Impact of protein binding on the analytical detectability and anticancer activity of thymoquinone.
Impact of protein binding on the analytical detectability and anticancer activity of thymoquinone.
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DOI：
10.1007/s12154-010-0052-4
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发表时间：
2011-07-01
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影响因子：
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通讯作者：
Vuorela, Heikki
中科院分区：
文献类型：
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作者：
El-Najjar, Nahed;Ketola, Raimo A;Vuorela, Heikki
未标记：百里醌（TQ），黑种草的活性成分，已知具有抗癌和抗炎作用;然而，尚未发表关于其在血清中的分析检测及其蛋白结合的研究。用高效液相色谱法测定血清中TQ的平均回收率，10 μ g/ml TQ为2.5%，100 μ g/ml TQ为72%。TQ从血清中的低回收率是由于其与血浆蛋白的广泛结合，因为超过99%的TQ在孵育30分钟内结合。研究了TQ与主要血浆蛋白牛血清白蛋白（BSA）和α-1酸性糖蛋白（AGP）的结合，发现BSA和AGP的结合率分别为94.5±1.7%和99.1±0.1%。质谱分析显示TQ共价结合到BSA上，特别是在Cyst-34上。使用WST-1增殖试验，我们表明BSA对TQ诱导的细胞死亡起保护作用;与BSA预孵育阻止TQ对DLD-1和HCT-116人结肠癌细胞发挥其抗增殖作用。另一方面，TQ与AGP的结合并不改变其对两种细胞系的抗增殖活性。当TQ在加入BSA之前与AGP预孵育时，TQ对DLD-1的活性保持不变，表明AGP阻止了TQ与BSA的结合。这是第一次记录BSA对TQ的共价结合和抑制作用。这些数据为TQ未来的体内药代动力学分析提供了新的依据。电子补充材料：本文的在线版本（doi：10.1007/s12154-010-0052-4）包含补充材料，可供授权用户使用。
UNLABELLED: Thymoquinone (TQ), an active component of Nigella sativa L., is known to have anti-cancer and anti-inflammatory effects; however, no studies on its analytical detection in serum and its protein binding have been published. Using high performance liquid chromatography analysis, we show that the average recovery of TQ from serum is 2.5% at 10mug/ml of TQ and 72% at 100mug/ml. The low recovery of TQ from serum is due to its extensive binding to plasma proteins, as more than 99% of TQ was bound within 30min of incubation. The binding of TQ to the major plasma proteins, bovine serum albumin (BSA) and alpha -1 acid glycoprotein (AGP), was studied and found to be 94.5±1.7% for BSA and 99.1±0.1% for AGP. Mass spectrometric analysis revealed that TQ was bound covalently to BSA, specifically on Cyst-34. Using WST-1 proliferation assay, we showed that BSA plays a protective role against TQ-induced cell death; pre-incubation with BSA prevented TQ from exerting its anti-proliferative effects against DLD-1 and HCT-116 human colon cancer cells. On the other hand, binding of TQ to AGP did not alter its anti-proliferative activity against both cell lines. When TQ was pre-incubated with AGP prior to the addition of BSA, the activity of TQ against DLD-1 was maintained, suggesting that AGP prevented the binding of TQ to BSA. This is the first time the covalent binding and inhibitory effect of BSA on TQ is documented. These data offer new grounds for TQ future pharmacokinetic analysis in vivo.ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s12154-010-0052-4) contains supplementary material, which is available to authorized users.