Identification of Quality Markers in Viola kunawaresis Royel Using HPLC Fingerprints, Chemometric Analysis, and Network Pharmacology.

Violae tianshanicae herba (VTH), a widely used crude drug in Uyghur medicine in China, is renowned for its immunomodulatory, anti-inflammatory, antibacterial, antiviral, and antidiabetic activities, with notable efficacy in treating asthma. However, systematic research on its quality markers (Q-markers) remains limited, underscoring the need to analyze its chemical composition to enable effective quality control. A high-performance liquid chromatography (HPLC) fingerprint method was developed for 21 VTH batches in order to identify the common peaks of 23 flavonoids and esculetin, which were verified using liquid chromatography-mass spectrometry (LC-MS) and reference standards. In this study, chemometric methods were employed to differentiate samples from four geographical origins and to screen for differential components. In addition, a thorough examination of virtual target prediction in network pharmacology, in conjunction with molecular docking validation, has identified Q-markers with potential antiasthmatic effects. A group of six representative Q-markers (including kaempferol 3-O-sophorosyl-7-O-glucoside, esculetin, kaempferol 3-O-rutinosyl-7-O-glucoside, nicotiflorin, narcissoside, and astragalin) was selected for the purpose of developing a method to quantitatively analyze these components simultaneously in VTH samples. This study provides important references for the comprehensive quality control of VTH and establishes the foundation for researchers to explore the chemical basis and mechanisms of action of VTH.