BIOCIDE PROTECTORS

PROTECTEURS DE BIOCIDES

English Abstract

A process for achieving sustained biocidal
effectiveness of organic biocides detrimentally
affected by the presence of iron metal, using one or
more biocide protectors selected from the group
consisting of molybdates, chromates, zinc salts, copper
salts, cadmium salts, dialkylthioureas, alkoxylated
rosin amines, certain azoles, certain phosphonates and
zinc dust. Biocidal compositions comprising one or
more such biocidal protectors are also described.

Claims

The embodiments of the invention in which an exclusive
property or privilege is claimed are defined as follows:
1) A process for treating an aqueous fluid in contact
with iron metal to sustain the biocidal effectiveness of
an organic biocide contained in the fluid and
detrimentally affected in untreated fluid by the presence
of the iron metal, comprising the step of adding to the
fluid at least one biocide protector selected from the
group consisting of molybdates, chromates, zinc salts,
copper salts, cadmium salts, dialkylthioureas,
alkoxylated rosin amines, azole biocide protectors,
phosphonate biocide protectors, and zinc dust, or a
mixture thereof, in an amount effective to inhibit the
loss of biocidal effectiveness and wherein the weight
ratio of biocide protector to biocide is in the range of
from 0.1:1 to 30,000:1.
2) The process of Claim 1 wherein the weight ratio of
biocide protector to biocide is in the range of from 1:1
to 20,000:1.
3) The process of Claim 1 wherein at least two of said
biocide protectors are added.
4) The process of Claim 1 wherein the biocide protector
is water insoluble.
5) The process of Claim 4 wherein the biocide protector
comprises zinc dust.
6) The process of Claim 1 wherein the biocide protector
comprises a zinc salt.
7) The process of Claim 6 wherein the zinc salt is zinc
dibenzylcarbamate or zinc sulphate.
49

8) The process of Claim 6 wherein the zinc salt is zinc
sulphate and wherein the dosage is between 10 ppm and
2,000 ppm.
9) The process of Claim 1 wherein the biocide protector
is a chromate.
10) The process of Claim 9 wherein the chromate is
potassium chromate.
11) The process of Claim 9 or Claim 10 wherein at least
20 ppm of the chromate is added.
12) A process according to Claim 1 wherein the biocide
protector comprises molybdate.
13) The process of Claim 12 wherein the molybdate is
sodium molybdate in a dosage of at least 150 ppm.
14) The process of Claim 1 wherein the copper salt is
copper sulfate in a dosage of at least 2 ppm.
15) The process of Claim 1 wherein the cadmium salt is
cadmium nitrate in a dosage of at least 5 ppm.
16) The process of Claim 1 wherein the biocide protector
is a phosphonate biocide protector having the formula:
<IMG>
wherein m is an integer from 1 to 10; R4 is hydrogen or an
alkyl group having from 1 to 4 carbons, and R5 is hydroxy,
amino, hydrogen or an alkyl group having from 1 to 4
carbons.

17) The process of Claim 1 wherein the organic biocide
comprises bis-trichloromethyl-sulphone, bis-tributyltin-
oxide, methylene bis-thiocyanate, 2,2-dibromo-3-nitrilo-
propionamide, and/or isothiazolone.
18) The process of Claim 1 wherein the organic biocide
comprises an isothiazolone having the general formula:
<IMG>
wherein R1 is a hydrogen atom, an alkyl group of 1 to 18
carbon atoms, alkenyl or alkynyl group of 2 to 18 carbon
atoms, and preferably 2 to 4 carbon atoms, a cycloalkyl
group of 3 to 12 carbon atoms, preferably having a 3 to 8
carbon atom ring, an aralkyl group of up to 10 carbon
atoms, or an aryl group of up to 10 carbon atoms; and R2
is a hydrogen atom, a halogen atom, or an alkyl group,
preferably having 1 to 4 carbon atoms; and R3 is a
hydrogen atom, a halogen atom, or an alkyl group,
preferably having 1 to 4 carbon atoms; or R2 and R3, taken
together, complete a benzene ring, optionally substituted
with at least one member selected from the group
consisting of halogen atoms, nitro groups, alkyl groups
having 1 to 4 carbon atoms, cyano groups, and alkoxy
groups having 1 to 4 carbon atoms.
19) The process of any one of claims 1 to 10 or 12 to 18
wherein the organic biocide concentration is between 0.1 ppm
and 10,000 ppm.
20) The process of Claim 17 wherein the organic biocide
comprises 5-chloro-2-methyl-4-isothiazolin-3-one and/or
methyl-4-isothiazolin-3-one.
51

21) A process for treating an aqueous metal working
fluid in contact with iron metal to sustain the biocidal
effectiveness of an organic biocide contained in the
fluid and detrimentally affected by the presence of the
iron metal, comprising the step of adding to the fluid at
least one biocide protector selected from the group
consisting of molybdates, chromates, zinc salts, copper
salts, cadmium salts, dialkylthioureas, alkoxylated rosin
amines, azole biocide protectors, phosphonate biocide
protectors, and zinc dust, or a mixture thereof, in an
amount effective to inhibit the loss of biocidal
effectiveness and wherein the weight ratio of biocide
protector to biocide is in the range of 0.1:1 to
30,000:1.
22) The process of Claim 21 wherein the aqueous metal
working fluid is a cutting oil.
23) The process of any one of claims 1 to 10, 12 to 18 or 20
to 22 wherein the aqueous fluid has an alkaline pH.
24) The process of claims 19 wherein the aqueous fluid has
an alkaline pH.
25) A biocidal composition for use in treatment of
aqueous fluids in contact with iron metal comprising an
organic biocide which is detrimentally affected by the
presence of iron metal and at least two biocide
protectors selected from the group consisting of
molybdates, chromates, zinc salts, copper salts, cadmium
salts, dialkylthioureas, alkoxylated rosin amines, azole
biocide protectors, phosphonate biocide protectors, and
zinc dust and wherein the weight ratio of biocide
protector to biocide is in the range of from .1:1 to
30,000:1.
52

26) The composition of Claim 25 wherein at least one of
the biocide protectors is a zinc salt.
27) The composition of Claim 25 wherein the organic
biocide comprises bis trichloromethyl-sulphone, bis
tributyltin-oxide, methylene bis-thiocyanate and/or 2,2
dibromo-3-nitrilo-propionamide.
28) The composition of Claim 27 wherein the organic
biocide comprises an isothiazolone having the general
formula:
<IMG>
wherein R1 is a hydrogen atom, an alkyl group of 1 to 18
carbon atoms, alkenyl or alkynyl group of 2 to 18 carbon
atoms, and preferably 2 to 4 carbon atoms, a cycloalkyl
group of 3 to 12 carbon atoms, preferably having a 3 to 8
carbon atom ring, an aralkyl group of up to 10 carbon
atoms, or an aryl group of up to 10 carbon atoms; and R2
is a hydrogen atom, a halogen atom, or an alkyl group,
preferably having 1 to 4 carbon atoms; and R3 is a
hydrogen atom, a halogen atom, or an alkyl group,
preferably having 1 to 4 carbon atoms; or R2 and R3, taken
together, complete a benzene ring, optionally substituted
with at least one member selected from the group
consisting of halogen atoms, nitro groups, alkyl groups
having 1 to 4 carbon atoms, cyano groups, and alkoxy
groups having 1 to 4 carbon atoms.
53

29) The composition of Claim 25 the two biocide
protectors are zinc sulphate and hydroxyethylidene
diphosphonic acid; wherein the weight ratio of zinc
sulphate to the organic biocide is from 0.5:1 to 1,000:1;
and wherein the weight ratio of hydroxyethylidene
diphosphonic acid to the organic biocide is from 0.25:1
to 1,000:1.
30) The composition of Claim 25 comprising zinc sulphate
and a chromate; wherein the weight ratio of zinc sulphate
to the organic biocide is from 0.5:1 to 1,000:1; and
wherein the weight ratio of the chromate to the organic
biocide is from 0.5:1 to 1,000:1.
31) The composition of Claim 25 wherein the organic
biocide comprises 5-chloro-2-methyl-4-isothiazolin-3-one.
32) The composition of Claim 25 wherein the organic
biocide comprises methyl-4-isothiazolin-3-one and 5-
chloro-2-methyl-4-isothiazolin-3-one in a weight ratio of
approximately 8.6:2.6.
33) A biocidal composition for use in treatment of
aqueous fluids in contact with iron metal comprising an
organic biocide which is detrimentally affected by the
presence of iron metal and at least one biocide protector
selected from the group consisting of molybdates,
chromates, zinc salts, copper salts, cadmium salts,
dialkylthioureas, alkoxylated rosin amines, azole biocide
protectors, phosphonate biocide protectors, and zinc
dust, or a mixture thereof, in an amount effective to
inhibit the loss of biocidal effectiveness and wherein
the weight ratio of biocide protector to biocide is in
the range of 0.1:1 to 30,000:1.
54

34) The composition of Claim 33 wherein the organic
biocide comprises an isothiazolone and the biocide
protector is molybdate in a weight ratio of isothiazolone
to molybdate of from 50:1 to 30,000:1.
35) The composition of Claim 33 wherein the organic
biocide comprises an isothiazolone and the biocide
protector is chromate in a weight ratio to isothiazolone
to chromate of from 4:1 to 3,000:1.
36) The composition of Claim 33 wherein the organic
biocide is an isothiazolone and the biocide protector is
selected from diethylthiourea in a weight ratio to
isothiazolone from 0.5:1 to 1,000:1, ethyoxylated rosin
amine in a weight ratio to isothiazolone from 0.1:1 to
1,000:1, benzotriazole in a weight ratio of isothiazolone
from 2:1 to 10,000:1, and 1-hydroxyethylidene-1,1-
diphosphonic acid in a weight ratio to isothiazolone from
2.5:1 to 5,000:1.
37) The composition of Claim 33 wherein the biocide
protector is water insoluble.
38) The composition of Claim 37 wherein the biocide
protector is zinc dust.
39) The composition of Claim 38 wherein the organic
biocide is isothiazolone; the biocide protector is zinc
dust; and wherein the weight ratio of zinc dust to
isothiazolone is from 5:1 to 5,000:1.
40) The composition of Claim 33 wherein the organic
biocide comprises methyl-4-isothiazolin-3-one and 5-
chloro-2-methyl-4-isothiazolin-3-one in a weight ratio of
approximately 8.6:2.6.

41) The process of claim 11 wherein the organic biocide
concentration is between 0.1 ppm and 10,000 ppm.
42) The process of claim 11 wherein the aqueous fluid has an
alkaline pH.
56

Description

-
1 33482~
FIELD OF THE lNV~I~lION
This invention relates in general to treatment of
aqueous fluids to protect the effectiveness of biocides
contained therein, and more particularly, to treating
an aqueous fluid in contact with iron metal to sustain
the biocidal effectiveness of organic biocides
contained in the fluid and detrimentally affected in
untreated fluid by the presence of the iron metal.
BACKGROUND OF THE l~v~NlION
Biocides have long been used to prevent the
proliferation of bacteria and other microorganisms in
aqueous solutions. They are often instrumental for the
efficient operation of industrial processes. Much work
has been directed to developing organic biocides such
as various isothiazolones, sulphones, thiocyanates and
nitrilopropionamides which are now known to be
effective in aqueous solution. Certain 3-isothiazolones
have been particularly useful as broad spectrum
microbicides. For example, U.S. Patent No. 4,539,071
to Clifford et al. describes their use in certain
cooling waters and paper-making process waters with
glutaraldehyde to inhibit bacterial growth and slime
formation. However, the tendency of 3-isothiazolones
to chemically decompose in solution requires the
addition of stabilizers for many applications. U.S.
Patent No. 4,031,055 to DuPont et al. describes use of
compounds containing zinc, molybdenum, copper, lead, or
- 2 - ~

1 334824
, .
mercury to stabilize formulations for mildew resistant
coatings. U.S. Patent No. 3,870,795 to Miller et al.
describes a method of stabilizing various
isothiazolone-containing solutions against chemical
decomposition by adding nitrates and nitrites of
selected metals such as zinc. In any case
isothiazolones, as well as other organic biocides, have
been developed and adapted for use in aqueous solution.
SUMMARY OF THE INVENTION
It has been found that when exposed to iron metal,
solutions containing isothiazolones or certain other
organic biocides can rapidly lose their biocidal
effectiveness, and that addition of certain protective
agents to the solution can inhibit such loss. The
biocide protectors of this invention comprise a group
consistinq of molybdates, chromates, sulphates, zinc
salts, zinc dust, copper salts, cadmium salts,
dialkylthioureas, alkoxylated rosin amines, certain
azoles, and certain phosphonates and mixtures thereof.
Preferably, the biocide protectors are water soluble.
It is an object of this invention to sustain the
biocidal effectiveness of organic biocides contained in
aqueous fluids in contact with iron metal. It is a
specific object to provide biocide protectors which
inhibit the loss of biocidal effectiveness in aqueous
solutions in contact with iron metal and containing
organic biocides detrimentally affected by the presence
of the iron metal. Other objects and advantages of the

~ I 334~32~
invention will be apparent from the following detailed
description of the invention.
DETAILED DESCRIPTION OF THE lNV~NlION
The present invention is directed toward
protecting the biocidal effectiveness of aqueous
solutions of organic biocides in general, and
isothiazolones (i.e. isothiazolinones) in particular,
which come into contact with iron metal. As used in
this specification, isothiazolones are intended to
include, in general, those which lose their
effectiveness upon exposure to iron metal and have the
formula:
S C- R3
11
C/
o
wherein Rl is a hydrogen atom, an alkyl group of 1 to
18 carbon atoms, alkenyl or alkynyl group of 2 to 18
carbon atoms, and preferably 2 to 4 carbon atoms, a
cycloalkyl group of 3 to 12 carbon atoms, preferably
having a 3 to 8 carbon atom ring, an aralkyl group of
up to 10 carbon atoms, or an aryl group of up to 10
carbon atoms; and R2 is a hydrogen atom, a halogen
atom, or an alkyl group, preferably having 1 to 4
carbon atoms; and R3 is a hydrogen atom, a halogen
atom, or an alkyl group, preferably having 1 to 4
carbon atoms; or R2 and R3, taken together, complete a

1 33~824
benzene ring, optionally substituted with one or more
halogen atoms, nitro groups, alkyl groups having 1 to 4
carbon atoms, cyano groups, alkoxy groups having 1 to 4
carbon atoms, or the like.
Examples of these biocides are presented in U.S.
Patent No. 3,870,795 to Miller et al.Cwhich is hereby
incorporated in this specification by reference. In a
commercial preferenc~, R1 is methyl, R2 is hydrogen,
and R3 is either hydrogen or a halogen, most preferably
hydrogen or chlorine. However, protection of other
isothiazolones as well as other classes of organic
biocides which are found to lose effectiveness in the
presence of metal iron is also contemplated within the
~ scope of this invention. Indeed, as discussed below,
the invention can be practiced with a broad range of
other organic biocides which are detrimentally
influenced by metal iron.
The effect of iron metal on isothiazolones is
illustrated by the non-limiting examples, numbered I
through VI, which follow.
EXAMPLE I
Isothiazolone solutions were prepared using Kathon*
886 marketed by Rohm & Haas Co. of Philadelphia,
Pennsylvania and having as active ingredients 8.6
weight percent 5-chloro-2-methyl-4-isothiazolin-3-one,
and 2.6 weight percent methyl-4-isothiazolin-3-one; and
Dearborn 6102*marketed by Dearborn Chemical Co. of
Mississauga, Onatrio and having as active ingredients
0.86 weight percent 5-chloro-2-methyl-4-isothiazolin-
* Trademark
-- 5 --
.~
- -

-- 1 3 ~
3-one, and 0.26 weight percent methyl-4-isothiazolin-
3-one. Dearborn 6012 also contained a plant extract
with saponins which was not believed to influence
biocidal effectiveness.
A mixed bacterial population was cultivated on a
tryptone glucose extract agar plate which had been
innoculated with 50 microliters of industrial cooling
water. The bacteria was harvested and suspended in tap
water which had been dechlorinated by passage through a
carbon filter.
The biocidal effectiveness of isothiazolones over
time in the presence of mild steel was tested by adding
100 milliliters (mls) of solution containing 100 parts
per million (ppm) Kathon 886 to sterile plastic
containers, with and without a mild steel corrosion
coupon. The containers were then incubated. After the
containers containing isothiazolone solution had been
incubated for two days, 1.0 ml aliquots of the
bacterial suspension were distributed into each
container. The containers were further incubated for
six days. The comparitive amount of living bacteria in
each container was then measured by filtering a one ml
sample from each container, and measuring the
micro-organisms retained on the filter by extracting
and analyzing adenosine triphosphate by the firefly
luciferase assay method. As described in Standard Test
Method for Adenosine Triphosphate (ATP) Content of
Microorganisms in Water, Designation D4012-81 (1981,
reapproved 1985), American Society for Testing and
Materials (ASTM), the firefly luciferase assay method
measures the luminescence produced by luciferase
reagent in the presence of adenosine triphosphate, a

- 1 334824
compound which can be related to viable biomass, or
metabolic activity. The degree of luminescence is
dependent upon the ATP present and can be measured
through the use of a photometer such as the one used in
these examples which is an ATP photometer produced by
SAI Technology Co. of San Diego, California. It is
well-known that when bacteria are killed their ATP
content decreases. Thus, the extent of reduction in
ATP in a biocide-treated sample is a measure of the
degree of biocide effectiveness. The detection limit
of the ATP assay used ranged from about 0.01 nanograms
per milliliter (ng/ml) of sample to about .001 ng/ml
depending upon the amount of sample filtered, the
volume of ATP extract, and the specific activity of the
luciferase reagent used in the analysis.
Parallel runs were made using 1000 ppm of Dearborn
6102 instead of the Kathon 886, and a control run was
made using no biocide. The results of these runs are
presented in Table I below:
TABLE I
ISOTHIAZOLONE PRESENCE OF MILD STEEL ATP AFTER
TREATMENT CORROSION COUPON 6 DAYS (ng/ml)
11.2 ppm (1) No 0.013
11.2 ppm (1) Yes 0.16
11.2 ppm (2) No 0.02
11.2 ppm (2) Yes 0.4
None (control) No 0.28
(1) Added as 100 ppm Kathon 886
(2) Added as 1000 ppm Dearborn 6102

8~
EXAMPLE II
This example used the same general procedures as
Example I, modified as follows: 1000 ppm of ferric
oxide (Fe2O3) was added as a generally insoluble powder
to sterile plastic containers, with and without
Kathon 886 (50 ppm). A bacterial suspension was added
to each container three days later. After a further
three day incubation, the ATP assays were performed.
The results of these runs are presented in Table II
below:
TABLE II
TREATMENT ATP AFTER 3 DAYS (ng/ml)
5.6 ppm Isothiazolones <0.01
+ 1000 ppm Fe2O3
1000 ppm Fe2O3 1.1
EXAMPLE III
This example used the same general procedures as
Example I, modified as follows: A mild steel corrosion
coupon, a stainless steel corrosion coupon, and steel
wool were added to separate sterile plastic containers,
each with 1000 ppm Dearborn 6102. A mild steel
corrosion coupon run was also made without the biocide.
A bacterial suspension was added to each container
three days later. After a further three days
incubation, the ATP assays were performed. The results
of these runs are presented in Table III below:

, ~
`- 1 334824
TABLE III
TEST DOSAGE OF ATP AFTER
METAL ISOTHIAZOLONES 3 DAYS (ng/ml)
Mild Steel Corrosion
Coupon None 3.3
Mild Steel Corrosion
Coupon 11.2 ppm 0.33
Stainless Steel
Corrosion Coupon 11.2 ppm 0.057
Steel Wool 11.2 ppm 1.6
*The one-half inch steel wool strips used for these
experiments were cut from a pad of #0000 grade steel
wool marketed by Thamesville Metal Products Limited
of Thamesville, Ontario under the name Bull Dog*Brand.
EXAMPLE IV
This example used the same general procedures as
Example I, modified as follows: Aluminum corrosion
coupons were added to sterile plastic containers, with
and without Dearborn 6102 (1000 ppm). A bacterial
suspension was added to each container one day later.
After a further 11 days incubation, the ATP assays were
performed. A control run was also made without biocide
~ and without coupons. The results of these runs are
presented in Table IV below:
* Trademark
, J",,!~

1 33~824
TABLE IV
ISOTHIAZOLONE PRESENCE OF ALUMINUM ATP AFTER
TREATMENT CORROSION COUPONS 11 DAYS (ng/ml)
11.2 ppm No 0.09
11.2 ppm Yes 0.07
None (control) No 1.0
The kinetics of the reduction of biocidal activity
was examined in the following example.
EXAMPLE V
100 ml of carbon-filtered tap water were added to
a series of six sterile whirl pack plastic bags.
One-half inch strips of steel wool were placed in five
of the bags, along with 500 ppm Dearborn 6102. The
sixth bag was a control. The steel wool strips were
removed from the bags after various exposure time
intervals ranging from 10 minutes to 24 hours. When
the last strip was removed a bacterial suspension,
prepared as in Example I, was added to each of the six
bags. After a one-day incubation, the ATP assays were
conducted. The results of these runs are presented in
Table V below:
-- 10 --

1 334~24
TABLE V
DURATION OF EXPOSURE DOSAGE OF ATP AFTER
TO STEEL WOOL ISOTHIAZOLONES ONE DAY (ng/ml)
None (control) NONE 1.3
10 minutes 5.6 ppm 0.11
] hour 5.6 ppm 0.11
3 hours 5.6 ppm 0.13
5 hours 5.6 ppm 1.9
24 hours 5.6 ppm 2.1
The effect of iron surface area upon the reduction
of biocidal activity was monitored in the following
example.
EXAMPLE VI
Iron dust (minimum assay: 95% iron) was
distributed in various amounts ranging from 0.01 grams
to 1.0 grams into a series of 250 ml Erylenmeyer
flasks. 100 ml of carbon-filtered tap water containing
1000 ppm of Dearborn 6102 were added to each flask, and
to a control flask to which no iron had been added.
The flasks were placed overnight on orbit shaker
operating at 205 rpm. A bacterial suspension, prepared
as in Example I, was then added to each container.
After a further two-day incubation with shaking, the
ATP assays were performed. The results of these runs
are presented in Table VI below:

-
1 334824
TABLE VI
AMOUNT OF IRON DOSAGE OF ATP AFTER
DUST (gm) ISOTHIAZOLONES TWO DAYS (ng/ml)
None 11.2 ppm 0.077
0.01 11.2 ppm 0.088
0.05 11.2 ppm 0.051
0.1 11.2 ppm 0.047
0.5 11.2 ppm 0.81
1.0 11.2 ppm 0.32
In sum, the Examples above demonstrate that the
biocidal effectiveness of isothiazolones can be
substantially reduced by exposure to iron metal,
whether it be in the form of mild steel, steel wool, or
iron dust. However, no substantial detrimental affect
on biocidal effectiveness was evident by contact with
stainless steel. The term "iron metal" is used in this
specification to signify the forms thereof which do
detrimentally affect organic biocides, and
consequently, does not include stainless steel.
The invention described herein provides for
protection of biocidal activity of isothiazolones when
iron metal is present. The general benefits achieved
by using various protectors is illustrated in the
non-limiting examples numbered VII through XIII which
follow.
EXAMPLE VII
100 ml of carbon-filtered tap water were added
along with 1000 ppm Dearborn 6102 to a series of

1 3 3 ~
sterile plastic containers. Sodium molybdate (NaMoO4.2H2O)
was added to two containers at dosages of 1000 ppm and
3000 ppm respectively, while the third container
contained no molybdate. A fourth container having
neither biocide nor molybdate was used as a control. A
mild steel corrosion coupon was added to each
container. After two days a bacterial suspension,
prepared as in Example I, was added to each container.
After an additional four-day incubation, ATP assays
were conducted. The results of these runs are
presented in Table VII below:
- TABLE VII
ISOTHIAZOLONE PRESENCE OF MILD NaMoO .2H2O ATP AFTER
TREATMENT STEEL CORROSION COUPON AD~ED 4 DAYS (ng/ml)
11.2 ppm Yes None 0.72*
11.2 ppm Yes 1000 ppm 0.12
11.2 ppm Yes 3000 ppm 0.015
None (control) Yes None 0.90
*Significant corrosion observed. It is noted that corrosion
occurred in many samples listed herein. It was also observed,
however, that corrosion was not a prerequisite to biocide
protection. Consequently, the degree of corrosion observed
is not generally reported.
It is evident that in the samples treated with
molybdate, the biocide remains effective even in the
presence of iron. In another experiment using only 100
ppm sodium molybdate (data not shown), biocide
effectiveness was not achieved and corrosion was
evident.

- ` 1 334~24
Zinc, which like molybdate is a well-known agent
used in water treatment to prevent corrosion, was also
tested as a biocide protector.
EXAMPLE VIII
100 ml of carbon-filtered tap water were added
along with mild steel corrosion to a series of sterile
plastic containers. 200 ppm zinc sulphate (ZnSO4.H2O)
were added to the containers, with and without Kathon
886 ~100 ppm). For comparison, 1000 ppm sodium
molybdate were added to other containers, again with
and without Kathon 886 (100 ppm). A control was run
with neither biocide nor protector, but with a mild
steel coupon. After three days incubation a bacterial
suspension, prepared as in Example I, was added.
After an additional eight-day incubation, the ATP
assays were conducted. The results of these runs are
presented in Table VIII below:
TABLE VIII
ISOTHIAZOLONE PRESENCE OF MILD STEEL TEST BIOCIDE ATP AFTER 8
TREATMENT CORROSION COUPONPROTECTOR DAYS (ng/ml)
11.2 ppm Yes 1000 ppm <0.01
4 20
None Yes 1000 ppm 1.0
4 2
11.2 ppm Yes 200 ppm (0.01
Z SO4. 2
None Yes 200 ppm 0.1
Zns4 H2
None Yes None 0.58

- 1
1 334824
It is evident from Table VIII that zinc sulphate
was effective in protecting the biocidal activity of
isothiazolones. However, in contrast to the molybdate
treatment, significant corrosion occurred in the
zinc-treated samples. This may indicate a different
mechanism of isothiazolone protection from that
achieved with molybdates.
The effect of zinc sulphate concentration on the
isothiazolone biocidal activity in the presence of iron
was also monitored.
EXAMPLE IX
100 ml of carbon-filtered tap water were added
along with a mild steel corrosion coupon to a series of
sterile plastic containers. 50 ppm Kathon 886 was
added to all but a control container, and zinc sulphate
(ZnSO4.H2O) was added to the containers in varying
concentrations ranging from 50 ppm to 200 ppm. Runs
with Kathon 886, but without zinc sulphate, were also
made. After one day a bacterial suspension, prepared
as in Example I, was added to each container. After
further incubation ranging from two days to nine days,
the ATP assays were performed. The results of these
runs are presented in Table IX below:

- ` 1 334824
-
TABLE IX
DOSAGE OF PRESENCE OF DOSAGE OF ATP (ng/ml) AFTER
ISOTHIAZOLONES CORROSION COUPON S 1--2 2 days/5 days/9 days
5.6 ppm Yes None 0.064 0.28 0.68
5.6 ppm Yes 50 0.030 0.050 0.27
5.6 ppm Yes 100 0.023 0.024 0.020
5.6 ppm Yes 150 0.023 0.018 0.015
5.6 ppm Yes 200 0.023 0.016 <0.010
None Yes None 2.3 1.9 2.1
It is evident from Table IX that where retention
of biocidal activity in the presence of iron is desired
for more than one week, more than 50 ppm of zinc
sulphate monohydrate should be added.
As shown in Examples X, XI, XII, and XIII below, a
wide variety of water treatment chemicals were tested
for suitability as biocide protectors.
EXAMPLE X
100 ml of carbon-filtered tap water was added
along with a mild steel corrosion coupon to each of a
series of sterile plastic containers. Chemicals to be
tested for their protective qualities were added to the
containers in the amounts shown in Table X, and
isothiazolones (as 1000 ppm of Dearborn 6102) was added
where shown in the table. Control runs without a
coupon were also run with and without biocide. After
three days a bacterial suspension, prepared as in
Example I, was added to each container. After a
- 16 -

~ 1 334824
further three-day incubation, ATP assays were
performed. The results are shown in Table X below:
- 17 -

`' 1 334824
TABLE X
PRESENCE OF
MILD STEEL DOSAGE OF ATP (ng/ml)
DOSAGE OF CORROSION TEST BIOCIDE TEST BIOCIDE AFTER 3
ISOTHIAZOLONES COUPON PROTECTOR PROTECTOR DAYS
None No None None 3.8
None Yes None None 3.1
11.2 ppm No None None 0.1
11.2 ppm Yes None None 4.9
None Yes 4' 2 200 ppm 2.1
11.2 ppm Yes SO4 2 200 ppm 0.12
None Yes (CH3COO)2Zn.2H2O 280 ppm 2.0
11.2 ppm Yes (CH3COO)2Zn.2H2O 280 ppm 0.24
None Yes Zinc Dust 75 ppm 2.4
11.2 ppm Yes Zinc Dust 75 ppm 0.18
None Yes Na2SiO3.9H2O1000 ppm 1.9
11.2 ppm Yes Na2SiO3.9H2O1000 ppm 1.9
None Yes Al2(SO4)3100 ppm 6.5
11.2 ppm Yes Al2(SO4)3100 ppm 2.5
None Yes NiS04.6 2110 ppm 3.5
11.2 ppm Yes i 4. 2 110 ppm 1.9
None Yes K2CrO4 300 ppm 1.8
11.2 ppm Yes K2CrO4 300 ppm 0.06
None Yes SnCl2 160 ppm 4.1
11.2 ppm Yes SnCl2 160 ppm 3.8
None Yes Co(NO3)2.6H2O100 ppm 2.2
11.2 ppm Yes Co(NO3)2.6H2O100 ppm 3.3
11.2 ppm Yes Na2B4O7.5H2O600 ppm 2.8
None Yes Cd(NO3)2 4H2)150 ppm 1.9
11.2 ppm Yes Cd(NO3)2 4H2O)150 ppm 0.22
None Yes KH2A2O4 50 ppm 2.9
11.2 ppm Yes KH2A2O4 50 ppm 2.3
11.2 ppm Yes MgSO4 500 ppm 3.2
11.2 ppm Yes MnSO4. 2 60 ppm 3.1

1 334824
EXAMPLE XI
Additional chemicals were tested in another series
of runs conducted in accordance with the procedure used
in Example X. The results of these runs are shown in
Table XI below:
TABLE XI
PRESENCE OF
MILD STEEL TEST DOSAGE OF ATP (ng/ml)
DOSAGE OF CORROSION BIOCIDE TEST BIOCIDE AFTER 3
ISOTHIAZOLONES COUPON PROTECTOR PROTECTOR DAYS
None No None None 1.2
15 None Yes None None 3.0
11.2 ppm Yes None None 2.0
11.2 ppm No None None 0.09
11.2 ppm Yes 4 2200 ppm 0.14
None Yes ZnSO4 H2O200 ppm 1.9
11.2 ppm Yes Pb(NO3)2 30 ppm 1.8
None Yes Pb(NO3)2 30 ppm 0.9
None Yes CuSO4 50 ppm 0.7
11.2 ppm Yes CuSO4 S0 ppm 0.06
11.2 ppm Yes EDTA (1)100 ppm 2.04
None Yes EDTA (1)100 ppm 4.6
None Yes BaCl2 150 ppm 2.3
11.2 ppm Yes BaC12 150 ppm 1.8
(1) Ethylene diamine tetracetic acid
-- 19 --

- 1 334824
r
EXAMPLE XII
Other chemicals were tested in yet another series
of runs conducted in accordance with the prodecure used
in Example X. The results of these runs are shown in
Table XII below:
TABLE XII
PRESENCE OF
MILD STEEL TEST DOSAGE OF ATP (ng/ml)
DOSAGE OF CORROSION BIOCIDE TEST BIOCIDE AFTER 3
ISOTHIAZOLONES COUPON PROTECTOR PROTECTOR DAYS
None Yes None None 1.0
11.2 ppm No None None 0.02
11.2 ppm Yes None None 0.64
11.2 ppm Yes Canarad 0515 (1) 10 ppm 0.35
None Yes Canarad 0515 (1) 10 ppm 0.84
11.2 ppm Yes Rodine 95 (2)10 ppm 0.23
None Yes Rodine 95 (2)10 ppm 1.2
11.2 ppm Yes Dequest 2010 (3) 100 ppm 0.63
None Yes Dequest 2010 (3) 100 ppm 1.9
11.2 ppm Yes K2CrO4 100 ppm 0.04
None Yes K2CrO4 100 ppm 0.37
11.2 ppm Yes K2CrO4 50 ppm 0.64
None Yes K2CrO4 50 ppm 0.90
11.2 ppm Yes K2CrO4 25 ppm 1.1
(1) Ethoxylated rosin amine, 15%, marketed by Diamond Shamrock, Canada,
Ltd. of Hamilton, Ontario.
(2) Proprietary Blended Corrosion Inhibitor, containing detergent and30marketed by Amchem Products, Inc. of Ambler, Pennsylvania.
(3) Hydroxyethylidine diphosphonic acid, 60%, marketed by Monsanto ofSt. Louis, Missouri.
- 20 -

~ 334~24
EXAMPLE XIII
Several more chemicals were tested in still
another series of runs conducted in accordance with the
procedure used in Example X. The results of these runs
are shown in Table XIII below:
- 21 -

1 334824
~ U~
e~: ~ O a~ .
o o In~ oo o ~~1
~ ,., . . . . . . . . .
--~ O O O ~ O ~ O
P~
P~
~n
o ..
'J
O P~
H--
OE~ O ~ e e o
o ~ a) a) a) ~ ~ ~ Q ~ ~
~: ~ O O O O O O O O O
C~ ~ Z Z Z ~U~ O O
V 0
o ~
`J
a) a) 5~
H - ~ ~; a) ~ a) ~ a) `I a) Cl O a)
H 'j -I ~: ~ S
o ~ a ~ ~ a
H ~a)G~ a) ~ e ~ e ~ e ~ e , ~ x
O O O O~ ~
ir E~ ~;Z Z Z ~ ~ ~ e
VJ ~ U 1~ 0 1~ C) ~ ~ ~
a) a)~D
a
N
J~ Z
O
H )U~ a UiUl U~
~: oa) a) o a~a) a)a) a) a
~ ~ z ~ ~ ~ ~ ~ ~ ~
~ z i~
oo
- ~
~ n
c~ c ~
z ~ ~
-~
no c
Y'
o
~V~ e e e e e
Z
C~ ~ ~ Q~ ~ ~ b~
~ c a) ~ ~ ~ a) ~ a) ~ a~
o ~
o ~ ~ ~ o ~ o ~ o ~
~: - Z ., ~ ~ Z ~ Z ~ Z a
I ~
~ - ~
v. o
o u~
G ~ ~c
-- 22 --

- -. 1 33~2~
Some of the chemicals tested above as protectors
changed the pH of the water. In those cases where the
pH was substantially altered from 7.5, sodium hydroxide
or hydrochloric acid was added to ad~ust the pH to 7.5.
It is evident from the Examples above that sodium
molybdate, zinc sulphate, zinc acetate, zinc dust, zinc
dibenzylcarbamate, the zinc-phosphonate formulation of
Dearborn 909, potassium chromate, cadmium nitrate,
copper sulphate, and diethylthiourea were clearly
effective in protecting biocidal activity in the
presence of iron metal. The effectiveness of Canarad*
0515, Dequest*2010, and Rodine*95 is also evident but
to a somewhat lesser extent.
~ From the foregoing results, it is also concluded
that protective activity will be generally demonstrated
by other molybdates, chromates, zinc salts, copper
salts, cadmium salts, dialkylthioureas, alkoxylated
rosin amines, and phosphonates or a mixture of agents
which effectively protect biocidal activity.
Preferably, the biocide protectors are water soluble to
achieve effective dispersion throughout the aquoues
fluid containing the biocide. However, the zinc dust
results demonstrate that water solubility is not
essential in every case.
As used in this invention, the term "phosphonate
biocide protector" means an organophosphonic acid
having one of the following formulae, A, B, or C:
* Trademark
- 23 -
"~
. ~

FORMULA A 1 3 3 4 8 2 4
HO O R O OH
11 14 1~"~
~P C P
HO 5 m OH
or
FORMULA B
R Y O OH
6~
N C- P
R7 Y' OH
or
FORMULA C
O O
(H)2=PCH2 OH CH2P=(OH)2
N -(CH2- CH-CH2- N)q~CH2PI=(OH)2
(HO)2=PCH2~ O
o
wherein m is an integer from 1 to 10; R4 is hydrogen,
or an alkyl group having from 1 to 4 carbons; R5 is
hydroxyl, amino, hydrogen or an alkyl group having from
1 to 4 carbons; R6 is a member selected from the group
consisting of hydrogen, hydroxyl, hydroxy alkyl groups
containing from 1 to 4 carbon atoms, aliphatic groups
containing from 1 to 30 carbon atoms, and
- 24 -

1 334824
Y O OH
11 ~
- C P ~
Y OH
R7 is a member selected from the group consisting of
hydrogen, aliphatic groups containing from 1 to 30
carbon atoms,
Y O OH Y
C P / and C N - Z
y~ OH ~ Y ~ n
wherein n is an integer from 1 to 30; Y and Y' are
members selected from the group consisting of hydrogen
and lower alkyl groups containing from 1 to 4 carbon
atoms; Z is a member selected from the group consisting
of hydrogen and
Y O OH
- C- P
Y OH
and Z' is a member selected from the group consisting
of hydrogen,
Y O OH Y Y O OH
C p < and - (C)n N C7 U"~
Y' OH Y' Z P y~ OH
wherein p is an integer from 1 to 30; with at least one
of the groups represented by R6 and R7 containing at
least one
- 25 -

1 334824
Y O OH
11
- C P~
Y OH
group; q is an integer from 1 to 10; and the water-
soluble salts and esters thereof; or mixtures thereof.
The preferred organophosphonic acid compound for
use in this invention is an alkylene diphosphonic acid
having the foregoing Formula A, such as those disclosed
in U.S. Pat. Nos. 3,214,454 and 3,297,578, the entire
disclosures of which are incorporated herein by
reference. Also suitable is an organophosphonic acid
having the foregoing Formula B such as those disclosed
in U.S. Pat. No. 3,298,956, the entire disclosure of
which is incorporated herein by reference. Suitable
acids of this type include methylenediphosphonic acid;
ethylidenediphosphonic acid; isopropylidenediphosphonic
acid; 1-hydroxy, ethylidenediphosphonic acid;
hexamethylenediphosphonic acid; trimethylenediphosphonic
2d acid; decamethylenediphosphonic acid; 1-hydroxy,
propylidenediphosphonic acid; 1,6-dihydroxy,
1-6-dimethyl, hexamethylenediphosphonic acid;
1,4-dihydroxyl, 1,4-diethyl, tetramethylenediphosphonic
acid; 1,3-dihydroxy 1,3-dipropyl, trimethylenediphos-
phonic acid; 1,4-dibutyl, tetramethylenediphosphonic
acid; dihydroxy, diethyl, ethylenediphosphonic acid;
4-hydroxy, 6-ethyl, hexamethylenediphosphonic acid;
l-hydroxy, butylidenediphosphonic acid; butylidene-
diphosphonic acid; 1-aminoethane-1,1-diphosphonic acid;
1-aminopropane-1,1-diphosphonic acid; 1-aminoethane-1,1-
- 26 -

-` 1 334824
diphosphonic acid monoethyl ester, amino tri(methyl
phosphonic acid), amino tri(ethylidene phosphonic
acid), amino tri(isopropylidene phosphonic acid), amino
tri(butylidene phosphonic acid), amino tri(isopen-
tylidene phosphonic acid, ethylene diamine tetra(methyl
phosphonic acid), ethylene diamine tri(methyl
phosphonic acid), ethylene diamine di(methyl phosphonic
acid), hexamethylene diamine tetra(methyl phosphonic
acid), diethylene triamine penta(methyl phosphonic
acid), N-(2-hydroxy-ethyl) nitrilo N,N-di(methyl
phosphonic acid), and 2-hydroxy propylene 1,3-diamine
tetra(methvl phosphonic acid). The water-soluble salts
of these acids such as the alkali metal, alkaline earth
metal, zinc, cobalt, lead, tin, nickel, ammonium, or
amine and lower alkanol amine salts can be used. Also,
esters of these acids with an aliphatic alcohol having
from 1 to 4 carbons, or mixtures of the above acids,
salts or esters can be used. Use of mixtures of any of
the general types of organophosphonic acid compounds
described above is also contemplated within the scope
of this invention. Hydroxy ethylidenediphosphonic acid
is particularly preferred.
The foregoing results also demonstrate that the
amount of each substance which is efficiently added can
vary according to the nature and amount of iron metal
present in the system. In general, however, the
following dosages are preferred. Where molybdates are
used as the sole protective agent at least about 150
ppm is used, with the preferred concentration range
being from about 150 ppm to about 10,000 ppm; most
preferably from about 500 ppm to about 3,000 ppm.
- 27 -

`- 1 334824
Where chromates are used as the sole protective agent
at least about 20 ppm is preferably added, with the
most preferred range being from about 40 ppm to about
300 ppm. Where zinc salts are used as the sole
protective agent at least about 10 ppm is used, with
the preferred concentration range being from about 10
ppm to about 2000 ppm; and most preferably from about
30 ppm to about 100 ppm. Where copper salts are used
as the sole protective agent at least about 2 ppm is
preferably used, with the preferred range being between
about 20 ppm and about 100 ppm. When cadmium salts are
used as the sole protective agent at least about 5 ppm
is preferably added, with the most preferred range
being between about 25 ppm and about 250 ppm. Where
dialkylthioureas are used as the sole protective
agents, at least about 1 ppm is preferably added, with
the most preferred range being between about 5 ppm and
about 100 ppm. Where alkoxylated rosin amines are used
as the sole protective agent, at least about 0.15 ppm
is preferably added, with the most preferred range
being between about 1 ppm and about 100 ppm. Where
phosphonate biocide protectors are used as a sole
protective agent at least about 5 ppm is preferably
added, with the most preferred range being between
about 25 ppm and about 500 ppm. Where zinc dust is
used as the sole protective agent at least about 5 ppm
is preferably added, with the most preferred range
being between about 50 ppm and about 500 ppm.
The addition of these protective agents may be
made separately or together with the organic biocide.
Indeed, various compositions containing the biocide in
- 28 -

- ` 1 334824
combination with biocide protectors are within the
scope of this invention. Weight ratios of total
protectors to biocide between about 0.1:1 and about
30,000:1 are generally preferred. Representative
compositions exemplified by the above Examples comprise
isothiazolones and, alternatively, molybdates;
chromates; zinc salts; dialkylthioureas; alkoxylated
rosin amines; phosphonate biocide protectors; and zinc
dust. In these compositions the most preferred weight
ratio range of molybdate to isothiazolone is from about
50:1 to about 30,000:1; that of chromate to
isothiazolone is from about 4:1 to about 3,000:1; that
of zinc salts to isothiazolone is from about 1:1 to
about 20,000:1; that of dialkylthiourea to
isothiazolone is from about 0.5:1 to about 1,000:1;
that of ethoxylated rosin amine to isothiazolone is
from about 0.1:1 to about 1,000:1; that of phosphonate
biocide protectors to isothiazolone is from about 2.5:1
to about 5,000:1; and that of zinc dust to
isothiazolone is from about 5:1 to about 5,000:1.
Mixtures, of course, may contain at least
proportionately lower ratios of each protector to
biocide. These compositions may be added to the
aqueous fluid in dry form such as powder or pellets,
where feasible. Such dry compositions preferably
contain between 0.1 weight percent and 100 weight
percent total active biocide plus protector, with the
weight ratio of the components being that identified
above.
_ 29 -

- 1 334824
The compositions may also be added as aqueous
solutions. Such aqueous solutions are preferably
containing between about 0.1 and 50 weight percent
total active biocide plus protector. The biocide
concentration after addition is preferably kept within
its normal effective range; generally for organic
biocides between about 0.1 ppm and about 10,000 ppm.
A mixture of the protective agents discussed above
can be used advantageously. Indeed, many of the
effective biocide protectors are also corrosion
inhibitors; and it is well known that some combinations
of corrosion inhibitors, such as zinc and phosphonates,
work together synergistically to prevent corrosion.
Synergism in protecting biocide effectiveness in the
presence of iron was tested in the following
non-limiting example.
EXAMPLE XIV
100 ml of carbon-filtered tap water was added
along with a mild steel corrosion coupon to each of a
series of sterile plastic containers. As shown in
Table XIV below, zinc sulphate (ZnSO4.H2O) was added to
some of the containers in various concentrations
ranging from 50 ppm to 100 ppm; and Dequest 2010
(containing l-hydroxyethylidene-1,1-diphosphonic acid)
was added to some of the containers in various
concentrations ranging from 25 ppm to 100 ppm.
Isothiazolones (as 1000 ppm Dearborn 6102) were also
added where shown in the Table. Control runs without a
coupon, without a protector, and without either biocide
- 30 -

1 334824
or protector were also run as shown. After three days
a bacterial solution, prepared as in Example I, was
added to each container. After a further three-day
incubation, ATP assays were performed on some of the
containers. As seen in the Table, it was evident that
the biocide was relatively effective in all the samples
and synergism would be difficult to observe. Thus, a
second dosage of bacterial solution was added on the
day following the initial ATP assays. After an
additional nine-day incubation, ATP assays were
repeated on the samples. In some cases an ATP assay
was also conducted 11 days after the second bacterial
dosage. The results are shown in Table XIV below:

1 334824
~_
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--u~ao
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~f~O
E~ a c~
f~ ~
a~ _
f~ r
~ 1~ J El O ~ ~1 (~ ~1 ~ ~1 0 ~ ~) ~1 ~ ~1
--E~ fI ~ u~ o ~D ~D ~1 0 ~ o o O ~1 o 1~ 0 ~ O ~ o ~ o 1--o t`l
~a f -------...............
~f~ ~ ~0~0~0~0~0~0~0000000~
-- a~
u, z a
~ ~ o
~fU
f~ a ~
U~
-
o ~ a~ o
o o:) o r--o ~ o u~ o o
~f~ ~000~0~0~000
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f~~ 0000000~00~00~oO~OO
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~n ooooooooooooooooooooooo
Z Z Z ~ U~ O O O O O O O O O O
a~ _
a~ z
H O
~n
~ O
O P~
1~; Z
~, ~, 2
E~ O
4~n ~
~n ~ O
Z Q ~ Q Q Q~ Z
~I O ~ I ~1 0 ~ O ~ O _I O _I O _I O -1 0 ~1 0 _I O _I O ::~
~, z~z~z-lz-~z-lz,læ,~z,~z-lz~z ~
fl-
~ f~
O ~1:
a-
-- 32 --

~ 1 334824
,
It is evident from the Table, and the comparative
low level of protection of Dequest 2010 evident from
Table XII, that synergism in biocide protection does
occur between zinc sulphate and phosphonate. Thus, as
little as 25 ppm of Dequest 2010 used in combination
with as little as 50 ppm zinc sulphate, afforded
substantial protection of the biocidal activity of
isothiazolones in the presence of mild steel over
comparatively long periods of time. Preferably, at
least 10 ppm of zinc sulfate is combined with at least
5 ppm 1-hydroxyethylidene-1,1-diphosphonic acid.
Compositions containing zinc sulphate in a weight ratio
to isothiazolone of about 0.5:1 to about 1,000:1, and
1-hydroxyethylidene-1,1-diphosphonic acid in a weight
ratio to isothiazolone of about 0.25:1 to about 1,000:1
are preferable for use in this synergistic manner.
While the invention herein described may be
utilized in many aqueous systems such as cooling
towers, pulp and paper mill process water, oil field
water treatment, and preservatives for various
industrial products, it is believed that the invention
is particularly suited for application in aqueous
systems with long retention times, such as in metal
working systems. Bacteria tend to accumulate in these
systems and the obvious presence of iron in some of
these applications can threaten the effectiveness of
isothiazolone biocide.
To test the protectors in aqueous metal working
fluids, a sample of cutting oil fluid containing
endogenous bacteria was obtained from a General Motors
transmission plant in Windsor, Ontario Canada. The

334824
protection afforded metal working fluids by practice of
this invention is illustrated by the non-limiting
examples numbered XV and XVI which follow.
EXAMPLE XV
100 ml of cutting oil fluid were added to each of
a series of sterile plastic containers. As shown in
Table XV below, a mild steel corrosion coupon was added
to some of these containers; and 200 ppm zinc sulphate
(ZnSO4.H2O) was added to some of the containers. As
also shown in the Table, isothiazolones were added to
some of the containers in various amounts (ranging from
200 ppm to 1000 ppm Dearborn 6102.) After eight days
of incubation, ATP assays were performed. The assays
showed that the biocide was functioning except where
used in low dosages without protector. Another ATP
assay was performed on same samples after 15 days of
incubation. The biocide evidently functioned longer in
the cutting oil fluid than in tap water. The
biocide-treated samples were thus re-innoculated after
16 days with 0.5 ml of an untreated cutting oil fluid
which contained 10 ng/ml ATP. Additional ATP assays
were then performed on some samples after 22 days, 26
days, and 31 days. The results are shown in Table XV
below:
- 34 -

TABLE XV
ISOTHIAZOLONE PRESENCE OF DOSAGE OF ATP (ng/ml) AFTER:
DOSAGE CORROSION COUPON ZnSOq.H2O8 days15 days 22 days26 days 31 days
None No No 4.2 6 0 8 5 * *
None Yes No 3.5 * * * *
None No 200 ppm 13.0 21 20 * *
None Yes 200 ppm 5.0 * * * *
2.24 ppm No No 2.3 * * * *
2.24 ppm Yes No 3.2 * 4 5 * *
2.24 ppm No 200 ppm(0.01 ~0.01 * * *
I 2.24 ppm Yes 200 ppm<0.01 * 16.5 * *
w 5.6 ppm No No <0.01 * * 17,0 *5.6 ppm Yes No ~0.0l. ~0.01 13.0 9.8 *
1 5.6 ppm No 200 ppm<0.01 ~0.01 * 20 *5.6 ppm Yes 200 ppm<0.01 * 4.1 50 *
11.2 ppm No No ~0.01 * 0.45 * 25
: 11.2 ppm Yes No ~0.01 ~0,01 2.4 28 100
11.2 ppm No 200 ppm<0.01 ~0.01 0.01 * 0.43
11.2 ppm Yes 200 ppm~0.01 * O,07 1.0 20
- * Analysis Not Done
, r~
,
-

1 334824
It is evident from the Table that the presence of
iron can reduce the duration of biocidal effectiveness
in cutting oil fluids. The presence of 200 ppm zinc
sulphate monohydrate increased the length of biocidal
effectiveness of every dosage of Dearborn 6102 tested.
The length of effectiveness was increased even when
corrosion coupons were not added to the test samples.
However, because particles of iron were present in the
cutting fluid sample as it was received, the same
mechanism of protection of biocidal activity was
evidently occurring in each sample. In any case, the
samples containing corrosion coupons appeared to lose
biocidal activity more quickly.
A variety of chemicals were tested for use as
protectors in metal working fluids using commercial
cutting oil stock.
EXAMPLE XVI
A stock cutting oil was diluted to 6% in tap
water, and 100 ml of the diluted stock cutting oil
fluid was added to each of a series of sterile plastic
containers. Chemicals to be tested for their
protective qualities were added to the containers in
the amounts shown in Table XVI. One-half inch strips
of steel wool were placed in each of the containers
along with Dearborn 6102 (either 500 ppm or 1000 ppm).
Control runs were also made without biocide, without
protector, and without iron. After two days, S ml of a
microbiologically contaminated cutting oil sample from
the General Motors transmission plant in Windsor was
- 36 -

1 334824
added to each container. ATP assays were performed
weekly for four weeks and no growth in any of the
biocide-treated samples was observed. After each ATP
assay, an additional dosage of contaminated cutting oil
was added. After four weeks, the steel wool strips
were removed and replaced by mild steel corrosion
coupons, and the dosage of cutting oil containing
micro-organisms was repeated. The results of the assay
two weeks after the corrosion coupons were introduced
are given for Table XVI.

1 334824
o
x
Z
o
H 3
E~
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C~
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H
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D O ~r> O
~ 8 u~ ~ ~ o ~. o ~ ~ a~ o ~
X IL~ 3 ~r N O O O O ~ a~ 11') ~C) CO H ~1 ~ ~ ~r O ~ O
~ U~
3 2
O
~ ~ E~
--:~ U~
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0 3
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,¢ U 4 E~ C4 P~ 4
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H ~ O O O O O O O O 11'1 If) O O O O O
O O O O O O 10 U~
X ~:t Z Z Z Z
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a o .. ~ ... .... ....... ..
: -- Z u) ~ ~ u~ ~ u~ u
C ~
a ~
~ 38 --

1 334824
No growth in any of the biocide treated samples was
observed until after the corrosion coupons had been
added.
This Example also demonstrates that benzotriazole
can be an effective protector of biocide effectiveness.
From these results, it is concluded that other suitable
azoles may be selected from a group consisting of
triazoles, pyrazoles, imidazoles, isoxazoles, oxazoles,
isothiazoles, thiazoles, and mixtures thereof as
disclosed in U.S. Pat. Nos. 2,618,608, 2,742,369, and
2,941,953. As used in this invention, the term "azole
biocide protector" means an azole having one of the
following Formulae D, E, F, G, H, I, or J, including
the indicated substituted forms thereof.
The triazoles which can be employed in this
invention are water-soluble 1,2,3-triazoles such as
1,2,3-triazole itself or a substitute 1,2,3-triazole
where the substitution takes place in either the 4 or 5
position (or both~ of the triazole ring, as shown here
by the structural Formula D:
FORMULA D
NH
~\
4 3~
- 39 -
!

` 1 334824
Suitable triazoles include benzotriazole (the preferred
triazole); 4-phenyl-1,2,3-triazole; 1,2-naphthotriazole
and 4-nitrobenzotriazole; and the like.
The pyrazoles which can be used in this invention
include water-soluble pyrazoles such as pyrazole itself
or a substituted pyrazole where the substitution takes
place in the 3, 4, or S position (or several of these
positions) of the pyrazole ring, as shown by the
structural Formula E:0
FORMULA E
NH
/1\
HC5 2-
HC4 3~H
Suitable pyrazoles include pyrazole; 3,5-dimethyl
pyrazole; 6-nitroindazole; 4-benzyl pyrazole; 4,S-
dimethyl pyrazole; and 3-allyl pyrazole; and the like.
Imidazoles which can be used in this invention
include water-soluble imidazoles such as imidazole
2S itself or a substituted imidazole where the
substitution takes place in the 2, 4, or S position (or
several of these positions) of the imidazole ring, as
shown here by the structural Formula F:
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~ --`
- 1 334824
FORMULA F
NH
/1\
HC5 2CH
HC4 3N
Suitable imidazoles which can be employed in this
invention include imidazole; adenine; quanine;
benzimidazole; 5-methyl benzimidazole; 2-phenyl
imidazole; 2-benzyl imidazole; 4-allyl imidazole;
4-(betahydroxy ethyl)-imidazole; purine;
4-methylimidazole; xanthine; hypoxanthene; 2-methyl
imidazole; and the like.
Isoxazoles which can be used in this invention
include water-soluble isoxazoles such as isoxazole
itself or a substituted isoxazole where the
substitution takes place in the 3, 4, or 5 position (or
several of these positions) of the isoxazole ring, as
shown here by the structural Formula G:
FORMULA G
HC5 2
HC 3CH
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1 334824
-
Suitable isoxazoles include isoxazole; 3-mercaptoisox-
azole; 3-mercaptobenzisoxazole; benzisoxazole; and the
like.
The oxazoles which can be used in this invention
include water-soluble oxazoles such as oxazole itself
or a substituted oxazole where the substitution takes
place in the 2, 4, or 5 position (or several of these
positions) of the oxazole ring, as shown here by the
structural Formula H:
1~ FORMULA H
HC5 2~H
HC4 3~
Suitable oxazoles include oxazole; 2-mercaptoxazole;
2-mercaptobenzoxazole; and the like.
The isothiazoles which can be used in this
invention include water-soluble isothiazoles such as
isothiazole itself or a substituted isothiazole where
the substitution takes place in the 3, 4, or 5 position
(or several of these positions) of the isothiazole
ring, as shown here by the structural Formula I:
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i,_,

1 334824
FORMULA I
HC
HC4 3C~
Suitable isothiazoles include isothiazole; 3-mercapto-
isothiazole; 3-mercaptobenzisothiazole; benzisothiazole;
and the like.
The thiazoles which can be used in this invention
include water-soluble thiazoles such as thiazole itself
or a substituted thiazole where the substitution takes
place in the 2, 4, or 5 position (or several of these
positions) of the thiazole ring, as shown here by the
structural Formula J:
FORMULA J
S
/ 1 \
HC5 2CH
H~4 3~
Suitable thiazoles include thiazole; 2-mercaptothiazole;
2-mercaptobenzothiazole; benzothiazole; and the like.
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In the above azole compounds, the constituents
substituted in the azole rings can be alkyl, aryl,
aralkyl, alkylol, and alkenyl radicals so long as the
substituted azole is water-soluble. Typically,
substituted members have from 1 to about 12 carbon
atoms. The triazoles are the preferred azoles, with
benzotriazole and tolyltriazole particularly preferred.
Azoles in general, and benzotriazole in particular, are
preferably used in concentrations of at least 10 ppm
with the preferred range being from about 20 ppm to
about 1,000 ppm. They may be added together with an
isothiazolone in dry or liquid form, as explained
above, with the preferred weight ratio of azole to
isothiazolone being~from about 2:1 to about 10,000:1.
Example XVI also demonstrates effective use of
zinc sulphate in combination with chromate to protect
the effectiveness of isothiazolone. Preferably in such
a combination, at least 10 ppm of zinc sulfate is
combined with at least 10 ppm of chromate; the weight
ratio of zinc sulfate to organic biocide is from about
0.5:1 to about 1,000:1; and the weight ratio of
chromate to organic biocide is about 0.5:1 to about
1 , 000 : 1 .
A test of the influence of ferrous sulphate on the
effectiveness of isothiazolones was also conducted
using the contaminated cutting oil.
EXAMPLE XVII
One gram of ferrous sulphate (FeSO4) was dissolved
in 10 grams of Dearborn 6102 and allowed to incubate
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1 334~24
overnight. Samples of the cutting oil contaminated
with micro-organisms were treated in sterile plastic
containers with either 500 ppm or 1000 ppm of this
solution. As control runs, contaminated cutting oil
samples were run with no treatment, and with treatment
with Dearborn 6102 alone. ATP assays were performed
after a five-hour incubation and after a one-day
incubation. The results are shown in Table XVII below:
TABLE XVII
ATP ( ng/ml)
AFTER 5 HOUR AFTER 1 DAY
TREATMENT INCUBATION INCUBATION
No treatment 10 3.9 -
500 ppm Dearborn 6102 + FeSO4 1.1 0.12
1000 ppm Dearborn 6102 + FeSO4 0.40 0.091
500 ppm Dearborn 6102 0.59 0.12
1000 ppm Dearborn 61020.35 0.11
It is evident from the Table that, although there
may be a slight interaction between the ferrous ion and
the isothiazolones, this interaction is not comparable
at the dosages tested with the influence observed with
the mild steel corrosion coupons.
Other known non-oxidizing type organic biocides
were also studied to determine whether their activity
was reduced in the presence of iron. The effect of
iron on these biocides is illustrated by the following
non-limiting example.

- ` 1 334824
EXAMPLE XVIII
100 ml of carbon-filtered tap water was added to
each of a series of sterile plastic containers. The
biocides to be tested in the presence of iron were
added in the amounts shown in Table XVIII. As shown,
one-half inch strips of steel wool were added to some
of the containers, and steel wool strips together with
200 ppm zinc sulphate (ZnSO4.H2O) were added to other
containers. A control run was made using steel wool
without either biocide or protector. After two days
bacterial solution, prepared as in Example I, was added
to each container. After a further three-day
incubation, ATP assays were performed. The results are
shown in Table XVIII.
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.
TABLE XVI I I
ATP (ng/ml) AFTER 3 DAYS INCUBATION
NO STEEL WOOL STEEL WOOL STEEL WOOL +
BIOCIDE ADDED ADDED200 ppm ZnSO4.H2O
5 None ------- 2.6 --------
15 ppm bis- 0.012 2.0 0.04
trichloro-methyl-
sulphone plus 4.5 ppm
bis-tributyltin-oxide (1)
30 ppm bis- 0.0034 0.038 0.0033
10 trichloro-methyl-
sulphone plus 9 ppm
bis-tributyltin-oxide (1)
10 ppm methylene 0.017 2.8 0.0089
~ bis-thiocyanate (21
20 ppm methylene 0.0032 0.00840.0064
bis-thiocyanate (2)
5 ppm 2,2 dibromo- 0.015 1.8 0.071
3-nitrilopropion-
amide (3)
(1) added as Dearcide*703 marketed by Dearborn Chemical Co.
of Mississauga, Ontario
(2) added as Dearcid~ 709 marketed by Dearborn Chemical Co.
of Mississauga, Ontario
(3) added as Dearcide*723 marketed by Dearborn Chemical Co.
of Mississauga, Ontario
* Trademark
~ ~, L~

1 334824
It is evident from the Table that other organic
biocides can be detrimentally affected by iron and that
they can also be protected from the influence of this
metal. Indeed, the results indicate that a broad range
of organic biocides are succeptible to losing
effectiveness in the presence of iron metal. Whether a
particular organic biocide is detrimetally affected in
untreated fluid by the presence of iron metal can be
simply determined by the test procedures used herein
(eg. Example I). If a loss of effectiveness is
confirmed, the biocide protectors of this invention may
be used advantageously to inhibit the affect of the
iron metal. Of course, compositions of such biocides
with biocide protectors similar to those disclosed
above using isothiazolone may be prepared.
No attempt was made to maintain the same source or
strength of bacteria throughout the examples. The
examples describe particular embodiments of the
invention. Other embodiments will be apparent to those
skilled in the art from a consideration of the
specification or practice of the invention disclosed
herein. It is understood that modifications and
variations may be practiced without departing from the
spirit and scope of the novel concepts of this
invention. It is further understood that the invention
is not confined to the particular formulations and
examples herein illustrated, but it embraces such
modified forms thereof as come within the scope of the
following claims.
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