Dextromethorphan Analgesia in a Human Experimental Model of Hyperalgesia.

BACKGROUND: Central pain sensitization is often refractory to drug treatment. Dextromethorphan, an N-methyl-D-aspartate receptor antagonist, is antihyperalgesic in preclinical pain models. The hypothesis is that dextromethorphan is also antihyperalgesic in humans. METHODS: This randomized, double-blind, placebo-controlled, crossover study explores the antihyperalgesic effect of single and repeated 30-mg dose of oral dextromethorphan in 20 volunteers, using the freeze-injury pain model. This model leads to development of primary and secondary hyperalgesia, which develops away from the site of injury and is associated with central sensitization and activation of N-methyl-D-aspartate receptor in the spinal cord. The primary outcome was antihyperalgesia calculated with the area under the curve of the percentage change in mechanical pain threshold (electronic von Frey) on the area of secondary hyperalgesia. The secondary outcomes were mechanical pain threshold on the area of primary hyperalgesia and cognitive (reaction time) effect. RESULTS: Single 30-mg results are reported. Antihyperalgesia (% · min) is significantly higher on the area of secondary hyperalgesia with dextromethorphan than placebo (median [interquartile range]: 3,029 [746; 6,195] vs. 710 [-3,248; 4,439], P = 0.009, Hedge's g = 0.8, 95% CI [0.1; 1.4]). On primary hyperalgesia area, mechanical pain threshold 2 h after drug intake is significantly higher with dextromethorphan (P = 0.011, Hedge's g = 0.63, 95% CI [0.01; 1.25]). No difference in antinociception is observed after thermal painful stimuli on healthy skin between groups. Reaction time (ms) is shorter with placebo than with dextromethorphan (median [interquartile range]: 21.6 [-37.4; 0.1] vs. -1.2 [-24.3; 15.4], P = 0.015, Hedge's g = 0.75, 95% CI [0.12; 1.39]). Nonserious adverse events occurrence (15%, 3 of 20 volunteers) was similar in both groups. CONCLUSIONS: This study shows that low-dose (30-mg) dextromethorphan is antihyperalgesic in humans on the areas of primary and secondary hyperalgesia and reverses peripheral and central neuronal sensitization. Because dextromethorphan had no intrinsic antinociceptive effect in acute pain on healthy skin, N-methyl-D-aspartate receptor may need to be sensitized by pain for dextromethorphan to be effective.

Determination of total and unbound concentrations of lopinavir in plasma using liquid chromatography-tandem mass spectrometry and ultrafiltration methods.

Lopinavir is an HIV protease inhibitor with high protein binding (98-99%) in human plasma. This study was designed to develop an ultrafiltration method to measure the unbound concentrations of lopinavir overcoming the non-specific binding issue. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of total concentrations of lopinavir in plasma was developed and validated, and an adaptation was also optimized and validated for the determination of unbound concentrations. The chromatographic separation was performed with a C18 column (100 mm × 2.1mm i.d., 5 µm particle size) using a mobile phase containing deionized water with formic acid, and acetonitrile, with gradient elution at a flow-rate of 350 µL min(-1). Identification of the compounds was performed by multiple reaction monitoring, using electrospray ionization in positive ion mode. The method was validated over a clinical range of 0.01-1 µg/mL for human plasma ultrafiltrate and 0.1-15 µg/mL in human plasma. The inter and intra-assay accuracies and precisions were between 0.23% and 11.37% for total lopinavir concentrations, and between 3.50% and 13.30% for plasma ultrafiltrate (unbound concentration). The ultrafiltration method described allows an accurate separation of the unbound fraction of lopinavir, circumscribing the loss of drug by nonspecific binding (NSB), and the validated LC-MS/MS methodology proposed is suitable for the determination of total and unbound concentrations of lopinavir in clinical practice.

[Use of illicit substances in the workplace]. / Consommation de substances illicites en milieu professionnel.

The development of addictive behaviors is a source of worry and concern for workplace and occupational physicians. To estimate the prevalence of behaviors, two types of surveys can be carried out: self-assessment surveys and biological testing in the workplace. For the latter, when a settlement is within the company, the prevalence is often lower compared to those enterprises that have not adapted this policy. Very few investigations have been published in France to date. Data published by the United Nations Office against Drugs and Crime (UNODC) shows a stable consumption of illicit substances in recent years. They reported consumption in the world among the general population (all subjects aged 15 to 64). For France, were described a prevalence estimated in 2005 to 8.6%, 0.6% and 0.2% for cannabis, cocaine and amphetamine derivatives, respectively, and in 2007 to 4.6% for opiates. Some prevalence in the workplace have been reported in Europe in chemical, petrochemical, metallurgical, automotive, in the transport sector and in medical and military fields. However, it appears that few surveys in the workplace have been published in France, this lack may be explained by a desire for anonymity on the subject at the level of company management and doctors work that focus on individual support with the problem of addiction. Screening for illicit substances is necessary because these psychotropic substances affect alertness and pose risks in the workplace, especially such that the association cannabis-alcohol further increases the risk. Knowledge of consumption is, moreover, an important factor in job security. It may be acquired if reliable methods, inexpensive to allow routine screening. Publication of results will reveal the extent of the problem and implement more effective campaigns of information and prevention in the workplace.

[Assessment of occupational exposure to ortho-toluidine using gas chromatography-mass spectrometry]. / Evaluation d'une exposition professionnelle à l'ortho-toluidine par chromatographie phase gazeuse couplée a la spectrométrie de masse.

Ortho-toluidine is a carcinogen aromatic amine. It is in part eliminated as unchanged form and its urine determination allows biologic monitoriing of occupational exposure. We propose a new method simple and fast in gas chromatography mass spectrometry. In the Nord-Pas-de-Calais region, a company initiated destruction and depollution of an old industrialsite.The GS-MS method permits exposition evaluation of workers employed in demolition of a liquid SO2 plant polluted with ortho-toluidine. This plant has been stopped twenty years ago. These results are compared with results of workers without any exposure in the same company. A 5 mL urine sample spiked with internal standard (ortho-toluidine D9) is extracted with hexane. Derivatisation is achieved with anhydrous pentafluoropropionic acid during 30 min at 60 degrees C. Chromatographic separation is performed on a BPX5MS column (25 m x 0.25 mm, 0.25 microm; SGE). Initial column temperature (60 degrees C) is hold 3 min then is raised to 300 degrees C at 25 degrees C/min. Detection is performed with mass spectrometry with negative chemical ionisation with methane. Acquisition is performed in single ion monitoring. Identification ions are 233 ion (m/z) and 213 ion (m/z) with 233 (m/z) used for quantification. Linearity of the method is verified between 0.1 and 100 microg/L. The limit of detection is 0.02 micro/L. Repeatability and intermediate fidelity are satisfactory (CV < 9%). For unexposed workers, urinary concentrations of ortho-toluidine ranged between 0.17 microg/L and 2.46 microg/g creatinine. Urinary concentrations for exposed workers ranged between 26.14 and 462.00 microg/g creatinine and after new action of protection between 2.35 et 20.11 microg/g creatinine. This new GC-MS method is specific and sensitive and allows for urinary determination of ortho-toluidine. Results showed that this method is adapted for biomonitoring as much for unexposed workers to this aromatic amine as for exposed workers.

A fatal case of mercuric cyanide poisoning.

A 57-year-old pharmacist was found dead 11 days after his disappearance. At the autopsy, samples of blood, urine, gastric content were obtained. Presence of ethanol, cyanide and mercury were detected in some samples. Cyanide and mercury were identified and quantified using high-performance liquid chromatography with diode array detector (HPLC) in fluorescence mode and ICP with mass selective detector (ICP-MS) respectively. Whole blood concentrations of ethanol was 1.72 g/L. Cyanide and mercury concentrations in whole blood were respectively 0.16 and 3.8 mg/L. Concentrations of cyanide (27 mg/L) and mercury (150 mg/L) in gastric contents prove a massive oral ingestion of mercuric cyanide or mercuric oxycyanide occurred. In this case report, the death was attributed to the combined toxicity of cyanide and mercury.

Exposure to glycol ethers in a population of French men evaluated by measurement of urinary alkoxycarboxylic acids.

OBJECTIVES: Glycol ethers are solvents that are present in a large number of products used commercially and domestically. During recent years, ethylene glycol ether derivatives, in particular ethylene glycol methyl ether and ethylene glycol ethyl ether, have been progressively replaced by propylene glycol ether derivatives, which are less toxic. The aim of this study was to estimate the level of exposure to glycol ethers in a sample population of French men employed by the Paris Municipality by measuring the amount of alkoxycarboxylic acid metabolites in their urine. METHODS: Urine samples were collected at the end of two different working weeks from 109 men, 54 of whom were judged to be occupationally exposed to glycol ether-containing products. Five alkoxyacetic acids (methoxyacetic, ethoxyacetic, n-propoxyacetic, phenoxyacetic, butoxyacetic acids) from ethylene glycol derivatives, and one alkoxypropionic acid (2-methoxypropionic) from a propylene glycol derivative, were simultaneously analysed by gas chromatography coupled to electron-capture detection. RESULTS: 2-Methoxypropionic was the most frequently found alkoxycarboxylic acid. The concentration of this metabolite reached 5.6 mmol/mol creatinine. The second most common alkoxycarboxylic acid was phenoxyacetic (up to 2.3 mmol/mol creatinine). The concentrations of the other alkoxycarboxylic acids were less than 1 mmol/mol creatinine. Although the concentration of alkoxycarboxylic acids was higher among men occupationally exposed to glycol ether-containing products than among unexposed men, the difference was significant only for butoxyacetic acid. CONCLUSIONS: Our data suggest that the use and exposure levels of glycol ethers have qualitatively and quantitatively changed dramatically over recent years. Particular attention should be paid in the future to alkoxypropionic acids derived from minor isomers of propylene glycol ether derivatives.

Rapid and simple determination of selenium in blood serum by inductively coupled plasma-mass spectrometry (ICP-MS).

An inductively coupled plasma mass spectrometer (ICP-MS) with a rapid sample-preparative procedure was used for the determination of selenium in blood serum. Blood serum was prepared by dilution in an acidic solution consisting of nitric acid (1%), X-triton (0.1%) and 1-butanol (0.8%). A calibration curve was established for 1-40 microg mL(-1) (r(2)>0.99). The limit of detection was 0.5 microg mL(-1). Repeatability and intermediate precision were satisfactory with relative standard deviations (RSD) of 2.0% and 3.2%, respectively. This method was easily applied to reference materials with satisfactory accuracy. Good correlation (r(2)=0.96) was observed between ICP-MS and atomic absorption spectrometry (AAS) for the determination of (82)Se in blood serum from 23 patients. These results suggest that the sample preparative procedure coupled with ICP-MS can be used for the routine determination of (82)Se in human blood serum.

Separation of new antidepressants and their metabolites by micellar electrokinetic capillary chromatography.

Selective serotonin reuptake inhibitors (SSRIs), serotonin noradrenergic reuptake inhibitors (SNaRIs) and noradrenergic and specific serotoninergic antidepressant (NaSSA) are widely used in the treatment of depression. An increase in antidepressant intoxications led to the development of reliable analytical methods for their analysis. A new determination procedure for these compounds (milnacipran, venlafaxine, desmethylvenlafaxine, mirtazapine, desmethylmirtazapine, citalopram, desmethylcitalopram, fluvoxamine, paroxetine, sertraline and fluoxetine) was developed by micellar electrokinetic capillary chromatography (MEKC) with diode array detection (DAD). Separation and determination were optimised on an uncoated fused-silica capillary (600 mm, 75 microm I.D.). The migration buffer consisted of 20 mM sodium borate, pH 8.55, with 20 mM SDS and 15% isopropanol, at an operating voltage of 25 kV. The column temperature was maintained at 40 degrees C. Injection in the capillary was performed in the hydrodynamic mode (0.5 p.s.i., 15 s). In these conditions, the migration time of the antidepressants was less than 11 min. In most cases, calibration curves were established for 30 - 2000 ng/ml (r > 0.995). The limit of detection and the limit of quantification were ranged between 10 and 20 and between 20 and 30 ng/ml, respectively, for all the molecules. This method allowed the determination of some of these compounds in biological fluids (blood, urine) in post-mortem cases. Samples (1 ml) were extracted with diethyl ether (5 ml) at pH 9.6 and reconstituted in diluted migration buffer. Similar results were obtained by a HPLC-DAD determination, performed as a reference method. These results suggest that this MEKC method can be useful for the determination of new antidepressants in post-mortem cases.

Spectrophotometric, spectrofluorimetric, HPLC and CZE determination of mirtazapine in pharmaceutical tablets.

Four analytical methods have been developed for the quality control of tablets containing mirtazapine: spectrophotometry, spectrofluorimetry, high performance liquid chromatography (HPLC) and capillary zone electrophoresis (CZE). All the methods only require a simple extraction procedure of mirtazapine from the tablets before analysis. The concentration of mirtazapine in solutions was determined in the linearity range of 5-25 microg/ml at lambda=315 nm for spectrophotometry and at lambda=220 nm for HPLC and CZE. Spectrofluorimetric determinations were achieved at lambda(excitation)=328 nm and lambda(emission)=415 nm in the linearity range of 2-25 ng/ml. All the methods gave similar results and were validated for selectivity, linearity, precision and sensitivity. Spectrometric methods gave slightly higher RSD values (up to 2.54%). The four methods were directly and easily applied to the pharmaceutical preparation with accuracy, resulting from recovery experiments between 99.72% in HPLC and 101.47% in spectrofluorimetry.

Comparison of high-performance liquid chromatography and capillary zone electrophoresis for the determination of parabens in a cosmetic product.

A high-performance liquid chromatographic method (HPLC) and a capillary zone electrophoresis method (CZE) have been developed for the analysis of methylparaben, ethylparaben, propylparaben and butylparaben in a commercial cosmetic product. A very simple extraction procedure with acidified diethylether was developed. The HPLC method involved a C18 reversed-phase column and a gradient of methanol and water-acetic acid (1%). Electrophoretic separation was performed on a fused-silica capillary with a mixed 15 mM tetraborate buffer (pH 9.2) and methanol (85:15, v/v). The calibration curves were linear from 1 to 40 microg/ml in HPLC and from 5 to 200 microg/ml in CZE. The limit of detection in CZE (0.21 microg/ml) was higher than in HPLC (0.05 microg/ml). Repeatability and intermediate precision were satisfactory for both methods (RSD values < 3.23% in HPLC and < 3.26%, in CZE). Only HPLC allowed the separation of butylparaben isomeric forms when CZE analysis was less time and reagents consuming. These results suggest that HPLC and CZE coupled with a simple extraction process are both suitable for parabens determination in cosmetic products.

Determination of urea, allantoin and lysine pyroglutamate in cosmetic samples by hydrophilic interaction chromatography.

A new HPLC method using a Polyhydroxyethyl A column involving hydrophilic interaction chromatography (HILIC) is described for the simultaneous determination of urea, allantoin and lysine pyroglutamate in a cosmetic cream. Validation of the method was accomplished with respect to linearity, repeatability and limits of detection/quantification. Compound recoveries approach 100% with acceptable RSD values. The method is very simple since no derivatisation is necessary. Furthermore, it allows the rapid and direct chromatographic analysis of urea and hence could provide an alternative to other methods used to determine this compound in biological or cosmetic samples.

Relationship between lipophilicity and binding to human serum albumin of arylpropionic acid non-steroidal anti-inflammatory drugs.

A possible relationship between lipophilicity and binding to human serum albumin was investigated for 11 arylpropionate non-steroidal anti-inflammatory drugs. The lipophilic parameter was determined by a reversed-phase high-performance liquid chromatographic procedure as the capacity factor (k'). The binding of arylpropionic acids to human serum albumin was studied in vitro by equilibrium dialysis. For each compound, a Scatchard analysis was performed considering two classes of binding sites characterized by high- and low-affinity constants, K1 and K2, respectively. A linear relationship was found between lipophilicity and binding parameters, n1K1 (r = 0.88, P < 0.0005) and n2K2 (r = 0.96, P < 0.0002). These results suggest the role of hydrophobic interactions in the binding of arylpropionic acids to human serum albumin.

Monocyte/macrophage recruitment and expression of endothelial adhesion proteins in human atherosclerotic lesions.

Since mononuclear cells are recruited in atherosclerotic lesions, the expression of adhesion proteins by the arterial endothelium may play a major role in atherogenesis. The relationships between ICAM-1, E-selectin, and VCAM-1 expression on the arterial endothelium and the presence and degree of maturation of intimal macrophages in human atherosclerotic lesions was investigated. By quantitative double immunostaining with a pan-macrophage-specific monoclonal antibody, HAM-56, and a recently developed monoclonal antibody that is specific for mature macrophages, 3MA-B38, arterial sections were classified as (I) normal, (II) thickened without macrophage infiltration, (III) atherosclerotic with recent macrophage infiltration or (IV) atherosclerotic with infiltration of mature differentiated macrophages. A marked increase in the expression of ICAM-1, E-selectin, and VCAM-1 was observed on endothelial cells adjacent to recently recruited macrophages. Endothelial cells overlying differentiated macrophages exhibited a lower but significant increase in VCAM-1 expression, with no difference in ICAM-1 and E-selectin expression with respect to that observed in endothelium of normal arteries. These findings indicate that the endothelium covering the human arterial wall exhibits different states of activation as reflected by the expression of adhesion proteins, and that intimal monocyte/macrophage recruitment appears to depend on the level of expression of adhesion proteins.

Central analgesic effects of aspirin-like drugs.

Aspirin-like drugs mainly include paracetamol, salicylates and other non-steroidal anti-inflammatory drugs, and metamizole. Their analgesic effect is classically ascribed to a peripheral site of action, within the pain-processing site. There is, however, convincing evidence that a central component contributes to the overall analgesia provided by these agents. Experimental and clinical studies referring to this challenging proposal are reviewed here. The exact site and mode of action of aspirin-like drugs within the central nervous system remains controversial. It is likely that supraspinal mechanisms play an important role. Some experiments lend support to the involvement of monoaminergic control systems. Other data indicate that these drugs act centrally through the inhibition of cyclo-oxygenase activity. The interactions between prostaglandins and various neurotransmitters suggest that both mechanisms may be linked.

Determination of flunitrazepam in plasma by liquid chromatography.

A reversed-phase liquid chromatographic method has been used to determine flunitrazepam in plasma. Extraction was simple and there was no need to hydrolyse the drug. Separation was achieved on a 150 x 3.9 mm i.d. column packed with 4-microns Nova Pack C18 using a mobile phase of water-acetonitrile-triethylamine (700:300:4, v/v/v) (adjusted to pH 7.5 with orthophosphoric acid). The method was shown to be rapid and reliable with a lower limit of detection of 5 ng ml-1. Results are reported of simple experiments on the effects of temperature and light on the stability of flunitrazepam in plasma kept on the laboratory bench.

Column-switching high-performance liquid chromatographic determination of vancomycin in serum.

Concentrations of vancomycin in serum were measured by an automatic high-performance liquid chromatographic (HPLC) micromethod. Vancomycin is a glycopeptide antibiotic with broad application in the therapy of gram-positive infections. As this drug is potentially nephro- and ototoxic, a method to maximize its therapeutic benefit while minimizing the risk of toxicity is desirable. This fully automated HPLC method did not involve a sample pretreatment step. The configuration of the apparatus permitted a solid phase extraction of the serum sample on two precolumns filled with a reversed-phase material, followed by a chromatographic separation of the sample constituents on an analytical column. The reversed phase analytical column (muBondapak C18) was flushed with a mobile phase of water-acetonitrile-triethylamine, 870: 130: 4 (vol/vol/vol); the pH was adjusted to 3.0 with orthophosphoric acid. Precision was expressed as the coefficient of variation (CV), which was always < or = 4.13% for intra- and inter-assays (n = 10) in the range 2-50 micrograms/ml. We compared this specific HPLC determination to an enzyme-multiplied immunoassay (EMIT). Fifty clinical samples obtained from patients under vancomycin therapy were assayed by each method and results compared using a linear regression analysis. There was a significant correlation between results from HPLC and EMIT: EMIT = 0.51 + 1 x HPLC (r = 0.963; p < 0.0001). The rapidity and specificity of this HPLC micromethod make it suitable for use in the monitoring of serum levels of vancomycin and for use in pharmacokinetic studies of this antibiotic.

[Side-effects during treatment of rheumatoid arthritis with methotrexate]. / Effets indésirables observés au cours du traitement de la polyarthrite rhumatoïde par le méthotrexate.

Methotrexate is the drug with the highest long-term continuation rate in rheumatoid arthritis patients. However, toxicity is the main reason for methotrexate withdrawal. Most adverse effects are mild abnormalities, such as digestive symptoms, stomatitis, elevations in transaminase levels, and moderate decreases in peripheral blood cell counts. Potentially life-threatening effects include hypersensitivity pneumonitis and pancytopenia. Cirrhosis is less common than in patients with psoriasis. Opportunistic infections and Epstein-Barr virus-related lymphomas have been reported. Neurological disorders, cutaneous reactions and renal lesions have been ascribed to low-dose methotrexate. Prior renal dysfunction and concomitant administration of a number of drugs, including cotrimoxazole, have been shown to increase methotrexate toxicity. However, susceptibility to the toxic effects of methotrexate varies widely across individuals. The effectiveness of folate supplementation in preventing methotrexate toxicity remains controversial.

Cefpiramide kinetics and plasma protein binding in cholestasis.

Cefpiramide is a new parenteral cephalosporin mainly excreted in the bile. Eight patients with cholestasis and 11 healthy subjects received a single 1 g i.v. dose. Cefpiramide concentrations in plasma and urine were measured by h.p.l.c. and plasma binding was determined by ultrafiltration. Total clearance of cefpiramide (mean +/- s.d.) was 15.5 +/- 7.1 ml min-1 in patients and 25.6 +/- 4.6 ml min-1 in healthy subjects. As a result, the terminal elimination half-life was longer in patients (12.0 +/- 2.9 h vs 5.3 +/- 0.9 h). Owing to impaired biliary elimination of cefpiramide in cholestasis, the urinary recovery of unchanged drug in patients was about five times greater than in healthy subjects (85.1 +/- 10.3% vs 16.2 +/- 3.9%). Plasma binding was significantly lower in cholestasis (fu = 0.23 +/- 0.13 vs 0.02 +/- 0.004 in healthy subjects). Accordingly, the dosage regimen of cefpiramide should be modified in patients with cholestasis.