Background
Brief Introduction to Mass Spectrometry
Mass spectrometry is the measurement of a molecule’s mass - the instruments are very precise scales but they work a little differently to a normal balance. Molecules are first ionised to form a charged species, which can then be separated, isolated, fragmented and detected. The most common type of ionisation in environmental chemistry is known as electrospray ionisation (ESI). Here the sample is sprayed into a hot chamber using high voltage to impart the charge to as many molecules as possible. The ions can enter the vacuum of the mass spectrometer where a sequence of electromagnetic fields can be used to manipulate the ions and control which ones make it to the detector. This way, you can measure the ions with very good accuracy and precision. High resolution mass spectrometry instruments are usually accurate to within 0.005 Da.
Summary of Non-Targeted Screening
The production of new substances is exceeding our capability to adequately assess the risk of each of these substances before they are added to the market (Persson et al., 2022). Many of these substances will then enter the environment when they are disposed of, for example in the discharge of wastewater from treatment plants. An important step in assessing the risk of chemicals in the environment is the detection of these substances. Traditional targeted analytical techniques can only detect a limited number of chemicals at once, and this requires knowledge of which substances to test and availability of reference standards. Advances in high-resolution mass spectrometry (HRMS) have allowed the simultaneous detection of tens of thousands of features* from a single sample in the same amount of time as targeted analysis. This includes naturally occurring substances, and other chemicals that do not pose a risk to environmental health. Non-targeted and suspect screening (NTS/SS) are data analysis techniques to filter and highlight substances of concern from this overwhelming number of chemicals (Szabo et al., 2024).
Suspect screening is a simple approach to prioritization that matches the m/z measured by the instrument with the known m/z from a list of known substances. For example, if you want to see if PFOS is in your sample, then you can calculate the exact mass is from the molecular formula (C8HF17O3S = 499.9374 Da), then look for the [M-H]- at the m/z = 498.9297. You can submit a list of as many chemicals as you like in this way, as long as you know the molecular formula for each substance. However, matches with candidates from a list of m/z does not always provide a confident structural match. Notice that we can only match against a molecular formula, so there could be isomers – that are different substances with the same mass. To increase the confidence in a structural match, you can use the retention time, and tandem mass spectrum (MS2) to provide more evidence for the match (Schymanski et al., 2014).
Non-targeted screening (AKA non targeted analysis) is an unbiased data analysis methodology when you do not know anything about the substances that may be present in the sample. These methods use statistics to group and filter HRMS features first, then try to identify the structure afterwards. The most commonly used type of NTS is differential analysis. Here you compare the presence and abundance of each feature between two or more samples. For example, you could filter for only those features that increase in peak area in a river before and after a sewage discharge point. This will highlight all those substances that are likely coming from that environmental source. A more complex NTS method is known as Feature Based Molecular Networking (FBMN) (Nothias et al., 2020). Here the relationship between each feature is computed by evaluating the similarity of the MS2. If the fragments of two features are similar, then its likely that they have a similar structure and they will be grouped together in a network. This is a very useful analysis to look for metabolites and transformation products.
* A feature is a data tuple – which means it’s a peak of a specific mass to charge ratio (m/z) and retention time (RT) (Samanipour et al., 2019). Note: Sometimes one m/z can elute at two retention times, so it’s important to distinguish between these.
Recorded Lecture
Drew Szabo who is an analytical chemist at the University of York, describes how high-resolution mass spectrometry (HRMS) can be used to study chemical contaminants in the environment.
Key Reading
Persson, L., et al., (2022) Outside the Safe Operating Space of the Planetary Boundary for Novel Entities. Environmental Science & Technology, 56(3):1510–1521. https://doi.org/10.1021/acs.est.1c04158
Szabo, D., et al., (2024) Online and Offline Prioritization of Chemicals of Interest in Suspect Screening and Non-targeted Screening with High-Resolution Mass Spectrometry. Analytical Chemistry, 96(9):3707–3716. https://doi.org/10.1021/acs.analchem.3c05705
Schymanski, E.L., et al., (2014) Identifying Small Molecules via High Resolution Mass Spectrometry: Communicating Confidence. Environmental Science & Technology, 48(4):2097–2098. https://doi.org/10.1021/es5002105
Nothias, L.-F., et al., (2020) Feature-based molecular networking in the GNPS analysis environment. Nature Methods, 17(9):905–908. https://doi.org/10.1038/s41592-020-0933-6
Samanipour, S., et al., (2019) Letter to the Editor: Optimism for Nontarget Analysis in Environmental Chemistry. Environmental Science & Technology, 53(10):5529–5530. https://doi.org/10.1021/acs.est.9b01476