Table_1_Screening and Identification of Cardioprotective Compounds From Wenxin Keli by Activity Index Approach and in vivo Zebrafish Model.DOCX

Wenxin Keli (WXKL) is a widely used Chinese botanical drug for the treatment of arrhythmia, which is consisted of four herbs and amber. In the present study, we analyzed the chemical composition of WXKL using liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) to tentatively identify 71 compounds. Through typical separate procession, the total extract of WXKL was divided into fractions for further bioassays. Cardiomyocytes and zebrafish larvae were applied for assessment. In vivo arrhythmia model in Cmlc2-GFP transgenic zebrafish was induced by terfenadine, which exhibited obvious reduction of heart rate and occurrence of atrioventricular block. Dynamic beating of heart was recorded by fluorescent microscope and sensitive camera to automatically recognize the rhythm of heartbeat in zebrafish larvae. By integrating the chemical information of WXKL and corresponding bioactivities of these fractions, activity index (AI) of each identified compound was calculated to screen potential active compounds. The results showed that dozens of compounds including ginsenoside Rg1, ginsenoside Re, notoginsenoside R1, lobetyolin, and lobetyolinin were contributed to cardioprotective effects of WXKL. The anti-arrhythmic activities of five compounds were further validated in larvae model and mature zebrafish by measuring electrocardiogram (ECG). Our findings provide a successful example for rapid discovery of bioactive compounds from traditional Chinese medicine (TCM) by activity index based approach coupled with in vivo zebrafish model.

文心颗粒（WXKL）作为一种广泛应用于治疗心律失常的中医草本药物，其组成为四种草药与琥珀。在本研究中，我们采用液相色谱联用高分辨率质谱（LC-HRMS）技术对WXKL的化学成分进行了分析，以初步鉴定出71种化合物。通过常规分离处理，WXKL的总提取物被分成若干部分，以进行进一步的生物活性检测。心肌细胞和斑马鱼幼鱼被用于评估。通过诱导Cmlc2-GFP转基因斑马鱼的体内心律失常模型，使用特非那定诱导，表现为心率显著降低和房室传导阻滞。采用荧光显微镜和敏感相机记录心脏的动态跳动，以自动识别斑马鱼幼鱼的心跳节律。通过整合WXKL的化学信息和相应分部的生物活性，计算了每个鉴定化合物的活性指数（AI），以筛选潜在的活性化合物。结果显示，包括人参皂苷Rg1、人参皂苷Re、三七皂苷R1、洛贝林和洛贝林素在内的数十种化合物对WXKL的心脏保护作用做出了贡献。五种化合物的抗心律失常活性通过测量电生理图（ECG）在幼虫模型和成熟斑马鱼中进一步得到验证。我们的发现为通过活性指数方法结合体内斑马鱼模型，从传统中医药（TCM）中快速发现生物活性化合物提供了一个成功的范例。