EXPLORING THE VOLATOME OF NONCOMMUNICABLE DISEASES AS A PROMISING, INNOVATIVE AND INTEGRATING APPROACH FOR THEIR RAPID DIAGNOSTICS. THE CASE STUDY OF CANCER AND NEURODEGENERATIVE DISEASES

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Exploring the potential of NTME/GC-MS, in the establishment of urinary volatomic profiles. Lung cancer patients as case study

Publication . Porto-Figueira, Priscilla; Pereira, Jorge; Miekisch, Wolfram; Câmara, José S.

The growing cancer incidence and mortality worldwide claims for the development of novel diagnostic strategies. In this study we aimed to explore the potential of an innovative methodology, based on a needle trap microextraction (NTME), combined with gas chromatography-mass spectrometry (GC-MS), as new approach to isolate and profile urinary volatile organic metabolites (VOMs) from lung cancer (LC) patients and healthy individuals (CTRL). In this context, different experimental parameters with influence of NTME extraction efficiency including, temperature, equilibration time, headspace volume, ionic strength, pH, effects of sample volume and stirring, were investigated and optimized. For the DVB/CarX/Car1000 needle trap device (NTD), the best results were obtained using 40 mL headspace of a 4-mL acidified (pH = 2) urine sample with 20% NaCl and an extraction temperature of 50 °C for 40 min of equilibration time. The stability of the isolated VOMs was investigated up to 72 h after extraction. From the VOMs identified, belonging namely to ketones, sulphur and benzene derivatives, 98 presented a frequency of occurrence above 90%. Data were processed by discriminant analysis, retrieving differentiated clusters for LC and CTRL groups. As far we are aware, this is the first study using NTME/GC-MS to establish urinary volatomic profiles. Preliminary results are very promising, as broad and comprehensive volatile profiles were obtained. Moreover, the extended storage stability of the NTD devices opens new opportunities for sampling other matrices in a wide range of applications.

2018Journal article

Open access

Untargeted urinary 1H NMR-based metabolomic pattern as a potential platform in breast cancer detection

Publication . Silva, Catarina L.; Olival, Ana; Perestrelo, Rosa; Silva, Pedro; Tomás, Helena; Câmara, José S.

: Breast cancer (BC) remains the second leading cause of death among women worldwide. An emerging approach based on the identification of endogenous metabolites (EMs) and the establishment of the metabolomic fingerprint of biological fluids constitutes a new frontier in medical diagnostics and a promising strategy to differentiate cancer patients from healthy individuals. In this work we aimed to establish the urinary metabolomic patterns from 40 BC patients and 38 healthy controls (CTL) using proton nuclear magnetic resonance spectroscopy (1H-NMR) as a powerful approach to identify a set of BC-specific metabolites which might be employed in the diagnosis of BC. Orthogonal partial least squares-discriminant analysis (OPLS-DA) was applied to a 1H-NMR processed data matrix. Metabolomic patterns distinguished BC from CTL urine samples, suggesting a unique metabolite profile for each investigated group. A total of 10 metabolites exhibited the highest contribution towards discriminating BC patients from healthy controls (variable importance in projection (VIP) >1, p < 0.05). The discrimination efficiency and accuracy of the urinary EMs were ascertained by receiver operating characteristic curve (ROC) analysis that allowed the identification of some metabolites with the highest sensitivities and specificities to discriminate BC patients from healthy controls (e.g. creatine, glycine, trimethylamine N-oxide, and serine). The metabolomic pathway analysis indicated several metabolism pathway disruptions, including amino acid and carbohydrate metabolisms, in BC patients, namely, glycine and butanoate metabolisms. The obtained results support the high throughput potential of NMR-based urinary metabolomics patterns in discriminating BC patients from CTL. Further investigations could unravel novel mechanistic insights into disease pathophysiology, monitor disease recurrence, and predict patient response towards therapy.

2019Journal article

Open access

Exploring the potential of needle trap microextraction combined with chromatographic and statistical data to discriminate different types of cancer based on urinary volatomic biosignature

Publication . Porto-Figueira, Priscilla; Pereira, Jorge A. M.; Câmara, José S.

The worldwide high cancer incidence and mortality demands for more effective and specific diagnostic strategies. In this study, we evaluated the efficiency of an innovative methodology, Needle Trap Microextraction (NTME), combined with gas chromatography-mass spectrometry (GC-MS), for the establishment of the urinary volatomic biosignature from breast (BC), and colon (CC) cancer patients as well as healthy individuals (CTL). To achieve this, 40 mL of the headspace of acidified urine (4 mL, 20% NaCl, pH = 2), equilibrated at 50 °C during 40 min, were loaded through the DVB/Car1000/CarX sorbent inside the NTD, and subjected to a GC-MS analysis. This allowed the identification of 130 VOMs from different chemical families that were further processed using discriminant analysis through the partial least squares method (PLS-DA). Several pathways are over activated in cancer patients, being phenylalanine pathway in BC and limonene and pinene degradation pathway in CC the most relevant. Butanoate metabolism is also highly activated in both cancers, as well as tyrosine metabolism in a lesser extension. In BC the xenobiotics metabolism by cytochrome P450 and fatty acid biosynthesis are also differentially activated. Different clusters corresponding to the groups recruited allowed to define sets of volatile organic metabolites (VOMs fingerprints) that exhibit high classification rates, sensitivity and specificity in the discrimination of the selected cancers. As far as we are aware, this is the first time that NTME is used for isolation urinary volatile metabolites, being the obtained results very promising.

2018Journal article

Restricted access

Volatilomic insight of head and neck cancer via the effects observed on saliva metabolites

Publication . Taware, Ravindra; Taunk, Khushman; Pereira, Jorge A. M.; Shirolkar, Amey; Soneji, Dharmesh; Câmara, José S.; Nagarajaram, H. A.; Rapole, Srikanth

Head and neck cancer (HNC) is a heterogeneous malignant disease with distinct global distribution. Metabolic adaptations of HNC are significantly gaining clinical interests nowadays. Here, we investigated effects of HNC on differential expression of volatile metabolites in human saliva. We applied headspace solid phase microextraction coupled with gas chromatography-mass spectrometry analysis of saliva samples collected from 59 human subjects (HNC - 32, Control - 27). We identified and quantified 48 volatile organic metabolites (VOMs) and observed profound effects of HNC on these metabolites. These effects were VOM specific and significantly differed in the biologically comparable healthy controls. HNC induced changes in salivary VOM composition were well attributed to in vivo metabolic effects. A panel of 15 VOMs with variable importance in projection (VIP) score >1, false discovery rate (FDR) corrected p-value < 0.05 and log2 fold change (log2 FC) value of ≥0.58/≤-0.58 were regarded as discriminatory metabolites of pathophysiological importance. Afterwards, receiver operator characteristic curve (ROC) projected certain VOMs viz., 1,4-dichlorobenzene, 1,2-decanediol, 2,5-bis1,1-dimethylethylphenol and E-3-decen-2-ol with profound metabolic effects of HNC and highest class segregation potential. Moreover, metabolic pathways analysis portrayed several dysregulated pathways in HNC, which enhanced our basic understanding on salivary VOM changes. Our observations could redefine several known/already investigated systemic phenomenons (e.g. biochemical pathways). These findings will inspire further research in this direction and may open unconventional avenues for non-invasive monitoring of HNC and its therapy in the future.

2018Journal article

Open access

Fingerprint targeted compounds in authenticity of sugarcane honey: an approach based on chromatographic and statistical data

Publication . Silva, Pedro; Silva, Catarina L.; Perestrelo, Rosa; Nunes, Fernando M.; Câmara, José S.

ugarcane honey (SCH) is a black syrup recognized by its excellent quality, being produced in Madeira Island using the regional sugarcane cultivars and following a traditional and peculiar manufacturing and storage processes. However, some low-quality commercial products have been labeled as SCH but do not respect its criteria, revealing the need of develop powerful strategies in order to detect and prevent adulterations. The knowledge of furanic derivatives (FDs) profile, produced during browning reactions that occurs during food processing and storage, emerged as a promising strategy in food quality and fraud prevention. Therefore, the aim of this study was to establish the FDs profiling of typical SCH produced by certified and non-certified producers, in different geographical regions (Madeira and Brazil), based on microextraction by packed sorbent (MEPS) combined with ultra-high performance liquid chromatography (UHPLC) as a useful approach to define its typicality and authenticity. These parameters are defined through the differentiation and discrimination of FDs profiles among other sugarcane-derived products using multivariate statistical analysis (ANOVA with post-hoc Tukey, principal components analysis, partial least square, linear discriminant analysis and hierarchical clustering). The results demonstrated that SCH samples from non-certified producers present the highest levels of FDs. In addition, SCH samples from Brazil present higher levels of FDs than samples from Madeira region. The obtained results revealed that the proposed approach is a valuable strategy to establish the typicality of SCH, ensuring its quality, authenticity, safety control and a useful support regarding the application of SCH from Madeira Island to EU certification.

2018Journal article

Restricted access