Determination of polyphosphates

This determination may be of interest in near-shore and estuarine water possibly contaminated with traces of polyphosphate-based detergents. In the method, any polyphosphate is converted into phosphate by acid hydrolysis. 

To 100 mL of the sample is added 0.5 mL of sulphuric add (reagent 1 of the phosphate determination. Section 10.2.5.4). Two 50 mL portions are taken from this solution and poured into two oxidation flasks. After heating and transferring to phosphate reaction flasks, add to both flasks 1 mL of ascorbic add solution (see Section 10.2.5.4 reagent 4) and to only cme flask 1 mL of the mixed reagent (see Section 10.2.5.4 reagent 6). 

Continue as for phosphate, but measure against a reference (or subtract the absorbance) of the sample with ascorbic acid only. 

This determination on an unfiltered sample yields the amount of total hydrolysable phosphorus (polyphosphate) plus inorganic phosphate in true solution and also the fraction of these ions adsorbed onto particles. The latter fraction is not included when analyses are performed using filtered samples. 

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The information given above concerning the chemical and physico-chemical properties of the inorganic polyphosphates will assist in the better understanding and prediction of the behaviour of these compounds during their extraction from cells and their subsequent fractionation. Knowledge of these properties will facilitate the development and use of efficient and reliable biochemical procedures for the isolation, purification, identification and determination of polyphosphates. 

Determination of polyphosphates using ion chromatography with suppressed conductivity detection. Application Note 71, Dionex, Sunnyvale, CA, USA, 2002. 

N. Yoza, Y. Kurokawa, Y. Hirai, and S. Ohashi, Flow Injection Determinations of Polyphosphates Based on Coloured Metal Complexes of Xy-lenol Orange and Methylthymol Blue. Anal. Chim. Acta, 121 (1980) 281. 

W.R. Biggs, J.T. Gano, and R.J. Brown. Determination of polyphosphate distribution by liquid chromatographic separation with direct current plasma-atomic emission spectrometric detection. Anal. Chem., 56(14), 2653 (1984). 

Ions which do not have a rare gas configuration appear not to fit into this rule. They are much more firmly bound than the alkali and alkaline earth cations. A dependence on radius is not observed and for Hg(I) the binding is stronger with the diphosphate anion than with triphosphate 366). (See, also the binding of cations by highmolccular polyphosphates, Section IV,E,/,d.) For the analysis and determination of triphosphates, see Section VII,B. 

Highly condensed polyphosphates, which cannot be separated by paper chromatography may be separated into groups by paper electrophoresis 255). Paper chromatography allows the complete separation and quantitative determination of all condensed phosphates with n up to 10, but its use is limited to 7-quantities. 

Robinson, N.A. Clark, J.E. Wood, H.G. Polyphosphate kinase from Pro-pionibacterium shermanii. Demonstration that polyphosphates are primers and determination of the size of the synthesized polyphosphate. J. Biol. Chem., 262, 5216-5222 (1987)... 

Tzeng, C.M. Kornberg, A. The multiple activities of polyphosphate kinase of Escherichia coli and their subunit structure determined by radiation target analysis. J. Biol. Chem., 275, 3977-3983 (2000)... 

The whole question of the specificity was reopened with the discovery that E. coli phosphatase, contrary to an earlier statement (114), hydrolyzed a variety of polyphosphates including metaphosphate of average chain length 8 (97). It was subsequently reported that partially purified phosphatases from several mammalian tissues had appreciable PPi-ase activity at pH 8.5 (115). This was confirmed (116) and extended to include ATPase and fluorophosphatase activities (117). Proof that the same enzyme is responsible for the monoesterase and PPi-ase activities was afforded by heat inactivation studies, cross inhibition experiments, and inhibition of PPi-ase activity by L-phenylalanine, a specific inhibitor of intestinal phosphatase. It was also found that calf intestinal phosphatase couid be phosphorylated by 32P-PP and the number of sites so labeled agreed with the number of active sites determined with a monoester substrate using a stopped-flow technique (118). It would seem that the main reason for the confusion with regard to the PPi-ase activity results from the inclusion of Mg2+ in the assay. This stimulates the monoesterase activity but almost completely inhibits PPi-ase activity (117). 

Figure 1.6 Changes in the viscosities of solutions of polyphosphates of different chain lengths on keeping for 12 h, where the abscissa represents the mean chain length as determined by end-group titration (a) immediately after solution (b) after keeping for 12 h (Strauss and Treitler, 1955b).

The first method of enzymatic PolyP assay was proposed by Clark et al. (1986). In this technique, PolyPs were determined by polyphosphate glucokinase obtained from Pro-pionibacterium shermanii. Glucose-6-phosphate dehydrogenase reduced NADP through utilization of the formed glucose-6-phosphate, and the increase in NADPH concentration was measured. 

J. E. Clark and H. G. Wood (1987). Preparation of standards and determination of sizes of long-chain polyphosphates by gel electrophoresis. Anal. Biochem., 161, 280-290. 

T. H. Kowalczyk and N. F. B. Phillips (1993). Determination of endopolyphosphatase using polyphosphate glucokinase. Anal. Biochem., 212, 194-205. 

B. Lorenz, J. Munkner, M. P. Oliveira, J. M. Leitao, W. G. E. Muller and H. C. Schroeder (1997a). A novel method for determination of inorganic polyphosphates using the fluorescent dye Fura-2. J.Anal. Biochem., 246, 176-184. 

Determination of the role of polyphosphate in transport-coupled phosphorylation in the yeast... 

If a water sample contains both soluble and insoluble manganese (Mn) compounds and ions, and it is filtered to separate the dissolved and insoluble fractions, and the filtrate and insoluble residue are analyzed separately, the results can be expressed as total dissolved Mn and total suspended or insoluble Mn. Phosphorus (P) can be determined colorimet-rically as the ortho-phosphate ion, P043, in aqueous samples after a reaction that forms an intensely blue-colored derivative. However polyphosphate ions and other ions and compounds containing P do not form this derivative. Total P in a sample can be determined with the same colorimetric procedure after acid hydrolysis and oxidation of all ions and compounds containing P to P04 3. In some elemental analyses the sample is treated with reagents designed to make available for measurement some fraction of an element or elements but not the total amount. For example, a soil sample may be treated with water at pH 3 to simulate the leaching process of acid rain. A total elemental analysis of the filtrate provides information about just those elements solubilized by the mild acid treatment. This can be called the determination of total mild acid leachable elements. 

Standard methods [2] used for the determination total phosphorus include several methods. Orthophosphates (PO and associated forms) are determined by colorimetry, ion chromatography or spectrophotometry (UV-visible). Acid hydrolysis allows the transformation of polyphosphates in orthophosphates, which are measured by one of the mentioned methods. Finally, a chemical digestion, followed by the determination of... 

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