LC-MS-MS Method for the Determination of Antidepressants and Benzodiazepines in Meconium.

An LC-MS-MS method for the determination of 14 benzodiazepines (BZDs) (alprazolam, α-hydroxyalprazolam, clonazepam, bromazepam, diazepam, nordiazepam, lorazepam, lormetazepam, oxazepam, flunitrazepam, 7-aminoflunitrazepam, triazolam, midazolam and zolpidem) and 15 antidepressants (ADs) (amitriptyline, nortriptyline, imipramine, desipramine, clomipramine, norclomipramine, fluoxetine, norfluoxetine, sertraline, norsertraline, paroxetine, venlafaxine, desmethylvenlafaxine, citalopram and desmethylcitalopram) in meconium was developed and validated. Meconium samples (0.25 ± 0.02 g) were homogenized in methanol and subjected to mixed-mode cation exchange solid-phase extraction. Chromatographic separation was performed in reversed phase, with a gradient of 0.1% formic acid in 2 mM ammonium formate and acetonitrile. Two different chromatographic gradient methods were employed, one for the separation of ADs and another for BZDs. Analytes were monitored by tandem mass spectrometry employing electrospray positive mode in MRM mode (2 transitions per compound). Method validation included: linearity [n = 5, limit of quantification (LOQ) to 400 ng/g], limits of detection (n = 6, 1-20 ng/g), LOQ (n = 9, 5-20 ng/g), selectivity (no endogenous or exogenous interferences), accuracy (n = 15, 90.6-111.5%), imprecision (n = 15, 0-14.6%), matrix effect (n = 10, -73 to 194.9%), extraction efficiency (n = 6, 35.9-91.2%), process efficiency (n = 6, 20.1-188.2%), stability 72 h in the autosampler (n = 3, -8.5 to 9%) and freeze/thaw stability (n = 3, -1.2 to -47%). The method was applied to four meconium specimens, which were analyzed with and without hydrolysis (enzymatic and alkaline). The authentic meconium samples tested positive for alprazolam, α-hydroxyalprazolam, clonazepam, diazepam, nordiazepam, fluoxetine, norfluoxetine, clomipramine and norclomipramine. Therefore, the present LC-MS-MS method allows a high throughput determination of the most common BZDs and ADs in meconium, which could be useful in clinical and forensic settings.

LC-MS/MS method for the determination of nine antidepressants and some of their main metabolites in oral fluid and plasma. Study of correlation between venlafaxine concentrations in both matrices.

In this paper, a fast, sensitive and selective LC-MS/MS method is described for the simultaneous determination of amitriptyline, imipramine, clomipramine, fluoxetine, paroxetine, sertraline, fluvoxamine, citalopram and venlafaxine, as well as some of their main metabolites (nortriptyline, desipramine, norclomipramine and norfluoxetine), in oral fluid and plasma. The sample (0.2 mL) was extracted with an automated solid-phase extraction system (ASPEC XL), using mixed mode OASIS MCX cartridges. Chromatographic separation was performed in a Sunfire C18 IS column (20 mmx2.1 mm, 3.5 microm), using a gradient of acetonitrile and ammonium formate (pH 3; 2 mM) as mobile phase, which allowed the elution of all the compounds in less than 5 min. The method has been fully validated in both specimens. This method was initially applied to the analysis of oral fluid and plasma samples from patients on antidepressant treatment in order to assess for which compounds it was likely to find a good correlation between both matrices. The best results were obtained for venlafaxine, so the study was extended for this compound, comparing the ratio between oral fluid and plasma concentrations (ROF/PL) in five patients on venlafaxine treatment when both samples were collected simultaneously on four different occasions. An important inter and intraindividual variability was found in oral fluid concentrations for 150 mg dose (mean=287.5 ng/m, range 58.8-531.2 ng/mL) and for 75 mg dose (mean=186.3 ng/mL, range=82.1-289.2 ng/mL). R(OF/PL) was calculated for each patient on the four different occasions, showing also a high variability (CV=24.2-69.6%).

[Shigella spp. infections in Getafe Hospital between 2001 and 2006]. / Infección por Shigella spp. en el Hospital de Getafe entre 2001 y 2006.

INTRODUCTION: Shigella spp. is a bacterium that frequently causes diarrhoea in underdeveloped countries, but not so much in Spain, where it has been published that its incidence has decreased in the last few years. OBJECTIVE: The aim of this study has been to asses changes in the epidemiology of Shigella spp. infection over the last 5 years in children of Getafe, as well as the subspecies of Shigella spp. that cause illness in this area (area 10, Madrid), the need for hospitalisation, the complications and the treatment. METHOD: A retrospective study was carried out, based on a review of the medical charts of children less than fifteen years old and had a diagnosis of Shigella spp. infection during 2000-2006. They were identified through the Getafe Hospital Microbiology Service. RESULTS: Nineteen patients were identified, of which only four needed to be hospitalised. There were no differences between males and females. The Shigella spp. subspecies identified were S. sonnei and S. flexneri. The most frequent symptoms were vomiting, abdominal pain, diarrhoea, and fever. Only one had dehydration as a complication. The treatment was symptomatic, except in the four hospitalised patients, who required fluid therapy and antibiotics. The outcome was good in all patients. CONCLUSION: In this study, there appears to be an increase in the Shigella spp. incidence over the last few years in Area 10 of Madrid. Due to the fact that Shigella spp. is not a frequent bacterium in Spain, there are not enough studies on it, therefore it would be of interest to carry out prospective studies, in order to confirm this increase in incidence.

Infección por Shigella spp. en el Hospital de Getafe entre 2001 y 2006 / Shigella spp. infections in Getafe Hospital between 2001 and 2006

Introducción: Shigella spp. es un patógeno que produce gastroenteritis aguda de forma frecuente en países en desarrollo, pero no tan habitual en España, donde se ha publicado que está disminuyendo su incidencia en los últimos años Objetivo: El objetivo de este estudio fue determinar cambios en la epidemiología de Shigella spp. en los últimos 5 años en la población infantil de Getafe, así como el tipo de Shigella spp. que produce enfermedad en el área 10 de Madrid (área que depende de nuestro hospital), la necesidad de ingreso, sus complicaciones y tratamiento. Método: Se llevó a cabo un estudio descriptivo, retrospectivo, basado en revisión de historias clínicas de los pacientes, menores de 15 años, afectados por shigellosis entre los años 2001 y 2006, que fueron identificados a partir del Servicio de Microbiología del Hospital Universitario de Getafe. Resultados: Se identificaron 19 pacientes, de los cuales sólo 4 precisaron ingreso. Las subespecies de Shigella spp. identificadas fueron S. sonnei y S. flexneri. La clínica más frecuente fue diarrea, que se acompañaba de vómitos, dolor abdominal tipo cólico y fiebre elevada. Sólo uno de los pacientes presentó deshidratación hiponatrémica como complicación. El tratamiento fue sintomático, salvo en los pacientes ingresados en los que se pautó fluidoterapia intravenosa y antibioterapia, con resolución de la enfermedad. Conclusión: En el estudio realizado parece existir un aumento de la incidencia de Shigella spp. en los últimos años en el área 10 de Madrid. Al no ser un patógeno frecuente en España, no existen muchos estudios, con lo que sería interesante realizar estudios prospectivos para confirmar este aumento de incidencia (AU)

Introduction: Shigella spp. is a bacterium that frequently causes diarrhoea in underdeveloped countries, but not so much in Spain, where it has been published that its incidence has decreased in the last few years. Objective: The aim of this study has been to asses changes in the epidemiology of Shigella spp. infection over the last 5 years in children of Getafe, as well as the subspecies of Shigella spp. that cause illness in this area (area 10, Madrid), the need for hospitalisation, the complications and the treatment. Method: A retrospective study was carried out, based on a review of the medical charts of children less than fifteen years old and had a diagnosis of Shigella spp. infection during 2000-2006. They were identified through the Getafe Hospital Microbiology Service. Results: Nineteen patients were identified, of which only four needed to be hospitalised. There were no differences between males and females. The Shigella spp. subspecies identified were S. sonnei and S. flexneri. The most frequent symptoms were vomiting, abdominal pain, diarrhoea, and fever. Only one had dehydration as a complication. The treatment was symptomatic, except in the four hospitalised patients, who required fluid therapy and antibiotics. The outcome was good in all patients. Conclusion: In this study, there appears to be an increase in the Shigella spp. incidence over the last few years in Area 10 of Madrid. Due to the fact that Shigella spp. is not a frequent bacterium in Spain, there are not enough studies on it, therefore it would be of interest to carry out prospective studies, in order to confirm this increase in incidence (AU)

Recurrent tuberculosis from 1992 to 2004 in a metropolitan area.

The proportion of recurrent tuberculosis (TB) cases caused by re-infection has varied widely in previous studies. The aim of the present study was to determine the relative frequency of relapse and exogenous re-infection in patients with second episodes of TB, using DNA fingerprinting. A population-based retrospective longitudinal descriptive study was conducted in Madrid (Spain) during 1992-2004. The study consisted of 645 patients with culture-confirmed TB. Of these, 20 (3.1%) were retained because they presented with a second isolate of Mycobacterium tuberculosis. Finally, 12 of these cases were excluded because they did not complete the full treatment prescribed. All strains were typed by restriction fragment length polymorphism analysis and some by mycobacterial interspersed repetitive unit-variable number of tandem repeats analysis. The patients with recurrent TB were compared with those without recurrent TB. For seven out of the eight patients, the restriction fragment length polymorphism patterns of the Mycobacterium tuberculosis strains from the episodes of recurrent disease showed identical initial and final genotypes, indicating relapse; the remaining recurrent case showed different genotypes, suggesting exogenous re-infection. Re-infection is possible among people in developed countries, but the rates are lower than those occurring in high-risk areas. The risk factors for recurrent tuberculosis should be taken into account in the follow-up of treatment and tuberculosis control strategies.

Ten-year population-based molecular epidemiological study of tuberculosis transmission in the metropolitan area of Madrid, Spain.

SETTING: The epidemiology of tuberculosis (TB) in urban populations is changing. Combining conventional epidemiological techniques with DNA fingerprinting of Mycobacterium tuberculosis can improve our understanding of how TB is transmitted. OBJECTIVE: To improve the definition of molecular epidemiology of TB over 10 years in an area of Europe not previously studied. DESIGN: A population-based retrospective study was conducted in the Autonomous Community of Madrid, Spain, from 1992 to 1998; from 1999 to 2001, the study was prospective. The study population consisted of all patients for whom positive culture and full clinical and demographic data were available. All strains were typed by RFLP. Non-clustered patients were compared with clustered patients and studied using univariate analysis and a logistic regression model. RESULTS: Of 448 patients studied, 228 (50.7%) were clustered. Youth was the strongest risk factor associated with clustering. Pleural effusion was also found to be associated with clustering. An epidemiological link was found in only 85 (37.4%) of the 228 patients belonging to a cluster. CONCLUSION: Youth and pleural effusion were identified as risk factors for clustering. These findings may help adjust TB control and contact tracing strategies.

Liquid chromatography-electrospray ionisation mass spectrometry for the determination of nine selected benzodiazepines in human plasma and oral fluid.

A new simple and rapid liquid chromatographic-mass spectrometric technique was designed for the determination of nine benzodiazepines in plasma and oral fluid. Benzodiazepines were extracted from alkalinised spiked and clinical plasma and oral fluid samples using a single step, liquid-liquid extraction procedure with diethyl ether. The chromatographic separation was performed with a Xterra RP18, 5 microm (150 x 2.1 mm i.d.) reversed-phase column using deuterated analogues of the analytes as internal standard. The recovery ranged from 70.3 to 86.9% for plasma and 63.9 to 77.2% for oral fluid. The limits of detection ranged from 0.5 to 1 ng/ml in plasma and 0.1 to 0.2 ng/ml for oral fluid. The method was validated for all the compounds, including linearity and the main precision parameters. The procedure, showed to be sensitive and specific, was applied to real plasma and oral fluid samples. The method is especially useful to analyse saliva samples from drivers undergoing roadside drug controls.

Sumisión química: Antecedente, situación actual, y perspectivas. Protocolos de actuación para estudios multicéntricos / No disponible

No disponible

Development and validation of a method for the quantitation of Delta9tetrahydrocannabinol in oral fluid by liquid chromatography electrospray-mass-spectrometry.

Analysis of Delta(9)tetrahydrocannabinol (Delta(9)THC) and its metabolites in biological samples is of great relevance for forensic purposes. In the case of oral fluid (OF), the analysis should determine Delta(9)THC, whereas in urine, it detects the inactive metabolite tetrahydrocannabinol carboxylic acid (THC-COOH). Most laboratories analyze Delta(9)THC in such samples using GC-MS methods, but these procedures are time-consuming and involve unavoidable previous extraction and derivatization. No data is yet available on the application of liquid chromatography-mass-spectrometry to detect Delta(9)THC in oral fluid. We report a validation method in which the Delta(9)THC is isolated from oral fluid by a simple liquid-liquid extraction with hexane and subsequently analyzed by liquid chromatography-mass-spectrometry. The method here reported for the determination of Delta(9)THC in oral fluid only requires 200 microl of sample and achieves limits of detection of 2 ng/ml, and has been used to analyze oral fluid samples collected from current drug users.

Utilización y rendimiento de los hemocultivos en una unidad de cuidados intensivos medicoquirúrgica / Utilization and utility of blood cultures in a medical-surgical intensive care unit

Objetivo. Estimar la utilización y el rendimiento de los hemocultivos en una unidad de cuidados intensivos (UCI) medicoquirúrgica y evaluar los factores que pueden influir en su rendimiento.Diseño. Estudio prospectivo de cohortes.Ámbito. Unidad de cuidados intensivos medicoquirúrgica de hospital de tercer nivel.Intervenciones. Ninguna.Variables de interés. Tasas de incidencia acumulada de extracción de hemocultivos, porcentaje de hemocultivos positivos y factores asociados a la obtención de un hemocultivo positivo.Resultados. En los 2 períodos de estudio se extrajeron 953 hemocultivos en los 2.663 pacientes que ingresaron en la UCI. Se extrajeron a los 10 (15) días (mediana, 5 días; P25, 1; P75, 13) desde el ingreso en la UCI. La tasa de incidencia acumulada obtenida fue de 36 hemocultivos por 100 pacientes y de 55 hemocultivos por 1.000 pacientes/día. Los hemocultivos positivos fueron 155 (16 por ciento; intervalo de confianza [IC] del 95 por ciento, 1419), los hemocultivos contaminados fueron 57 (6 por ciento; IC del 95 por ciento, 5-8) y los indeterminados 22 (2 por ciento; IC del 95 por ciento, 1-3). Los factores asociados a obtener un hemocultivo positivo fueron: no estar recibiendo antibióticos (odds ratio [OR], 2,16), ni descontaminación digestiva selectiva (OR, 1,52) en el momento de extraer el hemocultivo y si el hemocultivo se obtiene cuando el paciente lleva ingresado en la UCI más de 2 semanas (OR, 2,65).Conclusiones. En nuestro estudio la tasa de extracción de hemocultivos fue menor a la descrita en enfermos críticos. El rendimiento de los hemocultivos se asoció con la administración de antibióticos sistémicos y tópicos y con el momento de su extracción (AU)

Intoxicaciones agudas en Santiago de Compostela, en un periodo de cuatro años / Acute poisoning in Santiago de Compostela during a four year period

El objetivo de este trabajo fue evaluar las características de un grupo de pacientes intoxicados en el área sanitaria de Santiago de Compostela. Para ello, se estudiaron 1192 intoxicaciones agudas atendidas en el Servicio de Urgencias y Unidad de Cuidados Intensivos del Hospital Clínico Universitario de esta ciudad, desde el 1 de enero de 1993 al 31 de diciembre de 1996. Se han considerado solo aquellos casos en los que se solicitó un análisis toxicológico a este Instituto de Medicina Legal, desconociéndose el número de casos excluidos. Se diseñó una ficha clínica con todos los parámetros estudiados para su cumplimentación por los médicos en el momento del ingreso. Se observó un predominio del sexo masculino (61,4 por ciento) y una edad media, en la población estudiada, de 32,3 ñ 12,5 años (rango: 1593 años). Los tóxicos más frecuentemente implicados fueron las drogas de abuso (681 casos), seguidas de los fármacos (545 casos) y de "otros agentes" (88 casos). Las intoxicaciones voluntarias estaban asociadas mayoritariamente con mujeres y con ingesta de fármacos, en contra de las accidentales, asociadas con hombres y con consumo de drogas de abuso. Existen antecedentes psiquiátricos en 332 casos, tratamiento farmacológico previo en 219 casos, alcoholismo en 189 casos y drogadicción por vía parenteral en 144 casos. Se ha constatado que un 76 por ciento de los pacientes presentaban alguna clínica al ingreso, con un predominio de las manifestaciones neurológicas (682 casos). Se utilizó algún tipo de tratamiento específico en 574 sujetos, destacando la aplicación de lavado gástrico (444 casos) y la administración de antídotos (358 casos). La mayoría de los pacientes evolucionó satisfactoriamente, con solo un 2 por ciento de mortalidad. Esta información permite conocer los patrones de abuso y la eficacia del tratamiento aplicado con el fin de mejorar el control del paciente intoxicado. (AU)

Pneumonia in patients with severe burns : a classification according to the concept of the carrier state.

OBJECTIVE: To establish baseline values of pneumonia incidence and mortality and to distinguish primary endogenous from secondary endogenous and exogenous pneumonias in a homogeneous patient population with severe burns. DESIGN: Cohort study. SETTING: A six-bed burn ICU. PATIENTS: All patients of > or = 14 years admitted to the ICU between January 1995 and June 1996 with a total body surface area burn of > or = 20%. INTERVENTION: Collection of data on surveillance samples from throat and rectum on admission and twice weekly afterward, and pneumonias during the ICU stay. MEASUREMENTS AND RESULTS: Fifty-six patients fulfilled the criteria of the study. Mean age was 43 +/- 19.8 years; total body surface area burn, 41 +/- 18.2%; the area of full-thickness burn was 24 +/- 17.7%. Forty-one patients required mechanical ventilation. Twenty-seven patients (48%) experienced 37 episodes of pneumonia. Twenty-one pneumonias were of primary endogenous development, ie, caused by potential pathogens carried in the admission flora. There were 14 secondary endogenous and 2 exogenous infections caused by microorganisms acquired on the burn unit. Inhalation injury was identified in 26 patients. The pneumonia rate was two times higher in the subset of patients with inhalation injury compared with the group of patients without inhalation injury (p < 0.001). Overall mortality was 25%. CONCLUSIONS: This study shows that pneumonia in burn patients is mainly an endogenous problem. Interventions that prevent the development of endogenous infections deserve prospective evaluation in patients with severe burns.