Jiangsu Key Laboratory for TCM Formulae Research, Nanjing University of Chinese Medicine, Nanjing, China.
Peptides. 2010 May;31(5):786-93. doi: 10.1016/j.peptides.2010.02.016. Epub 2010 Mar 3.
Cornu Bubali (water buffalo horn, WBH) has been used in Traditional Chinese Medicine (TCM) for thousands of years. In the present study, three peptides with antioxidant properties were purified from aqueous extract of Cornu Bubali (water buffalo horn, WBH) by consecutive chromatographic methods including gel filtration chromatography, ion-exchange chromatography and high performance liquid chromatography. The sequences of the three peptides were identified to be Gln-Tyr-Asp-Gln-Gly-Val (WBH-1, 708Da), Tyr-Glu-Asp-Cys-Thr-Asp-Cys-Gly-Asn (WBH-2, 1018Da) and Ala-Ala-Asp-Asn-Ala-Asn-Glu-Leu-Phe-Pro-Pro-Asn (WBH-3, 1271Da) by matrix assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry (MALDI-LIFT-TOF/TOF MS). The antioxidant activity of these peptides was tested using 1-diphenyl-2-picrylhydrazyl (DPPH) scavenging assay directly. Methylthiazol tetrazolium (MTT) assay and lactate dehydrogenase (LDH) release assay were also used to evaluate the protection of peptides against hydrogen peroxide (H(2)O(2)) induced injury. The results showed that these peptides could reduce the DPPH radical and protect rat cerebral microvascular endothelial cells (rCMECs) against H(2)O(2)-induced injury, thus demonstrating that these peptides had antioxidant activity. These results suggest that WBH-1, WBH-2 and WBH-3, isolated from the aqueous extract of water buffalo horn are natural antioxidants and may contribute to the efficacy of WBH.
水牛角（Cornu Bubali，WBH）在中医药中已有数千年的应用历史。在本研究中，通过连续的色谱方法，包括凝胶过滤色谱、离子交换色谱和高效液相色谱，从水牛角的水提物中纯化出具有抗氧化活性的三种肽。通过基质辅助激光解吸电离飞行时间/飞行时间质谱（MALDI-LIFT-TOF/TOF MS）鉴定这三种肽的序列分别为 Gln-Tyr-Asp-Gln-Gly-Val（WBH-1，708Da）、Tyr-Glu-Asp-Cys-Thr-Asp-Cys-Gly-Asn（WBH-2，1018Da）和 Ala-Ala-Asp-Asn-Ala-Asn-Glu-Leu-Phe-Pro-Pro-Asn（WBH-3，1271Da）。直接使用 1,1-二苯基-2-苦基肼基（DPPH）清除实验来测试这些肽的抗氧化活性。还使用甲基噻唑四唑（MTT）测定法和乳酸脱氢酶（LDH）释放测定法来评估肽对过氧化氢（H2O2）诱导损伤的保护作用。结果表明，这些肽可以清除 DPPH 自由基，保护大鼠脑微血管内皮细胞（rCMECs）免受 H2O2 诱导的损伤，从而表明这些肽具有抗氧化活性。这些结果表明，从水牛角的水提物中分离出的 WBH-1、WBH-2 和 WBH-3 是天然抗氧化剂，可能对 WBH 的功效有贡献。
