Nanomaterials (NMs) can be produced in plenty of variants posing several challenges for NM hazard and risk assessment. Metabolomic profiling of NM-treated cells and tissues allows for insights into underlying Mode-of-Action (MoA) and offers several advantages in this context. It supports the description of Adverse Outcome Pathways (AOPs) and, therefore, tailored AOP-based hazard testing strategies. Moreover, it bears great potential for biomarker discovery supporting toxicity prediction. Here, we applied metabolomics profiling to cells treated with four well-selected SiO₂ variants, differing in structure, size and surface charge. TiO₂ NM-105 served as a benchmark. Responses were studied in vitro in rat lung epithelial cells (RLE-6TN) and alveolar macrophages (NR8383) and compared to in vivo responses in rat lung tissues obtained from in vivo instillation and short-term inhalation studies (STIS). Time- and concentration-dependent changes were observed in both in vitro models but with cell-type specific responses. Overall, the levels of lipids and biogenic amines (BAs) tended to increase in epithelial cells but decreased in macrophages. Many identified metabolites like Met-SO, hydroxy-Pro and spermidine were related to oxidative stress, indicating that oxidative stress contributes to the MoA for the selected NMs. Several biomarker candidates such as Asp, Asn, Ser, Pro, spermidine, putrescine and LysoPCaC16:1 were identified in vitro and verified in vivo. In this study, we successfully applied a metabolomics workflow for in vitro and in vivo samples, which proved to be well suited to identify potential biomarkers, to gain insights into NM structure–activity relationship and into the underlying MoA.

纳米材料（NMs）的生产形式多样，对NM危害和风险评估提出了若干挑战。NM处理过的细胞和组织的代谢组学谱分析有助于深入了解潜在的作用模式（MoA），并在此方面提供了许多优势。它支持对不良结果途径（AOP）的描述，因此支持基于AOP量身定制的危害测试策略。此外，它在支持毒性预测的生物标志物发现方面具有巨大潜力。在这里，我们将代谢组学图谱应用于经过四个精心选择的SiO ₂变体处理的细胞，这些结构在结构，大小和表面电荷方面都不同。TiO ₂ NM-105为基准。反应进行了体外研究在大鼠肺上皮细胞（RLE-6TN）和肺泡巨噬细胞（NR8383）中进行了比较，并与通过体内滴注和短期吸入研究（STIS）获得的大鼠肺组织中的体内反应进行了比较。在两个体外模型中均观察到时间和浓度依赖性变化，但是具有细胞类型特异性反应。总体而言，脂质和生物胺（BAs）的水平在上皮细胞中倾向于增加，但在巨噬细胞中则降低。许多鉴定出的代谢产物（如Met-SO，羟脯氨酸和亚精胺）与氧化应激有关，表明氧化应激有助于所选NM的MoA。在体外鉴定了几种生物标志物候选物，如Asp，Asn，Ser，Pro，亚精胺，腐胺和LysoPCaC16：1并进行体内验证。在这项研究中，我们成功地将代谢组学工作流程应用于体外和体内样品，事实证明，该代谢组学工作流程非常适合识别潜在的生物标志物，以深入了解NM结构与活性之间的关系以及潜在的MoA。