Untargeted Metabolomics Dataset from Rheumatic Heart Disease, Degenerative Aortic Stenosis, and Healthy Controls

<b>Study Summary</b>This dataset contains untargeted LC–MS/MS metabolomics profiles of serum samples collected from patients with rheumatic heart disease (RHD, n = 33), degenerative aortic stenosis (AS, n = 9), and healthy controls (n = 15). The study aimed to characterize metabolic alterations associated with valvular heart diseases relative to healthy individuals.Serum samples were analyzed using ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UHPLC–QTOF–MS) operated in both positive and negative electrospray ionization (ESI⁺/ESI⁻) modes. The dataset includes processed peak tables, and metadata describing the clinical samples.<b>Study Design</b><b>Study groups:</b>Rheumatic Heart Disease (RHD): n = 33Degenerative Aortic Stenosis (AS): n = 9Healthy Controls: n = 15<b>Quality control and blanks:</b><i>Pooled QC:</i> prepared by combining equal aliquots of all serum samples (RHD, AS, and controls). Multiple aliquots were stored to avoid repeated freeze–thaw cycles.<i>Long-term QC:</i> NIST SRM 1950 plasma reference standard (NIST, USA) was included in each batch as a long-term reference.<i>Blanks:</i> extraction blanks were processed alongside study samples to monitor background contamination.<b>Sample Collection</b>Venous blood was collected from all participants after written informed consent. For RHD and AS patients, samples were obtained prior to general anaesthesia. Blood was drawn into VACUETTE® CAT serum clot activator tubes (Greiner Bio-One, Austria), allowed to clot for 30 minutes at room temperature, and centrifuged at 2,000 × g for 15 minutes at 4 °C. Serum aliquots were stored at −80 °C until analysis (within 1–2 years of collection).<b>Sample Preparation and Extraction</b>Frozen serum samples were thawed on ice. Metabolites were extracted from 100 µL of each sample (including QC and reference materials) by adding 400 µL of ice-cold acetonitrile:methanol (9:1, v/v) containing 1% formic acid (for ESI⁺) or 1 mM acetic acid (for ESI⁻). The mixture was vortexed for 2 minutes and centrifuged at 14,000 × g for 15 minutes at 4 °C. Supernatants were dried under nitrogen gas, reconstituted in 100 µL of deionized water with 0.1% formic acid (ESI⁺) or 1 mM acetic acid (ESI⁻), and centrifuged again before LC–MS analysis.<b>Chromatographic Separation</b>Chromatographic separation was performed on an ExionLC™ AD UPLC system (SCIEX, USA) using an Omega Polar C18 column (1.6 µm, 100 Å, 3 × 100 mm; Phenomenex, USA) maintained at 40 °C.<b>Mobile phase A:</b> HPLC-grade water<b>Mobile phase B:</b> acetonitrile:methanol (1:1, v/v)<b>Modifiers:</b> 2 mM ammonium formate and 0.1% formic acid (ESI⁺) or 1 mM acetic acid (ESI⁻)<b>Gradient:</b> 60 minutes at 0.4 mL/min<b>Injection volume:</b> 5 µL<b>Autosampler temperature:</b> 15 °C<b>Needle rinsing:</b> water:methanol:isopropanol (6:3:1, v/v/v) between injections<b>Mass Spectrometry</b>Mass spectrometric analysis was performed on a SCIEX X500R Q-TOF equipped with a dual ESI source and operated in data-dependent acquisition (DDA) mode using SCIEX OS 1.4.<b>Source parameters:</b>Ion source gas 1: 40 psiIon source gas 2: 65 psiCurtain gas: 25 psiSource temperature: 500 °CIon spray voltage: +5000 V (ESI⁺)Collision gas (CAD): 7 psi<b>Acquisition settings:</b>Mass range: m/z 50–1200TOF–MS accumulation time: 200 msTOF–MS/MS accumulation time: 60 msMaximum candidate ions per cycle: 7Intensity threshold: 50 cpsCollision energy: 35 ± 15 VQ1 resolution: unitMS1 rate: 4 scans/sMS2 rate: 8 scans/s<b>Data Processing</b>Raw LC–MS/MS data were converted to <code>.mzML</code> using MSConvert (ProteoWizard 3.0.1908) and processed in MS-DIAL 4.38.<b>Peak detection and alignment:</b> performed using MS-DIAL.<b>Missing value filtering:</b> features with &gt;30% missing values overall or within any study group were removed.<b>Drift correction:</b> QC-based LOESS normalization using pooled QC samples injected every five samples, with additional QCs at the start and end of each batch.<b>Background filtering:</b> features retained if sample/blank intensity ratio &gt; 5.<b>Features annotations:</b> Features were annotated by MS and MS/MS spectral matching to HMDB, KEGG, and PubChem (similarity &gt;70%), yielding <i>putative identifications</i> consistent with MSI Level 2.<b>Output files:</b> Putative annotated and aligned peak tables as obtained from MSDIAL for both ESI⁺ and ESI⁻ modes.<b>Quality Control</b><b>Pooled QC:</b> used to assess analytical reproducibility and applied for normalization.<b>NIST SRM 1950:</b> served as a long-term performance reference across analytical batches.<b>Blank samples:</b> used to identify and filter out background or contaminant signals.<b>Ethics</b>All participants provided written informed consent. The study was approved by the Faculty of Health Sciences Human Research Ethics Committee, University of Cape Town (HREC Ref: 574/2018 and 061/2018).<b>Data Availability</b><b>Processed data:</b> MS-DIAL aligned and annotated peak tables (<code>.csv</code>) positive and negative mode.<b>Metadata:</b> includes sample information.