HERIPENES WITH PAIN-RELIEVING EFFECT, ACTIVE SUBSTANCES OF Hericium erinaceus MYCELIUM AND THE PREPARATION METHOD THEREOF, AND PHARMACEUTICAL COMPOSITION CONTAINING THE HERIPENES OR ACTIVE SUBSTANCES

Abstract

The present invention is related to an active substance of Hericium erinaceus having a pain-relieving effect, and a pharmaceutical composition including the active substance. The active substance is prepared using the following steps: (a) inoculating a mycelium of H. erinaceus on an agar plate and incubating at 15-32° C. for 8-16 days; (b) inoculating the incubated H. erinaceus mycelia from step (a) into a medium in a flask and incubating at 20-30° C. and pH 4.5-6.5 for 3-5 days; (c) inoculating the incubated H. erinaceus mycelia from step (b) into a medium in a fermentation tank and incubating at 24-32° C. and pH 4.5-5.5 for 8-16 days to obtain a fermented medium of the H. erinaceus mycelia; and (d) desiccating the fermented medium of the H. erinaceus mycelia from step (c) to obtain the powder of the H. erinaceus mycelia, which is further purified and isolated to obtain a novel compound of H. erinaceus.

1 .- 24 . (canceled) 
     
     
         25 . A preparation method for an active substance of a Hericium  erinaceus  for relieving a pain, comprising:
 (a) inoculating an  H. erinaceus  mycelium on an agar plate to be incubated;   (b) inoculating the incubated mycelium in step (a) into a first medium on a small scale to be incubated; and   (c) inoculating the incubated mycelium in step (b) into a second medium on a large scale to be incubated to obtain a fermented medium containing the active substance.   
     
     
         26 . The preparation method according to  claim 25 , wherein the incubation in step (a) is performed at 15-32° C. for 8-16 days. 
     
     
         27 . The preparation method according to  claim 25 , wherein the incubation in step (b) is performed at 20-30° C., pH 4.5-6.5, and a shaking rate of 100-250 rounds per minutes (rpm) for 3-5 days. 
     
     
         28 . The preparation method according to  claim 25 , wherein in step (c) the second medium on the large scale is accommodated in a fermentation tank having a tank pressure of 0.8-1.2 kg/cm 2  and a stirring rate of 10-150 rpm, a gas is introduced into the fermentation tank at an aeration rate of 0.5-1 volume per volume per minute (vvm), the gas is one selected from a group consisting of air, oxygen, carbon dioxide, nitrogen gas and a combination thereof, and the incubation is performed at 24-32° C. and pH 4.5-5.5 for 8-16 days. 
     
     
         29 . The preparation method according to  claim 25 , wherein the first and the second media are the same. 
     
     
         30 . The preparation method according to  claim 29 , wherein each of the first and the second media comprises one selected from a group consisting of a complex carbon and nitrogen source, animal or plant sources of one of a protein and a hydrolyzate thereof, an inorganic salt, a saccharide, a yeast extract, a malt extract, a defoaming agent and a combination thereof, and the complex carbon and nitrogen source is one of a grain and a legume, and the inorganic salt is one of a sulfate and a phosphate. 
     
     
         31 . The preparation method according to  claim 25 , wherein the preparation method further comprises (d) desiccating the fermented medium to obtain a powder of the  H. erinaceus  mycelium. 
     
     
         32 . The preparation method according to  claim 31 , wherein the powder is further extracted with an alcohol solution to obtain an alcohol extract of the  H. erinaceus  mycelium, and the alcohol solution is an ethanol solution of 30-100 volume-volume percentage (v/v %) or a methanol solution of 30-100 (v/v %). 
     
     
         33 . The preparation method according to  claim 32 , wherein the alcohol extract is further extracted with water-ethyl acetate, followed by a column chromatography to obtain a heripene having a formula (I) as follows: 
       
         
           
           
               
               
           
         
         where R is one selected from a group consisting of hydrogen, hydroxyl, C 1 -C 10  alkyl, C 2 -C 10  alkenyl and C 2 -C 10  alkynyl, each of which is optionally substituted with a substitute bonding with one selected from a group consisting of halogen, oxygen, nitrogen, phosphorus and sulfur, and where the stereocenters at C-5, C-6, C-9, C-13, C-14, C-21, C22 and C-23 are one of an R configuration and an S configuration. 
       
     
     
         34 . The preparation method according to  claim 33 , wherein the heripene is an erinacine S having a formula (II) as follows: 
       
         
           
           
               
               
           
         
         when R included in the formula (I) is hydrogen. 
       
     
     
         35 . The preparation method according to  claim 33 , wherein the water-ethyl acetate has a ratio of 1:4 (v/v). 
     
     
         36 . The preparation method according to  claim 25 , wherein the pain is one of a neuropathic pain and a cancer pain. 
     
     
         37 . The preparation method according to  claim 36 , wherein the pain involves a signaling pathway with regard to a P2-purinoceptor (P2R). 
     
     
         38 . An active substance of a  Hericium erinaceus  for relieving a pain as obtained according to  claim 25 . 
     
     
         39 . The active substance according to  claim 38 , wherein the active substance is obtained in a form of a powder. 
     
     
         40 . The active substance according to  claim 39 , wherein the powder is extracted as one of an alcohol extract and a heripene having a formula (I) as follows: 
       
         
           
           
               
               
           
         
         where R is one selected from a group consisting of hydrogen, hydroxyl, C 1 -C 10  alkyl, C 2 -C 10  alkenyl and C 2 -C 10  alkynyl, each of which is optionally substituted with a substitute bonding with one selected from a group consisting of halogen, oxygen, nitrogen, phosphorus and sulfur, and where the stereocenters at C-5, C-6, C-9, C-13, C-14, C-21, C22 and C-23 are one of an R configuration and an S configuration. 
       
     
     
         41 . The active substance according to  claim 40 , wherein the heripene is an erinacine S having a formula (II) as follows: 
       
         
           
           
               
               
           
         
         when R included in the formula (I) is hydrogen. 
       
     
     
         42 . A pharmaceutical composition for a pain-relieving effect, comprising an active substance of a  Hericium erinaceus  according to  claim 36 , and one selected from a group consisting of a biologically acceptable carrier, an excipient, a diluent and an adjuvant. 
     
     
         43 . A heripene compound for a pain-relieving effect, including a formula (I) as follows: 
       
         
           
           
               
               
           
         
         where R is one selected from a group consisting of hydrogen, hydroxyl, C 1 -C 10  alkyl, C 2 -C 10  alkenyl and C 2 -C 10  alkynyl, each of which is optionally substituted with a substitute bonding with one selected from a group consisting of halogen, oxygen, nitrogen, phosphorus and sulfur; and
 where the stereocenters at C-5, C-6, C-9, C-13, C-14, C-21, C22 and C-23 are one of an R configuration and an S configuration. 
 
       
     
     
         44 . The heripene compound according to  claim 43 , wherein the heripene compound is an erinacine S having a formula (II) as follows: 
       
         
           
           
               
               
           
         
         when R included in the formula (I) is hydrogen.