Polysaccharide extract of dioscorea sp. and an orally active pharmaceutical composition comprising the same

Abstract

This invention provides a Dioscorea polysaccharide extract prepared by the method of this invention, which enhances the immunological activities by regulating the gene expression of cytokines. This invention further provides a Dioscorea polysaccharide extract, which has a synergistic effect with an oral vaccine and is capable of eliminating oral tolerance.

1 - 27 . (canceled)  
     
     
         28 . A method of inducing mucosal IgA antibody responses in a subject comprising orally co-administering to the subject an oral vaccine and a composition comprising a polysaccharide extract of  Dioscorea  sp.  
     
     
         29 . The method according to  claim 28 , wherein the polysaccharide extract of  Dioscorea  sp. in the oral composition is prepared by a process comprising the steps of: 
 (1) extracting a tuber of  Dioscorea  sp. using a 40% methanol solution in the presence of 1 % acetic acid, followed by separation, so as to obtain an insoluble solid portion;    (2) subjecting the insoluble solid portion obtained in step (1) to an aqueous solvent in the presence of a starch hydrolyzing enzyme to obtain an aqueous solution; and    (3) treating the aqueous solution obtained in step (2) with a 75% alcohol based solvent to obtain a precipitated solid portion; and    (4) treating the precipitated solid portion obtained in step (3) with a deproteinizing agent to remove proteinaceous substances therefrom.    
     
     
         30 . The method according to  claim 29 , wherein in step (2) the starch hydrolyzing enzyme used is α-amylase.  
     
     
         31 . The method according to  claim 30 , wherein in step (2), the amount of α-amylase used is 0.6% by weight.  
     
     
         32 . The method according to  claim 29 , wherein step (2) is carried out at an elevated temperature.  
     
     
         33 . The method according to  claim 29 , wherein in step (4) the solvent is a mixture of chloroform and 1-butanol.  
     
     
         34 . The method according to  claim 28 , wherein the oral co-administration to 
 a subject of the oral vaccine and the composition containing a polysaccharide extract of  Dioscorea  sp. induces expression of a gene encoding a cytokine in intestinal epithelium lymphocytes, lamina propria, and Peyer's patch.    
     
     
         35 . The method according to  claim 34 , wherein the cytokine encoded by the gene is selected from the group consisting of IL-2, IFN-γ, IL-4, IL-5, IL-6, IL-10, and TGF-β.  
     
     
         36 . The method according to  claim 28 , wherein the oral co-administration of the oral vaccine and the composition containing a polysaccharide extract of  Dioscorea  sp. enhances the systemic IgG antibody responses of a subject.  
     
     
         37 . The method according to  claim 29 , wherein the oral vaccine contains a microbial antigen.  
     
     
         38 . The method according to  claim 29 , wherein the oral vaccine is Pneumovax 23.  
     
     
         39 . A method for inhibiting oral tolerance of an oral vaccine induced by a protein antigen contained in the oral vaccine, comprising orally co-administering the oral vaccine containing the protein antigen, with a composition containing a polysaccharide extract of  Dioscorea  sp.  
     
     
         40 . The method according to  claim 39 , wherein the polysaccharide extract of  Dioscorea  sp. containing composition is prepared by a process comprising the steps of: 
 (1) extracting a tuber of  Dioscorea  sp. using a 40% methanol solution in the presence of 1% acetic acid, followed by separation, so as to obtain an insoluble solid portion;    (2) subjecting the insoluble solid portion obtained in step (1) to an aqueous solvent in the presence of a starch hydrolyzing enzyme to obtain an aqueous solution; and    (3) treating the aqueous solution obtained in step (2) with a 75% alcohol based solvent to obtain a precipitated solid portion; and    (4) treating the precipitated solid portion obtained in step (3) with a deproteinizing agent to remove proteinaceous substances therefrom.    
     
     
         41 . The method according to  claim 40 , wherein in step (2) the starch hydrolyzing enzyme used is α-amylase.  
     
     
         42 . The method according to  claim 41 , wherein in step (2) the amount of α-amylase used is 0.6% by weight.  
     
     
         43 . The method according to  claim 40 , wherein step (2) is carried out at an elevated temperature.  
     
     
         44 . The method according to  claim 40 , wherein in step (4) the solvent is mixture of chloroform and 1-butanol.  
     
     
         45 . The method according to  claim 39 , wherein the oral co-administration of the oral vaccine containing the protein antigen with the composition containing a polysaccharide extract of  Dioscorea  sp. induces the expression of a gene encoding a cytokine in intestinal epithelium lymphocytes, lamina propria, and Peyer's patch.  
     
     
         46 . The method according to  claim 45 , wherein the cytokine encoded by the gene is selected from the group consisting of IL-2, IFN-γ, IL-4, IL-5, IL-6, IL-10, and TGF-β.  
     
     
         47 . The method according to  claim 39 , wherein the oral co-administration of the oral vaccine containing the protein antigen and the composition containing a polysaccharide extract of  Dioscorea  sp. enhances systemic IgG and mucosal IgA antibodies responses.  
     
     
         48 . The method according to  claim 39 , wherein the oral vaccine contains a microbial antigen.