Mechanism of fructosamine assay: evidence against role of superoxide as intermediate in nitroblue tetrazolium reduction

Baker, John; Zyzak, David V.; Thorpe, Suzanne R.; Baynes, John W.

doi:10.1093/clinchem/39.12.2460

Cited by 38 publications

(12 citation statements)

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“…A proposed direct interaction between reduced quinone species such as LY 83583 and NBT resulting in formazan production. demonstrated in some assays using NBT as a final electron acceptor (Baker et al, 1993). The effect of SOD instead suggests that superoxide acts as the intermediary between reduced LY 83583, which undergoes auto-oxidation by molecular 02, and NBT reduction ( Fig.…”

Section: Discussionmentioning

confidence: 98%

Histochemical Detection of Quinone Reductase Activity In Situ Using LY 83583 Reduction and Oxidation

Murphy¹,

Vincent³

1998

Journal of Neurochemistry

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The application of enzymatic staining techniques, using tetrazolium dyes, to aldehyde‐treated brain sections has revealed the presence of NADPH‐diaphorase activity attributed to nitric oxide synthase. When evaluating the specificity of the putative guanylyl cyclase inhibitor LY 83583, a robust and novel staining pattern was noted in epithelial, endothelial, and astrocytic cells when LY 83583 was included in the NADPH‐diaphorase histochemical reaction. This LY 83583‐dependent staining could be blocked by the NAD(P)H:quinone oxidoreductase inhibitor dicumarol. Based on its quinone structure, we hypothesized that LY 83583 was a substrate for the enzyme NAD(P)H:quinone oxidoreductase. Transfection of human embryonic kidney 293 cells with the rat liver isoform of NAD(P)H:quinone oxidoreductase resulted in robust NADPH‐ and LY 83583‐dependent staining that was completely blocked by dicumarol and was not observed in untransfected cells. Analysis of transfected cell extracts and brain homogenates indicated that LY 83583 was a substrate for NAD(P)H:quinone oxidoreductase, with a Km similar to the well‐characterized substrate menadione. Sensitivity of the nitroblue tetrazolium reduction to superoxide dismutase indicated that the reduction of LY 83583 by NAD(P)H:quinone oxidoreductase leads to superoxide generation. The localization of NAD(P)H:quinone oxidoreductase activity to astrocytic cells suggests a role for glia in combating oxidative insults to brain and in activating quinone‐like drugs such as LY 83583.

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Section: Discussionmentioning

confidence: 98%

Histochemical Detection of Quinone Reductase Activity In Situ Using LY 83583 Reduction and Oxidation

Murphy¹,

Vincent³

1998

Journal of Neurochemistry

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“…Protein carbonyls were quantitated by spectrophotometric measurement of their 2,4 dinitrophenylhydrazone derivatives ( ε 370 nm = 22000 M-1 cm-1), while nitrotyrosine (NTY) was measured according to its spectral contribution at 370 nm. Fructosamine assay was based on nitroblue tetrazolium (NBT) reduction to the purple monoformazan dye in alkaline conditions, where the absorbance change at 530 nm was measured in the intervals of 10 and 15 min after the start of the reaction at 37 ° C. The corrected albumin fructosamine value was expressed as the amount of fructosamine per gram of proteins, taking into consideration the variation in protein concentrations in plasma samples [29] . Plasma RNase activity against double-stranded forms poly (I:C) and poly(A:U) was measured in heparinised plasma according to the methods explained in our previous study [33] .…”

Section: Methodsmentioning

confidence: 99%

Possible Impact of Impaired Double-stranded RNA Degradation and Nitrosative Stress on Immuno-inflammatory Cascade in Type 2 Diabetes

Kocić¹,

Kocić²,

Pavlović³

et al. 2009

Exp Clin Endocrinol Diabetes

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The immune response can be triggered by molecules derived from microorganisms (PAMP) or from molecules derived from damaged or dead host cells, known as the damage-associated molecular-pattern molecules (DAMP). Their immune effects are accompanied by altered redox environment. The level of stable end products of nitric oxide (NO)- plasma nitrate and nitrite (NOx), carbonyl groups (PCO) and nitrotyrosine (NTY), in relation to the metabolism of dsRNAs (poly I:C and poly A:U) and xanthine oxidase (XO activity), in plasma of type2 diabetic patients was determined. Thirty-six patients with type 2 diabetes (age group 34-66 years, 19 male and 17 female) were allocated to the study. Diabetic patients had a significantly higher level of plasma NOx products, NTY and PCO, fructosamine (FA) and XO activity indicating about altered redox environment. The concentration of circulating ribonucleic acids (CNAs) was significantly higher in type 2 diabetic patients, which was accompanied by a significantly decreased activity of RNase against double stranded RNA forms (poly I:C and poly A:U), compared to control samples. To determine whether CNAs, as possible DAMP molecules, are capable of exerting effect on inflammatory and host antiviral response, the effect of isolated CNAs on NF-kappaB, Bcl-2, Bax, MDA-5 and IRF-3 regulation was evaluated in culture of fresh isolated thymocytes. Circulating nucleic acids isolated from type 2 diabetic patients were able to upregulate NF-kappaB more than control RNA samples. In the same experimental conditions the mild Bcl-2 upregulation, followed by the marked Bax upregulation, was demonstrated. Since the Bcl-2/Bax ratio was lower in type 2 diabetic samples, obtained results may implicate that CNAs may exert proapoptotic response in type 2 diabetes. The CNAs isolated from diabetic patients were able to downregulate MDA-5 and IRF-3, very important subjects of the surveillance and cellular anti-viral response. The major findings of the present study are that impaired dsRNA metabolism may lead to increased level of different sized RNAs in type 2 diabetic patients. Acting as possible DAMP molecules, they may contribute to higher susceptibility of immune cells to inflammatory cascade via NF-kappaB activation, and possible MDA-5/IRF-3 axis downregulation, what may have an influence on further ineffective response against different pathogens.

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“…Non‐glycated BSA was also prepared using the same method except for the addition of glucose. The amount of fructosamine in the synthesized glycated BSA was determined with the alkaline‐nitroblue tetrazolium method using a LiquiTech fructosamine kit (Roche, Basel, Switzerland).…”

Section: Methodsmentioning

confidence: 99%

Electrochemical sensing system employing fructosamine 6‐kinase enables glycated albumin measurement requiring no proteolytic digestion

Kameya

Tsugawa

Yamada-Tajima

et al. 2016

Biotechnology Journal

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Currently available enzymatic methods for the measurement of glycated proteins utilize fructosyl amino acid/peptide oxidases (FAOXs/FPOXs) as sensing elements. FAOXs/FPOXs oxidize glycated amino acids or glycated dipeptides but they are not able to accept longer glycated peptides or intact glycated proteins as substrates. Therefore, pretreatment via proteolytic digestion is unavoidable with the current enzymatic methods, and there remains a need for simpler measurement methods for glycated proteins. In this study, in order to develop a novel sensing system for glycated albumin (GA), a marker for diabetes, with no requirement for proteolytic digestion, we created an electrochemical sensor based on fructosamine 6-kinase (FN6K) from Escherichia coli. Uniquely, FN6K can react directly with intact GA unlike FAOXs/FPOXs. The concentration of GA in samples was measured using a carbon-printed disposable electrode upon which FN6K as well as two additional enzymes, pyruvate kinase and pyruvate dehydrogenase were overlaid. A clear correlation between the response current and the concentration of GA was observed in the range of 20-100 µM GA, which is suitable for measurement of GA in diluted blood samples from both healthy individuals and patients with diabetes. The sensing system reported here could be applied to point-of-care-testing devices for measurement of glycated proteins.

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Mechanism of fructosamine assay: evidence against role of superoxide as intermediate in nitroblue tetrazolium reduction

Cited by 38 publications

References 0 publications

Histochemical Detection of Quinone Reductase Activity In Situ Using LY 83583 Reduction and Oxidation

Histochemical Detection of Quinone Reductase Activity In Situ Using LY 83583 Reduction and Oxidation

Possible Impact of Impaired Double-stranded RNA Degradation and Nitrosative Stress on Immuno-inflammatory Cascade in Type 2 Diabetes

Electrochemical sensing system employing fructosamine 6‐kinase enables glycated albumin measurement requiring no proteolytic digestion

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