United States Patent (19) 11 4,259,103

Malek et al. 45) Mar. 31, 1981

54 METHOD OF REDUCING THE NUMBER OF 56) References Cited
MICROORGANSMS IN A MEDIA AND A
METHOD OF PRESERVATION U.S. PATENT DOCUMENTS
3,227,614 l/1966 Scheuer.................................. 424/25
(75) Inventors: James R. Malek; John L. Speier, both 3,624,224 11/1971 Watchung... ... 424/25
of Midland, Mich. 3,794,736 2/974. Abbott et al. .......................... 424/78
(73) Assignee: Dow Corning Corporation, Midland, Primary Examiner-Delbert R. Phillips
Mich. Assistant Examiner-Blondel Hazel
Attorney, Agent, or Firm-Robert L. McKellar
21 Appl. No.: 38,997 57 ABSTRACT
22) Filed: May 14, 1979 What is disclosed is a method of reducing microorgan
isms by contacting the microorganisms with the surface
of a substrate which has been altered by contacting a
Related U.S. Application Data substrate which develops a negative charge in water
63 Continuation-in-part of Ser. No. 19,532, Mar. 12, 1979, with a substance that ionizes in water to form cations
abandoned. and anions, which cations absorb to the substrate by a
process of ion exchange in which protons are replaced
51 Int. Cl........................ A01N 9/20: A01N 17/00; by the cations of the ionizing substance. The altered
A61 K9/00 surfaces can include hospital sheeting, clothing and a
(52) U.S.C. .......................................... 71/67; 424/16; wide variety of surfaces which come into contact with
424/25; 424/287 microorganisms.
58 Field of Search ....................... 424/16, 25, 26, 27,
424/28, 78, 31, 287; 71/67 26 Claims, No Drawings
 1
4,259,103
METHOD OF REDUCING THE NUMBER OF
MICROORGANISMS IN A MEDIA AND A
METHOD OF PRESERVATION
BACKGROUND OF THE INVENTION
This application is a continuation-in-part application
of Ser. No. 19,532, filed Mar. 12, 1979 now abandoned.
Almost concurrently with the notion that microor
ganisms cause infection and disease came the discovery
that the microorganisms could be controlled either by
killing all of them or reducing their numbers signifi
cantly.
Many methods have been devised that would accom
plish the end result including reducing all known in
fected things to ashes. As a practical matter, because all
such infected things could not be summarily subjected
to such treatment, other, more specific methods were
devised to control the microorganisms. One such spe
cific method was the treatment of milk and similar prod
ucts, at elevated temperatures high enough to destroy
objectional organisms but not high enough to alter the
chemical characteristics of the milk. This method was
developed by Louis Pasteur during the mid-nineteenth
century. Sterilization where foodstuffs were not in 25
volved constituted the use of chemicals such as phenol.
Phenol is effective but is a very harsh chemical in terms
of its acidic properties. Similar microbial poisons have
been commercially available for a number of years.
Certain amines and their quaternary salts, boric and 30
carboxylic acids and their salts and a host of other
chemicals have been used. These materials were used
neat or in aqueous solutions and then placed on the
substrate to be sterilized.
The utility and commercial potential of bacteriostatic 35
properties of certain cationic nitrogen compounds is
well known and seems first to have been recognized
about 1935 in U.S. Pat. No. 2,108,765 and, G. Domagk,
"A New Class Of Disinfectant Materials”, Deut. Med.
Wochn., 61, 829 (1935). Domagk recognized that if at
least one group in an amine was sufficiently large and
properly hydrophobic to give the molecule surfactant
properties in water, such a molecule then had bacterio
static or bacteriocidal properties. Following these pub
lications, scores of cationic surfactants were found to 45
have antimicrobial properties, and the teaching of Do
magk was proved to be correct.
It is observed that surfactant properties and antimi
crobial activity are found in common among com
pounds with at least one positively charged nitrogen 50
atom and at least one hydrophobic group. The hydro
phobic group should be equivalent to an aliphatic group
of at least C8H17- and not larger than C22H45-.
An excellent review of the antimicrobial behavior of
cationic substances in solution is to be found in "Cati 55
onic Surfactants', Marcel Dekker, Inc., New York,
1970, especially Chapter 14, Carl A. Lawrence-"Germi
cidal Properties of Cationic Surfactants'; and Chapter
11, Martin E. Ginn, "Adsorption of Cationic Surfac
tants on Miscellaneous Solid Substrates'. 60
Researchers have investigated ways in which sub
stances which were proven to be useful antimicrobials
in Solution could be made more persistent on substrates.
One such way was to attach an antimicrobial agent to a
substrate by some means. 65
In one method, an antimicrobial agent was mixed
with a curable coating. This method was quite satisfac
tory for prolonging the residence of the antimicrobial
2
agent but problems were still encountered as the antimi
crobial agent leached from the coating and was washed
away.
A better method evolved around 1965 when re
searchers discovered that antimicrobial organofunc
tionalsilanes could be chemically bound to certain sub
strates by what were believed to be Si-O linkages. The
method was described as orienting the organofunctional
10
silane in such a way that hydrolyzable groups on the
silicon atom were hydrolyzed to silanols and the sila
nols formed chemical bonds with the substrate and the
antimicrobial end of the molecule, for example a quater
nary nitrogen, was oriented away from the substrate; P.
A. Walters, E. A. Abbott and A. J. Isquith, Applied
15
Microbiology, Feb. 1973, pages 253–256. Theories abid
ing at the time alleged that any molecule having a reac
tive silyl group to bind to a surface and which also
displayed antimicrobial activity in solution would func
tion this way. In other words, if the molecule had activ
20
ity in solution, this activity could be fixed to a surface
by means of the silanol groups. The fixation would be so
strong that little or no material would be removed from
the surface by water. Thus, such a surface would affect
microorganisms in water without loss of the active
structure from the surface.
The theory could not be substantiated because no
method existed by which the theory could be tested.
The methods available were designed to measure the
activity of an agent in solution, and this fact is important
because the present invention deviates significantly
from that theory and greatly increases the number and
variety of substances that can be useful to render sur
faces antimicrobial.
A clear statement of the theory that a totally bound
substance could not affect microorganisms can be found
in L. A. Volf, "Imparting Antimicrobial Properties to
Fibers', Tekstilnaya Promnyshlenmost No. 8, 9 (1965),
U.S.S.R. (Translation by Techtron Corp.), who stated,
"Imparting antimicrobial activity to fibers can be car
ried out in two ways: (1) by sorption of a chemothera
peutic preparation or antiseptic agent (with subsequent
desorption) and (2) by chemical linking of the fiber with
compounds which impart to the fibers an anti-bacterial
effect (with their subsequent cleavage).” Speaking of
method (2) Volf goes on to state, "The problem of the
fixation of antimicrobial substances on the fiber by
means of a chemical bond contains at first sight, two
contradictory postulates. Actually on the one hand it is
dictated by the effort to prevent the removal of group
ings which are responsible for the antimicrobial effect
from the fiber during use and on the other hand it is
necessary to assume the transportation (diffusion) of
these groups within the microbial cells for otherwise,
the main goal of destroying the pathogenic microflora
at a distance will not be achieved.' He also adds, ". . .
the necessary condition for the manifestation of antibac
terial action is the presence of moisture. It is precisely
this which accomplishes the transport of active ingredi
ents to the microorganisms.'
In 1967, Canadian Pat. No. 774,529 was issued and
was completely contrary to Volf's opinion. This patent
stated that antimicrobial activity could in fact be put on
a substrate by binding an antimicrobial organosilicon
compound to a substrate but this patent showed no
example of how this should be done, nor an example
that this had been done. The first example in which this
was thought to be demonstrated was taught clearly by
 3
4,259,103
Isquith et al. later in a publication; A. J. Isquith, E. A.
Abbott and P. A. Walters, Applied Microbiology, De
cember, 1972, Pages 859-863. In this publication they
taught that only cationic antimicrobial agents which
were substituted by a (CH3O)3Si-group could display
antibacterial activity when deposited on a surface.
P. A. Walters, E. A. Abbott and A. J. Isquith, Ap
plied Microbiology, 25, No. 2, p. 253-256, Feb. 1973
amplified this teaching. They taught that 3-(trimethox
ysilyl)propyldimethylalkylammonium chlorides, with
alkyl chain lengths from 6 to 22 carbons in solution
were capable of killing algae. They state, "these com
pounds retained biological activity when durably at
tached to a surface'. They further teach that this was a
"unique property' of these compounds and that such
closely similar compounds as the commercial Benzalko
nium chlorides (Winthrop Laboratories, Division of
Sterling Drug Inc., New York, NY) did not display this
property.
The teaching was further amplified by E. A. Abbott
and A. J. Isquith, U.S. Pat. No. 3,794,736, Feb. 26, 1974.
In this patent they teach that a variety of siloxane-sub
stituted amines and amine salts which display antibacte
rial and antifungal properties in solution, continue to
display these properties when the substances were de
posited on surfaces. They claimed a method of inhibit
ing growth of bacteria and fungi by contacting said
organisms with a surface that had been coated with a
variety of siloxane-substituted organic amines or their
salts.
One substance favored by these investigators has
been adopted for commercial use to prepare antimicro
bial surfaces. This substance is essentially a methanol
solution of (CH3O)3Si(CH2)3N(CH3)3C18H37+Cl-.
The use of a filler, e.g. silica or titanium dioxide,
which had been treated with an organosilicon-sub
stituted quaternary ammonium salt, to inhibit the
growth of micro-organisms in or on the surface of a
polymeric organic matrix into which the filler had been
incorporated was taught in Canadian Pat. No.
1,010,782, issued May 24, 1977, to Roth.
This is a new way to use a surface treated as taught by
Isquith, et al. and Roth was surprised to discover that a
filler treated with 10% by weight of (CH3O)3.
Si(CH2)3NC18H37(CH3)3 + Cl- was effective, unexpect
edly contrary to the teaching of Isquith, et al. in that this
substance is not antimicrobial in solution.
Thus, it can be observed that all prior art indicates
that antimicrobial systems can be prepared by (1) treat
ment of surfaces with harsh chemicals; (2) chemical
bonding of chemical agents to substrates.
The Invention
It has now been discovered that there is a signifi
cantly better way to reduce the number of viable micro
organisms. By changing the surface characteristics of a
substrate and then contacting microorganisms with the
changed surface of the substrate, the microorganisms
are killed.
Thus what is disclosed is a method of reducing the
number of viable microorganisms in media by physi
cally contacting the microorganisms with a surface
which has been altered in a manner which comprises
contacting a substrate, which develops a negative
charge in water, with a substance that ionizes in water
to form cations and anions, which substance consists of
organic amines having the formula RR2R3N in which
R, R2 and R3 are independently hydrogen, alkyl or
4.
aralkyl groups wherein there is a total of less than 30
carbon atoms in the molecule; an organic quaternary
ammonium salt of the formula R4R5R6R7N-X-
wherein R, R5, R6 and R7 are independently alkyl or
aralkyl groups wherein there is a total of less than 30
carbon atoms in the molecule and X is a water soluble
monovalent anion; a sulfonium salt of the formula
R8R9R1OS+X- in which R8, R9, and R10 are indepen
dently alkyl groups or aralkyl groups wherein there is a
O total of less than 30 carbon atoms in the molecule or a
silylsubstituted alkyl radical of the formula
YSiCH2
15 Qb
in which Y is a hydrolyzable group, Q is an alkyl radical
of 1 or 2 carbon atoms of (CH3)3SiO, a has an average
value of 0-3; b has an average value of 0-3, n is an
integer of 1 or greater and X is a water soluble mono
valent anion; an isothiuronium salt of the formula
RSC(NH2)2X- in which R is independently alkyl or
aralkyl groups wherein there is a total of less than 20
carbon atoms in the molecule or a silyl substituted alkyl
25 radical of the formula
YSiCH2
Qb
30
in which Y is a hydrolyzable group, Q is an alkyl radical
of 1 or 2 carbon atoms or (CH3)3SiO, a has an average
value of 0-3; b has an average value of 0-3, and n is an
integer of 1 or greater and X is a water soluble mono
35 valent anion; a phosphonium salt of the formula
R4R5R6R7P+X- in which R4, R5, R6 and R7 are inde
pendently alkyl groups or aralkyl groups wherein there
is a total of less than 30 carbon atoms in the molecule or
a silyl substituted alkyl radical of the formula
Yich
Qb,
45 in which Y is a hydrolyzable group, Q is an alkyl radical
of 1 or 2 carbon atoms or (CH3)3SiO, a has an average
value of 0-3; b has an average value of 0-3, n is an
integer of 1 or greater and X is a water soluble mono
valent anion; a sulfonium salt of the formula O
50 Si(CH3)2CdH2dS+(R1)2X-2 in which R1 is indepen
dently an alkyl group or aralkyl group wherein there is
a total of less than 60 carbon atoms in the molecule, d is
an integer of 1 or greater and X is a water soluble
monovalent anion; an isothiuronium salt of the formula
55
OSi(CH3)2CdhdS+ C(NH2)2X-2 in which d is an
integer of 1 or greater and X is a water soluble mono
valent anion; a phosphonium salt of the formula O
Si(CH3)2CH2dP+(R12)3X-2 in which R12 is indepen
dently an alkyl group or aralkyl group wherein there is
60
a total of less than 60 carbon atoms in the molecule, d is
an integer of 1 or greater and X is a water soluble
monovalent anion and, an organic amine of the formula
65
OSCH):CH-CH-NH): t
H
 5
4,259,103
in which d is an integer of 1 or greater.
The value of n is normally 1-10 carbon atoms. For all
practical purposes, d generally has a value of 1-10 car
bon atoms.
As can be observed, the size of the alkyl or aralkyl
groups is dependent on the type of substance that is
being utilized. Thus, if the material is an organic amine
of the formula RR2RN, a quaternary ammonium salt
of the formula RR5R6RTN+X-, a sulfonium salt of the
formula R8R9R10SX or a phosphonium salt of the
formula R4R5R6R7P+X-, the total number of carbon
atoms in the molecule can be as large as 30 while the
isothiuronium salts of the formula RSC(NH2)2X
require a total of 20 carbon atoms or less. When the
substance is a sulfonium salt of the formula O
Si(CH3)2CH2dSt(R)2X2 or a phosphonium salt of
the formula O(Si(CH3)2CH2dP+(R12)3X-2, the total
number of carbon atoms in the molecule can be as large
as 60. It should be understood that as regards the latter
sulfonium and phosphonium salts, this invention does
not contemplate alkyl or aralkyl groups of 60 carbon
atoms but instead contemplates a disiloxane which con
tains alkyl or aralkyl groups on each sulfur orphospho material as later defined in this specification, for exam
rus atom such that there are less than 30 carbon atoms ple, a cationic surfactant, the surface is altered to the
per each sulfur or phosphorus per molecule. The size 25 extent that hydrogen ions of adsorbed water are re
and type of the alkyl or aralkyl group is significant in moved from the surface and moved to the bulk water
that the larger groups contribute to the permanence of
the surface alteration which in turn contributes to the
permanence of the antimicrobial effect.
Bulky organic cations adsorb very strongly to sub
strates. If the organic groups have straight chain alkyl
configurations then they tend to lay closely packed, side
by side, extending essentially upward from the substrate
surface. This closer proximity leads to bonding between
the alkyl groups through Van der Waals forces or some
similar effect. This increased bonding mechanism tends
to enhance the permanence of the cationic surface.
For purposes of this invention, generally one or two
large alkyl or aralkyl groups on the molecule are used.
For all practical purposes, molecules containing more
than two large alkyl or aralkyl groups are hard to pre
pare and further they tend to be too bulky to give the
optimum bonding between groups necessary for this
invention. Preferred for this invention are those com
pounds that contain at least one alkyl or aralkyl group
having greater than 10 carbon atoms.
X for purposes of this invention is a water soluble
monovalent anion. Examples of preferred anions are
Cl-, Br-, - and CH3COO-.
The advantages of the newly discovered method are
that a very large variety of surfaces, which include all
which develop a negative charge in contact with water,
can be altered to have antimicrobial activity by treat
ment with a very large variety of substances which need
have no biocidal activity in solution, but which need
only to ionize in solution in water to form a cation other
than a proton.
The most critical aspect of this invention is the fact
that the interface between surfaces of the substrates of
this invention and water must be altered. It should be
understood that the substrate is not chemically altered.
The interface between the surface of the substrate and
water is altered in that adsorbed water on the surface is
altered. Substrates that fall within the scope of this
invention are any substrates which develop a negative 65
charge on their surfaces in the presence of water. Such
things as fibers, textiles, particulate materials such as
sand, earth, concrete and masonry surfaces, wood, plas
6
tics and polymers all develop negative charges on their
surfaces in the presence of water.
The adsorption of cations upon mineral surfaces from
water has been studied extensively for many years and
a carefully detailed theory of the physical chemistry of
the process has been developed. The adsorption of cati
ons upon organic surfaces has also been extensively
studied, although the theory in this case is not as com
pletely developed. The concept that an electrically
O charged layer develops at a solid-liquid boundary or
interface has been accepted for at least as long as 160
years (F. Reuss, 1809, R. Porret, 1816). That a process
of ion exchange occurs in this charged layer has been
believed for at least 100 years (H. von Helmholtz, 1879).
15 A mathematical semi-quantitative description of the
process was well developed at least 50 years ago. (O.
Stern, 1924). Any good textbook on the physical chem
istry of surfaces describes these phenomena. See S.
Glasstone, Text-Book of Physical Chemistry, D. Van
20 Nostrand Company, Inc. New York, NY (1940) pp.
1194 et seq.
When surfaces are subjected to solutions of a cationic
and the cations move to the surface. It is theorized, but
the inventors do not wish to be held to such a theory,
that the cations replace hydrogen ions of water ad
30 sorbed on the surface of the substrate until the cations
are packed very closely together. Further, if the cati
onic material also contains appropriate groups in the
molecule (as explained later in this specification), the
alteration is permanent and the substrate surface re
35 mains antimicrobial.
Water adsorbed on the substrate is highly ionized,
thus H30 OH. When such a surface is treated with a
cationic material, the hydrogen ions of the adsorbed
water preferentially move to the surrounding bulk
water and the cation of the cationic material moves
preferentially to the surface. Thus, it is theorized that
there is an equilibrium at the surface i.e. Hi OH ---
H2O2HO --- H3O+. When the cations from the cati
onic material approach the surface the equilibrium is
45 upset and shifted to the right. There then exists on the
surface of the substrate, a tightly held, adsorbed system
consisting of the new cation and the adsorbed water
hydroxyl. The major discovery of this invention is that
surfaces so modified by cations are antimicrobial to any
50 microorganism that physically contacts them.
The second critical aspect of this invention is the fact
that the microorganisms must physically contact the
altered surface. By way of example, if the substrate is
glass beads which have been treated as discussed above
55 and the media is aqueous, for example, milk, and the
two are mixed together and stirred with a spatula for a
few minutes, then this would be sufficient force to get
the desired effect. Similarly, flowing a thin film of beer
across an altered surface, such as polyethylene, would
be enough force to get the desired effect. A third exam
ple would be stirring a solution containing the appropri
ate materials with a powered stirrer with enough speed
to make a vortex in the solution. A fourth example
would be blowing contaminated air through a filter,
such as a furnace filter, which has been treated to make
its surface antimicrobial, according to this invention.
The amount of force required is a function of the partic
ular system being used.
 7
4,259,103 8
Once the surface of the substrate has been altered, The time required for rinsing the surface is not criti
microorganisms that contact the surface are destroyed. cal. Generally water is used for this purpose but water
In certain examples there is no transfer of cationic mate and organic solvents can be used. For example, a rinse
rial into the media and since the adsorbed surface is not with a mixture of alcohol and water and then a clear
susceptible to erosion or leaching, the substrate can be rinse with water is generally sufficient.
used over and over without destroying its effectiveness. Some of the advantages of the method of this inven
Contemplated within the scope of this invention is tion are: Subtances chosen for use in this invention may
any substrate which develops a negative charge in wa be inoccuous compounds that are not toxic, corrosive,
ter, or hazardous to any appreciable extent. They do not
A surface suitably treated is essentially impervious to 10 have to be biologically hazardous substances.
attack by microorganisms. A surface in a medium Exceedingly small quantities of the substances are
which supports the growth of microorganisms has no capable of killing an apparently limitless number of
apparent effect upon organisms in the medium, but organisms that contact a properly treated surface.
organisms that contact the surface are killed. If the It can be noted, for example, that in Test No. 13 of
medium is stirred to cause organisms to contact the 15 Example 1, as low as 0.03 g. ion/100A2 was sufficient to
surface, the organisms can all be killed. In the absence give 100% reduction of microorganisms in only 1 treat
of stirring, no noticeable number of organisms are ment.
killed, but the surface is protected from attack by the No substance needs to enter media in which the mi
organisms, thus, there is a preservative effect contem croorganisms are found. Thus a medium need not be
plated within the scope of this invention. 20 contaminated by an antimicrobial agent to obtain a very
Substrates that develop negative charges in water at a strong antimicrobial effect.
pH below about 10 are suitable for the method of this A properly treated surface is capable of killing most
invention. These include all known textile fibers, such diverse spectra of microorganisms.
as cotton, cellulose acetate, polyesters, nylon, wool, Now, so that those skilled in the art can better under
rayon, acrylon, etc.; all known organic surfaces, such as 25 stand and appreciate this invention, the following exam
paints, polystyrene, silicone polymers, wood, rubber, ples are given. The examples following hereafter give
etc.; almost all inorganic surfaces, such as silica, sand, those skilled in the art the guidance necessary to prac
earth, silicate minerals, alumina, glass, including con tice this invention.
crete and masonry surfaces etc. which are not soluble in
Water. 30 EXAMPLE 1.
Substances capable of making these substrates antimi Many substrates were coated with materials by a
crobial are those that ionize in water to form cations to “Paint on Technique'. By this method a known amount
exchange with protons in the layer of water adsorbed of a chemical in solution was used to wet a solid. The
onto the substrate. These include such diverse materials solvent was evaporated and the amount of chemical in
as sodium chloride, ammonium chloride, every type of 35 solution was deposited on the surface of the solid.
amine, primary, secondary or tertiary, as well as quater Table I shows surprising results obtained when finely
nary ammonium salts, diamines, isothiuronium salts, divided powders are mixed mechanically with cultures
sulfonium salts, and phosphonium salts. No antimicro of microorganisms in what is called "The Powder
bial activity of the substances in solution is required nor Test'.
is the activity in solution of any influence on the level of 40 In the Powder Test, a quantity between 0.1 and 0.5g.,
antimicrobial activity shown by surfaces of substrates depending upon the particle size and density of a finely
treated with these substances. divided solid, was weighed into a clean Pyrex test tube.
Cations that contain bulky organic substituents are A sample treated with a chemical by the Paint on Tech
adsorbed to surfaces more strongly. They do not give nique, and an equal sized sample of untreated sample
rise to more active surfaces, but they do give rise to 45 were each mixed thoroughly (on a Vortex mixer) for 15
surfaces that are more resistant to loss of activity seconds and placed in an incubator for 30 minutes at 37
through loss of the cation from the surface. C.
Cations that contain silyl substituted organic substitu Each sample was then agitated for 10 seconds with 10
tents have a unique advantage over other cations. Cati ml. of sterile broth. A 0.1 ml. portion of the broth was
ons having substituents such as (CH3O)3Si- hydrolyze 50 agar plated and incubated 18 hours. After this time the
and polymerize and become essentially irreversibly number of viable organisms extracted from a treated
bound. sample was compared with the number extracted from
The manner of putting these substances on the sur an untreated sample. The percent reduction of viable
face of a substrate is critical to a degree. It is possible to cells was calculated from the difference between these
spray, dip, paint or roll on the cationic material or in the 55 values, i.e. (No. of cells from untreated sample-No. of
case of particulate solids dispersions may be used or in cells from treated sample divided by No. of cells from
the case of fibers soaking techniques may be used. The untreated sample)x100-% Reduction.
length of time that contact between the cationic mate If no living organisms were extracted, the sample was
rial and the substrate is necessary depends on the sub said to have achieved 100% kill.
strate and the type of cationic material being used. Fi 60 This technique permits the evaluation of the antimi
bers for example should not be treated for long periods crobial activity of a large surface area in relatively small
of time unless the surface of the internal fibers is to be volumes.
treated. Times of contact can range from a few seconds The data in Table I was obtained by use of 0.1 ml. of
to a few hours and the time required is that time neces 1:100 dilution of an 18 hour culture of Escherichia coli
sary to exchange the hydrogen ions of the adsorbed 65 B.
water on the surface for cations. In many cases the sample was washed and extracted
In actual practice, it is sometimes desirable to rinse with water before being subjected to the powder test.
the surface of the substrate after being treated. Typically, about 1 g of sample was mixed thoroughly
 4,259,103
9 10
with about 10 ml. of distilled water and the mixture was TABLE I-continued
filtered or centrifuged and sucked dry in a vacuum
filter. Sometimes samples were washed in this way Perce SUTCCS
EdinCaCO Yale Ellicial
tyle all-O
P By
CCIICUCl
many times. % Reduction
In Table I, the following compositions are shown 5 est g. ion/ Un
wherein the following Test Nos. correspond to the Test No. Substrate Solvent 100 A washed Washed
Nos. in the Table: Acetate()
5 (RLudox() FF 100 100
'; 6 S/ANOe) Water 8.2(b) 52
Test Substance 10 7 f 14(b) 38
No. tested 8 (CH3SiO3/2)0) f 100 100
1 (CH3)4NCI- 9 (CH3SiO3/2)(g) " (g) 100 37
2 10 Minusil FA 10.5 62 5.
3 11 Minusil Ethanol 9.4 100
4 f 13 ff 0.03(h) 100(h)
5 15 14 SAAN t 15. 100 100
6 15 (ECabosil) Hexane 0.1 70
7 16 Minusil Methanol 0 100 100
8 wn 17 FF Water 10 100 24
9 NaCl- 18 S/AN Water 10 44 18
10 NH4+Cl- 19 Minusil Methanol 10 99 90
11 C18H3N(CH3)3 I 20 20 p FF O 100 100
13 2 f 10 100 93
4. v 22 F. 10 53 38
15 C18H37NCH3)2 23 S/AN 10 64 45
16 Zephiran) 24 Minusil AF 0 23 O
17 n-C6H13NC5H5C1 25 S/AN FF 0. 100 46
18 25 26 Minusi Methanol 10 25 14
19 n-C4H9N(CH3)3Cl. 27 S/AN 10 24 17
20 (n-C4H9)2N(CH3)2Cl 28 Minusil Water 10 99 90
21 n-C4H9n-C8H17N(CH3)2Cl 29 Cellulose f 10 95 00
22 (C2H5)3N Acetate
23 30 S/AN AA 10 84 30
24 n-C4H9NH2 30 31 Minusil f 10 9.
25 wn 32 P Methanol 10 87 43
26 (C2H5)2NH 33 re t 10 69 33
27 34 f 10 100
28 (CH3O)3Si(CH2)3N(CH3)3tcl- 35 t Hexane 130 100 97
29 (CH3O)3Si(CH2)N(CH3)3Cl. 36 Cellulose Water 10 100 100
30 35 Acetate
31 (CH3O)3Si(CH2)3N(CH3)3toH- 37 Minusil Hexane 10 100 100
32 (CH3O)3Si(CH2)3N(C2H5)3t I 38 Methanol 10 100 100
33 (CH3O)3Si(CH2)3N(C2H5)3Cl. 39 f Water 10 60 21
34 (CH3O)3Si(CH2)3N(C2H5)2 40 Ludox O 100 100(k)
35 (CH3O)3SiCH2)3NHCH2CH2NH2 4. S/AN AF O 97 14
s {(CH3)3SiO3Si(CH2)3NHCH2CH2NH2 40 FF 10 63 57
7 43 inusi ex 10 98 96
38 (CHO)Si(CH2NCHst Cl- 44 Missil in 18 100 100
45 Ethanol 75
4142 (CH3O)3Si(CH2)3NCCH3)2C8H17Cl
(CH3O)3SiCH2)3N(CH3)2C4H9Cl 48
G
F
water 100
t 8
100
g20
43 (CH3O);Si(CH);N(CH3)2ClsHy7+CI- 45 49 f 10 100 96
: . . 50 FF 10 100 9
46 5 p , 10 96 83
47 (CH3O)3Si(CH2)3P(n-C4H9)3 I G f 8 8 65
48 . (CH3O)3Si(CH2)3P(C6H5)3 I 54 Hexane 10 100 48
49 (CH3O)3Si(CH2)3SC(NH2)2+Cl 50 55 A. 164 100 100
50 .. .. CHSC(NH2)2+Cl 56 f 170 100 100
51 (CH3O)3Si(CH2)3S(CH3)C18H37+1-
52 t (199.9 + 9% silica, 10 particle size, 1.1 m/g surface area (Pennsylvania Glass Sand,
--- Co.)
: ESSESSEE Chample was prepared with radioactive (CH3)3CHN C1 by Substantive
55 O{Si(CH3)2(CH2)3NHCH2CH2NH2)2 55 Rose Acetate fiber, 10 denier 0.1 m/g surface area (Celanese Cor
56 {(CH3)3SiO2SiCH3(CH2)3NHCH2CH2NH2
Zephiran (B) is a mixture of alkyl (40% C, 50% Cis, 10% CI) benzyldime- size.
ESSEE St. Marticle
thylammonium chlorides. (Winthrop Laboratories) “S/AN, Styrene-Acrylonitrile microcapsules, 1 to 20pt particle size, 0.63 m/g
surface area (Dow Chemical Co.)
Methyltrimethoxysilane (i.7 g., 0.1 mole) and tetramethylammonium chloride
TABLE I (1.2g, 0.015 mole), was dissolved in 5 mi. of water and methanol and water was
60 distilled from the solution as a polymeric gel formed. The gel was dried to an
Percent Reduction Of Viable Cells Of Escherichia coli-B By insoluble white solid which was ground to a fine powder. The powder was washed
Surfaces Treated By The "Paint-on' Technique exhaustively with water until the wash water contained no detectable chloride ion
by test with silver nitrate.
% Reduction Same as (f) with sodium chloride. Unwashed powder contained 2.6% Na', 0.79%
Test g. ion/ Un- Cl, washed powder contained 0.07% Na, nil C.
No. Substrate Solvent 100 A2 washed Washed Prepared with Cish N(CH3)2CH 'I applied at a low level and washed with
65 water, ethanol and dilute hydrochloric acid. Concentration of the radioactive cation
1 (E)Minusil(e) Methanol 10 100 on the solid was measured by radioactive assay.
2 . . 8.2 90 85 Cabosil MS-75, silica aerogel, 12-15 m. particle size, 274 m/g (Cabot Corp).
3 ' F. 02(b) 94 “washed exhaustively with water until free of detectable chloride ion.
4. Cellulose Water 540 30 77
 11
4,259,103
12
The data shown in Table I indicates a number of (CH3O)3Si(CH2)3NC5H5+Cl-. The salt of No. 40 has
unexpected effects. Most surprising is the fact that an N/Cl ratio of unity, but elemental analysis of the
every substance tested by the Powder Test displayed washed Ludox indicated an N/C ratio of about 296/1.
antimicrobial behavior. This was most surprising since Although essentially all of the chloride ion was not on
many of the substances have little or no antimicrobial the substrate, the surface was highly anti-microbial.
activity in solution.
The most widely standardized method of measuring EXAMPLE 2
antimicrobial activity of any substance is probably that To gain an analytical capability to show what oc
of Serial Tube Dilution. In this technique, 1 g of sub curred on a surface that caused it to become antimicro
stance is mixed with 9 ml. of sterile broth in a sterile 10 bial, typical substances were made radioactive. Radio
glass tube. One ml. of this mixture is transferred asepti tracer techniques were then applied to measure accu
cally to a second tube containing 9 ml. of broth to create rately very low concentrations of the substances and to
a 1/100 dilution of the substance being tested. This is locate the substances. (A) (CH3)3NCH3Cl was
repeated to form a series of dilutions each increasing by prepared by sealing C-methylchloride (1.96 mg., 0.039
a factor of 10. Thus 1/10, 1/100, 1/1000, 1/10,000 etc. 15 m mole), (0.252 m Ci) with methylchloride (218.3 mg.,
The broth is selected to contain vitamins, amino acids, 4.5 m mole) in methanol (0.83 g) and trimethylamine
sugars, etc. selected for optimum growth of the micro (1.8118 g., 30.71 m mole) in a flask at room temperature.
organism to be used for the test. Crystals formed until the mixture solidified. Vacuum
Each of the series of tubes is inoculated with about
0.1 ml. of a test organism and incubated for 24 hours at 20
was applied to remove all volatile materials leaving
37 C. for bacterial growth or at 25° C. for fungal 2.961.6 g. of fine white crystals of radioactive tetra
growth. After incubation the most dilute concentration methylammonium chloride with an activity of 0.025
of substance to inhibit growth of the organism is re mCi/g. (B) C18H37NCCH3)214CH3+. I was made in
corded as the Minimum Inhibitory Concentration essentially the same way from C18H37NCH3)2, CH3
(MIC). This is usually expressed as parts per million 25
and CH3 in methanol. The product was recrystal
(PPM) or g/ml., i.e. in parts by weight of broth. lized from ethanol to obtain 520.9 mg. with an activity
In most of the tests of Table I, the amount of the of 3.28 mCi/g. (C) (CH3O)3.
substance tested in the Powder-test was insufficient to Si(CH2)3.N(CH3)214CH2C17H35+Cl- was made from
kill the E. coli-B by dissolving from the substrates to octadecyl-1-4C chloride in hexane with (CH3O)3-
enter the culture. Calculations of the ug of substances 30
Si(CH2)3.NCCH3)2 in methanol in a sealed glass tube at
on the unwashed samples, if completely dissolved in 0.1 110 for 70 hours. The product was washed with hexane
ml. of culture, indicate that such solution could not and dried to give 10 mg. of crystals with an activity of
equal the MIC of the substance. The amount of sub 1 mCi/m mole. The hexane was used to dissolve and
stance after washing is unknown, but it was necessarily recrystallize 967.7 fing. of (CH3O)3-
less. 35
Si(CH2)3NCH3)2C18H37+Cl to obtain 1.023 g. of
Table II indicates the MIC, for E. coli-B for examples radioactive crystals with a specific activity of 0.36
for which this value is known and lists the maximum mCi/m mole. (D) (CH3O)3.
concentration of the substances that could be reached if Si(CH2)3.N(CH3)2CH3+ Cl- was prepared from 3
all the substances were dissolved from the surfaces of dimethylaminopropyltrimethoxysilane (2.44 g., 12 m
the substrates. The test nos. refer to Table I. 40 moles) in methanol (1.0 g) and CH3Cl (3.89 mg., 0.5
TABLE II
mCi) at 100 C. for 2 hrs in a sealed ampoule. All vola
tile materials were removed from the product under
Test MIC
Max.
ug/ml.
vacuum leaving white crystals (2.98 g., 98% yield).
No. Substance ug/ml in test Solutions of (B), (C), (D) were prepared in distilled
3 (CH3)4NCE > 10,000 < 175 45
water, each 10-3M concentrations. Each of these solu
O NH4+ Cl > 10,000 <250
tions was stirred with sufficient Minusil so that if the
3 C18H37NCH3)3 I 100 <3 solutions were depleted of (B), (C) or (D), the Mipusil
19 n-C4H9N(CH3)3Cl. > 10,000 (2700 should have 10 molecules of radio-cations per 100 A2 of
28
38
(CH3O)3SiOH2)3N(CH3)3Cl.
(CH3O)3SiCH2)3.NC5Hs Cl
> 10,000
> 10,000
<300
<3200
the surface of the Minusil. The stirring was for the time
42 (CH3O)3SiOH2)3N(CH3)2n-C4Hot Cli > 10,000 <2000 50 and at the temperatures indicated in Table III. The
41 (CH3O)3Si(CH2)3.N(CH3)2C8H17Cl > 10,000 (2600 Minusil was removed from the solutions, washed re
45 (CH3O)3Si(CH2)3N(CH3)2C18H37 Cl 1,000 <200 peatedly with water and assayed.
TABLE III
The examples of Table I indicated that every sub Rate and Level of Adsorption on Minusill
stance that dissolved in water and ionized in water to 55 Substance °C. Hours g-ion/100 A
form cations other than protons were capable of making (B) 25 2
surfaces antimicrobial as measured by the Powder-Test. 25 24 1
Every salt was effective and every amine. Amines are 60
60
2
24
1
l
well known to dissolve in water to form ammonium
(C) 25 2. 3.8
hydroxides, e.g. R3N + H2Oa2R3NH+ --OH-. 60 25 24 4.8
The fact that certain substrates that had been treated 25 168 7.2
with chloride salts and were washed until free or almost 25 336 7.4
free of chloride ion, indicated that the cation from each 60 2 6.5
60 4. 7.8
substance was the essential activating species that 60 24 9.62
caused surfaces to exhibit antimicrobial behavior. 65 (D) 25 Seconds 7
This was most surprising and also most evident with 25 Seconds l3(a)
examples such as No. 9, Na+Cl; No. 7, “Cellulose Acetate fibers, 0.12 m/g surface area.
(CH3)4N+Cl; No. 13, C18H37NCH3)3+I-; or No. 40,
 4,259,103 14
13
These data indicate that the adsorption of (B) was TABLE VI
limited to one molecule/100 A2, but that (C) was ad Substantive Treatment of Fibers with
sorbed without limit until the solution was depleted. (CH2O)2Si(CH2)3N(CH3)2CH2C17H35' Cl
This method in which the cations are adsorbed by a Decompositions/min/g of Solution
surface from a very dilute solution is referred to as the Cellulose Acetate Polyester Nylon-66
Substantive Method. This method is not like that of the Minutes pH 5 pH 7.5 pH 10 pH 7.5 pH 7.5
Paint-on Technique by which all of a substance is 0.0 159 163 167 169 167
coated onto or mixed with surfaces. Both methods pro 0.5
5
121
18
125
116
14
02 126 102
duce active surfaces, but the surfaces are not necessarily 10 15 00 107 97 102 104
the same. 30 94 99 93 81 94
By the Paint-on Technique (D) was put onto Minusil 1440 52 56 71 89
and Cellulose Acetate fiber and the effect of water Count on
Washed
washing, level of treatment and the effect of drying the Samples
solids before they were washed was studied. 15 dec/min/g 14,581 16,317 20,808 18,483 14,719
Cations/00
TABLE IV A2 of
Surface 1.3 1.4 1.8 1.6 0.8
(D) (CH3O)3Si(CH2)3N(CH3)2'CH3t Cl- on Minusi
Temp. of g-ion/100 A.
Drying C. initially wash 2 wash 20 Cellulose acetate fibers in a 1% solution as above
-25 2.3 1 1 adsorbed 210 cations/100 A2 of surface.
-25 9.5 1.8 1 Fibers (10 g) of Cellulose Acetate, Polyester, Cotton
70
100
9.3
4.6
.4
1.1
and Fiberglass were stirred in 100 ml. of 0.4%
-25 4.8 1.1 (CH3O)3Si(CH2)3N(CH3)214CH2C17H35+C in water
-25 25.5 2.4 1.3
25 at room temperature. The stirring was for a short time
-25 20a 9a until they were thoroughly wetted by the solution.
'Cellulose Acetate They were then taken from the solution, pressed as dry
as possible on filter paper and assayed for radioactivity.
The Substantive Method was employed at room tem The samples were then washed with distilled water
perature by stirring powders for.2 hours with enough 30 for five hours with a change of water every 10 minutes
(C) to apply 10 molecules/100 A2. After 2 hours, the for the first hour and every hour for four hours.
powders were separated in a centrifuge. They were TABLE VII
assayed and then water washed 3 times in a centrifuge Initial Activity After Washing
and assayed again. 35 Fiber mCi/g mCi/g
Polyester 49 15
TABLE V Cotton .82 69
Substantive Treatment With (C) At Room Temperature Cellulose
g. ion/100 A2 of surface Acetate 45 .44
Fiberglass 50 16
Substrate As Prepared Washed 40
Minusi 3.8 3.7
S/AN 7.6 6.9 The polyester fiber (0.5 g) was soaked in 0.01% con
Teflond 2.5 2.3 centration of above solution for 24 hrs, and assayed.
The fiber had adsorbed 3 cations/100 A2 of surface.
Finely powdered Teflon, 0.012 m/g surface area.
45 A second (0.5 g) sample was soaked in the same solu
These data indicate that such diverse surfaces as silica, tion for 24 hours. This sample adsorbed 2 cations/100
polystyrene-acrylonitrile, cellulose acetate and Teflon A2 of surface.
adsorb cations. The rate at which the ions are adsorbed A third sample (0.5 g) was soaked in the same solution
is very fast for the first molecular layer of ions. Com 74 hours. This sample adsorbed 5 cations/100 A2 of
surface. Sr.
pounds (B) and (D) are adsorbed very strongly up to 50
A fourth sample (0.5 g) was soaked in the same solu
approximately a limit of one molecular layer. Com
pound C is adsorbed slowly after about the first molecu tion 24 hpurs. This sample adsorbed less than 1 ca
tion/100 A2 of surface.
lar layer, but it is adsorbed without limit until the solu The solution was then assayed and had no detectable
tion is depleted. If excess (B) and (D) is applied by the 55 radioactivity, indicating that the solution had been de
paint-on technique, excess is removable by water until pleted of all radio cations.
the level near one molecular level remains.
Every sample in Tables III, IV aind V have near EXAMPLE 3
100% reduction of E. coli-B in the powder test. Minusil was treated with C18H37NCCH3)3 +36Cl- as
Fibers of Cellulose Acetate, Polyester and Nylon-66 60 nearly identically as possible to the sample in Table III.
(0.47 to 0.49 g) were stirred in 100 ml. of 0.001% A solution in water that contained 21.28X 10-6 moles of
(CH3O)3Si(CH2)3.N(CH3)214CH2C17H35+ Cl- in water chloride at a specific activity of 77.823 dpm/umol of
at room temperature. Periodically 0.1 ml. of the solution Cl was used to treat 1.0073 g of 10u Minusil.
was withdrawn and assayed in a liquid scintillation Unwashed powder indicated only 0.16 Cl- per 100
spectrometer to measure the rate at which the solution 65 A2 of surface. One wash with 15 ml. of water reduced
was depleted of radioactive cations. After 24 hours the the Cl- level to zero within the limits of detection of
fibers were washed thoroughly with distilled water and Cl-, which we estimate to be about 0.005 Cl/100
assayed. A2.
 15
4,259,103
The activity of the water remained unchanged during very little effect upon these values. Washing with etha
this experiment indicating all of the chloride ion re nol reduced them to 13.4 cations and 1.0 anions/100A2.
mained in the water and none became adsorbed onto the A mixture
Minusil although 1 cation/100 A2 of surface was ad
sorbed.
This is taken as compelling evidence that the cation
was adsorbed onto the surface by a process of ion ex
change, most likely described by an equation such as
O
H2O
Surface. H2O + Mt (a) +X" (aq) + H(aq)ed
Surface. OH M.
+ X (ag)
15
In this equation, M is a cation other than Ht, and an
H+ is displaced from water adsorbed onto the substrate
to become hydrated and to enter the aqueous phase as
Hit (ag).
Our data indicates that this equation is quantitatively 20
accurate up to the point that the concentration of M
near the surface of the substrate reaches a limit of about
one M+ ion/100 A2 of surface area.
A sample of Minusil with one non-radioactive
Me3NC18H37+ cation per 100 A2 was stirred in a dilute 25 face was subjected to sufficient washing, most or all of
aqueous solution that contained a large excess of the anions were absent from the surface.
C18H37NCCH3)2CH3+ cations. After two hours at Precipitation of the polymeric hydrolyzate of
room temperature, the Minusil assayed as having 1.0 (CH3O)3Si(CH2)3N(CH3)2C18H37+ Cl from dilute
radioactive cations/100 A2 of surface.
This indicated that the process of ion exchange took 30
place between radioactive and non-radioactive cations
on the surface and that an equilibrium was established
rapidly in this example. Thus:
Surface-OH.M. (a)+M+- 35
Surface-OH.M + M. (a).
This further indicates that simple cations do not ad
sorb beyond one cation per 100 A2 by the Substantive 40
Method from dilute solutions.
A sample of Minusil with one non-radioactive
Me3NC18H37+ cation per 100 A2 was stirred with a
dilute aqueous solution containing a large excess of
(CH3)3NCH3+Cl as above. Subsequent assay 45
showed no activity on the surface.
This indicated that a small cation such as (CH3)4N
cannot measurably displace a large one according to the
above equation.
This agrees with commonly accepted theory that 50
cations that associate in solution, e.g., form micelles at
suitable concentrations, adsorb to solids much more
strongly than do smaller cations.
EXAMPLE 4
55
The experiments of Example 3 which used (CH3O)3-
Si(CH2)3.N(CH3)214CH2C17H35+ Cl- were repeated
with (CH3O)3SiOH2)3.N(CH3)2C18H3s +36Cl-,
Minusil in Table III which had adsorbed 3.8 ca
tions/ 100A2 was assayed for 36Cl- and 1.89 36Cl 60 in Table VIII.
ions/100A2 was found. The sample was washed further
and the 36Cl- was reduced to 1.1 36Cl- ions/100 A2.
Minusil (1 g) was stirred at 60° C. for 24 hours with
2X 10-3 Molar solutions (100 ml) of (CH3O)3.
Si(CH2)3.N(CH3)214CH2C17H35+Cl- and of (CH3O). 65
Si(CH2)3.N(CH3)2C18H37+36Cl-. Assay of these sam
ples showed 16.3 cations and 4.3 anions per 100 A2 of
surface. Exhaustive water washing of these samples had
16
of 92 mg (CH3O)3.
Si(CH2)3.N(CH3)214CH2C17H35+ Cl- and 2.1851 g.
(CH3O)3Si(CH2)3.N(CH3)2C18H37 Cl- was hydro
lyzed with water and freed of all volatiles under vac
uum at room temperature or below to obtain a powder
essentially (HO)3Si(CH2)3.N(CH3)2C18H37+Cl with a
specific activity of 1.4719x 108 dpm/g.
The experiment was repeated with 3.1 g of (CH3O)3.
Si(CH2)3N(CH3)2C18H37+36Cl-,
Solutions of these salts, 7X 104 Molar and 0.40 g. of
Minusil were stirred 2 hours at room temperature and
water washed. The Minusi adsorbed 3.5 cations/100
A2, nearly duplicating the value in Table III under
nearly the same conditions. The Minusil had adsorbed
no detectable level of 36Cl.
These data indicate that the adsorption of the silyl
substituted salt differed from the adsorption of a simple
ammonium salt, by not being limited to one cation/100
A2 of surface. Adsorption beyond this level included
some part which was adsorbed as an ion pair. If adsorp
tion was from sufficiently dilute solution, or if the sur
solution in water in the absence of a substrate occurs
slowly. The precipitate was filtered from the water,
dried and analyzed for chloride ion. The level was too
low to measure by non-radiotracer techniques. The
dried precipitate was ground to a powder and gave
100% reduction of E. coli-B in the powder test. The
precipitate when dried at 100° C. is polymeric and no
longer soluble in water.
These data indicate that the silyl substituted salts can
polymerize in the absence of a substrate to form an
insoluble product which is formed by a process in
which ion exchange was only one part. The polymer
was a polysiloxane containing polymer units approxi
mately described 2S HO-O3/-
2SiOH2)3N+(CH3)2C18H37Cl-.
EXAMPLE 5
A sample (10 mg., 100 cm2) of Minusil having 3.8
(CH3O)3Si(CH2)3N(CH3)214CH2C17H35+ cations per
100 cm2 of surface was added to 2 ml of resting state E.
coli-B culture containing 3750 cells/ml in a small closed
vial, Vial-l.
A sample (10 mg) of untreated Minusil was added to
2 ml of the cells in Vial-2.
Only the culture was added to Vial-3.
The three vials were fastened to a wheel at a 45
angle and tipped end for end 24 times/min. as the wheel
rotated in an incubator at 37° C. Samples were removed
from the vials periodically and counts were made of the
viable cells/ml. at various times. The results are shown
TABLE VIII
Viable Cells/ml E. coli-B
Minutes Via-3 Vial-2 Wia.
O
5
3750
4370
3750
3900
3750
1600
5 5000 3700 29
30 4200 4300
45 400 3
 4,259,103
17 18
TABLE VIII-continued TABLE IX-continued
Viable Cells/ml E. coli-B Viable Cells of
Minutes Via-3 Vial-2 Vial-1 E. coli-B- Minusil
cm/ml : So t K(ml/cm/min)
60 4400 0 5 7 0.002(b)
8,243 3.6 0.004
Vials 3 and 2 showed an increase in the number of cells. . 3,750
1459
3.3
1.7
0.004
0.008
Vial-1 contained no more than about 4 pug of active 992 3.3(a) 0.004(a)
substance per ml. of culture, or less than 4% of the 10
100 7,502 1.75 0.004
(MIC) for the substance. Under the conditions of this 1,314 1.68 0.004
test, this small amount caused rapid reduction in the (surface of S/AN
'substance (CHO)Si(CH2)N(CH3). “Cl
number of viable organisms in the culture.
A plot of the logarithm of the number of viable cells
versus time for the cells in Vial-1 gave a straight line. The samples of Minusil and S/AN shown in Table V
See FIG. 1, which is a plot of the number of viable cells 15 were studied to see what effect the level of treatment of
on a log10 scale versus minutes on a linear scale. a surface has upon the specific rate constant K. The
This plot indicates that the organisms were killed by results are shown in Table X.
physical contact with the treated surface and that the (CH3O)3Si(CH2)3NCH3)2C18H37+Cl- (25 g) was
rate of decrease in the number that were viable under dissolved in 34.5 g. of methanol. Distilled water (15g.)
standardized conditions of agitation followed a mathe 20 was added to the solution which was then heated to.
matical equation very like one for the equation for the reflux and then all volatiles were removed under vac
kinetics of a second order reaction. Thus the rate of uum leaving a solid polymer, essentially {O3/.
decrease is: 2Si(CH2)3N(CH3)2C18H37Cl). This solid was
ground to a fine powder and washed with water. It was
dS
25 essentially insoluble in water. It was washed with etha
dt K (area of surface/ml}{S} nol and isopropanol and lost less than 2% by weight
during these washings. The washed powder was dried
in which and ground again and sieved. Particles smaller than 150
S=Concentration of viable organisms/ml um were tested. The area/g of this powder was not
K= A specific rate constant 30 accurately known, so K for this powder could only be
t=time approximated as the 10 mg. of this powder had an area
During an experiment the area remains constant. Inte equal to 10 mg. of Minusil.
gration therefore gives: TABLE X
molecules/ K(ml/cm/min
-Ln S=K{Area/ml)t+constant 35
Substance 100 A2 Substrate Ecoli-B P Aeruginosa
a plot of Ln S versus t called for by this equation is a C(a) Minusil 0.002 0.003
S/AN 0.001 0.001
straightline with a slope=K{area/ml). 7 Minusil 0.006 O.OO
A convenient way to determine K is from the equa SAN: 0.004 0.005
tion not applicable One 0.008
B(b) Minusil 0.008 0.002
S/AN 0.003 0.001
7 Minusil. 0,004
S/AN 0.002
where Sois the concentration of organisms at to and Sis (C is (CHO)Si(CH2)N(CH3)2CH7 Cl"
the concentration of organisms at time, t. 45 B is C8H3N(CH3)3C
The time required to kill half of the organisms can be
t1/2 and These data in Tables IX and X indicate that the sur
faces have remarkably similar effects upon organisms.
Multiple layers of the organic cation may slightly re
duce the activity of a surface. Data of Table III suggests
From FIG. 1, t1/2 is about 3.25 minutes and that layers of the organic cation would be removed
area/ml=50 cm2/ml. Then K=0.004 ml/cm2/min. during the test until only one layer remained. Thus the
This calculation says that the treated surface under activity measured in this manner for a simple cation,
these conditions of agitation killed half of the organisms most probably, is that of an adsorbed level one cation
every 3.25 minutes and it continued to do this until the 55 thick on the substrate.
viable organisms were very few. Experimentally, viable Multiple layers of the silyl-substituted cation seem to
organisms vanished in about 3 hr. although mathemati slightly increase the activity of the surface. The solid
cally this should require an infinite amount of time. polymeric substance had an activity similar to that of a
Experiments were repeated with a range of concen substrate with numerous layers of adsorbed cations.
trations of cells, So and surface areas/ml, see Table IX. 60 This suggests that as the number of layers of these cati
ons increase, the effects of the substrate vanish, so that
TABLE IX a substrate coated with multiple layers of a substance
Viable Cells of and the polymeric substance alone present very similar
E. coli-B- Minusil- . . . . . effects upon contact with organisms.
cm2/ml So it K(ml/cm/min); 65
25 10,270 5.6 0.005
EXAMPLE 6
50 4,639,000 3.2
3.0(a)
0.004
0.005(a)
Nylon fibers were treated with sufficient dilute solu
tion of (CH3O)3SiOH2)3NCCH3)2C18H37 - Cl- in water
 4,259, 103
19 20
to deposit 0.1% by weight upon the fiber if the solution coccus faecalis. In both cases a 50% reduction in viable
were depleted. The fibers were washed with water and cells was obtained over that recovered from a sample of
submitted to the test described in Example 5. the untreated soil.
This time 3 mi. of a tryptic soy broth was inoculated Fine sand (2 g) was sterilized with steam and wet
with 1 ml. of an 18 hour culture of Staphylococcus aureus 5 with 2 19% solution of (CH3O)3.
FDA-209 to give 4 m. of liquid in the vials which were Si(CH2)N(CH3)2C18H37 - Cl. Sample 1 was sucked dry
rotated 29 times/minute and approximately 10 mg. of on a filter. Sample 2 was heated to 70 until it was dry.
fiber was tested. The surface area per gram of this fiber These samples were tested by the Powder Test with
was not known, but it was much less than that of the 500,000 cells of Staphylococcus faecalis, and compared
samples in Example 5. O with untreated sterilized sand. Recovered viable cells
TABLE X from Sample 1 were 5.6% and from Sample 2, 5.9% of
the number recovered from untreated sand.
Minutes Vial-3 Vial-2 Vill Diatomaceous earth (Celite-281) was treated in the
() 9,000 9,000 19,000 5 same way. The Celite was divided into three samples.
30 38,000 20,000 16,000 Sample 1 was not washed. Sample 2 was washed with
6)
9() 45,000
25,000 11,000
7,000
100 ml. of distilled water. Sample 3 was washed five
2) 100,000 4,600 times with 100 ml. of water. These samples were tested
50 50,000 60,000 1,200 by the Powder Test with 500,000 cells of E. coli-B or of
18) 90,000 S. 700
20
S. faecalis and retested with 2,500,000 cells of each.
Untreated Celite permitted recovery of viable cells
The logarithm of the number of viable cells in Vial-l too numerous to count. Samples 1, 2 and 3 gave no
plotted versus time was a slightly concave curve. This is recovery of viable cells.
due to the fact that in the nutrient medium, cells were Sand and Celite are both silica. The superior activity
multiplying during this test, so that the net rate of their 25 of treated Celite was due to its having much more sur
decrease was the difference between the rates at which face area per gram than the fine sand.
they were dying and at which they were multiplying. The Celite (0.11 g) was stirred with 2 ml. of 1%
The experiment was repeated with 3 ml. of sterile (CH3O)3SiOCH2)3.N(CH3)24CH2C18H37 - C - in water
brine replacing the soy broth. and allowed to dry atroom temperature. It then assayed
TABLE X 30 as 69 molecules/100 A2 of the radioactive cation. It was
Viable cells of S. aureus in Serie Brine washed with 15 ml. of distilled water 3 times and then
Minutes Via-2 Wi
assayed as 10 molecules/100 A2.
15,000 15,000
Celite treated with only enough substance to put 1.8,
30 4,000 880
2.5, 2.8, 2.5 molecules/100 A2 was washed as above and
60 2,000 15 35. assayed as having 1.7, 2.3, 26, 2.5 molecules/100 A2.
90. , ,500 . 2
20 1,500 () EXAMPLE 8
The Celite from Example 7 having 2.5 cations/100
The logarithm of the number of viable cells in Vial A2 (1.5% by weight) was milled into a natural rubber,
in this case plotted versus time was a straight line as in 40 heavy duty tire tread stock and cured at 300 F. for 30
FIG. 1. minutes to prepare Sample 1.
The Celite (1.5% by weight) was also milled into a
EXAMPLE 7 passenger car-sidewall synthetic (SBR)-rubber tire
A 'X7' piece of polyethylene tubing was filled with stock and cured at 300 F., for 30 minutes to prepare
6' of fine sand, 6.920 g., (0,012 m2/g, total area 0.083 45 Sample 2.
m2). A solution of 0.00538 g., sufficient for 8 molecu Samples of these cured rubbers with and without the
les/100 A2 of (CH3O)3- Celite were washed thoroughly with water and placed
Si(CH2)3.N(CH3)214CH2C18H37 + Cl- in 10 ml. of water into petri dishes containing tryptic soy broth. Each dish
was permitted to drip onto the column of sand during 15 was then sprayed with a mixture of fungal spores of A.
minutes, followed by 50 ml. of distilled water. The 50 niger, A. flavus, A. versicolor, C. globosum and P.
column of sand was then assayed. It was uniformly funiculosum and incubated at 25 C. for one week. At
treated to a level of 0.52 to 0.56 molecules/100 A2. the end of one week visual inspection of the samples
The experiment was repeated with 5.218 g. of soil, showed that fungi had grown extensively upon the
which was mostly sand gently swirled in 3 ml. of the control samples that contained no treated Celite. Sam
solution for hour. It then had an activity of 0.86 55 ple 1 had fungi upon about 7% of its surface. Sample 2
mCi/g. had fungi growing upon about 2% of its surface.
Untreated soil (4,084 g) was put into the polyethyl EXAMPLE 9
ene tube and treated soil (0.9757g) was put on top of it.
Water was then trickled thru the column very slowly, A verythin film of Cellulose Acetate was dipped into
275 ml. in 120 hrs. The water was assayed and contained 60 2 2% solution of (CH3O)3.
less than 0.8% of the radioactivity introduced as treated Si(CH2)3.NCCH3)2C18H37 Cl- and hung up to dry.
soil. The column of soil was assayed. All the radioactiv 1'X "pieces of the film were submerged in cultures of
ity had remained in the top portion of treated soil. No E. coli-B and S. Faecalis which each contained
radioactivity could be detected in the untreated portion 1,000,000 cells/ml. They were removed and then
of the soil, indicating that the active substance had not 65 warmed to 37 C. for 30 minutes and submerged into
migrated during the long elution with water. tubes of sterile broth and warmed to 37 C. for 18 hrs.
The treated soil both before and after it had been No viable organisms were detected in the broth after
washed was tested by the Powder Test with Staphyllo this period of incubation with the treated film. Un
 4,259,103
21 22
treated film gave rise to growth of organisms in the and (B) (CH3O)3.
broth. The treated films had killed every organism they
had encountered when they were submerged in the Untreated pieces and pieces treated with (A) and (B)
cultures. were placed on sterile Sabouraud agar and sprayed with
EXAMPLE 10 a mixture of spores of A. niger, A. flavus, A. versicolor,
Six inch squares of thin Silastic (E) sheet, Medical C. globosum, and P. funiculosum and incubated at 27
Grade Silicone Rubber, were swirled about in 100 ml. for 21 days. Untreated pine was completely overgrown
of toluene for 1.5 hours at room temperature. The tolu with fungi in three days. Pine treated with (A) had very
ene contained 0.2% by weight (A) (CH3O)3- 10 little fungi on it in three days and no more than of its
Si(CH2)3N(CH3)214CH2C17H35+ Cl- to give Sample 1. surface had fungi after 21 days. Pine treated with (B)
The toluene contained 0.1% by weight (B) (CH3O)3. had about of its surface with fungi in 3 days. Fungi
Si(CH2)3.N(CH3)214CH3+Cl to give Sample 2. spread on this sample very slowly and it had portions
The samples were removed from the toluene and still free of fungi after 21 days.
hung up to dry at room temperature. One half of each 15 Fungi fluorished in the agar microscopically close to
sample was swirled about in 100 ml. of distilled water the treated wood. These samples showed no zone of
for 2 minutes. This was repeated 3 times with the fresh inhibition for growth of fungi near the wood.
water each time. The washed half of Sample 1 became EXAMPLE 13
Sample 3. The washed half of Sample 2 became Sample 20
4. These samples were assayed for the radioactive cati An experiment was carried out to determine if multi
ons. Sample 1 assayed as 0.4 mg(A)/g. of Silastic (R) ple washings with various liquid media would decrease
Rubber. Sample 3 assayed as 0.4 mg(A)/g. of Silastic (E) the activity on a treated substrate.
Rubber. Sample 2 assayed as 0.5 mg(B)/g. of Silastic (E) A commercial cellulose sponge was washed exhaus
Rubber. Sample 4 assayed as 0.4 mg(B)/g. of Silastic (E) 25 tively in tap water and boiled for 45 minutes in distilled
Rubber. water and dried to constant weight at 80° C. Cylinders
Six inch squares of the Silastic (R) Rubber sheet were were cut from the dry sponge with a cork borer. Each
swirled about in a 2% by weight solution of (CH3O)3. cylinder weighed about 1.05 g. and was about 3" in
Si(CH2)3N(CH3)2C18H37+C in water for a few min diameter and 1' long.
utes, removed from the water and dried on sterile filter 30 Cylinders were wet with aqueous solutions, about 35
paper in sterile petri dishes. Half of each square was ml. that contained 10 mg. of O3/-
washed with water as described immediately above. 2Si(CH2)3.N(CH2)3C18H37+ Cl- salt. The solution was
A piece of each of the samples were placed in 20 ml. squeezed and worked into a cylinder which swelled,
of sterile nutrient solution or in tryptic soy broth. Each softened and absorbed most of the liquid and was then
example was then sprayed with a mixture of spores of 35 labeled cylinders A. All cylinders were sterilized prior
A. niger, A. flavus, A. versicolor, C. globosum, and P. to washing in order to overcome any affect from bac
funiculosum and incubated for one week at 25 C.
At the end of the week the samples were examined teria in the sponge.
visually. Untreated Silastic (R) Rubber was overgrown A cylinders were heated to 80 for 0.5 hr. and were
completely with fungi. Unwashed samples had no fungi 40 labeled cylinders A'.
growing on them. Washed samples had a few spots on A and A cylinders were forced into a glass tube and
them, about 5% of the surfaces, upon which fungi were about 345 liters of tap water/g. of dry sponge was
Seein. forced thru them during 4 days.
With these examples fungiflourished microscopically These cylinders were labeled respectively B and B'.
close to the treated Silastic (R) Rubber but did not grow 45 An A cylinder was forced into a glass tube and one
on the surfaces. liter of homogenized milk was forced through it slowly.
EXAMPLE 11 This was labeled Cylinder C;
An A cylinder was forced into a glass tube and one
A typical latex paint was prepared with f60 g. of liter of Gallo brand Hardy Burgundy was forced
Rhoplex (R) AC35X latex paint and 135 g. of a pigment 50 through it slowly. This cylinder was labeled Cylinder
slip. Paper squares were dipped into the paint and hung D;
up to dry. (Sample A). An A cylinder was forced into a glass tube and one
Two ml. of a 50% solution of (CH3O)3. liter of Budweiser (E) Beer was forced through it slowly
Si(CH2)3.N(CH3)2C18H37+Cl- in methanol was stirred
into the paint and paper squares were dipped into the 55 andAnthis cylinder was labeled Cylinder E;
A cylinder was forced into a glass tube and one
paint and hung up to dry (Sample B).
These paper squares were then placed on the surface liter of sea water was forced through it slowly and this
cylinder was labeled Cylinder F.
of malt agar in petri dishes and inoculated with spores
of Pullularia pullulans ATC-9348 and incubated one were inoculated cylinder
An untreated and cylinders A, A, B and B'
with P. aeruginosa ATCC 870, eluted
month at 28-30° C. 60
At the end of this time the fungus had grown over with four 10 ml. portions of sterile water, pour plated in
much of the squares of Sample A. Fungus was found on letheen growth agar, incubated at 37 and the viable
2% of the surfaces of Sample B. cells in the agar were counted and shown in Table XIII
EXAMPLE 12
as number per ml.
65 Sponge cylinders A, B, C, D, and E were sterilized at
Square pieces of Ponderosa pine wood, 15 psig, 120 for 15 minutes in steam. An untreated
(1.5' x 1.5' x 0.25") were soaked for five minutes in 5% sponge cylinder and A, B, C, D, E and F were inocu
by weight solutions of (A) (CH3O)3. lated heavily and tested as above.
 23
4,259,103 24
TABLE XIII These were exceedingly small particle size disper
P. aeruginosa Recovered from 1 g. of Sponge sions with the appearance of a slightly hazy, slightly
No. of viable cells/ml. of culture
blue liquid. ' ' w
Sponge Sponge 5 The standard serial tube dilution test was used with
Not Sterilized Sterilized these colloidal dispersions. No agitation was required to
Control 232,700 2,075,000 keep these surfaces dispersed and Brownian motion was
A 380 86,750 sufficient to cause contact with organisms.
A' 200 Sample 2 was active at 100 ug/ml. 100 ug. of disper
B
B'
122,000
184,000
954,000
O
sion could contain no more than 2 ug. of
C 934,000 C12H25NCCH3)3 + Cl-, so that the apparent MIC for
D 352,000 this substance was decreased 100 to 1000 fold by ad
E S81,000 sorption on colloidal particles.
F 62,250
EXAMPLE 16
15
These data indicate that the cellulose sponge became A colloidal dispersion of methylsilsesquioxane
remarkably antimicrobial treated at a very low level. (CH3SiO3/2) particles 5% by weight was prepared in
Heating the sponge had very little effect upon its activ water with 0.7% dodecylbenzyldimethylammonium
ity. The activity was reduced but remained high after chloride as the dispersing agent. The colloidal particles
washing exhaustively with diverse liquids. 20 were about 125 Ain diameter.
The serial tube dilution test indicated that the M.I.C.
EXAMPLE 14 of this colloidal dispersion was 100 ppm for E. coli and
Pieces of cloth were treated by the Paint-on tech 1000 ppm for S. aureus. This would correspond to con
nique with (CH3O)3Si(CH2)3.N(CH3)2C18H37 + Cl- in centrations of the cationic dispersing agent of 0.7 ppm
water to treat the cloth to a level of 1% by weight. 25 for E. coli and 7 ppm for S. aureus.
Some pieces were sterilized after treatment in an auto The MIC of the dispersing agent in solution is 750
clave with steam for fifteen minutes. ppm for both E. coli and S. aureus. This experiment
A piece of Nylon cloth was treated with (CH3O)3. indicated that particles so smail that they move by
Si(CH2)3.N(CH3)3 + Cl- to give a level of 0.1% by Brownian motion as colloidal particles in water contact
weight. 30 and kill microorganisms. The activity of the cation in
The pieces of cloth were tested by AATCC-Test this example was increased by absorption on the particle
Method 100-1970, for evaluation of Antimicrobial Fin by 1000 fold for E. coli and 100 fold for S. aureus.
ishes on Fabrics with a contact time of 30 minutes. Compare, for example, page 54, Table 22 of Dekker,
supra page 3.
TABLE XIV
35 EXAMPLE 17
Type of Type of % Reduction
Fabric Organism Initial Sterilized Cotton thread was treated with a 0.5% solution of
Cotton S. aureus 93 92 (CH3O)3Si(CH2)3N(CH3)2C18H37+ Cl- in water and
K. pneumoniae 81.5 air dried. The treated thread and untreated thread were
Polyester S. aureus 99.1 40
K. pneumoniae 68.6 buried in soil inoculated with A. niger ATCC-9642; A.
Nylon S. aureus 100 flavus, ATCC 9643; A. versicolor ATCC 11730; P.
funiculosum, ATCC 9644; C. globosum ATCC 6205 as
The MIC of (CH3O)3SiOH2)3.N(CH3)3 + Cl is very in Mil. Std. 810B. The tensile strength of the thread was
high. The compound showed no biocidal activity in measured as in ASTM D 1682.
45
solution at concentrations as high as 10% by weight
against the organisms in Table XIV. Pounds/sq.in. Tensile Strength
Initial After 7 days
EXAMPLE 15 C) treatinct 5. 0.7
A colloidal dispersion of solid methylsilsesquioxane 50 treated cotton 5.8 5.6
(CH3SiO3/2) particles, 10% by weight, was prepared
by hydrolyzing an emulsion of methyltrimethoxysilane The treatment very effectively reduced or prevented
(10 g) in water (37.1 g) with 0.75 g. of (CH3O)3. fungal attack upon cotton.
Si(CH2)3.N(CH3)2C18H37 Cl as an emulsifier and 0.25
ml. of 1 N NH4OH as a catalyst for hydrolysis. 55 EXAMPLE 18
A second dispersion was made with (0.75 g) Nylon cloth was treated with a 0.1% solution of
C12H25NCCH3)3 + Cl- as the emulsifier. (CH3O)3Si(CH2)3.N(CH3)2C18H37 F.C. in water and
Methanol liberated by hydrolysis of the methoxysi air dried. The Nylon was then sprayed with a culture of
lanes was removed from a portion of each dispersion by Candida albicans ATCC 10231, and the number of via
distillation at 90-95 without disturbing the stability of 60 ble organisms recoverable from the cloth was deter
the colloidal dispersions. mined. ‘. . . . -
During these preparations the pH of the aqueous
phase decreased slowly from pH 11 due to the NH4OH,
to a pH of 6.5 for the first example and pH 8 in the Organisms recovered/ml
second. This change in pH indicated the ion-exchange 65 three runs Average % Reduction
described in Example 3 had occurred between the cati O treatment 200/210/205 205 ()
onic emulsifying agents and the surfaces of the colloidal treated. Nylon 22/24/18 . 21 9()
particles.
 25 4,259,103
26
Cotton cloth was treated in the same way and tested then it was allowed to cool to room temperature with
with Trichophyton interdigitale ATCC 9533. . . . . . . out stirring.
The cold cream was evaluated by a method set forth
% surface growth in 28 days in CTFA Cosmetic Journal, Vol. 5, No. 1, "Evaluation
5 of
Average % Methods for Determining Preservative Efficacy".
() treatment 100/100/100 Briefly, aliquats of the cream were challenged with the
treated cotton
100. organisms and any growth was observed at day inter
20/20/10 17
vals during which time the challenged plates were incu
bated at 37 C. The results of this test can be found on
A polyester-cotton cloth was treated with a 0.33%. 10 Table XV.
TABLE XV
"Evaluation of Treated Silica in Cold Cream Formulation"
CFTA Challenge Test - Challenge Organism
Pseudomonas Aeroginosa ATCC 15442
Bacterial Count
Sample Day 0 Day 1 Day 7 Day 14 Day 21 Day 28
control
(no treated
powder) 7,250,000 10,200,000 615,000 450,000 500,000 1825,000
5.0% powder
with 1.0%
active
compound 9,500 <13 <10 <10 <10 <10

That which is claimed is:

solution in the same way and sprayed with cultures of 1. A method of reducing the number of viable bac
the following organisms and the numbers recoverable teria, fungi, algae and yeast in media by physically con
were measured. :
tacting the bacteria, fungi, algae and yeast with a sur
30 face which has been altered in a manner which com
No. of Organisms/ml after 6 hrs. prises contacting a substrate, which develops a negative
Organism Untreated...si. Treated. . .2% Reduction charge in water, with an amount effective to inhibit the
Micrococcus sp. 215,500 . . . . 2,700 99 growth of said microorganisms of a substance that io
Staph. epidermidis 58,000 3,000 95 nizes in water to form cations and anions which sub
Enterobacter
aglomerans 1,355,000 16,500 90 35
stance is selected from the group consisting of sulfo
Acinetobacter. nium salts of the formula R8R9RIOS+X-,
salts of the formula ' '.
sulfonium
calcoaceticus 3,500 1,000 72
Micrococcus sp. 305,000 O 100
395,000 200 99
Staph. aureus 200,000 200 99
40

EXAMPLE 19 isothiuronium salts of the formula RSC(NH2)2X,

Aerosil-200 silica manufactured by the Degussa Co., isothiuronium salts of the formula
West Germany was treated in aqueous solution by mix 45
ing (CH3O)3SiOCH2)3SC(NH2)+2Cl- into the water otichs' C(NH2)2X
and adding the Aerosil-200 to the mixture. It was ho (CH3)2
mogenized for 5 minutes using a blender. The material
was then dried at 90° C. in an oven for 10-15 minutes.
The silica was then formulated into a cold cream in the 50 phosphonium salts of the formula R“R5R6R7P+X-,
following manner. Two parts were prepared. and phosphonium salts of the formula
Part A- 2.5gms Stearyl Alcohol
4.0 gms White petrolatum
otch +X-3
0.5gms Sorbitan Monooleate 55
(CH3)2 (R)
2.5gms Isopropyl Myristate
Part B Propylene glycol 3.5gms wherein R8, R9 and R10 are independently alkyl groups
*Polyoxyl-40 Stearate 1.25 gms or aralkyl groups wherein there is a total of less than 30
Water 35.8 gms
Preservative Powder 5% powder based on carbon atoms in the molecule or a silylsubstituted alkyl
Prepared Above total weight of B radical of the formula
with 1% active 60
perservative thereon
Polyoxyl-40 Stearate is a polyethylene oxide stearate ester having 40 CHCHO
units per stearic acid residue. Yich
Qh

The parts A and B were heated separately to 75° C. 65 where Y is a hydrolyzable group, Q is an alkyl radical of
and then B was added to A with agitation until the 1 or 2 carbon atoms or (CH3)3SiO, a has an average
material became creamy and smooth. The material was
slowly cooled with stirring until it began to set up and value of 0-3, b has an average value of 0-3, n is an
integer of 1 or greater and X is a water soluble mono
 4,259,103
27
valent anion, R is independently alkyl or aralkyl groups
wherein there is a total of less than 20 carbonatoms in
the molecule or a silylsubstituted alkyl radical of the
formula
Yich
Qi,
as defined above, R', R5, R6 and R7 are as defined above
for R8, R9 and R10; R1 and R12 are independently alkyl
groups or aralkyl groups wherein there is a total of less
than 60 carbon atoms in the molecule and d is an integer
of 1 or greater.
2. The method of claim 1 in which the microorgan
isms are gram-positive or gram-negative bacteria.
3. The method of claim 1 in which the microorgan
isms are fungi.
4. The method of claim 1 in which the microorgan
isms are yeasts.
5. A method as claimed in claim 1 wherein the sulfo
nium salt is a sulfonium halide.
6. A method as claimed in claim 5 wherein the halide
is chloride.
7. A method as claimed in claim 5 wherein the halide 25 26. A method of reducing the number of viable bacte
is iodide.
8. A method as claimed in claim 5 wherein the halide
is bromide. .
9. A method as claimed in claim 1 wherein in the 30
groups R, R9 and R10, at least one such group has at
least 10 carbon atoms.
10. A method as claimed in claim 5 wherein the sulfo
nium salt is (CH3O)3SiOH2)3S (CH3)C18H37.I.
11. A method as claimed in claim 5 wherein the sulfo
nium salt is (CH3O)3Si(CH2)3S+(CH3)C2H5I.
12. A method as claimed in claim 1 wherein the iso
thiuronium salt is an isothiuronium halide.
13. A method as claimed in claim 12 wherein the
isothiuronium salt is an isothiuronium chloride.
28
14. A method as claimed in claim 12 wherein the
isothiuronium salt is an isothiuronium iodide.
15. A method as claimed in claim 12 wherein the
isothiuronium salt is an isothiuronium bromide.
16. A method as claimed in claim 12 wherein the
isothiuronium salt is (CH3O)3SiOCH2)3S-C(NH2)2Cl.
17. A method as claimed in claim 1 wherein the phos
phonium salt is a phosphonium halide.
18. A method as claimed in claim 17 wherein the
O phosphonium salt is a phosphonium chloride.
19. A method as claimed in claim 17 wherein the
phosphonium salt is phosphonium iodide.
20. A method as claimed in claim 17 wherein the
phosphonium salt is a phosphonium bromide.
15 21. A method as claimed in claim 17 wherein the
phosphonium salt is (CH3O)3SiOH2)3P+(n-C4H9)3I.
22. A method as claimed in claim 21 wherein the
phosphonium salt is (CH3O)3SiCH2)3P+(C6H5)3I.
23. A method as claimed in claim 1 wherein the sulfo
20 nium salt is a sulfonium carboxylate.
24. A method as claimed in claim 1 wherein the iso
thiuronium salt is an isothiuronium carboxylate.
25. A method as claimed in claim 1 wherein the phos
phonium salt is a phosphonium carboxylate.
rial, fungi, algae and yeast in media by physically con
tacting the bacteria, fungi, algae and yeast with a sur
face which has been altered in a manner which com
prises contacting a substrate, which develops a negative
charge in water, with an amount effective to inhibit the
growth of said microorganism of a substance that io
nizes in water to form cations and anions which sub
stance consists of an organic amine of the formula
35
olich-CH-NH).
in which d in each case
it is
is anis integer
a
of 1 or greater.
45
50
55
 UNITED STATES PATENT AND TRADEMARK OFFICE
CERTIFICATE OF CORRECTION
DATED March 31, 1981
NVENTOR(S) James R. Malek and John L. Speier
It is certified that error appears in the above-identified patent and that said Letters Patent
are hereby Corrected as shown below
In Column 6, line 37; the formula reading "H30 OH" should
read "Hit OH".

In Column 9, line 64; the heading reading "g. ion/"

2
should
lOO A
read "g. ion/".
d
100 A2

In Column l0, line 6; the heading reading "g. ion/" should

100 A2
read "g. ion/".
100 A2
In Column l2, line 55; the heading reading "g-ion/l (00 A2.
d

should read "g-ion/100 A2".

In Column 13, line l8; the heading reading "g-ion/100 A2."
O
should read "g-ion/100 A2".
In Column l3, line 38; the heading reading
"g. ion/100 A2 of surface" should read
O
"g. ion/l.00 A2 of surface".
 UNITED STATES PATENT AND TRADEMARK OFFICE
CERTIFICATE OF CORRECTION
PATENT NO. : 4, 259,103 Page 2 of 2
ODATED : March 31, 1981
INVENTOR(S) : James R. Malek and John L. Speier
It is Certified that error appears in the above-identified patent and that said Letters Patent
are hereby corrected as shown below:
OIn Column l2, line 43; the line reading "Solutions of (B), (C),
(D) were prepared in distilled" should read "Solutions of (B),
(C), and (D) were prepared in distilled".
O
In Column l4, line l7; the words reading ti A2 of" should read
o
"A1 of ".

OIn Column l8, line 36; the heading reading "molecules/"

2
should read "molecules/".
o
OO A
100 A2

O Column 24, line 30; the word reading "absorption" should

read "adsorption".
eigned and escaled this
O sixth Da y of October 1981
SEAL
Attest:

O GERALD J. MOSSINGHOFF
Attesting Officer Commissioner of Patents and Trademarks