Studies On Chiral Stationary Phases Drived From N-cyclohexaneformylated, N-butyrylated And N-hexanoylated Chitosan Derivatives

The significant difference between a pair of enantiomers of chiral drugs in pharmacologic action and physiological activity makes enantioseparation be one of the research hotspots in the fields of separation and analysis. High-performance liquid chromatography(HPLC) based on chiral stationary phase(CSP) has become the best technique to separate and detect enantiomers. Therefore, it is very important to investigate the preparation and the structural dependence on the properties of novel CSPs. In order to broaden the application scope of CSPs, and to find CSPs with good chiral recognition ability and durability, in this work, a series of CSPs based on the chitosan with an ultra-high degree of deacetylation(D.D.) were designed and prepared. The enantioseparation capability and tolerability against organic solvents were evaluated. In addition, the structural dependence of chitosan derivatives on chiral recognition, enantioseparation and tolerability to organic solvent was disclosed by investigating the enantioseparation capability of the chitosan derivatives with various structures. The works are summarized as follows:(1) By changing the reaction time, two kinds of chitosan with different molecular weights were prepared in n-pentanol-NaOH reaction system. The viscosity-average molecular weights(M?) of the chitosan were 2.4×105 and 2.8×105 respectively. The chitosan with the M? of 1.4×105 was obtained by the degradation reaction of the chitosan with the M? of 2.4×105 in the presence of hydrogen peroxide. The chitosan whose M? were 1.4×105 and 2.4×105 were modified with an excess of cyclohexanecarboxylic anhydride yielding N-cyclohexaneformylated chitosan. The N-cyclohexaneformylated chitosan were characterized by 1H NMR and elemental analysis confirming the amino group on chitosan was selectively and almost completely acylated. Three polymers of chitosan bis(arylcarbamate)-(cyclohexanecarboxamide) were prepared with methyl-containing phenyl isocyanates and the N-cyclohexaneformylated chitosan that was derived from the chitosan with the M? of 1.4×105, and four polymers were prepared with chloro-containing phenyl isocyanates and the N-cyclohexaneformylated chitosan that was derived from the chitosan with the M? of 2.4×105. The chitosan derivatives were used as the chiral selectors and were coated on 3-aminopropyl silica gel to obtain seven new CSPs. The chiral selectors were characterized by FT-IR, 1H NMR and elemental analysis. The chiral recognition and enantioseparation capabilities of these CSPs were evaluated via HPLC. Moreover, the effect of the number and position of the substituent(s) located on phenyl group on chiral recognition ability of the CSPs was studied. The results showed that the CSPs had better enantioseparation capability when the modifying agents contained electron withdrawing group(s). The CSP prepared from chitosan bis(3,4-dichlorophenylcarbamate)-(cyclohexylcarboxamide) showed the strongest enantioseparation capability among this series of the CSPs derived from N-cyclohexaneformylated chitosan derivatives. The tolerability of the chitosan derivatives against organic solvents was observed. These chitosan derivatives were nearly insoluble in acetone, chloroform and ethyl acetate, etc., yet soluble in tetrahydrofuran(THF). Compared to the CSPs prepared by coating cellulose tris(3,5-dimethylphenylcarbamate)(CDMPC) and amylose tris(3,5-dimethylphenylcarbamate)(ADMPC) on silica gel, the above CSPs can work in the eluents of chloroform, acetone and ethyl acetate, suggesting a wider range of mobile phase suitable for these CSPs.(2) The amino groups on 2-position of the chitosans with the M?s of 2.4×105 and 2.8×105 were selectively and completely modified by hexanoic anhydride. The two yielded N-hexanoylated chitosans, respectively, reacted with four methyl-containing phenyl isocyanates and four chloro-containing phenyl isocyanates affording 8 chiral selectors of chitosan bis(arylcarbamate)-(hexanamide). These chiral selectors were coated on 3-aminopropyl silica gel to obtain 8 new CSPs. The chitosan derivatives were characterized by FT-IR, 1H NMR and elemental analysis. The chiral recognition and enantioseparation capability of the CSPs were investigated by HPLC. Chromatographic separation results exhibited most CSPs possessed a better enantioseparation capability. Under the same chromatographic conditions, the chiral recognition capability of these CSPs was generally equivalent to or even better than those of the CSPs of CDMPC and ADMPC. The tolerability of chitosan bis(3-methylphenylcarbamate)-(hexanamide) against organic solvents was studied. The CSP can work in the mobile phases of 100% acetone, 100% chloroform and 100% THF. Compared with N-cyclohexylformylated chitosan derivatives, N-hexanoylated chitosan derivatives had better tolerability towards organic solvents. These derivatives were not only insoluble in acetone and other organic solvents, but also insoluble in THF. Therefore, compared to the CSPs of N-cyclohexaneformylated chitosan derivatives, the applicable scope of the mobile phases for the CSPs of N-hexanoylated chitosan derivatives was further broadened.(3) Ten kinds of chitosan bis(arylcarbamate)-(butyramide) were characterized by FT-IR, 1H NMR and elemental analysis. These chitosan derivatives were coated on 3-aminopropyl silica gel to obtain new CSPs. The chiral recognition and enantioseparation capabilities of these CSPs were evaluated by HPLC. Most CSPs possessed a better enantioseparation capability. Comparing swelling and dissolution of N-hexanoylated and N-butyrylated chitosan derivatives in organic solvents, and the chiral recognition and enantioseparation capability of their corresponding CSPs, it was found that N-hexanoylated chitosan derivatives with a relatively lower molecular weight had lower swelling and solubility than the N-butyrylated chitosan derivatives. Therefore, the CPS of N-hexanoylated chitosan derivatives should have a relatively more stable separation capability. Under the same chromatographic conditions, the CSPs of N-butyrylated chitosan derivatives had better chiral separation capability than the CSPs of N-hexanoylated chitosan derivatives as a whole. Compared with the CSPs of CDMPC and ADMPC, the CSPs prepared with chitosan bis(3-methylphenylcarbamate)-(butyamide) and chitosan bis(3,4-dichlorophenylcarbamate)-(butyramide) significantly showed a stronger chiral separation capability.