Texas A&M University, Department of Chemistry, College Station, TX 77843, USA.
Metallomics. 2013 Mar;5(3):232-41. doi: 10.1039/c3mt00009e.
The presence of labile low-molecular-mass (LMM, defined as <10 kDa) metal complexes in cells and super-cellular structures such as the brain has been inferred from chelation studies, but direct evidence is lacking. To evaluate the presence of LMM metal complexes in the brain, supernatant fractions of fresh mouse brain homogenates were passed through a 10 kDa cutoff membrane and subjected to size-exclusion liquid chromatography under anaerobic refrigerated conditions. Fractions were monitored for Mn, Fe, Co, Cu, Zn, Mo, S and P using an on-line ICP-MS. At least 30 different LMM metal complexes were detected along with numerous P- and S- containing species. Reproducibility was assessed by performing the experiment 13 times, using different buffers, and by examining whether complexes changed with time. Eleven Co, 2 Cu, 5 Mn, 4 Mo, 3 Fe and 2 Zn complexes with molecular masses <4 kDa were detected. One LMM Mo complex comigrated with the molybdopterin cofactor. Most Cu and Zn complexes appeared to be protein-bound with masses ranging from 4-20 kDa. Co was the only metal for which the "free" or aqueous complex was reproducibly observed. Aqueous Co may be sufficiently stable in this environment due to its relatively slow water-exchange kinetics. Attempts were made to assign some of these complexes, but further efforts will be required to identify them unambiguously and to determine their functions. This is among the first studies to detect low-molecular-mass transition metal complexes in the mouse brain using LC-ICP-MS.
细胞和超细胞结构（如大脑）中不稳定的低分子量（LMM，定义为 <10 kDa）金属配合物的存在已从螯合研究中推断出来，但缺乏直接证据。为了评估大脑中是否存在 LMM 金属配合物，将新鲜小鼠脑匀浆的上清液通过 10 kDa 截止膜，并在厌氧冷藏条件下进行大小排阻液相色谱。使用在线 ICP-MS 监测 Mn、Fe、Co、Cu、Zn、Mo、S 和 P 的馏分。至少检测到 30 种不同的 LMM 金属配合物，以及许多含有 P 和 S 的物质。通过使用不同的缓冲液进行 13 次实验，以及检查配合物是否随时间变化来评估重现性。检测到 11 种 Co、2 种 Cu、5 种 Mn、4 种 Mo、3 种 Fe 和 2 种 Zn 配合物，分子量 <4 kDa。一种 LMM Mo 配合物与钼喋呤辅因子共迁移。大多数 Cu 和 Zn 配合物似乎与蛋白质结合，分子量范围为 4-20 kDa。Co 是唯一可重复观察到“游离”或水合配合物的金属。由于其相对较慢的水交换动力学，水溶液中的 Co 可能在此环境中足够稳定。尝试对其中一些配合物进行了分配，但需要进一步努力才能明确识别它们并确定它们的功能。这是使用 LC-ICP-MS 首次在小鼠大脑中检测到低分子量过渡金属配合物的研究之一。
