Untargeted metabolomics is the study of all detectable small molecules, and in geroscience, metabolomics has shown great potential to describe the biological age—a complex trait impacted by many factors. Unfortunately, the sample sizes are often insufficient to achieve sufficient power and minimize potential biases caused by, for example, demographic factors. In this study, we present the analysis of biological age in ~10,000 toxicologic routine blood measurements. The untargeted screening samples obtained from ultra‐high pressure liquid chromatography‐quadruple time of flight mass spectrometry (UHPLC‐ QTOF) cover + 300 batches and + 30 months, lack pooled quality controls, lack controlled sample collection, and has previously only been used in small‐scale studies. To overcome experimental effects, we developed and tested a custom neural network model and compared it with existing prediction methods. Overall, the neural network was able to predict the chronological age with an rmse of 5.88 years ( r ² = 0.63) improving upon the 6.15 years achieved by existing normalization methods. We used the feature importance algorithm, Shapley Additive exPlanations (SHAP), to identify compounds related to the biological age. Most importantly, the model returned known aging markers such as kynurenine, indole‐3‐aldehyde, and acylcarnitines along with a potential novel aging marker, cyclo (leu‐pro). Our results validate the association of tryptophan and acylcarnitine metabolism to aging in a highly uncontrolled large‐s cale sample. Also, we have shown that by using robust computational methods it is possible to deploy large LC‐MS datasets for metabolomics studies to reduce the risk of bias and empower aging studies.

GHB is an endogenous short-chain organic acid presumably also widely applied as a rape and knock out drug in cases of drug-facilitated crimes or sexual assaults (DFSA). Due to the endogenous nature of GHB and its fast metabolism in vivo, the detection window of exogenous GHB is however narrow, making it challenging to prove use of GHB in DFSA cases. Alternative markers of GHB intake have recently appeared though none has hitherto been validated for forensic use. UHPLC-HRMS based screening of blood samples for drugs of abuse is routinely performed in several forensic laboratories which leaves an enormous amount of unexploited data. Recently we devised a novel metabolomics approach to use archived data from such routine screenings for elucidating both direct metabolites from exogenous compounds, but potentially also regulation of endogenous metabolism and metabolites. In this paper we used UHPLC-HRMS data acquired over a 6-year period from whole blood analysis of 51 drivers driving under the influence of GHB as well as a matched control group. The data were analyzed using a metabolomics approach applying a range of advanced analytical methods such as OPLS-DA, LASSO, random forest, and Pearson correlation to examine the data in depth and demonstrate the feasibility and potential power of the approach. This was done by initially detecting a range of potential biomarkers of GHB consumption, some that previously have been found in controlled GHB studies, as well as several new potential markers not hitherto known. Furthermore, we investigate the impact of GHB intake on human metabolism. In aggregate, we demonstrate the feasibility to extract meaningful information from archived data here exemplified using GHB cases. Hereby we hope to pave the way for more general use of the principle to elucidate human metabolites of e.g. new legal or illegal drugs as well as for applications in more global and large scale metabolomics studies in the future.

Mass spectrometry is used to determine infectious microbial species in thousands of clinical laboratories across the world. The vast amount of data allows modern data analysis methods that harvest more information and potentially answer new questions. Here, we present an end-to-end deep learning model for predicting antibiotic resistance using raw matrix assisted laser desorption ionization mass spectrometry (MALDI-MS) data. We used a 1-dimensional convolutional neural network to model (almost) raw data, skipping conventional peak-picking and directly predict resistance. The model's performance is state-of-the-art, having AUCs between 0.93 and 0.99 in all antimicrobial resistance phenotypes and validates across time and location. Feature attribution values highlight important insights into the model and how the end-to-end workflow can be improved further. This study showcases that reliable resistance phenotyping using MALDI-MS data is attainable and highlights the gains of using end-to-end deep learning for spectrometry data.

Questioned document examination aims to assess if a document of interest has been forged. Spectroscopy-based methods are the gold standard for this type of evaluation. In the past 15 years, Matrix-Assisted Laser Desorption Ionisation–Mass Spectrometry Imaging (MALDI-MSI) has emerged as a powerful analytical tool for the examination of finger marks, blood, and hair. Therefore, this study intended to explore the possibility of expanding the forensic versatility of this technique through its application to questioned documents. Specifically, a combination of MALDI-MSI and chemometric approaches was investigated for the differentiation of seven gel pens, through their ink composition, over 44 days to assess: (i) the ability of MALDI MSI to detect and image ink chemical composition and (ii) the robustness of the combined approach for the classification of different pens over time. The training data were modelled using elastic net logistic regression to obtain probabilities for each pen class and assess the time effect on the ink. This strategy led the classification model to yield predictions matching the ground truth. This model was validated using signatures generated by different pens (blind to the analyst), yielding a 100% accuracy in machine learning cross-validation. These data indicate that the coupling of MALDI-MSI with machine learning was robust for ink discrimination within the dataset and conditions investigated, which justifies further studies, including that of confounders such as paper brands and environmental factors.