Compositions for inactivating pathogenic microorganisms, methods of making the compositions, and methods of use thereof

Abstract

Nanoemulsion compositions with low toxicity that demonstrate broad spectrum inactivation of microorganisms or prevention of diseases are described. The nanoemulsions contain an aqueous phase, an oil phase comprising an oil and an organic solvent, and one or more surfactants. Methods of making nanoemulsions and inactivating pathogenic microorganisms are also provided.

1 . A composition comprising a nanoemulsion, the nanoemulsion comprising: 
 an aqueous phase;    an oil phase comprising an oil and an organic solvent; and    one or more surfactants;    wherein the nanoemulsion comprises nanoemulsion particles having an average diameter of from less than 150 nm    
     
     
         2 . The composition of  claim 1 , wherein the nanoemulsion particles have an average diameter of less than or equal to about 100 nm or less than or equal to about 50 nm.  
     
     
         3 . The composition of  claim 1 , wherein the organic solvent comprises a C 1 -C 12  alcohol, diol, or triol, a dialkyl phosphate, a trialkyl phosphate or a combination thereof.  
     
     
         4 . The composition of  claim 1 , wherein the alcohol comprises ethanol, isopropyl alcohol, glycerol or a combination thereof.  
     
     
         5 . The composition of  claim 2 , wherein the trialkyl phosphate is tri-n-butyl phosphate.  
     
     
         6 . The composition of  claim 1 , wherein the oil comprises soybean oil, mineral oil, avocado oil, squalene oil, olive oil, canola oil, corn oil, rapeseed oil, safflower oil, sunflower oil, fish oils, flavor oils, cinnamon bark, coconut oil, cottonseed oil, faxseed oil, pine needle oil, silicon oil, essential oils, water insoluble vitamins, or a combination thereof.  
     
     
         7 . The composition of  claim 6 , wherein the oil is soybean oil.  
     
     
         8 . The composition of  claim 1 , wherein the surfactant is a nonionic surfactant.  
     
     
         9 . The composition of  claim 8 , wherein the nonionic surfactant is TWEEN® 20, Triton® X-100, nonoxynol-9, or a combination thereof.  
     
     
         10 . The composition of  claim 1 , wherein the surfactant is a cationic surfactant.  
     
     
         11 . The composition of  claim 10 , wherein the cationic surfactant is cetyl pyridium chloride, benzalkonium chloride or a combination thereof.  
     
     
         12 . The composition of  claim 1  comprising: 
 about 5 vol. % to about 50 vol. % of aqueous phase;    about 30 vol. % to about 90 vol. % of oil phase; and    about 3 vol. % to about 15 vol. % of surfactant.    
     
     
         13 . The composition of  claim 1 , further comprising an additive selected from the group consisting of activity modulators, gelling agents, auxiliary surfactants, and a combination comprising one or more of the foregoing additives.  
     
     
         14 . The composition of  claim 13 , wherein the activity modulator is an interaction enhancer, a germination enhancer, a therapeutic agent, or a combination comprising one or more of the foregoing enhancers.  
     
     
         15 . The composition of  claim 14 , wherein the germination enhancer comprises glucose, fructose, asparagine, sodium chloride, ammonium chloride, calcium chloride, and potassium chloride.  
     
     
         16 . The composition of  claim 14 , wherein the germination enhancer comprises L-alanine, inosine, PBS, and ammonium chloride.  
     
     
         17 . The composition of  claim 14 , wherein the therapeutic agent is an antimicrobial agent, an antifungal agent, an antiviral agent, an anti-mold agent, an anti-mildew agent, or a combination thereof.  
     
     
         18 . The composition of  claim 14 , wherein the therapeutic agent is a penicillin, a cephalosporin, cycloserine, vancomycin, bacitracin, miconazole, ketoconazole, clotrimazole, polymyxin, colistimethate, nystatin, amphotericin B, chloramphenicol, the tetracyclines, erythromycin, clindamycin, an aminoglycoside, a rifamycin, a quinolone, trimethoprim, a sulfonamide, zidovudine, gangcyclovir, vidarabine, acyclovir, phenylphenol, propyl paraben, poly(hexamethylene biguanide) or a combination comprising one or more of the foregoing therapeutic agents.  
     
     
         19 . The composition of  claim 1 , wherein the composition comprises from about 0.01% to about 90% nanoemulsion per milliliter of composition.  
     
     
         20 . The composition of  claim 1 , wherein the composition comprises greater than about 0.25%, about 1.0%, about 5%, about 10%, about 20%, about 35%, about 50%, about 65%, about 80%, about 90%, or about 95% nanoemulsion per milliliter of composition.  
     
     
         21 . The composition of  claim 1 , further comprising a pharmaceutically acceptable carrier, an auxiliary surfactant, a suds suppressor, a detergent builder, or a combination thereof.  
     
     
         22 . A method of reducing the average nanoemulsion particle size of a composition comprising a nanoemulsion, comprising treating a nanoemulsion comprising an aqueous phase, an oil phase comprising an oil and an organic solvent, and a surfactant, and having nanoemulsion particles of an average diameter of greater than or equal to about 250 nm, so as to reduce the average diameter of the nanoemulsion particles to less than 150 nm.  
     
     
         23 . A method of making a nanoemulsion, comprising passing a first nanoemulsion comprising 
 an aqueous phase,    an oil phase comprising an oil and an organic solvent, and    one or more surfactants, wherein the nanoemulsion particles have an average diameter of greater than or equal to about 250 nm, through a microfluidizer or high pressure homogenizer under conditions effective to reduce the average diameter of the nanoemulsion particles to less than 150 nm.    
     
     
         24 . The method of  claim 22 , wherein the ratio of oil phase to aqueous phase is from about 1:9 to about 5:1, from about 5:1 to about 3:1, or from about 4:1.  
     
     
         25 . The method of  claim 23 , wherein the ratio of oil phase to aqueous phase is from about 1:9 to about 5:1, from about 5:1 to about 3:1, or from about 4:1.  
     
     
         26 . The method of claims  22 , wherein the nanoemulsion particles have an average diameter of less than or equal to about 100 nm or less than or equal to about 50 nm.  
     
     
         27 . The method of claims  23 , wherein the nanoemulsion particles have an average diameter of less than or equal to about 100 nm or less than or equal to about 50 nm.  
     
     
         28 . A method of inactivating a microorganism, comprising contacting the microorganism with a composition comprising a nanoemulsion for a time effective to inactivate the microorganism, wherein the nanoemulsion comprises: 
 an aqueous phase;    an oil phase comprising an oil and an organic solvent;    and one or more surfactants; and wherein the nanoemulsion particles have an average diameter of less than 150 nm.    
     
     
         29 . The method of claims  28 , wherein the nanoemulsion particles have an average diameter of less than or equal to about 100 nm or less than or equal to about 50 nm.  
     
     
         30 . The method of  claim 28 , wherein the composition further comprises an activity modulator, wherein the activity modulator is an interaction enhancer, a germination enhancer, a therapeutic agent, or a combination comprising one or more of the foregoing.  
     
     
         31 . The method of  claim 28 , wherein the composition further comprises a pharmaceutically acceptable carrier.  
     
     
         32 . The method of  claim 28 , wherein the microorganism is a bacteria, a fungus, a protozoa, a virus, or a combination of one or more of the foregoing microorganisms.  
     
     
         33 . The method of  claim 32 , wherein the bacteria is a vegetative bacteria, a bacterial spore, or a combination thereof.  
     
     
         34 . The method of  claim 32 , wherein the bacteria comprises a Gram negative bacteria, a Gram positive bacteria, an acid fast bacilli, or a combination thereof.  
     
     
         35 . The method of  claim 33 , wherein the bacterial spore is  B. anthracis.    
     
     
         36 . The method of  claim 32 , wherein the bacteria comprises comprises  B. anthracis, B. cereus, B. circulans, B. megatertium, B. subtilis, C. botulinum, C. tetani, C. perfringens, H. influenzae, N. gonorrhoeae, S. agalactiae, S. pneumonia, S. pyogenes, V. cholerae, S. aureus, Yersinia species, G. vaginalis, G. mobiluncus, M. hominis, Salmonellae  species,  Shigellae  species,  Pseudomonas  species,  Eschericia  species,  Klebsiella  species,  Proteus  species,  Enterobacter  species,  Serratia  species,  Moraxella  species,  Legionella  species,  Bordetella  species,  Helicobacter  species,  Arthobacter  species,  Micrococcus  species,  Listeria  species,  Corynebacteria  species,  Planococcus  species,  Nocardia  species,  Rhodococcus  species,  Mycobacteria  species, or a combination thereof.  
     
     
         37 . The method of  claim 32 , wherein the virus belongs to a family selected from the group consisting of Orthomyxoviridae, Retroviridae, African Swine Fever Viruses, Papovaviridae, Hepadnaviridae, Coronaviridae, Flaviviridae, Togaviridae, Picornaviridae, Filoviridae, Paramyxoviridae, or Rhabdoviridae.  
     
     
         38 . The method of  claim 37 , wherein the Orthomyxovirdae virus is influenza virus, herpes simplex, herpes zoster, sendai virus, sindbis virus, pox virus, small pox or vaccinia virus.  
     
     
         39 . The method of  claim 37 , wherein the Retroviridae is human immunodeficiency virus, west nile virus, hanta virus, or human papilloma virus.  
     
     
         40 . The method of  claim 32 , wherein the fungus is a yeast or a filamentous fungus.  
     
     
         41 . The method of  claim 40 , wherein filamentous fungus is selected from the group consisting of an  Aspergillus  species or a dermatophyte.  
     
     
         42 . The method of  claim 41 , wherein the dermatophyte is selected from the group consisting of  Trichophyton rubrum, Trichophyton mentagrophytes, Microsporum canis, Microsporum gypseum  and  Epidermophyton floccosum.    
     
     
         43 . The method of  claim 32 , wherein the fungus comprises  Cladosporium, Fusarium, Alternaria, Curvularia, Aspergillus Penicillium  or a combination thereof.  
     
     
         44 . A method of inactivating a pathogenic microorganism comprising contacting a subject infected with the microorganism with a composition comprising a nanoemulsion comprising: 
 an aqueous phase;    an oil phase comprising an oil and an organic solvent; and    one or more surfactants;    wherein the nanoemulsion comprises particles having an average diameter less than 150 nm.    
     
     
         45 . The method of  claim 44 , wherein the particles have an average diameter of less than or equal to about 100 nm or less than or equal to about 50 nm.  
     
     
         46 . The method of  claim 44 , wherein the subject is a human, an animal, or a plant.  
     
     
         47 . The method of  claim 44 , wherein the composition further comprises an activity modulator, wherein the activity modulator is an interaction enhancer, a germination enhancer, a therapeutic agent, or a combination comprising one or more of the foregoing.  
     
     
         48 . The method of  claim 44 , wherein the composition further comprises a pharmaceutically acceptable carrier.  
     
     
         49 . The method of  claim 44 , wherein the microorganism is a bacteria, a fungus, a filamentous fungus, a protozoa, a virus, or a combination of one or more of the foregoing microorganisms.  
     
     
         50 . The method of  claim 49 , wherein the bacteria is a vegetative bacteria, a bacterial spore, or a combination thereof.  
     
     
         51 . The method of  claim 49 , wherein the fungus is a yeast.  
     
     
         52 . The method of  claim 49 , wherein the bacteria comprises a Gram negative bacteria, a Gram positive bacteria, an acid fast bacilli, or a combination thereof.  
     
     
         53 . The method of  claim 50 , wherein the bacterial spore is  B. anthracis.    
     
     
         54 . The method of  claim 49 , wherein the bacteria is  B. anthracis, B. cereus, B. circulans, B. megatertium, B. subtilis, C. botulinum, C. tetani, C. perfringens, H. influenzae, N. gonorrhoeae, S. agalactiae, S. pneumonia, S. pyogenes, V. cholerae, S. aureus, Yersinia  species,  G. vaginalis, G. mobiluncus, M. hominis, Salmonellae  species,  Shigellae  species,  Pseudomonas  species,  Eschericia  species,  Klebsiella  species,  Proteus  species,  Enterobacter  species,  Serratia  species,  Moraxella  species,  Legionella  species,  Bordetella  species,  Helicobacter  species,  Arthobacter  species,  Micrococcus  species,  Listeria  species,  Corynebacteria  species,  Planococcus  species,  Nocardia  species,  Rhodococcus  species,  Mycobacteria  species, or a combination thereof.  
     
     
         55 . The method of  claim 49 , wherein the virus belongs to a family selected from the group consisting of Orthomyxoviridae, Retroviridae, African Swine Fever Viruses, Papovaviridae, Hepadnaviridae, Coronaviridae, Flaviviridae, Togaviridae, Picomaviridae, Filoviridae, Paramyxoviridae, or Rhabdoviridae.  
     
     
         56 . The method of  claim 55 , herein the Orthomyxovirdae virus is influenza virus, herpes simplex, herpes zoster, sendai virus, sindbis virus, pox virus, small pox or vaccinia virus.  
     
     
         57 . The method of  claim 55 , herein the Retroviridae is human immunodeficiency virus, west nile virus, hanta virus, or human papilloma virus.  
     
     
         58 . The method of  claim 49 , wherein the filamentous fungus comprises  Aspergillus  species or dermatophytes such as  Trichophyton rubrum, Trichophyton mentagrophytes, Microsporum canis, Microsporum gypseum  or  Epidermophyton floccosum.    
     
     
         59 . The method of  claim 49  wherein the fungus is selected from the group consisting of  Cladosporium, Fusarium, Alternaria, Curvularia, Aspergillus  and  Penicillium.    
     
     
         60 . A method of preventing an infected state caused by a microorganism, comprising administering to a subject, either before or after exposure to a microorganism, a composition comprising a nanoemulsion, wherein the nanoemulsion comprises: 
 an aqueous phase;    an oil phase comprising an oil and an organic solvent; and one or more surfactants;    wherein the nanoemulsion comprises particles having an average diameter less than 150 nm.    
     
     
         61 . The method of  claim 60 , wherein the infected state is a sexually transmitted genital infection.  
     
     
         62 . The method of  claim 61 , wherein the sexually transmitted genital infection is selected from the group consisting of genital herpes, human papilloma virus, human immunodeficiency virus, trichomoniasis, gonorrhea, syphilis, and Chlamydia.  
     
     
         63 . The method of  claim 60 , wherein the microorganism is selected from the group consisting of a pox virus,  B. anthracis,  and  Yersinia  species.  
     
     
         64 . The method of  claim 60 , wherein the step of administering comprises application of the composition to the mucosa of a subject.  
     
     
         65 . An immunogenic composition comprising a nanoemulsion, wherein the nanoemulsion comprises: 
 an aqueous phase;    an oil phase comprising an oil and an organic solvent; and a surfactant;    and wherein the nanoemulsion comprises nanoemulsion particles having an average diameter of less than or equal to 250 nm and a microorganism or a portion thereof.    
     
     
         66 . The immunogenic composition of  claim 65 , wherein the particles have an average diameter of less than or equal to about 200 nm, less than or equal to about 150 nm, less than or equal to about 100 nm, or less than or equal to about 50 nm.  
     
     
         67 . The immunogenic composition of  claim 65 , wherein the composition further comprises a pharmaceutically acceptable carrier.  
     
     
         68 . The immunogenic composition of  claim 65 , wherein the microorganism or portion thereof comprises at least a portion of a bacteria, a fungus, a protozoa, a virus, or a combination of one or more of the foregoing microorganisms.  
     
     
         69 . The immunogenic composition of  claim 68 , wherein the bacteria is a vegetative bacteria, a bacterial spore, or a combination thereof.  
     
     
         70 . The immunogenic composition of  claim 68 , wherein the bacteria is a Gram negative bacteria, a Gram positive bacteria, an acid fast bacilli or a combination thereof.  
     
     
         71 . The immunogenic composition of  claim 69 , wherein the bacterial spore is  B. anthracis.    
     
     
         72 . The immunogenic composition of  claim 68 , wherein the bacteria is a respiratory transmitted bacteria.  
     
     
         73 . The immunogenic composition of  claim 68 , wherein the virus is selected from the group consisting of herpes virus, sendai virus, sindbis virus, vaccinia virus, human immunodeficiency virus, west nile virus, hanta virus, human papilloma virus, vaccinia virus, SARS, respiratory transmitted viruses and combinations thereof.  
     
     
         74 . The immunogenic composition of  claim 65 , wherein the microbial antigen is mixed with the nanoemulsion alone or with an adjuvant.  
     
     
         75 . The immunogenic composition of  claim 74 , wherein the adjuvant is a CpG oligonucleotide.  
     
     
         76 . The immunogenic composition of  claim 65 , wherein administration of the immunogenic composition to a subject raises antibodies in the subject capable of neutralizing or inactivating the microorganism.  
     
     
         77 . The immunogenic composition of  claim 65 , wherein the microorganism or portion thereof is a PA of  B. anthracis.    
     
     
         78 . A method of vaccinating against a microorganism comprising administering to a subject the a composition comprising a nanoemulsion, wherein the nanoemulsion comprises: 
 an aqueous phase;    an oil phase comprising an oil and an organic solvent; and    one or more surfactants;    wherein the nanoemulsion comprises particles having an average diameter of less than 250 nm,    wherein the microorganism is inactivated by the composition and an immunological response by the subject is elicited.    
     
     
         79 . A kit comprising a composition comprising the nanoemulsion of  claim 1 , wherein the composition is provided in a single formulation or a binary formulation, wherein the binary formulation is mixed prior to using the composition.  
     
     
         80 . The kit of  claim 79 , further comprising instructions for using the composition.  
     
     
         81 . The kit of  claim 79 , wherein the composition is formulated for topical administration, said kit further including means for applying the composition topically.