Chromatographic fingerprinting has been perceived as an essential tool for assessing quality and chemical equivalence of traditional Chinese medicine. However, this pattern-oriented approach still has some weak points in terms of chemical coverage and robustness. In this work, we proposed a multiple reaction monitoring (MRM)-based fingerprinting method in which approximately 100 constituents were simultaneously detected for quality assessment. The derivative MRM approach was employed to rapidly design MRM transitions independent of chemical standards, based on which the large-scale fingerprinting method was efficiently established. This approach was exemplified on QiShenYiQi Pill (QSYQ), a traditional Chinese medicine-derived drug product, and its robustness was systematically evaluated by four indices: clustering analysis by principal component analysis, similarity analysis by the congruence coefficient, the number of separated peaks, and the peak area proportion of separated peaks. Compared with conventional ultraviolet-based fingerprints, the MRM fingerprints provided not only better discriminatory capacity for the tested normal/abnormal QSYQ samples, but also higher robustness under different chromatographic conditions (i.e., flow rate, apparent pH, column temperature, and column). The result also showed for such large-scale fingerprints including a large number of peaks, the angle cosine measure after min-max normalization was more suitable for setting a decision criterion than the unnormalized algorithm. This proof-of-concept application gives evidence that combining MRM technique with proper similarity analysis metrices can provide a highly sensitive, robust and comprehensive analytical approach for quality assessment of traditional Chinese medicine.

色谱指纹图谱已被认为是评估中药质量和化学当量的重要工具。但是，这种面向模式的方法在化学覆盖率和鲁棒性方面仍存在一些弱点。在这项工作中，我们提出了一种基于多反应监测（MRM）的指纹方法，其中同时检测了约100种成分以进行质量评估。派生MRM方法用于独立于化学标准品快速设计MRM过渡，在此基础上有效地建立了大规模指纹识别方法。此方法已在以中药为原料的七神益气丸（QSYQ）上得到例证，并通过四个指标系统地评估了其稳健性：通过主成分分析进行聚类分析，通过同余系数，分离的峰数以及分离的峰的峰面积比例进行相似性分析。与常规的基于紫外线的指纹相比，MRM指纹不仅为测试的正常/异常QSYQ样品提供了更好的区分能力，而且在不同色谱条件（即流速，表观pH，柱温和色谱柱）下均具有更高的鲁棒性。结果还显示，对于包含大量峰的此类大规模指纹，与非归一化算法相比，最小-最大归一化后的角度余弦量度更适合于设置决策标准。