Characterization of Sulfate Groups and Assessment of Anti-Coagulant Activity of Glucomannan Sulfate Prepared from Konjac Glucomannan

Purpose: To determine the structure of Konjac glucomannan sulfate (KOGMS) including homogeneity, sulfate group composition as well as its anti-coagulant activity in vitro.
Methods: KOGMS was prepared using chlorosulfonic acid and N, N-dicyclohexyl carbodiimide (DCC) from konjac oligo-glucomannan (KOGM). Homogeneity analysis was confirmed by cellulose acetate membrane electrophoresis. Fourier transform infrared FT-IR spectra, Laser Raman spectra and 13C Nuclear Magnetic Resonance NMR spectra were obtained and used to analyze the sulfate groups. The anti-coagulant activity of KOGM and KOGMS was evaluated in vitro using activated partial thromboplastin time (APTT), prothrombin time (PT) and thrombin time (TT) as indicators.
Results: KOGMS was shown to be a homogenous carbohydrate with a relative molecular mass ranging from 5.8 × 10³ to 6.2 × 10³ Da and various degrees of substitution (DS) ranging from 1.28 to 1.81. The sulfate groups were inserted at C-2, C-6 and C-3 positions of KOGM. The APTT of human platelet-poor plasma containing KOGMS was 47.2 s which was close to that of heparin at a concentration of 0.5 mg/mL. Furthermore, the APTT of human platelet-poor plasma containing KOGM was only 15.6 s which is close to that of the negative control group. For KOGMS, the values obtained in all three tests were significantly higher than those of KOGM and the negative control group (p < 0.05).
Conclusion: Hydroxyl groups can be substituted efficiently by sulfated groups at C-2, C-6 and C-3 positions of KOGM with little degree of degradation. In vitro anti-coagulant activity data indicates that KOGMS has a significantly stronger anti-coagulant activity than KOGM due probably to the sulfated groups in the main chain of its molecule.

Keywords
Konjac; Oligo-glucomannan sulfate; Laser Raman spectra; Anticoagulant activity