Note: Descriptions are shown in the official language in which they were submitted.
BACRGROU~D OF l~E INVENTION
1. Field of the Invention
This invention relates to the use of certain hydroxy-substituted esters
of thiolsulfonic acids to control the growth in aqueou~ systems f ~
pneumophila, the causative bacterium for Legionnaires Dlsease. More particu-
larly, the compounds 2~hydroxyethyl methanethiolsulfonate and 2-hydroxypropyl
methanethiolsulfonate are useful as bactericides to inhibit the growth of
Legionella Pneumophila in eommercial and industrial cooling water systems,
cooling towers, evaporative condensers, air-washing systems, and other aqueous
systems in which this pathogenic organism might grow.
2. Description of the Prior Art
~egionnaires Disease is a serious and sometimes fatal pneumonia-like
respiratory illness. The causative organlsm for this disease was identified
after an outbreak of the disease among the people ~ho attended the July 1976
American Legion Convention at the Bellevue-Stratford Hotel in Philadelphia.
; This epidemic resulted in the deaths of 29 of the 182 people who became ill.
After the 1976 epidemic among the Legionnaires who went to Philadelphia, the
I U.S. Public Health Service Center for Disease Control (CDC)~ Atlanta, Georgia,initiated an intensive effort to determine the cause of this respiratory
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illness, which has since become known as "Legionnaires Disease." Subsequently,¦
the CDC identified the causative organism as a gram-negative bacterium to
which they gave the name Legioneila pneumophila. Later investigations by the
ji CDC showed that L. pneumophila ~as not new but could be connected to lllnesses
'I that occurred as far back as the late 1940s. For a review of the epidemiologyof this disease, see Eickhoff, "Epidemiology of Legionnaires' Disease," Ann.
I Inter. Med., Vol. 90, pp. 499-502 (1979).
i In the course of the CDC investigations, L. pneumophila was recovered
- ~' from the water ln cooling towers and evaporative condensers of air-conditioning;
~ or cooling water systems of buildings or insta11ations associated with out-
breaks of Legionnaires Disease. See, for eYample, Eickhoff in the work cited
in the foregoing. The epidemiological data indicated that infection may have
been the result of inhaling aerosols of contaminated water generated from
these sources. ~lthough the exact route of infection has not yet been defi-
nitely confirmed, the CDC deems it prudent to take measures to control or
prevent the growth of this pathogenic organism in the water in cooling water
systems, cooling towers, evaporative condensers, air-washing systems, and
other aqueous systems that might generate aerosols wh~ch may accidentally be
inhaled by people in the vicinity of these systems.
It is well-known that bacteria and other microorganisms grow in cooling
towers and other aqueous systems, causing slime formation, fouling of equip-
ment, corrosion, odors, and other problems. Therefore it,is customary to use
chemical treatments or other techniques in order to control the growth of
these troublesome organisms. However, such control procedures have not
usually been designed to completely eradicate the organisms but only to keep
the microbial growth at a tolerable level where slime and other prpblems are
mlnimal. ~n the other hand, in the case of pathogenic organisms like L.
pneumophila, it is desirable to have treatment chemicals and ~ethods that can
completely eradicate ~hese organisms and still be economically feasible to
apply and safe to use.
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'I Since it first became known that Legionnaires Disease was associated with
I the presence of L. pneumophila in cooling water systems and other aqueous
i~' systems that might generate ~erosols, studies have been made to determine the
1 efficacy of various commercial microbicides commonly used to control other
1l microorganisms in wa~er systems of these types. However, while these products
j have generally been adequate for the de~ree of control required for the
¦l troublesome but usually nonpathogenic organisms that cause slime and operating I
problems, most of those tested cannot provide, at safe and economical use
~ rates, the degree of control required to eliminate L. pneumophila as a hazard
in ~ater. Por most of the commonly used products, the concentration of
chemical required to completely eradicate L. Pneumophila in reasonable contact
times has been several times that required to control other organisms, and the
; cost of treatment at these concentrations has been economically prohibitive.
Other products that could inhibit the growth of this organism at reasonable
lS concentrations have had other disadvantages such as relatively high toxicities
to fish and other aquatic organisms or severe foaming tendencies. These
latter properties make them hazardous to ~he enYironment or cause problems in
operating the cooling systems.
For example, the product poly~oxyethylene (dimethyliminio) ethylene
~dimethyliminio) ethylene dichloride] is useful for controlling the growth of
algae and bacteria in industrial water systems. See, for example, Pera et
al., U.S. Pat. No. 3,771,989, ~here it is reported that 2.0 parts per million
of this microbicide completely inhibited the growth of several algal and
cyanobacterial species commonly encountere~ in cooling water systems. How-
ever, it required 120 parts per million of this microbicide to ob~ain complete
inhibition or complete kill of L. pneumophila in standar~ in vitro tests ~ith
2 hours contact time. See Hollis et al., "Resistance of Legionella pne_ ophila
to Microbicides," Dev. Ind. Microbiol., Vol. 21, pp. 265-271 (1980).
Halogenated amides such as 2,2-dibromo-3-nitrilopropionamide ~DBNPA) are
also useful as antimicrobials in cooling water and other aqueous systems.
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See, for example, Wolf, U.S. Pat. No. 3,6479610, and Wolf et al., "2,2-Dibromo-
3-nitrilopropionamide, a Compound with Slimicidal Activity," Appl. Microbiol.,
Vol. 24, No. 4, pp. 581-584 (1972). DBNPA has demonstrated a higher degree of
lieffectiveness than many other microbicides in inhibiting the growth of L.
l pneumophila, slnce it gave a complete kill with 4 parts per million of the
compound with 2 hours contact time. See Hollis e~ al. in the ~ork cited in
l the foregoing. ~owever, the cost of this compound, which i8 currently manu-
¦ factured and sold by The Dow Chemical Co., Midland, Michigan under the trade-
t~s
~ ¦ n~es DOW Antimicrobial 72B7 (20% active DBNPA) and DOW Antimicrobial 8536 (5%
~ active DBNPA), makes lts use costly even at this concentration. In addition,
this compound like many others used for cooling wa~er treatment has a rela-
tively high degree of toxicity to fish.
It has been demonstrated that L. pneumo~hila is hi~hly susceptible to
free chlorine. See, for example, Skaliy et al., "Laboratory Studies of
Disinfectants Against Legionella pneumophila," Appl. Envir. Microbiol., Vol.
40, No. 4, pp. 697-700 (1980). These workers found that 3.3 parts per million
chlorine from calcium hypochlorite gave a 100% kill of L. pneumophila.
However, as they pointed out, chlorine is especially unstable in the presence
of or~anic matter, and its effects may rapidly be neutralized because of this.
To get the levels of free chlorine necessary to inhibit L. pneumophila under
actual use conditions, excessive amounts of calcium hypochlorite would be
required, which would result in a severely corrosive situation as ~ar as
the materials of construction used in the coolins water system are concerned.
Also effluents with high chlorine residuals could be harmful to the environment.Another microbicide that is reported to be effective in controlling L.
pneumophlla i6 the combination of N-alkyl dimethyl benzyl ammonium chloride
(12.5%) and bis(tri-n-butyltin) oxide (2.25%). See, for example, Grace et al.
"Susceptibility of Leglonella pneumophila to Three Cooling Tower Microbicides,"
Appl. Environ. Microbiol., Vol. 41, No. 1, pp. 233-236 (1981), where it is
reported tha~ 2 parts per million of this combination provides a complete kill
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'of L. Pneumophlla in an aqueous medium with a 3-hour contact time. Quaternary
, am,monium compounds of this type, however, are ~urface active and cause fsam
; I problems in recirculating cooling water systems as well as in many other
Illaqueous systems. Moreover, quaternary ammonium compounds generally have a
, high toxicity to fish, and heavy metal compounds such as the organlc tin
i!
'compound in this combination are not always acceptable-ln effluents going into
' lakes, streams, or other public waters.
- 1 Grace et al. in the work cited in the foregoing also tested another
compound that is widely used as a cooling tower biocide, that is, methylene
bis(thiocyanate). They found that the minimum lethal concentration of this
comp~und against L. pneumophila was 62 parts per million at a 3-hour contact
time, which makes the use of this compound economically prohibitive for the
control of this organism.
In addition to the disadvantages previously outlined for prior art micro-
bicides, these products also all have relatively high degrees of toxicity to
fish. For example, DBNPA, which, as discussed in the foregoing9 shows rela-
tively good effectiveness in inhibiting the growth of L. ~ , has
96-hour LC~o values in rainbow trout and bluegill sunfish in the range of 0.9
to 1.8 parts per million of the compound, according to the "Technical Handbook
for DOW Antimicrobial 7287 and DOW Antimicrobial 8536" published by The Dow
Chemical Co. (1980). This means that concentrations of the compound in this
range cause a 50% kill of these fish species with a 96-hour exposure in static
fish toxicity tests. Of course, since this compound is rapidly degraded and
is not persistent in aqueous systems, it generally causes no fish-kill problems
in actual use. However, where cooling tower water effluents flow, for example,
directly into fish bearing lakes or streams without golng through a waste
treatment system, it would be possible for fish kills to occurl especially
~here this microbicide or others with similarly high fish toxicities must be
used in the higher concentratioDs required for the control of L. pneumophila.
~0 It is, therefore, an object of the present inventiun to provide a method
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I of controlling the growth of L. pneumophila in aqueous ~ystems that obviates
the disadvantages of the prior art metbods. This and oth~r objects, features,
and advantages of the invention will become apparent as the description
l proceeds.
' Summary of the Invention
¦, To the accomplishment of the foregoing and related ends, this invention
Il then comprises the features hereinafter fully described and particularly
! pointed out in the claims, the following description setting forth in detail
certain illustrative embodiments of the $nvention, these beln~ indicative,
however, of but a few of the various ways in which the principles of this
invention may be employed.
In brief, we have discovered that the foregoing objects and advantages
are attained by employing as a bactericide 2-hydroxyethyl methanethiolsulfonate
(HEMTS) or 2-hydroxypropyl methanethiolsulfonate (HPMTS), alone or in combi-
nation with each other or with other microbicides. When these compounds are
used to treat aqueous systems, the biocidal amount providing complete inhibi-
tion (100% kill) of Legionella pneumophila bacteria is much lower than that of
other microbicides used in prior art methods of controlling this organism. In
addition, at least one of these two substances has been shown to be relatively
nontoxic to fish. Moreover, these compounds are not corrosive to mater-lals of
conseruction when used in the low concentrations required for the control of
L. pneumophila.
Detailed Description of the Invention
The bactericides of this invention are compounds o~ the type that may be
described as 2-hydroxyalkyl esters of thiolsulfonic acids. One method for the
preparation of such compounds is disclosed by Buckman et al. in U.S. Patent
3,859,322. The method of preparation comprlses reacting an alkali metal salt
¦ of a thiolsulfonic acid in the presence of an organic acid with a 1,2-epoxide.The preferred compounds of the present invention can be prepared sccording to
the teachings of this patent by reacting sodium methanethiolsulfonate with
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ethylene oxide or propylene oxidc in the presence of acetic acld, These
compounds may further be characterized by the names 2-hydroxyethyl methane-
~thiolsulfonate and 2-hydroxypropyl methanethiolsulfonate, respectively, which
have the following formulas:
O OH
11 1 ;
CH~- S- S - CH2- CHa
. 1,,. o
~ 2-~ydroxyethyl methanethiolsulfonate
O OH
1 10 CH3- S - S- CH2 -CH - CH3
2-Hydroxypropyl methanethiolsulfonate
When these compounds are used to inhibit the growth of Legionella
pneumophila accordin~ to the teachings of our invention, suitable quantities
vary from 0.1 to 25 parts per million parts of water. Preferred quantities
vary from 0.25 to 10 parts per million parts of water. It will be understood,
of course, that larger quantities of the compounds may be used with no detri-
mental effect, but such larger quanti~ies increase the cost of operation with
limited material benefit. Both of these compounds are water-soluble and are
compatible with many other water-soluble microbicides co~monly used to treat
aqueous systems such as recirculating cooling ~ater systems~ Thus, they can
easily be formulated wi~h other compatible bactericides, algicides, or fungi-
cides to provide a broad range of microorganism control with a single formu-
lated product.
One of these compounds, 2-hydroxypropyl methanethiolsulfonate, has been
found to be relatively nontoxic to fish. A solution containing 80 percent of
2-hydroxypropyl methanethiolsulfonate, for example, had a 96-hour LC~o of 78
parts per m~llion in bluegill sunfish and a 96-hour LC~o greater than 56 parts
per million in rainbow trout. Such concentrations ar~ much greater than those
required to accomplish comple~e inhibition of L. pneumophila. Thus, the use
~109)
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of this compound for the treatment of aqueous sy6tems presents ~ery little
~hazard to ~he aquatic environment compared to that presented by other micro-
I bicides commonly used to treat ~uch systems.
Il In order to disclose the nature of the present invention still more
clearly, the following illustrative examples will be given. It is understood,
however, that the invention is not to be limited to the specific conditions or
details set forth in these examples, except insofar as such limitations are
'~pecified in the appended claims.
EXAMPLE 1
The effecti~eness of the compound 2-hydroxyethyl methanethiolsulfonate in
inhibiting the growth of the bacterium Legionella pneumophila ~as tested by
the following testing protocol. Le~ionella pneumoPhila (Philadelphia strain,
Serogroup 1), provided by the Center for Disease Control, Atlanta, Georgia, to
Methodist Hospita~, Memphis, Tennessee, where this work W2S performed, was
grown initially and maintained on Mueller-Hinton Agar supplemented by 1%
IsoVitaleX (BBL) and 1% hemoglobin, and subsequently on FG sgar at 35C, pH
6.9, under 2.5% CO2. For the preparation of an inoculum for testing, the
organism was grown on FG agar plates for 72 hours and resuspended in 0.85%
saline in distilled water at pH 6.9. The concentration of cells was adjusted
to 6 x 10~/uL. A dilute stock solution of 2-hydroxyethyl methane thiolsulfo-
nate was prepared fresh immediately before testing by weighing a sufficient
quantity to prepare a stock solution in sterile 0.85% saline containing 1.000
mg/mL of the compound. The stock solution was added to sterile 0.85% saline
so that when 1.0 mL of the inoculum was added to the contact bottle, it would
contain a total of 100 mL. Tests were run with con~act times of 2 and 8 hours
at a contact temperature of 24C. Then 4-mm loops from each contact bottle
were streaked on FG agar plates, which were then incubated at 35C for 10 days
under 2.5% C02. At the end of the lncubation perlod, the number of colonies
on ~ach plate was counted. The results are summarized ~n Table 1 with the
following key for interpreting the results:
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4 = More than 300 colonies on recovery plate
3 = 151 to 300 c~lonies
j 2 - 51 to 150 colonies
~I 1 = l to 50 colonies
5 1, 0 ~ No growth
! ! TABLE 1
I Inhibition of L. pneumophila by 2-hydroxyethyl methanethiolsulfonate
¦, Concentration, Contact Time
~ i parts per
l~ million 2 Hours 8 Hours
0 4 4
0.25
0.5 0 0
1.0 0 o
2.0 0 o
These results` show that addition of only 0.25 part per million 2-hydroxy-
ethyl methanethiolsulfonate provided a high degree of inhibition of L. pneumo-
phila, and 0.5 part per million of the compound gave complete inhibition even
with only 2 hours contact time.
EXA~LE 2
The-compound 2-hydroxypropyl methanethiolsulfonate was evaluated for its
efficacy in controlling the growth of L. pneumophila by use of the same proce-
dure described in Example 1. The results are shown in Table 2. The key for
the interpretation of the degree of inhibition is the same as used for Table 1.
TABLE 2
25 Inhibition of L. pneumoDhila by 2-hydroxypropyl methanethiolsulfonate
Concentration, Contact Time
parts per
million 2 Hours 8 Hours
0 4 4
0.5 1 0
l.o o o
2.0
6.0 o o
_ g _
ll
1 ~ 7 7 5 ~ og)
!
l~ The results of these tests show that a concentration of only 0.5 part per
`million 2-hydroxypropyl methanethiolsulfonate provided a high de~ree of inhi-
bition of the growth of L. pneumophila with just 2 hours contact ~ime and
1, complete inhibition ~ith 8 hours of ~ontact time~ A c~ncentration of 1.0 part
l per million 2-hydroxypropyl methanethiolsulfonate provlded a 100% kill of L.
j'pneumophila with a contact time of only 2 hours.
¦I EXAMPLE 3
1 The following compounds or mixtures of compounds that are commonly used
, .
in cooling water treatment were evaluated by the same procedure described in
Example 1. The minimum inhibitory concentrations of each required to kill
Le~ionella pneumophila with only 2 hours contact time are shown in Table 3.
The results for the compounds of this invention, 2-hydroxyethyl methanethiol-
sulfonate (HEMTS) nd 2-hydroxypropyl methanethiolsulfonate (HPMTS), are
included in the same table for comparison purposes.
Microbicide A: Potassium N-methyldithiocarbamate, 14.7 parts
Disodi~ cyanodithioimidocarbonate, 20.3 parts
Microbicide B: Potassium dimethyldithiocarbamate
- Microbicide C: 2,2-Dibromo-3-nitrilopropionamide
Microbicide D: Polyloxyethylene (dimethyliminio) ethylene
(dimethyliminio) ethylene dichloride]
TABLE 3
Minimum inhibitory concentrations of cooling-water microbicides
required to kill L. ~neumophila in 2 hours contact time
Microbicide Minimum Inhibitory C_ncentration
Parts per million
A 70.
~5 B 100.
C 4.0
D 120.
HE~TS 0.5
HPMTS 1.0
These tests clearly indicate the great superiority of 2-hydroxypropyl
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methanethlolsulfonate and 2-hydroxyethyl methanPthiolsulf~nate over other
j commonly used microbicides in controlling the growth of Legionella pneumo-
i; phila in aqueous media.
,, While particular embodiments of the in~ention have been described, it
I will be understood, of course, that the invention is not limited thereto since,~ ~any modifications may be made and it is, therefore, contemplated to cover by~i the appended claims any such modifications as fall within the true spirit and I scope ~f the invention.
~ The invention having thus been described, what is claimed and desired to
be secured by Letters Patent is:
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