Table 1.
Yield, chemical compositions and contents of carbohydrate, protein, MW and sulfonic acid in the C. fluminea protein-bound polysaccharide CFPS-1.
Table 2.
The amino acid composition of the protein-bound polysaccharide CFPS-1 from C. fluminea.
Fig 1.
Molecular and morphological characterization of C. fluminea protein-bound polysaccharide CFPS-1.
(A and B) The atomic force microscopy (AFM) images of CFPS-1. The CFPS-1 concentration was 1 μg/mL (A; B1), 10 μg/mL (B2) and 100 μg/mL (B3), respectively. Arrow: (a) single linear chain; (b) branched chain; (C) FTIR spectra of CFPS-1 (4000 cm−1 to 400 cm−1) were obtained from solid samples by KBr disc method using a Nicolet 5700 FT-IR spectrophotometer. FTIR spectra showed that CFPS-1 mainly contained three types of groups, namely hydroxyl group, amino group and sulfate group.
Table 3.
GC-MS of alditol acetate derivatives from the methylated product of the C. fluminea protein-bound polysaccharide CFPS-1.
Table 4.
1H and 13C NMR chemical shifts for the protein-bound polysaccharide CFPS-1 isolated from C. fluminea in D2O a.
Fig 2.
Structural elucidation of the C. fluminea protein-bound polysaccharide CFPS-1.
(A) 1H NMR and (B) 13C NMR spectra of CFPS-1 in D2O. Chemical shifts were reported relative to internal acetone at δH 2.23 ppm for 1H spectrum and δC 32.45 ppm for 13C spectrum; (C) A diagram showing the partial structures of polysaccharide portions of the protein-bound polysaccharide CFPS-1 from C. fluminea. (a): 1,6-linked -α-D-Glc; (b): 1,4,6-linked-α-D-Glc; (c): 1-linked-α-D-Man.
Fig 3.
Effect of the C. fluminea protein-bound polysaccharide CFPS-1 on growth of human breast cancer MCF-7 and MDA-MB-231 cells.
The inhibitory effect of CFPS-1 on the cell growth of MCF-7 (A) and MDA-MB-231(C) cells at 24, 48, 72 and 96 h. Proliferation and cytotoxicity against MCF-7 (B) and MDA-MB-231 (D) cells by the CFPS-1 at 72 h. CK, Control of proliferation or cytotoxicity. All data were expressed as mean ± SD of three experiments and each experiment included triplicate repeats. Values marked with * are significantly different from the control (p < 0.05).
Fig 4.
Effect of the C. fluminea protein-bound polysaccharide CFPS-1 on MCF-7 and MDA-MB-231 cell cycle progression.
Cells were cultured for 24 h with CFPS-1 (0, 50, 150 and 250 μg/mL, respectively). Untreated cells (0 μg/mL) were used as control. Cell cycle distribution (%) of MCF-7 (A) and MDA-MB-231 (C) cells were determined by flow cytometry after CFPS-1 treatment; Apoptotic rate (%) of MCF-7 (B) and MDA-MB-231 (D) cells treated with CFPS-1. Apoptotic cells were identified with a TUNEL technique and counted with a light microscope (magnification, ×40). All data were expressed as mean ± SD of three experiments and each experiment included triplicate repeats. Values marked with * are significantly different from the control (p < 0.05).
Fig 5.
Effects of the C. fluminea protein-bound polysaccharide CFPS-1 on the expression of p21, Cyclin D1, Cdk4, p53, Bcl-2, Bax and caspase-7 proteins associated with cell cycle and apoptosis in MCF-7(A, B, C and D) and MDA-MB-231 (E, F, G and H) cells.
The relative expression of protein was quantified densitometrically (%). Western blot analysis was performed in triplicate per experimental point; β-actin was used as reference control. Values marked with * are significantly different from the control (p < 0.05).