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Analysis of extracellular metabolome by HS-SPME/GC–MS: optimization and application in a pilot study to evaluate galactosamine-induced hepatotoxicity

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dc.contributor.authorAraújo, Ana Margarida
dc.contributor.authorMoreira, Nathalie
dc.contributor.authorLima, Ana Rita
dc.contributor.authorBastos, Maria de Lourdes
dc.contributor.authorCarvalho, Félix
dc.contributor.authorCarvalho, Márcia
dc.contributor.authorGuedes de Pinho, Paula
dc.date.accessioned2021-07-02T09:37:53Z
dc.date.available2021-07-02T09:37:53Z
dc.date.issued2018
dc.description.abstractTwo methods based on headspace solid-phase microextraction (HS-SPME) coupled to gas chromatography-mass spectrometry (GC-MS) were developed to study in vitro the volatile exometabolome, which were then further tested in a pilot study to evaluate galactosamine-induced hepatotoxicity. The analysis of volatile organic compounds (VOCs) was carried out directly in the headspace of the cell culture medium, while some other volatile organic compounds such as volatile carbonyl compounds (VCCs) (aldehydes and ketones) were determined in the headspace of the cell culture medium after a previous derivatization step with O-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine hydrochloride (PFBHA). Fiber selection was performed using a univariate mode, whereas a central composite design (CCD) was used in the optimization of several other parameters that affect the extraction conditions. VOCs showed optimal extraction results using a DVB/CAR/PDMS fiber, by adding 0.43 g of NaCl to a sample volume of 2 mL and allowing the sample to equilibrate for 10 min at 45 °C with a subsequent extraction for 39 min at the same temperature. For VCCs, the best extraction response was achieved after in-solution (2 mL) derivatization with 0.94 g L-1 of PFBHA (final concentration), followed by an incubation period of 6 min and an extraction time of 37 min at 53 °C, using a PDMS/DVB fiber. The applicability of both optimized methods was then tested, through a untargeted study, on cell culture medium samples obtained from primary mouse hepatocytes (PMH) exposed to three low concentrations (LC01, LC10 and LC30) of the well-known hepatotoxic agent galactosamine (GalN). The results obtained by both methods showed that volatile compounds from GalN exposed cells are separated from controls in a concentration-dependent manner. Several volatile compounds, namely aldehydes, ketones and alcohols, suffered significant alterations, suggesting that GalN induces marked metabolic alterations in cells even at low, non-toxic concentrations. Although preliminary, this metabolomics approach proved its potential to be used in future studies to evaluate toxicity of different xenobiotics.pt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.doi10.1016/j.toxlet.2018.05.028pt_PT
dc.identifier.issn0378-4274
dc.identifier.urihttp://hdl.handle.net/10284/10014
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherElsevierpt_PT
dc.relationThis work received financial support from the European Union (FEDER funds POCI/01/0145/FEDER/07728) and National funds (FCT/MEC, Fundação para a Ciência e a Tecnologia and Ministério da Educação e Ciência) under the Partnership Agreement PT2020 UID/ MULTI/04378/2013. We also thank to the ‘Comissão de Coordenação e Desenvolvimento Regional do Norte (CCDRN)/NORTE2020/Portugal2020 through the project: DESignBIOtecHealth – New Technologies for three Health Challenges of Modern Societies: Diabetes, Drug Abuse and Kidney Diseases for its financial support. Ana Margarida Araújo, Ana Rita Lima and Nathalie Moreira thank FCT for their PhD and postgraduate fellowships (SFRH/BD/107708/ 2015, SFRH/BD/123012/2016 and SFRH/BPD/109668/2015, respectively). Márcia Carvalho also acknowledges FCT through project UID/ MULTI/04546/2016.pt_PT
dc.subjectHeadspace solid-phase microextractionpt_PT
dc.subjectGas chromatography–mass spectrometrypt_PT
dc.subjectCentral composite designpt_PT
dc.subjectExometabolomept_PT
dc.subjectPrimary mouse hepatocytespt_PT
dc.subjectGalactosaminept_PT
dc.titleAnalysis of extracellular metabolome by HS-SPME/GC–MS: optimization and application in a pilot study to evaluate galactosamine-induced hepatotoxicitypt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.citation.endPage31pt_PT
oaire.citation.startPage22pt_PT
oaire.citation.titleToxicology Letterspt_PT
oaire.citation.volume295pt_PT
person.familyNameCarvalho
person.givenNameMarcia
person.identifier2017111
person.identifier.ciencia-id8B10-171E-E63E
person.identifier.orcid0000-0001-9884-4751
person.identifier.ridD-5999-2013
person.identifier.scopus-author-id7201413997
rcaap.rightsclosedAccesspt_PT
rcaap.typearticlept_PT
relation.isAuthorOfPublication3837b828-ba57-47f7-a811-cce65e4922c6
relation.isAuthorOfPublication.latestForDiscovery3837b828-ba57-47f7-a811-cce65e4922c6

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