scholarly journals A colorimetric determination of inositol monophosphates as an assay for d-glucose 6-phosphate–1l-myoinositol 1-phosphate cyclase

1970 ◽  
Vol 119 (2) ◽  
pp. 183-186 ◽  
Author(s):  
J. E. G. Barnett ◽  
R. E. Brice ◽  
D. L. Corina
Keyword(s):  
Inorganic Phosphate ◽  
Periodic Acid ◽  
Competitive Inhibitor ◽  
Phosphate Esters ◽  
Colorimetric Determination ◽  
Radiochemical Method ◽  
Chemical Assay

A rapid and convenient chemical assay for the enzyme d-glucose 6-phosphate–1l-myoinositol 1-phosphate cyclase is described. The 1l-myoinositol 1-phosphate formed enzymically was oxidized with periodic acid liberating inorganic phosphate, which was assayed. myoInositol 2-phosphate can be assayed in the same way. Glucose 6-phosphate and other primary phosphate esters gave only very small quantities of inorganic phosphate under the conditions described. The Km of the enzyme for d-glucose 6-phosphate, 7.5±2.5×10−4m, was identical with that measured by the radiochemical method. 2-Deoxy-d-glucose 6-phosphate was a powerful competitive inhibitor, Ki 2.0±0.5×10−5m, but was not a substrate for the enzyme.

COMPARISON OF VARIOUS METHODS FOR THE CHEMICAL ASSAY OF CORTICOIDS IN URINE

1955 ◽  
Vol 18 (4) ◽  
pp. 374-378
Author(s):  
Mogens Sprechler
Keyword(s):  
Calibration Curve ◽  
Periodic Acid ◽  
Reducing Power ◽  
Standard Technique ◽  
Phosphomolybdic Acid ◽  
Florisil Column ◽  
Chemical Assay ◽  
Chromatographic Procedure ◽  
Acid Reagent

SUMMARY Since 1949 about 10,000 urinary corticoid analyses have been performed routinely in our laboratory. The method used for this purpose was described in 1950 (Sprechler). We determine the corticoids which can be extracted from the urine with chloroform immediately after acidification to pH 1. The extract is washed with sodium hydroxide and water, a Girard separation is performed, and finally the reducing power of the ketonic fraction is measured by means of the phosphomolybdic acid reagent reaction. During the last few years two other chemical reactions have been used for comparison: The formaldehyde and the Porter-Silber method. After a thorough examination of the above methods a standard technique was followed. In the formaldehyde method a microdiffusion in a Conway unit was used instead of distillation of the formaldehyde following the oxidation with periodic acid. The calibration curve was corrected for loss of material by taking the standard doses of DOC through all the procedures of the method. A micromodification of the Porter-Silber method was chosen. Furthermore attempts were made to determine how specific the chromatographic procedure is in the determination of steroids in urinary extracts. For this purpose the Florisil column was used, and the technique described by Nelson & Samuels was followed. Finally we have investigated the glucuronide-bound corticoids in urine in a smaller series of objects.

Interference of phenothiazine compounds in the colorimetric determination of inorganic phosphate

1972 ◽  
Vol 47 (2) ◽  
pp. 329-336 ◽  
Author(s):  
Hamza F.A. El-Dorry ◽  
Heitor Medina ◽  
Metry Bacila
Keyword(s):  
Inorganic Phosphate ◽  
Colorimetric Determination

scholarly journals The inhibition of mitochondrial dicarboxylate transport by inorganic phosphate, some phosphate esters and some phosphonate compounds

1974 ◽  
Vol 138 (2) ◽  
pp. 171-175 ◽  
Author(s):  
R. N. Johnson ◽  
J. B. Chappell
Keyword(s):  
Succinate Dehydrogenase ◽  
Inorganic Phosphate ◽  
Potent Inhibitor ◽  
Competitive Inhibitor ◽  
Phosphate Esters ◽  
Alternative Substrate ◽  
Succinate Oxidation ◽  
Phenyl Phosphate ◽  
Dicarboxylate Transport ◽  
Phosphate Carrier

1. Pi competitively inhibited succinate oxidation by intact uncoupled mitochondria in the presence of sufficient N-ethylmaleimide to block the phosphate carrier, with a Ki of 2.5mm. 2. Of a large number of phosphate esters and phosphonate compounds, phenyl phosphate and phenylphosphonate were found to inhibit competitively uncoupled succinate oxidation by intact but not broken mitochondria. By comparison, benzoate was a relatively weak competitive inhibitor of succinate oxidation by intact mitochondria but a relatively potent inhibitor of succinate dehydrogenase. 3. Phenyl phosphate and phenylphosphonate were non-penetrant, and inhibited Pi-dependent swelling of mitochondria suspended in isosmolar ammonium malate in a manner non-competitive with Pi. The inhibitors did not affect mitochondrial swelling when tested with Pi alone. 4. It is concluded that: (i) phenyl phosphate and phenylphosphonate behaved as non-penetrant analogues of Pi, since their inhibitory properties were in strict contrast with those of benzoate; (ii) phenyl phosphate and phenylphosphonate interacted with the dicarboxylate carrier but not with the phosphate carrier; (iii) Pi was effective as a competitive inhibitor of succinate oxidation because of its being either an alternative substrate for the dicarboxylate carrier or competitive with succinate for the intramitochondrial cations as proposed by Harris & Manger (1968).

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