Analysis of microperoxidases using liquid chromatography, post-column substrate conversion and fluorescence detection

Haselberg, Rob; Hempen, Christel; Leeuwen, S.M. van; Vogel, Martin; Kärst, Uwe

doi:10.1016/j.jchromb.2005.10.013

Cited by 4 publications

(1 citation statement)

References 26 publications

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“…Microperoxidases are enzymes used in analytical chemistry to catalyze the oxidation of organic substrates by hydrogen peroxide [ 70 ]. Haselberg et al [ 71 ] developed a dual substrate system for the detection of microperoxidases in proteolytic digests of cytochrome c. 4-( N -methylhydrazino)-7-nitro- 2,1,3-benzooxadiazole is converted by microperoxidases to 4-( N -methylamino)-7-nitro-2,1,3-benzooxadiazole, which fluoresces in the presence of hydrogen peroxide. Coupling this BCD system to reversed phase HPLC enabled identification of fractions with microperoxidase activity.…”

Section: Hplc Coupled On-line To Ead and Rad Assays And Their Applmentioning

confidence: 99%

Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

Malherbe

Beer

Joubert

2012

IJMS

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Biochemical detection (BCD) methods are commonly used to screen plant extracts for specific biological activities in batch assays. Traditionally, bioactives in the most active extracts were identified through time-consuming bio-assay guided fractionation until single active compounds could be isolated. Not only are isolation procedures often tedious, but they could also lead to artifact formation. On-line coupling of BCD assays to high performance liquid chromatography (HPLC) is gaining ground as a high resolution screening technique to overcome problems associated with pre-isolation by measuring the effects of compounds post-column directly after separation. To date, several on-line HPLC-BCD assays, applied to whole plant extracts and mixtures, have been published. In this review the focus will fall on enzyme-based, receptor-based and antioxidant assays.

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Section: Hplc Coupled On-line To Ead and Rad Assays And Their Applmentioning

confidence: 99%

Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

Malherbe

Beer

Joubert

2012

IJMS

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NBD Methylhydrazine (Mnbdh)

Sabnis¹

2015

Handbook of Fluorescent Dyes and Probes

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Enzymatic Assays Coupled with Mass Spectrometry with or without Embedded Liquid Chromatography

et al. 2015

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This article reviews monitoring strategies for enzymatic assays coupled with mass spectrometric detection. This coupling has already been shown to be helpful in providing versatile and detailed knowledge about enzyme kinetics. Various available publications address two general approaches. 1) The continuous-flow setup allows real-time determination of substrate degradation. Simultaneously, resulting product or potential intermediates can be detected. 2) The online coupled continuous-flow mixing assay allows the direct coupling of an enzymatic assay to chromatographic separation of complex mixtures. The latest efforts in improving the methodology have been made with regard to miniaturization. This is especially advantageous with regard to reducing costly consumption of chemicals. Finally, these developments are applicable for diverse bioanalytical purposes in the realms of pharmaceutical, biotechnological, food, and environmental research.

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Analysis of microperoxidases using liquid chromatography, post-column substrate conversion and fluorescence detection

Cited by 4 publications

References 26 publications

Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

NBD Methylhydrazine (Mnbdh)

Enzymatic Assays Coupled with Mass Spectrometry with or without Embedded Liquid Chromatography

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