Metabolomics and chemometrics approaches to investigate storage impact on phenolic compounds stability and their antioxidant activities in baby spinach

• Untargeted analysis of baby spinach putatively identified 117 metabolites, which shows the diversity of phytochemicals in baby spinach. • Refrigeration storage retains the maximum bioactive compounds, while storage at 25 °C significantly declines total phenolics, flavonoids, and antioxidant activities of baby spinach. • Chemometrics and kinetics analysis clearly show compositional transitions and degradation trends, suggesting flavonoids are broken down into simpler phenolic acids. Improper storage of baby spinach can affect the composition of its phenolic compounds. Most studies have examined only total phenolics or total flavonoids, without comprehensive compound-level analysis across various storage times and temperatures. To gain a comprehensive understanding of the impact of storage conditions, this study used a liquid chromatography mass spectrometry hyphenated electrospray ionisation and quadrupole time of flight (LC-ESI-QTOF-MS/MS) based metabolomics approach to achieve the phenolic profile of the baby spinach stored at 4 °C or 25 °C for nine days. Baby spinach stored at 4 °C showed minimal changes in TPC, TFC, and antioxidant activity, whereas storage at 25 °C for six days caused significant declines. We tentatively identified 117 metabolites, of which 23 were semi-quantified. It was further evidenced that betaine, 5,4′-dihydroxy-3,3′-dimethoxy-6,7-methylenedioxyflavone 4′-glucuronide, patuletin 3-(2′'-apiosylgentiobioside), rutin, and p -coumaric acid were the main contributors to baby spinach's bioactive potential. Phenolic acids like benzoic acid and p -coumaric acid were consistent at refrigeration temperature, potentially due to the degradation of flavonoids into these phenolic acids. Chemometric analysis (heatmap clustering and correlations) of metabolite degradation kinetics in baby spinach reveals a transition in the composition of bioactive compounds and their antioxidant activities during storage. This study helps us better understand changes in spinach phenolics during storage.