Boron Schiff bases derived from α-amino acids as nucleoli/cytoplasm cell-staining fluorescent probes in vitro

Abstract

The size, shape, and number of nucleoli in a cell's nucleus might help to distinguish a malignant from a benign tumor. Cellular biology and histopathology often require better visualization to understand nucleoli-related processes, thus organelle-specific fluorescent markers are needed. Here, we report the design, synthesis, and fully chemo-photophysical characterization of fluorescent boron Schiff bases (BOSCHIBAs), derived from α-amino acids (i.e., phenylalanine, tyrosine and tryptophan), with nucleoli- and cytoplasm-specific staining in cells. It is the first time that Boron Schiff bases derived from α-amino acids act as notorious dual (nucleoli and cytoplasm) cell-staining fluorescent probes. The boron derivatives not only showed good photostability and acceptable quantum yields (∼5%) in solution, but also exhibited low cytotoxicity (>90% cell viability at 0.1 and 1 μg mL^-1), which make them good candidates to be used in medical diagnosis.

Scheme 1. Fluorescent dyes for in vitro nucleoli staining.

Scheme 2. Mono and binuclear Boron Schiff bases.

Scheme 3. Synthesis of Schiff bases by ultrasound Lig1–3 and BOSCHIBAs 1a–4a.

Fig. 1. Crystal structure of BOSCHIBA 3a. Anisotropic displacement parameters are depicted at the 30% probability level. Hydrogen atoms were omitted for clarity. Distances: B(1)–N(1) 1.631, B(1)–O(1) 1.495, B(1)–C(17) 1.614, B(1)–C(23) 1.618, C(11)–N(1) 1.299 Å. Bond angles: C(11)–B(1)–N(1) 118.64, O(1)–B(1)–N(1) 104.22, O(1)–B(1)–C(17) 110.89, N(1)–B(1)–C(23) 105.98, C(2)–C(11)–N(1) 122.85, C(11)–N(1)–C(12) 119.38°.

Fig. 2. Intermolecular hydrogen interactions in BOSCHIBA 3a.

Fig. 3. Boron out of the plane in half chair conformation.

Fig. 4. Absorption spectra of BOSCHIBAS 1a–4a (short dash dot) and (inset) amino acid Schiff base Lig1–3 (solid lines) in methanol.

Fig. 5. Emission spectra of BOSCHIBAs 1a–4a and (inset) α-amino acid Schiff bases Lig1–3 in MeOH.

Fig. 6. Cytotoxicity effect of BOSCHIBAs 1a–3a. B16F10 melanoma cells were treated with 10 mg mL⁻¹ (soft blue bars), 5 mg mL⁻¹ (purple bars), 2.5 mg mL⁻¹ (green bars), 1 mg mL⁻¹ (red bars) or 0.1 mg mL⁻¹ (sharp blue bars) for 24 hours.

Fig. 7. Confocal fluorescence images of B16F10 melanoma cells stained with BOSCHIBAs 1a–3a (10 μg mL⁻¹). (A–C) Untreated cells; (D–F) compound 1a; (G–I) compound 2a; (J–L) compound 3a (scale bar shown represents 20 mm).

Fig. 8. Confocal fluorescence images of B16F10 melanoma cells stained with 10 μg mL⁻¹ of BOSCHIBAs 1a (A), 2a, (B), and 3a (C).

Fig. 9. Photostability of 1a–4a (1 mg/50 mL) in methanol at the absorption maximum wavelength with the irradiation time.