目的 建立超高效液相色谱-（1，1-二苯基-2-三硝基苯肼）-二极管阵列检测器-电喷雾飞行时间质谱联用技术（UPLC-DPPH-PAD-ESI-TOF/MS）快速筛选鉴别天然产物中抗氧化成分的方法，并用于丹参中抗氧化成分的快速发现。方法 采用WatersACQUITY UPLC HSS T3柱（100 mm×3.0 mm，1.8 μm），以乙腈-0.2%甲酸水进行梯度洗脱，体积流量0.35 mL/min实现丹参提取物的快速分析。柱后流出液与另一路DPPH溶液（0.3 mL/min）通过三通阀注入PEEK反应线圈（内径0.25 mm，长度10 m）混合反应后流入检测器检测，检测波长为254和517 nm，通过517 nm下产生的负峰所对应的样品色谱峰来筛选丹参中的抗氧化成分。结果 采用UPLC技术在低体积流量下实现样品的高效分离，避免了高体积流量引起的基线噪音、显著提高了在线筛选方法的分辨率和灵敏度，降低了溶剂消耗。采用该方法在丹参提取物中共筛选出31个抗氧化活性成分，采用PAD-ESI-TOF/MS初步鉴定出11种，活性验证实验表明，咖啡酸、丹酚酸B、丹酚酸A、丹参素均具有较好抗氧化活性，其中，咖啡酸的DPPH半数清除浓度（IC₅₀）值为29.00 μmol/L，明显优于阳性对照维生素C。结论 该方法分离速度快、分辨率和灵敏度较高、溶剂消耗低，为复杂天然产物中抗氧化成分的快速发现研究提供了方法和技术支持。

Objective To establish a method for rapid screening and identifying antioxidants in natural products by ultra high performance liquid chromatography-(1,1-dipheny l-2-trinitrophenylhydrazine)-photo diode array-electrospray-time-of-flight mass spectrometry (UPLC-DPPH-PDA-ESI-TOF/MS) and apply it to the rapid discovery of antioxidants from Salvia miltiorrhiza extracts. Methods The Waters ACQUITY UPLC system and a Waters ACQUITY UPLC HSS T3 column (100 mm×3.0 mm, 1.8 μm; Waters Technologies, USA) were used to elute the sample at a low flow rate of 0.35 mL/min, which could achieve the rapid separation of S. miltiorrhiza extracts. The elution was joined with DPPH solution (0.3 mL/min) and reacted in the PEEK coil, and then flew into the detector. The anti-oxidation components in S. miltiorrhiza extracts were quickly screened by comparing the chromatograms at 517 nm and 254 nm. Results The UPLC technology was adopted to realize the efficient separation of samples at low volume flow rate, which avoided the baseline noise caused by high volume flow rate, significantly improved the resolution and sensitivity of the online screening method, and reduced the solvent consumption. The results showed that 31 antioxidant active compounds were screened from S. miltiorrhiza extracts, and 11 of them were identified by PAD-ESI-TOF/MS. Activity verification experiments showed that caffeic acid, salvianolic acid B, salvianolic acid A and danshensu had better antioxidant activity. The IC₅₀ value of caffeic acid was 29.00 μmol/L, which was significantly better than the positive control (vitamin C). Conclusion The method is rapid and efficient, which provides methods and technical support for the rapid discovery of antioxidant components from complex natural products.

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山东省自然基金资助项目（ZR2017LH071）；山东省自然基金资助项目（ZR2017LH069）；现代农业产业技术体系建设专项资金资助（CARS-21）；"十三五"重大专项生态种植项目（SQ2017YFC170295）；"十三五"重点研发计划（2017YFC1702701）：山东省泰山学者岗位资助项目（郭兰萍）；山东省科学院先导专项（郭兰萍）