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Space is a hostile environment characterized by high vacuum, extreme temperatures, meteoroids, space debris, ionospheric plasma, microgravity and space radiation, which all represent risks for human health. A deep understanding of the biological consequences of exposure to the space environment is required to design efficient countermeasures to minimize their negative impact on human health. Recently, proteomic approaches have received a significant amount of attention in the effort to further study microgravity-induced physiological changes. In this review, we summarize the current knowledge about the effects of microgravity on microorganisms (in particular Cupriavidus metallidurans CH34, Bacillus cereus and Rhodospirillum rubrum S1H), plants (whole plants, organs, and cell cultures), mammalian cells (endothelial cells, bone cells, chondrocytes, muscle cells, thyroid cancer cells, immune system cells) and animals (invertebrates, vertebrates and mammals). Herein, we describe their proteome's response to microgravity, focusing on proteomic discoveries and their future potential applications in space research., Biological Significance: Space experiments and operational flight experience have identified detrimental effects on human health and performance because of exposure to weightlessness, even when currently available countermeasures are implemented. Many experimental tools and methods have been developed to study microgravity induced physiological changes. Recently, genomic and proteomic approaches have received a significant amount of attention. This review summarizes the recent research studies of the proteome response to microgravity inmicroorganisms, plants, mammalians cells and animals. Current proteomic tools allow large-scale, high-throughput analyses for the detection, identification, and functional investigation of all proteomes. Understanding gene and/or protein expression is the key to unlocking the mechanisms behind microgravity-induced problems and to finding effective countermeasures to spaceflight-induced alterations but also for the study of diseases on earth. Future perspectives are also highlighted., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Exposure assessment of health care workers to antineoplastic drugs (ADs) is still an open issue since new, critical, and emerging factors may put pharmacists who prepare hazardous drugs or nurses who administer anti cancer agents to an increased risk of developing adverse health effects. Overall, eight pharmacies and nine patient areas have been surveyed in this study. Wipe and pad samples were experienced during the surveillance program in four Italian health care settings. Urine samples were collected from workers handling ADs. Cyclophosphamide (CP), ifosfamide (IF), and gemcitabine (GEM) were detected in all the work environments by using a LC-MS/MS method-based capable of analysing all the three drugs simultaneously. In total, 54% of wipe samples were positive for at least one drug and 19% of pad samples were shown to be contaminated by cyclophosphamide. Pharmacies were generally more contaminated than patient areas with the exception of one site where a nurse had an acute exposure during the cleaning-up of an hazardous drug solution spill. In total, 22 urine samples collected from pharmacists and 78 urine samples from nurses had no detectable concentrations of any antineoplastic drugs. Despite the adherence to the recommended safety practices residue contamination on surfaces and floors has continued to be assessed in all the investigated sites., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

The use of antineoplastic agents for the treatment of cancer and other non-neoplastic diseases is an increasingly common practice in hospitals. As a result, workers involved with handling antineoplastic drugs may be accidentally exposed to these agents, placing them at potential risk for long term adverse effects. To date, the challenge of protecting workers' health is persisting and expanding, with an increasing number of publications demonstrating that contamination of antineoplastic drugs (ADs) is still present on work surfaces after cleaning procedures are concluded. In this paper, five workplaces were selected for surveillance of professional exposure to ADs. Hospital pharmacies involved in the study were set in the North (Units A1 and B2), Center (Units C3 and D4) and South (Unit E5) of Italy. Contamination levels on a number of work surfaces and trends over a 10-year period are presented. Environmental and biological levels were obtained by high performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (MS/MS). A strong reduction of surface contamination was evidenced since 2003, when the recommended procedures for the safe handling of antineoplastic drugs started to be followed by health care workers. Employers' adherence to these recommendations allowed risk characterisation to achieve other important goals. The percentage of positive urine samples was found to be around 30% in the 1990s and 2% in the 2000s. Moreover, no positive samples were detected in 2006 or 2007. In conclusion, our study emphasized that one helpful strategy to reduce risk to all potentially exposed workers is also provided by a data-storage system that allows potential risks of working to be rapidly identified and controlled., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Polychlorinated biphenyls (PCBs) and organochlorine pesticides (OCPs) have been widely used in industry and agriculture. Due to their persistence and bioaccumulation, they were globally spread in the environment and may still be found in environmental and biological media, despite the international restrictions on production and use. The main aim of our study was to develop a simple and fast method suitable for the establishment of the reference values for 15 PCB congeners and 16 OCPs in general population subgroups. A cost- and time-saving screening procedure using gas chromatography coupled with low-resolution mass spectrometry, was improved and validated before application to the analysis of real samples. The overall method was validated including uncertainty measurement. Preliminary field data were collected from 95 volunteers living in two Italian areas. HCB, p,p'-DDE, PCB 153, PCB 138 and PCB 180 were the most frequently detected compounds. Age and residence area were found to be significant variables for the most abundant compounds, while no correlation between serum concentrations and gender was observed. Our results suggest that long-banned substances, including PCBs and the pesticides HCB and DDT's breakdown product, are still detectable in the general population., (Copyright 2009 Elsevier Ireland Ltd. All rights reserved.)

Two analytical methods (HPLC-fluorimeter [HPLC-FLD] and tandem mass spectrometry LC/MS/MS) are available to assay phenyl-hydroxyethylmercapturic acids (PHEMAs), the mercapturic acids of styrene in humans. In the past, each method was used to check different populations of subjects, but until now no attempt has been made to compare the two methods. This study was designed to verify whether the two methods actually give comparable results. The influence of different conditions of sample storage in altering the concentration of PHEMAs was also investigated. Urine samples were collected at the beginning and at the end of the workshift from 10 workers exposed to different levels of styrene. Each sample was analysed both by LC/MS/MS after storage under different conditions (respectively, at -20 and +4 degrees C, and after repeated freezing-thawing cycles), and by HPLC-FLD (in the same conditions of storage). Strong correlations were found between the two methods both for total PHEMAs and for each of the isomers measured, including the minor (S,R)-M1. Also an alternative approach, the Bland-Altman test, confirmed the agreement between the two methods. The different storage conditions tested did not decrease the concentration of PHEMAs but, surprisingly, a clear trend to increase was shown, particularly for (R,R)-M1, (S,R)-M2 and (R,R)-M2 in samples stored at +4 degrees C for 1 week. In conclusion, the study demonstrates that the methods give comparable results. Indirectly, this confirms also the main characteristics of PHEMAs, showed in the previous experiments: low biotransformation rates of styrene into PHEMAs; large inter-individual variability; and the presence of a clear preference in the excretion of the isomers deriving from (S)-styrene oxide. PHEMAs appear stable under different storage conditions, but further studies are needed to explain the increase of levels that occurs when samples are not kept frozen. To avoid pre-analytical errors, samples collected for biomonitoring or research purposes should be frozen as soon as possible, and thawed only one time just before the analysis.

An LC-MS method is described for the determination of urinary sorbic acid (SA), a common food additive, which allows to measure down to 4 microg/L of the compound. The method involves an acidic hydrolysis followed by solid-phase extraction. The method was applied to two volunteers who ingested SA and to 36 individuals with no dietary restriction. The results confirm that a little aliquot of ingested SA is found in urine also in humans. The significant correlation found between urinary levels of SA and trans,trans-muconic acid (MA) seems to indicate that the measurement of SA in urine could allow to estimate the amount of MA excreted following a dietary intake of SA and, consequently, to enhance the specificity of MA as a biomarker of benzene exposure. A point of clarification in future studies will be the actual chemical form of SA excreted, since our results clearly demonstrate that without hydrolysis only a very little amount of SA can be found even in subjects heavily exposed to SA.

A new high-performance liquid chromatographic (HPLC) method is described for the determination of urinary N-acetyl-S-(N-methylcarbamoyl)cysteine (AMCC), the final product of the conjugation reaction between a metabolic intermediate of N,N-dimethylformamide (DMF) and glutathione. Urine samples were purified by C(18) solid-phase extraction and then directly analysed by HPLC with an Aminex Ion Exclusion HPX-87H column maintained at 25 degrees C and a UV detector set at 196 nm. Under isocratic conditions (2.4 mM sulphuric acid, flow-rate=0.6 ml/min) AMCC eluted at 20.2 min. The reproducibility (C.V.%) was 1.3-2.7% (intra- and inter-assay, N = 5); the accuracy was 98.0+/-1.7% at 10 mg/l and 101.9+/-1.5% at 800 mg/l (mean+/-SD, N = 3). AMCC was measured in urine from 22 exposed subjects. A strong correlation was found between AMCC and environmental DMF [AMCC (mg/g creatinine)=3.40xDMF (mg/m(3)) + 3.07; r=0.95], while in the urine of 20 unexposed subjects the concentration of AMCC was constantly below the detection limit of the method (0.9 mg/l in urine). The method described appears to be useful for the biological monitoring of DMF exposure., (Copyright 2002 Elsevier Science B.V.)

S-phenylmercapturic acid (S-PMA) was measured in urine from 145 subjects exposed to low benzene concentrations in the air (C(I), benzene). The 8-h, time-weighted exposure intensity of individual workers was monitored by means of charcoal tubes and subsequent gas-chromatographic analysis after desorption with CS2. S-PMA excretion level in urine was determined by high-performance liquid chromatography with fluorescence detection. The following linear correlation was found between S-PMA concentrations in urine and benzene concentrations in the breathing zone: log(S-PMA, microg/g creatinine) = 0.712 log (C(I)-benzene, ppm) + 1.644 (n = 145, r = 0.74, P < 0.001). The geometric mean (GSD) of S-PMA concentrations in urine from 45 subjects occupationally not exposed to benzene but smoking more than 20 cigarettes/day was 7.8 microg/g creatinine (2.11), the corresponding value among non-smokers being 1.0 microg/g creatinine (2.18). It is concluded that the urinary level of S-PMA can be regarded as a useful indicator of exposure to benzene.

Styrene (S) has been shown to be responsible for neurotoxic effects, including behavioural changes and neuroendocrine disturbances. The initial step of S metabolism is conversion to styrene 7,8-epoxide (SO), which is present in two enantiomeric forms [(R)(+)-SO and (S)(-)-SO]; this electrophilic intermediate is considered to be directly responsible for most toxic effects of S. The major urinary metabolites derived from the biotransformation of SO in man are mandelic acid (MA) and phenylglyoxylic acid (PGA). In rats an alternative pathway has been demonstrated, which involves the conjugation of SO to glutathione (GSH), leading to the excretion of two specific mercapturic acids, N-acetyl-S-(-(1-phenyl-2-hydroxyethyl)-cysteine [M1] and N-acetyl-S-(2-phenyl-2-hydroxy-ethyl)-cysteine [M2]; a close relationship has been found between exposure to S and urinary excretion of M1 and M2 in rats. As a consequence of the chiral nature of SO, both M1 and M2 consist of two diastereoisomers (M1-'R', M1-'S', M2-'R' and M2-'S'). Early reports have shown that the conversion of S to mercapturic acids is much lower in man (below 1% of the absorbed dose) than in rats (about 10%). We propose an analytical method for the determination of urinary M1 and M2 in man, which involves a urine clean-up by a chromatographic technique with a short reversed-phase pre-column; purified samples are then deacetylated with porcine acylase and deproteinized by centrifugal ultrafiltration. A derivatization is then performed with o-phthaldialdehyde and 2-mercaptoethanol and the fluorescent derivatives are separated on a reversed-phase analytical column. The mobile phase consists of acetate buffer and methanol mixed at variable proportions, the fluorescence detector is set at 330 nm (exc.) and 440 nm (em.). M1-'S' and M1-'R' are separated (retention times = 52.8 and 73.7 min, respectively) while the diastereoisomers of M2 coelute as a single peak at 70.5 min. The detection limit is about 7 micrograms/l, the coefficients of variation are below 7% and the error percentages are less than 6%. The method was applied to 25 urine samples from workers exposed to S: significant correlations were found between mercapturic acids and MA and PGA, the best correlation being between M2 and PGA (r = 0.79). Urine samples form unexposed subjects showed no detectable amounts of the analytes. A high stereoselectivity is shown by the enzymes involved in the metabolism of S to mercapturic acids: M1-'S', which derives from (S)-SO, is excreted in much higher amounts than M1-'R', which derives from (R)-SO.

'In the field' application of the measurement of urinary excretion of unmodified solvent for the biological monitoring of exposed workers has been investigated in many recent papers. The results obtained for several solvents are reviewed. The values of correlation coefficients (r) and regression lines obtained for benzene, toluene, xylene, styrene, n-hexane, cyclohexane, 2- and 3-methylpentane, methyl chloride, tetrachloroethylene, carbon tetrachloride, methyl chloroform, p-dichlorobenzene, nitrous oxide, halothane, isoflurane, enflurane, acetone, methyl ethyl ketone and methyl isobutyl ketone are presented. The correlations observed were generally good: r values range from 0.50-0.97, and the majority are between 0.84 and 0.90. The regression lines reported for the same solvent in different studies present some variability: this is possibly due to an inadequate control of factors influencing the relationship between external dose and absorption, such as differences in body burden, work load, individual characteristics, etc. These factors are discussed. As a whole, results reported in the literature show that measuring of urinary excretion of unmodified solvents provides a highly sensitive and specific exposure index, and can also be applied for the biological monitoring of occupational exposure to low levels of solvents or to solvent mixtures. Nevertheless, for an adequate assessment of biological limit values, further studies evaluating the reproducibility of regression lines are needed, given that the aspects influencing the correlation between external dose and urinary excretion are fully controlled. Another crucial aspect is the correlation with early effects: even though this has yet to be evaluated for several solvents, for others such as styrene and perchloroethylene a good correlation was obtained, further supporting the usefulness of the measurement of urinary excretion of solvent for the biological monitoring of occupational exposure.

A method for the determination of urinary N-acetyl-S-(1-phenyl-2-hydroxyethyl)-L-cysteine (M1) and N-acetyl-S-(2-phenyl-2-hydroxyethyl)-L-cysteine (M2) in man was developed. Clean-up of urine samples was obtained by a chromatographic technique, using a short reversed-phase precolumn; purified samples were then deacetylated with porcine acylase I for 16 h at 37 degrees C and deproteinized by centrifugal ultrafiltration. Derivatization was performed with o-phthaldialdehyde and 2-mercaptoethanol and the fluorescent derivatives were separated on a reversed-phase analytical column with a gradient mobile phase consisting of 50 mM acetate buffer (pH 6.5) and methanol. The retention times of the diastereoisomers of M1 (M1-"S" and M1-"R") were 52.8 and 73.7 min, respectively: M2 diastereoisomers eluted as a single peak at 70.5 min. The fluorescence detector was set at 330 nm (excitation) and 440 nm (emission). The detection limit (at a signal-to-noise ratio of three) was about 7 micrograms/1. The method was applied to 25 urine samples from workers exposed to styrene. A relationship was found between urinary mandelic and phenylglyoxylic acids and mercapturic acids specific for styrene. Urine samples from ten non-exposed subjects showed no detectable amounts of analytes.

A sensitive method for the determination of free and total urinary 2,5-hexandione (2,5-HD) using high-performance liquid chromatography with fluorescence detection was developed. After purification of urine with a disposable C18 cartridge, 2,5-HD was derivatized with dansylhydrazine; 1,3-diacetyl benzene (1,3-DAB) was added to the samples, as internal standard, prior to extraction. The resulting fluorescent adducts were separated on a reversed-phase column with a gradient mobile phase of 25 mM phosphate buffer (pH 6.4) and acetonitrile. The retention times of the 2,5-HD and 1,3-DAB derivatives were 9.4 and 13.7 min, respectively. The derivatives were detected by a fluorescence detector (excitation 340 nm, emission 525 nm). The mean recoveries of 2,5-HD and 1,3-DAB were 92.0 and 94.0%, respectively; the detection limit of 2,5-HD (signal-to-noise ratio of 3) was 5 micrograms/l in urine without hydrolysis and ca. 12 micrograms/l in hydrolyzed samples. The method was applied to 39 urine samples from workers exposed to n-hexane; the mean values were 2.597 mg/l (S.D. = +/- 0.758) for total 2,5-HD and 0.179 mg/l (S.D. = +/- 0.086) for free 2,5-HD. Urine samples of 22 non-exposed subjects showed a mean concentration of 0.437 mg/l (S.D. = +/- 0.109) and 0.022 mg/l (S.D. = +/- 0.011) for total and free 2,5-HD, respectively.

Carbon disulfide-induced neurobehavioral effects are well known and do not need further evidence. Carbon disulfide vasculopathy and the syndromic complex resulting in depression, loss of memory and concentration, and behavior disturbances have been widely demonstrated. Less known is the evolution of the symptomatology when the environmental conditions are consistently improved, that is, the reversibility or the progression of the dysfunctions observed. This paper reports on a neurobehavioral follow-up in a viscose rayon factory carried out, in intervals, from 1974 to 1990. Several successive improvements were implemented in the plant through the years, until finally, the most radical changes were made at the end of the Seventies and these resulted in exposure levels far below the current Threshold Limit Values. A total of 493 subjects were examined and some of them were reexamined up to six times. The last examination was completed in September, 1990. In this paper, studies by our group over the 15 years of monitoring are discussed. The results show that the general mental state, as measured by neurobehavioral methods, reflects past and current exposure. This point was explored by dividing the subjects into six groups on the basis of their length of exposure and year of examination and by comparing their performances. The results show that even exposure to levels of carbon disulfide not exceeding 8 mg/m3 may induce absentmindedness and difficulties in perceptive abilities.