Comparing two metabolic profiling approaches (liquid chromatography and gas chromatography coupled to mass spectrometry) for extra-virgin olive oil phenolic compounds analysis: A botanical classification perspective.

Over the last decades, the phenolic compounds from virgin olive oil (VOO) have become the subject of intensive research because of their biological activities and their influence on some of the most relevant attributes of this interesting matrix. Developing metabolic profiling approaches to determine them in monovarietal virgin olive oils could help to gain a deeper insight into olive oil phenolic compounds composition as well as to promote their use for botanical origin tracing purposes. To this end, two approaches were comparatively investigated (LC-ESI-TOF MS and GC-APCI-TOF MS) to evaluate their capacity to properly classify 25 olive oil samples belonging to five different varieties (Arbequina, Cornicabra, Hojiblanca, Frantoio and Picual), using the entire chromatographic phenolic profiles combined to chemometrics (principal component analysis (PCA) and partial least square-discriminant analysis (PLS-DA)). The application of PCA to LC-MS and GC-MS data showed the natural clustering of the samples, seeing that 2 varieties were dominating the models (Arbequina and Frantoio), suppressing any possible discrimination among the other cultivars. Afterwards, PLS-DA was used to build four different efficient predictive models for varietal classification of the samples under study. The varietal markers pointed out by each platform were compared. In general, with the exception of one GC-MS model, all exhibited proper quality parameters. The models constructed by using the LC-MS data demonstrated superior classification ability.

Exploratory analysis of urinary tract infection using a GC-APCI-MS platform.

Urinary tract infection (UTI) is among the most common bacterial infections worldwide. The understanding of the physiological mechanisms affected by UTI may need modern integrative '-omics' technologies, and metabolomics in particular. Here we present the first GC-APCI-MS-based explorative metabolomics study of UTI, using MS and FID detectors simultaneously. This provides high quality mass spectral data as well as semi-quantitative information demonstrating the feasibility of the GC-APCI-MS platform for non-targeted approaches. The work is part of a bigger project aiming at providing a comprehensive overview of UTI-induced changes in urine. Taking advantage of a fully clinically characterized cohort that offers the possibility of both case-control and longitudinal modelling, we can define UTI-induced change as a list of urinary metabolites which distinguish E. coli UTI patients from the subjects with no signs of an active infection. The list of molecular descriptors includes compounds related to bacterial activity such as lactic acid and lactose while other molecules show an association with the physiological status (inositol, citric acid).

Evaluation of gas chromatography-atmospheric pressure chemical ionization-mass spectrometry as an alternative to gas chromatography-electron ionization-mass spectrometry: avocado fruit as example.

Although GC-APCI-MS was developed more than 40 years ago this coupling is still far from being a routine technique. One of the reasons explaining the limited use of GC-APCI so far is the lack of spectral database which facilitates the identification of the compounds under study. The first application of a very recently developed GC-APCI database to identify as many compounds as possible in a complex matrix such as avocado fruit is presented here. The results achieved by using this database has been checked against those obtained using traditional GC-EI-MS and a comparison of the MS signals observed in both ionization sources has been carried out. 100 compounds belonging to different chemical families were identified in the matrix under study. Considering the results of this study, the wide range of application (in terms of polarity and size of analytes) and the robustness of APCI as interface, the high quality of TOF spectra, and our library as a publicly available resource, GC-APCI-TOF MS is definitively a valuable addition to the "metabolomics toolbox".

Online spectral library for GC-atmospheric pressure chemical ionization-ToF MS.

BACKGROUND: Invented more than three decades ago by Horning, GC-MS under atmospheric pressure chemical ionization (GC-APCI-MS) has only recently emerged from years of obscurity. However, the general acceptance of GC-APCI-MS is certainly constrained by the lack of spectral libraries, which make the traditional GC-MS approaches so powerful. RESULTS: Here we present a concept of a GC-APCI-QqToF spectral library. The library is web-based, fully searchable and at moment includes spectra of 150 compounds from the most common chemical families. The fragmentation pattern of some chemical families is explained and a protocol for de novo identification has been provided in order to facilitate the identification of unknown compounds. CONCLUSION: A library for GC-APCI-QqToF is now publicly available online.

(1)H NMR-based metabolic profiling of urinary tract infection: combining multiple statistical models and clinical data.

Urinary tract infection (UTI) encompasses a variety of clinical syndromes ranging from mild to life-threatening conditions. As such, it represents an interesting model for the development of an analytically based scoring system of disease severity and/or host response. Here we test the feasibility of this concept using (1)H NMR based metabolomics as the analytical platform. Using an exhaustively clinically characterized cohort and taking advantage of the multi-level study design, which opens possibilities for case-control and longitudinal modeling, we were able to identify molecular discriminators that characterize UTI patients. Among those discriminators a number (e.g. acetate, trimethylamine and others) showed association with the degree of bacterial contamination of urine, whereas others, such as, for instance, scyllo-inositol and para-aminohippuric acid, are more likely to be the markers of morbidity. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11306-012-0411-y) contains supplementary material, which is available to authorized users.

Metabolomic investigations of human infections.

Metabolomics has a special place among other 'omics' disciplines (genomics, transcriptomics and proteomics) as it describes the most dynamic level of biological regulation and, as such, provides the most direct reflection of the physiological status of an organism. Quick development of the analytical technologies in the first place - MS and NMR - has enabled the metabolomics analysis of such complex biological phenomena as host-pathogen interactions in the development of infection. In this review, an overview of the metabolomics studies of infectious diseases carried out on human material is provided. The relevant papers on the metabolomics of human infectious diseases are comprehensively summarized in a table, including, for example, information on the study design, number of subjects, employed technology and metabolic discriminator. Future considerations, such as importance of the time-resolved study designs and the embedment of metabolomics in large-scale epidemiological studies are discussed.

Fibrinogen alpha chain O-glycopeptides as possible markers of urinary tract infection.

Urinary tract infection (UTI) is the most common bacterial infection leading to substantial morbidity and considerable health care expenditures across all ages. Here we present an exploratory UPLC-MS study of human urine in the context of febrile, complicated urinary tract infection aimed to reveal and identify possible markers of a host response on infection. A UPLC-MS based workflow, taking advantage of Ultra High Resolution (UHR) Qq-ToF-MS, and multivariate data handling were applied to a carefully selected group of 39 subjects with culture-confirmed febrile Escherichia coli UTI. Using a combination of unsupervised and supervised multivariate modeling we have pinpointed a number of peptides specific for UTI. An unequivocal structural identification of these peptides, as O-glycosylated fragments of the human fibrinogen alpha 1 chain, required MS2 and MS3 experiments on two different MS platforms: ESI-UHR-Qq-ToF and ESI-ion trap, a blast search and, finally, confirmation was achieved by matching experimental tandem mass spectra with those of custom synthesized candidate-peptides. In conclusion, exploiting non-targeted UPLC-MS based approach for the investigation of UTI related changes in urine, we have identified and structurally characterized unique O-glycopeptides, which are, to our knowledge, the first demonstration of O-glycosylation of human fibrinogen alpha 1-chain.

Ultra high performance liquid chromatography-time of flight mass spectrometry for analysis of avocado fruit metabolites: method evaluation and applicability to the analysis of ripening degrees.

We have developed an analytical method using UHPLC-UV/ESI-TOF MS for the comprehensive profiling of the metabolites found in the methanolic extracts of 13 different varieties of avocado at two different ripening degrees. Both chromatographic and detection parameters were optimized in order to maximize the number of compounds detected and the sensitivity. After achieving the optimum conditions, we performed a complete analytical validation of the method with respect to its linearity, sensitivity, precision, accuracy and possible matrix effects. The LODs ranged from 1.64 to 730.54 ppb (in negative polarity) for benzoic acid and chrysin, respectively, whilst they were found within the range from 0.51 to 310.23 ppb in positive polarity. The RSDs for repeatability test did not exceed 7.01% and the accuracy ranged from 97.2% to 102.0%. Our method was then applied to the analysis of real avocado samples and advanced data processing and multivariate statistical analysis (PCA, PLS-DA) were carried out to discriminate/classify the examined avocado varieties. About 200 compounds belonging to various structural classes were tentatively identified; we are certain about the identity of around 60 compounds, 20 of which have been quantified in terms of their own commercially available standard.

Gas chromatography-atmospheric pressure chemical ionization-time of flight mass spectrometry for profiling of phenolic compounds in extra virgin olive oil.

A new analytical approach based on gas chromatography coupled to atmospheric pressure chemical ionization-time of flight mass spectrometry was evaluated for its applicability for the analysis of phenolic compounds from extra-virgin olive oil. Both chromatographic and MS parameters were optimized in order to improve the sensitivity and to maximize the number of phenolic compounds detected. We performed a complete analytical validation of the method with respect to its linearity, sensitivity, precision, accuracy and possible matrix effects. The LODs ranged from 0.13 to 1.05ppm for the different tested compounds depending on their properties. The RSDs for repeatability test did not exceed 6.07% and the accuracy ranged from 95.4% to 101.5%. To demonstrate the feasibility of our method for analysis of real samples, we analyzed the extracts of three different commercial extra-virgin olive oils. We have identified unequivocally a number of phenolic compounds and obtained quantitative information for 21 of them. In general, our results show that GC-APCI-TOF MS is a flexible platform which can be considered as an interesting tool for screening, structural assignment and quantitative determination of phenolic compounds from virgin olive oil.

Evaluation of GC-APCI/MS and GC-FID as a complementary platform.

With a development of the metabolomics field, complementary cross-platform approaches started to attract attention, as none of the contemporary analytical methods had the capacity to cover the entire space of the human metabolome. In the current manuscript, we have evaluated an online coupling of gas chromatography (GC)-mass spectrometry (MS) and flame ionization detector (FID) as ways of cross-detector analysis. The possible value of this combination was recognized from the very first days of GC-MS history but was never explored in detail. We have compared the basic analytical parameters of both detectors, such as limit of detection (LOD) and limit of quantification, with intra- and interday reproducibility. We show that for the majority of the tested compounds, MS detector demonstrates lower LOD. At the same time, FID appeared to be more robust, showing lower relative standard deviations (RSDs) for intra- and interday reproducibility. We conclude that the gain of this dual detector acquisition appears to be most evident for complex biological samples, where wide dynamic range and predictable response of FID are useful for an initial quantitative overview of sample composition and estimation of molar proportions of different metabolites. MS provides reliable, structural information and superior, at least in the case of atmospheric pressure chemical ionization, sensitivity. Taken together, both detectors represent a flexible tool for explorative studies and if supported by a powerful data-processing algorithm, would appear to be useful in any metabolic profiling study.

Monitoring quinolone antibacterial residues in bovine tissues: extraction with hot water and liquid chromatography coupled to a single- or triple-quadrupole mass spectrometer.

A rapid and sensitive procedure for determining residues of seven quinolone antibacterials in bovine muscle, kidney and liver is presented. The method is based on the matrix solid-phase dispersion (MSPD) technique with hot water as extractant followed by liquid chromatography/single quadrupole mass spectrometry (LC/MS) or triple-quadrupole mass spectrometry (LC/MS/MS). After dispersing tissue samples on hydrazine sulfate treated sand, target compounds were eluted from the MSPD column by passing through it 4 mL of water heated at 100 degrees C. After pH adjustment and filtration, 200 and 5 microL of the aqueous extracts were respectively injected into the LC/MS and LC/MS/MS instruments. With the former instrument, MS data were acquired in the three-ion selected ion monitoring mode, while MS/MS data acquisition was performed in the multi-reaction monitoring mode by selecting two precursor ion to product ion transitions for each target compound. Hot water appeared to be an efficient extracting medium, since absolute recoveries of the analytes were 84-102%. Using norfloxacin (a quinolone not used in veterinary medicine) as surrogate internal standard, the accuracy of the method at three concentration levels equal to 0.5, 1 and 1.5 times the maximum residue limits (MRLs) set by the european union was 88-109% with relative standard deviations (RSDs) not higher than 7%. The use of LC/MS/MS allowed detection and quantification of the analytes in any tissue considered to be performed at concentrations by far lower than half of their MRLs. Vice versa, the single-quadrupole MS arrangement, while succeeding in monitoring quinolones in muscle tissue at the 0.5 MRL level, showed to be not sufficiently selective for unambiguous identification of some quinolones in kidney and liver.