Toenail zinc and risk of prostate cancer in the MCC-Spain case-control study.

BACKGROUND: Some researchers have suggested that zinc (Zn) could reduce the risk of prostate cancer (PC). However, research from observational studies on the relationship between PC risk and biomarkers of Zn exposure shows conflicting results. OBJECTIVES: To evaluate the association between toenail Zn and PC, considering tumour extension and aggressiveness, along with a gene-environment approach, exploring the interaction of individual genetic susceptibility to PC in the relationship between toenail Zn and PC. METHODS: In MCC-Spain study we invited all incident PC cases diagnosed in the study period (2008-2013) and recruited randomly selected general population controls. In this report we included 913 cases and 1198 controls with toenail Zn determined by inductively coupled plasma mass spectrometry. To measure individual genetic susceptibility, we constructed a polygenic risk score based on known PC-related single nucleotide polymorphisms. The association between toenail Zn and PC was explored with mixed logistic and multinomial regression models. RESULTS: Men with higher toenail Zn had higher risk of PC (OR quartile 4 vs.1: 1.41; 95% CI: 1.07-1.85). This association was slightly higher in high-grade PC [(ISUP≤2 Relative risk ratio (RRR) quartile 4 vs.1: 1.36; 1.01-1.83) vs. (ISUP3-5 RRR quartile 4 vs.1: 1.64; 1.06-2.54)] and in advanced tumours [(cT1-cT2a RRR quartile 4 vs.1: 1.40; 95% CI: 1.05-1.89) vs. (cT2b-cT4 RRR quartile 4 vs.1: 1.59; 1.00-2.53)]. Men with lower genetic susceptibility to PC were those at higher risk of PC associated with high toenail Zn (OR quartile 4 vs.1: 2.18; 95% CI: 1.08-4.40). DISCUSSION: High toenail Zn levels were related to a higher risk for PC, especially for more aggressive or advanced tumours. This effect was stronger among men with a lower genetic susceptibility to PC.

Untargeted Metabolomic Study of Lung Cancer Patients after Surgery with Curative Intent.

Lung cancer (LC) is a leading cause of mortality, claiming more than 1.8 million deaths per year worldwide. Surgery is one of the most effective treatments when the disease is in its early stages. The study of metabolic alterations after surgical intervention with curative intent could be used to assess the response to treatment or the detection of cancer recurrence. In this study, we have evaluated the metabolomic profile of serum samples (n = 110) from preoperative (PRE) and postoperative (POST) LC patients collected at two different time points (1 month, A; 3-6 months, B) with respect to healthy people. An untargeted metabolomic platform based on reversed phase (RP) and hydrophilic interaction chromatography (HILIC), using ultra-high performance liquid chromatography (UHPLC) and mass spectrometry (MS), was applied (MassIVE ID MSV000092213). Twenty-two altered metabolites were annotated by comparing all the different studied groups. DG(14,0/22:1), stearamide, proline, and E,e-carotene-3,3'-dione were found altered in PRE, and their levels returned to those of a baseline control group 3-6 months after surgery. Furthermore, 3-galactosyllactose levels remained altered after intervention in some patients. This study provides unique insights into the metabolic profiles of LC patients after surgery at two different time points by combining complementary analytical methods.

Metallomic Signatures of Lung Cancer and Chronic Obstructive Pulmonary Disease.

Lung cancer (LC) is the leading cause of cancer deaths, and chronic obstructive pulmonary disease (COPD) can increase LC risk. Metallomics may provide insights into both of these tobacco-related diseases and their shared etiology. We conducted an observational study of 191 human serum samples, including those of healthy controls, LC patients, COPD patients, and patients with both COPD and LC. We found 18 elements (V, Al, As, Mn, Co, Cu, Zn, Cd, Se, W, Mo, Sb, Pb, Tl, Cr, Mg, Ni, and U) in these samples. In addition, we evaluated the elemental profiles of COPD cases of varying severity. The ratios and associations between the elements were also studied as possible signatures of the diseases. COPD severity and LC have a significant impact on the elemental composition of human serum. The severity of COPD was found to reduce the serum concentrations of As, Cd, and Tl and increased the serum concentrations of Mn and Sb compared with healthy control samples, while LC was found to increase Al, As, Mn, and Pb concentrations. This study provides new insights into the effects of LC and COPD on the human serum elemental profile that will pave the way for the potential use of elements as biomarkers for diagnosis and prognosis. It also sheds light on the potential link between the two diseases, i.e., the evolution of COPD to LC.

Untargeted Gut Metabolomics to Delve the Interplay between Selenium Supplementation and Gut Microbiota.

Selenium (Se) is an essential trace element with important health roles due to the antioxidant properties of selenoproteins. To analyze the interplay between Se and gut microbiota, gut metabolomic profiles were determined in conventional (C) and microbiota depleted mice (Abx) after Se-supplementation (Abx-Se) by untargeted metabolomics, using an analytical multiplatform based on GC-MS and UHPLC-QTOF-MS (MassIVE ID MSV000087829). Gut microbiota profiling was performed by 16S rRNA gene amplicon sequencing. Significant differences in the levels of about 70% of the gut metabolites determined, including fatty acyls, glycerolipids, glycerophospholipids, and steroids, were found in Abx-Se compared to Abx, and only 30% were different between Abx-Se and C, suggesting an important effect of Se-supplementation on Abx mice metabolism. At genus level, the correlation analysis showed strong associations between metabolites and gut bacterial profiles. Likewise, higher abundance of Lactobacillus spp., a potentially beneficial genus enriched after Se-supplementation, was associated with higher levels of prenol lipids, phosphatidylglycerols (C-Se), steroids and diterpenoids (Abx-Se), and also with lower levels of fatty acids (Abx-Se). Thus, we observed a crucial interaction between Se intake-microbiota-metabolites, although further studies to clarify the specific mechanisms are needed. This is the first study about untargeted gut metabolomics after microbiota depletion and Se-supplementation.

Levels and determinants of urinary cadmium in general population in Spain: Metal-MCC-Spain study.

BACKGROUND: Cadmium is a ubiquitous and persistent metal, associated with different harmful health effects and with increased morbidity and mortality. Understanding the main sources of exposure is essential to identify at risk populations and to design public health interventions. OBJECTIVE: To evaluate cadmium exposure in a random-sample of general adult population from three regions of Spain, assessed by the urinary cadmium (U-Cd) concentration, and to identify its potential determinants and sex-specific differences, including sociodemographic, lifestyle and dietary factors. MATERIALS AND METHODS: We measured U-Cd (µg/g creatinine) in single urine spot samples from 1282 controls enrolled in the multicase-control study in common tumors in Spain (MCC-Spain) with inductively coupling plasma-mass spectrometry equipped with an octopole reaction systems (ICP-ORS-MS). The association between sociodemographic, lifestyle, and dietary characteristics and U-Cd concentrations was evaluated using geometric mean ratios (GMR) estimated by multiple log-linear regression models. RESULTS: Overall, geometric mean U-Cd concentration was 0.40 (95%CI: 0.38, 0.41) µg/g creatinine. Levels were higher in women than in men (GMR]: 1.19; 95%CI: 1.07, 1.32), and increased with age in males (ptrend< 0.001). Cigarette smoking was clearly associated to U-Cd levels (GMRformer vs non-smokers: 1.16; 95%CI: 1.05, 1.29; GMRcurrent vs non-smokers: 1.42; 95%CI: 1.26, 1.60); the relationship with secondhand tobacco exposure in non-smokers, was restricted to women (pinteraction = 0.02). Sampling season and region also seemed to influence U-Cd concentrations, with lower levels in summer (GMRsummer vs average: 0.79; 95%CI: 0.71, 0.88), and higher levels in North-Spain Asturias (GMRAsturias vs average: 1.13; 95%CI: 1.04, 1.23). Regarding diet, higher U-Cd concentration was associated with eggs consumption only in men (pinteraction = 0.04), just as rice intake was associated in women (pinteraction = 0.03). CONCLUSION: These results confirmed that tobacco exposure is the main modifiable predictor of U-Cd concentrations, and remark that the role of dietary/sociodemographic factors on U-Cd levels may differ by sex.

Toxic metals in toenails as biomarkers of exposure: A review.

Toenails have been used as biomarkers of exposure to toxic metals, but their validity for this purpose is not yet clear and might differ depending on the specific agent. To evaluate this issue, we reviewed the literature on: a) the time-window of exposure reflected by toenails; b) the reproducibility of toenail toxic-metal levels in repeated measures over time; c) their relationship with other biomarkers of exposure, and; d) their association with potential determinants (i.e. sociodemographic, anthropometric, or lifestyle characteristics) or with sources of exposure like diet or environmental pollution. Thus, we performed a systematic review, searching for articles that provided original data for levels of any of the following toxic metals in toenails: aluminum, beryllium, cadmium, chromium, mercury, nickel, lead, thallium and uranium. We identified 88 articles, reporting data from 67 different research projects, which were quite heterogeneous with regard to population profile, sample size and analytical technique. The most commonly studied metal was mercury. Concerning the time-window of exposure explored by toenails, some reports indicate that toenail cadmium, nickel and lead may reflect exposures that occurred 7-12 months before sampling. For repeated samples obtained 1-6 years apart, the range of intraindividual correlation coefficients of aluminum, chromium and mercury was 0.33-0.56. The correlation of toxic metal concentrations between toenails and other matrices was higher for hair and fingernails than for urine or blood. Mercury levels were consistently associated with fish intake, while other toxic metals were occasionally associated with specific sources (e.g. drinking water, place of residence, environmental pollution, and occupation). The most frequently evaluated health endpoints were cardiovascular diseases, cancer, and central nervous system diseases. Available data suggest that toenail mercury levels reflected long-term exposures and showed positive associations with fish intake. The lack of standardization in sample collection, quality control, analytical techniques and procedures - along with the heterogeneity and conflicting results among studies - mean it is still difficult to conclude that toenails are a good biomarker of exposure to toxic metals. Further studies are needed to draw solid conclusions about the suitability of toenails as biomarkers of exposure to toxic metals.

Insights into cancer and neurodegenerative diseases through selenoproteins and the connection with gut microbiota - current analytical methodologies.

Introduction: Selenium plays many key roles in health especially in connection with cancer and neurodegenerative diseases. However, it needs to be appreciated that the essentiality/toxicity of selenium depends on both, a narrow range of concentration and the chemical specie involved. In this context, selenoproteins are essential biomolecules against these disorders, mainly due to its antioxidant action. To this end, analytical methodologies may allow identifying and quantifying individual selenospecies in human biofluids and tissues. Areas covered: This review focus on the role of selenoproteins in medicine, with special emphasis in cancer and neurodegenerative diseases, considering the possible link with gut microbiota. In particular, this article reviews the analytical techniques and procedures recently developed for the absolute quantification of selenoproteins and selenometabolites in human biofluids and tissues. Expert commentary: The beneficial role of selenium in human health has been extensively studied and reviewed. However, several challenges remain unsolved as discussed in this article: (i) speciation of selenium (especially selenoproteins) in cancer and neurodegenerative disease patients; (ii) supplementation of selenium in humans using functional foods and nutraceuticals; (iii) the link between selenium and selenoproteins expression and the gut microbiota and (iv) analytical methods and pitfalls for the absolute quantification of selenoproteins and selenometabolites.

Metabolic Impairments Caused by a "Chemical Cocktail" of DDE and Selenium in Mice Using Direct Infusion Triple Quadrupole Time-of-Flight and Gas Chromatography-Mass Spectrometry.

Among organic contaminants, pesticides are one of the most important groups of chemicals due to their persistent character and toxicity. However, the biological systems are exposed to a complex environment in which the contaminants can interact in a synergistic/antagonistic fashion, and for this reason, the study of "chemical cocktails" is of great interest to fully understand the final biological effect. In this way, selenium is known for its antagonistic action against several toxicants. In this paper, metabolic impairments caused by the joint exposure of p,p'-dichloro diphenyl trichloroethane (DDE) and selenium (Se) have been issued for the first time. A metabolomic workflow was applied to mice fed DDE and DDE with Se diet, on the basis of the complementary use of two organic mass spectrometric techniques, combining direct infusion mass spectrometry (DI-ESI-QqQ-TOF MS) and gas chromatography-mass spectrometry (GC-MS). The results show a good classification between the studied groups caused by about 70 altered metabolites in the liver, kidney, or brain, including the pathways of energy metabolism, degradation of phospholipidic membrane, ß-oxidation, and oxidative stress, which confirm the potential of combined metabolomic platforms in environmental studies.

Toenails as biomarker of exposure to essential trace metals: A review.

Health problems associated with essential trace metals can result from both inadequate (i.e., low intake) and excessive exposures (i.e., from environmental and/or occupational source). Thus, measuring the exposure level is a real challenge for epidemiologists. Among non-invasive biomarkers that intend to measure long-term exposure to essential trace metals, the toenail is probably the biological matrix with the greatest potential. This systematic review collects the current evidence regarding the validity of toenail clippings as exposure biomarker for trace metals such as boron, cobalt, copper, iron, manganese, molybdenum, selenium, silicon, vanadium and zinc. Special attention was paid to the time-window of exposure reflected by the toenail, the intraindividual variability in exposure levels over time in this matrix, and the relationship of toenail with other biomarkers, personal characteristics and environmental sources. Our search identified 139 papers, with selenium and zinc being the most studied elements. The variability among studies suggests that toenail levels may reflect different degrees of exposure and probably correspond to exposures occurred 3-12 months before sampling (i.e., for manganese/selenium). Few studies assessed the reproducibility of results over time and, for samples obtained 1-6 years apart, the correlation coefficient were between 0.26 and 0.66. Trace metal levels in toenails did not correlate well with those in the blood and urine and showed low-moderate correlation with those in the hair and fingernails. Available data suggests that for some elements (Se, Mn, Zn) toenail concentrations reflect long-term external exposures in fairly reproducible levels, while for other metals, this association has not yet been assessed. Among dietary factors, only toenail selenium showed clear associations with the intake of supplements or specific foods. The toenail levels could also represent occupational exposure, for instance, Mn exposure in welders. The scarcity of information on other essential trace elements, together with the great heterogeneity among studies makes the validation of the usage of toenails as biomarkers of exposure to these elements difficult. Standardization of sample collection, quality control, analytical techniques and reporting procedures might facilitate further research focused on the clear understanding of the significance of essential levels in this promising matrix and would enhance its utility in epidemiological research.

Metabolomic study of bioactive compounds in strawberries preserved under controlled atmosphere based on GC-MS and DI-ESI-QqQ-TOF-MS.

INTRODUCTION: The storage of the vegetables products in a controlled atmosphere (CA) with low O2 and high CO2 concentrations, reduces respiration rates and delays the ripening process, and in some cases, improves their quality and organoleptic properties. OBJECTIVE: To obtain deep insight into strawberry fruit metabolic changes caused by these CA treatments. METHODOLOGY: Freshly harvested strawberries were preserved under different atmospheres enriched with 10%, 20% and 30% of CO2 , for 2 days at 0°C, containing in all the cases 5% of O2 and were subjected to a metabolomic analysis based on gas chromatography-mass spectrometry (GC-MS) and direct-infusion with electrospray ionisation source equipped with triple quadrupole coupled to time of flight mass spectrometry (DI-ESI-QqQ-TOF-MS). Partial least square discriminant analysis (PLS-DA) was employed to compare the control and treated samples for the identification of altered metabolites. RESULTS: Several metabolites related to CA treatment could be identified by databases and literature, which are mainly sugars, organic acids and phenolic compounds (bioactive compounds). CONCLUSIONS: Good correlation coefficients were obtained between discriminant metabolites and fruit quality parameters. These results suggest that treated strawberries under CA could be considered as bioactive healthy compounds, suggesting that treated strawberries under CA could be used as raw material for the preparation and formulation of food supplements and nutraceutical products.

The Metallome of Lung Cancer and its Potential Use as Biomarker.

Carcinogenesis is a very complex process in which metals have been found to be critically involved. In this sense, a disturbed redox status and metal dyshomeostasis take place during the onset and progression of cancer, and it is well-known that trace elements participate in the activation or inhibition of enzymatic reactions and metalloproteins, in which they usually participate as cofactors. Until now, the role of metals in cancer have been studied as an effect, establishing that cancer onset and progression affects the disturbance of the natural chemical form of the essential elements in the metabolism. However, it has also been studied as a cause, giving insights related to the high exposure of metals giving a place to the carcinogenic process. On the other hand, the chemical species of the metal or metallobiomolecule is very important, since it finally affects the biological activity or the toxicological potential of the element and their mobility across different biological compartments. Moreover, the importance of metal homeostasis and metals interactions in biology has also been demonstrated, and the ratios between some elements were found to be different in cancer patients; however, the interplay of elements is rarely reported. This review focuses on the critical role of metals in lung cancer, which is one of the most insidious forms of cancer, with special attention to the analytical approaches and pitfalls to extract metals and their species from tissues and biofluids, determining the ratios of metals, obtaining classification profiles, and finally defining the metallome of lung cancer.

Combination of HPLC with organic and inorganic mass spectrometry to study the metabolic response of the clam Scrobicularia plana to arsenic exposure.

Arsenic is a toxic element extensively studied in the marine environment due to differential toxicological effects of inorganic and organic species. In the present work, the bivalve Scrobicularia plana was exposed to AsV (10 and 100 µg/L) for 14 days to evaluate the metabolic perturbations caused by this element. Arsenic speciation and metabolomic analysis were performed in the digestive gland of the bivalve using two complementary analytical platforms based on inorganic and organic mass spectrometry. It has been observed the greater presence of the innocuous specie arsenobetaine produced in this organism as defense mechanism against arsenic toxicity, although significant concentrations of methylated and inorganic arsenic were also present, depending on the level of arsenic in aqueous media. Complementarily, a metabolomic study based on mass spectrometry and statistical discriminant analysis allows a good classification of samples associated to low and high As(V) exposure in relation to controls. About 15 metabolites suffer significant changes of expression by the presence of As(V): amino acids, nucleotides, energy-related metabolites, free fatty acids, phospholipids and triacylglycerides, which can be related to membrane structural and functional damage. In addition, perturbation of the methylation cycle, associated with the increase of homocysteine and methionine was observed, which enhance the methylation of toxic inorganic arsenic to less toxic dimethylarsenic.

Environmental Metallomics.

Metallomics is the new paradigm about the metallobiomolecules related to living organisms, considering the interactions between toxic and essential metals, transport through biological fluids, passing across biological membranes and interfaces, synergic and antagonist actions among metal species, and alterations in metabolic pathways triggered by overexpression or inhibition of these metallobiomolecules. These challenging studies require the development of new analytical approaches in order to get suitable information of these species close to their native environment which has promoted the application of new tools based in mass spectrometry under the double focus of elemental (ICP-MS) and molecular (Qq-TOF-MS) mass spectrometry, generally arranged with chromatography in multidimensional platforms. The driving force for the design of these new analytical instrumental arrangements is the analyst imagination who adapts the new metallomic methodology to the new problems. In this work the most recent metallomic approaches proposed have been considered, deepening their application to the most frequent problems related to metal toxicity in environmental issues, such as exposure experiments of mice to toxic metals, interactions and homeostasis of metals, metal imaging, metabolic alterations caused by metallobiomolecules over- or down-expressed, and more interestingly real-life consequences of metal species expression in environmental field studies. In this way, the application of two-dimensional chromatographic approaches with ICP-MS detection, the use of multidimensional chromatography-column-switching-ICP-MS devices, metal imaging with LA-ICP-MS, combined application of metallomics and metabolomics for environmental toxicological appraisal, and the application of these metallomic techniques in environmental field studies have been reviewed.

Selenium, selenoproteins and selenometabolites in mothers and babies at the time of birth.

The deficiency of Se, an essential micronutrient, has been implicated in adverse pregnancy outcomes. Our study was designed to determine total serum Se, selenoproteins (extracellular glutathione peroxidase (GPx-3), selenoprotein P (SeP)), selenoalbumin (SeAlb) and selenometabolites in healthy women and their newborns at delivery. This cross-sectional study included eighty-three healthy mother-baby couples. Total Se and Se species concentrations were measured in maternal and umbilical cord sera by an in-series coupling of two-dimensional size-exclusion and affinity HPLC. Additional measurements of serum SeP concentration and of serum GPx-3 enzyme activity were carried out using ELISA. Total Se concentration was significantly higher in maternal serum than in cord serum (68·9 (sd 15·2) and 56·1 (sd 14·6) µg/l, respectively; P<0·01). There were significant correlations between selenoprotein and SeAlb concentrations in mothers and newborns, although they also showed significant differences in GPx-3 (11·2 (sd 3·7) v. 10·5 (sd 3·5) µg/l; P<0·01), SeP (42·5 (sd 9·5) v. 28·1 (sd 7·7) µg/l; P<0·01) and SeAlb (11·6 (sd 3·6) v. 14·1 (sd 4·3) µg/l; P<0·01) concentrations in maternal and cord sera, respectively. Serum GPx-3 activity and concentration were positively correlated in mothers (r 0·33; P=0·038) but not in newborns. GPx-3 activity in cord serum was significantly correlated with gestational age (r 0·44; P=0·009). SeAlb concentration was significantly higher in babies, whereas SeP and GPx-3 concentrations were significantly higher in mothers. The differences cannot be explained by simple diffusion; specific transfer mechanisms are probably involved. GPx-3 concentrations in mothers, at delivery, are related to maternal Se status, whereas the GPx-3 activity in cord serum depends on gestational age.

Region-specific metabolic alterations in the brain of the APP/PS1 transgenic mice of Alzheimer's disease.

Alzheimer's disease (AD) is the most common neurodegenerative disorder worldwide, but its etiology is still not completely understood. The identification of underlying pathological mechanisms is becoming increasingly important for the discovery of biomarkers and therapies, for which metabolomics presents a great potential. In this work, we studied metabolic alterations in different brain regions of the APP/PS1 mice by using a high-throughput metabolomic approach based on the combination of gas chromatography-mass spectrometry and ultra-high performance liquid chromatography-mass spectrometry. Multivariate statistics showed that metabolomic perturbations are widespread, affecting mainly the hippocampus and the cortex, but are also present in regions not primarily associated with AD such as the striatum, cerebellum and olfactory bulbs. Multiple metabolic pathways could be linked to the development of AD-type disorders in this mouse model, including abnormal purine metabolism, bioenergetic failures, dyshomeostasis of amino acids and disturbances in membrane lipids, among others. Interestingly, region-specific alterations were observed for some of the potential markers identified, associated with abnormal fatty acid composition of phospholipids and sphingomyelins, or differential regulation of neurotransmitter amino acids (e.g. glutamate, glycine, serine, N-acetyl-aspartate), not previously described to our knowledge. Therefore, these findings could provide a new insight into brain pathology in Alzheimer's disease.

Metabolomics reveals significant impairments in the immune system of the APP/PS1 transgenic mice of Alzheimer's disease.

Inflammatory processes and other failures related to the immune system are common features associated with Alzheimer's disease (AD), in both brain and the peripheral system. Thus, the study of the main organs of the immune system may have a great potential for the elucidation of pathological mechanisms underlying these abnormalities. This is the first metabolomic investigation performed in spleen and thymus from transgenic mice of AD. Tissues were fingerprinted using a metabolomic platform comprising GC-MS and ultra-HPLC-MS. Multivariate statistics demonstrated significant differences in numerous metabolites between the APP/PS1 mice and wild-type controls, and it was proven that multiple biochemical pathways are disturbed in these organs including abnormal metabolism of phospholipids, energy deficiencies, altered homeostasis of amino acids, oxidative stress, and others. Therefore, these findings highlight the importance of the proper metabolic functioning of peripheral immune system in the development of neurodegenerative disorders such as AD.

Environmental metabolomics: Biological markers for metal toxicity.

Environmental metabolomics is an emerging field referred to the application of metabolomics to characterize the interactions of living organisms with their environment. In this sense, the importance of monitoring the effects of toxic metals on living organisms has increased as a consequence of natural changes and anthropogenic activities that have led to an increase of toxic metal levels in terrestrial and aquatic ecosystems. For this purpose, the use of metabolomics based on MS to study metal toxicity is gaining importance in recent years. Environmental metabolomics can be used to: discover the mode of action (MOA) of toxic metals through controlled laboratory experiments; evaluate toxicity (biological adverse response to a substance), that may be useful in risk assessment; and develop new biomarkers (based in metabolome shifts discovered through controlled laboratory experiments) that may be applied in environmental biomonitoring (environmental realistic scenario). In this review, it is discussed how metabolomics based on MS can be applied to study metal toxicity, considering the most important hallmarks related to metabolomic experiments.

High throughput multiorgan metabolomics in the APP/PS1 mouse model of Alzheimer's disease.

Metabolomics has demonstrated a great potential for the study of pathological mechanisms occurring in brain from Alzheimer's disease patients and transgenic models. However, its application to peripheral samples is not so common, although it can provide interesting information about systemic abnormalities underlying to disease. This work represents the first metabolomic investigation of multiple peripheral organs (liver, kidney, spleen, and thymus) from the APP/PS1 mice by using a high-throughput approach based on direct infusion MS. Our findings demonstrated that these organs suffer significant metabolic impairments related to energy metabolism (e.g. glycolysis, Krebs cycle, ß-oxidation), lipid homeostasis (e.g. cellular membrane breakdown and fatty acid metabolism), degradation of nucleotides, oxidative stress, hyperammonemia, and metabolism of amino acids. It is noteworthy that many of these alterations have been previously described in brain, confirming the systemic character of this neurodegenerative disorder and the utility of peripheral samples to understand its pathogenesis.

Metabolomic profiling of serum in the progression of Alzheimer's disease by capillary electrophoresis-mass spectrometry.

There is high interest in the discovery of early diagnostic biomarkers of Alzheimer's disease, for which metabolomics exhibits a great potential. In this work, a metabolomic approach based on ultrafiltration and analysis by CE-MS has been used to obtain representative fingerprints of polar metabolites from serum samples in order to distinguish between patients with Alzheimer's disease, mild cognitive impairment, and healthy controls. By the use of partial least squares discriminant analysis it was possible to classify patients according to the disease stage and then identify potential markers. Significant increase was observed with progression of disease in levels of choline, creatinine, asymmetric dimethyl-arginine, homocysteine-cysteine disulfide, phenylalanyl-phenylalanine, and different medium chain acylcarnitines. On the other hand, asparagine, methionine, histidine, carnitine, acetyl-spermidine, and C5-carnitine were reduced in these serum samples. In this way, multiple essential pathways were found implicated in the underlying pathology, such as oxidative stress or defects in energy metabolism. However, the most interesting results are related to the association of several vascular risk factors with Alzheimer's disease.

Development of a metabolomic approach based on urine samples and direct infusion mass spectrometry.

The analysis of urine by direct infusion mass spectrometry suffers from ion suppression due to its high salt content and inter-sample variability caused by the differences in urine volume between persons. Thus, urine metabolomics requires a careful selection of the sample preparation procedure and a normalization strategy to deal with these problems. Several approaches were tested for metabolomic analysis of urine samples by direct infusion electrospray mass spectrometry (DI-ESI-MS), including solid phase extraction, liquid-liquid extraction, and sample dilution. In addition, normalization of results based on conductivity values and statistical treatment was performed to minimize sample variability. Both urine dilution and solid phase extraction with mixed mode sorbent considerably reduced the salt content in urine, providing comprehensive metabolomic fingerprints. Moreover, statistical data normalization enabled the correction of inter-sample physiological variability, improving the quality of results obtained. Therefore, high-throughput DI-ESI-MS fingerprinting of urine samples can be achieved with simple pretreatment procedures allowing the use of this noninvasive sampling in metabolomics. Finally, the optimized approach was tested in a pilot metabolomic investigation of urine samples from transgenic mice models of Alzheimer's disease (APP/PS1) in order to illustrate the potential of the methodology.