A malachite green procedure for orthophosphate determination and its use in alkaline phosphatase-based enzyme immunoassay

Baykov, Alexander A.; Evtushenko, Olga A.; Avaeva, S.M.

doi:10.1016/0003-2697(88)90484-8

Cited by 769 publications

(631 citation statements)

References 9 publications

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“…Hence a study of the estimation process was done whereby the preconcentration of Fe and PO4 ions was carried out by the protonation of the samples using various concentrations of HCl and sulphuric acid. The use of sulphuric acid and hydrochloric acid in the estimation of PO4 and Fe respectively is traditional [27][28][29][30][31][32][33] but our experiments confirmed that the concentration of acids used in the conventional methods is insufficient to break the strong Fe-O-P bond as hydrochloric acid used is mostly for acid hydrolysis in the estimation of iron. Fe from various sources like plant body, the human body, drinking water or any other source is estimated by using HCl for the process of acid hydrolysis.…”

Section: Resultsmentioning

confidence: 59%

INVESTIGATION AND ALLEVIATION OF MUTUAL INTERFERENCE OF Fe2+AND PO43-IONS BY PROTONATION

2017

RJC

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Mutual interference of iron and phosphates in the spectrophotometric estimation process is a result of the strong bond in the iron phosphate complex. The interference of these two ions can be successfully overcome by the isolation of the two ions. The present investigation was carried to study the mutual interferences of iron and phosphates in the spectrophotometric analysis. Complexing both Fe and PO4 ions in the form of Fe-P complex were done for isolating the Iron and phosphate radicals from other interferences. Whereas to avoid the interference of Fe in PO4 estimation, AlPO4 complex was formed. This triggered the idea to pre-concentrate the two ions by protonation using a strong acid. The results obtained justified the need for isolation of Fe 2+ and PO4 3-.This study provides an exact estimation of Fe 2+ at pH 1.5. The results obtained were statistically correlated by Pearson's correlation coefficient and the results were statistically significant at p value 0.01.

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Section: Resultsmentioning

confidence: 59%

INVESTIGATION AND ALLEVIATION OF MUTUAL INTERFERENCE OF Fe2+AND PO43-IONS BY PROTONATION

2017

RJC

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“…The activities of CaMKP, CaMKP-N, and PP2C were determined using phosphopeptide pp10 (YGGMHRQET(p)VDC) as a substrate [19,20].…”

Section: Production Of Antibodiesmentioning

confidence: 99%

Inhibitors of the Ca2+/calmodulin-dependent protein kinase phosphatase family (CaMKP and CaMKP-N)

Sueyoshi

Takao

Nimura

et al. 2007

Biochemical and Biophysical Research Communications

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multifunctional CaMKI, II, and IV. However, the lack of specific inhibitors of these phosphatases has hampered studies on these enzymes in vivo. In an attempt to obtain specific inhibitors, we searched inhibitory compounds and found that Evans Blue and Chicago Sky Blue 6B served as effective inhibitors for CaMKP. These compounds also inhibited CaMKP-N, but inhibited neither protein phosphatase 2C, another member of PPM family phosphatase, nor calcineurin, a typical PPP family phosphatase. The minimum structure required for the inhibition was 1-amino-8-naphthol-4-sulfonic acid.When Neuro2a cells cotransfected with CaMKIV and CaMKP-N were treated with these compounds, the dephosphorylation of CaMKIV was strongly suppressed, suggesting that these compounds could be used as potent inhibitors of CaMKP and CaMKP-N in vivo as well as in vitro.

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“…The reaction was stopped by adding 20 pA of 50% trichloroacetic acid and then centrifuged at 12 500 × g for 10 min. Aliquots of the supernatant (10-20 Izl) were withdrawn, diluted to 300 ixl with distilled water and then 75 Ixl of the Malachite green reagent [10] were added in order to determine the released inorganic phosphate. A Pi calibration curve was determined by using standard solutions of potassium dihydrogenphosphate.…”

Section: Polyacrylamide Gel Electrophoresismentioning

confidence: 99%

“…The developed colour was read at 600 nm using an EIA reading photometer. The release of Pi from D-myo-inositol 1,4-bisphosphate, D-myo-inositol 1,4,5-triphosphate, [3-glycerophosphate, T-AMP, and 2'-GMP was assayed by the Malachite green reagent [10], as described for myo-inositol l-phosphate. The determination of K m and Vma x on myo-inositol 1-phosphate was performed by measuring the initial hydrolysis rates by the method described by Gee et al [11].…”

Section: Polyacrylamide Gel Electrophoresismentioning

confidence: 99%

Identity of zinc ion-dependent acid phosphatase from bovine brain and myo-inositol 1-phosphatase

Caselli

Cirri

Bonifacio

et al. 1996

Biochimica et Biophysica Acta (BBA) - General Subjects

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A 62 kDa Zn2+-dependent acid phosphatase has been purified from bovine brain. The protein was carboxymethylated and then cleaved by endoproteinase GIu-C, trypsin and CNBr. Several fragments were subjected to structural analysis either by using mass spectrometry or automated peptide sequencing. The four sequenced peptides were compared with the known protein sequences contained in the EMBL Data Bank. All four peptide sequences were identical to the corresponding amino-acid sequences present in myo-inositol 1-phosphatase from bovine brain. Furthermore we found that the amino-acid composition of Zn2+-dependent acid phosphatase purified in our laboratory is very similar to that of myo-inositol 1-phosphatase, and that several peptide fragments have molecular weights (measured by mass spectrometry techniques) identical to those expected for cleavage-fragments originated from the authentic myo-inositol l-phosphatase. This is one of the key enzymes in the receptor-stimulated inositol phospholipid metabolism and it has been considered as the probable target of Li ÷ ion during LiC1 therapy in manic-depressive patients. The comparison of the Zn2+-dependent acid phosphatase and the Mg 2 +-dependent myo-inositol-1-phosphatase activities, measured at different purification steps, shows that the ratio between the two activities was remarkably constant during enzyme purification. We also demonstrated that in the presence of Mg 2+ this enzyme efficiently catalyses the hydrolysis of myo-inositol l-phosphate, and that the Li + ion inhibits this activity. Furthermore, the thermal treatment of the enzyme causes a time-dependent parallel decrease of both Zn-dependent p-nitrophenyl phosphatase (assayed at pH 5.5) and Mg2+-dependent myo-inositol-l-phosphatase (assayed at pH 8.0) activities, suggesting the hypothesis that the same protein possesses both these activities.

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A malachite green procedure for orthophosphate determination and its use in alkaline phosphatase-based enzyme immunoassay

Cited by 769 publications

References 9 publications

INVESTIGATION AND ALLEVIATION OF MUTUAL INTERFERENCE OF Fe2+AND PO43-IONS BY PROTONATION

INVESTIGATION AND ALLEVIATION OF MUTUAL INTERFERENCE OF Fe2+AND PO43-IONS BY PROTONATION

Inhibitors of the Ca2+/calmodulin-dependent protein kinase phosphatase family (CaMKP and CaMKP-N)

Identity of zinc ion-dependent acid phosphatase from bovine brain and myo-inositol 1-phosphatase

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