Fluorometric Microdetermination of Alpha-Keto Acids.

Spikner, J. E.; Towne, Jack C.

doi:10.1021/ac60191a035

Cited by 50 publications

(11 citation statements)

References 15 publications

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“…120 150 0-5mM the fluorescence intensity decreases more markedly with increasing pyruvate concentrations. No condensation occurs in the presence of 0-1 Mphosphate buffer, pH8 3 (Spikner & Towne, 1962). Effect of time of incubation, pH and concentration of enzyme on the assay of D-amino acid-oxida8e activity.…”

Section: Resultsmentioning

confidence: 99%

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Fluorimetric determination of d-amino acid oxidase

Verity¹,

Gallagher²,

Brown³

1967

Biochemical Journal

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1. A sensitive fluorimetric procedure for the assay of d-amino acid oxidase has been developed. 2. The method depends on the formation of a fluorescent derivative, 2-hydroxy-3-methylquinoxaline, on condensation of pyruvate with o-phenylenediamine in acid medium. 3. 2-Hydroxy-3-methylquinoxaline fluoresces strongly in 50% (v/v) sulphuric acid after excitation at 375mmu. A single emission peak is observed at 480mmu. 4. Formation of the quinoxaline is dependent on time, temperature, acidity and relative concentration of reactants. 5. A particulate preparation from mouse kidney required FAD for optimum activity at pH8.5; K(m) was 3.8x10(-3)m; K(FAD) was 1.4x10(-7)m and the reaction was strongly inhibited by p-chloromercuribenzoate and phenylmercuric acetate. 6. Subcellular fractionation on a sucrose density gradient confirmed that the d-amino acid oxidase was localized on small granules.

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Section: Resultsmentioning

confidence: 99%

“…The method reported below fulfils these criteria and depends on the formation of fluorescent quinoxalines on condensation of a-oxo acids with o-phenylenediamine in acid solution. Such a derivative of pyruvic acid, 2-hydroxy-3-methylquinoxaline, is highly fluorescent and stable in concentrated acid (Spikner & Towne, 1962).…”

mentioning

confidence: 99%

Fluorimetric determination of d-amino acid oxidase

Verity¹,

Gallagher²,

Brown³

1967

Biochemical Journal

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“…Fluorogenic reagents may be used for detection of α-keto acids in very dilute solutions because of their quantitative conversion into the corresponding substituted quinoxalines (Spiker and Towne, 1962;Hara et al, 1985a). Because sialic acids are α-keto acids, Hara et al (1985b) tried several of these reagents and found that 1,2-diamino-4,5-dimethoxybenzene reacts with Neu5Ac and Neu5Gc under conditions different than those for other α-keto acids and forms fluorescent compounds.…”

Section: Background Informationmentioning

confidence: 99%

Analysis of Monosaccharides

Manzi

Powell

Varki

1995

CP Molecular Biology

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This unit presents methods for assaying sialic acids, reducing sugars, and hexosamines. The BCA assay detects free reducing terminii in sugars released from glycoconjugates by appropriate treatments. Assays employing Ehrlich reagent (DMAB) detect hexosamines and N-acetylhexosamines, including a method for hydrolyzing the glycosidic linkages of the hexosamines and a method for re-N-acetylation. The TBA and DMB assays can be used to quantitate and fractionate free forms of many types of sialic acids. Techniques for liberating the sialic acids from the parent glycoconjugates are also provided.

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“…The advantages of the OPD derivatization include distinct labeling selectivity, high trapping efficiency, and low analytical artifacts caused under mild conditions. 1,7,20,21 By doing so, a well-established LC−MS/MS multimethod 6 has been reported to extend the in vivo spectrum of 17 target α-DCs. Nevertheless, as a growing number of α-DCs (Table 1) are disclosed from the reaction cascades, this picture is becoming more complex and thus there is a clear requirement for wide-scope analysis of more α-DC species in a single sample.…”

Section: ■ Introductionmentioning

confidence: 99%

Isotope-Coding Derivatization for Quantitative Profiling of Reactive α-Dicarbonyl Species in Processed Botanicals by Liquid Chromatography–Tandem Mass Spectrometry

Wang

Liu

Guo

et al. 2021

J. Agric. Food Chem.

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α-Dicarbonyls (α-DCs) are key reactive Maillard intermediates with structural diversity and are widely found in foods and in vivo, but little is known regarding the complete molecular profiles of these potentially harmful electrophiles. Herein, we reported a novel isotope-coding derivatization (ICD) strategy for the broad-spectrum, quantitative profiling of (non)target α-DC species in natural foodstuffs. It utilized differential isotope labeling (DIOL) with a reagent pair o-phenylenediamine (OPD)/OPD-d 4 (deuterated) to form stable quinoxalines for class-specific fragmentation-dependent acquisition using liquid chromatography–hybrid quadrupole linear ion trap mass spectrometry (LC-QqLIT). A combination of facile one-pot quantitative labeling and convenient cleanup protocol afforded satisfactory sensitivity, linearity, accuracy (81–116%), and process recovery (86–109% with RSDs < 10%) by matrix-matched ICD-internal standard calibration, without significant matrix interference (−9 to 5%), isotopic effect (<0.5%), and cocktail effect. A more generic DIOL-based LC-QqLIT algorithm integrated double precursor ion and neutral loss scan to trigger enhanced product ions with the unique isobaric doublet tags (4 Da shift), enabling simultaneous screening and relative quantitation of nontarget α-DC analogues in a single analysis. This study has widened the vision on complex α-DC profiles in traditional botanicals, which revealed a wide occurrence of α-DCs in such processed sugar-rich products, yet their abundance varied greatly among different samples.

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Fluorometric Microdetermination of Alpha-Keto Acids.

Cited by 50 publications

References 15 publications

Fluorimetric determination of d-amino acid oxidase

Fluorimetric determination of d-amino acid oxidase

Analysis of Monosaccharides

Isotope-Coding Derivatization for Quantitative Profiling of Reactive α-Dicarbonyl Species in Processed Botanicals by Liquid Chromatography–Tandem Mass Spectrometry

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