State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, Jiangxi 330047, China.
Analyst. 2010 Aug;135(8):2059-68. doi: 10.1039/c0an00161a. Epub 2010 Jun 11.
Beta-agonists such as ractopamine (RAC) and clenbuterol (CLEN), have similar effects as anabolic steroids i.e. they promote growth of muscular tissue and reduce body fat. They have been used successfully with animals and humans but have also been banned in many countries principally, because of their serious side effects. However, their illegal use persists. Thus, their interaction with biomolecules such as bovine serum albumin (BSA) is of significance, especially the co-operative reaction of mixed ligands with the protein. Fluorescence and UV-vis spectra of complex mixtures of individual ligands, binary and ternary complexes with BSA resulted in significantly overlapping spectral profiles. Qualitative and quantitative information about the various complex ligand-protein species formed, was obtained with the resolution of the excitation-emission fluorescence three-way data matrices by chemometrics methods-MCR-ALS and PARAFAC. Individual spectra of the ligands, their binary complexes with BSA and their ternary complexes were extracted, and quantitative concentration profiles for each species in a particular interaction were constructed. Such analyses made it possible to interpret the role and behaviour of each reaction component. It was found that both ligands, RAC and CLEN, bound co-operatively in site I of the BSA. This was confirmed with the use of site markers such as warfarin (site I) and ibuprofen (site II). However, CLEN formed a 1:1 CLEN-BSA complex, while RAC formed a 2:1 RAC(2)-BSA binary species. Interestingly, when CLEN or RAC was added to RAC(2)-BSA or CLEN-BSA, respectively, ternary complexes were produced such as RAC(2)-BSA-CLEN. Significantly, the presence of the second ligand in such an interaction in excess, appeared to increase the affinity of the added ligand for BSA. This may have consequences on the amount of steroid required to achieve a desired tissue growth effect.
β-激动剂，如莱克多巴胺（RAC）和克伦特罗（CLEN），具有与合成代谢类固醇类似的作用，即促进肌肉组织生长和减少体脂肪。它们在动物和人类中已成功使用，但也因严重的副作用而在许多国家被禁止。然而，它们的非法使用仍然存在。因此，它们与生物分子如牛血清白蛋白（BSA）的相互作用具有重要意义，特别是混合配体与蛋白质的协同反应。单独配体、二元和三元复合物与 BSA 的荧光和紫外可见光谱导致光谱轮廓显著重叠。通过化学计量学方法-MCR-ALS 和 PARAFAC-解析激发-发射荧光三向数据矩阵，获得了关于形成的各种配体-蛋白质配合物的定性和定量信息。提取了配体的个体光谱、它们与 BSA 的二元复合物及其三元复合物，并构建了每种特定相互作用中每种物质的定量浓度分布。这种分析使得解释每个反应成分的作用和行为成为可能。结果发现，两种配体 RAC 和 CLEN 均在 BSA 的 I 位协同结合。这一点通过使用华法林（I 位）和布洛芬（II 位）等位点标记物得到了证实。然而，CLEN 形成了 1:1 的 CLEN-BSA 复合物，而 RAC 形成了 2:1 的 RAC(2)-BSA 二元物种。有趣的是，当 CLEN 或 RAC 分别添加到 RAC(2)-BSA 或 CLEN-BSA 中时，会产生 RAC(2)-BSA-CLEN 等三元复合物。重要的是，在这种相互作用中过量存在第二种配体似乎增加了添加配体与 BSA 的亲和力。这可能会对达到所需组织生长效果所需的类固醇量产生影响。
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