed tolerance to propofol's sedative effect within the first hours of administration related to changes to the drug's metabolic clearance.

Solid phase microextraction gas chromatographic analysis of organophosphorus pesticides in biological samples.

Headspace-solid phase microextraction (HS-SPME) was studied and optimised for the determination of four common organophosphorus pesticides (OPPs) in biological samples. Various parameters controlling SPME were studied: choice of SPME fiber, type and content of salt added, preheating and extraction time, desorption time, extraction temperature. Capillary gas chromatographic analysis with nitrogen phosphorus detection (GC-NPD) facilitates sensitive and selective detection of the OPPs: malathion, parathion, methyl parathion and diazinon. Fenitrothion was used as the internal standard. The method was applied to the determination of the pesticides in human biological specimens: whole blood, blood plasma, urine, cerebrospinal fluid, liver and kidney. Limits of detection ranged from 2 to 55 ng/ml depending on pesticide and type of specimen. The developed methodology overcomes limitations and obstacles of conventional methods such as the use of organic solvents, the formation of emulsions and the tedious-cumbersome procedures. The proposed protocol is seen as an attractive alternative to be used in routine toxicological analysis.

Determination of amphetamines in human urine by headspace solid-phase microextraction and gas chromatography.

Solid-phase microextraction (SPME) is under investigation for its usefulness in the determination of a widening variety of volatile and semivolatile analytes in biological fluids and materials. Semivolatiles are increasingly under study as analytical targets, and difficulties with small partition coefficients and long equilibration times have been identified. Amphetamines were selected as semivolatiles exhibiting these limitations and methods to optimize their determination were investigated. A 100- micro m polydimethylsiloxane (PDMS)-coated SPME fiber was used for the extraction of the amphetamines from human urine. Amphetamine determination was made using gas chromatography (GC) with flame-ionization detection (FID). Temperature, time and salt saturation were optimized to obtain consistent extraction. A simple procedure for the analysis of amphetamine (AMP) and methamphetamine (MA) in urine was developed and another for 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxy-N-methamphetamine (MDMA) and 3,4-methylenedioxy-N-ethylamphetamine (MDEA) using headspace solid-phase microextraction (HS-SPME) and GC-FID. Higher recoveries were obtained for amphetamine (19.5-47%) and methamphetamine (20-38.1%) than MDA (5.1-6.6%), MDMA (7-9.6%) and MDEA (5.4-9.6%).

Development and validation of a high-performance liquid chromatography method for the evaluation of niflumic acid cross-reactivity of two commercial immunoassays for cannabinoids in urine.

Niflumic acid is a nonsteroidal, anti-inflammatory drug widely prescribed in Greece. We recently noticed that this drug cross-reacts for cannabinoids in a kinetic interaction of microparticles in a solution (KIMS) immunoassay method but does not in an enzyme multiplied immunoassay technique (EMIT) immunoassay method. The objective of the study was to develop and validate a high-performance liquid chromatographic method in order to evaluate niflumic acid cross-reactivity in two commercial immunoassays for cannabinoids in urine, both in niflumic acid standards as well as in urine specimens obtained from subjects receiving niflumic acid. Urine niflumic acid standards were prepared in drug-free urine at 13 concentrations ranging from 1.25 to 1000 microg/mL. The standards gave presumptive positive cannabinoids results when analyzed by the KIMS immunoassay method when the concentration was above 2.5 microg/mL. None of the prepared standards gave a false-positive cannabinoid result when analyzed by the EMIT immunoassay method. By applying a 50 ng/mL cutoff for cannabinoids in these assays, all 55 urine specimens collected from the 5 subjects who participated gave negative results by the EMIT and false-positive results by the KIMS immunoassay method. It is concluded that KIMS is more prone to cross-reactions by niflumic acid compared to EMIT. Therefore, all positive screening tests for cannabinoids obtained by KIMS should be confirmed by another technique.