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

Abstract

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.

Reactive Secondary Sequence Oxidative Pathology Polymer Model and Antioxidant Tests

“…To better understand discrepancies between measured vitamin antioxidant potentials and clinical failure perhaps the antioxidant tests should be considered. Current antioxidant tests that created the indices comparing different vitamins with activities of other antioxidants are primarily based on changes between the oxidation of a sample and free-radicals from a stable source, metal ions through a Fenton reaction or a physiologic-relevant reactive species such as the hydroxyl radical [74][75][76][77]. The original antioxidant testing by Burton and Ingold in 1981 used thermal changes as an indication for free-radical complexing with a sample [15].…”

“…The original antioxidant testing by Burton and Ingold in 1981 used thermal changes as an indication for free-radical complexing with a sample [15]. The most current common antioxidant tests use colorimetric assays with uv-vis spectroscopy that examines optical adsorption at a characteristic maximum peak in the visible region of the electromagnetic spectrum between about 400-700 nm [74][75][76][77]. In the assay sample-freeradical complex, adsorption occurs at a maximum peak that can be measured from a control standard.…”

Quality-In(Process)Line (QuIProLi) process intensification for a micro-flow UV-photo synthesis enabled by online UHPLC analysis

“…37 Therefore, process miniaturization requires an orders of magnitude smaller volume sampling, which in micro process engineering would mean easily most of the volume of the whole reactor. Some analytical techniques, which include sampling, fulfil that criterion such as Raman, 38 liquid chromatography, 39 or capillary electrophoresis. 40,41 However, for small molecule analysis, high-performance liquid chromatography (HPLC) is commonly preferred and is quasi the unique method used for process monitoring applications in micro-channels offline.…”

Multiple on‐line HPLC coupled with biochemical detection methods to evaluate bioactive compounds in Danshen injection

“…Acetylcholinesterase (AChE) is an important enzyme owing to its relation with AD, so the screening of AChE inhibitors from complex extracts has attracted much attention owing to most AD drugs inhibiting AChE. At present, the on‐line analysis methods for rapid identification of active compounds in complex extracts have overcome to some extent the problems in the traditional separation and screening processes of active compounds of complex extracts, such as time, labor and money consumption for useless compounds (Peng, Tan, Huang, & Ding, ; Niederländer, van Beek, Bartasiute, & Koleva, ; Shi et al, ; Li, Qian, & Li, ; Malherbe, de Beer, & Joubert, ). Some novel analysis techniques, on‐line HPLC coupled with different detectors such as UV detection nearby 400 nm (Fabel, Niessner, & Weller, ), fluorescence (FL) detection based on fluorogenic substrate 7‐acetoxy‐1‐methyl quinolinium iodide ( λ ex 406 nm; λ em 505 nm) (Rhee, Appels, Luijendijk, Irth, & Verpoorte, ; Marques et al, ; Heus et al, ) and MS detection (Ingkaninan et al, ; de Jong, Derks, Bruyneel, Niessen, & Irth, ), have been developed to quickly screen AChE inhibitors in complex extracts.…”