Bioaffinity mass spectrometry for screening and identification of contaminants
Our environment is constantly threatened by large amounts and variations of man-made chemicals and natural substances. Parts of these substances accumulate and contaminate soil and surface water, affecting the organisms living in it and eventually contaminate the food chain. The European Union (EU) has imposed regulations and obliged EU member states to monitor for possible contaminants in the environment and food. For this, highly sophisticated mass spectrometry (MS) techniques, which can nowadays screen >100 contaminants in a single run, are applied. For rapid and inexpensive screening of contaminants, bioactivity-based screening assays are applied, however, identification of compounds based on their chemical-physical properties is not possible. As both methods cannot identify emerging and unknown bioactive contaminants, there is a need for new tools and concepts. In this thesis, new bioaffinity MS (BioMS) concepts, using an antibody, transport proteins and a receptor, are presented for the screening and identification of contaminants. In the first concept, monoclonal antibodies (Mabs) against ochratoxins were coupled to fluorescent labeled paramagnetic microbeads for high-throughput flow cytometric screening of ochratoxins in wheat and cereal. The identification of ochratoxins with nano-ultra performance liquid chromatography-quadrupole-time-of-flight-MS (nano-UPLC-Q-ToF-MS) was achieved in full scan accurate mass mode. In the second BioMS approach, the flow cytometer was replaced by UPLC-triple quadrupole (QqQ)-MS for rapid screening of thyroid transporter ligands. For this, thyroid transport protein transthyretin (TTR) was immobilized onto inexpensive non-colored paramagnetic microbeads and a stable isotopic thyroid hormone was used as label in the competitive inhibition format. For the identification of TTR-binding endocrine disrupting chemicals (EDCs) in process water and urine, nano-UPLC-Q-ToF-MS was used. In order to perform high-throughput screening, a microtiter plate-based high-throughput BioMS approach was developed with the same beads but coupled with recombinant human sex hormone-binding globulin (rhSHBG) for the detection of designer steroids in dietary supplements. Following the screening with rhSHBG-based BioMS using LC-QqQ-MS, the rhSHBG bioaffinity extracts were injected onto chip-UPLC-Q-ToF-MS operated in full scan mode and a wide range of steroids were identified. The same approach was applied with the estrogen receptor α (ERα) in which LC-QqQ-MS, instead of the commonly applied GC-MS, was used for the screening of estrogens with a suitable LC-MS-compatible label. The identification of estrogens in ERα-purified supplement extracts was achieved with UPLC-ion mobility (IM)-Q-ToF-MS. These new BioMS concepts present new tools for the screening and identification of emerging yet unknown food and environmental contaminants to ensure consumer’s health and fair play in sports.
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| Other Authors: | |
| Format: | Doctoral thesis biblioteca |
| Language: | English |
| Published: |
Wageningen University
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| Subjects: | analytical chemistry, contaminants, mass spectrometry, analytische scheikunde, besmetters, massaspectrometrie, |
| Online Access: | https://research.wur.nl/en/publications/bioaffinity-mass-spectrometry-for-screening-and-identification-of |
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| Summary: | Our environment is constantly threatened by large amounts and variations of man-made chemicals and natural substances. Parts of these substances accumulate and contaminate soil and surface water, affecting the organisms living in it and eventually contaminate the food chain. The European Union (EU) has imposed regulations and obliged EU member states to monitor for possible contaminants in the environment and food. For this, highly sophisticated mass spectrometry (MS) techniques, which can nowadays screen >100 contaminants in a single run, are applied. For rapid and inexpensive screening of contaminants, bioactivity-based screening assays are applied, however, identification of compounds based on their chemical-physical properties is not possible. As both methods cannot identify emerging and unknown bioactive contaminants, there is a need for new tools and concepts. In this thesis, new bioaffinity MS (BioMS) concepts, using an antibody, transport proteins and a receptor, are presented for the screening and identification of contaminants. In the first concept, monoclonal antibodies (Mabs) against ochratoxins were coupled to fluorescent labeled paramagnetic microbeads for high-throughput flow cytometric screening of ochratoxins in wheat and cereal. The identification of ochratoxins with nano-ultra performance liquid chromatography-quadrupole-time-of-flight-MS (nano-UPLC-Q-ToF-MS) was achieved in full scan accurate mass mode. In the second BioMS approach, the flow cytometer was replaced by UPLC-triple quadrupole (QqQ)-MS for rapid screening of thyroid transporter ligands. For this, thyroid transport protein transthyretin (TTR) was immobilized onto inexpensive non-colored paramagnetic microbeads and a stable isotopic thyroid hormone was used as label in the competitive inhibition format. For the identification of TTR-binding endocrine disrupting chemicals (EDCs) in process water and urine, nano-UPLC-Q-ToF-MS was used. In order to perform high-throughput screening, a microtiter plate-based high-throughput BioMS approach was developed with the same beads but coupled with recombinant human sex hormone-binding globulin (rhSHBG) for the detection of designer steroids in dietary supplements. Following the screening with rhSHBG-based BioMS using LC-QqQ-MS, the rhSHBG bioaffinity extracts were injected onto chip-UPLC-Q-ToF-MS operated in full scan mode and a wide range of steroids were identified. The same approach was applied with the estrogen receptor α (ERα) in which LC-QqQ-MS, instead of the commonly applied GC-MS, was used for the screening of estrogens with a suitable LC-MS-compatible label. The identification of estrogens in ERα-purified supplement extracts was achieved with UPLC-ion mobility (IM)-Q-ToF-MS. These new BioMS concepts present new tools for the screening and identification of emerging yet unknown food and environmental contaminants to ensure consumer’s health and fair play in sports. |
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