Note: Descriptions are shown in the official language in which they were submitted.
- Dx.26110
~ .-
,, ., j
04S5~S
..i
j ~hiB invention relates to blocidal compo6itions and
'' more pRrtioularly to combinations and complexes of lunown biocide6
having improved effectivenes6.
UK Patent Speeirication No. 884,541 relates to a
process for the protection of ~queou~ media against infectionby mioro-organisms by adding to the aqueou6 media a 1,2-benziso-
thiazolin-3-one which may be substituted in the benzene ring
.I by chlorine or brominet or salts thereof.
~ ~t ~
Dx.26110
~04SSgLS
Also known to have biocidal activity iare quaternary
ammonium compvunds in which one at least of the substituent6
on the nitrogen atom i6 a long~chain alkyl group or an alkyl
phenoxyethoxyethyl group.
It has now been ~ound that ,certain 192-benz isothiazolin~3-one
- compound6 and quaternary a~m~nium compounds in admixture
- are particularly e~fective bioci~esO
According to the pre~ent invention there are provided ::
. . . - _ .... - . . .
blocldal compositions comprising: -
A(i) a quaternary ammonium compound of the general ~ormula
R R1R2R3N + X Formula I
wherein R represents a C10_C22 alkyl group or a C1- Cg
alkylphenoxyethoxyethyl group, or a phenoxyethyl group~
¦ R1 repre8ent6 a C10- C22 alkyl group, a C1- C4 alkyl
,~ ~5 group or a benzyl group~
.,
.~ R2 and R3 each independently represent a C1 - C~ alkyl
group and -
X~ repre6ents an anion
~, ~
or (ii~ a quaternary ammonium compound of the general formula
.
.
~ - 4 x Formula II
wherein R4 represents an optionally substituted C~O - C22 alkyl group :~
represent6 an an~on,~
,~, , . ~ .
~ 3 _ . :
''~': ; , , ' ': ~
,
, .. - . - , ., . ., .. ; . . . . . . . . .. .. . . .
lO~;L5545 Dxo261 10
~- ~` and wherein the heterocyclic nucleu~ may be fu6ed with a benzene
nucleus adjacent to the nitrogen atom and which may carry an
amino substituent; and
B a ~ubstituted or unsub~tituted 1~2~benz i60thiazolin-3-one
oo~po~md having the gener~l formula
~ ao
NE
~J\S/ Formula III
!
~herein the benzene nucleus may carry substi~uents 6elected from
halo~en atom~ C1-C4 aIkyl groups, C1-Cl~ alkoxy groups, the nitro
group and the cyano ~roup.
; 10 ~8 examples of the alkyl ~roups whioh are represented by
R and R1 in the compou~ds of Formula I there may be mentioned the
dodeoyl group and the hexadecyl group.
As an example of the alkylphenoxyethoxyethyl group which
R represents there may be mentioned the p-octylphenoxyethoxy ethyl
grou~.
The ~roups R2 and R3 ~ay be, for example, ethyl, propyl,
isopropyl or the lsomeric butyl groups but are pra~erably methyl
'' groups.
The groups R4 in the compound~ o~ Formula II may be~ ~or
example~ a dodecyl group or a hexadeoyl group~ and as an example o~
j a substituent whioh ma~ be prese~t in the group R4 there may be
mentioned a substituted or unsubstituted pyridinium radical as defined
in ~ormula II.
As example~ of the anions which are represented by X there
may be mentioned the ohloride, bromide, acetate or methosulp~ate ion~
Speoi~ic examples of the quatern~ry ammonium compounds o~
Formula I and II are dodec~l benæ~l dimethyl ammorium chloride~
4 -
:
;. ~. :. , ,: ~ :~: ,:.:, .,.,:: , : .,, , , , ,: . ; ,. . , : , .. . . . ...
~ .
DX fff 26110
~LO~S5~5
- hexadecyl trimethyl a~nonium bromide, hexadecyl pyridinium chloride,
f4-ar~noquinaldinium dodecyl acetate~f didecyl dimethyl ammonium
chloride and dioctyl dimethyl ammo~it~ chloride. The preferred compound
is a dodecyl (lauryl) benzyl dimethyl ammonium salt~
g As examples of 1,2-benz isothiazolin-3-one6 which ma~ be
emplo~ed to~ether with the quaternary ammonium compounds of Formulae
I and II there may be mentioned 1~2-benz isothiazolirt-3-one itself
-~- and the 5- and 6-chloro and 5-methyl derivatives thereo~. -
The molar ratio of substituted or unsubstituted 1,2-benz
isotbiazolin-3 one as bereinbefore described, to qt~ternary ammonium
compound in the compositions of the pre6ent invention may be varied
widely, but will eenerally be in the rAngef 1.0 : 0.04 ~o 1.0 : 1Ø
The combination of the quaternary aMmonit~ compound and the
benzisothiazolinone has a biocidal actiYity greater than that lfrhich
would be expected as the sum of the aotivities of the indiYidual
components.
It i-g preferred tbat the composition co~prising the
quaternary a~monium compound and the benzisothiazolinone is in the
form of a complex of the two compounds in which there is one
benzisothiazolinone group for each quaternary ammoni-un group i~ the
quaternary~ammom um compoùnd.
~I These comple~eff~ according to the present invention may be
prepared by mixing a~ aqueous solution of an alkali metalt pre~erably
odium, salt of~the benz i60thiazolone with an aqueous solution of
25f the quaternary a~moniuIf compound in 6toichiometric proportion~f. The
oomplex~6eparat6s ~d may be~purified by~dissolving it i~ an organic
solvent~ 6eparRting an~ insolubl6~impurities, wash;n~-the ~olution ;~
f
~ 5 -
~, ~
~;
~;
, Dx.26110
~6~45545
! with water, drying the solution and removing the solvent by evaporation
or distillation under reduced pressure.
'~he biocidal compositions of the present invention find
I application as bactericides and un~icides, p~rticularly in aqueous
~edia, for example~ tYater-oil emulsions, in-can preservation of
water-based paints and adhesives and in coolinO water systems; also
; as paint-film fungicides and in preventing fungal attack on woodO
The compositions are also effective in the presence of anionic compounds
- for example, soap, which are not compatible with quaternary ammonium
- 10 biocides alone.
Thus the present invention also provides a method Por p~r~cring~
aqueous media against infection by micro-organi~ms, and ~ontrolling
or preventing the proliferation o~ micro-or~anisms in aqueous media
already lnfeoted thereby~ which comprises adding to the aqueous media
from 1 to 1,000 part~ per million of a biocidal composition comprising
in admixture a quaternary amm~nium compound and a substituted or
unsllbstituted 1,2-benzlsothiazolin-~one as hereinbefore described.
A further feature of the invention provides a method for the
protectior. of paint ~ilms against fungal attack wherein there is
added to the paint before its application to a surface, a biocidal
composition comprising in admixture a quaternar~ ammonium compoumd
and Q substituted or unsubstituted 1~2-benzisothiazolin-~-one a~
~herei~before desoribed, in amount to provide a concentration in the
.. .
in the paint o~ 500 to 10,000 parts per million.
A still ~urther featura of the invention provides a method
for the protection of wood against iungal attack wherein the wood
(, iR treated with a solution of a composition comprisin~
. ',~ ' . ' ' .
~;
. ~ ~
.. . , . , , , . ~, ~ ", . , . .. , , ,, .. ", . ... ..
104554S DX . 26110
~ in admixtule a quaternar~ ammonium
- compound and a substituted or unsub&tituted 1,2-benzisothiazolin-3-
one a6 hereinbefore described9 the solution containing frol~ 50 to
20,00~ paste per nillion of the compo&itionO
The treatment of the wood may be by dipping, brushin~ or
~praying with a solution of the composition~ in which case the
solution may be aqueous or substantially aqueous. For example,
a 50~ 601utio~ of the composition in propylene ~lycol may be added
to ~ater to give a substantially aqueous solution of the complex
having the desired stren~th. The wood fiO treated may then be
I allowed to dry at normal temperatures or the dr~ing may be accelerated
i by heatin~ the treated wood for some time at an elevated temperature,
for example, up to 50C for 6everal hour60
Alternatively the wood may be treated by vacuum or pressure
impre~nation, in which case the composition may be applied from a
non aqueous rnedium, for example, a hydrocarbon mediu~l such as toluene~
petroleum spirit or mixtures thereof.
'rhe invention i8 illu~trated but not limited by the
following Fxamples in which parts and percenta~efi are by wei6ht
Ullle~8 otherwise 5tated~ the ratio of parts by wei~ht to parts by
volume being that of the kilogram to the litre.
, ~a~ .
~To 6.o6 parts of 1~2-benzisothiazolin-~-one is added
40.62 parts of a solution of sodium hydroxide prepared ~y di3solving
2~056 p~rt~ o~ sodium hydroxide in 50 parts of wateri
7~
` 1,;:
.~r .~ !
1045S45 DX, 26i10
The rnixture is warmed to effect solution and there i~ then
added, with stirring, 28.10 parts of a 48.596 aqueous solution
of lauryl benzyl dimethyl ammonium chlorideO An immediate
phase separation occurs and the two-phase system i8 warmed
for 2 hours at 90-100C.
The lower layer (20.09 parts) is separated and is dried
overnight in a vacuum oven at room temperature. The product
is dissolved in 250 parts of benzene and the solution i8
heated under reflux using a Dean and Stark separator to
. .
- 10 remove water from the refluxin~ mixture. The benzene solution
is cooled and filtered. The filtrate is washed with four
separate portions of 50 parts of water and is then dried over
anhydrous magnesium sulphate. The drying agent is removed
by filtration and the benzene solution is evaporated to dryness
under vacuum. The residual product, 16.7 parts, containg 0.1~6
of chloride ion and it~i infra-red spectrum indicates the prosence
of both benzisothiazolin-3-one and quaternary ammonium
components.
.- . ~ .
Comparison between the bactericidal activity o~ 1.2-
.
benzisothiazolin-3-one ~BIT) and the complex from the latter
compound and lauryl benzyl dimethyl ammonium chloride (LBDAC).
1 ml. of an overnight broth culture of the baoterium
.. . . .
, Pieudomonas aeru~inosa is introduced into each of 100 ml
25 Yolume6 of test solutio;ns of the biocides in 250 mlO conical
flasks, at concentrations oIn 1000, 500, 250 and 100 p.p.m.
'., ' respectiYely.
, ` , . :; . ,
- .
~ . . ,
,.,,, :
,8~
;:
s
Dx. ~6110
;iS9LS
.
The test solutions are prepared from a soIution of 1.0 g.
of the biocide in 100 ml. methanol/water 1 : 1 v/v by dilution
with water to give a test solution of the desired strength.
Thus a test solution containins 1000 p.p.m. of biocide also
contains 5~ methanol, and lower concentrations of bioclde
proportionally less methanol.- ~ach culture is then incubated
on a rotary shaker at 25 C and samples are removed a~ter 2
'~ hours and 6 hours in order to determi~e the number o~ surviving
- cells. Result~ are as follows:-
Concentration Survivins_~ells/ml.,after:
~,' of biocide
(p p.m.) 2 hours 6 hours
BIT 1000 8.0 x 106 0
5 ~ 3.0 x 107 0
250 ~ 3.0 x 107~ 3.0 x 107
100 ~ 3.0 x 107~ 3.0 x 107
Equimolecular1000 0 0
complex of
BIT and LBDAC 500 0 0
250 0 0
~~ ~ 100 1.0 x 103 0
3; ~
, 20 Control
, ~ ` (no biocide) 7 3.0 x 1077 3.0 x 107
Control
(5~ aqueous
methanol) ~73.0 x 107;7 3.0 x 107
It is clear from tkle above that the complex has a
~ bigher bacter1cidal actlvitg than the same amount of 1.2
be~zisothiazolin-~-one alD~e~
Dx.26110
, '
The 5,~6 aqueous methanol control represents the highest
concentration of methanol in any of the tests, and shows that
,, the methanol has no detectable effect on the resultfi.
, ~ ample
.
~` 5 Comparison of the fungicidal activity of 1.2-
benzi60thiazolin-3-orle (BIT) and the complex from the latter
compound and lauryl ~enzyl dimethyl ammonium chloride (LBDAC~.
Four 1 cm. square nylon meshes contaminat~d with overnig~ht
growth of Aspergillu6 ~G~ are introduced into 100 ml. Yolumes
of test solutions of the biocides in 250 ml. conical flasks,
prepared as de6cribed in Example 2. Each culture i8 then
incubated at 25C on A rotary shaker. Two nylon squares are
~' removed from eaoh fla6k after a contact time of 3 hours and
the remainlng two squares after 6 hours. On removal from
the solution each nylon 6quare ~ gorously shaken in 10 ml.
, of sterile water for 5 minute6 and tran6ferred on to a malt
agar medium in a Petri dish. The latter is incubated for
3 days at 25C, after which the 6urvival or kill of the fungal
.,.. '1 : ~ myoelium i6 determined accordi~g to the presence or absence
; 20 of fungal ~rowth on the malt agar medium from the nylon mesh.
Pesults are as follows:-
~' , ' .
,. ~.
~' `, ' ' , ~ '
~ , ~
. ~
~ 45~45 Dx.26110
Concentr~tion Survival of fun ~ s after contact
of biocide ti~e of:
(p.p.m.) ~ h~ 6 hour~
BIT 1000 j - -
500 _ _
250 _ _
'
. CoMplex 1000 - - ~
5C0 _ _ ~.
. , .
~. 250
.
. 10 100 _ _
~` Control
- (no biocide) ~++ +++
. Control ~5
Aqueous methanol) +++ ++~
., .
. 15 In the above qualitative assessment +++ indicates
vigorous growth, while - indicates total kill. It is clear :~
. `! that the complex has a higher fungicidal activity than 1.2-
~ benzisothiazolin-3-one a1one.
¦ ~ 20 : Comparison of the bacteriostatio and fungistatic activity
:, 1: -
o~ 1.2-benzisothiazolin-3-one (BIT) and the complex ~rom the
latter compound and lauryl benzyl-dimethyl ammonium chloride
(LBDAC). .
. 1.2 Bénzisothiazolin 3-one and the complex are each
dissolved in a 1:1~v~v methanol/water solvent, and the
solutions are~:~added to nutrient:agar and to malt agar to
: provide conoentratlons of 20, 40, 60, oO, 100 and 200 p.p.m.
of the active ingredien~. Ihe test bacteria, ~scherichia ooli,
D~,261 10
1~4S5~5
Stap~loco cus aureus and Pseudomonas g~s~gy~ are
streak i~oculated on to the 6urface of nutrient agar in
Petri dishes at the concentrations indicated aboYe. I'he
te6t fungi Altenaria tenuis, Pullularia pullulans, ~h~~1Ym
~lobo~um, Pol~tictus ver~ic~lor,r~2~,hoderm~ viride and
A~e~ n ~er are similarly inoculated (point inoculum)
on to the malt agar medium. The nutrient agar plates are
- incubated for 1 day at 37C and the malt agar plates are
incubated for 7 days at 28C, and then examined for the
. _ .,,, -. . -- . .
presence or absence of growth~ Results are as follows:-
Minimal inhibitory concentr _ o
, against
Ps. aeru~inos~ E. coli Staph.aureus
~ BI'r 40 20 20
', , 15 Complex ~00 20 20
'~ ' Minimal inhibitory concentration (p.~.m.)
~, against
,; A,tenuis P. Ch. P- rr. A
t~ 3~ ~lobosum versicolor viride ni~e.r
20 BIT ~ 20 ~0~0 < 20 ho 200
` ComplexC 20 ~ 20< 20 ~ 20 40 60
LBDAC 80 ~ 20200 < 20 200 ~200
: , ' '
From the above it can be seen that the ~ungistatic
~` ~ activity of the complex`is, in general, fiomewhat greater than
that of either component of the complex used alone.
~ ~ .
,. . ~ ~ '
:
45~ Dx. 26110
am~le ~
Comparison of the fungicidal activity of 1~2 - benziso-
, thiazolin-3-one (BIT) alone and together with the cationic
,-~ compounds lauryl benzyl dimethyl ammonium chloride (LBDAC),
cetyl trimethyl ammonium bron~de (CThB) and cetyl pyridinium
chloride (CPC), thenonionic adduct of nonylphenol with 8 mols
~ of ethylene oxide and the anionic sodium dioctyl
;~ - sulphosuccinate (SDSS).
~ ~ ~ Sterilised 1 cm. square nylon meshès on malt agar are
.. . .
~' 10 seeded with a spore suspension of Asper~illus nige~ and
incubated for 24 hours at 25C, after which time the mesh
.... .
~ ~ is covered with fungal mycelium while no spore formation has
,.; . :
commenced. Each test biocidal composition is prepared at
-, the required concentration in a 100 ml. volume of water in a
250 ml. conical flask. Four fun~us-seeded squ~res are tran6~
ferred into each flask and the flasks are then rotated on an
, , .
orbital shaker. Two meshes are removed from each flask after
i ~ 3 hours and the remaining t~lo meshes after 6 hours of incubation.
,', .
~, Each mesh is vigorously shaken in 15 ml. of sterile water
: 20 before being transferred on to a malt agar plate. The latter
' is inoubated for 3 days at 25C after which the nylon meshes
~i are examined for the presenae or absence of fun~al growth.
A oomplete kill of the fungal myceliu~ is indicated by the
absence of any growth on or around the nylon mesh on the malt
25agar p}ate. ~ ~ ;
p ' . ; .
~ Resulta are ae followa:-
,,~1;, I
:.,: : :
.,' ', ' , ' . , . . ' ,, . ' ~ '
. .:
DX.26110
- 10455~S
Survival of fun~us after contact time of
Biocide (~.p.~ k~ 6 hours
Control (no biocide) +~ ~++
BI~ (1000) -~++
BIT~ (100) ++,+ +~
BIT (100) ~ LBDAC (20) - -
BIT (100) + LBDAC (10) +
BIT ( 100) ~ CTAB (20) ~ ~ -
BIT (100~ + CTAB (10) . ~ -
., ~ - - . :
BIT (100) + CPC (20)
BIT (100) + CPC (10) ++ +
BIT (100) + adduct
. (10) ~+~ ~++
BIT ~100) + SDSS (10) ~+~ +++
LBDAC t20) +~+ ~+
CTAB (20) ~++ +++
CPC (20~ ~ +~+ ~+~ :
Adduct (10) +~+ ~++ : :
SDSS (10) +~+ . +~+
,~: . ''
~ ' 20 ~ In this ¢ase the BIT was used in the form of a solution of
~ l
; ~ BIT (33.3~) in ethylene diamine t24.~h) and water (42.7~o)
: ~l (percentages by weight).
, In the above table +~+, ~+ and ~ indicate vigorous growth, -
. ~ i ~ . , : , ~
. moder~te growth and slight growth respectively, and - indicate~ :.
a total kill~
;, ~ .~ .
; j ,. , - .
14
Dx. 26110
1~5545
The above tests show that, in the presence of the cationic
compounds lauryl benzyl dimethyl ammonium chloride, cetyl
trimethyl ammonium bromide a~d cetyl pyridinium chloride,
1.2-benzisothiazolin-3-one is more effective as a fungicide
~ 5 than when used alone. It is also apparent that the non-ior~c
- nonylphenol ethylene oxide adduct and the anio~ic sodium dioctyl
sulpho6uccinate surfactants do not haYe a similar efEect with
the 1.2-benzisothiazolin-3-one.
The experiment described below shows in greater detail
; 10 the effect obtained by using 1.2-benzisothiazolin-3-one and
lauryl benzyl dimethyl ammo~ium chloride in combination. All
other details are as described above.
~; 5urvival of~ un~us ~fter contaot time of:
1 hour ~ E~ 4 hours 6 hours
BIT~1000 ~++ + +
500 +++ ~++ ++~ ~++
,, ~ ,1 100 +++ +,,,+
i l 50 ++~ ~++ ++~ +++
~1 LBDAC 50 ++-~ +++ +++ +t+
.~ ,
" 20 +++ ~ +~ +++
' " 10 +++ +~+ ~+ +++
,, , :
BIT(100) ~ LBDAC(20) *++ +
BIT(100) + LBDAC(10) +++ +++
BIT (50j + LBDACt10) ~ +
~The BIT in this case was used in the form of a solution
in agueous ethylene diamQne as described~in the first part of
this Example.
15 -
~5~5 DX. ~6110
- Example 6
Comparison of the bactericidal activity of 1.2-benr~iso-
thiazolin-3-one (BIT) alone and together with lauryl benzyl
- dimethyl ammonium chloride (LBDAC), the adduct of nonylphenol
with o mols of ethylene oxide and sodium dioctyl sulphosuccinate
(SDSS).
Test biocide solutions are prepared at the required
concentration in 100 ml. volumes of water in 250 ml. conical
~; flasks. rThe control consisted of 100 ml~ of sterile water.
A 1 ml. suspension of a 24 hour culturè of Pseudomonas ~ -
aeru~inosa is transferred into each flask, and the flasXs
are then incubated at 25C in an orbital shaker. r~be numbers
., .
- of surviving bacteria ~re determined after 1, 2 and 6 hours.
Resiults are as follo~G:-
~urviK, I ~f b~c-~ri~ Icount per ml.) after:
, Biocide ( ~ 1 hour 2 hours 6 hour~ -
/ BIrP~ ~1000)~ 300 x 1057300 x 105 ~-10
'' ~BIrr~ (1C0~7 3O x 105~300 x 105 ~ 300 x 105
, . ,
LBDAC ~10) 30 x 105 42 x 10 120 x 103
'~ 20 Adduct (10) ~300 x 105~30G x 105 ~ 300 x 105
i ~ .
SDS~ ~10) ~3G0 x 105~300 x 105 ~ 300 x 105
BIT~100) + LBDACt10) ~10 x 10 57 x 103 C 10
, l ~ BIrr~100) ~ Adduct(10) >300 x 1~ > 300 x 105 ~ ~300 x 105
~` BIT(100) + SVSS (10) ~ 300 x 105 ~ 300 x 105~ ~300 x 105
Control > 300 x 105>3O x 105 ~ 300 x 10
The BIT in this ~ase was used i~ the form of a solution
t ~ iD aqueous~ethyleDe diamine~as described in the first~part of
Example 5~
16 -
. j : ..,
Dx.26110
~O~SS45
The benzisothiazolone-lauryl benzyl dimethyl ammonium chloride
combination9 in the ratio 100 ppm : 10 ppm sho~s activity after 6 hours
whilst the other combinations do not, and is AS active as 1000 ppm of
the benzisothiazolone alone.
ample 7
Comp~rison of 1.2-benzisothiazolin-3-one (BIT) and its
complex with lauryl benzyl dimethyl ammonium chloride (LBDAC)
~ as paint film fungicides.
; ~ The test biocides are mixed into an e~ulsion paint to
~ provide the concentrations (w/w) in wet paint as shown below.
. .
A commercially availablé fungicide, 2 -(4'-thia~olyl) - benzimidazole,
; is used as a control. The paint in the can is then stored
at 50 C for 2 weeks in order to simulate prolonged storage.
The treated paint is applied to one side Qf a filter paper
"
5.5 cm. in diameter, and after 24 hours a second coat of paint
; 15 is ~pplled to the si~me ~ide.
A Gecond set of filter papers coated with two layers of
treated paint is subjected to a weathering procedure consisting
of a 24 hour leaching period in a standard spray cabinet
(BS 3900 F4) and 24 hour heatin~ in an oven at 65C~
The filter papers are then transferred on to malt asar
in a Petri dish (paint side uppermost) and inocùlated with one of
. j .
; the two test fungi Pullularia pullulans and Altern_ria tenuis.
... .
The inoculi~ consists of a visible suspension of the test
organi m sprayed over the painted filter paper and surrounding
border of agar in the Petri dish. The inoculated paint films
are incubated at 25C for 5 days, after which the filter
papers are examined ~or the presence or absence of fungal
growth. Re`sults are a~ follows:- ;
17 _
Dx 26110
~ 5545
Minimal inhibitor~__evel of _iocide in ~int
,~ ~
; Compound (p.p.m.) Storage Weathering
Alt. tenuisP. Pullula=6 is P. Pullulanus
.. , . ,.~_ .. _ .. _~ -
Complex of BIT and
LBDAC (1000,3000)1000 1000 3000 1000
BIT(500,1000,
2000, 3) 500 500 ~000 3000
2-(4'-thiazolyl)
benzimidazole
(1000,3000) 3000 1000 3000 1000
....~ ._ ... _
_ . :
: `:
Since only one tllird of the weight of the complex is BI~,
it i6 clear that the BIT i8 more active as a paint film fungicide
when in the form of its complex with LBDAC than when used
~, J alone,
~,
~( Example 8
;~ 15 The aotivity of the 1,2-benzisothiazolin-~-one (BIT)/ lauryl
, benzyl dimethyl ammonium chloride ~LBDAC) complex as an antifungal
agent in prevent1ng the growth ofcellulc1ytic fungi on wood. ~`
Malt agar plates were spra~ed with a suspension of ~
versiaolor or Chr~sospori~um li~norum- Three cubical blocks of beeoh
wood (sides 1 cm.) were soaked for 5 minutes in a 0.3~ w/v aqueou6
~olution of BIT/LBDAC oomplex, (i.e. containing 0.3 g. complex in 100 ml.
solution; prepared by adding a 50~ solution of compIex in propylene
.~,.;
glycol to water), dried at 50C for 4 hours and eqiulibrated at room
temperature for 20 hours.
A PY 900 tpoly (vin~l chloride))soreen mesh whioh had previously
beer. soaked in~70~ ethanol for 36 hour~ and allowed to dry was txansferred
on ~o the innoculated agar surfaoe ir~ each plate. The three biocide-
treated beeohwood blocks were placed on the screen mesh and innoculated
t 25C for three week~. The~blocks were then e~a~ined for the presence
1?~ 4 5 5 4 S Dx~26110
,
or absence of fun~al growth.
Results are as follow~s:- -
,~ ~ Growth o~ beech blocks of
Polxstictus ~,ersicolor~
' 5 Control (no biocide) ~ *~
BIT/LBDAC complex
~1 .
~, 0,3~ solution
:i -.. - - -' :'' ' -
Key: ~+~ indicates YigOrOUs growth
~ oderate
; 10 _ " no " ttotal kill)
.; . , , :
' It is clear from theæe re~ults that the BI~/IBDAC complex
'~ at tho,above-mentioned ooncentration presents the ~rowth o~ the
fungi on beeoh wood.
, .. . . .
.' ~a~ . -
'''~ 15 ; The effeot of BIT/IBDAC complex in retarding the oxidation
~ , of nitrite by Nitrobacter in a model cooli~g tower system.
,, ,l A model cooling tower 6ystem WRS constructed, consisting
i, of a wooden frame 6upporting six wooden slat6~ the fxa~e being
mous~ted over a reservoir holding 10 litre6 of water containing
20 sodium nitrite as a corrosion inhibitor~ The water is then
~, co~tRmsinated with 1 g. o~ oooling tower 61ime lunown to harbour
Nitrobacterl and the w~ter ln the reservoir is directed to the top
o~ the wooden slats by means~of a Churchill pump, so that the water
tri¢klea down~oYer the~slats and returns to the reservoir. If no
27 biocide is added to the water the'wooden slats become colonised'by a
,, . , ~
?~ ~ ~
~, Nitrobacter - containi~lg slime which rapidly ovidi~es nitrite to
' nitrabe.
Initi~lly9 the rate of los~of nitrite ~rom the ~ystem was i'
;, :, , ~ , i:
19 - ?
~ 5~
: `\
~ Dx.26110 ~:)4S5~5
determaned in the absence of biocide by removing water samples ,-~nd
- analysinF for nitrite content. 'Ihe nitrite level was then adiusted
to a hieher concentration before 150 ppm. of BIT~LBDAC complex was
added to the ~later. The ~coolirgtower' water was then circulated - --
for the necessary period until the concentration of nitrite was
appreciably reduced, the water in the ~ystem ~as drained o~f and
fresh nitrite - containin~ water was added. Be~orè the next biocide
- for test w~s added~ the rate of loss of nitrite in the biocide-~ree
~ystem was checke3d, to ensure that the nitrite-oxidising actiYity
had normalised, i.e. all biocides were compared In the same system
consecutively.
Re~ults are as ~ollows:-
B ide and concentration Circulation Ni~trate leveladded to water initially _me ~ detected ln water (ppmi)
Control (no biocide) 0 965
i 2 450
3 4 517
3 8 316
I LBDAC, 250 ppM. 49 4
2 550
i 3 L~96
6 550
7 47 -
Il 8 470
1 25 10 522
14 360
17 317
Poly(hexamcthylene biguanide) 0 ô40
, bydrochloride, 85 ppmO 4 705 1
I ~ 30 11 260
~ 18 125
-3~ 5 o
; BII/LBDAC co~plex, 150 ppm. 2 690
; ` ~ ; ; 16 655
- 23 538
580
20 -
DX.26110
~LS~
These results indicate that under the experimental
conditions the BIT/LBDAC complex is markedly more effective than
either LBDAC alone or poly(hexamethylene biguanide) hydrochloride
(a ~ell-known bactericide) ~e~eas these two compounds ~aintained
the initial level of nitrite concentration for approximately 10 days
and 4 days respectively, there ~Jas no significant fall in the nitrite
level of the water treated with BIT/LBDAC complex a~ter 58 days.
~`~~~~ ~ am~le 10
- .
The efficiency o~ BIT/LBDAC complex in retarding the
oxidation of nitrite by Nitrobacter in a cooling tower.
An all-wood cooling tower system~ 25,000 gallons in
capacity ~ th a ~aximum temperature of ~2C. was lsed for this trial.
Before the biocide was added the cooling water wa~ bled so that
addition of nitrite to ~ive a concenkration of 500 ppm the total
dissolved solids were not more than 1,000 ppm. The biocide (BIT/
L~DAC complex) was added at the distribution troughs at th~ top of
the tower to give an initial concentration of 150 ppm. active
ingredient .
The single dose of biocide controlled microbial activity
and hence ~atisfactorily prevented lo~æ of nitrite from the system
for a period up to ei~ht weeks. Comparison with quaternary ammonium
compounds showed that BI'~/LBDAC complex provides protection for àt
lcast 7 times longer than 250 ppm. of LBDAC alone,
In this test, coMbinations of BIT with LBDAC or Dequadi~
were co~pared for bactericidal activity,
_ 21 -
. . , .; . .,: . ., :, .: :., . :- . .. . : . . ... . .:, . ... .
. . , . ". . , . , . ... , . . . ,.: . .: : . .
.. . . .
5S4S ~
.
Results are as follows:
Biocide
Control (no biocide) 1~3 x 107
BI~, 200 ppm. 8-o x 108
LBDAC, Z0 ppm. ,B.3 x 103
Dequadin, 20 ppm. 1.0 x 103
BI~ (100) ~ LB~AC (10) < 10 ~ -
... . ~ .
BIT (100) ~ Dequadin (10) 2.o x 102
All conce~trations re~er to active ingredient.
The synergi~m shown by BIT and LBDAC acting together
i~ clearly seen. A lower order of ~ynergism is also shown between
BIT and Dequadin.
Example 12
In this test, oombi~ation~of BIT and Domiphen~ BIT and
Dequadin, and BIT and Phemeride were tested ~or fungicidal ac~ivity
against ~ J~ , the test method being that described in
Example 5.
Results are as follows:
~ Survival of ~un~us after contact times
~ hourfi ~ , '' '
Co~trol (no biocide) ~t ~++
BIT, 100 ppm. ~ +~
Dequadin, 20 ppm. ~+~ ~+
Domdphen~ 20 ppm. +++ ~++
Phemerideg 20 ppm. ~+~ ~+~ j;
BIT (100) ~ Dequadin (10) +~
BIl' (100) + Domiphen ~iiO) ~ ~+ ~ ~ ¦
BIT ~100) ~ Phemeride t10) +~ +
!:
~45545 ~2611Q
These result~ indicate the presence of synergi6tic
activity between BIT and Dequadin, BIT and Domiphen9 and to a lesser
extent betwe~n BIT and Phemeride~
Dequadin, Domiphen and Phemerid~ (Registered Trade Marks)
are all quaternary ammonium compounds having phar~aceutical applications,
and have the following structures:-
2 ~ 2
: ~ CN3 ~ 2X- De~u~din
CH3
C ~ - N~ C~I C~ O ~ . X~ Domiphen
C12H2S
X~ . CN~ ~ 2CN2CCH2CN2 ~ - CN2 C(C~l3~3 Phe~eride
'
' ~
~;
I ' .
I ~.
-: ; : : . ; ~ : . : .
