Phosphate photometrical determination

Photometric determinations with p-chloromercuribenzoate (3) were carried out in phosphate at pH 8 27 1SH groups were found per mole of unmodified and also of acetyl glutamate-activated carbamyl-P synthetase. With cold-inactivated enzyme, however, only 19 to 20 groups could be detected. The same results were obtained by amperometric titration in aqueous solution (2) and in 8M urea (5) after sulfitolysis on the cold-inactivated enzyme, an average of 23 SH groups per mole of enzyme was determined. The cases thus far examined, of substrate-induced enzyme inactivation, show also a change in SH group content (1, 14). 

L6. Leyton, L., Phosphate interference in the flame-photometric determination of calcium. Analyst 79, 497-500 (1954). 

Yl. Yofe, J., and Firrkelstein, R., Elimination of anionic interference in flame photometric determination of calcium in the presence of phosphate and sulfate. Anal. Chim. Acta 19, 166-173 (1958). 

Polyphosphonic acids can be separated on a special latex-based anion exchanger. They are photometrically determined after complexation with ferric nitrate [27] (see Section 3.3.5.2). This method enables a qualitative analysis of inorganic and organic phosphates that are contained in conditioners. Fig. 8-27 illustrates this with the chromatogram of a commercial product. Clearly, it is a multicomponent mixture with two main components and several minor components. 

A variant of the photometric method for determining chlorine dioxide in water using chlorophenol red (CPR method) was described by I.J. Fletcher and P. Hammings in ANALYST, June 1985. (Further literature on this method is also listed there.) They show how it is possible to optimize the procedure for determining chlorine dioxide in water and eliminate the effect of free available chlorine when using chlorophenol red by adding sodium cyclamate solution. A phosphate buffer is used to adjust to pH 7, after which a practically undisturbed reaction in the presence of thioacetamide permits the photometric determination of chlorine dioxide. 

Unsaturated lipids in blood serum can be spectro-photometrically determined by using sulfuric acid and vanillin phosphate, which produces a red substance of unknown composition whose absorbance is measured at 520-530 nm. 

Flame photometric determination cannot be recommended except for solutions containing upwards of 10 p.p.m. of magnesium because the resonance line occurs at 2852 A in the middle of an intense OH molecular band system and correction for this background is difficult even when a recording spectrophotometer is used. A hot flame is required and it has been reported by Knutson that some increase in sensitivity is obtained if an oxy-acetylene flame containing 55 per cent by volume of acetylene is used instead of the stoichiometric level of 29 per cent by volume. The resonance line suffers no serious spectral interference from other elements present in the sample solution but aluminium, chromium and molybdenum cause varying degrees of depression. Silicate, phosphate and sulphur ions depress the emission seriously and other anions less seriously and so standards should be made up to contain the same acids in the same concentrations as the samples. 

The insecticide fenitrothion (0,0-dimethyl-0-4-nitro-3-methylphenyl thio-phosphate) can be measured in sea water and sediments by gas chromatography, using a flame photometric detector to determine P and S [387]. The degradation products of the organophosphorus insecticides can be concentrated from large water by collection on Amberlite XAD-4 resin for subsequent analysis [383]. 

Ishikawa et al. [3] developed procedures to determine the trialkyl and triaryl phosphate esters in sediment, involving extraction with dichloromethane (for water) or acetone (for sediment), followed by gas chromatography using a flame photometric detector and gas chromatography-mass spectrometry after clean-up through a Florisil column. 

Solyom has conducted an intercalibration of methods used for the determination of phosphorus in sludges [37]. The methods used to determine phosphorus were that of Koroleff [83] in which the sample is digested with potassium peroxydisulphate and phosphate determined spectro-photometrically. Alternatively a reducing Kjeldahl digestion was used followed by determination of phosphate using molybdate and ascorbic acid. The former method gives somewhat low results. The reducing Kjeldahl method is therefore recommended. 

NADH is determined UV-photometrically as a measure of the amount of the substrate galactose-1-phosphate present in the reaction [14]. 

Z F6P + ATP + 2 NADH + H+ <-> 2 glycerol-3-phosphate + ADP + 2 NAD+ where F6P is fructose-6-phosphate, FDP is fructose-1,6-diphosphate, DHA-P is dihydroxyacetone phosphate, TIM is triosephosphate isomerase, and GDH is glycerol-3-phosphate dehydrogenase. The oxidation of NADH is a measure of the 6-PFK activity and is determined photometrically (decrease of OD per minute) [4]. 

Fig. 3. Trace determination of some inorganic and organic anions in pure water, after an electrophoretic enrichment at 5 kV for 45 s with an addition of 75 p,M octanesulfonate to the sample [42]. The electrolyte 10 mM sodium chromate and 0.5 mM OFM-BT (a surfactant used as the EOF modifier), adjusted to pH 8 with sulfuric acid 15 kV 60 cmX75 fjim I.D. capillary, distance to detector, 52 cm UV photometric detection at 254 nm. Anions (concentrations in mg/1) 1, chloride (3.5) 2, sulfate (4.8) 3, nitrate (6.2) 4, oxalate (5) 5, fluoride (1.9) 6, formate (5) 7, phosphate (3.2) 8, acetate (5) 9, propionate (5).

Phosphorus can serve as a benehcial adjunct or as a deleterious agent. There are several test methods for the determination of phosphorus. In addition to the three test methods described here, reference should also be made to multielement analysis methods such as inductively coupled plasma atomic emission spectroscopy (ICPAES) (ASTM D-4951, ASTM D-5185) and X-ray fluorescence (XRF) (ASTM D-4927, ASTM D-6443) described above in this guide. Phosphorus can also be determined by a photometric procedure (IP 148) or by a test method (ASTM D-1091) in which the organic material in the sample is destroyed, phosphorus in the sample is converted to phosphate ion by oxidation with sulfuric acid, nitric acid, and hydrogen peroxide, and the magnesium pyrophosphate is determined gravimetrically. Another method (ASTM D-4047, IP 149) in which the phosphorus is converted to quinoline phosphomolybdate is also available. 

Another procedure to determine oxidative phosphorylation of intact mitochondria or of submitochondrial particles is to measure, in a single photometric experiment, the respiration at 436 nm and the formation of NADPH at 334 nm. NADPH results from the action of hexokinase and glucose-6-phosphate dehydrogenase on ATP (the product of oxidative phosphorylation) and an excess of glucose and NADP+ (Figure 7). 

D.G. Themelis, A. Economou, A. Tsiomlektsis, P.D. Tzanavaras, Direct determination of phosphate in urine by sequential-injection analysis with single on-line dilution-calibration method and photometric detection, Anal. Biochem. 330 (2004) 193. 

The use of mass spectrometry as a detector in g.l.c. has the advantage that isomeric materials with the same retention times may still be distinguished, e.g. a phosphate and phosphonate. Mono-, di-, and tri-butyl phosphates may be separated by g.l.c. after conversion of the acidic components into their silyl esters. A dual flame photometric detector has been described for the simultaneous determination of phosphorus-, sulphur-, and chlorine-containing compounds. The method is based on the measurement of the... 

G. Wei and H. Ma, Determination of Total Phosphate in Samples from Trees by Flow Injection Analysis—Molybdenum Yellow Photometric Method [in Chinese]. Linye Daxue Xuebao, 3 (1986). 

A method of phosphate determination described in the German Standard Methods and in DIN 38405 similarly uses molybdate ions, in the presence of antimony ions, to form a complex which is reduced to phosphorus molybdenum blue by ascorbic acid and can be measured photometrically. Experience with this method has so far been good. 

The analysis of phosphorus in waters has historically been based on the photometric measurement of 12-phosphomolybdate or the phosphomolybdenum blue species, which are produced when phosphomolybdate is reduced. The majority of manual and automated methods of phosphate determination are based on the spectrophotometric determination of phosphorus as phosphomolybdenum blue, i.e.. 

Phosphate is mostly determined photometrically through the reaction with ammonium heptamolyb-date in acidic medium in the presence of potassium antimonyl tartrate as a catalyst. 12-Molybdophos-phoric acid is formed that upon reduction with, for example, ascorbic acid has a blue color. The lower limit of detection is lOpgl. Chromium(VI), nitrite, and high concentrations of chloride and iron interfere. The lower recoveries caused by iron are eliminated by addition of hydrogensulfite. 

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