Cargando…

Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics

Regulatory bodies have started to recognise the value of in vitro screening and metabolomics as two types of new approach methodologies (NAMs) for chemical risk assessments, yet few high-throughput in vitro toxicometabolomics studies have been reported. A significant challenge is to implement automa...

Descripción completa

Detalles Bibliográficos
Autores principales: Malinowska, Julia M., Palosaari, Taina, Sund, Jukka, Carpi, Donatella, Lloyd, Gavin R., Weber, Ralf J. M., Whelan, Maurice, Viant, Mark R.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8778710/
https://www.ncbi.nlm.nih.gov/pubmed/35050173
http://dx.doi.org/10.3390/metabo12010052
_version_ 1784637389559300096
author Malinowska, Julia M.
Palosaari, Taina
Sund, Jukka
Carpi, Donatella
Lloyd, Gavin R.
Weber, Ralf J. M.
Whelan, Maurice
Viant, Mark R.
author_facet Malinowska, Julia M.
Palosaari, Taina
Sund, Jukka
Carpi, Donatella
Lloyd, Gavin R.
Weber, Ralf J. M.
Whelan, Maurice
Viant, Mark R.
author_sort Malinowska, Julia M.
collection PubMed
description Regulatory bodies have started to recognise the value of in vitro screening and metabolomics as two types of new approach methodologies (NAMs) for chemical risk assessments, yet few high-throughput in vitro toxicometabolomics studies have been reported. A significant challenge is to implement automated sample preparation of the low biomass samples typically used for in vitro screening. Building on previous work, we have developed, characterised and demonstrated an automated sample preparation and analysis workflow for in vitro metabolomics of HepaRG cells in 96-well microplates using a Biomek i7 Hybrid Workstation (Beckman Coulter) and Orbitrap Elite (Thermo Scientific) high-resolution nanoelectrospray direct infusion mass spectrometry (nESI-DIMS), across polar metabolites and lipids. The experimental conditions evaluated included the day of metabolite extraction, order of extraction of samples in 96-well microplates, position of the 96-well microplate on the instrument’s deck and well location within a microplate. By using the median relative standard deviation (mRSD (%)) of spectral features, we have demonstrated good repeatability of the workflow (final mRSD < 30%) with a low percentage of features outside the threshold applied for statistical analysis. To improve the quality of the automated workflow further, small method modifications were made and then applied to a large cohort study (4860 sample infusions across three nESI-DIMS assays), which confirmed very high repeatability of the whole workflow from cell culturing to metabolite measurements, whilst providing a significant improvement in sample throughput. It is envisioned that the automated in vitro metabolomics workflow will help to advance the application of metabolomics (as a part of NAMs) in chemical safety, primarily as an approach for high throughput screening and prioritisation.
format Online
Article
Text
id pubmed-8778710
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-87787102022-01-22 Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics Malinowska, Julia M. Palosaari, Taina Sund, Jukka Carpi, Donatella Lloyd, Gavin R. Weber, Ralf J. M. Whelan, Maurice Viant, Mark R. Metabolites Article Regulatory bodies have started to recognise the value of in vitro screening and metabolomics as two types of new approach methodologies (NAMs) for chemical risk assessments, yet few high-throughput in vitro toxicometabolomics studies have been reported. A significant challenge is to implement automated sample preparation of the low biomass samples typically used for in vitro screening. Building on previous work, we have developed, characterised and demonstrated an automated sample preparation and analysis workflow for in vitro metabolomics of HepaRG cells in 96-well microplates using a Biomek i7 Hybrid Workstation (Beckman Coulter) and Orbitrap Elite (Thermo Scientific) high-resolution nanoelectrospray direct infusion mass spectrometry (nESI-DIMS), across polar metabolites and lipids. The experimental conditions evaluated included the day of metabolite extraction, order of extraction of samples in 96-well microplates, position of the 96-well microplate on the instrument’s deck and well location within a microplate. By using the median relative standard deviation (mRSD (%)) of spectral features, we have demonstrated good repeatability of the workflow (final mRSD < 30%) with a low percentage of features outside the threshold applied for statistical analysis. To improve the quality of the automated workflow further, small method modifications were made and then applied to a large cohort study (4860 sample infusions across three nESI-DIMS assays), which confirmed very high repeatability of the whole workflow from cell culturing to metabolite measurements, whilst providing a significant improvement in sample throughput. It is envisioned that the automated in vitro metabolomics workflow will help to advance the application of metabolomics (as a part of NAMs) in chemical safety, primarily as an approach for high throughput screening and prioritisation. MDPI 2022-01-08 /pmc/articles/PMC8778710/ /pubmed/35050173 http://dx.doi.org/10.3390/metabo12010052 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Malinowska, Julia M.
Palosaari, Taina
Sund, Jukka
Carpi, Donatella
Lloyd, Gavin R.
Weber, Ralf J. M.
Whelan, Maurice
Viant, Mark R.
Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics
title Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics
title_full Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics
title_fullStr Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics
title_full_unstemmed Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics
title_short Automated Sample Preparation and Data Collection Workflow for High-Throughput In Vitro Metabolomics
title_sort automated sample preparation and data collection workflow for high-throughput in vitro metabolomics
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8778710/
https://www.ncbi.nlm.nih.gov/pubmed/35050173
http://dx.doi.org/10.3390/metabo12010052
work_keys_str_mv AT malinowskajuliam automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics
AT palosaaritaina automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics
AT sundjukka automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics
AT carpidonatella automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics
AT lloydgavinr automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics
AT weberralfjm automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics
AT whelanmaurice automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics
AT viantmarkr automatedsamplepreparationanddatacollectionworkflowforhighthroughputinvitrometabolomics