# Patent

## Title
Microbicidal composition

## Abstract

A synergistic microbicidal composition having two components. The first component is a nonionic surfactant with structure: R2O(CH2CH(CH3)O)3(CH2CH2O)5H, where R2 is a mixture of C8-C14 linear alkyl groups. The second component is 2-bromo-2-nitropropane-1,3-diol. The weight ratio of the nonionic surfactant to 2-bromo-2-nitropropane-1,3-diol is from 1:0.08 to 1:0.12 or 1:0.2 to 1:1.8286.

## Background
This invention relates to microbicidal compositions containing 2-bromo-2-nitropropane-1,3-diol (BNPD), tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione (DAZOMET) or BIOBAN P-1487 (ca. 81:5 mixture of 4-(2-nitrobutyl)morpholine and 4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine, respectively) and a surfactant.
A composition containing 5-chloro-2-methylisothiazolin-3-one, 2-methylisothiazolin-3-one and a nonionic dispersant is disclosed in U.S. Pat. No. 4,295,932. The composition contains a 3:1 mixture of 5-chloro-2-methylisothiazolin-3-one and 2-methylisothiazolin-3-one, and a copolymer of ethylene oxide and propylene oxide which appears to have the same composition as PLURONIC L61 or TERGITOL L61 dispersant. However, there is a need for combinations of microbicides having synergistic activity against various strains of microorganisms to provide effective control of the microorganisms. Moreover, there is a need for such combinations containing lower levels of individual microbicides for environmental and economic benefit. The problem addressed by this invention is to provide such synergistic combinations of microbicides.
STATEMENT OF THE INVENTION
The present invention is directed to a synergistic microbicidal composition comprising: (a) a nonionic surfactant with structure:

R2O(CH2CH(CH3)O)3(CH2CH2O)5H

where R2 is a mixture of C8-C14 linear alkyl groups; and (b) 2-bromo-2-nitropropane-1,3-diol; wherein a weight ratio of said nonionic surfactant to 2-bromo-2-nitropropane-1,3-diol is from 1:0.08 to 1:0.12 or 1:0.2 to 1:1.8286.

The present invention is further directed to a synergistic microbicidal composition comprising: (a) a nonionic surfactant with structure:

R2O(CH2CH(CH3)O)3(CH2CH2O)5H

where R2 is a mixture of C8-C14 linear alkyl groups; and (b) tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione; wherein a weight ratio of said nonionic surfactant to tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione is from 1:0.0024 to 1:0.4571.

The present invention is further directed to a synergistic microbicidal composition comprising: (a) a nonionic surfactant with structure:

R1O(CH2CH(CH3)O)5(CH2CH2O)9H

where R1 is a C8 alkyl group; and (b) a mixture of 4-(2-nitrobutyl)morpholine and 4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine in a weight ratio of 81:5; wherein a weight ratio of said nonionic surfactant to said mixture is from 1:0.0006 to 1:0.2743.

The present invention is further directed to methods for inhibiting the growth of microorganisms in aqueous media by adding to an aqueous medium a nonionic surfactant as described herein and 2-bromo-2-nitropropane-1,3-diol, tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione or a ca. 81:5 mixture of 4-(2-nitrobutyl)morpholine and 4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine in the ratios described herein.


DETAILED DESCRIPTION OF THE INVENTION
“BNPD” is 2-bromo-2-nitropropane-1,3-diol, “DAZOMET” has as an active ingredient tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione (CAS No. 533-74-4) and “BIOBAN P-1487” is an approximately 81:5 (weight) mixture of 4-(2-nitrobutyl)morpholine and 4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine. As used herein, the following terms have the designated definitions, unless the context clearly indicates otherwise. The term “microbicide” refers to a compound capable of inhibiting the growth of or controlling the growth of microorganisms; microbicides include bactericides, fungicides and algaecides. The term “microorganism” includes, for example, fungi (such as yeast and mold), bacteria and algae. The following abbreviations are used throughout the specification: ppm=parts per million by weight (weight/weight), mL=milliliter. Unless otherwise specified, temperatures are in degrees centigrade (° C.), references to percentages are percentages by weight (wt %) and amounts and ratios are on an active ingredient basis, i.e., total weight of BNPD, DAZOMET or BIOBAN P-1487 active ingredients and the nonionic surfactant. Numbers of polymerized units of propylene oxide or ethylene oxide are number averages.
Preferably, the weight ratio of the nonionic surfactant with structure:

R2O(CH2CH(CH3)O)3(CH2CH2O)5H

where R2 is a mixture of C8-C14 linear alkyl groups to BNPD is from 1:0.2286 to 1:1.18286.

The present invention is further directed to a method for inhibiting the growth of mold, preferably C. albicans, in an aqueous medium by adding: (a) a nonionic surfactant with structure:

R2O(CH2CH(CH3)O)3(CH2CH2O)5H

where R2 is a mixture of C8-C14 linear alkyl groups; and (b) BNPD; wherein a weight ratio of said nonionic surfactant to BNPD is from 1:0.08 to 1:0.12 or 1:0.2 to 1:1.8286, preferably from 1:0.2286 to 1:1.18286.

Preferably, the weight ratio of the nonionic surfactant with structure:

R2O(CH2CH(CH3)O)3(CH2CH2O)5H

where R2 is a mixture of C8-C14 linear alkyl groups to DAZOMET is from 1:0.0114 to 1:0.4571.

The present invention is further directed to a method for inhibiting the growth of S. aureus, in an aqueous medium by adding: (a) a nonionic surfactant with structure:

R2O(CH2CH(CH3)O)3(CH2CH2O)5H

where R2 is a mixture of C8-C14 linear alkyl groups; and (b) DAZOMET; wherein a weight ratio of said nonionic surfactant to DAZOMET is from 1:0.0114 to 1:0.4571.

Preferably, the weight ratio of the nonionic surfactant with structure:

R1O(CH2CH(CH3)O)5(CH2CH2O)9H

where R1 is a C8 alkyl group to BIOBAN P-1487 is from 1:0.0160 to 1:0.2743.

The present invention is further directed to a method for inhibiting the growth of fungi, preferably A. niger, in an aqueous medium by adding: (a) a nonionic surfactant with structure:

R1O(CH2CH(CH3)O)5(CH2CH2O)9H

where R1 is a C8 alkyl group to BIOBAN P-1487 is from 1:0.0160 to 1:0.2743.

R2 is a mixture of C8-C14 linear alkyl groups. Preferably, the C8-C14 linear alkyl groups comprise from 50 to 85 wt % C8-C10 linear alkyl groups and 15 to 50 wt % C12-C14 linear alkyl groups, preferably from 60 to 75 wt % C8-C10 linear alkyl groups and 25 to 40 wt % C12-C14 linear alkyl groups, preferably about 70 wt % C8-C10 linear alkyl groups and about 30 wt % C12-C14 linear alkyl groups. Preferably, the linear alkyl groups are derived from seed oil. Preferably, R1 is 2-ethylhexyl.
Preferably, each of the compositions is substantially free of microbicides other than the nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487, i.e., it has less than 1 wt % of microbicides other than the nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487 based on total weight of active ingredients, preferably less than 0.5 wt %, preferably less than 0.2 wt %, preferably less than 0.1 wt %. Preferably, when the nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487 are added to an aqueous medium, the medium is substantially free of other microbicides, i.e., it has less than 1 wt % of microbicides other than the nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487 based on total weight of active ingredients, preferably less than 0.5 wt %, preferably less than 0.2 wt %, preferably less than 0.1 wt %.
The compositions of this invention may contain other ingredients, e.g., defoamers and emulsifiers. The microbicidal compositions of the present invention can be used to inhibit the growth of microorganisms or higher forms of aquatic life (such as protozoans, invertebrates, bryozoans, dinoflagellates, crustaceans, mollusks, etc) by introducing a microbicidally effective amount of the compositions into an aqueous medium subject to microbial attack. Suitable aqueous media are found in, for example: industrial process water; electrocoat deposition systems; cooling towers; air washers; gas scrubbers; mineral slurries; wastewater treatment; ornamental fountains; reverse osmosis filtration; ultrafiltration; ballast water; evaporative condensers; heat exchangers; pulp and paper processing fluids and additives; starch; plastics; emulsions; dispersions; paints; latices; coatings, such as varnishes; construction products, such as mastics, caulks, and sealants; construction adhesives, such as ceramic adhesives, carpet backing adhesives, and laminating adhesives; industrial or consumer adhesives; photographic chemicals; printing fluids; household products, such as bathroom and kitchen cleaners; cosmetics; toiletries; shampoos; soaps; personal care products such as wipes, lotions, sunscreen, conditioners, creams, and other leave-on applications; detergents; industrial cleaners; floor polishes; laundry rinse water; metalworking fluids; conveyor lubricants; hydraulic fluids; leather and leather products; textiles; textile products; wood and wood products, such as plywood, chipboard, flakeboard, laminated beams, oriented strandboard, hardboard, and particleboard; petroleum processing fluids; fuel; oilfield fluids, such as injection water, fracture fluids, and drilling muds; agriculture adjuvant preservation; surfactant preservation; medical devices; diagnostic reagent preservation; food preservation, such as plastic or paper food wrap; food, beverage, and industrial process pasteurizers; toilet bowls; recreational water; pools; and spas.
The specific amount of the microbicidal compositions of this invention necessary to inhibit or control the growth of microorganisms in an application will vary. Amounts of the compositions of the present invention are given on the basis of active ingredients (i.e., nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487). For BNPD/surfactant used in paper applications, preferably the concentration is 2 to 50 ppm, preferably 5 to 40 ppm. For BNPD/surfactant used in oil and gas drilling applications, preferably the concentration is 10 to 500 ppm, preferably 20 to 350 ppm. For BNPD/surfactant used in metalworking fluids, preferably the concentration is 150 to 2000 ppm, preferably 300 to 1500 ppm. For BIOBAN P-1487/surfactant used in metalworking fluids, preferably the concentration is 600 to 1800 ppm, preferably 800 to 1500 ppm. For BIOBAN P-1487/surfactant used in fuel applications, preferably the concentration is 300 to 4000 ppm, preferably 500 to 3000 ppm. Fuel applications include the preservation of hydrocarbons for fuel, e.g., bioethanol applications, diesel storage tank preservation and general fuel tank preservation. In a method of this invention, a composition is treated to inhibit microbial growth by adding, together or separately, the nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487, in amounts that would produce the concentrations indicated above.
EXAMPLES
Surfactants and biocides were evaluated for synergy by determining the synergy index (S.I.) of the combination. Synergy index was calculated based on minimum inhibitory concentrations (MIC) of two antimicrobial compounds (A and B) alone and in combinations. The tests organisms were Gram negative bacteria (Pseudomonas aeruginosa ATCC #15442), Gram positive bacteria (Staphylococcus aureus ATCC #6538), yeast (Candida albicans ATCC #10203) and mold (Aspergillus niger ATCC #16404). Contact time for the bacteria was 24 and 48 hours, yeast was 48 and 72 hrs, and 3 and 7 days for mold. The test was carried out in 96 well microtiter plates.

    Surf. A R1O(CH2CH(CH3)O)5(CH2CH2O)9H, where R1 is 2-ethylhexyl
    Surf. D R2O(CH2CH(CH3)O)3(CH2CH2O)5H
    Surf. E R2O(CH2CH(CH3)O)3(CH2CH2O)7H

In Surf. D and Surf. E, R2 is a mixture of C8-C14 linear alkyl groups (70% C8-C10 linear alkyl and 30% C12-C14 linear alkyl)

Inoculums Used













Inoculum Size of organisms (CFU/ml)












Staphylococcus
Pseudomonas
Aspergillus
Candida



aureus
aeruginosa
niger
albicans



ATCC#
ATCC#
ATCC#
ATCC#


Surfactants
6538
15442
16404
10203





Surf. A
1.156E+06
8.134E+07
1.156E+06
 1.156+06


Surf. D
1.808E+05
1.156E+08
1.156E+06
5.726E+05


Surf. E
1.808E+06
5.727E+07
5.726E+05
1.808E+06
















TABLE 4







Media Used


Media Used for testing










Staphylococcus
Pseudomonas
Aspergillus
Candida


aureus
aeruginosa
niger
albicans


ATCC#
ATCC#
ATCC# 
ATCC# 


6538
15442
16404
10203





10% Tryptic soy
10% Tryptic soy
Potato dextrose
Potato dextrose


broth
broth
broth
broth









The pH of the Triptic soy broth was 7.3 and the Potato dextrose broth was 5.1.
The test results for demonstration of synergy of the MIC combinations are shown in the tables below. Each table shows the results for combinations of two components against the microorganisms tested with incubation times; the end-point activity in ppm measured by the MIC for compound A alone (CA), for component B alone (CB), and the mixture (Ca) and (Cb); the calculated SI value; and the range of synergistic ratios for each combination tested. SI is calculated as follows:

Ca/CA+Cb/CB=Synergy Index(“SI”)

Wherein:

    
    
        CA=concentration of compound A in ppm, acting alone, which produced an end point (MIC of Compound A).
        Ca=concentration of compound A in ppm, in the mixture, which produced an end point.
        CB=concentration of compound B in ppm, acting alone, which produced an end point (MIC of Compound B).
        Cb=concentration of compound B in ppm, in the mixture, which produced an end point.

When the sum of Ca/CA and Cb/CB is greater than one, antagonism is indicated. When the sum is equal to one, additivity is indicated, and when less than one, synergism is demonstrated.

    
    


The ratio ranges at which BNPD, DAZOMET or BIOBAN P-1487 and the surfactants were tested are as summarized in the following tables:










BNPD with Surf. A











Organism
ATCC #
From
To
Ratio Range














Staphylococcus
6538
0.02:20,000
 100:218.75
1:0.000001-


aureus



1:0.45714


Aspergillus
16404
 0.2:20,000
1,000:218.75
1:0.00001-


niger



1:4.5714


Candida
10203
 0.2:20,000
1,000:218.75
1:0.00001-


albicans



1:4.5714


Pseudomonas
15442
0.02:20,000
 100:218.75
1:0.000001-


aeruginosa



1:0.45714



















BNPD with Surf. E











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.02:20,000
 100:218.75
1:0.000001-


aureus



1:0.45714


Aspergillus
16404
 0.2:20,000
1,000:218.75
1:0.00001-


niger



1:4.5714


Candida
10203
 0.2:20,000
1,000:218.75
1:0.00001-


albicans



1:4.5714


Pseudomonas
15442
0.02:20,000
 100:218.75
1:0.000001-


aeruginosa



1:0.45714



















BNPD with Surf. D











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.02:20,000
 100:218.75
1:0.000001-


aureus



1:0.45714


Aspergillus
16404
 0.2:20,000
1,000:218.75
1:0.00001-


niger



1:4.5714


Candida
10203
 0.2:20,000
1,000:218.75
1:0.00001-


albicans



1:4.5714


Pseudomonas
15442
0.02:20,000
 100:218.75
1:0.000001-


aeruginosa



1:0.45714



















BIOBAN P-1487 with Surf. A











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.002:20,000 
 10:218.75
1:0.0000001-


aureus



1:0.0457143


Aspergillus
16404
0.02:20,000
100:218.75
1:0.000001-


niger



1:0.45714


Candida
10203
0.02:20,000
100:218.75
1:0.000001-


albicans



1:0.45714


Pseudomonas
15442
0.02:20,000
100:218.75
1:0.000001-


aeruginosa



1:0.45714



















BIOBAN P-1487 with Surf. E











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.02:20,000
100:218.75
1:0.000001-


aureus



1:0.45714


Aspergillus
16404
0.02:20,000
100:218.75
1:0.000001-


niger



1:0.45714


Candida
10203
0.02:20,000
100:218.75
1:0.000001-


albicans



1:0.45714


Pseudomonas
15442
0.02:20,000
100:218.75
1:0.000001-


aeruginosa



1:0.45714



















BIOBAN P-1487 with Surf. D











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.02:20,000
 100:218.75
1:0.000001-


aureus



1:0.45714


Aspergillus
16404
 0.2:20,000
1,000:218.75
1:0.00001-


niger



1:4.5714


Candida
10203
0.02:20,000
 100:218.75
1:0.000001-


albicans



1:0.45714


Pseudomonas
15442
 0.2:20,000
1,000:218.75
1:0.00001-


aeruginosa



1:4.5714



















DAZOMET with Surf. A











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.2:20,000
1,000:218.75
1:0.00001-


aureus



1:4.5714


Aspergillus
16404
0.2:20,000
1,000:218.75
1:0.00001-


niger



1:4.5714


Candida
10203
0.2:20,000
1,000:218.75
1:0.00001-


albicans



1:4.5714


Pseudomonas
15442
0.2:20,000
1,000:218.75
1:0.00001-


aeruginosa



1:4.5714



















DAZOMET with Surf. E











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.2:20,000
1,000:218.75
1:0.00001-


aureus



1:4.5714


Aspergillus
16404
0.2:20,000
1,000:218.75
1:0.00001-


niger



1:4.5714


Candida
10203
0.2:20,000
1,000:218.75
1:0.00001-


albicans



1:4.5714


Pseudomonas
15442
0.2:20,000
1,000:218.75
1:0.00001-


aeruginosa



1:4.5714



















DAZOMET with Surf. D











Organism
ATCC#
From
To
Ratio Range














Staphylococcus
6538
0.2:20,000
1,000:218.75
1:0.00001-


aureus



1:4.5714


Aspergillus
16404
0.2:20,000
1,000:218.75
1:0.00001-


niger



1:4.5714


Candida
10203
0.2:20,000
1,000:218.75
1:0.00001-


albicans



1:4.5714


Pseudomonas
15442
0.2:20,000
1,000:218.75
1:0.00001-


aeruginosa



1:4.5714









    A: Surf. A
    B: BNPD
    Media: 1/10 TSB
    Inoculum size: 8.134E+07 CFU/ml















PPM AI MIC




Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















Ps. aeruginosa
>20000
10
20000
8
<1.80
1:0.0004


ATCC# 15442
>20000
10
10000
8
<1.30
1:0.0008



>20000
10
5000
8
<1.05
1:0.0016



>20000
10
2500
8
<0.93
1:0.0032



>20000
10
1750
8
<0.89
1:0.0046



>20000
10
875
8
<0.84
1:0.0091



>20000
10
437.5
8
<0.82
1:0.0183



>20000
10
218.75
8
<0.81
1:0.0366









No Synergy: A. niger, C. albicans, S. aureus 

    A: Surf. D
    B: BNPD
    Media: PDB
    Inoculum size: 1.156E+06















PPM AI MIC




Values (3rd day)












Test
Alone
Combination

Ratio













Organism
CA
CB
Ca
Cb
S.I.
(Ca:Cb)
















A. niger
>20000
1000
20000
500
<1.50
1:0.0250


ATCC# 16404
>20000
1000
10000
400
<0.90
1:0.0400



>20000
1000
10000
500
<1.00
1:0.0500



>20000
1000
5000
500
<0.75
1:0.1000



>20000
1000
5000
600
<0.85
1:0.1200



>20000
1000
5000
800
<1.05
1:0.1600



>20000
1000
2500
500
<0.63
1:0.2000



>20000
1000
2500
600
<0.73
1:0.2400



>20000
1000
2500
800
<0.93
1:0.3200



>20000
1000
1750
600
<0.69
1:0.3429



>20000
1000
1750
800
<0.89
1:0.4571



>20000
1000
875
600
<0.64
1:0.6857



>20000
1000
875
800
<0.84
1:0.9143









    A: Surf. D
    B: BNPD
    Media: PDB
    Inoculum size: 5.726E+05 CFU/ml















PPM AI MIC




Values (48 hrs)












Test
Alone
Combination

Ratio













Organism
CA
CB
Ca
Cb
S.I.
(Ca:Cb)
















C. albicans
>20000
1000
20000
400
<1.40
1:0.0200


ATCC# 10203
>20000
1000
10000
400
<0.90
1:0.0400



>20000
1000
10000
500
<1.00
1:0.0500



>20000
1000
5000
400
<0.65
1:0.0800



>20000
1000
5000
500
<0.75
1:0.1000



>20000
1000
5000
600
<0.85
1:0.1200



>20000
1000
5000
800
<1.05
1:0.1600



>20000
1000
2500
400
<0.53
1:0.1600



>20000
1000
2500
600
<0.73
1:0.2400



>20000
1000
2500
800
<0.93
1:0.3200



>20000
1000
1750
400
<0.49
1:0.2286



>20000
1000
1750
600
<0.69
1:0.3429



>20000
1000
1750
800
<0.89
1:0.4571



>20000
1000
875
500
<0.54
1:0.5714



>20000
1000
875
600
<0.64
1:0.6857



>20000
1000
875
800
<0.84
1:0.9143



>20000
1000
437.5
800
<0.82
1:1.8286









    A: Surf. D
    B: BNPD
    Media: 1/10 TSB
    Inoculum size: 1.16E+08















PPM AI MIC




Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















Ps. aeruginosa
>20000
10
875
8
<0.84
1:0.0091


ATCC# 15442
>20000
10
437.5
8
<0.82
1:0.0183









    A: Surf. D
    B: BNPD
    Media: 1/10 TSB
    Inoculum size: 1.808E+05















PPM AI MIC




Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















S. aureus
>20000
20
20000
4
<1.20
1:0.0002


ATCC# 6538
>20000
20
10000
3
<0.65
1:0.0003



>20000
20
10000
6
<0.80
1:0.0006



>20000
20
10000
8
<0.90
1:0.0008



>20000
20
10000
10
<1.00
1:0.0010



>20000
20
5000
2
<0.35
1:0.0004



>20000
20
5000
4
<0.45
1:0.0008



>20000
20
5000
8
<0.65
1:0.0016



>20000
20
5000
10
<0.75
1:0.0020



>20000
20
5000
2
<0.35
1:0.0004



>20000
20
5000
6
<0.55
1:0.0012



>20000
20
5000
10
<0.75
1:0.0020



>20000
20
1750
2
<0.19
1:0.0011



>20000
20
1750
8
<0.49
1:0.0046



>20000
20
1750
10
<0.59
1:0.0057



>20000
20
875
2
<0.14
1:0.0023



>20000
20
875
4
<0.24
1:0.0046



>20000
20
875
6
<0.34
1:0.0069



>20000
20
875
10
<0.54
1:0.0114



>20000
20
437.5
6
<0.32
1:0.0137



>20000
20
437.5
8
<0.42
1:0.0183



>20000
20
437.5
10
<0.52
1:0.0229



>20000
20
218.75
8
<0.41
1:0.0366



>20000
20
218.75
10
<0.51
1:0.0457









    A: Surf. E
    B: BNPD
    Media: PDB
    Inoculum size: 1.808E+06 CFU/ml















PPM AI MIC




Values (48 hrs)












Test
Alone
Combination

Ratio













Organism
CA
CB
Ca
Cb
S.I.
(Ca:Cb)
















C. albicans
>20000
1000
20000
200
<1.20
1:0.0100


ATCC# 10203
>20000
1000
10000
200
<0.70
1:0.0200



>20000
1000
10000
300
<0.80
1:0.0300



>20000
1000
10000
500
<1.00
1:0.0500



>20000
1000
5000
200
<0.45
1:0.0400



>20000
1000
5000
400
<0.65
1:0.0800



>20000
1000
5000
600
<0.85
1:0.1200



>20000
1000
5000
800
<1.05
1:0.1600



>20000
1000
2500
400
<0.53
1:0.1600



>20000
1000
2500
600
<0.73
1:0.2400



>20000
1000
2500
800
<0.93
1:0.3200



>20000
1000
1750
500
<0.59
1:0.2857



>20000
1000
1750
800
<0.89
1:0.4571



>20000
1000
875
800
<0.84
1:0.9143









    A: Surf. E
    B: BNPD
    Media: 1/10 TSB
    Inoculum size: 5.727E+07 CFU/ml















PPM AI MIC




Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















Ps. aeruginosa
>20000
10
5000
8
<1.05
1:0.0016


ATCC# 15442
>20000
10
2500
8
<0.93
1:0.0032



>20000
10
1750
8
<0.89
1:0.0046



>20000
10
875
8
<0.84
1:0.0091



>20000
10
437.5
8
<0.82
1:0.0183



>20000
10
218.75
8
<0.81
1:0.0366









    A: Surf. E
    B: BNPD
    Media: 1/10 TSB
    Inoculum size: 1.808E+06 CFU/ml















PPM AI MIC




Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















S. aureus
>20000
20
20000
8
<1.40
1:0.0004


ATCC#
>20000
20
10000
8
<0.90
1:0.0008


6538
>20000
20
5000
8
<0.65
1:0.0016



>20000
20
5000
10
<0.75
1:0.0020



>20000
20
2500
8
<0.53
1:0.0032



>20000
20
2500
10
<0.63
1:0.0040



>20000
20
1750
8
<0.49
1:0.0046



>20000
20
1750
10
<0.59
1:0.0057



>20000
20
875
8
<0.44
1:0.0091



>20000
20
875
10
<0.54
1:0.0114









    No synergy: A. niger 
    A: Surf. A
    B: DAZOMET
    Media: 1/10 TSB
    Inoculum size: 8.13E+07
    S. aureus ATCC 6538 No Synergy
    A: Surf. A
    B: DAZOMET
    Media: PDB
    Inoculum size: 1.156E+06















PPM AI MIC




Values (3rd day)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)

















>20000
60
10000
0.6
<0.51
1:0.0001


A. niger
>20000
60
10000
0.8
<0.51
1:0.0001


ATCC# 16404
>20000
60
10000
1
<0.52
1:0.0002



>20000
60
10000
2
<0.53
1:0.0003



>20000
60
10000
3
<0.55
1:0.0004



>20000
60
10000
4
<0.57
1:0.0005



>20000
60
10000
5
<0.58
1:0.0006



>20000
60
10000
6
<0.60
1:0.0008



>20000
60
10000
8
<0.63
1:0.0010



>20000
60
10000
10
<0.67
1:0.0020



>20000
60
10000
20
<0.83
1:0.0030



>20000
60
10000
30
<1.00
1:0.0040



>20000
60
10000
40
<1.17
1:0.0050



>20000
60
10000
50
<1.33
1:0.0008



>20000
60
5000
4
<0.32
1:0.0016



>20000
60
5000
8
<0.38
1:0.0020



>20000
60
5000
10
<0.42
1:0.0040



>20000
60
5000
20
<0.58
1:0.0060



>20000
60
5000
30
<0.75
1:0.0080



>20000
60
5000
40
<0.92
1:0.0100



>20000
60
5000
50
<1.08
1:0.0020



>20000
60
2500
5
<0.21
1:0.0024



>20000
60
2500
6
<0.23
1:0.0032



>20000
60
2500
8
<0.26
1:0.0040



>20000
60
2500
10
<0.29
1:0.0080



>20000
60
2500
20
<0.46
1:0.0120



>20000
60
2500
30
<0.63
1:0.0160



>20000
60
2500
40
<0.79
1:0.0200



>20000
60
2500
50
<0.96
1:0.0029



>20000
60
1750
5
<0.17
1:0.0020



>20000
60
1750
6
<0.23
1:0.0024



>20000
60
1750
8
<0.26
1:0.0032



>20000
60
1750
10
<0.29
1:0.0040



>20000
60
1750
20
<0.46
1:0.0080



>20000
60
1750
30
<0.63
1:0.0120



>20000
60
1750
40
<0.79
1:0.0160



>20000
60
1750
50
<0.96
1:0.0200



>20000
60
875
20
<0.17
1:0.0029



>20000
60
875
30
<0.19
1:0.0034



>20000
60
875
40
<0.22
1:0.0046



>20000
60
875
50
<0.25
1:0.0057



>20000
60
437.5
30
<0.42
1:0.0114



>20000
60
437.5
40
<0.59
1:0.0171



>20000
60
437.5
50
<0.75
1:0.0229



>20000
60
218.75
40
<0.92
1:0.0286



>20000
60
218.75
50
<0.38
1:0.0229









    A: Surf. A
    B: DAZOMET
    Media: PDB
    Inoculum size: 1.156E+06















PPM AI MIC




Values (48 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















C. albicans
>20000
50
2500
40
<0.93
1:0.0160


ATCC# 10203
>20000
50
1750
40
<0.89
1:0.0229



>20000
50
437.5
40
<0.82
1:0.0914



>20000
50
218.75
40
<0.81
1:0.1829









    A: Surf. A
    B: DAZOMET
    Media: 1/10 TSB
    Inoculum size: 8.134E+07















PPM AI MIC




Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















Ps. aeruginosa
>20000
80
2500
4
<0.18
1:0.0016


ATCC# 15442
>20000
80
2500
5
<0.19
1:0.0020



>20000
80
2500
6
<0.20
1:0.0024



>20000
80
2500
8
<0.23
1:0.0032



>20000
80
2500
10
<0.25
1:0.0040



>20000
80
2500
20
<0.38
1:0.0080



>20000
80
2500
30
<0.50
1:0.0120



>20000
80
2500
40
<0.63
1:0.0160



>20000
80
2500
50
<0.75
1:0.0200



>20000
80
2500
60
<0.88
1:0.0240



>20000
80
1750
6
<0.16
1:0.0034



>20000
80
1750
8
<0.19
1:0.0046



>20000
80
1750
10
<0.21
1:0.0057



>20000
80
1750
20
<0.34
1:0.0114



>20000
80
1750
30
<0.46
1:0.0171



>20000
80
1750
40
<0.59
1:0.0229



>20000
80
1750
50
<0.71
1:0.0286



>20000
80
1750
60
<0.84
1:0.0343



>20000
80
437.5
60
<0.77
1:0.1371



>20000
80
218.75
60
<0.76
1:0.2743









    A: Surf. E
    B: DAZOMET
    Media: PDB
    Inoculum size: 1.156E+06
    C. albicans ATCC 102031 No synergy
    A: Surf. E
    B: DAZOMET
    Media: 1/10 TSB
    Inoculum size: 5.7E+07
    P. aeruginosa ATCC 15442 No synergy
    A: Surf. E
    B: DAZOMET
    Media: 1/10 TSB
    Inoculum size: 1.8E+06
















PPM AI MIC Values




(24 hrs)










Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)





S. aureus
>20000
200
437.5
100
<0.52
1:0.2286


ATCC# 6538









    A: Surf. E
    B: DAZOMET
    Media: PDB
    Inoculum size: 1.156E+06
















PPM AI MIC Values (3rd day)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















A. niger
>20000
30
10000
2
<0.57
1:0.0002


ATCC#
>20000
30
10000
4
<0.63
1:0.0004


16404
>20000
30
10000
6
<0.70
1:0.0006



>20000
30
10000
8
<0.77
1:0.0008



>20000
30
10000
10
<0.83
1:0.0010



>20000
30
10000
20
<1.17
1:0.0020



>20000
30
5000
0.4
<0.26
1:0.0001



>20000
30
5000
0.5
<0.27
1:0.0001



>20000
30
5000
0.6
<0.27
1:0.0001



>20000
30
5000
0.8
<0.28
1:0.0002



>20000
30
5000
1
<0.28
1:0.0002



>20000
30
5000
2
<0.32
1:0.0004



>20000
30
5000
3
<0.35
1:0.0006



>20000
30
5000
4
<0.38
1:0.0008



>20000
30
5000
5
<0.42
1:0.0010



>20000
30
5000
6
<0.45
1:0.0012



>20000
30
5000
8
<0.52
1:0.0016



>20000
30
5000
10
<0.58
1:0.0020



>20000
30
5000
20
<0.92
1:0.0040



>20000
30
1750
0.4
<0.10
1:0.0002



>20000
30
1750
0.5
<0.10
1:0.0003



>20000
30
1750
0.6
<0.11
1:0.0003



>20000
30
1750
0.8
<0.11
1:0.0005



>20000
30
1750
1
<0.12
1:0.0006



>20000
30
1750
2
<0.15
1:0.0011



>20000
30
1750
3
<0.19
1:0.0017



>20000
30
1750
4
<0.22
1:0.0023



>20000
30
1750
5
<0.25
1:0.0029



>20000
30
1750
6
<0.29
1:0.0034



>20000
30
1750
8
<0.35
1:0.0046



>20000
30
1750
10
<0.42
1:0.0057



>20000
30
1750
20
<0.75
1:0.0114



>20000
30
875
3
<0.14
1:0.0034



>20000
30
875
4
<0.18
1:0.0046



>20000
30
875
5
<0.21
1:0.0057



>20000
30
875
6
<0.24
1:0.0069



>20000
30
875
8
<0.31
1:0.0091



>20000
30
875
10
<0.38
1:0.0114



>20000
30
875
20
<0.71
1:0.0229



>20000
30
437.5
0.2
<0.03
1:0.0005



>20000
30
437.5
0.3
<0.03
1:0.0007



>20000
30
437.5
0.4
<0.04
1:0.0009



>20000
30
437.5
0.5
<0.04
1:0.0011



>20000
30
437.5
0.6
<0.04
1:0.0014



>20000
30
437.5
0.8
<0.05
1:0.0018



>20000
30
437.5
1
<0.06
1:0.0023



>20000
30
437.5
2
<0.09
1:0.0046



>20000
30
437.5
3
<0.12
1:0.0069



>20000
30
437.5
4
<0.16
1:0.0091



>20000
30
437.5
5
<0.19
1:0.0114



>20000
30
437.5
6
<0.22
1:0.0137



>20000
30
437.5
8
<0.29
1:0.0183



>20000
30
437.5
10
<0.36
1:0.0229



>20000
30
437.5
20
<0.69
1:0.0457



>20000
30
218.75
5
<0.18
1:0.0229



>20000
30
218.75
6
<0.21
1:0.0274



>20000
30
218.75
8
<0.28
1:0.0366



>20000
30
218.75
10
<<0.34
1:0.0457



>20000
30
218.75
20
<0.68
1:0.0914









    A: Surf. D
    B: DAZOMET
    Media: PDB
    Inoculum size: 1.16E+08
    P. aeruginosa ATCC 15442 No synergy
    A: Surf. D
    B: DAZOMET
    Media: PDB
    Inoculum size: 5.72E+05
















PPM AI MIC Values (48 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















C. albicans
>20000
50
10000
10
<0.70
1:0.0010


ATCC# 10203
>20000
50
10000
20
<0.90
1:0.0020



>20000
50
10000
30
<1.10
1:0.0030



>20000
50
10000
40
<1.30
1:0.0040



>20000
50
5000
30
<0.85
1:0.0060



>20000
50
5000
40
<1.05
1:0.0080



>20000
50
2500
5
<0.23
1:0.0020



>20000
50
2500
10
<0.33
1:0.0040



>20000
50
2500
20
<0.53
1:0.0080



>20000
50
2500
30
<0.73
1:0.0120



>20000
50
2500
40
<0.93
1:0.0160



>20000
50
1750
6
<0.21
1:0.0034



>20000
50
1750
10
<0.29
1:0.0057



>20000
50
1750
20
<0.49
1:0.0114



>20000
50
1750
30
<0.69
1:0.0171



>20000
50
1750
40
<0.89
1:0.0229



>20000
50
875
30
<0.64
1:0.0343



>20000
50
875
40
<0.84
1:0.0457



>20000
50
437.5
40
<0.82
1:0.0914



>20000
50
218.75
40
<0.81
1:0.1829









    A: Surf. D
    B: DAZOMET
    Media: PDB
    Inoculum size: 1.56E+06
















PPM AI MIC Values (3rd day)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















A. niger
>20000
40
10000
0.2
<0.51
1:0.0002


ATCC#
>20000
40
10000
0.3
<0.51
1:0.0003


16404
>20000
40
10000
0.4
<0.51
1:0.0004



>20000
40
10000
0.5
<0.51
1:0.0005



>20000
40
10000
0.6
<0.52
1:0.0006



>20000
40
10000
0.8
<0.52
1:0.0008



>20000
40
10000
1
<0.53
1:0.0010



>20000
40
10000
2
<0.55
1:0.0020



>20000
40
10000
3
<0.58
1:0.0030



>20000
40
10000
4
<0.60
1:0.0020



>20000
40
10000
5
<0.63
1:0.0030



>20000
40
10000
6
<0.65
1:0.0000



>20000
40
10000
8
<0.70
1:0.0008



>20000
40
10000
10
<0.75
1:0.0001



>20000
40
10000
20
<1.00
1:0.0001



>20000
40
5000
0.2
<0.26
1:0.0037



>20000
40
5000
0.3
<0.26
1:0.0046



>20000
40
5000
0.4
<0.26
1:0.0000



>20000
40
5000
0.5
<0.26
1:0.0001



>20000
40
5000
0.6
<0.27
1:0.0001



>20000
40
5000
0.8
<0.27
1:0.0002



>20000
40
5000
1
<0.28
1:0.0002



>20000
40
5000
2
<0.30
1:0.0004



>20000
40
5000
3
<0.33
1:0.0006



>20000
40
5000
4
<0.35
1:0.0008



>20000
40
5000
5
<0.38
1:0.0003



>20000
40
5000
6
<0.40
1:0.0010



>20000
40
5000
8
<0.45
1:0.0020



>20000
40
5000
10
<0.50
1:0.0030



>20000
40
5000
20
<0.75
1:0.0060



>20000
40
5000
30
<1.00
1:0.0020



>20000
40
2500
0.2
<0.13
1:0.0001



>20000
40
2500
0.3
<0.13
1:0.0001



>20000
40
2500
0.4
<0.14
1:0.0002



>20000
40
2500
0.5
<0.14
1:0.0002



>20000
40
2500
0.6
<0.14
1:0.0002



>20000
40
2500
0.8
<0.15
1:0.0003



>20000
40
2500
1
<0.15
1:0.0004



>20000
40
2500
2
<0.18
1:0.0008



>20000
40
2500
3
<0.20
1:0.0012



>20000
40
2500
4
<0.23
1:0.0016



>20000
40
2500
5
<0.25
1:0.0020



>20000
40
2500
6
<0.28
1:0.0024



>20000
40
2500
8
<0.33
1:0.0032



>20000
40
2500
10
<0.38
1:0.0040



>20000
40
2500
20
<0.63
1:0.0080



>20000
40
2500
30
<0.88
1:0.0120



>20000
40
1750
0.2
<0.09
1:0.0001



>20000
40
1750
0.3
<0.10
1:0.0002



>20000
40
1750
0.4
<0.10
1:0.0002



>20000
40
1750
0.5
<0.10
1:0.0003



>20000
40
1750
0.6
<0.10
1:0.0003



>20000
40
1750
0.8
<0.11
1:0.0005



>20000
40
1750
1
<0.11
1:0.0006



>20000
40
1750
2
<0.14
1:0.0011



>20000
40
1750
3
<0.16
1:0.0017



>20000
40
1750
4
<0.19
1:0.0023



>20000
40
1750
5
<0.21
1:0.0029



>20000
40
1750
6
<0.24
1:0.0034



>20000
40
1750
8
<0.29
1:0.0046



>20000
40
1750
10
<0.34
1:0.0057



>20000
40
1750
20
<0.59
1:0.0114



>20000
40
875
0.2
<0.05
1:0.0002



>20000
40
875
0.3
<0.05
1:0.0003



>20000
40
875
0.2
<0.05
1:0.0002



>20000
40
875
0.4
<0.05
1:0.0005



>20000
40
875
0.5
<0.06
1:0.0006



>20000
40
875
0.6
<0.06
1:0.0007



>20000
40
875
0.8
<0.06
1:0.0009



>20000
40
875
1
<0.07
1:0.0011



>20000
40
875
2
<0.09
1:0.0023



>20000
40
875
0.2
<0.05
1:0.0002



>20000
40
875
0.3
<0.05
1:0.0003



>20000
40
875
0.4
<0.05
1:0.0005



>20000
40
875
0.5
<0.06
1:0.0006



>20000
40
875
0.6
<0.06
1:0.0007



>20000
40
875
0.8
<0.06
1:0.0009



>20000
40
875
1
<0.07
1:0.0011



>20000
40
875
2
<0.09
1:0.0023



>20000
40
875
3
<0.12
1:0.0034



>20000
40
875
4
<0.14
1:0.0046



>20000
40
875
5
<0.17
1:0.0057



>20000
40
875
6
<0.19
1:0.0069



>20000
40
875
8
<0.24
1:0.0091



>20000
40
875
10
<0.29
1:0.0114



>20000
40
875
20
<0.54
1:0.0229



>20000
40
437.5
0.4
<0.03
1:0.0009



>20000
40
437.5
0.5
<0.03
1:0.0011



>20000
40
437.5
0.6
<0.04
1:0.0014



>20000
40
437.5
0.8
<0.04
1:0.0018



>20000
40
437.5
0.4
<0.03
1:0.0009



>20000
40
437.5
1
<0.05
1:0.0023



>20000
40
437.5
2
<0.07
1:0.0046



>20000
40
437.5
3
<0.10
1:0.0069



>20000
40
437.5
4
<0.12
1:0.0091



>20000
40
437.5
5
<0.15
1:0.0114



>20000
40
437.5
6
<0.17
1:0.0137



>20000
40
437.5
8
<0.22
1:0.0183



>20000
40
437.5
10
<0.27
1:0.0229



>20000
40
437.5
20
<0.52
1:0.0457



>20000
40
218.75
0.6
<0.03
1:0.0027



>20000
40
218.75
0.8
<0.03
1:0.0037



>20000
40
218.75
1
<0.04
1:0.0046



>20000
40
218.75
2
<0.06
1:0.0091



>20000
40
218.75
3
<0.09
1:0.0137



>20000
40
218.75
4
<0.11
1:0.0183



>20000
40
218.75
5
<0.14
1:0.0229



>20000
40
218.75
6
<0.16
1:0.0274



>20000
40
218.75
8
<0.21
1:0.0366



>20000
40
218.75
10
<0.26
1:0.0457



>20000
40
218.75
20
<0.51
1:0.0914



>20000
40
20000
0.2
<1.01
1:0.0000









    A: Surf. D
    B: DAZOMET
    Media: 1/10 TSB
    Inoculum size: 5.72E+05
















PPM AI MIC Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















S. aureus
>20000
200
10000
20
0.60
1:0.0020


ATCC#
>20000
200
10000
30
0.65
1:0.0030


6538
>20000
200
10000
40
0.70
1:0.0040



>20000
200
10000
50
0.75
1:0.0050



>20000
200
10000
60
0.80
1:0.0060



>20000
200
10000
80
0.90
1:0.0080



>20000
200
10000
100
1.00
1:0.0100



>20000
200
10000
20
0.60
1:0.0020



>20000
200
5000
4
0.27
1:0.0008



>20000
200
5000
5
0.28
1:0.0010



>20000
200
5000
6
0.28
1:0.0012



>20000
200
5000
8
0.29
1:0.0016



>20000
200
5000
10
0.30
1:0.0020



>20000
200
5000
20
0.35
1:0.0040



>20000
200
5000
30
0.40
1:0.0060



>20000
200
5000
8
0.29
1:0.0016



>20000
200
5000
10
0.30
1:0.0020



>20000
200
5000
20
0.35
1:0.0040



>20000
200
5000
30
0.40
1:0.0060



>20000
200
5000
40
0.45
1:0.0080



>20000
200
5000
50
0.50
1:0.0100



>20000
200
5000
60
0.55
1:0.0120



>20000
200
5000
80
0.65
1:0.0160



>20000
200
5000
100
0.75
1:0.0200



>20000
200
2500
20
0.23
1:0.0080



>20000
200
2500
30
0.28
1:0.0120



>20000
200
2500
40
0.33
1:0.0160



>20000
200
2500
50
0.38
1:0.0200



>20000
200
2500
60
0.43
1:0.0240



>20000
200
2500
80
0.53
1:0.0320



>20000
200
2500
100
0.63
1:0.0400



>20000
200
1750
0.2
0.09
1:0.0001



>20000
200
1750
0.3
0.09
1:0.0002



>20000
200
1750
0.4
0.09
1:0.0002



>20000
200
1750
0.5
0.09
1:0.0003



>20000
200
1750
0.6
0.09
1:0.0003



>20000
200
1750
0.8
0.09
1:0.0005



>20000
200
1750
1
0.09
1:0.0006



>20000
200
1750
2
0.10
1:0.0011



>20000
200
1750
3
0.10
1:0.0017



>20000
200
1750
4
0.11
1:0.0023



>20000
200
1750
5
0.11
1:0.0029



>20000
200
1750
6
0.12
1:0.0034



>20000
200
1750
8
0.13
1:0.0046



>20000
200
1750
10
0.14
1:0.0057



>20000
200
1750
20
0.19
1:0.0114



>20000
200
1750
40
0.29
1:0.0229



>20000
200
1750
60
0.39
1:0.0343



>20000
200
1750
80
0.49
1:0.0457



>20000
200
1750
100
0.59
1:0.0571



>20000
200
875
0.2
0.04
1:0.0002



>20000
200
875
0.3
0.05
1:0.0003



>20000
200
875
0.4
0.05
1:0.0005



>20000
200
875
0.5
0.05
1:0.0006



>20000
200
875
0.6
0.05
1:0.0007



>20000
200
875
0.8
0.05
1:0.0009



>20000
200
875
1
0.05
1:0.0011



>20000
200
875
2
0.05
1:0.0023



>20000
200
875
3
0.06
1:0.0034



>20000
200
875
4
0.06
1:0.0046



>20000
200
875
5
0.07
1:0.0057



>20000
200
875
6
0.07
1:0.0069



>20000
200
875
8
0.08
1:0.0091



>20000
200
875
10
0.09
1:0.0114



>20000
200
875
20
0.14
1:0.0229



>20000
200
875
30
0.19
1:0.0343



>20000
200
875
40
0.24
1:0.0457



>20000
200
875
50
0.29
1:0.0571



>20000
200
875
60
0.34
1:0.0686



>20000
200
875
80
0.44
1:0.0914



>20000
200
875
100
0.54
1:0.1143



>20000
200
437.5
30
0.17
1:0.0686



>20000
200
437.5
40
0.22
1:0.0914



>20000
200
437.5
50
0.27
1:0.1143



>20000
200
437.5
60
0.32
1:0.1371



>20000
200
437.5
80
0.42
1:0.1829



>20000
200
437.5
100
0.52
1:0.2286



>20000
200
218.75
50
0.26
1:0.2286



>20000
200
218.75
60
0.31
1:0.2743



>20000
200
218.75
80
0.41
1:0.3657



>20000
200
218.75
100
0.51
1:0.4571









    A: Surf. A
    B: BIOBAN P-1487
    Media: PDB
    Inoculum size: 1.156E+06 CFU/ml
















PPM AI MIC Values (3rd day)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















A. niger
>20000
80
20000
5
<1.06
1:0.0003


ATCC# 16404
>20000
80
10000
6
<0.58
1:0.0006



>20000
80
10000
10
<0.63
1:0.0010



>20000
80
10000
20
<0.75
1:0.0020



>20000
80
10000
40
<1.00
1:0.0040



>20000
80
5000
30
<0.63
1:0.0060



>20000
80
5000
40
<0.75
1:0.0080



>20000
80
5000
60
<1.00
1:0.0120



>20000
80
2500
30
<0.50
1:0.0120



>20000
80
2500
40
<0.63
1:0.0160



>20000
80
2500
60
<0.88
1:0.0240



>20000
80
1750
30
<0.46
1:0.0171



>20000
80
1750
40
<0.59
1:0.0229



>20000
80
1750
60
<0.84
1:0.0343



>20000
80
875
30
<0.42
1:0.0343



>20000
80
875
60
<0.79
1:0.0686



>20000
80
437.5
50
<0.65
1:0.1143



>20000
80
437.5
60
<0.77
1:0.1371



>20000
80
218.75
50
<0.64
1:0.2286



>20000
80
218.75
60
<0.76
1:0.2743









    A: Surf. A
    B: BIOBAN P-1487
    Media: PDB
    Inoculum size: 1.156E+06 CFU/ml
















PPM AI MIC Values (48 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















C. albicans
>20000
30
20000
10
<1.33
1:0.0005


ATCC# 10231
>20000
30
10000
10
<0.83
1:0.0010



>20000
30
2500
8
<0.39
1:0.0032



>20000
30
2500
10
<0.46
1:0.0040



>20000
30
2500
20
<0.79
1:0.0080



>20000
30
1750
10
<0.42
1:0.0057



>20000
30
1750
20
<0.75
1:0.0114



>20000
30
437.5
20
<0.69
1:0.0457



>20000
30
218.75
20
<0.68
1:0.0914









    A: Surf. A
    B: BIOBAN P-1487
    Media: 1/10 TSB
    Inoculum size: 8.134E+07 CFU/ml
















PPM AI MIC Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















Ps. aeruginosa
>20000
80
20000
60
<1.75
1:0.0030


ATCC# 15442
>20000
80
5000
60
<1.00
1:0.0120



>20000
80
2500
40
<0.63
1:0.0160



>20000
80
2500
60
<0.88
1:0.0240



>20000
80
1750
50
<0.71
1:0.0286



>20000
80
1750
60
<0.84
1:0.0343



>20000
80
875
50
<0.67
1:0.0571



>20000
80
875
60
<0.79
1:0.0686



>20000
80
437.5
40
<0.52
1:0.0914



>20000
80
437.5
60
<0.77
1:0.1371



>20000
80
218.75
60
<0.76
1:0.2743









    No Synergy: S. aureus 
    A: Surf. E
    B: BIOBAN P-1487
    Media: PDB
    Inoculum size: 1.156E+06 CFU/ml
















PPM AI MIC Values (3rd day)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















A. niger
>20000
60
20000
30
<1.50
1:0.0015


ATCC# 16404
>20000
60
10000
30
<1.00
1:0.0030



>20000
60
5000
30
<0.75
1:0.0060



>20000
60
5000
40
<0.92
1:0.0080



>20000
60
5000
50
<1.08
1:0.0100



>20000
60
2500
30
<0.63
1:0.0120



>20000
60
2500
40
<0.79
1:0.0160



>20000
60
2500
50
<0.96
1:0.0200



>20000
60
1750
30
<0.59
1:0.0171



>20000
60
1750
40
<0.75
1:0.0229



>20000
60
1750
50
<0.92
1:0.0286



>20000
60
875
30
<0.54
1:0.0343



>20000
60
875
40
<0.71
1:0.0457



>20000
60
875
50
<0.88
1:0.0571



>20000
60
437.5
30
<0.52
1:0.0686



>20000
60
437.5
40
<0.69
1:0.0914



>20000
60
437.5
50
<0.86
1:0.1143



>20000
60
218.75
30
<0.51
1:0.1371



>20000
60
218.75
40
<0.68
1:0.1829



>20000
60
218.75
50
<0.84
1:0.2286









    A: Surf. E
    B: BIOBAN P1487
    Media: PDB
    Inoculum size: 1.808E+06
















PPM AI MIC Values (48 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















C. albicans
>20000
30
20000
0.04
<1.00
1:0.0000


ATCC# 10203
>20000
30
10000
0.04
<0.50
1:0.0000



>20000
30
10000
0.4
<0.51
1:0.0000



>20000
30
10000
4
<0.63
1:0.0004



>20000
30
10000
10
<0.83
1:0.0010



>20000
30
10000
20
<1.17
1:0.0020



>20000
30
5000
4
<0.38
1:0.0008



>20000
30
5000
5
<0.42
1:0.0010



>20000
30
5000
10
<0.58
1:0.0020



>20000
30
5000
20
<0.92
1:0.0040



>20000
30
2500
4
<0.26
1:0.0016



>20000
30
2500
10
<0.46
1:0.0040



>20000
30
2500
20
<0.79
1:0.0080



>20000
30
1750
5
<0.25
1:0.0029



>20000
30
1750
10
<0.42
1:0.0057



>20000
30
1750
20
<0.75
1:0.0114



>20000
30
875
8
<0.31
1:0.0091



>20000
30
875
10
<0.38
1:0.0114



>20000
30
875
20
<0.71
1:0.0229



>20000
30
437.5
8
<0.29
1:0.0183



>20000
30
437.5
20
<0.69
1:0.0457



>20000
30
218.75
10
<0.34
1:0.0457



>20000
30
218.75
20
<0.68
1:0.0914









    A: Surf. E
    B: BIOBAN P-1487
    Media: 1/10 TSB
    Inoculum size: 5.727E+07 CFU/ml
















PPM AI MIC Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















Ps. aeruginosa
>20000
60
20000
50
<1.83
1:0.0025


ATCC#15442
>20000
60
10000
50
<1.33
1:0.0050



>20000
60
5000
50
<1.08
1:0.0100



>20000
60
2500
50
<0.96
1:0.0200



>20000
60
1750
50
<0.92
1:0.0286



>20000
60
875
50
<0.88
1:0.0571



>20000
60
437.5
50
<0.86
1:0.1143



>20000
60
218.75
50
<0.84
1:0.2286









    A: Surf. E
    B: BIOBAN P-1487
    Media: 1/10 TSB
    Inoculum size: 1.808E+06 CFU/ml
















PPM AI MIC Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















S. aureus
>20000
30
2500
20
<0.79
1:0.0080


ATCC# 6538
>20000
30
437.5
20
<0.69
1:0.0457









    A: Surf. D
    B: BIOBAN P-1487
    Media: PDB
    Inoculum size: 1.156E+06
















PPM AI MIC Values (3rd day)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















A. niger
>20000
6
20000
0.6
<1.10
1:0.0000


ATCC# 16404
>20000
6
10000
3
<1.00
1:0.0003



>20000
6
5000
5
<1.08
1:0.0010



>20000
6
2500
3
<0.63
1:0.0012



>20000
6
1750
2
<0.42
1:0.0011



>20000
6
875
5
<0.88
1:0.0057



>20000
6
437.5
3
<0.52
1:0.0069



>20000
6
218.75
4
<0.68
1:0.0183









    A: Surf. D
    B: BIOBAN P-1487
    Media: PDB
    Inoculum size: 5.726E+05 CFU/ml
















PPM AI MIC Values (48 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















C. albicans
>20000
30
20000
3
<1.10
1:0.0002


ATCC# 10231
>20000
30
10000
2
<0.57
1:0.0002



>20000
30
10000
3
<0.60
1:0.0003



>20000
30
10000
10
<0.83
1:0.0010



>20000
30
10000
20
<1.17
1:0.0020



>20000
30
5000
5
<0.42
1:0.0010



>20000
30
5000
10
<0.58
1:0.0020



>20000
30
5000
20
<0.92
1:0.0040



>20000
30
2500
0.5
<0.14
1:0.0002



>20000
30
2500
10
<0.46
1:0.0040



>20000
30
2500
20
<0.79
1:0.0080



>20000
30
1750
4
<0.22
1:0.0023



>20000
30
1750
10
<0.42
1:0.0057



>20000
30
1750
20
<0.75
1:0.0114



>20000
30
875
20
<0.71
1:0.0229



>20000
30
437.5
5
<0.19
1:0.0114



>20000
30
437.5
10
<0.36
1:0.0229



>20000
30
437.5
20
<0.69
1:0.0457



>20000
30
218.75
8
<0.28
1:0.0366



>20000
30
218.75
10
<0.34
1:0.0457



>20000
30
218.75
20
<0.68
1:0.0914









    A: Surf. D
    B: BIOBAN P-1487
    Media: 1/10 TSB
    Inoculum size: 1.808E+05
















PPM AI MIC Values (24 hrs)











Test
Alone
Combination














Organism
CA
CB
Ca
Cb
S.I.
Ratio (Ca:Cb)
















S. aureus
>20000
30
20000
20
<1.67
1:0.0010


ATCC# 6538
>20000
30
10000
20
<1.17
1:0.0020



>20000
30
5000
20
<0.92
1:0.0040



>20000
30
2500
20
<0.79
1:0.0080



>20000
30
1750
20
<0.75
1:0.0114



>20000
30
218.75
20
<0.68
1:0.0914









    No synergy: P. aeruginosa 

The following biocides had no synergy against any organism tested when paired with the following surfactants:

    Surf. A
    Sodium Benzoate, TRIS NITRO
    Surf. E
    DMDMH
    Surf. D
    CS-1246, OPP, DMDMH

In the following combinations, the ratio of surfactant to biocide where synergy was observed were not commercially relevant, i.e., a ratio of 1:0.2 or greater (less biocide relative to surfactant). At these ratios, the biocide levels in a formulated product would be too low to be practical:

    Surf. A
    DIDAC, IPBC
    Surf. E
    CMIT/MIT, IPBC, OIT, TTPC, WSCP
    Surf. D
    CMIT/MIT, OIT, DIDAC
    (MBIT, IPBC, WSCP were synergistic only at 1:0.05 or worse except for one data point)

## Claims

1. A synergistic microbicidal composition comprising: (a) a nonionic surfactant with structure:


R2O(CH2CH(CH3)O)3(CH2CH2O)5H


where R2 is a mixture of C8-C14 linear alkyl groups; and (b) 2-bromo-2-nitropropane-1,3-diol; wherein a weight ratio of said nonionic surfactant to 2-bromo-2-nitropropane-1,3-diol is from 1:0.08 to 1:0.12 or 1:0.2 to 1:1.8286.

2. The synergistic microbicidal composition of claim 1 in which said mixture of C8-C14 linear alkyl groups comprises from 60 to 75 wt % C8-C10 linear alkyl groups and 25 to 40 wt % C12-C14 linear alkyl groups.