Reduced solvent and reagent amounts: effect on carbonyl dinitrophenylhydrazone measurements at low concentrations.

This work aims to advance towards a more affordable laboratory procedure for sample treatment to determine carbonyl compounds by derivatization with 2,4-dinitrophenylhydrazine (DNPH). The proposal is based on reducing the amount of DNPH and solvents. A simple addition of standard carbonyls in a solution containing DNPH to prepare hydrazone standards is described and evaluated. Tedious recrystallization steps are avoided. Formaldehyde, acetaldehyde, acetone, tolualdehyde and hexanal, as carbonyl models, were quantified using a DNPH concentration of 400 µg mL-1 and 3.8 mM H2SO4 and by keeping for 24 hours at room temperature. Analytical coefficients of variation between 10 and 25% were found from the analysis of blanks under intermediate conditions (two different devices, very different concentrations of DNPH and analysis on two days). From these values of relative standard deviations and background levels, quantification limits were estimated between 15 and 40 ng mL-1. The reduction of reagent amounts allows the operator to better control the background levels in the use of DNPH, as well as making the method more cost-effective and easy to use. In short, it leads to a more sustainable adaptation of the classical method. The versatility in analytical application was tested to estimate the levels of formaldehyde, acetaldehyde and acetone in very different types of environmental samples. In particular, outdoor and indoor samples were collected in filters and impregnated cartridges, respectively. Moreover, tars in 2-propanol and particulate matter from gasification processes were also tested.

Characterization of organic aerosol at a rural site influenced by olive waste biomass burning.

Biomass burning is a major air pollution problem all around the world. However, the identification and quantification of its contribution to ambient aerosol levels is a difficult task due to the generalized lack of observations of molecular markers. This paper presents the results of a yearlong study of organic constituents of the atmospheric aerosol at a rural site in southern Spain (Villanueva del Arzobispo, Jaén). Sampling was performed for PM10 and PM2.5, and a total of 116 and 115 samples, respectively, were collected and analyzed by GC/MS, quantifying 77 organic compounds. Higher levels of organic pollutants were recorded from November to March, coinciding with the cold season when domestic combustion is a common practice in rural areas. This jointly with adverse meteorological conditions, e.g. strong atmospheric stability, produced severe pollution episodes with high PMx ambient levels. High daily concentrations of tracers were reached, up to 26 ng m-3 for B(a)P and 6065 ng m-3 for levoglucosan in PM2.5, supporting that biomass burning is a major source of pollution at rural areas. A multivariate statistical study based on factor and cluster analysis, was applied to the data set with the aim to distinguish sources of organic compounds. The main resulting sources were related with biomass combustion, secondary organic aerosol (SOA), biogenic emissions, lubricating oil and soil organic components. A preliminary organic source profile for olive wastes burning was evaluated, based on cluster results, showing anhydrosacharides and xylitol are the main emitted compounds, accounting for more than 85% of the quantified compounds. Other source compounds were fatty acids, diacids, aliphatics, sugars, sugar alcohols, PAHs and quinones.

Organic Analysis of Environmental Samples Using Liquid Chromatography with Diode Array and Fluorescence Detectors: An Overview.

This overview is focused to provide an useful guide of the families of organic pollutants that can be determined by liquid chromatography operating in reverse phase and ultraviolet/fluorescence detection. Eight families have been classified as the main groups to be considered: carbonyls, carboxyls, aromatics, phenols, phthalates, isocyanates, pesticides and emerging. The references have been selected based on analytical methods used in the environmental field, including both the well-established procedures and those more recently developed.

Validating Analytical Protocols to Determine Selected Pesticides and PCBs Using Routine Samples.

This study aims at providing recommendations concerning the validation of analytical protocols by using routine samples. It is intended to provide a case-study on how to validate the analytical methods in different environmental matrices. In order to analyze the selected compounds (pesticides and polychlorinated biphenyls) in two different environmental matrices, the current work has performed and validated two analytical procedures by GC-MS. A description is given of the validation of the two protocols by the analysis of more than 30 samples of water and sediments collected along nine months. The present work also scopes the uncertainty associated with both analytical protocols. In detail, uncertainty of water sample was performed through a conventional approach. However, for the sediments matrices, the estimation of proportional/constant bias is also included due to its inhomogeneity. Results for the sediment matrix are reliable, showing a range 25-35% of analytical variability associated with intermediate conditions. The analytical methodology for the water matrix determines the selected compounds with acceptable recoveries and the combined uncertainty ranges between 20 and 30%. Analyzing routine samples is rarely applied to assess trueness of novel analytical methods and up to now this methodology was not focused on organochlorine compounds in environmental matrices.

Sequential Inoculation of Native Non-Saccharomyces and Saccharomyces cerevisiae Strains for Wine Making.

The use of non-Saccharomyces yeast for wine making is becoming a common trend in many innovative wineries. The application is normally aimed at increasing aromas, glycerol, reducing acidity, and other improvements. This manuscript focuses on the reproduction of the native microbiota from the vineyard in the inoculum. Thus, native selected yeasts (Hanseniaspora uvarum, Metschnikowia pulcherrima, Torulaspora delbrueckii, Starmerella bacillaris species and three different strains of Saccharomyces cerevisiae) were inoculated sequentially, or only S. cerevisiae (three native strains together or one commercial) was used. Inoculations were performed both in laboratory conditions with synthetic must (400 mL) as well as in industrial conditions (2000 kg of grapes) in red winemaking in two different varieties, Grenache and Carignan. The results showed that all the inoculated S. cerevisiae strains were found at the end of the vinifications, and when non-Saccharomyces yeasts were inoculated, they were found in appreciable populations at mid-fermentation. The final wines produced could be clearly differentiated by sensory analysis and were of similar quality, in terms of sensory analysis panelists' appreciation.

Exploring petroleum hydrocarbons in groundwater by double solid phase extraction coupled to gas chromatography-flame ionization detector.

This work proposes an analytical procedure for measuring aliphatic and aromatic hydrocarbons fractions present in groundwater. In this method, hydrocarbons are solid phase extracted (SPE) twice from the groundwater and the resulting fractions are analyzed by gas chromatography with flame ionization detection. The first SPE disposes the hydrocarbons present in groundwater in organic solvents and the second SPE divides them into aliphatic and aromatic hydrocarbons. The validation study is carried out and its uncertainties are discussed. Identifying the main sources of uncertainty is evaluated through applying the bottom-up approach. Limits of detection for hydrocarbons ranges are below 5 µg L(-1), precision is not above of 30%, and acceptable recoveries are reached for aliphatic and aromatic fractions studied. The uncertainty due to volume of the sample, factor of calibration and recovery are the highest contributions. The expanded uncertainty range from 13% to 26% for the aliphatic hydrocarbons ranges and from 14% to 23% for the aromatic hydrocarbons ranges. As application, the proposed method is satisfactorily applied to a set of groundwater samples collected in a polluted area where there is evidence to present a high degree of hydrocarbons. The results have shown the range of aliphatic hydrocarbons >C21-C35 is the most abundant, with values ranging from 215 µg L(-1) to 354 µg L(-1), which it is associated to a contamination due to diesel.

Harmonization in hemolysis detection and prevention. A working group of the Catalonian Health Institute (ICS) experience.

BACKGROUND: Hemolysis is the main cause of non-quality samples in clinical laboratories, producing the highest percentage of rejections in the external assurance programs of preanalytical quality. The objective was to: 1) study the agreement between the detection methods and quantification of hemolysis; 2) establish comparable hemolysis interference limits for a series of tests and analytical methods; and 3) study the preanalytical variables which most influence hemolysis production. METHODS: Different hemoglobin concentration standards were prepared using the reference method. Agreement was studied between automated methods [hemolytic indexes (HI)] and reference method, as well as the interference according to hemolysis degree in various biochemical tests was measured. Preanalytical variables which could influence hemolysis production were studied: type of extraction, type of tubes, transport time, temperature and centrifugation conditions. RESULTS: Good agreement was obtained between hemoglobin concentrations measured using the reference method and HI, for the most of studied analyzers, particularly those giving quantitative HI. The hemolysis interference cut-off points obtained for the majority of tests studied (except LDH, K) are dependent on the method/analyzer utilized. Furthermore, discrepancies have been observed between interference limits recommended by the manufacturer. The preanalytical variables which produce a lower percentage of hemolysis rejections were: centrifugation at the extraction site, the use of lower volume tubes and a transport time under 15 min at room temperature. CONCLUSIONS: The setting of interference limits (cut-off) for each used test/method, and the study of preanalytical variability will assist to the results harmonization for this quality indicator.

A chromatographic method to analyze products from photo-oxidation of anthropogenic and biogenic mixtures of volatile organic compounds in smog chambers.

A method for quantifying secondary organic aerosol compounds (SOA) and water soluble secondary organic aerosol compounds (WSOA) produced from photo-oxidation of complex mixtures of volatile organic compounds (VOCs) in smog chambers by gas chromatography/mass spectrometry (GC/MS) has been developed. This method employs a double extraction with water and methanol jointly to a double derivatization with N,O-bis (trimethylsilil) trifluoroacetamide (BSTFA) and O-(2,3,4,5,6)-pentafluorobenzyl-hydroxylamine hydrochloride (PFBHA) followed by an analysis performed by GC/MS. The analytical procedure complements other methodologies because it can analyze SOA and WSOA compounds simultaneously at trace levels. As application, the methodology was employed to quantify the organic composition of aerosols formed in a smog chamber as a result of photo-oxidation of two different mixtures of volatile organic compounds: an anthropogenic mixture and a biogenic mixture. The analytical method allowed us to quantify up to 17 SOA compounds at levels higher than 20 ng m(-3) with reasonable recovery and a precision below 11%. Values found for applicability, selectivity, linearity, precision, recovery, detection limit, quantification limit and sensitivity demonstrated that the methodology can be satisfactorily applied to quantify SOA and WSOA.

Retinol binding protein-4 circulating levels were higher in nonalcoholic fatty liver disease vs. histologically normal liver from morbidly obese women.

OBJECTIVE: We aimed to analyze the retinol binding protein-4 (RBP4) messenger RNA (mRNA) expression profiles in adipose tissues and liver of morbidly obese (MO) women with or without nonalcoholic fatty liver disease (NAFLD), and to study the relationships with other pro- and anti-inflammatory adipokines in vivo and in vitro. DESIGN AND METHODS: We performed a cross-sectional analysis of subcutaneous adipose tissue (SAT), visceral adipose tissue (VAT) and liver samples from four lean and 45 MO women with or without NAFLD by enzyme-linked immunosorbent assay and real-time reverse transcription-PCR. We also studied RBP4 expression in HepG2 hepatocytes under various inflammatory stimuli. RESULTS: Circulating RBP4 levels were higher in MO women, and specifically, in MO subjects with NAFLD compared with normal liver controls (lean and MO). RBP4 liver expression was higher in nonalcoholic steatohepatitis (NASH)-moderate/severe than in NASHmild. Overall RBP4 gene expression was higher in liver than in adipose tissues. Among them, the higher expression corresponded to SAT. VAT expression was lower in the MO cohort. In HepG2, RBP4 mRNA expression was reduced by tumor necrosis factor (TNF)-α and increased by adiponectin treatment. CONCLUSIONS: The results obtained in MO women with NAFLD, brings up the use of RBP4 and other adipokines as a panel of noninvasive molecular biomarkers when NAFLD is suspected. Further studies are needed with other obesity groups.

An evaluation of uncertainty associated to analytical measurements of selected polycyclic aromatic compounds in ambient air.

This paper presents an evaluation of uncertainty associated to analytical measurement of 18 polycyclic aromatic compounds (PACs) in ambient air by liquid chromatography with fluorescence detection (HPLC/FD). The study was focused on analyses of PM(10), PM(2.5) and gas phase fractions. Main analytical uncertainty was estimated for 11 polycyclic aromatic hydrocarbons (PAHs), four nitro-polycyclic aromatic hydrocarbons (nitro-PAHs) and two hydroxy-polycyclic aromatic hydrocarbons (OH-PAHs) based on the analytical determination, reference material analysis and extraction step. Main contributions reached 15-30% and came from extraction process of real ambient samples, being those for nitro-PAHs the highest (20-30%). Range and mean concentration of selected PACs measured in gas phase and PM(10)/PM(2.5) particle fractions during a full year are also presented. Concentrations of OH-PAHs were about 2-4 orders of magnitude lower than their parent PAHs and comparable to those sparsely reported in literature.

Estimation of measurement uncertainty of pesticides, polychlorinated biphenyls and polyaromatic hydrocarbons in sediments by using gas chromatography-mass spectrometry.

The evaluation of the uncertainty associated to analytical methods is essential in order to demonstrate quality of a result. However, there is often lack of information about uncertainty of methods to estimate persistent organic pollutants concentration in complex matrix. Current work has thoroughly evaluated uncertainty associated to quantification of several organochloride pesticides, PCBs and PAHs in sediments. A discussion of the main contributions to the overall uncertainty is reported, allowing authors to establish the accuracy of results and plan future improvements. Combined uncertainties ranged between 5-9% (pesticides), 4-7% (PCBs) and 5-10% (PAHs), being uncertainty derived of calibration the main contribution. Also, the analytical procedure was validated analysing a standard reference material (IAEA-408).

Sampling of tar from sewage sludge gasification using solid phase adsorption.

Sewage sludge is a residue from wastewater treatment plants which is considered to be harmful to the environment and all living organisms. Gasification technology is a potential source of renewable energy that converts the sewage sludge into gases that can be used to generate energy or as raw material in chemical synthesis processes. But tar produced during gasification is one of the problems for the implementation of the gasification technology. Tar can condense on pipes and filters and may cause blockage and corrosion in the engines and turbines. Consequently, to minimize tar content in syngas, the ability to quantify tar levels in process streams is essential. The aim of this work was to develop an accurate tar sampling and analysis methodology using solid phase adsorption (SPA) in order to apply it to tar sampling from sewage sludge gasification gases. Four types of commercial SPA cartridges have been tested to determine the most suitable one for the sampling of individual tar compounds in such streams. Afterwards, the capacity, breakthrough volume and sample stability of the Supelclean™ ENVI-Carb/NH(2), which is identified as the most suitable, have been determined. Basically, no significant influences from water, H(2)S or NH(3) were detected. The cartridge was used in sampling real samples, and comparable results were obtained with the present and traditional methods.

Assessment uncertainty associated to the analysis of tar from gasification of sewage sludge.

The uncertainty evaluation associated with the quantification of tar from gasification of sewage sludge is present. Each of the sources of uncertainty associated with the wet type sampling method and GC-MS analysis was identified to determine the critical stages of the analytical methodology in order to reduce them. The study shows that major contributions to the overall uncertainty are related to extraction steps. High expanded uncertainties were found for all compounds, due to the segregation of the tar in different samples because of the sampling method. However, the analytical method used was successfully applied for the evaluation of the tar cleaning filter in a real gasification plant.

Quality indicators and specifications for key analytical-extranalytical processes in the clinical laboratory. Five years' experience using the Six Sigma concept.

BACKGROUND: The results of 5 years of experience (2004-2008) with process-based quality management using quality indicators for key laboratory processes (analytic and extra-analytic) in a group of clinical laboratories affiliated with the Catalan Health Institute are presented. Our purpose was to analyze the evolution of the indicators, identify processes that require corrective measures, and obtain specifications that are more robust than the preliminary ones proposed in a previous study by the same group. METHODS: The yearly average was recorded for each indicator in each laboratory, the yearly interlaboratory median was calculated, and the changes occurring were studied to determine their continuity in the 5-year period. For each indicator, the average of the yearly medians was calculated and the results transformed to the Six Sigma scale to estimate the degree of control over the related process. It was suggested to establish the yearly interlaboratory median as the desirable specification for each indicator. RESULTS: The medians for most indicators were stable during the period studied. Thus, the specifications proposed in the first study were considered robust in these cases. The Six Sigma statistic provided added value in this study because it enabled detection of processes that should be improved, in which case the specifications proposed were considered provisional despite their stability. After identifying processes that have the greatest impact on patient safety, the group set a specification of 0%, regardless of the actual specification obtained, although the members are conscious of the difficulty in attaining this level of quality. Certain processes that are in a period of change obtained specifications that are considered in a phase of consolidation. CONCLUSIONS: The results for indicators related with sample collection indicate that the process is stable and well controlled. However, based on the results for Hemolyzed serum sample, the group saw the need for installing centrifuges in all phlebotomy centers and established a recommendation to unify the system for measuring hemolysis. The indicator External control exceeds acceptance limit clearly highlighted the need to rigorously monitor the analytic phase of the clinical laboratory. The values obtained for the indicator Reports from referred tests exceed delivery time show that there is considerable deviation regarding the expected report delivery time, whereas for in-house laboratory reports, delivery time is satisfactory.

Association of retinol-binding protein-4 (RBP4) with lipid parameters in obese women.

BACKGROUND: Although the adipokine retinol-binding protein-4 (RBP4) has been implicated in the development of obesity-related insulin resistance, its role in human obesity is still unclear. Our objectives were to find out the effect on RBP4 systemic levels of a weight loss induced by gastric bypass surgery and to analyze RBP4 relationships with insulin resistance, parameters of body composition, lipid metabolism, and inflammation. METHODS: Sixty-three obese women were analyzed before and 12 months after surgery of systemic concentrations of RBP4, fasting glucose, insulin, lipid profile molecules, and inflammation-related proteins (C-reactive protein, tumor necrosis factor-alpha receptors 1 and 2, interleukin-18, and adiponectin), and waist and hip circumference measurements, body mass index calculation, and insulin resistance index by homeostasis model assessment were also made. RESULTS: We found that RBP4 levels were lower after weight reduction by gastric bypass surgery (p < 0.0001). We found RBP4 associated with triglycerides before (beta = 0.37, p = 0.02) and after surgery (beta = 0.59, p < 0.0001) and negatively with weight loss after surgery (beta = -0.37, p = 0.003). When expressed as a percentage of change, the decrease of RBP4 was related to the reduction in the levels of triglycerides and with the increase in HDL-cholesterol (beta = 0.73, p = 0.02 and beta = 0.62, p = 0.04, respectively). Others parameters analyzed, including inflammatory markers, were not related to RBP4. CONCLUSIONS: This study shows that, in obese women and after a substantial weight loss due to bariatric surgery, RBP4 was related to weight status and lipid parameters rather than to insulin sensitivity or inflammatory markers.

Indicators and quality specifications for strategic and support processes related to the clinical laboratory: four years' experience.

BACKGROUND: Quality specifications for indicators of the key analytic processes have been defined by international consensus. However, only preliminary specifications for laboratory-related strategic and support processes have been developed. The present study attempts to increase the robustness of the preliminary proposed specifications. METHODS: Recovering records and incidences occurred over a 4-year follow-up period, for 12 indicators, used in all laboratories from this group regarding strategic and support processes. RESULTS AND CONCLUSIONS: The results obtained indicate that it is better to establish an interval rather than a fixed value for the majority of indicators. Longer studies are needed to properly assess some quality specifications, and data recording system must be standardized in others. Additional, multicenter studies are needed to establish more robust specifications and determine the state of the art of laboratories in other settings.

Assessment uncertainty associated to the analysis of organic compounds linked to particulate matter of atmospheric aerosols.

Current work has evaluated uncertainty associated to quantification of several organic compounds present in particulate matter of atmospheric aerosols, setting out the stages of analytical procedure that contribute most to the global uncertainty. Several sources of uncertainty have been identified, which were clustered into five main contributions: sampling, extraction, clean-up, derivatization and analysis. A discussion of the main contributions to the overall uncertainty is reported, allowing authors to locate the largest ones and plan future improvements. Combined uncertainties ranged between 10-18% (alkanes), 12-16% (PAHs), 10-18% (alcohols) and 9-21% (acids). The analytical procedure was validated by analysing a standard reference material (SRM1649a, urban dust). Also, the proposed method was applied to the analysis of four samples of particulate matter.

Large Evaluation of Anti-Cardiolipin and anti-Beta2Glycoprotein I Assays: Results from the AutoimmunityWorkshop of the Spanish Society of Immunology

Despite their clinical utility and the importance that laboratory testshave in APS diagnosis, probably the most important drawback of suchtests is the elevated intra- and inter-laboratory variation. The aim of thepresent work was to assess the multilaboratory performance of aCL (..) (AU)

A pesar de la indudable utilidad clínica y de la importancia de laspruebas de laboratorio en el diagnóstico del síndrome antifosfolípido(APS), probablemente el mayor defecto de dichas pruebas es su elevadavariabilidad intra- e inter-laboratorio. El objetivo del presente trabajo fueevaluar el comportamiento de los ensayos (..) (AU)

Parallel downregulation of retinol-binding protein-4 and adiponectin expression in subcutaneous adipose tissue of non-morbidly obese subjects.

CONTEXT AND OBJECTIVE: Adipokines are involved in the etiopathology of obesity-related disorders. Since the role of adipokine retinol-binding protein-4 (RBP4) in obesity remains uncertain and its relationship with other adipokines and inflammatory markers has not been examined in detail, we investigated the relationships of RBP4 mRNA expression and circulating protein levels with obesity, anthropometric and metabolic variables, as well as with obesity-related inflammatory markers adiponectin and C-reactive protein. SUBJECTS AND METHODS: One-hundred and twenty-five subjects participated, 36 lean (body mass index (BMI): <25 kg/m(2)) and 89 obese (overweight/obese; BMI: > or =25<40) whose anthropometric and metabolic variables were assessed. mRNA expression was quantified by real-time PCR in subcutaneous adipose tissue (s.c.-AT) of 46 subjects. RESULTS: There was a tendency for circulating RBP4 levels to positively correlate with waist circumference (beta=0.29, P=0.08; R(2)=0.08), but there was no significant association with the obesity-related parameters analysed. RBP4 and adiponectin mRNA expression levels were similarly downregulated in the s.c.-AT of obese subjects (0.5-fold); however, RBP4 downregulation did not affect its circulating protein levels. The expression of RBP4 and adiponectin was positively correlated even after controlling for confounding factors (beta=0.59, P<0.0001; R(2)=0.40). CONCLUSIONS: In our population, RBP4 circulating levels were not significantly correlated with obesity-related parameters, although a tendency to correlate with waist circumference suggests a relationship with insulin resistance and other metabolic disorders. In addition, our results suggest that the production of RBP4 by other tissues such as liver, rather than s.c.-AT, may be involved in regulating RBP4 circulating levels.

Quality indicators and specifications for strategic and support processes in laboratory medicine.

BACKGROUND: This work is the second part of a study regarding indicators and quality specifications for the non-analytical processes in laboratory medicine. Five primary care and five hospital laboratories agreed on the indicators for two strategic processes (quality planning and project development) and various support processes (client relationships, instrument and infrastructure maintenance, safety and risk prevention, purchases and storage, personnel training). METHODS: In the majority of cases, the median values recorded over 1 year is considered to be the state-of-the-art in our setting and proposed as the quality specification for the indicators stated. Values have been stratified according to primary care and hospital laboratory for referred tests and group of personnel for training. In some cases, the specifications have been set equal to zero events, such as serious incidents in the infrastructure maintenance process and number of work accidents in the safety and risk prevention process. RESULTS AND CONCLUSIONS: In light of this study, an effort is needed to optimize decisions regarding corrective actions and to move from a subjective individual criterion to systematic and comparative management. This preliminary study provides a comprehensive vision of a subject that could motivate further research and advances in the quality of laboratory services.