Fluorimetric Properties of 3-Aminoflavone Biomolecule (3-AF). X-ray Crystal Structure of New Polymorph of 3-AF

Abstract

The crystal structure of the new polymorphic form of 3-aminoflavone (3-AF) has been determined by single crystal X-ray diffraction. This report presents results of fluorimetric studies on 3-AF in methanol and aquatic solvents. Based on 3D fluorescence emission spectra, optimal values for excitation (λ_ex) and emission/analytical (λ_em) wavelength, the analytical concentration range as well as the range of concentration quenching for the studied compound were established. Moreover, the limit of detection (LOD) and the limit of quantification (LOQ) were determined. The results were compared with those obtained using the standard UV-Vis absorption spectrophotometric method. The effect of acidity (pH) and the concentration of halide anions (chlorides, bromides, iodides and fluorides) on fluorescence quenching were analysed.

Keywords: 3-aminoflavone; LOD (limit of detection); LOQ (limit of quantification); X-ray crystallography; concentration quenching; fluorescence.

Figure 1

Structural formula of 3-aminoflavone.

Figure 2

A view of 1o with atom numbering scheme. Displacement ellipsoids for non-hydrogen atoms were draw with 50% probability level—(a); superposition of 3-AF molecules from crystal 1o (red) and centrosymmetric 1m (dashed green)—(b).

Figure 3

Hirshfeld surfaces mapped with electrostatic potential calculated with B3LYP/6-; 311++G(d,p) and the fingerprint plots for 1o—(a) and 1m—(b).

Figure 4

UV-Vis absorption spectrum of 3-aminoflavone in methanol at a concentration of 100 μg/mL, in quartz cuvette with l = 0.2 cm, at temperature t = 20 °C, with methanol as the reference.

Figure 5

Three-dimensional spectrum of 3-aminoflavone in methanol. Intensity of fluorescence emission I_f in RFU (relative fluorescence unit). Instrument parameters. Measurement type: 3D scan; data mode: Fluorescence; EX (Excitation) sampling interval: 3.0 nm; EM (Emission) sampling interval: 3.0 nm; scan speed: 1200 nm/min; EX slit: 5.0 nm; EM slit: 5.0 nm; PMT (Photomultiplier Tube) voltage: 700 V; sensitivity: 1; threshold: 1.0.

Figure 6

Emission spectrum of 3-aminoflavone in methanol at a concentration c = 0.1 μg/mL in quartz cuvette with a thickness l = 1 cm. Fluorescence excitation λ_ex = 365 nm; intensity of fluorescence emission I_f is presented in RFU (relative fluorescence unit).

Figure 7

Intensity of fluorescence emission I_f as a function of 3-aminoflavone concentration in methanol. The fluorescence was measured in cuvette with a path length l of 1 cm. For excitation and emission, filters with wavelengths of 350 nm and 470 nm were used, respectively.

Figure 8

Fluorescence emission intensity dependence as a function of 3-aminoflavone concentration in methanol fitted with a linear function. The cuvette thickness l was 1 cm. For excitation and emission, filters with wavelengths of 350 nm and 470 nm were used, respectively.

Figure 9

Dependence of absorbance A at λ = 364 nm as the function of 3-aminoflavone concentration in methanol. The quartz cuvette thickness l was 0.2 cm.

Figure 10

Relationship between fluorescence intensity I_f of 3-aminoflavone and pH. For excitation and emission, filters with wavelengths of 350 nm and 470 nm were used, respectively.

Figure 11

UV-Vis spectra of 3-aminoflavone (c = 5 μg/mL) in HCl solutions at pH = 1, 2, 3, 4, 5. l = 1 cm.

Figure 12

The relationship between fluorescence intensity I_f of 3-aminoflavone and the concentration of chloride, bromide, iodide and fluoride ions. The concentration of 3-aminoflavone was 5 µg/mL in cuvette with a path length of 1 cm. For excitation and emission, filters with wavelengths of 350 nm and 470 nm were used, respectively.