Tuning a 96-Well Microtiter Plate Fluorescence-Based Assay to Identify AGE Inhibitors in Crude Plant Extracts
AbstractAdvanced glycation end-products (AGEs) are involved in the pathogenesis of numerous diseases. Among them, cellular accumulation of AGEs contributes to vascular complications in diabetes. Besides using drugs to lower blood sugar, a balanced diet and the intake of herbal products potentially limiting AGE formation could be considered beneficial for patients’ health. The current paper presents a simple and cheap high-throughput screening (HTS) assay based on AGE fluorescence and suitable for plant extract screening. We have already implemented an HTS assay based on vesperlysines-like fluorescing AGEs quickly (24 h) formed from BSA and ribose under physiological conditions. However, interference was noted when fluorescent compounds and/or complex mixtures were tested. To overcome these problems and apply this HTS assay to plant extracts, we developed a technique for systematic quantification of both vesperlysines (λexc 370 nm; λem 440 nm) and pentosidine-like (λexc 335 nm; λem 385 nm) AGEs. In a batch of medicinal and food plant extracts, hits were selected as soon as fluorescence decreased under a fixed threshold for at least one wavelength. Hits revealed during this study appeared to contain well-known and powerful anti-AGE substances, thus demonstrating the suitability of this assay for screening crude extracts (0.1 mg/mL). Finally, quercetin was found to be a more powerful reference compound than aminoguanidine in such assay.
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Séro, L.; Sanguinet, L.; Blanchard, P.; Dang, B.T.; Morel, S.; Richomme, P.; Séraphin, D.; Derbré, S. Tuning a 96-Well Microtiter Plate Fluorescence-Based Assay to Identify AGE Inhibitors in Crude Plant Extracts. Molecules 2013, 18, 14320-14339.
Séro L, Sanguinet L, Blanchard P, Dang BT, Morel S, Richomme P, Séraphin D, Derbré S. Tuning a 96-Well Microtiter Plate Fluorescence-Based Assay to Identify AGE Inhibitors in Crude Plant Extracts. Molecules. 2013; 18(11):14320-14339.Chicago/Turabian Style
Séro, Luc; Sanguinet, Lionel; Blanchard, Patricia; Dang, Bach T.; Morel, Sylvie; Richomme, Pascal; Séraphin, Denis; Derbré, Séverine. 2013. "Tuning a 96-Well Microtiter Plate Fluorescence-Based Assay to Identify AGE Inhibitors in Crude Plant Extracts." Molecules 18, no. 11: 14320-14339.