Patent Publication Number: US-2009238898-A1

Title: Method of inhibiting the proliferation and migration of helicobactor pylori

Description:
TECHNICAL FIELD 
     The present invention relates to a method for inhibiting the proliferation and migration of  Helicobacter pylori  strains that are considered a cause of gastric ulcer, duodenal ulcer, stomach cancer, etc. 
     More specifically, the present invention relates to a method for inhibiting the proliferation and migration of  Helicobacter pylori  strains in a manner not producing resistant bacteria to, or harmful side effects from treatment drugs of,  Helicobacter pylori  strains that are indigenous to the host. 
     For clarification purposes, in the context of the present invention a method for inhibiting the proliferation and migration of  Helicobacter pylori  strains refers to both a method for inhibiting the proliferation of  Helicobacter pylori  strains and a method for inhibiting the active migration of  Helicobacter pylori  strains. 
     Pumping of deep sea water began in Japan in 1989 off the coast of Cape Muroto in Kochi Prefecture using a deep sea water collection facility constructed on land. Today, deep sea water is collected in various parts of Japan including Toyama Prefecture and Okinawa Prefecture. It is well-known that deep sea water has various beneficial characteristics such as containing a lot of nutrients, being clean, and remaining stable at low temperature. Accordingly, many products that use deep sea water have been developed in recent years. Among others, there is a trend of active utilization of deep sea water in the food industry in the production of drinking water, processed seafood products, fermented products, etc., and a number of products using deep sea water are now available on the market. Many of these products are drawing the attention as offering the benefits of deep sea water such as cleanliness and high mineral content, and utilized as a pure material used in product manufacturing or a source of quality minerals. 
     On the other hand, medical and scientific studies are gradually finding the functions and mechanisms embodied in the human body by drinking water that uses deep sea water and thereby contains a lot of minerals. Although deep sea water is expected to offer numerous benefits, such as regulating the actions of the intestines, having positive effects on the immune strength of the body, and preventing lifestyle-related diseases, the specific actions and effects of deep sea water including those mentioned above are not yet understood fully. 
     On the other hand,  Helicobacter pylori  strains infect and colonize in the human stomach and are reported to have a hand in the development of diseases of the upper gastrointestinal tracts as well as autoimmune diseases, acute coronary diseases, and lifestyle-related diseases, among others. In particular, it is a well-known fact that  Helicobacter pylori  strains are closely related to the causes of gastric ulcer, duodenal ulcer, stomach cancer, etc. 
     For this reason, various therapies are used to remove  Helicobacter pylori  strains, such as those using various antibacterial agents, proton pump inhibitors (PPIs) that inhibit secretion of gastric acid, and antibiotics. As a specific example, a local oral agent that contains a bismuth compound, such as bismuth citrate, to kill  Helicobacter pylori  strains in the oral cavity is disclosed in Patent Literature 1 (Japanese Patent Laid-open No. Hei 8-20543). Also, Patent Literature 2 (Japanese Patent Laid-open No. Hei 8-48629) and Patent Literature 3 (Japanese Patent Laid-open No. Hei 9-208578) disclose anti-pylori agents exhibiting excellent antibacterial activity against  Helicobacter pylori  strains, where the effective ingredient is pyridone carboxylic acid, its ester, or salt thereof. 
     In addition to the above, Patent Literature 4 (Japanese Patent Laid-open No. Hei 9-295938) discloses an ulcer treatment drug, whose effective ingredients include a carbapenem compound or its pharmacologically acceptable form of salt having strong antibacterial activity against  Helicobacter pylori  strains that colonize and grow in the internal membranes of the stomach, for use as a drug to treat digestive ulcers and chronic superficial gastritis as well as prevent their recurrence. 
     In addition, Patent Literature 5 (Japanese Patent Laid-open No. 2000-63280) and Patent Literature 6 (Japanese Patent Laid-open No. Hei 10-109942) also disclose drugs that combine coptis root, cork tree bark or other galenical preparation in powder or extract form having antibacterial activity against  Helicobacter pylori  strains, as an effective medicinal composition to treat or prevent gastrointestinal diseases by effectively inhibiting the proliferation of, or removing and discharging,  Helicobacter pylori  strains that are considered a cause of the so-called digestive ulcers such as gastritis, gastric ulcer and duodenal ulcer, with at least one ingredient selected from histamine H 2  receptor antagonists, proton pump inhibitors, gastric-mucosa protective drugs for gastritis and digestive ulcers, gastric anti-acid drugs, and anti-diarrheal drugs. 
     Furthermore, Patent Literature 7 (Japanese Patent Laid-open No. Hei 11-180888) discloses a polyphenol obtained from fruits of rosaceous plants as an effective ingredient for use in drugs, anti-infection agents, or food, having an antibacterial action against  Helicobacter pylori  strains, while Patent Literature 8 (Japanese Patent Laid-open No. Hei 11-292788) discloses a Fe-bonded lactoferrin for use in anti-infection agents, drugs or beverages/food. Similarly, Patent Literature 9 (Japanese Patent Laid-open No. 2002-68992) discloses a beverage/food or food additive containing tea polyphenol as an effective ingredient along with a proton inhibitor, while Patent Literature 10 (Japanese Patent Laid-open No. 2005-68014) discloses an antibacterial agent against  Helicobacter pylori  strains, constituted by one or two or more extracts selected from the group that includes  Apocynum venetum  L.,  Vaccinium myrtillus  L. and  Acanthopanax senticosus  Harms. 
     However, the types of  Helicobacter pylori  strains present in the body are different from one host to another, and accordingly the effects of general antibacterial agents are not clear and cases of unsuccessful removal of  Helicobacter pylori  due to emergence of resistant bacteria and side effects are increasing. 
     For the aforementioned reasons, a number of simple methods for detecting and identifying  Helicobacter pylori  strains have been reported, including: an infection diagnosis probe useful in the detection and identification of  Helicobacter pylori  strains themselves, as disclosed in Patent Literature 11 (Japanese Patent Laid-open No. Hei 10-33179); a real-time infection inspection method based on the principles of “analysis of expired gas using  13 C labeled urea,” as disclosed in Patent Literature 12 (Japanese Patent Laid-open No. Hei 10-87512; a primer based on  Helicobacter pylori  strain genes and a method for examining clarithromycin resistance of  Helicobacter pylori  strains, as disclosed in Patent Literature 13 (Japanese Patent Laid-open No. Hei 10-285099) and Patent Literature 14 (Japanese Patent Laid-open No. 2001-321197); and a detection method using an enzyme urease derived from  Helicobacter pylori  strains, as disclosed in Patent Literature 15 (Japanese Patent Laid-open No. Hei 11-318490). However, utilization of deep sea water in the inhibition of proliferation or migration of  Helicobacter pylori  strains has not been reported to date. 
     Patent Literature 1: Japanese Patent Laid-open No. Hei 8-20543
 
Patent Literature 2: Japanese Patent Laid-open No. Hei 8-48629
 
Patent Literature 3: Japanese Patent Laid-open No. Hei 9-208578
 
Patent Literature 4: Japanese Patent Laid-open No. Hei 9-295938
 
     Patent Literature 5: Japanese Patent Laid-open No. 2000-63280 
     Patent Literature 6: Japanese Patent Laid-open No. Hei 10-109942
 
Patent Literature 7: Japanese Patent Laid-open No. Hei 11-180888
 
Patent Literature 8: Japanese Patent Laid-open No. Hei 11-292788
 
     Patent Literature 9: Japanese Patent Laid-open No. 2002-68992 
     Patent Literature 10: Japanese Patent Laid-open No. 2005-68014 
     Patent Literature 11: Japanese Patent Laid-open No. Hei 10-33179
 
Patent Literature 12: Japanese Patent Laid-open No. Hei 10-87512
 
Patent Literature 13: Japanese Patent Laid-open No. Hei 10-285099
 
     Patent Literature 14: Japanese Patent Laid-open No. 2001-321197 
     Patent Literature 15: Japanese Patent Laid-open No. Hei 11-318490 
     SUMMARY OF THE INVENTION 
     Problems to Be Solved by the Invention 
     As mentioned above, methods for preventing or treating gastrointestinal diseases are being sought, which are able to accommodate the diversity of  Helicobacter pylori  strains closely related to the causes of gastric ulcer, duodenal ulcer, stomach cancer, etc., and thereby suppress side effects of antibacterial agents and emergence of resistant bacteria. In the meantime, inhibition of proliferation and migration of  Helicobacter pylori  strains itself provides a means for prevention and treatment of diseases caused by  Helicobacter pylori  strains. When prevention is considered, it is safer for the health of human body and also provides an easier method of intake to continuously ingest as part of meals a food matter having the effect of inhibiting the proliferation and migration of  Helicobacter pylori  strains, instead of using a drug regularly. 
     In light of the above, it is an object of the present invention to achieve inhibition of proliferation and migration of  Helicobacter pylori  strains through utilization of inexpensive materials, such as drinking water and other substances we consume daily, because such is an ideal way to inhibit the proliferation and migration of  Helicobacter pylori  strains. 
     It is another object of the present invention to establish a technology for inhibiting the proliferation and migration of  Helicobacter pylori  strains that colonize in and infect the human stomach and are also reported to have a hand in diseases of the upper gastrointestinal tracts as well as autoimmune diseases, acute coronary diseases, and lifestyle-related diseases, among others, by focusing on deep sea water, which is rich in nutrients and minerals and inexpensive and therefore used widely in the food industry for use in the production of drinking water, processed seafood products, fermented products, etc., because deep sea water can be consumed easily in everyday life as drinking water. 
     Means for Solving the Problems 
     In light of the above, the present invention establishes a technology for inhibiting the proliferation and migration of  Helicobacter pylori  strains by utilizing drinking water and other substances consumed daily, based on a discovery of a method for inhibiting the proliferation and migration of  Helicobacter pylori  strains through substances containing minerals. To be specific, the present invention represents a newly discovered method for inhibiting the proliferation and migration of  Helicobacter pylori  strains by utilizing a liquid that contains at least one or more of magnesium, calcium, sodium and potassium, based on the results of examination with focus on magnesium, calcium, sodium and potassium that exist in particularly high quantities in deep sea water among the various minerals contained in deep sea water, and also with focus on liquids that can be used as a way to ingest these minerals easily in everyday life, and consequently the present invention establishes a technology for inhibiting the proliferation and migration of  Helicobacter pylori  strains by utilizing drinking water and other substances consumed daily. The minerals examined in connection with the present invention are derived from deep sea water subject to minimal contamination by harmful substances, etc., and thus can be used favorably as materials for beverages and food. However, it is not necessary to limit the source of these minerals to deep sea water. If the mineral content per kilogram is 40 mg or more, proliferation and migration of  Helicobacter pylori  strains can be inhibited. 
     If the mineral content per kilogram is 5,900 mg or more, proliferation and migration of all  Helicobacter pylori  strains can be inhibited. From the viewpoint of cost effectiveness, there is no need to increase the mineral content to beyond 5,900 mg per kilogram. 
     EFFECTS OF THE INVENTION 
     The greatest benefit of the present invention is that by consuming mineral-rich water obtained from deep sea water that is conveniently available, proliferation and migration of  Helicobacter pylori  strains indigenous to the host can be inhibited to prevent and treat diseases of the upper gastrointestinal tracts, among others, without being concerned about emergence of resistant bacteria and side effects of treatment drugs. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1 . Condition of colonies that grew in an agar culture medium 
     
    
    
     BEST MODE FOR CARRYING OUT THE INVENTION 
     [Method for Testing Inhibition of Proliferation of  Helicobacter Pylori  Strains] 
     The method for testing the inhibition of proliferation of  Helicobacter pylori  strains used in the present invention is explained below. 
     Each type of  Helicobacter pylori  strains corresponding to approx. 1 platinum loop was taken from an agar culture medium (Table 1) on the second day of culture and suspended in 1 ml of 2.8%  Brucella  broth culture solution so that O.D. 600  became 0.3. 
     The prepared solution was then diluted to 10 −4 , 10 −5  and 10 −6  times, respectively, using sterilized water, and each bacteria-suspended solution was dripped by 10 μl each into agar culture mediums (Table 3) containing mineral-rich waters A, B, C, D, E, F and G (test waters) prepared from deep sea water as shown in Table 2. Next, the culture mediums were cultured for three days in an environment of 37° C. and 10% CO 2 , after which the CFU (colony forming unit=viable bacterial cell count) of each culture medium was measured. 
     For your reference, Milli-Q water was used as a control water. 
     
       
         
           
               
             
               
                 TABLE 1 
               
             
            
               
                   
               
               
                 Composition of agar culture medium 
               
            
           
           
               
               
               
            
               
                   
                 Composition 
                 Content 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                   Brucella  Broth 
                 2.8 
                 g 
               
               
                   
                 Milli-Q water 
                 90 
                 ml 
               
               
                   
                 Agar powder 
                 1.4 
                 g 
               
               
                   
                 Horse serum 
                 10 
                 ml 
               
               
                   
                 Vancomycin solution (100 mg/ml) 
                 10 
                 μl 
               
               
                   
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 2 
               
             
            
               
                   
               
               
                 Compositions of mineral-rich waters (mg/kg) 
               
            
           
           
               
               
               
            
               
                 Mineral-rich 
                 Ion type 
                   
               
            
           
           
               
               
               
               
               
            
               
                 water 
                 Mg 
                 Ca 
                 Na 
                 K 
               
               
                   
               
               
                 A 
                 100 
                 200 
                 50 
                 50 
               
               
                 B 
                 150 
                 150 
                 50 
                 50 
               
               
                 C 
                 200 
                  70 
                 65 
                 65 
               
               
                 D 
                 400 
                 — 
                 — 
                 — 
               
               
                 E 
                 — 
                 400 
                 — 
                 — 
               
               
                 F 
                 — 
                 — 
                 400  
                 — 
               
               
                 G 
                 — 
                 — 
                 — 
                 400  
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 3 
               
             
            
               
                   
               
               
                 Composition of agar culture medium 
               
            
           
           
               
               
               
            
               
                   
                 Composition 
                 Content 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                   Brucella  Broth 
                 2.8 
                 g 
               
               
                   
                 Test water or control water 
                 90 
                 ml 
               
               
                   
                 Agar powder 
                 1.4 
                 g 
               
               
                   
                 Horse serum 
                 10 
                 ml 
               
               
                   
                 Vancomycin solution (100 mg/ml) 
                 10 
                 μl 
               
               
                   
                   
               
            
           
         
       
     
     [ Helicobacter Pylori  Strains Used] 
     In the test, mainly clinically isolated strains were used. 
     KMT series: Clinically isolated strain obtained from the Medical School of Kochi University
 
NY series: Strain resistant to antibiotics (metronidazole and clarithromycin) (made in Japan)
 
26695: Clinically isolated strain analyzed in details to gene level (obtained from Europe)
 
NCTC11637: Clinically isolated strain (obtained from the U.S.)
 
HPK5: Clinically isolated strain (obtained in Japan)
 
HPKT510: Broken strain of cdrA gene (gene relating to cell division) of HPK5 prepared by means of gene recombination
 
     [Method for Evaluating Proliferation Inhibition] 
     The effects of inhibiting the proliferation of  Helicobacter pylori  strains in test waters A, B, C, D, E, F and G were determined based on the CFU (colony forming unit=viable bacterial cell count) of the control water being 100%. 
     By considering the variations resulting from the operations carried out in the test, test waters whose CFU was smaller than 80% of the CFU of the control water were considered effective. 
     The test was conducted three times and if effectiveness was found in two test runs, the test water was judged effective in inhibiting  Helicobacter pylori  proliferation (indicated by ◯). 
     [Method for Testing Inhibition of Migration of  Helicobacter Pylori  Strains] 
     Using a sterilized toothpick, bacteria of each type were taken from agar culture mediums in which  Helicobacter pylori  strains had been cultured, and then transferred to  Brucella  broth agar culture mediums (Table 4) using mineral-rich waters A, B, C, D, E, F and G prepared from deep sea water and also using Milli-Q water as a control. Next, the culture mediums were cultured for three days in an environment of 37° C. and 10% CO 2 , after which the diameters of colonies in each culture medium were measured. 
     
       
         
           
               
             
               
                 TABLE 4 
               
             
            
               
                   
               
               
                 Composition of agar culture medium 
               
            
           
           
               
               
               
            
               
                   
                 Composition 
                 Content 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                   Brucella  Broth 
                 2.8 
                 g 
               
               
                   
                 Test water or control water 
                 90 
                 ml 
               
               
                   
                 Agar powder 
                 0.35 
                 g 
               
               
                   
                 Horse serum 
                 10 
                 ml 
               
               
                   
                 Vancomycin solution (100 mg/ml) 
                 10 
                 μl 
               
               
                   
                   
               
            
           
         
       
     
     [Method for Evaluating Migration Inhibition] 
     In the evaluation, test waters whose colonies had a diameter smaller than the diameters of colonies in the control water were considered effective. The test was conducted three times and if effectiveness was found in two test runs, the test water was judged effective in inhibiting  Helicobacter pylori  migration (indicated by ⊙). 
     For your reference, colonies with a diameter less than 2 mm in the control were considered not sufficiently grown and not counted. 
     The present invention is explained below using examples with respect to test waters A, B, C, D, E, F and G. It should be noted, however, that mineral-rich waters prepared form deep sea water are not at all limited to these waters. 
     Example 1 
     Table 5 shows the effects of test waters in inhibiting the proliferation of various types of  Helicobacter pylori  strains. 
     
       
         
           
               
             
               
                 TABLE 5 
               
             
            
               
                   
               
               
                 Effects of inhibiting proliferation of various 
               
               
                 types of  Helicobacter pylori  strains 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                   
                   
               
               
                 2 
                 KMT31 
                   
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 3 
                 KMT40 
                   
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 4 
                 KMT44 
                   
                 ◯ 
                   
                 ◯ 
                   
                   
                 ◯ 
               
               
                 5 
                 KMT45 
                 ◯ 
                 ◯ 
                   
                 ◯ 
                   
                 ◯ 
               
               
                 6 
                 KMT46 
                   
                   
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 7 
                 KMT47 
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 8 
                 KMT50 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 9 
                 NY1 
                   
                   
                   
                   
                   
                 ◯ 
                 ◯ 
               
               
                 10 
                 NY8 
                   
                   
                 ◯ 
                 ◯ 
               
               
                 11 
                 NY11 
               
               
                 12 
                 NY31 
                   
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 13 
                 26695 
               
               
                 14 
                 HPK5 
                   
                   
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 15 
                 HPKT510 
                 ◯ 
                   
                   
                   
                 ◯ 
               
               
                 16 
                 11637 
                 ◯ 
                 ◯ 
                   
                 ◯ 
               
               
                   
               
            
           
         
       
     
     As shown in Table 5, whether proliferation of a given type of bacterial strain was inhibited or not varied depending on the type of test water. 
     Also, it is evident from Table 5 that test water D inhibited the proliferation of many types of bacterial strains, where clearly the effect of inhibiting  Helicobacter pylori  proliferation varies according to the type of bacterial strain. 
     Based on the above results, it is found that by consuming mineral-rich water conveniently available and obtained from deep sea water having the effect of inhibiting the proliferation of various types of  Helicobacter pylori  strains, diseases of the upper gastrointestinal tracts, among others, can be prevented and treated without being concerned about emergence of resistant bacteria and side effects of treatment drugs. 
     Example 2 
     Table 6 shows the effects of inhibiting the migration of various types of  Helicobacter pylori  strains cultured in agar culture mediums using mineral-rich waters obtained from deep sea water. 
     
       
         
           
               
             
               
                 TABLE 6 
               
             
            
               
                   
               
               
                 Effects of inhibiting migration of various 
               
               
                 types of  Helicobacter pylori  strains 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                   
                   
                   
                   
                   
                   
                   
               
               
                 2 
                 KMT31 
                 ⊙ 
                   
                   
                   
                 ⊙ 
               
               
                 3 
                 KMT40 
               
               
                 4 
                 KMT44 
                   
                 ⊙ 
                 ⊙ 
                 ⊙ 
                 ⊙ 
               
               
                 5 
                 KMT45 
                   
                 ⊙ 
                 ⊙ 
                 ⊙ 
                 ⊙ 
               
               
                 6 
                 KMT46 
                   
                 ⊙ 
                 ⊙ 
                 ⊙ 
                 ⊙ 
               
               
                 7 
                 KMT47 
               
               
                 8 
                 KMT50 
               
               
                 9 
                 NY1 
               
               
                 10 
                 NY8 
                 ⊙ 
                 ⊙ 
                   
                   
                 ⊙ 
               
               
                 11 
                 NY11 
                 ⊙ 
                 ⊙ 
                   
                   
                 ⊙ 
               
               
                 12 
                 NY31 
               
               
                 13 
                 26695 
               
               
                 14 
                 HPK5 
                   
                 ⊙ 
                 ⊙ 
                   
                 ⊙ 
               
               
                 15 
                 HPKT510 
                   
                   
                   
                   
                 ⊙ 
               
               
                 16 
                 11637 
               
               
                   
               
            
           
         
       
     
     As shown in Table 6, test waters B, C, D, and E are effective in inhibiting the migration of KMT44, 45 and 46 strains. 
     Based on the above results, it is found that by selecting proper mineral-rich waters, migration of  Helicobacter pylori  strains infecting and colonized in the host can be inhibited. 
     Example 3 
     Starting from mineral-rich waters each having a total mineral content of 400 mg/kg, the total mineral content of each water was changed within a range of 20 to 11,800 mg/kg based on the percentages of ion contents shown in Table 7, and the effects of varying total mineral concentrations on the inhibition of proliferation of  Helicobacter pylori  strains were examined. 
     
       
         
           
               
             
               
                 TABLE 7 
               
             
            
               
                   
               
               
                 Percentages of ion contents in various mineral waters (%) 
               
            
           
           
               
               
               
            
               
                 Mineral-rich 
                 Ion type 
                   
               
            
           
           
               
               
               
               
               
            
               
                 water 
                 Mg 
                 Ca 
                 Na 
                 K 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 A 
                 25 
                 50 
                 12.5 
                 12.5 
               
               
                 B 
                 37.5 
                 37.5 
                 12.5 
                 12.5 
               
               
                 C 
                 50 
                 17.5 
                 16.25 
                 16.25 
               
               
                 D 
                 100 
                 0 
                 0 
                 0 
               
               
                 E 
                 0 
                 100 
                 0 
                 0 
               
               
                 F 
                 0 
                 0 
                 100 
                 0 
               
               
                 G 
                 0 
                 0 
                 0 
                 100 
               
               
                   
               
            
           
         
       
     
     By considering the variations resulting from the operations carried out in the test, test waters whose CFU was smaller than 80% of the CFU of the control water were considered effective. The test was conducted three times and if effectiveness was found in two test runs, the test water was judged effective in inhibiting  Helicobacter pylori  proliferation (indicated by $$◯$$). 
     For your reference, the strains used in the test were those  Helicobacter pylori  strains indicated in 1 through 16. 
     Tables 8 through 15 show the results of the examination where the total mineral concentration was gradually raised from 20 to 11,800 mg/kg. 
     
       
         
           
               
             
               
                 TABLE 8 
               
             
            
               
                   
               
               
                 Total mineral concentration = 20 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                   
                   
                   
                   
                   
                   
                   
               
               
                 2 
                 KMT31 
               
               
                 3 
                 KMT40 
               
               
                 4 
                 KMT44 
               
               
                 5 
                 KMT45 
               
               
                 6 
                 KMT46 
               
               
                 7 
                 KMT47 
               
               
                 8 
                 KMT50 
               
               
                 9 
                 NY1 
               
               
                 10 
                 NY8 
               
               
                 11 
                 NY11 
               
               
                 12 
                 NY31 
               
               
                 13 
                 26695 
               
               
                 14 
                 HPK5 
               
               
                 15 
                 HPKT510 
               
               
                 16 
                 11637 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 9 
               
             
            
               
                   
               
               
                 Total mineral concentration = 40 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                   
                   
                 ◯ 
                 ◯ 
                   
                   
                   
               
               
                 2 
                 KMT31 
               
               
                 3 
                 KMT40 
               
               
                 4 
                 KMT44 
               
               
                 5 
                 KMT45 
               
               
                 6 
                 KMT46 
               
               
                 7 
                 KMT47 
               
               
                 8 
                 KMT50 
               
               
                 9 
                 NY1 
               
               
                 10 
                 NY8 
               
               
                 11 
                 NY11 
               
               
                 12 
                 NY31 
                   
                 ◯ 
                   
                 ◯ 
               
               
                 13 
                 26695 
               
               
                 14 
                 HPK5 
               
               
                 15 
                 HPKT510 
               
               
                 16 
                 11637 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 10 
               
             
            
               
                   
               
               
                 Total mineral concentration = 100 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                   
                   
                 ◯ 
                 ◯ 
                   
                   
                   
               
               
                 2 
                 KMT31 
               
               
                 3 
                 KMT40 
                   
                   
                   
                   
                   
                 ◯ 
               
               
                 4 
                 KMT44 
                   
                   
                   
                   
                   
                   
                 ◯ 
               
               
                 5 
                 KMT45 
               
               
                 6 
                 KMT46 
               
               
                 7 
                 KMT47 
               
               
                 8 
                 KMT50 
                   
                 ◯ 
                 ◯ 
               
               
                 9 
                 NY1 
               
               
                 10 
                 NY8 
               
               
                 11 
                 NY11 
               
               
                 12 
                 NY31 
                   
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 13 
                 26695 
               
               
                 14 
                 HPK5 
               
               
                 15 
                 HPKT510 
               
               
                 16 
                 11637 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 11 
               
             
            
               
                   
               
               
                 Total mineral concentration = 1,200 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 2 
                 KMT31 
                   
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 3 
                 KMT40 
                   
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 4 
                 KMT44 
                 ◯ 
                 ◯ 
                   
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 5 
                 KMT45 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 6 
                 KMT46 
                   
                   
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 7 
                 KMT47 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 8 
                 KMT50 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 9 
                 NY1 
                   
                   
                   
                   
                   
                 ◯ 
                 ◯ 
               
               
                 10 
                 NY8 
                   
                   
                 ◯ 
                 ◯ 
               
               
                 11 
                 NY11 
               
               
                 12 
                 NY31 
                   
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 13 
                 26695 
               
               
                 14 
                 HPK5 
                   
                   
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 15 
                 HPKT510 
                 ◯ 
                   
                   
                   
                 ◯ 
               
               
                 16 
                 11637 
                 ◯ 
                 ◯ 
                   
                 ◯ 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 12 
               
             
            
               
                   
               
               
                 Total mineral concentration = 2,500 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 2 
                 KMT31 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 3 
                 KMT40 
                   
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 4 
                 KMT44 
                 ◯ 
                 ◯ 
                   
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 5 
                 KMT45 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 6 
                 KMT46 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 7 
                 KMT47 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 8 
                 KMT50 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 9 
                 NY1 
                   
                   
                   
                   
                   
                 ◯ 
                 ◯ 
               
               
                 10 
                 NY8 
                   
                   
                 ◯ 
                 ◯ 
               
               
                 11 
                 NY11 
               
               
                 12 
                 NY31 
                   
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 13 
                 26695 
                   
                   
                   
                   
                   
                   
                 ◯ 
               
               
                 14 
                 HPK5 
                   
                   
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 15 
                 HPKT510 
                 ◯ 
                   
                   
                   
                 ◯ 
               
               
                 16 
                 11637 
                 ◯ 
                 ◯ 
                   
                 ◯ 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 13 
               
             
            
               
                   
               
               
                 Total mineral concentration = 3,500 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 2 
                 KMT31 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 3 
                 KMT40 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 4 
                 KMT44 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 5 
                 KMT45 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 6 
                 KMT46 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 7 
                 KMT47 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 8 
                 KMT50 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 9 
                 NY1 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 10 
                 NY8 
                   
                   
                 ◯ 
                 ◯ 
               
               
                 11 
                 NY11 
                 ◯ 
                   
                   
                   
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 12 
                 NY31 
                   
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 13 
                 26695 
               
               
                 14 
                 HPK5 
                   
                   
                 ◯ 
                 ◯ 
                   
                 ◯ 
                 ◯ 
               
               
                 15 
                 HPKT510 
                 ◯ 
                   
                   
                   
                 ◯ 
               
               
                 16 
                 11637 
                 ◯ 
                 ◯ 
                   
                 ◯ 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 14 
               
             
            
               
                   
               
               
                 Total mineral concentration = 5,900 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 2 
                 KMT31 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 3 
                 KMT40 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 4 
                 KMT44 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 5 
                 KMT45 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 6 
                 KMT46 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 7 
                 KMT47 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 8 
                 KMT50 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 9 
                 NY1 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 10 
                 NY8 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 11 
                 NY11 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 12 
                 NY31 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 13 
                 26695 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 14 
                 HPK5 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 15 
                 HPKT510 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 16 
                 11637 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 15 
               
             
            
               
                   
               
               
                 Total mineral concentration = 11,800 mg/kg 
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 No. 
                 Bacteria name 
                 A 
                 B 
                 C 
                 D 
                 E 
                 F 
                 G 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
            
               
                 1 
                 KMT27 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 2 
                 KMT31 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 3 
                 KMT40 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 4 
                 KMT44 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 5 
                 KMT45 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 6 
                 KMT46 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 7 
                 KMT47 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 8 
                 KMT50 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 9 
                 NY1 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 10 
                 NY8 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 11 
                 NY11 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 12 
                 NY31 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 13 
                 26695 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 14 
                 HPK5 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 15 
                 HPKT510 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                 16 
                 11637 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
                 ◯ 
               
               
                   
               
            
           
         
       
     
     Increasing the total mineral concentration in mineral-rich waters prepared from deep sea water significantly increased the types of bacterial strains whose proliferation could be inhibited, and once the total mineral concentration reached 5,900 mg/kg, these mineral-rich waters were found effective in inhibiting the proliferation of all types of  Helicobacter pylori  strains. In addition, the effects of inhibiting the proliferation of  Helicobacter pylori  strains became particularly prominent when the magnesium concentration was high.