Abstract
Chromenone-derived natural products include chromones (flavone, isoflavone) and coumarins. Chromenone compounds not only exhibit impressive biological activities, but also are an important resource of experimentally used fluorophores, such as, 7-amino-4-methylcoumarin (AMC). Various chromenone compounds have reported to have weak fluorescence, and this has the potential to interfere with the measurements during AMC fluorogenic assays and result in non-robust assay readouts. Several flavones and isoflavones were found as SIRT1 activators, while fluorogenic sirtuin assays utilized AMC labelled peptides as the substrates. In this study we investigated whether the fluorescent properties of chromenone-derived natural products interrupt the measurement of SIRT1/2 modulated activities. We found that the reported SIRT1 activators: flavones were detected with the SIRT1 activation activity, but isoflavones were not detected with SIRT1 activation activity, and instead that they were found to be fluorogenic compounds. Another chromenone compound, osthole, exhibited a moderate SIRT2 inhibitory activity with an IC50 of 10 μM. In conclusion, the fluorescent properties of these chromenone compounds do affect the measurement of the sirtuin activities of both inhibitors and activators. However, if the possible fluorescence properties are mitigated in the assay readout, these fluorogenic assays enable the screening of activity modulators.
Highlights
Flavonoids are a large family of natural products, which are mainly found and extracted from plants [1,2,3]
Chromenone derived compounds are a class of privileged scaffolds within medicinal chemistry, with many chromenone compounds exhibiting impressive biological activities
Several chromenone compounds have been well recognized as commercial fluorophores, such as AMC and AMHC
Summary
Flavonoids are a large family of natural products, which are mainly found and extracted from plants [1,2,3]. Once the peptide was deacetylated by the sirtuin enzyme, the peptide was digested by trypsin to release the fluorescent tag and quantified. The sirtuin enzyme, the peptide was digested by trypsin to release the fluorescent tag and quantified In this assay, 7-amino-4-methylcoumarin (AMC) was used as the fluorophore. SIRT1 activators were initially screened from flavonoid-derived sirtuin inhibitors (Figure 3) [12,29,30,31,32,33,34,35,36,37,38]. The deacetylated peptide is digested by trypsin to release the free AMC, which has turn-on fluorescence. The measured fluorescence intensity can be the usedfree as aAMC, readout for the deacetylated peptide is digested by trypsin to release which has activity turn-on of fluorescence.
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