Figures & data
Figure 1. Synthesis of poly-hydroxy functionalized pyrimidine-fused heterocyclics (PHPFHs) using the reaction of d-(+)-glucose, barbituric acid and amines. The detailed description of the synthetic procedure is seen in the Materials and Methods (General procedure for the synthesis of compounds C1–C5 subsection). As shown in this figure, further substitutions (C1–C5) were also made at the position 10 (R-group) of the general structure of pyrimidine-fused heterocyclic (PFH) with a poly-hydroxy carbon chain to obtain either the aliphatic or aromatic derivatives.
![Figure 1. Synthesis of poly-hydroxy functionalized pyrimidine-fused heterocyclics (PHPFHs) using the reaction of d-(+)-glucose, barbituric acid and amines. The detailed description of the synthetic procedure is seen in the Materials and Methods (General procedure for the synthesis of compounds C1–C5 subsection). As shown in this figure, further substitutions (C1–C5) were also made at the position 10 (R-group) of the general structure of pyrimidine-fused heterocyclic (PFH) with a poly-hydroxy carbon chain to obtain either the aliphatic or aromatic derivatives.](/cms/asset/3557ecfd-fc73-4de6-ae4b-2f6d9bc14cd0/ienz_a_727812_f0001_b.gif)
Table 1. The IC50, Ki and inhibition mode of the synthetic compounds.
Figure 2. The Lineweaver–Burk plots derived from the inhibition of yeast and mouse α-glucosidases. (A) The yeast α-glucosidase (α-Gls) activity was measured as a function para-nitrophenyl-α-d-glucopyranoside (pNPG) concentration (0.1–2 mM) in the absence and presence of C3 compound (0–20 µM). The experiments performed in 100 mM phosphate buffer pH 7.0, at 25°C for 10 min. The different symbols represent the absence (diamonds) and presence of 5 µM (squares), 10 µM (triangles), and 20 µM (circles) of C3 inhibitor in the reaction mixtures. (B) The mouse α-Gls activity was measured as a function pNPG concentration (0.6–3 mM) in the absence and presence of C3 compound (0–200 µM). The experiments performed in 10 mM phosphate buffer pH 7.0, at 37°C for 30 min. The different symbols represent the absence (diamonds) and presence of 50 µM (squares), 100 µM (triangles), and 200 µM (circles) of C3 inhibitor in the reaction mixtures.
![Figure 2. The Lineweaver–Burk plots derived from the inhibition of yeast and mouse α-glucosidases. (A) The yeast α-glucosidase (α-Gls) activity was measured as a function para-nitrophenyl-α-d-glucopyranoside (pNPG) concentration (0.1–2 mM) in the absence and presence of C3 compound (0–20 µM). The experiments performed in 100 mM phosphate buffer pH 7.0, at 25°C for 10 min. The different symbols represent the absence (diamonds) and presence of 5 µM (squares), 10 µM (triangles), and 20 µM (circles) of C3 inhibitor in the reaction mixtures. (B) The mouse α-Gls activity was measured as a function pNPG concentration (0.6–3 mM) in the absence and presence of C3 compound (0–200 µM). The experiments performed in 10 mM phosphate buffer pH 7.0, at 37°C for 30 min. The different symbols represent the absence (diamonds) and presence of 50 µM (squares), 100 µM (triangles), and 200 µM (circles) of C3 inhibitor in the reaction mixtures.](/cms/asset/6d9aac2c-9b66-4ca3-acbd-89924ea4e9e6/ienz_a_727812_f0002_b.gif)
Table 2. The binding parameters of interaction between C3 inhibitor and α-Gls.
Figure 3. Fluorescence studies of the interaction between C3 and yeast α-glucosidase (α-Gls). (A) The intrinsic fluorescence quenching of α-Gls induced by C3 inhibitor. The enzyme was incubated with increasing concentration of C3 (0–50 µM) for 15 min at 37°C and 42°C. As the experiments performed in 100 mM phosphate buffer pH 7.0, the excitation wavelength was 295 nm and emission spectra were acquired by scanning from 300 to 400 nm. Only the emission spectra obtained at 37°C are shown in this figure. Also the Stern–Volmer plots based on the fluorescence quenching at 37 and 42°C are shown as the inset figures. (B) The 8-anilinonaphthalene 1-sulphonate (ANS)-fluorescence of α-Gls as function of concentration of C3 inhibitor. The ANS fluorescence measurements were performed in 100 mM phosphate buffer pH 7.0 while enzyme and ANS concentration were 2 and 30 µM, respectively. The enzyme was incubated with certain concentrations of C3 inhibitor (0–50 µM) for 15 min, at 37°C in the dark, and fluorescence of protein-bound dye measured by excitation at 365 nm and measuring the emission between 400 and 600 nm.
![Figure 3. Fluorescence studies of the interaction between C3 and yeast α-glucosidase (α-Gls). (A) The intrinsic fluorescence quenching of α-Gls induced by C3 inhibitor. The enzyme was incubated with increasing concentration of C3 (0–50 µM) for 15 min at 37°C and 42°C. As the experiments performed in 100 mM phosphate buffer pH 7.0, the excitation wavelength was 295 nm and emission spectra were acquired by scanning from 300 to 400 nm. Only the emission spectra obtained at 37°C are shown in this figure. Also the Stern–Volmer plots based on the fluorescence quenching at 37 and 42°C are shown as the inset figures. (B) The 8-anilinonaphthalene 1-sulphonate (ANS)-fluorescence of α-Gls as function of concentration of C3 inhibitor. The ANS fluorescence measurements were performed in 100 mM phosphate buffer pH 7.0 while enzyme and ANS concentration were 2 and 30 µM, respectively. The enzyme was incubated with certain concentrations of C3 inhibitor (0–50 µM) for 15 min, at 37°C in the dark, and fluorescence of protein-bound dye measured by excitation at 365 nm and measuring the emission between 400 and 600 nm.](/cms/asset/0e11d583-1b84-4705-b572-c18f8e2200e0/ienz_a_727812_f0003_b.gif)