The Optimisation of Analytical Parameters for Routine Profiling of Antioxidants in Complex Mixtures by HPLC Coupled Post‐column Derivatisation

Abstract

The study confirms that the commercial HPLC-coupled post-column derivatisation instruments are suitable for routine on-line detection of antioxidants in complex mixtures and the determination of their TE values. The analytical parameters optimised here appear to represent a ready-to-use toolbox for the food and pharmaceutical industries, enabling the monitoring of bioactive substances along the production line and during storage, and the characterisation of plant material by creating chromatographic profiles supplemented with antioxidant fingerprints.

The influence of roasting and additional processing on the content of bioactive components in special purpose coffees

“…The conditions of chromatographic separation of bioactive compounds present in coffee extracts were identical with those described above. The post-column derivatisation procedure was done according to Kusznierewicz et al, (2011).…”

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

“…Recently, an on-line radical scavenging assay, employing galvinoxyl radicals, has also been described [79]. Additionally, assays based on the redox reaction of antioxidants with a phosphomolybdate reagent [80], Folin-Ciocalteau reagent [81,82] and Cu(II)-neocuproine reagent [83], used in the cupric reducing antioxidant capacity (CUPRAC) batch assay, have been reported. An HPLC-chemiluminescence (CL) assay based on CL of acidic permanganate has also been reported [84–86], indicating reducing capacity of the antioxidants detected.…”

“…In both these cases, the reagent was prepared in hexane. Other novel applications include optimization of the assay for use with commercially available derivatization equipment [81,82] and use of the technique to obtain information on the antioxidant activity and the position of conjugation for hepatic phase II metabolites of aspalathin [105]. …”

“…Hyphenation of HPLC-ABTS with MS [110] and SPE-NMR [104,111] detection has also been reported. Novel methodology or modifications applied since 2008 include the following: NP-HPLC using ABTS •+ reagent prepared in phosphate buffered saline and diluted with methanol [88]; reaction temperature of 40 °C [108,112,113]; method validation [114]; assessment of the activity and position of conjugation for hepatic phase II metabolites of aspalathin [105]; and use of a commercially available derivatization equipment [81,82]. …”

“…No other reports mentioning this technique have been forthcoming until recent reports of an on-line antioxidant assay involving the related Folin-Ciocalteau reagent containing heteropolyphosphotungstates-molybdates [81,82]. Both these assays are believed to share a similar reaction mechanism involving electron-transfer from an antioxidant compound reducing Mo 6+ to Mo 5+ to form a blue-green complex detected at wavelengths higher than 580 nm [80,115].…”