Huskens J, Main M, Malloy C R, Sherry A D
University of Texas at Dallas, Department of Chemistry, Richardson 75083-0688, USA.
Biochim Biophys Acta. 1997 Oct 20;1336(3):434-44. doi: 10.1016/s0304-4165(97)00054-8.
The ligand 1,4,7-triazacyclononane-1,4,7-tris(methylene methylphosphinic acid), NOTMP, was used to measure free MgII levels in blood plasma by 31P MRS. Separate resonances were observed for the free ligand and the MgII complex and the ratio of their resonance areas was used to evaluate the free, ionized MgII concentration, [Mg]free. The CaII and the ZnII complexes gave rise to separate resonances in the 31P spectrum in an aqueous sample. In human blood plasma samples, however, these resonances were never observed thus excluding the interference of these metal ions. Heparin, up to 150 units/ml, had no influence on the Mg-NOTMP equilibrium. The 31P MRS methodology was applied to twenty human blood plasma samples. Total MgII ([Mg]total), as measured by atomic absorption spectroscopy, averaged 0.85 +/- 0.12 mM while free ionized MgII ([Mg]free) measured by 31P MRS was 0.66 +/- 0.09 mM. The 31P MRS method gave inherently larger values for free ionized MgII than that reported by ion-selective electrodes (ISE). This was traced to a redistribution of existing plasma MgII species after the addition of about 2 mM of NOTMP. Calculations using existing thermodynamic data show that the ionized MgII concentration (iMg) and the concentration of MgII weakly complexed to small anions (Mg(comp)) both drop after the addition of NOTMP, with Mg(comp) dropping to negligible levels. Thus, the 31P MRS method appears to be less sensitive to variations in the concentration of weakly binding anions (bicarbonate, carbonate, chloride, lactate, phosphate, etc.) than the ISE method. Our data indicates that the difference between Mg(total), as measured by atomic absorption spectroscopy, and Mg(free), as measured by 31P MRS, provides an direct estimate of the protein bound MgII fraction.
配体1,4,7-三氮杂环壬烷-1,4,7-三（亚甲基甲基膦酸），即NOTMP，用于通过31P磁共振波谱法测量血浆中的游离MgII水平。观察到游离配体和MgII络合物有单独的共振峰，其共振峰面积之比用于评估游离的、离子化的MgII浓度[Mg]free。CaII和ZnII络合物在水性样品的31P谱中产生单独的共振峰。然而，在人体血浆样品中从未观察到这些共振峰，因此排除了这些金属离子的干扰。高达150单位/毫升的肝素对Mg-NOTMP平衡没有影响。31P磁共振波谱法应用于20份人体血浆样品。通过原子吸收光谱法测得的总MgII（[Mg]total）平均为0.85±0.12 mM，而通过31P磁共振波谱法测得的游离离子化MgII（[Mg]free）为0.66±0.09 mM。31P磁共振波谱法测得的游离离子化MgII值比离子选择性电极（ISE）报道的值固有地更大。这可追溯到加入约2 mM的NOTMP后现有血浆MgII物种的重新分布。使用现有热力学数据进行的计算表明，加入NOTMP后，离子化的MgII浓度（iMg）和与小阴离子弱络合的MgII浓度（Mg(comp)）均下降，Mg(comp)降至可忽略不计的水平。因此，31P磁共振波谱法似乎对弱结合阴离子（碳酸氢根、碳酸根、氯离子、乳酸根、磷酸根等）浓度变化的敏感性低于ISE法。我们的数据表明，通过原子吸收光谱法测得的Mg(total)与通过31P磁共振波谱法测得的Mg(free)之间的差异可直接估计与蛋白质结合的MgII部分。
