School of Pharmacy, Shenyang Pharmaceutical University, Wenhua Road 103, Shenyang, Liaoning Province, PR China.
J Ethnopharmacol. 2020 May 23;254:112733. doi: 10.1016/j.jep.2020.112733. Epub 2020 Mar 4.
Zi-shen pill (ZSP), a traditional Chinese medicine, is widely used for the treatment of benign prostatic hyperplasia (BPH) and has remarkable curative effect.
To screen the potential 5-Lipoxygenase(5-LOX) inhibitors from ZSP extract.
A new approach based on affinity ultrafiltration-ultra performance liquid chromatography-mass spectrometry(UPLC-MS) was established and validated. Zileuton and glipizide were chosen as positive and negative control drug, respectively. For better screening result, the concentration of 5-LOX enzyme, incubation temperature and time, pH and ion strength were optimized. In addition, 5-LOX inhibitory assay in vitro and molecular docking technique were used for further verification.
20 compounds were characterized in the ultrafiltrate by high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and 16 ligands showed binding ability to 5-LOX. Among them, six ligands were deduced as high-potential 5-LOX inhibitors with their high specific binding values (>2.0). The inhibitory activities of anemarrhenasaponin I, timosaponin AI, nyasol and demethyleneberberine were confirmed by the 5-LOX inhibitory assay for validating the reliability of affinity ultrafiltration approach and the computer-simulated molecular docking technique further clarified the possible mechanism of action between the active compounds and the 5-LOX active sites.
紫参丸（ZSP）是一种中药，广泛用于治疗良性前列腺增生（BPH），具有显著的疗效。
从 ZSP 提取物中筛选潜在的 5-脂氧合酶（5-LOX）抑制剂。
建立并验证了一种基于亲和超滤-超高效液相色谱-质谱联用（UPLC-MS）的新方法。齐留通和格列吡嗪分别作为阳性和阴性对照药物。为了获得更好的筛选结果，优化了 5-LOX 酶浓度、孵育温度和时间、pH 值和离子强度。此外，还进行了体外 5-LOX 抑制试验和分子对接技术进一步验证。
通过高分辨率傅里叶变换离子回旋共振质谱（FT-ICR MS）对超滤物中的 20 种化合物进行了表征，其中 16 种配体显示出与 5-LOX 的结合能力。其中，6 种配体被推断为具有高潜力的 5-LOX 抑制剂，其高特异性结合值（>2.0）。通过 5-LOX 抑制试验证实了知母皂苷 I、知母皂苷 AI、菝葜皂苷元和去亚甲二氢小檗碱的抑制活性，验证了亲和超滤方法的可靠性，计算机模拟分子对接技术进一步阐明了活性化合物与 5-LOX 活性部位之间的可能作用机制。
