THE SPECIFIC PRECIPITATION OF ORTHOPHOSPHATE AND SOME BIOCHEMICAL APPLICATIONS.
THE SPECIFIC PRECIPITATION OF ORTHOPHOSPHATE AND SOME BIOCHEMICAL APPLICATIONS.
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发表时间：
1964-07
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通讯作者：
Y. Sugino;Y. Miyoshi
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作者：
Y. Sugino;Y. Miyoshi
已经报道了从磷酸酯和酸酐中分离无机正磷酸盐的各种方法。磷钼酸的溶剂萃取法最初由Berenblum和Chain（1）介绍，已广泛用于生物材料的分析（2，3）。为了最大限度地减少溶剂萃取过程中液滴堵塞造成的繁琐和错误，Hagihara和Lardy（4）最近开发了一种采用反相色谱法去除溶剂可溶性磷钼酸的新方法。然而，几种材料的制备，例如用Al 2 O3盘密封的柱、硅化硅藻土、用高氯酸和硫酸钠溶液的混合物平衡的己醇甲醇混合物等，仍然或多或少是繁琐和耗时的。用CaCl*-Ca（OH）z（5）、CaCl *-醇（6）或“镁混合物”（7）沉淀正磷酸盐是有用的，特别是用于分析含有极端酸不稳定的磷酸盐化合物的材料，如磷酸肌酸和乙酰磷酸。然而，这些沉淀方法的缺点是灵敏度相对较低，并且沉淀所需的高盐浓度干扰了对残留在上清液级分中的磷酸盐化合物的分析。活性炭处理（8，9）可用于从各种无机盐（包括Pi）中分离核苷酸及其衍生物，但不能有效地从焦磷酸盐、糖磷酸盐和其他不吸附在活性炭上的磷酸盐衍生物中分离Pi。在用三乙基碳酸氢铵通过离子交换柱色谱法（DEAE-纤维素）从ZPi中分离 *PPi的过程中，我们观察到，在酸性条件下向非常稀的Pi溶液中加入碳酸氢铵时，形成重沉淀。然而，在相同条件下，用PPi没有形成沉淀。这一观察使我们发现三乙胺是磷钼酸的有效和选择性沉淀剂。本文描述了为确定沉淀Pi的最佳条件而进行的实验的结果，
Various procedures for the separation of inorganic orthophosphate from phosphoric acid esters and anhydrides have been reported. The solvent estraction method for phosphomolybdic acid, originally introduced by Berenblum and Chain (l), has been widely adopted for the analysis of biological materials (2, 3). In order to minimize the tedium and error due to the droplet occlusion during the solvent extraction, Hagihara and Lardy (4) have recently developed a new procedure employing reversed phase chromatography to remove solvent-soluble phosphomolybdie acid. However, preparation of several materials, such as columns sealed with Alundum disks, siliconized Celite, a hexanolmethanol mixture equilibrated with a mixture of perchloric acid and sodium sulfate solution, and others, is still more or less tedious and time consuming. Precipitation of orthophosphate with CaCl*-Ca(OH)z (5), CaCl,-alcohol (6), or “magnesium mixture” (7) is useful, especially for the analysis of materials that contain extremely acidlabile phosphate compounds, such as creatine phosphate and acetyl phosphate. However, these precipitation procedures have the disadvantage that the sensitivity is relatively low and the high salt concentration needed for precipitation interferes in the analysis of phosphate compounds remaining in the supernatant fraction. Charcoal treatment (8,9), which is useful in separating nucleotides and their derivatives from various inorganic salts, including Pi, is not effective in separating Pi from pyrophosphate, sugar phosphates, and other phosphate derivatives that are not adsorbed on charcoal. In the course of separating ‘*PPi from ZPi by ion exchange column chromatography (DEAE-cellulose) with triethylammonium bicarbonate, we observed that on adding ammonium molybdate under acidic conditions to a very dilute solution of Pi, a heavy precipitate was formed. However, no precipitate was formed with PPi under the same conditions. This observation led us to the finding that triethylamine is a potent and selective precipitant of phosphomolybdic acid. This paper describes the results of experiments performed to determine the optimal conditions for the precipitation of Pi by