Better than bench top. High speed antioxidant screening via the cupric reducing antioxidant capacity reagent and reaction flow chromatography

This study is based upon a recently established method for quantification of the antioxidant capacity of natural samples via a HPLC separation and a hyphenated selective detection (post-column derivatization with cupric reducing antioxidant capacity reagent) technique. This protocol demonstrated the main improvements to transform the quantitative protocol into a high-speed qualitative automated assay to screen samples for their potential total antioxidant capacity, typically performed via manual mixing of the sample and derivatisation and measured on a 96 well plate reader/bench top UV–Vis spectrophotometer. This approach with automated mixing is a more informative alternative for total antioxidant capacity as the antioxidant peaks are profiled for each sample within four minutes. This antioxidant profile may be used for routine analysis of raw materials and/or a guide for targeted approaches for structure elucidation for laboratories interested in early drug discovery, natural product research and the search of alternative antioxidant additives in consumer goods/therapeutics. This technique could also be used to monitor the stability, alteration or adulteration of manufactured goods containing antioxidants.Fil: Suktham, Thirada. University of Western Sydney; AustraliaFil: Jones, Andrew. University of Western Sydney; AustraliaFil: Acquaviva, Agustín. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Química. Grupo Cromatografía; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; ArgentinaFil: Dennis, Gary R.. University of Western Sydney; AustraliaFil: Shalliker, R. Andrew. University of Western Sydney; AustraliaFil: Soliven, Arianne. University of Western Sydney; Australia. Universidad de la República; Urugua

Total Antioxidant Capacity with Peak Specificity via Reaction Flow Chromatography and the Ferric Reducing Antioxidant Power Assay

An established ferric reducing antioxidant power (FRAP) assay was optimised by preparation of the derivatisation reagent in 300 mM formate instead of 300 mM acetate conditions, resulting in increased sensitivity signal to noise responses by up to five to ten times. The quantitative protocol for selective detection of antioxidants via a HPLC post column derivatisation (PCD) technique using the 300 mM formate FRAP reagent conditions was then transformed into a high-speed qualitative screening protocol by utilizing an emerging technology ‘reaction flow (RF) chromatography’. Reaction flow chromatography’s ability to screen for total antioxidant capacity with additional peak specificity/profile information of active peaks could be achieved in under 2 min.Laboratorio de Investigación y Desarrollo de Métodos Analíticos (LIDMA

Better than bench top : High speed antioxidant screening via the cupric reducing antioxidant capacity reagent and reaction flow chromatography

This study is based upon a recently established method for quantification of the antioxidant capacity of natural samples via a HPLC separation and a hyphenated selective detection (post-column derivatization with cupric reducing antioxidant capacity reagent) technique. This protocol demonstrated the main improvements to transform the quantitative protocol into a high-speed qualitative automated assay to screen samples for their potential total antioxidant capacity, typically performed via manual mixing of the sample and derivatisation and measured on a 96 well plate reader/bench top UV–Vis spectrophotometer. This approach with automated mixing is a more informative alternative for total antioxidant capacity as the antioxidant peaks are profiled for each sample within four minutes. This antioxidant profile may be used for routine analysis of raw materials and/or a guide for targeted approaches for structure elucidation for laboratories interested in early drug discovery, natural product research and the search of alternative antioxidant additives in consumer goods/therapeutics. This technique could also be used to monitor the stability, alteration or adulteration of manufactured goods containing antioxidants.Laboratorio de Investigación y Desarrollo de Métodos Analíticos (LIDMA

Antioxidant Profiling of Ginger via Reaction Flow Chromatography

Reaction flow (RF) chromatography is a powerful and efficient approach that utilizes conventional high-performance liquid chromatography (HPLC)–ultraviolet (UV)–visible detection. This technique exploits a novel column end-fitting and an extra HPLC pump that delivers a reagent specific for selective detection, in particular the antioxidant profiling of natural products. This study employed RF for the first time to identify antioxidants in a commercial ginger sample. This demonstrated the previously validated assay’s ease and power to extract information about the natural product’s antioxidant properties. Due to the simplicity involved with data analysis and peak matching process, the following information was revealed between the chemical and antioxidant profiles: three of the strongest antioxidant activity peaks in the ginger sample (593 nm) did not correlate with the three most abundant chemical profile peaks (UV absorbance at 254 and 280 nm); the ratio of seven antioxidant peaks may be potentially used for food authenticity purposes, and future research should target these peaks for the early discovery of novel antioxidants sourced in ginger. Utilization of this previously validated assay provided the resolution of numerous peaks in the ginger extract and information associated with their antioxidant attributes and chemical abundance. This approach is more informative than total antioxidant assays that lack compound specificity information. Furthermore, it is superior to mass spectrometric (MS) assays that cannot evaluate each compound’s antioxidant strength, and does not involve the expense involved in the acquisition and maintenance of the MS detection hardware, and does not require the high level of expertise needed to conduct the MS data analysis.Laboratorio de Investigación y Desarrollo de Métodos Analíticos (LIDMA

The future of liquid chromatographic separations should include post column derivatisations : a discussion view point based on the perspective for the analysis of vitamin D

The coupling of high performance liquid chromatography (HPLC) with post column derivatisation (HPLC-PCD) will increase the sensitivity of an instrument to the target molecule, improve specificity by selective derivatisation to remove potential interferences from the matrix, and be cost-effective; all these factors may well prove that PCD is an alternative to existing techniques for routine analysis. Calcidiol (25(OH)D) concentrations are used in the determination of vitamin D deficiency in humans, and there is growing interest and demand in its analysis with some controversy surrounding the sensitivity and specificity of its detection. 25(OH)D analysis is currently performed using either competitive binding assays or HPLC analysis, coupled with detection by a variety of means, such as, UV–Vis absorption, electrochemical or mass spectroscopy (MS) detection. HPLC coupled with MS (HPLC-MS) detection is the gold standard for 25(OH)D analysis due to its sensitivity and selectivity compared to all other techniques, however, its utility is limited by its high cost. This article reviews the current status of HPLC based techniques in the detection of 25(OH)D (and its metabolites), and further explores the use of HPLC-PCD