Abstract:
This application relates to disinfectant formulations including calcium hydroxide and sodium hypochlorite and further relates to method of manufacturing the disinfectant formulations and methods of using the formulations.

Description:
CROSS REFERENCE TO PRIOR APPLICATIONS 
       [0001]    The present application claims priority under the Paris Convention to U.S. Application No. 61/698,076, filed Sep. 7, 2012, the entire contents of which are incorporated herein by reference. 
     
    
     FIELD OF THE INVENTION 
       [0002]    The present invention relates to disinfectant formulations and in particular to formulations comprising calcium hydroxide and sodium hypochlorite for disinfecting surfaces. 
       BACKGROUND OF THE INVENTION 
       [0003]    The health effects caused by bacteria, fungus, mold, viruses and other micro-organisms that can grow on surfaces is a serious concern, especially in the food service and preparation industry and in the medical field. 
         [0004]    Disinfectants are substances that are applied to non-living objects to destroy microorganisms that are living on the objects. Generally, disinfectants work by destroying the cell wall of microbes or interfering with the metabolism. Various disinfectants have been used with mixed results. Often the most efficacious disinfectants also have harmful side effects to the individuals who use them. 
         [0005]    For example, solutions containing sodium hypochlorite are commonly used as disinfectants however these products can release harmful chlorine vapours which can have negative health consequences such as impaired respiratory health. This is particularly the case where high concentrations of hypochlorite are used. Hypochlorite solutions are often combined with sodium hydroxide which acts to stabilize the hypochlorite and prevent decomposition to chlorine. However, the addition of the substantial amounts of sodium hydroxide required for stabilization, results in solutions that are highly caustic and therefore also harmful for those handling such solutions. 
         [0006]    There is a need for disinfecting agents that are effective in disinfecting surfaces but that are less harmful to those who have to use them. 
       SUMMARY OF THE INVENTION 
       [0007]    The present invention relates to a disinfectant formulation comprising calcium hydroxide (Ca(OH) 2 ) and sodium hypochlorite (NaClO). In one embodiment the disinfectant formulation is an aqueous solution comprising about 0.1% to about 5% v/v sodium hypochlorite (NaClO) and about 0.12% to about 0.18% v/v calcium hydroxide (Ca(OH) 2 ). 
         [0008]    In a particular embodiment the disinfectant formulation is prepared by forming a supernatant solution of calcium hydroxide in water having a concentration of about 0.12% to 0.18% and combining the supernatant with about 0.1% to 5% by volume of sodium hypochlorite. 
         [0009]    In a further aspect there is provided a method of disinfecting objects comprising applying the disinfecting formulation to said object to destroy microorganisms living on said object. Said microorganisms including bacteria, fungus, mold, viruses and/or other microbes. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0010]    The features of the invention will be described in relation to the appended drawings in which: 
           [0011]      FIG. 1  is a picture showing a typical zone of inhibition of 200 μl of Ca(OH)2 paste against 4 bacteria, overnight A)  S. Aureus,  B)  Salmonella,  C)  E. Coli  and D)  Pseudomonas aeruginosa.    
           [0012]      FIG. 2  shows a picture of a typical zone of inhibition of 200 μl of Ca(OH)2 paste against three fungi, A)  Aspergillus niger  (48 hr) B)  Auriobasidum pullulans  (48 hr) and C)  Chaetobium, glabosom  (72 hr). 
       
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
       [0013]    It has been found that Ca(OH) 2  acts to stabilize hypochlorite in solution. By stabilizing the hypochlorite, the release of toxic chlorine gas may be reduced. Further it has been found that less Ca(OH) 2  is required to stabilize hypochlorite in solution as compared to the amount of NaOH that would be required to achieve a similar result. Since less Ca(OH) 2  can be used, the resulting formulation can be less caustic and therefore less harmful to the user. 
         [0014]    It has also been found that the use of Ca(OH) 2  results in less precipitation than using NaOH to stabilize hypochlorite. This is particularly advantageous in the application of the disinfectant formulation as a spray disinfectant where the precipitates can cause spray nozzles to become clogged and malfunction. 
         [0015]    Ion channels in a cell serve many functions apart from electrical signal transduction: chemical signaling, (Ca 2+  as a second messenger), transepithelial transport, regulation of cytoplasmic or vesicular ion concentration and pH, and regulation of cell volume. Therefore, ion channel dysfunction can cause cell disruption and in the case of calcium channels, alter pH and cellular osmotic pressure. It is believed that the Ca(OH) 2  acts through the calcium channel to disrupt the normal cell function and ultimately kill the cell. Providing further antimicrobial action in the formulation. 
         [0016]    This mechanism of action of Ca(OH) 2  is believed to be different from the action of NaOH which acts on the cell membrane, lipids and proteins, thereby denaturing and weakening cell membrane surface structure and resulting in cell death. 
         [0017]    In an embodiment of the invention the disinfectant formulation comprises an aqueous solution with about 0.1% to about 5% v/v sodium hypochlorite (NaClO) and about 0.12% to about 0.18% v/v calcium hydroxide (Ca(OH) 2 ). In a further embodiment the solution comprises about 0.1% to about 3% v/v sodium hypochlorite. In another embodiment the solution comprises about 0.125% to about 2.5% sodium hypochlorite. In still another embodiment the solution comprises 0.125% sodium hypochlorite. In yet another embodiment the solution comprises 2.5% sodium hypochlorite. 
         [0018]    In a further aspect there is provided a method of manufacturing a disinfectant solution. In one embodiment the method comprises mixing 97% pure calcium hydroxide (Ca(OH) 2 ) dry powder with deionized water. In a particular embodiment the calcium hydroxide powder is 200 mesh size. In one embodiment the percentage ratio of calcium hydroxide to deionized water is about 2.5 to about 97.5 by weight, to produce a supernatant solution. The concentration of calcium hydroxide in the supernatant solution is controlled by temperature to be about 0.12% to about 0.18%. Sodium hypochlorite (NaClO) is added to the supernatant solution. The solution concentration of sodium hypochlorite is in the rage of from about 0.01% to about 5% by volume. In a further embodiment sodium hypochlorite is in a rage from about 0.1% to about 3% by volume. In a particular example the sodium hypochlorite concentration is about 0.125% by volume. In another example the sodium hypochlorite concentration is about 2.5% by volume. 
         [0019]    It has been further been found that incorporating a calcium hydroxide supernatant solution having a pH of about 12.1-12.6 in the disinfectant solution allows pH stability to be maintained over a longer period of time due to a high pH environment in the solution. 
         [0020]    In a further embodiment the disinfectant formulation is prepared in a compounding batch process using large mixing and storage tanks. The decantation of supernatant solution having solid concentration of about 0.12% to about 0.18% is controlled by temperature. From storage, the solution is pumped to a final mix tank where sodium hypochlorite solution is metered into the bottling lines. In a particular example, 0.125% of sodium hypochlorite is metered into the bottling line. 
         [0021]    In a further aspect the disinfectant formulation can be packaged in various types and sizes of containers for example 946 ml, 4 L and 10 L containers. 
         [0022]    In still a further aspect, the disinfectant formulation can be used to disinfect non-living objects. In a particular aspect the formulation can be used to disinfect hard surfaces by applying the formulation to the surface. In certain applications the formulation may be left on the surface for a specific period of time. For example, the disinfectant formulation may be left on the surface for a period ranging from about 5 minutes to about 24 hours. 
         [0023]    In a further aspect the disinfectant formulation may be used to kill microbes on an object to which it is applied. The microbes may be bacteria, fungus, mold, viruses and other micro-organisms. Examples of some micro-organisms which may be killed by the disinfectant formulation include Herpes Simplex Virus,  Staphylococcus aureus, Salmonella enteric, E. Coli, Pseudomonas aeruginosa, Aspergillus niger, Auriobasidium pullulans  and  Chaetobium globosom.  It will be understood by one of skill in the art that the above list of micro-organisms is merely provided to illustrate the invention and that the disinfecting formulation is expected to be active against other micro-organisms. 
         [0024]    Experiments 
         [0025]    A. Investigating the Antiviral Properties of Ca(OH) 2    
         [0026]    Experiments: 10 μL of virus stock was mixed with 100% saturated Ca(OH) 2  (Final Ca(OH) 2  concentration 99%) and incubated for different lengths of time. Then the mixture was diluted and the viable number of virus (Log TCID) was measured. 
         [0027]    Results: 
         [0028]    99% saturated Ca(OH)2 was used to treat 2 viruses using different incubation times (5 min to 1 hr). The preliminary result for HSV-1 is shown in table 1 below. 
         [0000]    
       
         
               
               
               
               
               
               
               
             
           
               
                 TABLE 1 
               
               
                   
               
               
                 Log 
                   
                   
                   
                   
                   
                   
               
               
                 TCID50/ 
               
               
                 100 ul 
                 0 min 
                 5 min 
                 10 min 
                 15 min 
                 30 min 
                 60 min 
               
               
                   
               
             
             
               
                 HSV-1 
                 6.5 
                 6 
                 5.5 
                 3.5 
                 3 
                 &lt;3 
               
               
                   
               
             
          
         
       
     
         [0029]    Conclusion: Ca(OH) 2  saturated solution is effective against Herpes Simplex Virus (HSV-1, &gt;3.5 log kill in 1 hr). 
         [0030]    B. Investigating the Biocidal Properties of Ca(OH) 2    
         [0031]    Experiments: After confirmation regarding the observed morphologies of  E. coli  and  Pseudomonas,  tests were conducted to investigate the biocidal properties of Ca(OH) 2 . An efficacy study against fungi was also completed. 
         [0032]    Results: 
         [0000]                                                    TABLE 2               Antimicrobial Susceptibility Testing Ca(OH) 2  paste (a complete study)                                    Zone of Inhibition (mm)                Paste                   Bacteria Organism   volume   16-18 hr   24 hr   48 hr                 Staphylococcus     0.2 ml   19, 19, 18   18, 17, 18   18, 17-18, 18         aureus  (# 6538)   0.3 ml   19, 18, 18-19   19, 18, 18   17, 18, 18         Salmonella enteric     0.2 ml   21, 21, 21   21, 20, 21   21, 20-21, 21       (# 6994)   0.3 ml   23, 21, 22   23, 22, 22   22, 21-22, 21         E. Coli  (#4157)   0.2 ml   20, 20, 19   20, 17, 18   17, 17, 17           0.3 ml   20, 20, 20   18, 20, 18   18, 19, 18         E. Coli  (# 12810)   0.2 ml   16, 16, 17   16, 16, 16   16, 16, 16           0.3 ml   17, 17, 17   17, 17, 16-17   17, 17, 16         Pseudomonas     0.2 ml   17, 17, 17   16, 17, 16   16, 16, 16         aeruginosa  (#13388)         Pseudomonas     0.3 ml   17, 17, 17   17, 17, 17   17, 16, 17         aeruginosa  (#13388)                   Paste       Fungi Organism   volume   24 hr   48 hr   72 hr                 Aspergillus     0.2 ml   —, ~35≠, —   30, 30, 28-29   22, 23, 21         niger (#9642)   0.3 ml   —, ~34≠, —   29, 30, 27-28   22, 23, 21         Auriobasidum     0.2 ml   27, 29, 29   23-24, 24-25, 23-24   22, 22, 20         pullulans (#15233)   0.3 ml   30, 30, 30   25-26, 26-27, 27   22, 22, 23         Chaetobium     0.2 ml   —, —, —   —, —, —   23, 22, 22         globosom (#6295)   0.3 ml   —, —, —   —, —, —   25, 24, —               “—” means patchy (partial) lawn because fungi grows much slower.       “≠” means very spotty growth, not yet a confluence lawn.            
Quality negative control: 200 μL of sterile water was added to 8 mm bore hole, all showed no sign of microbial inhibition (0 mm past 8 mm bore).
 
         [0033]    Antibacterial Activity Test 
         [0034]    A germicidal spray test was carried out to test the efficacy of a disinfectant formulation of the invention comprising a concentration of calcium hydroxide of 0.12% to 0.18% in water and 2.5%/w sodium hypochlorite. The efficacy was tested against  S. aureus.  The test was conducted on 30 sterile 18×36×1 mm glass slides as the carrier. No soil was added. The samples were incubated for 48±6 hours at an incubation temperature of 36.0±1° C. The samples were sprayed with 3 sprays of the disinfectant and left for a contact time of 5 minutes at ambient temperature. The neutralizer was 20 ml Modified Letheen Broth (w/0.1% sodium thoisulfate). 
         [0000]    
       
         
               
             
               
               
               
               
               
               
               
             
           
               
                 TABLE 3 
               
             
             
               
                   
               
               
                 Germicidal Spray Test Results 
               
             
          
           
               
                 Test 
                 Soil 
                 Contact 
                   
                 Average 
                 Carriers 
                 Carriers 
               
               
                 Substance 
                 load 
                 time 
                 Microorganism 
                 CFU/Carrier 
                 Positive 
                 negative 
               
               
                   
               
               
                 supernatant 
                 None 
                 5 minutes 
                 
                   S. Aureus 
                 
                 8.10E+05 
                 0 
                 30 
               
               
                 composition of 
                   
                   
                 ATCC 6538 
               
               
                 0.12-0.18% 
               
               
                 Ca(OH) 2  in 
               
               
                 water with 
               
               
                 2.5% sodium 
               
               
                 hypochlorite 
               
               
                   
               
             
          
         
       
     
         [0035]    Antiviral Activity Test 
         [0036]    An experiment was conducted to investigate the antiviral properties of a disinfecting formulation of the invention. A disinfecting formulation comprising 0.12-0.18% of calcium hydroxide and 0.125% sodium hypochlorite was mixed with 10 μL of virus stock. The sample was at a pH or 12.64 as measured and incubated for different time periods. The mixture was then diluted and the viable number of virus (Log TCID) was measured. 
         [0037]    Study Results 
         [0000]    
       
         
               
               
               
               
               
               
               
             
               
               
               
               
               
               
               
             
           
               
                   
               
               
                 Log 
                   
                   
                   
                   
                   
                   
               
               
                 TCID50/ 
               
               
                 100 μL 
                 0 min 
                 5 min 
                 10 min 
                 1 hr 
                 2 hr 
                 24 hr 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                 HSV-1 virus 
                 6.6 
                 &lt;3 
                 &lt;3 
                 &lt;3 
                 &lt;3 
                 &lt;3 
               
               
                 EMCV virus 
                 7 
                 5.3 
                 3.5 
                 3 
                 3 
                 &lt;3 
               
               
                   
               
             
          
         
       
     
         [0038]    The disinfectant formulation was found to be very effective against Herpes Simplex Virus (HSV-1), (&gt;3.5 log kill within 5 min.), and very effective (3.5 log kill within 10 min,) against Encephalomyocarditis (EMCV) virus.