Changes in the rhizosphere metabolome profile of wastewater-irrigated lettuces (Lactuca sativa L.) highlighted using non-target HRMS and ROIMCR chemometrics method

• Impact of reclaimed water irrigation in lettuce rhizosphere using non-target metabolomics. • Simultaneous analysis of MS1 and MS2 signals in positive and negative ionization modes using ROIMCR. • Similar rhizosphere metabolomics profiling when irrigating with different types of water. • Significant changes in metabolite concentration in the rhizosphere between fall and summer. In recent decades, alternative water resources have been explored to address water scarcity; the use of reclaimed water in agricultural practices has been particularly studied. To evaluate the impact of different irrigation waters on lettuce cultures, 18 plants were grown across two seasons (fall and summer), using tap water as control, and two types of reclaimed water as substitute irrigation water sources. Samples were collected from the rhizosphere soil of the lettuces and were analyzed using Liquid Chromatography – High-Resolution Mass Spectrometry (LC-HRMS) with Data-Independent Acquisition (DIA). Non-target metabolomic profiling was performed using the Regions of Interest procedure (ROI), followed by Multivariate Curve Resolution-Alternating Least Squares (MCR-ALS) for simultaneous analysis of MS1 and MS2 signals in positive and negative ionization modes. In total, 37 chemical compounds (endogenous and exogenous) were identified belonging to different classes and origins, such as several sesquiterpene lactones or flavonoids that play a key role in the secondary metabolism of plants. Subsequent statistical and chemometric analyses highlighted that the sampling campaign season had a greater effect on the metabolomic profile than the type of water used for irrigation. Based on these findings, reclaimed waters did not exhibit sufficient influence on the rhizosphere to be discarded as a potential alternative for use in the agricultural industry.