Note: Descriptions are shown in the official language in which they were submitted.
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W098/59108
PCT/DE 98/01689
OXIDATION AND BLEACHING SYSTEM WITH ENZYMATICALLY
PRODUCED OXH)IZING AGENTS
It is known from a number of literature references and review articles, for
example from
"Preparative Biotransformations" by S.M. Roberts, K. Wiggins and G. Casy, J.
Wiley &
Sons Ltd, that enzymes such as certain lipases are capable of forming epoxides
via the
1o formation of peroxy acids (perfatty acids). For example, in the lipase
system (from
Candida antarctica), with continuous addition of H202 and in the presence of
certain
fatty acids, for example tetradecanoic (myristic) acid or dodecanoic (lauric)
acid, from
cyclooctene the corresponding epoxide is produced. It is also known that
manganese
peroxidases + unsaturated fatty acids can generate peracids which in turn can
act as a
H202 source for manganese peroxidases (literatute: B.W. Bogan et al., Applied
and
Environmental Microbiology, vol. 62, No. 5, pp. 1788-1792).
There are also a few patents that describe the formation of peracids with the
aid of
haloperoxidases.
It is also known that dimethyldioxirane can be generated in situ from peracids
or salts of
peracids (such as Oxone) and acetone as the simplest ketone. Ketones other
than
acetone are also used. Dioxiranes can also be prepared as pure substances
before they
are used as oxidants, but their stability is problematical (WO 92/13993).
Moreover, it is known from Canadian Patent 1 129 162, US 5, 034 096 and WO
96/13634 that certain metal ions, for example Mo6+ + HZOZ and nitrilamides +
HZOZ
and dicyandiamides + H202 are capable of generating dioxiranes chemically from
H202. In this case, surprisingly, certain combinations are possible with the
enzyme
components system (ECS) of the present invention (see below), namely the
enzymatic
3o generation of activated oxygen species e.g. dioxiranes can be further
enhanced.
These very strong and highly selective oxidants can be used in many oxidation
reactions
(for example epoxide reactions etc). Recently, it has been proposed to use
them, in
particular, as bleaching agents in the cellulose /pulp industry. Because of
the dangerous
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preparation and high cost, this proposal has not found acceptance (WO
92/13993).
The object of the present invention is to provide a highly selective oxidation
or
bleaching system for use in cellulose /pulp bleaching or high yield wood pulp
bleaching,
in the oxidative treatment of wastewater of all kinds, in the preparation of
wood-based
composites, as an enzymatic deinking system, as an oxidant in organic
synthesis, in coal
liquefaction, as bleaching system in detergents and as a bleaching agent or
oxidant in
the textile industry (for example in stone washing and fabric bleaching). Said
system
does not present many of the drawbacks of purely chemical systems (for example
1o environmental pollution problems) or of enzymatie systems (which often show
inadequate performance and are very costly).
Surprisingly, we have now found, that an oxidation system which contains
certain
lipases, oxidants such as H202, certain fatty acids and certain ketones
resulted, for
example, in bleaching of cellulose/pulp while at the same time the kappa
number
(delignification) was markedly reduced, i.e. it could be clearly demonstrated,
surprisingly, that when the appropriate optimal components were present in an
optimum
proportion and concentration relative to each other, it was possible to
achieve in the
aforesaid cellulose/pulp bleaching a bleaching action comparable to that of
dioxirane-
forming chemical systems.
2o Surprisingly, it was also possible to demonstrate considerable bleaching
action in the
bleaching of high yield wood pulp, in the bleaching of pulps after a deinking
processes,
in oxidative polymerization of lignin and/or lignin-like substances and in the
oxidative
treatment of wastewater of all kinds, such as wastewater from high yield wood
pulp
preparation (groundwood, refiner pulp), from the cellulose/pulp industry and
dye-
contaminated wastewater, for example from the textile industry. For most of
these
wastewaters, besides the decolorization and oxidation and thus the
"destruction" of
environmental pollutants, the polymerization of lignins is the preferred
application,
because it causes a marked increase in molecular size and permits an easier
and
substantially less expensive precipitation of these polymers and thus their
elimination
3o from COD considerations.
Surprisingly, we were also able to demonstrate this oxidative polymerization
of lignin
and/or lignin-like substances in the preparation of wood-based composites
(binder
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and/or adhesive preparation) by oxidative polymerization of the
polyphenylpropanes
present. Moreover, surprisingly, we were able to demonstrate removal of
printing inks
in the deinking process (probably occurring by swelling of the lignin-
containing waste
paper fibers). Surprisingly, we were also able to observe coal liquefaction
properties in
the treatment of lignite or anthracite. Moreover, also surprisingly, we found
a marked
and selective oxidation power in the use as "oxidant" in organic synthesis,
high
bleaching power when used as bleaching additive to detergents, in the general
bleaching
of textile fabrics and as special bleach when used in the stone washing
processes,
namely as a replacement for mechanical color removal and/or as postbleach in
these
1o processes. It seems possible, that the responsible oxidants(s), for
example, generated
from the present ketones and peracids are dioxiranes which in the above-
indicated
applications serve as oxidants or bleaching agents either alone or in
combination with
the peracids formed.
The foregoing objective is reached also by providing an enzyme component
system
(ECS) according to the invention which contains one or more lipases,
preferably from
the group of triacylglycerol lipases (3.1.1.3), or one or more amidases,
preferably from
the group of amidases (3.5.1.4) ( system component 1) which from one or more
fatty
acids present (preferably C6- Cz6 and particularly Cg - C,6 fatty acids)
(system
component 2) and in the presence of an oxidant such as a peroxide, preferably
H202
(system component 3) can produce peracids and in the presence of ketones as
additional component (system component 4), for example, dioxiranes.
Description of Various Applications of the Enzyme Component S s~(ECS) of the
Invention
I) Use in the bleaching of cellulose/ wood pulp.
II) Use:
a) in the treatment of, primarily, wood pulp wastewater in the pulp and
paper industries, and
3o b) of wastewater in other industries.
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III) Use in the preparation of lignin solutions or gels, of the corresponding
binders/adhesives and of wood-based composites.
IV) Use as enzymatic deinking system.
V) Use as oxidation systems in organic synthesis.
VI) Use in coal liquefaction.
VII) Use as bleaching agent in detergents.
VIII) Use in the bleaching/decolorization of textile fabrics.
Use of the Enzyme Component System (ECS) of the Invention in the
1o Bleaching of Cellulose/Wood Pulp
Wood pulp is currently produced mainly by the sulfate and sulfite processes.
By both
processes, pulp is made by cooking at high temperature and under pressure. The
sulfate
process involves the addition of NaOH and Na2S, whereas the sulfite process
uses
Ca(HS03)2, + SO2, although the sodium and ammonium hydrogen sulfite salts are
currently used because of their higher solubility.
The main objective of all processes is the removal of lignin from the plant
material,
wood or annual plants used.
The lignin, which together with the cellulose and hemicellulose forms the main
constituent of the plant material (stalks and stems), must be removed, because
it is
otherwise not possible to produce nonyellowing, mechanically highly resistant
papers.
The processes for making high yield wood pulp involve the use of stone
grinders
(groundwood) or of refiners (TMP = thermomechanical pulp) which after an
appropriate pretreatment (chemical, thermal or thermechemical) defibrillate
the wood
by milling.
These wood pulps still contain most of the lignin. They are used primarily for
the
production of newspapers, magazines etc.
The possibilities of using enzymes for lignin degradation have been under
investigation
for several years. The mechanism of action of such lignolytic systems was
discovered
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only a few years ago, when it became possible to obtain sufficient amounts of
enzymes
from the white rotting fungus Phanerochaete chryosporium by use of proper
culturing
conditions and the addition of inductors. This is how the hitherto unknown
lignin
peroxidases and manganese peroxidases were detected. Because Phanerochaete
chryosporium is a very effective lignin degrader, attempts have been made to
isolate its
enzymes and use them in purified form for lignin degradation. This was
unsuccessful,
however, because it was found that the enzymes primarily cause
repolymerization of
lignin and not its degradation.
The same is true for other lignolytic enzyme species, such as the laccases
which degrade
lignin oxidatively with the aid of oxygen rather than hydrogen peroxide. It
was found
that similar processes are at work in all cases, namely that radicals are
formed which
then react with each other causing the mentioned polymerization.
Currently, there are only processes based on the use of in-vivo systems
(fungal
systems). Optimization attempts were directed mainly toward biopulping and
biobleaching.
By biopulping is meant the treatment of wood chips with live fungal systems.
There are
two kinds of application forms:
1 . Pretreating the wood chips before charging them to the ref iners or
milling for the
purpose of saving energy in high yield wood pulp production (for example, TMP
or
2o groundwood). Another advantage is the usually achieved improvement of
mechanical
properties of the stock, and a drawback is that the final brightness is worse.
2. Pretreating the wood chips (softwood/hardwood) before wood pulp cooking
(kraft
process, sulfite process). Here, the objective is to reduce the amount of
digestion
chemicals, to improve digestion capacity and extended cooking. The advantages
include
improved kappa number reduction following digestion compared to digestion
without
pretreatment.
The drawbacks of these processes are clearly their long treatment times
(several weeks)
and particularly the unsolved problem of risk of contamination during the
treatment, if it
is desired to omit the uneconomical sterilization of the wood chips.
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Biobleaching also uses in-vivo systems. Before bleaching, the digested pulp
(softwood/hardwood) is inoculated with the fungus and treated for a period of
days or
weeks. Only after such a long treatment time is it possible to observe a drop
in kappa
number and a significant improvement in brightness, so that the process is
uneconomical for implementation in current bleaching sequences.
Another application, mostly carried out with immobilized fungal systems, is
the
treatment of pulp production wastewaters, particularly bleaching plant
wastewaters, for
the purpose of decolorizing them and reducing the AOX value (reducing the
amount of
chlorinated compounds in the wastewater, compounds which were used for
chlorine or
1o chlorine dioxide bleaching). It is also known to use hemicellulases and
particularly
xylanases and mannases as bleach boosters.
These enzymes act mainly on the reprecipatated xylan, which after the cooking
process
partly covers the residual lignin, for the purpose of degrading it and thus
improving
accessibility to the lignin of the bleaching chemicals (primarily chlorine
dioxide) used
15 in the subsequent bleaching sequences. The savings in bleaching chemicals
demonstrated in the laboratory have been confirmed on a large scale only to a
limited
extent so that this type of enzyme must also be classified as a bleaching
additive.
Patent application PCTBP 87/00635 describes a system for removing lignin from
lignin-cellulose- containing material with simultaneous bleaching. The system
is based
20 on the use of lignolytic enzymes from white rotting fungi with the addition
of reducing
agents, oxidants and phenolic compounds as mediators.
According to DE 4 008 893 C2, in addition to the redox system, mimicking
substances
are added which simulate the active center (prosthetic group) of lignolytic
enzymes. In
this manner, a marked improvement in performance is achieved.
25 According to patent application PCT/EP 92/01086, additional improvement is
achieved
by use of a redox cascade with the aid of phenolic or nonphenolic aromatics
"balanced"
in terms of their oxidation potential.
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All three processes are limited in regard to their applicability on an
industrial scale in
that they must be used at low wood pulp consistency (up to a maximum of 4%)
and, in
the case of the last two applications also by the risk of leaching out metals
when
chelating agents are used, the metals possibly causing peroxide decomposition
in the
subsequent peroxide bleaching stages.
WO 94/12619, WO 94/12620 and WO 94/12621 disclose processes in which the
peroxidase activity is increased with enhancers. Enhancers are characterized
in WO
94/ 12619 in terms of their half life.
According to WO 94/12620, enhancers are characterized by the f ormula A= N-N =
B
to where N means nitrogene A and B are defined cyclic groups. According to WO
94/12620, enhancers are organic chemicals containing at least two aromatic
rings of
which at least one is substituted with defined groups.
All three patent applications concern dye transfer inhibition and the use of
enhancers
together with peroxidases as detergent additives or detergent compositions
used in the
15 detergent sector.
Although the applicability to lignin is mentioned in the specification of the
applications,
our own tests with the substances actually disclosed in these applications
have shown
that the claimed mediators are ineffective in increasing the bleaching action
of
peroxidases in the treatment of lignin-containing materials!
2o WO 94/29510 and WO 96/18770 describe a process for enzymatic
delignification
whereby the enzymes are used together with mediators. The mediators disclosed
are, in
general, compounds characterized by the structure NO-, NOH- or NRNOH.
Among the mediators disclosed in WO 94/29510 and WO 96/18770, 1-hydroxy-1H-
benzotriazole (HOBT) gave the best delignification results. HOBT, however, has
25 several drawbacks:
~ it is available only at a relative high price and in insufficient
quantities,
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_g_
~ under deliginification conditions, it reacts forming 1 H-benzotriazole and
other
colored products, this compound shows relatively low degradability and could,
in
large amounts, present a pollution problem,
~ to a certain degree, it harms the enzymes,
~ its delignification velocity is not very high.
Other mediator of the described NO-, NOH- and HRN-OH type do not show most of
these drawbacks, but still have the disadvantage that a relatively large
amount of
chemicals must be used and, particularly, that because of their physiological
reactivity
they may not be entirely harmless (mostly because of NO- radical formation).
1o It is therefore desirable to provide systems for modifying, degrading or
bleaching lignin,
lignin- containing materials or similar substances, which do not have the said
drawbacks
or present them only to a minor degree.
Quite surprisingly, we have now found that when the enzyme component system
(ECS)
of the invention is used, similar or better delignification and bleaching
results are
15 achieved compared to the abovesaid oxidoreductase-mediator systems, and the
said
drawbacks are negligible.
In other words, according the invention, the foregoing objective is reached by
providing
an enzyme component system (ECS) according which contains one or more lipases,
preferably from the group of triacylglycerol lipases (3.1.1.3), or one or more
amidases,
2o preferably from the group of amidases (3.5.1.4) ( system component 1) which
from one
or more fatty acids present (preferably C6- Czb and particularly C~ - C16
fatty acids)
(system component 2) and in the presence of an oxidant such as a peroxide,
preferably
H2O2 (system component 3) can produce peracids and in the presence of ketones
as
additional component (system component 4), for example, dioxiranes.
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IIa) Use of the Enzyme Component System (ECS) of the Invention in the
Enzymatic Treatment of Special Wastewaters (Paper Industry Wastewaters from,
for Example, Groundwood Plants or Refiner Plants)
Unlike most enzymes, oxidases and peroxidases exhibit low substrate
specificity,
namely they can convert a wide range of substances, usually of phenolic
nature.
Without mediators, oxidases as well as many peroxidases show the tendency to
polymerize phenolic substances via free radical-induced polymerization, a
property
which is attributed, for example, to laccase, belonging to the group of
oxidases, also in
nature. The ability to polymerize appropriate substances, for example lignins,
namely to
to increase the size of the molecules involved by "coupling reactions", can be
utilized, for
example, for the treatment of lignin- containing wastewaters in the paper
industry such
as TMP wastewater (wastewater from the production of thermomechanical pulp by
means of refiners) and of grinder wastewater from mechanical wood pulping
units.
The water-soluble lignin compounds (polyphenolpropanes) contained in these
15 wastewaters are mainly responsible for the high COD (chemical oxygen
demand) and
cannot be removed by conventional technology. In the water treatment plant and
in the
downstream waters, they are not degraded at all or they are degraded only very
slowly.
At very high concentrations, these compounds can even inhibit the bacteria in
a water
treatment plant and thus create problems.
2o In this application, the enzyme action can be observed immediately by a
rapid
development of turbidity in the wastewater being treated, caused by an
enlargement and
thus insolubilization of lignin molecules. The target molecules (polymerized
lignin) thus
enlarged in molecular weight by enzymatic catalysis can be removed by
appropriate
treatments (by flocculation, by precipitation with, for example, aluminum
25 sulfate/sodium aluminate, optionally in the presence of cationic or anionic
polyelectrolytes or by sedimentation). The wastewater then shows a markedly
reduced
COD. Upon disposal, such wastewater causes less pollution, namely it increases
the
certainty of remaining below the permissible COD limits. Thus is particularly
important
for not infrequently used "procedures" run at the limit.
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For this treatment, for example with laccase, the cost of removing the
reaction products
of the enzymatic treatment by flocculation, sedimentation or precipitation or
a
combination of several such methods constitutes by far the predominant part of
the
overall cost of the process.
We have now found, quite surprisingly, that when the enzyme component system
(ECS)
of the invention is used by employing a special combination of the components,
much
higher efficiency than be attained than with the above-described enzymatic
systems.
This means that the process according to the invention represents a
substantially
improved system compared to the aforesaid systems employing oxidoreductases
(such
1o as, for example, laccases) as oxidation catalysts, the advantages of which
are, in
particular, its higher oxidation power and the use of readily degradable fatty
acids and
ketones (for example, benzophenones) which, although raising the COD
temporarily,
are readily removed in the subsequent steps carried out in the water treatment
plant.
In other words, according the invention, the foregoing objective is reached by
providing
15 an enzyme component system (ECS) according which contains one or more
lipases,
preferably from the group of triacylglycerol lipases (3.1.1.3), or one or more
amidases,
preferably from the group of amidases (3.5.1.4) ( system component 1) which
from one
or more fatty acids present (preferably C6- C26 and particularly Cg - C16
fatty acids)
(system component 2) and in the presence of an oxidant such as a peroxide,
preferably
2o H202 (system component 3) can produce peracids and in the presence of
ketones as
additional component (system component 4), for example, dioxiranes.
To this system are added other special compounds ( polymerization catalysts)
which
serve as condensation nuclei and can substantially enhance lignin
polymerization so that
the main objective of this enzymatic wastewater treatment, which is to use the
lowest
25 possible amount of the cost-intensive precipitant, can be attained.
IIb) Use in the Enzymatic Treatment of Other Industrial Wastewaters
All other industrial wastewaters containing phenolic or, in general,
oxidizable
substances (for example, lignin, dyes etc) can, in principle, be treated with,
for example,
the abovesaid oxidoreductases. Such treatment can be applied to wastewaters
from
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grape presses, olive presses, dyeing plants in the textile industry,
wastewaters from
pulping plants etc. If at all possible, to attain maximum ei~iciency, polluted
streams
should be treated before they are combined with other wastewaters.
In this case, too, we found, surprisingly, that the use of the enzyme
component system
(ECS) of the invention is very well suited for the treatment of the abovesaid
wastewaters and in some cases shows performance advantages over the
oxidoreductase
systems. Here, too, the abovesaid special compounds, namely polymerization
catalysts, are used. These substances consist of phenols, phenol derivatives
or other
polycyclic phenolic compounds with a number of oxidizable hydroxyl groups.
to Preferably, such polymerization catalysts are, for example:
alizarin, 5-amino-2-hydroxybenzoic acid, 3-aminophenol, pyrocatechol, 2,2-
bis(4-
hydroxyphenyl)- propane, bis(4-hydroxyphenyl)methane, quinalizarin, 4-chloro-1-
naphthol, coniferyl alcohol, 2,4-di- aminophenol dihydrochloride, 3,5-dichloro-
4-
15 hydroxyaniline, 1,4-dihydroxyanthraquinone, 2,2-di- hydroxybiphenyl, 4,4-
dihydroxybiphenyl, 2,3-dihydroxynaphthalene, 2,6-diisopropylphenol, 3,5-di-
methoxy-
4-hydroxybenzhydrazine, 2,5-ditert.butylhydroquinone, 2,6-ditert.butyl-4-
methylphenol, 4-hydroxybiphenyl, 2-hydroxydiphenylmethane, 2-(2-
hydroxyphenyl)benzothiazole, 5-indanol, 2-iso- propoxyphenol, 4-isopropyl-3-
2o methylphenol, 5-isopropyl-2-methylphenol, 4-isopropylphenol, lauryl
gallate, 2-
naphthol, 4-nonylphenol, 3-(pentadecyl)phenol, 2-propylphenol, 4-propylphenol,
purpurine, pyrogallol, 4-(1,1,3,3-tetramethylbutyl)phenol, 1,2,4-
trihydroxybenzene,
2,4,6-trimethylphenol, 2,3,5-trimethylphenol, 2,3,6-trimethylphenol, 3,4,5-
trimethylphenol, 6,7-dihydroxy-4-methyl coumarin, 2-(2-
hydroxyethoxy)benzaldehyde,
25 1 -naphthol, nordihydroguaiaretic acid, octyl gallate, silibinin, 3,4,6-
trihydroxyben-
zoate-octylester, 2,4,6-tritert.butylphenol, 2,4-ditert.butylphenol, 2,6-
dichlorophenol,
indophenol, ethoxyquin, 1-aminoanthraquinone, 2-amino-5-chlorobenzophenone, 4-
aminodi- phenylamine, 7-amino-4-hydroxy-2-naphthalenesulfonic acid, 2-(4-
aminophenyl)-6-methylbenzothiazole, benzanthrone, trioctyl trimellitate, trans-
3o chalcone, bis(4-aminophenyl)amine sulfate, 2,2'- ethylidenebis (4,6-
ditert.butylphenol),
2,2-bis(2,6-dibromo-4-(2-hydroxyethoxyphenyl)propane, bis(3,5-ditert.butyl-4-
CA 02335253 2000-12-15
-12-
hydroxyphenyl)methane, 2,2-bis(3,5-dichloro-4-hydroxyphenyl)propane, Bismarck
Brown Y, I-bromophthalein, 4-butylaniline, 2-tert.butyl-5-methylphenol, 1-
chloro-
anthraquinone, 2-chloroanthraquinone, triallyl 1,3,5-benzenetricarboxylate,
1,1,1-
tris(hydroxymethyl)propane, tri-methacrylate, pentaerythrityl triacrylate,
1,2,4-
trivinylcyclohexane, trans,cis-cyclododeca-I 5,9-tri- ene, pentaerythritol
tetrabenzoate,
4,4'-methylenebis(2,6-ditert.butylphenol), 4,4'-isopropylidene-
bis(2,6-dichlorophenol), 4,4'-isopropylidene-bis(2,6-dibromophenol), 4,4'-
isopropylidene-bis[2-(2,6- dibromophenoxy)ethanol, 2,2'-ethylidene-bis(4,6-
ditert.butylphenol), 3-tert.butyl-4-hydroxy-5-methylphenol, 5-tert.butyl-4-
hydroxy-2-
lo methylphenol, syringaldazine, 4,4'-dimethoxytriphenylmethane and di-
sec.butylphenol.
Also particularly preferred are compounds with several hydroxyl groups, such
as:
ellagic acid, gallic acid, gallein, gallangin, myoinositol, morin, nitranilic
acid,
15 phenolphthalein, purpurin, purpurogallin, quinizarin, chrysazin, quercitin,
quinhydrone,
chloranilic acid, carmine, rhodizonic acid, croconic acid, meilitic acid,
hematoxylin, 9-
phenyl-2,3,7-trihydroxy-6-fluorene, 9-methyl-2,3,7- trihydroxy-6-fluorene,
tetrahydroxy-p-benzoquinone, 2,2',4,4'-tetrahydroxybenzophenone, Pyragallol
Red, 1-
nitrophloroglucinol, 1,4-dihydroxyanthraquinone, 5,8-dihydroxy-1,4-
naphthoquinone,
2o hexa- oxocyclohexane octahydrate, 5,7-dihydroxyflavanone, 3',4'-
dihydroxyflavanone,
glyoxal hydrate, 1,3,5-tris(2-hydroxyethyl)isocyanuric acid, quinalizarin and
2,4,5-
trihydroxybenzamine.
)(>~ Use of the Enzyme Component System of the Invention in the Preparation of
25 Lignin Solutions or Gels, of the Corresponding Binders/Adhesives and of
Wood-
Based Composites
The object of the present invention is to provide a process for enzymatic
polymerization
and/or modification of lignin or lignin-containing materials, for example for
use in the
3o production of wood compositions or wood-based composites such as, for
example, fiber
board from disintegrated wood or particle board from wood chips or wood pieces
(chipboard, plywood, wood composite beams).
CA 02335253 2000-12-15
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It is known from the literature and patents, for example WO 94/01488, WO
93/23477,
WO 93/25622 and DE 3 037 992 C2 that laccases, lignin peroxidases or
peroxidases can
be used for this purpose. The main drawbacks, particularly in the case of
laccases and
lignin peroxidases, are the difficulty of preparing these enzymes and the low
yields even
of genetically modified systems.
We have now found, quite surprisingly, that here, too, the enzyme component
system
(ECS) of the invention shows much better performance compared to the prior-art
enzymatic systems for the polymerization and/or modification of lignin and/or
lignin-
containing materials.
to In other words, according the invention, the foregoing objective is reached
by providing
an enzyme component system (ECS) according which contains one or more lipases,
preferably from the group of triacylglycerol lipases (3.1.1.3), or one or more
amidases,
preferably from the group of amidases (3.5.1.4) ( system component 1) which
from one
or more fatty acids present (preferably C6- C26 and particularly Cg - C16
fatty acids)
15 (system component 2) and in the presence of an oxidant such as a peroxide,
preferably
HZOZ (system component 3) can produce peracids and in the presence of ketones
as
additional component (system component 4), for example, dioxiranes.
To this end, the enzyme component system of the invention is brought together
with
20 lignin (for example, with lignosulfates and/or unevaporated or evaporated
sulfite waste
liquor and/or sulfate lignin --> kraft lignin, for example induline) and/or
with lignin-
containing material.The lignin and/or the lignin-containing material can
either be
preincubated at an elevated pH, namely above pH 8 and preferably at a pH
between 9.5
and 10.5, at 20 to 100 °C (preferably at 60 to 100 °C) after
which the pH is reduced to
25 below pH 7, depending on the optimum pH range for enzyme activity of the
component
system (ECS) or, if the activity optimum of the enzyme component system (ECS)
is on
the alkaline side, the ECS and the lignin and/or the lignin-containing
material are
brought together immediately, without pretreatment. The purpose of the
pretreatment or
treatment under alkaline pH conditions is to utilize the substantially easier
solubilization
30 of lignin at these higher pH values. This is a major advantage for the use
according to
the invention, because it is thus possible to work without an organic solvent.
CA 02335253 2000-12-15
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In other words, the main purpose of the described bringing together of the
enzyme
component system and the lignin and/or lignin-containing material is to
achieve
activation of the substrates (polyphenylpropanes) by oxidation, namely to
convert the
lignin and/or the lignin-containing material by free radical-induced
polymerization
(modification) into an activated and active binder which then when brought
together
with the wood fibers and/or wood parts to be bonded (cemented together) can be
cured
under the action of pressure and elevated temperature to give solid wood-based
composites such as the abovesaid wood products, for example fiber boards and
particle
boards. The main advantage consists of reducing, or producing savings in, the
amount
of urea-formaldehyde resins normally used, for example, for gluing in the
production of
chipboard, which resins, besides being toxic, have only limited moisture
resistance, or
of phenol-formaldehyde resins which exhibit unfavorabie swelling properties
and
require long pressing times (once again, besides the question of toxicity).
The polymerizing and/or modifying action of the enzyme component system can be
additionally enhanced by addition of certain chemical polymerization
catalysts, for
example polydiphenylmethane diisocyanate (PMDI) and other polymerization
catalysts
used also for the polymerization of lignin in lignin-containing wastewaters.
Such
substances consist of phenols, phenol derivatives or other polycyclic phenolic
compounds with a number of oxidizable hydroxyl groups, as already indicated
2o hereinabove (wastewater treatment).
IV) Use of the Enzyme Component System (ECS) as an Enzymatic Deinking
System
In principle, by deinking, which is currently always run in a conventional
manner by
flotation, is meant a two-step process.
Its objective is to remove printing ink and other dye particles from the waste
paper. The
waste paper used in most cases is paper collected domestically and consists
mainly of
newspapers and magazines.
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In the first treatment step, the dye particles adhering to the paper fibers
are removed
primarily by mechanical/chemical means. This is accomplished by "recycling"
the paper
as a uniform fibrous slurry, namely by disintegrating (comminuting) the waste
paper in
pulpers, drums or the like with simultaneous addition of chemicals capable of
enhancing
removal and preventing yellowing and thus also acting as bleaching chemicals,
namely
sodium hydroxide solution, fatty acid, water glass and hydrogen peroxide
(H202). Here,
the fatty acid acts as a fiber dye particle collector and in the second
treatment step, the
flotation, also as foaming agent.
After the waste paper has been disintegrated and the said chemicals have been
allowed
to to act for a certain length of time, the flotation is carried out in
special flotation vessels
by injecting air. During this process, the dye particles adhere to the foam
bubbles and
are removed together with the bubbles. The dye is thus separated from the
paper fibers.
Currently, this operation is preferably carried out at a neutral pH, which
makes it
necessary to use certain detergents in place of the fatty acids.
15 It is known from the literature (WO 91/14820, WO 92/20857) to use an
oxidoreductase
or laccase system characterized primarily by the addition of special
substances which
cause the optimum pH for the action of laccase obtained from Trametes
versicolor,
which normally is in the range of about pH 4-S, to shift into the slightly
alkaline range
(pH 8 to 8.7). This, on the one hand, is an important prerequisite for use in
the deinking
2o system because of the CaS04 problems arising below pH 7 and, on the other,
does not
optimize the action of laccase in the polymerizing or depolymerizing sense,
but only
produces a certain swelling of the fibers. Such swelling (which is one of the
main
actions of sodium hydroxide in purely chemical deinking systems) is a primary
performance characteristic of the dye-removing mechanism.
25 The only other additives required for this enzymatie system employing
oxidoreductases
are the detergents needed to produce foam. Nearly all suitable detergents also
exert a
dye-removing action. Moreover, in conventional deinking systems the use of
sodium
hydroxide and peroxide improves brightness as a result of the bleaching action
of these
chemicals. This bleaching action cannot be achieved with the prior-art enzyme
system
3o because of the nature of the system.
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We have now found, quite surprisingly, that by appropriate selection of the
components
the enzyme component system (ECS) of the invention exceeds the efficiency of
other
enzymatic deinking systems, particularly those with oxidoreductases and those
applied
to lignin-containing deinked pulp and at least in part compensates for the
advantage of
bleaching with purely chemical systems. This means that it is possible to
provide a
system offering environmentally friendly deinking under neutral pH conditions
and thus
better post-bleaching, better pulp properties etc and good performance similar
to that of
purely chemical systems.
In other words, according the invention, the foregoing objective is reached by
providing
1o an enzyme component system (ECS) according which contains one or more
lipases,
preferably from the group of triacylglycerol lipases (3.1.1.3), or one or more
amidases,
preferably from the group of amidases (3.5.1.4) ( system component 1) which
from one
or more fatty acids present (preferably C6- C26 and particularly Cg - C16
fatty acids)
(system component 2) and in the presence of an oxidant such as a peroxide,
preferably
15 H202 (system component 3) can produce peracids and in the presence of
ketones as
additional component (system component 4), for example, dioxiranes.
In this case, a further improvement of printing ink removal can be attained by
the
aforesald addition of special substances mostly of a phenolic nature and, in
particular,
2o containing several hydroxyl groups, which are also used as polymerization
catalysts in
enzymatic wastewater treatment and general polymerization reactions, as in the
production of binders/adhesives from lignin or lignin-containing substances
primarily
for the preparation of wood-based composites.
V) Use of the Enzyme Component System (ECS) of the Invention as an Oxidation
25 System in Organic Synthesis
Recently, enzymes have increasingly been used for chemical reactions in
organic
synthesis. A few examples showing a variety of oxidative reactions that can be
carried
out with enzymatic systems can be found in: Preparative Biotransformations
(Whole
Cell and Isolated Enzymes in Organic Synthesis), S.M. Roberts, K. Wiggins and
G.
3o Casy, J. Wiley & Sons Ltd, 1992/93; Organic Synthesis with Oxidative
Systems, H.L.
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Holland, VCH, 1992; and Biotransformations in Organic Chemistry, K. Faber,
Springer
Verlag [publisher], 1992:
1) Hydroxylation reactions
a) Synthesis
of alcohols
b) Hydroxylationof steroids
c) Hydroxylationof terpenes
d) Hydroxylationof benzenes
e) Hydroxylationof alkanes
to Hydroxylationof aromatic compounds
f)
g) Hydroxylationof double bonds
h) Hydroxylationof nonactivated methyl
groups
i) Dihydroxylation
of aromatie
compounds
2) Oxidation of unsaturated aliphatics
a) Preparation of epoxides
b) Preparation of compounds by
epoxidation
c) Preparation of arene oxides
d) Preparation of phenols
2o Preparation of cis-dihydrodiols
e)
3) Baeyer-Villiger oxidations
a) Baeyer-Villiger conversion of steroids
4) Oxidation of heterocycles
a) Transformation of organic sulfides
b) Oxidation of sulfur compounds
c) Oxidation of nitrogen compounds (formation of N-oxides etc.)
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d) Oxidation of other heteroatoms
5) Carbon-carbon dehydrogenation
a) Dehydrogenation of steroids
6) Other oxidation reactions
a) Oxidation of alcohols and aldehydes
b) Oxidation of aromatic methyl groups to aldehydes
c) Oxidative coupling of phenols
c) Oxidative degradation of alkyl chains (13-oxidation etc.)
e) Formation of peroxides or percompounds
1o f) Initiation of free-radical induced chain reactions.
Here, too, we found, quite surprisingly, that with the aid of the enzyme
component
system (ECS) of the invention it is possible to carry out many oxidation
reactions
exemplified hereinabove, namely that the aforesaid objective can be reached by
providing an enzyme component system (ECS) according to the invention which
contains one or more lipases, preferably from the group of triacylglycerol
lipases
(3.1.1.3), or one or more amidases, preferably from the group of amidases
(3.5.1.4)
( system component 1) which from one or more fatty acids present (preferably
C6- C26
and particularly Cg - C16 fatty acids) ( system component 2) and in the
presence of an
oxidant such as a peroxide, preferably H202 ( system component 3) can produce
peracids and in the presence of ketones as additional component (system
component
4), for example, dioxiranes.
VI) Use of the Enzyme Component System (ECS) in Enzymatic Coal Liquefaction
In this field, the prior art is as follows:
Preliminary studies show that, in principle, lignite and anthracite can be
attacked and
liquefied by in-vivo treatment with, for example, a white rotting fungus such
as
Phanerochaete chryosporium (incubation time: several weeksBioengineering 8, 4,
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1992). The possible structure of anthracite is a tridimensional network of
polycyclic,
aromatic ring systems with a "certain" similarity to lignin structures.
Assumed cofactors
besides the lignolytic enzymes are chelating agents (siderophors, such as
ammonium
oxalate) and biosurfactants.
1) Until now, effective coal liquefaction systems are known only as in-vivo
systems
(with lignin- degrading organisms, particularly white rotting fungi),
or as systems employing oxidoreductases plus mediators (laccase-mediator
system -
-> WO 94/29510; WO 96/18770).
2) It has been proven that, in principle, white rotting fungi that are capable
of
degrading lignin in vivo can also liquefy coal in culture.
3) Coal: Both lignite and anthracite were formed from wood by
chemical/physical
"actions"; hence, their chemical structures are at least similar to those
occurring in
lignin.
4) In coal liquefaction with white rotting fungi, we see, on the one hand, an
alkalinization of the pH during fungal growth "on coal" and, on the other, a
secretion of siderophor-like chelators, namely substances known to have a
positive
2o effect on coal liquefaction.
The main reason for economical, meaningful industrial coal liquefaction is the
industrial
demand for alternative liquid sources of energy, especially considering that
in the future
the quantities of other sources of fossil energy such as oil and gas will be
decreasing
while at the same time the demand for energy will be increasing, and that
other
alternatives such as nuclear fusion, among others, will not yet be available.
Here, too, we found, quite surprisingly, that with the aid of the enzyme
component
system (ECS) of the invention liquefaction of, for example, lignite is
possible with
better performance than with the conventional enzymatic oxidoreduetase
systems,
3o namely that the aforesaid objective can be reached by providing an enzyme
component
system (ECS) according to the invention which contains one or more lipases,
preferably
from the group of triacylglycerol lipases (3.1.1.3), or one or more amidases,
preferably
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from the group of amidases (3.5.1.4) ( system component 1) which from one or
more
fatty acids present (preferably C6- C26 and particularly Cg - C16 fatty acids)
( system
component 2) and in the presence of an oxidant such as a peroxide, preferably
HZOz
( system component 3) can produce peracids and in the presence of ketones as
additional component (system component 4), for example, dioxiranes.
VII) Use of the Enzyme Component System (ECS) as Bleaching Agent in
Detergents
1o Conventional bleaching systems in domestic detergents are unsatisfactory,
particularly
in the low- temperature range. Below a washing temperature of 60 °C,
the standard
bleaching agent, i.e., H202 / sodium perborate/sodium percarbonate must be
activated
by the addition of chemical bleach activators, such as TAED and/or SNOBS*.
Also, the
need exists for more highly biodegradable, bio-compatible bleaching systems
and
15 systems for low-temperature washing that can be used in small amounts.
Whereas
enzymes are already being used industrially for protein-starch-fat solution
and for fiber
treatment in the washing process, no enzymatic system is currently available
for
detergent bleaching. WO 1/05839 describes the use of different oxidative
enzymes
(oxidases and peroxidases) to prevent dye transfer. As is known, peroxidases
are
2o capable of "decolorizing" different pigments (3-hydroxyflavone and betaine
are
decolorized by horseradish peroxidase and carotene by peroxidase). Said patent
application describes the decolorization (also referred to as bleaching) of
textile dyes
removed from the laundered goods and present in the washing liquor (conversion
of a
colored substrate into a noncolored, oxidized substance). In this case, the
enzyme is said
25 to have the advantage over, for example, hypochlorite which attacks dyes in
or on the
fabric, in that the enzyme decolorizes only the dissolved dyes. Hydrogen
peroxide or an
appropriate precursor generating hydrogen peroxide in situ participates in the
catalysis
of the decolorization. The enzyme reaction can be enhanced somewhat by
addition of
other oxidizable enzyme substrates, for example metal ions such as Mn++,
halide ions,
such as Cl~ or Br or organic phenols, such as p-hydroxycinnamic acid and 3,4-
dichlorophenol. In this case, it is postulated that short-lived radicals or
other oxidized
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states of the added substrate are formed and are responsible for the bleaching
or other
modification of the colored substance.
US 4, 077 6768 describes the use of iron porphin, hemin chloride, iron
phthalocyanines
or derivatives thereof together with hydrogen peroxide for preventing dye
transfer.
These substances, however, are rapidly destroyed by excess peroxide, and for
this
reason hydrogen peroxide formation must occur in a controlled fashion.
Processes are known from WO 94/12619, WO 94/12620 and WO 94/12621 whereby
the activity of the peroxidase is enhanced by means of enhancers. Such
enhancers are
characterized in WO 94/12620 in terms of their half life. According to WO
94/12621,
1o enhancers are characterized by the formula A = N-N = B where N means
nitrogen and
A and B are defined cyclic groups. According to WO 94/12620, enhancers
* TAED = tetraacetylethylenediamine; SNOBS = sodium nonyloxybenzenesulfonate
are organic chemicals containing at least two aromatic rings of which at least
one is
substituted with defined groups.
All three patent applications concern dye transfer inhibition and the use of
enhancers
together with peroxidases as detergent additives or detergent compositions
used in the
detergent sector. The combination of these enhancers is limited to
peroxidases. The use
of mixtures containing peroxidases is also known from WO 92/18687. A special
system
of oxidases and of appropriate substrates such as hydrogen peroxide is
described in
German Unexamined Patent Application DE-42 31 761. German Unexamined Patent
Application DE 19 18 729 concerns another special detergent system consisting
of
glucose and glucose oxidase or of starch, aminoglucosidase and glucose oxidase
(GOD)
2s and of added hydroxylamine or a hydroxylamine compound, wherein the
hydroxylamine or the derivatives thereof serve to inhibit the catalase that is
often
present in GOD. Hydroxylamine and the derivatives thereof have definitely not
been
described as mediator additives.
Finally, WO 94/29425, DE 4445088.5 and WO 97/48786 concern multicomponent
3o bleaching systems for use with detergents and which consist of oxidation
catalysts and
oxidants and of aliphatic, cycloaliphatic, heterocyclic or aromatic compounds
containing NO-, NOH- or H-NR-OH groups.
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All hitherto known "enzymatically enhanced" detergent-bleaching systems have
the
drawback that their cleaning and bleaching action is still unsatisfactory and
that the
mediators must be used in excessive amounts which may cause environmental and
economic problems.
We have now found, quite surprisingly, that the enzyme component system (ECS)
of
the invention exceeds the performance of the aforesaid oxidoreductase-mediator
systems and does not have the said drawbacks of the prior art,
namely that the aforesaid objective can be reached by providing an enzyme
component
system (ECS) according to the invention which contains one or more lipases,
preferably
to from the group of triacylglycerol lipases (3.1.1.3), or one or more
amidases, preferably
from the group of amidases (3.5.1.4) ( system component 1) which from one or
more
fatty acids present (preferably C6- C26 and particularly Cg - C16 fatty acids)
( system
component 2) and in the presence of an oxidant such as a peroxide, preferably
H202
( system component 3) can produce peracids and in the presence of ketones as
15 additional component (system component 4), for example, dioxiranes.
VIII) Use of the Enzyme Component System of the Invention in the Bleaching
and/or Decolorization of Textile Fabrics
2o Enzymes are currently being used to an increasing extent in various
applications in the
textile industry, For example, the use of amylases in the desizing process is
of great
importance, because the use of strong acids, alkalies or oxidants is thereby
avoided.
Similarly, cellulases are used for biopolishing and biostoning, a process
which is mostly
employed together with conventional stone washing with pumice in the treatment
of
25 denim fabrics for jeans to remove the indigo dye. WO 94/29510, WO 96/18770,
DE 196
12 194 Al and DE 44 45 088 Al describe enzymatic delignification processes
which use
enzymes together with mediators. In general, the disclosed mediators are
compounds
with the NO-, NOH- or HRNOH structure. These systems, of course, are
restricted to
use in pulp bleaching. Because the mechanisms underlying lignin-removing pulp
3o bleaching, and this is the process involved here, are entirely different
from those
underlying the decolorization, removal and/or "destruction" of denim dyes,
particularly
indigo dyes, in the jeans producing sector, it is entirely surprising that a
number of
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substances of the said NO-, NOH- and HNROH types are also suitable for this
application.
WO 97/06244 describes systems for the bleaching of pulp, for dye transfer
inhibition
and for bleaching stains when used with detergents, which systems employ
enzymes
(peroxidases, laccases) and enzyme-enhancing (hetero-)aromatic compounds, such
as
nitroso compounds etc.. In this case, as in patents WO 94/12619, WO 94/12620
and
WO 94/12621, only the above-described use is intended. The mechanisms of stain
decolorization in detergent bleaching or of dye transfer inhibition are
entirely different
from those underlying the decolorization, removal and/or "destruction" of
indigo dyes,
1o as, for example, in denim treatment. Hence, it is quite surprising that a
number of
substances of the said NO-, NOH- and HNROH-types are also suitable for this
application.
Processes are known from said WO 94/12619, WO 94/12620 and WO 94/12621 in
which the activity of peroxidase is increased by use of enhancers. Such
enhancers are
characterized in WO 94/12620 in terms of their half life. According to WO
94/12621,
enhancers are characterized by the formula A=N-N=B where N means nitrogen and
A
and B are defined cyclic groups. According to WO 94/12621, enhancers are
organic
chemicals containing at least two aromatic rings of which at least one is
substituted with
defined groups. All three applications concern (as already stated) dye
transfer inhibition
2o and the use of enhancers together with peroxidases as detergent additives
or detergent
compositions for washing applications or in cellulose bleaching. The
combinations of
these enhancers are restricted to use with peroxidases.
Moreover, oxidoreductases, primarily laccases, but also peroxidases, are
currently used,
mainly for treating denim for jeans.
It is known from patent application WO 96/12846 that laccase and peroxidase +
certain
enhancers, mainly derivatives of phenothiazine or phenoxazine, are used in two
application forms in the treatment of cellulose-containing fabrics, such as
cotton,
viscose, rayon (artificial silk), ramie, linen, Tencelm, silk or mixtures
thereof or
mixtures of these fabrics with synthetic fibers, for example a mixture of
cotton and
3o spandex (stretch denim), but mainly denim fabrics (mainly for use in
jeans).
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On the one hand, the system (oxidoreductases + enhancers) is intended as a
replacement
for the conventional hypochlorite bleaching of denim, usually after a stone
washing
pretreatment, this enzymatic treatment providing only partial replacement of
hypochlorite, because the desired bleaching erect cannot be attained.
On the other hand, the system can be used together with cellulase in stone
washing in
place of the usual mechanical treatment with pumice, and this represents an
improvement over the "treatment with cellulase only".
The main drawbacks of the system described in WO 96/12846 are the following,
among
others:
I ) To achieve the desired goal, laccase must be used in considerable amounts
(about
10 international units [ICJ]/g of denim);
2) In some cases, optimum treatment requires 2-3 hours;
3) The preferred mediator (here phenothiazine-10-propionic acid) must be used
in an
amount of about 2 to about 14 mg per gram of denim, which represents a
considerable cost;
4) Buffer systems (about 0.1 mol/L) must be used, because otherwise no
performance can be achieved, and this also raises the cost of the system.
This, for
example, is not required for the system of the invention;
5) The color of the enhancer component (long-lived radical) causes "browning"
of
the fabric.
The general advantage of a laccase and/or oxidoreductase system with enzyme
action-
enhancing compounds (enhancers, mediators etc.) when used in the above-
described
treatment of textiles (for example, jeans fabrics) consists, in an improved
system over
the prior-art systems, in that fashion looks can be achieved, which is not
possible with
conventional hypochlorite bleaching.
The dyes normally used for jeans denim are vat dyes, such as indigo or indigo
derivatives, for example thioindigo, as well as sulfur dyes. By use of such
special
enzymatic systems, it is possible (as a result of the high specificity of such
systems),
3o when a mixed dye system such as an indigo dye and a sulfur dye system is
present, to
decolorize only the indigo dye, while the sulfur dye is not oxidized.
Depending on the
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enzyme action-enhancing compound used, this can produce almost any desired
fabric
color (for example, gray shades etc), which is often desirable.
An additional advantage is that the enzymatic treatment is substantially more
gentle
than bleaching with hypochlorite, and as a result fiber damage is reduced.
In the stone washing process, the ecological effect is of particular
importance (in
addition to the reduced fiber damage caused by enzymes) considering, for
example, that
this purely mechanical process produces about 1 kg of stone sludge per kg of
jeans
denim.
As can be seen from the prior art, for colored fabrics, in particular, the
textile industry
Io has a great need for alternative bleaching processes (alternatives to
conventional
hypochlorite bleaching) and/or treatment methods as alternatives to stone
washing to
achieve the bleached look, in the latter case also because of the
environmental pollution
problems.
The present invention has for an object to minimize or eliminate the drawbacks
of the
15 conventional processes: stone washing/bleaching after stone washing or
general
bleaching of dyed and/or undyed textile fabrics, particularly the pollution
problems and
fiber damage, as well as the drawbacks of the known oxidoreductase/enhancer
systems
(for example also NO-radical formation etc).
Entirely surprisingly, we have now found that the enzyme component System
(ECS) of
2o the invention exceeds the performance of the aforesaid oxidoreductase-
mediator sytems
and that it does not exhibit the said drawbacks of the prior art.
In other words, according the invention, the foregoing objective is reached by
providing
an enzyme component system (ECS) according which contains one or more lipases,
preferably from the group of triacylglycerol lipases (3.1.1.3), or one or more
amidases,
25 preferably from the group of amidases (3.5.1.4) ( system component 1) which
from one
or more fatty acids present (preferably C6- Cz~ and particularly Cg - C16
fatty acids)
(system component 2) and in the presence of an oxidant such as a peroxide,
preferably
HZOz (system component 3) can produce peracids and in the presence of ketones
as
additional component (system component 4), for example, dioxiranes.
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Description of Individual System Components of the Enzyme Component System
(ECS) of the Invention
System Component 1 (Liaases and Other Enzymes
Preferred are enzymes of group 3 (hydrolases), 3.1, 3.1.1, 3.1.2, 3.1.3, 3.1.4
and 3.1.7
according to the International Enzyme Nomenclature: Committee of the
International
Union of Biochemistry and Molecular Biology (Enzyme Nomenclature, Academic
Press, Inc., 1992, pp. 306-337).
1o Preferred are enzymes acting on ester bonds (3.1, particularly those acting
on
carboxylate esters (3.1.1):
A) Carboxylate
Ester Hydrolases
(3.1.1)
3.1. 1. Carboxylate esterase
l
3.1.1.2 Aryl esterase
3.1.1.3 Triacylglycerollipase
3.1.1.4 Phospholipase AZ
3.1.1.5 Lysophospholipase
3.1.1.6 Acetyl esterase
3.1.1.7 Acetylcholine esterase
3.1.1.8 Choline esterase
3.1.1.10 Tropine esterase
3.1.1.11 Pectin esterase
3.1.1.13 Sterol esterase
3.1.1.14Chlorophyllase
3.1.1.15 L-Arabinolactonase
3.1.1.17 Gluconolactonase
3.1.1.19 Uronolactonase
3.1.1.20 Tannase
3.1.1.21Retynil palmitate esterase
3.1.1.22 Hydroxybutyrate dimer
hydrolase
3.1.1.23 Acylglycerollipase
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3.1.1.24 3-Oxoadipate enol lactonase
3.1.1.25 1,4-Lactonase
3.1.1.26 Galactolipase
3.1.1.27 4-Pyrodoxolactonase
3.1.1.28 Acylcarnitine hydrolase
3.1.1.30 D-Arabinone lactonase
3 .1.1.316-Phosphogluconolactonase
3.1.1.32 Phospholipase Al
3.1.1.32 6-Acetylglycose deacetylase
l03.1.1.34 Lipoprotein lipase
3 .1.1.3 Dihydrocoumarin hydrolase
5
3.1.1.36 Limonine D-ring lactonase
3.1.1.37 Steroid lactonase
3 .1.1.3 Triacetate lactonase
8
153.1.1.39 Actinomycin lactonase
3.1.1.40 Orseilinate depside hydrolase
3.1.1.41 Cephalosporin C deacetylase
3.1.1.42 Chlorogenate hydrolase
3.1.1.43 oc-Amino acid esterase
203.1.1.44 Methyloxaloacetate esterase
3.1.1.45 Carboxymethylenebutenolidase
3.1.1.46 Deoxylimonate A-ring lactonase
3.1.1.47 1 -Alkyl-2-acetylglycerophosphocholine
esterase
3.1.1.48 Fusarinine C ornithine esterase
253.1.1.49 Sinapine esterase
3.1.1.50 Wax ester hydrolase
3.1.1.51 Phorbol diester hydrolase
3.1.1.52 Phosphatidylinositol deacetylase
3.1.1.53 Sialate O-acetyl esterase
303.1.1.54 Acetoxybutynylbithiophene deacetylase
3.1.1.55 Acetylsalicylate deacetylase
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3.1.1.56 Methylumbeiliferyl acetate deacetylase
3.1.1.57 2-Pyrone-4,6-dicarboxygallate lactonase
3.1.1.58 N-Acetylgalactosaminoglycan deacetylase
3.1.1.59 Juvenile hormone esterase
3.1.1.60 Bis(2-ethythexyl)phthalate esterase
3.1.1.61 Protein glutamate methylesterase
3.1.1.63 11 -cis-Retynil palmitate hydrolase
3.1.1.64 all-trans-Retynil paimitate hydrolase
3.1.1.65 L-Rhamnono-1,4-lactonase
3.1.1.665-(3,4-diacetoxybutynyl)-2,2'-bitiophene
deacetylase
3.1.1.67 Fatty acid ethyl ester synthase
3.1.1.68 Xylono-1,4-lactonase
3.1.1.69 N-Acetylglucosaminylphosphatidylinositol
deacetylase
3.1.1.70 Cetraxate benzyl esterase
Also preferred are:
B) Thiol ester hydrolases (3.1.2)
3.1.2.6 Hydroxyacylglutathione hydrolase
3.1.2.7 Glutathione thiol esterase
3.1.2.12 S-Formylglutathione hydrolase
3.1.2.13S-Succinylglutathione hydrolase
3.1.2.14 Oleoyl-(acyl carrier protein)
hydrolase
3.1.2.15 Ubiquitin thiol esterase
3 .1.2.16 (Citrate-(pro-3 S)-lyase)thiol esterase.
Also preferred are:
C) Phosphoric Acid Monoester Hydrolases (Phosphatases) (3.1.3)
3.1.3.1 Alkaline phosphatase
3.1.3.2 Acid phosphatase
3.1.3.3 Phosphoserine phosphatase
3.1.3.4 Phosphatidate phosphatase
CA 02335253 2000-12-15
-29-
3.1.3.8 3-Phytase
3.1.3.9 Glucose-6-phosphatase
3.1.3.10 Glucose-1-phosphatase
3.1.3.11 Fructose bisphosphatase
3.1.3.12 Trehalose phosphatase
3.1.3.13 Bisphosphoglycerate phosphatase
3.1.3.14 Methylphosphothioglycerate
phosphatase
3.1.3.15 Histidinol phosphatase
3.1.3.16 Phosphoprotein phosphatase
l0 3.1.3.17(Phosphorylase) phosphatase
3.1.3.18 Phosphoglycolate phosphatase
3.1.3.19 Glycerol-2-phosphatase
3.1.3.20 Phosphoglycerate phosphatase
3.1.3.21 Glycerol-1-phosphatase
~5 3.1.3.22Mannitol-1-phosphatase
3.1.3.23 Sugar phosphatase
3.1.3.24 Sucrose phosphatase
3.1.3.25 Myoinositol-1 (or 4)-monophosphatase
3.1.3.26 6-Phytase
20 3.1.3.27 Phosphatidylglycerophosphatase
3.1.3.36 Phosphatidylinositol bisphosphatase
3.1.3.37 Sedoheptulose bisphosphatase
3.1.3.38 3-Phosphoglycerate phosphatase
3.1.3.39 Streptomycin-6-phosphatase
25 3.1.3.40 Guanidinodeoxy-scyllo-inositol-4-phosphatase
3.1.3.41 4-Nitrophenyl phosphatases
3.1.3.42 (Glycogen synthase-D) phosphatase
3.1.3.43 (Pyruvate dehydrogenase (lipoamide) phosphatase
3.1.3.44 3-Deoxy-manno-octulosonante-8-phosphatase
30 3.1.3.46 Fructose-2,6-biphosphate-2-phosphatase
3.1.3.48 Protein-tyrosine phosphatase
CA 02335253 2000-12-15
-30-
3.1.3.49 (Pyruvate kinase) phosphatase
3.1.3.50 Sorbitol-6-phosphatase
3.1.3.51 Dolichyl phosphatase
3.1.3.52 3-Methyl-2-oxobutanoate dehydrogenase) (lipoamide)
phosphatase
3.1.3.53 Myosin light chain phosphatase
3.1.3.54 Fructose-2,6-bisphosphate-6-phosphatase
3.1.3.55 Caldesmon phosphatase
3 .1.3. Inositol-1,4, 5-triphosphate-B-phosphatase
56
3.1.3.57 Inositol-1,4-bisphosphate-1-phosphatase
3.1.3.58Sugar terminal phosphatase
3.1.3.59 Alkylacetylglycerophosphatase
3.1.3.60 Phosphoenolpyruvate phosphatase
3 .1.3 .61 Inositol-1,4, 5-trisphosphate-1-phosphatase
3 .1.3.62 Inositol-1, 3,4, 5-tetrakisphosphate-3-phosphatase
3.1.3.632-Carboxy-D-arabinitol-l-phosphatase
3.1.3.64 Phosphatidylinositol-3-phosphatase
3 .1.3.65 Inositol-1 3-bisphosphate-3-phosphatase
3 .1.3 .66 Inositol-3,4-bisphosphate-4-phosphatase.
Also preferred are:
2o D) Phosphoric Acid Diester Hydrolases (3.1.4)
3.1.4.1 Phosphodiesterase I
3.1.4.2 Glycerophosphocholine phosphodiesterase
3.1.4.3 Phospholipase C
3.1.4.4 Phosphollpase D
3.1.4.101 -Phosphatidylinositol phosphodiesterase
3.1.4.11 1 -Phosphatidylinositol-4,5-bisphosphate
phosphodiesterase
3.1.4.12 Sphingomyelin phosphodiesterase
3.1.4.13 Serine-ethanolamine phosphate phosphodiesterase
3.1.4.14 (Acyl carrier protein) phosphodiesterase
CA 02335253 2000-12-15
-31 -
3.1.4.36 1,2-Cyclie inositol phosphate phosphodiesterase
3.1.4.38 Glycerophosphocholine choline phosphodiesterase
3.1.4.39 Alkylglycerophosphoethanolamine phosphodiesterase
3.1.4.40 CMP-N-acylneuraminate phosphodiesterase
3.1.4.41Sphyngomyelin phosphodiesterase D
3.1.4.42 Glycerol-1,2-cyclic phosphate-2-phosphodiesterase
3.1.4.43 Glycerophosphoinositol inositol phosphodiesterase
3.1.4.44 Glycerophosphoinositol glycerophosphodiesterase
3.1.4.45 N-acetylglucosamine-1-phosphodiesterase
3.1.4.46Glycerophosphodiester phosphodiesterase
3.1.4.47 Variant surface glycoprotein phospholipase C
3.1.4.48 Dolochyl phosphate-glucose phosphodiesterase
3.1.4.49 Dolochyl phosphate-mannose phosphodiesterase
3.1.4.50 Glycoprotein phospholipase D
3.1.4.51 Glucose-1-phospho-D-mannosylglycoprotein phosphodiesterase.
Also preferred are:
E) Diphosphoric Acid Monoster Hydrolases (3.1.7)
3.1.7.1 Prenyl pyrophosphatase
3.1.7.3 Monoterpenyl pyrophosphatase
2o Particularly preferred among these are enzymes of group 3.1.1.3 lipases
(triacylglycerol
lipases, triglycerolacyl hydrolases) from organisms such as Candida
antarctica, Candida
rugosa, Candida lipolytica, Candida cylindracae, Candida spec., Geotrichum
candidum,
Humicula lanuginosa, Penicillium cambertii, Penicillium roqufortii,
Aspergillus spec.,
Mucor javanicus, Mucor mehei, Rhizopus arrhizus, Rhizopus niveus, Rhizopus
delamar,
Rhizopus spec., Chromobacterium viscosum, Pseudomonas cepacia and Pseudomonas
spec. from wheat seedlings or pancreas (pig or other sources) or other
sources.
CA 02335253 2000-12-15
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Enzymes salittin$ carbon/nitro~en (C/N) bonds (other than peutide bonds) can
also be used (3.5).
This subclass includes enzymes capable of splitting amides, amidines and other
C/N
bonds. Particularly preferred are enzymes of class 3.5.1 which act on linear
amides, of
class 3.5.2 which act on cyclic amides, of class 3.5.3 which act on linear
amidines, of
class 5.3.1 which act on nitrites and of class 3.5.99 which act on other
compounds.
Particularly
preferred
are enzymes
of class
3.5.1.
which act
on linear
amides:
3.5.1.1 Asparaginase
3.5.1.2 Glutaminase
l0 3.5.1.3 ~-Amidase
3.5.1.4 Amidase
3.5.1.5 Urease
3.5.1.6 13-Ureidopropionase
3.5.1.7 Ureidosuccinase
3.5.1.8 Formylaspartate deformylase
3.5.1.9 Arylformamidase
3.5.1.10 Formyltetrahydrofolate deformylase
3.5.1.11 Penicillin amidase
3.5.1.12 Biotinidase
3.5.1.13Arylacyl amidase
3.5.1.14 Aminoacylase
3.5.1.15 Aspartoacylase
3.5.1.16 Acetylornithine deacetylase
3.5.1.17 Acyl-lysine deacetylase
3.5.1.18Nicotinamidase
3.5.1.20 Citrullinase
3.5.1.22 Pantothenase
3.5.1.30 5-Aminopentanamidase
3.5.1.31 Formylmethionine deformylase
CA 02335253 2000-12-15
- 33 -
3.5.1.32 Hippurate hydrolase
3.5.1.39 Alkylamidase
3.5.1.40 Acylagmatin amidase
3.5.1.41 Chitin deacetylase
s 3.5.1.42Nicotinamide nucleotide amidase
3.5.1.49 Formamidase
3.5.1.50 Pentanamidase
3.5.1.55 Long-chain fatty acylglutamate
deacylase
3.5.1.56 N,N-Dimethylformamidase
l0 3.5.1.57Tryptophanamidase
3.5.1.58 N-Benzyloxycarbonylglycine
hydrolase
3.5.1.59 N-Carbamoylsarcosine amidase
3.5.1.72 D-Benzoylarginine-4-nitroanilide
amidase
3.5.1.73 Carnitine amidase
15 3.5.1.75Urethanase
Also preferred are enzymes of class 3.5.2 which act on cyclic amides, such as:
3.5.2.1 Barbiturase
3.5.2.2 Dihydropyrimidase
3.5.2.3. Dihydroorotase
20 3.5.2.4 Carboxymethylhydantoinase
3.5.2.5 Allantoinase
3.5.2.6 f3-Lactamase
3.5.2.10 Creatininase
Also particularly preferred are class 3.5.3 enzymes which act on linear
amidines, such
2s as:
3.5.3.1 Arginase
3.5.3.3 Creatinase
3.5.3.4 Allantoinase
CA 02335253 2000-12-15
-34-
3.5.3.6 Arginine deiminase
3.5.3.9 Allantoate deiminase
3.5.3.10 D-Arginase
3.5.3.14 Amidinoaspartase
3.5.3.15 Protein-arginine deiminase
Also particularly preferred are enzymes of class 3.5.4 which act on cyclic
amidines,
such as:
3.5.4.8 Aminoimidazolase
3.5.4.21 Creatinine deaminase
to
Preferred are also the enzymes of class 3.5.99 which act on other compounds,
such as:
3.5.99.1 Riboflavinase
3.5.99.2 Thaminase
Particularly preferred enzymes are especially those of class 3.5.5. l,
nitrilase (3.5.5.2 -
3.5.5.6, other nitrilases).
Also particularly preferred are enzymes of class 3.5.1, here particularly
those of class
3.5.1.4, amidases.
System Comuonent 2 of the Enzyme Comuonent System (ECS) of the Invention
Fatty acids which in the process according to the invention can be used as
sources
of peracids are, for example:
I) Saturated fatty acids
Butanoic acid (butyric acid)
Pentanoic acid (valeric acid)
Hexanoic acid (caproic acid)
Heptanoic acid (enanthic acid)
CA 02335253 2000-12-15
Octanoic acid (caprylic
acid)
Nonanoic acid (pelargonic
acid)
Decanoie acid (capric acid)
Undecanoic acid
Dodecanoic acid (lauric acid)
Tridecanoic acid
Tetradecanoic acid(myristic
acid)
Pentadecanoic acid
Hexadecanoic acid (palmitic
acid)
1o Heptadecanoic
acid
Octadecanoic acid (stearic acid)
Nonadecanoic acid
Eicosanoic acid (arachic acid)
Heneicosanoic acid
Docosanoic acid(behenic acid)
Tricosanoic acid
Tetracosanoic acid(lignoceric
acid)
Pentacosanoic acid
Hexacosanoic acid (cerotic acid)
2o Octacosanoic
acid
Triacontanoic acid(melissic
acid)
2) Unsaturated fatty acids
10-Undecenoic acid
9-cis-Dodecenoic acid (lauroleic acid)
9-cis-Tetradecenoic acid (myristoleic acid)
9-cis-Hexadecenoic acid (paimitoleic acid)
6-cis-Octadecenoic aeid (petroselic acid)
6-trans-Octadecenoic acid (petroselaidic acid)
9-cis-Octadecenoic acid (oleic acid)
9-trans-Octadecenoic acid (elaidic acid)
CA 02335253 2000-12-15
-36-
9-cis, 12 cis-Octadecadienoic acid (linoleic acid)
9-trans, 1 2-trans-Octadecadienoic acid(linolaidic acid)
9-cis, 12-cis,15-cis-Octadecatrienoic (linolenic acid)
acid
9-trans, 11 -trans, 1 3-trans- Octadecatrienoic(a-eleostearic
acid acid)
9-cis, 11 -trans, 1 3-trans-Octadecatrienoic(13-eleostearic
acid acid)
9-cis-Icosenic acid (gadoleic acid)
Icosa-5,8,11,14-tetraenoic acid (arachidic acid)
13-cis-Docosenoic acid (erucic acid)
13-trans-Docosenoic acid (brassidic acid)
4, 8,12,15,19-Docosapentaenoic acid (clupanodonic
acid)
3) Polyunsaturated fatty acids
9,12-Octadecadienoic acid (linoleic acid)
9,12,1 5-Octadecatrienoic acid (linolenic acid)
5,9,1 2-Octadecatrienoic acid
9,11,13-Octadecatrienoic acid (eleostearic acid)
9,11,13,15-Octadecatetraenoic acid (parinaric acid)
5,11,14-Icosatrienoic acid
5,8,11,1 4-Icosatetraenoic acid (arachidic acid)
4,8,12,1 S,1 8-Icosapentaenoie acid
4, 8,12,15,19-Docosapentaenoic acid (clupanodonic acid)
4,8,12,15,18,21 -Tetracosahexaenoic (nisinic acid)
acid
Particularly preferred are tetradecanoic acid (myristic acid) and dodecanoic
acid (lauric
acid).
System Component 3 (Oxidants: Peroxides or Per Comuounds) of the Enzvme
Comuonent Svstem (ECS) of the Invention
CA 02335253 2000-12-15
-37-
Preferred oxidants in the enzyme component system of the invention are
hydrogen
peroxide (H202) , organic peroxides and per-compounds such as perborates,
persulfates,
percarbonates, perphosphates, percarbamides, perchlorates etc.
Preferred organic peroxides are, for example:
3-chloroperoxybenzoic acid, monoperoxyphthalic acid Mg salt, di-tert.butyl
peroxide,
cumene hydroperoxide, lauroyl peroxide, chloroperoxybenzoic acid, dicumyl
peroxide,
methyl ethyl ketone peroxide, benzoyl peroxide, diperoxidododecandionic acid
Na salt
etc..
to Besides lipase-catalyzed peracid formation, combinations of bleach
activators, such as
TAED (tetraacetylethylenediamine), TAGU (tetraacetylglycoluril) and iso-NOBS
(sodium p-isononanoyl-oxybenzenesulfonate) and the like, which are also used
in
detergents, together with per-compounds such as perborates, percarbonates etc.
can
serve as additional sources of peracid generation.
15 The abovesaid per-compounds, as well as, for example, glucose + GOD, can be
used as
systems generating H202 for the corresponding lipase action. Substances such
as
nitrilamines or dicyandiamines or metal ions, e.g. Mo6+, Va6+ and W6-+ can be
used
together with a peroxide, for example HZO2.
2o System Component 4 (Ketones) of the Enzvme Comuonent Svstem (ECS) of the
Invention
Particularly preferred are carbonyl compounds of general formula I:
O
R~R2
3o The Rl and RZ groups can be equal or different and denote aliphatic or
aromatic groups.
Moreover, the Rl and Rz groups can form a ring containing besides carbon also
heteroatoms such as nitrogen, oxygen and sulfur.
CA 02335253 2000-12-15
-3g-
Particularly preferred are 1,2-diketones (formula II), 1,3-diketones (formula
III),
polyketones (polyketides) and the tautomeric enols (formula IV):
O
3~ R4 O O O OH
R ii ~ _
O R R6 ~_ R / R6
II III IV
1o wherein the R3 to R6 groups, once again, can be equal or different and
denote aliphatic
or aromatic groups. Moreover, groups R3 and R4 and groups RS and R6, together,
can
form a ring containing besides carbon also heteroatoms such as nitrogen,
oxygen or
sulfur. The possibility of tautomerization or formation of a resonance hybrid
is
particularly important in this case.
15 Besides general carbonyl compounds, particularly preferred are ketones,
such as, in
general hydroxyketones, oc,f3-unsaturated ketones, oxycarboxylic acids,
quinones and
halogenated ketones.
Particularly preferred among these are the following:
Acetone, methyl ethyl ketone, diethyl ketone, methyl n-butyl ketone, methyl
isobutyl
ketone, cyclohexanone, cyclopentanone, 2-methylcyclohexanone, 3-
methylcyclohexanone, 4-methylcyclohexanone, dihydroxyacetone, diacetyl
monohydrazone, diacetyl dihydrazone, acetophenone, p-hydroxyacetophenone,
1 -phenyl-3-butanone, 3-pentanone, 4-heptanone, 2-nonanone, cycloheptanone,
cyclooctanone, cyclodecanone, cyclododecanone, dimethyl ketone, ethyl propyl
ketone,
methyl amyl ketone, acetylacetone, pinacoline, methyl isopropyl ketone, methyl
isoamyl ketone, ethyl amyl ketone diisopropyl ketone, diisobutyl ketone,
methyl vinyl
ketone, methyl isopropenyl ketone, mesityl oxide, isophorone, hydroxyacetone,
CA 02335253 2000-12-15
-39-
methoxyacetone, 2,3-pentanedione, 2,3-hexanedione, phenylacetone,
propiophenone,
benzophenone, benzoin, benzil, 4,4'-dimethoxybenzil, 4'-methoxyacetophenone,
3'- methoxyacetophenone, O-ethylbenzoin, (2-methoxyphenyl)acetone, (4-
methoxyphenyl)acetone, methoxy-2-propanone, glyoxylic acid, benzyl glyoxylate,
benzylacetone, methyl benzyl ketone, methylcyclohexyl ketone, 2-decanone,
dicyclohexyl ketone, 3,3-dimethyl-2-butanone, methyl isobutyl ketone, methyl
isopropyl ketone,2-methyl-3-heptanone, 5-methyl-3-heptanone, 6-methyl-5-hepten-
2-
one, 5-methyl-2-hexanone, 3-nonanone, 5-nonanone, 2-octanone, 3-octanone, 2-
undecanone, 1,3- dichloroacetone, I-hydroxy-2-butanone, 3-hydroxy-2-butanone,
4-
to hydroxy-4-methyl-2-pentanone, 2-(1S)- adamanantone, anthrone,
bicyclo(3.2.0)hept-2-
en-6-one, cis-bieyclo(3.3.0)octan-3,7-dione, (1 S)- (-)-camphor, p-chloranil,
cyclobutanone, 1,3-cyclohexanedione, 1,4-cyclohexanedione monoethylene ketal,
dibenzosuberone, ethyl 4-oxocyclohexanecarboxylate, 9-fluorenone, 1,3-
indandione,
methyl- cyclohexanone, phenylcyclohexanone, 4-propylcyclohexanone, 1,2,3,4-
15 tetrahydro-1-naphthalenone, 1,2,3,4-tetrahydro-2-naphthalenone, 3,3,5-
trimethylcyclo-
hexanone, 3-acetoxy-2-cyclohexen-1-one, benzylideneacetone, (R)-(-)-carvone,
(S)-(-)carvone, curcumin, 2-cyclohexen-1-one, 2,3-diphenyl-2- cyclopropen-1-
one, 2-
hydroxy-3-methyl-2-cyclopentene-1-one, isophorone, a-ionone, 13-ionone, 3-
methoxy-2-
cyclohexen-1-one, 3-methyl-2-cyclopenten-1-one, 3-methyl-3-penten-2-one, (R)-
(+)-
2o pulegone, tetraphenyl-2,4-cyclopentadien-1-one, 2,6,6-trimethyl-2-
cyclohexen-1,4-
dione, 2-acetylbenzoic acid, 1-acetylnaphthalene, 2-acetylnaphthalene,
3'-aminoacetophenone, 4'-aminoacetophenone, 4'-cyclohexylacetophenone, 3',4'-
diacetoxyacetophenone, diacetylbenzene, 2',4'-dihydroxyacetophenone, 2',5'-
dihydroxyacetophenone, 2',6'-dihydroxyacetophenone, 3,4-dimethoxyacetophenone,
25 2'-hydroxyacetophenone, 4'-hydroxyacetophenone, 3'-methoxyacetophenone, 4'-
methoxyacetophenone, 2'-methylacetophenone, 4'-methylacetophenone,
2'-nitroacetophenone, 3'-nitroacetophenone, 4'-phenylacetophenone, 3,'4',5'-
trimethoxy-
acetophenone, 4'-aminopropiophenone, benzoylacetone, benzoylpropionic acid,
benzylideneacetophenone, cyclohexyl phenyl ketone, desoxybenzoin, 4',4'-
30 dimethoxybenzil, 1,3-diphenyl-1,3-propanedione, ethylbenzoyl acetate, ethyl
phenylglyoxylate, 4'- hydroxypropiophenone, 1,3-indandione, I-indanone,
isopropyl
phenyl ketone, 6-methoxy-1,2,3,4- tetrahydronaphthalen-1-one, methylphenyl
CA 02335253 2000-12-15
-40-
glyoxylate, phenylglyoxylonitrile, 1-phenyl-1,2-propanedione 2-oxime,
valerophenone,
2-acetyl-y-butyrolactone, 2-acetylpyrrole, 1-benzylpiperidin-4-one,
dehydroacetic acid,
3,4-dihydro-4,4-dimethyl-2H-pyran-2-one, 1,4-dihydro-4-pyridinone, N-eth-
oxycarbonyl-4-piperidinone, 2-methyl furyl ketone, 5-hydroxy-2-hydroxymethyl-
4H-
pyran-4-one, 3-hydroxy-2-methyl-4-pyranone, 3-indolyl methyl ketone, isatin, 1-
methyl-4-piperidinone, methyl 2-pyridyl ketone, methyl 3-pyridyl ketone,
methyl 4-
pyridyl ketone, methyl 2-thienyl ketone, phenyl 2-pyridyl ketone, phenyl 4-
pyridyl
ketone, tetrahydrofuran-2,4-dione, tetrahydro-4H-pyran-4-one, 2,2,6,6-
tetramethyl-4-
piperidone, xanthone, acenaphthene quinone, pyruvic acid, (1 R)-(-)-camphor
quinone,
to (1S)-(+)-camphor quinone, 3,5-ditert.butyl-o-benzoquinone, 1,2-dihydroxy-
3,4-
cyclobutendione, ethyl (2-amino-4-thiazolyl)glyoxylate, ethyl pyruvate, 2,3-
hexanedione, 3,4- hexanedione, 3-methyl-2-oxobutyric acid, 3-methyl-2-
oxovaleric
acid, 4-methyl-2-oxovaleric acid, 2- oxobutyric acid, 2,3-pentandione, 9,10-
phenanthrene quinone, acetoacetanilide,2-acetyl-y-butyrolactone, 2-acetylcyclo-
pentanone, allyl acetoacetate, benzoylacetone, tert.butyl acetoacetate, 1,3-
cyclopentanedione, diethyl 3-oxoglutarate, dimethyl acetylsuccinate, dimethyl
3-
oxoglutarate, 1,3-diphenyl-1,3-propanedione, ethyl acetoacetate, ethyl
benzoylacetate,
ethyl butyrylacetate, ethyl 2-oxocyclohexanecarboxylate, ethyl 2-
phenylacetoacetate,
methyl acetoacetate, 2-methyl-1,3- cyclohexanedione, 2-methyl-1,3-
cyclopentanedione,
2o methyl isobutyrylacetate, methyl 3-oxopentanoate, methyl pivaloylacetate, 3-
oxoglutaric acid, tetrahydrofuran-2,4-dione, 2,2,6,6-tetramethyl-3,5-
heptanedione, 3-benzoylpropionic acid, 1,4-cyclohexanedione, dimethyl
acetylsuccinate, ethyl levulinate, 2-aminoanthraquinone, anthraquinone, p-
benzoquinone, 1,4-dihydroxyanthraquinone, 1,8-dihydroxyanthraquinone,
2-ethylanthraquinone, methyl-p-benzoquinone, 1,4-naphthoquinone, tetramethyl-p-
benzoquinone, 2,2-dimethyl-1,3-dioxan-4,6-dione, 2-benzoylbenzoic acid, 3-
benzoyl-
propionic acid, 5,6-dimethoxyphthataldehydic acid, levulinic acid, methyl
traps-4-oxo-
2-pentenoate, phthalaldehydic acid, terephthalaidehydic acid, dibutyl maleate,
dibutyl
succinate, dibutyl phthalate, dicyclohexyl phthalate, diethyl
acetamidomalonate, diethyl
3o adipate, diethyl benzylmalonate, diethyl butylmalonate,
diethylethoxymethylene-
malonate, diethyl ethylmalonate, diethyl fumarate, diethyl glutarate, diethyl
isopropylidenemalonate, diethyl maleate, diethyl malonate, diethyl
methylmalonate,
CA 02335253 2000-12-15
-41 -
diethyl oxalate, diethyl 3-oxoglutarate, diethyl phenylmatonate, diethyl
phthalate,
diethyl pimelate, diethyl sebacate, diethyl suberate, diethyl succinate,
diisobutyl
phthalate, dimethyl acetylene- dicarboxylate, dimethyl acetylsuccinate,
dimethyl
adipate, dimethyl 2-aminoterephthalate, dimethyl fumarate, dimethyl
glutaconate,
dimethyl glutarate, dimethyl isophthalate, dimethyl malonate, dimethylmethoxy-
malonate, dimethyl methylenesuccinate, dimethyl oxalate, dimethyl 3-
oxoglutarate,
dimethyl phthalate, dimethyl succinate, dimethyl terephthalate, ethylene
glycol
diacetate, ethylene glycol dimethacrylate, monoethyl fumarate, monomethyl
malonate,
monoethyl adipate, monomethyl phthalate, monomethyl pimelate, monomethyl
to terephthalate, 1,2-propylene glycol diacetate, triethyl
methanetricarboxylate, trimethyl
1,2,3-propanetricarboxylate, 3-acetoxy-2-cyclohexen-1-one, allyl acetoacetate,
allyl
cyanoacetate, benzyl acetoacetate, tert.butyl acetoacetate, butyl
cyanoacetate,
chlorogenic acid hemihydrate, coumarin-3-carboxylic acid, diethyl ethoxy-
carbonylmethanephosphonate, dodecyl gallate, dodecyl 3,4,5-trihydroxybenzoate,
(2,3-
epoxypropyl) methacrylate, (2-ethoxyethyl) acetate, ethyl
acetamidocyanoacetate, ethyl
2-aminobenzoate, ethyl 3-aminopyrazol-4-carboxylate, ethyl benzoxylacetate,
ethyl
butyrylacetate, ethyl cyanoacetate, ethyl 2-cyano-3-ethoxyacrylate, ethyl
cyanoformate,
ethyl 2-cyanopropionate, ethyl 3,3-diethoxypropionate, ethyl 1,3-dithian-2-
carboxylate,
ethyl 2-ethoxyacetate, ethyl 2-furancarboxylate, ethyl levulinate, ethyl
mandelate, ethyl
2o gallate, ethyl 2-methyllactate, ethyl 4- nitrocinnamate, ethyl oxamate,
ethyl 2-
oxocyclohexanecarboxylate, ethyl 4-oxocyclohexane- carboxylate, ethyl 5-
oxohexanoate, ethyl 2-phenylacetoacetate, ethyl 4-piperidinecarboxylate,
ethyl 2-pyridinecarboxylate, ethyl 3-pyridinecarboxylate, ethyl 4-
pyridinecarboxylate, ethyl thioglycolate, ethyl 3,4,5-trihydroxybenzoate, 2-
hydroxyethyl methacrylate, 2-hydroxypropyl methacrylate, 3-indole acetate, 2-
methoxyethyl acetate, 1-methoxy-2-propyl acetate, methyl 2- aminobenzoate,
methyl 3-
aminocrotonate, methyl cyanoacetate, methyl 4-cyanobenzoate, methyl 4-
formylbenzoate, methyl 2-furancarboxylate, methyl isobutyrylacetate, methyl
methoxyacetate, methyl 2-methoxybenzoate, methyl 3-oxopentanoate, methyl
3o phenylglyoxylate, methyl phenyl- sulfinylacetate, methyl pivatoylacetate,
methyl 3-
pyridinecarboxylate, 5-nitrofurfurylidene diacetate, propyl gallate, propyl
3,4,5-
trihydroxybenzoate, methyl 3-methylthiopropionate, acetamide, acetani-
CA 02335253 2000-12-15
-42-
Tide, benzamide, benzanilide, N,N-diethylacetamide, N,N-dimethylformamide, N,N-
diethyl-3-methyl- benzamide, diethyltoluamide, N,N-dimethylacetamide, N,N-
diphenylacetamide, N-methylformamide, N-methylformanilide, N-acetylthiourea,
adipic
acid diamide, 2-aminobenzamide, 4-aminobenzamide, succinic acid diamide,
malonic
acid diamide, N,N'-methylene diacrylamide, oxalic acid diamide, pyrazine-2-
carboxamide, pyridine-4-carboxamide, N,N,N',N'-tetramethylsuccinic acid
diamide,
N,N,N',N'-tetramethylglutaric acid diamide, acetoacetanilide, benzohydroxamic
acid,
cyanoacetamide, 2-ethoxybenzamide, diethyl acetamidomalonate, ethyl
acetamidocyanoacetate, ethyl oxamate, hippuric acid Na salt, N-(hydroxy-
1o methyl)acrylamide, L-(-)-lactamide, 2'-nitroacetanilide, 3'-
nitroacetanilide, 4'-
nitroacetanilide, paracetamol, piperine, salicylanilide, 2-acetyl-y-
butyrolactone, y-
butyrolactone, s-caprolactone, dihydrocoumarin, 4-hydroxycoumarin, 2-(SH)-
furanone,
2,5-dihydro-S-methoxy-2-furanone, phthalide, tetrahydrofuran-2,4-dione, 2,2,6-
trimethyl-1,3-dioxin-4-one, y- valerolactone, 4-amino-1,3-dimethyluracil,
barbituric
15 acid, O-benzyloxycarbonyl-N-hydroxysuccinimide, succinimide, 3,6-dimethyl-
piperazin-2,5-dione, 5,5-diphenylhydantoin, ethyl 1,3-dioxoisoindoline-2-
carboxylate,
9-fluorenylmethylsuccinimidyl carbonate, hydantoin, maleimide, 3-methyl-1-
phenyl-2-
pyrazolin-5-one, 1-methyl-2-pyrrolidone, methyluracil, 6-methyluracil,
oxindole,
phenytoin, 1-(2H)-phthalazinone, phthalimide, 2,5-piperazinedione, 2-
piperidinone, 2-
2o pyrrolidone, rhodanine, saccharin, 1,2,3,6-tetrahydrophthalimide, 1,2,3,4-
tetrahydro-
6,7-dimethoxyquinazolin-2,4-dione, 1,5,5-trimethyl-hydantoin, 1-vinyl-2-
pyrrolidone,
ditert.butyl dicarbonate, diethyl carbonate, dimethyl carbonate, dimethyl
dicarbonate,
diphenyl carbonate, 4,5-diphenyl-1,3-dioxol-2-one, 4,6- diphenylthieno-(3,4-d)-
1,3-
dioxo[-2-one 5,5-dioxide, ethylene carbonate, magnesium methoxide methyl
carbonate,
25 monomethyl carbonate Na salt, propenyl carbonate, N-allylurea,
azodicarbonamide, N
benzylurea, biuret, l, l'-carbonyldiimidazol, N,N-dimethylurea, N-ethylurea, N
formylurea, urea, N-methylurea, N-phenylurea, 4-phenylsemicarbazide,
tetramethylurea, semicarbazide hydrochloride, diethyl azodicarboxylate, methyl
carbamate, 1-(4-methoxyphenyl)-2-(2- methoxyphenoxy)ethanone and 1-(4-
3o methoxyphenyl)-2-(2-methoxyphenoxy)ethanol.
Also preferred are anhydrides, such as the following:
CA 02335253 2000-12-15
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Benzoic anhydride, benzene-1,2,4,5-tetracarboxylic acid-1,2,4,5-dianhydride,
3,3',4,4'-
benzophenonetetracarboxylic anhydride, succinic anhydride, butyric anhydride,
crotonic
anhydride, cis-1,2-cyclo- hexanedicarboxylic anhydride, ditert.butyl
dicarbonate,
dimethyl dicarbonate, dodecenylsuccinic anhydride, Epicon B 4400, acetic
anhydride,
glutaric anhydride, hexanoic anhydride, isatoic anhydride, isobutyric
anhydride,
isovaleric anhydride, malefic anhydride, 1,8-naphthalenedicarboxylic
anhydride, 3-
nitrophthalic anhydride, 5-norbornene-2,3-dicarboxylic anhydride, phthalic
anhydride,
2-phenylbutyric anhydride, pivalic anhydride, propionic anhydride, cis-1,2,3,6-
tetrahydrophthalic anhydride and valeric anhydride.
Particularly preferred are benzophenones such as the following:
Benzophenone, 4-aminobenzophenone, 2-amino-5-chlorobenzophenone,
benzophenone-2-carboxylic acid, (S)-(-)-2-(N-benzopropyl)aminobenzophenone,
4,4'-
bis(dimethylamino)benzophenone, 4,4'- bis(diethylamino)benzophenone, 3,4-
dimethoxybenzophenone, 4,4'-dihydroxybenzophenone, 2,4- dihydroxybenzophenone,
4-hydroxybenzophenone, 2-hydroxy-4-methoxybenzophenone, 4-methoxybenzo-
phenone, 4,4'-dimethoxybenzophenone, 2,2',4,4'-tetrahydroxybenzophenone and 2-
chlorobenzophenone.
25
CA 02335253 2000-12-15
44
Figure 1 shows schematically a possible reaction cycle involving all
components.
Figure 1:
O-O ~ O
R~ R <----- R R + R R
(D = dioxirane) (C = ketone) B: e.g.: per-fatty acid
(R A = OOH)
(B) R - OOOH <------ R - OOH
A: fatty acid
HZO <______ HZOZ (Oxi)
lipase/amidase
component 1) = enzyme/ hydrolase: e.g.: lipase/amidase
component 2) = fatty acid (A)
component 3) = oxidation agent, e.g.: (Oxi)
component 4) = ketone (C)
B) = e.g.: per-fatty acid
D) = dioxir~ne
CA 02335253 2000-12-15
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A more detailed description of the enzyme component system (ECS) of the
invention in
terms of various applications follows:
1) Use in Wood Pulp Bleaching
One of the components of the enzyme component system (ECS) of the invention
used is
an enzyme, preferably lipase from, for example, Humicola lanuginosa, used at a
concentration of 0.05 to 5 mg per gram of wood pulp, preferably from 0.05 to 2
mg of
enzyme per gram of wood pulp (which corresponds to about 250 to 10,000 IU per
gram
of wood pulp) (1 IU hydrolyzes 1 g equivalent of a triglyceride fatty acid in
1 hour at pH
7.7 and 37 °C).
Preferably, the delignification (bleaching) with the enzyme component system
of the
invention is carried out in the presence of oxygen or air at atmospheric
pressure or at a
slight positive Oz pressure and at pH from 2 to 11, preferably at pH 3-9, at a
temperature of 20 to 95 °C, preferably 40-95 °C and a pulp
consistency of 0.5 to 40%.
An unusual and surprising finding concerning the use of enzymes for wood pulp
bleaching is that when the enzyme component system of the invention is used,
the
consistency of the material can be increased and the kappa value thus markedly
reduced.
For economic reasons, the process according to the invention is carried out at
a pulp
2o consistency from 4 to 35% and particularly from 4 to 15%.
Another component is the oxidant, preferably H20z, which is used at a
concentration
from 0.05 to 20 mg/g of wood pulp (100% basis) and preferably from 0.05 to 10
mg/g
of wood pulp.
Another component consists of one or more fatty acids, preferably C6 to Cz6,
and
particularly C6 to C16 fatty acids and more particularly tetradecanoic or
dodecanoic acid
at a concentration from 0.05 to 20 mg/g of wood pulp and preferably at a
concentration
from 0.05 to 10 mg/g of wood pulp.
Another component are the ketones, preferably, for example, benzophenone at a
concentration from 0.05 to 20 mg/g of wood pulp and preferably at a
concentration from
0.05 to 10 mg/g of wood pulp.
When the enzyme component system of the invention is used, for example, in a
process
for treating lignin, the chosen components are mixed with an aqueous
suspension of the
CA 02335253 2000-12-15
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lignin-containing pulp simultaneously or in any order. The reaction is
preferably started
by adding the oxidant or the enzyme.
Besides the abovesaid main components of the enzyme component system (ECS) of
the
invention, namely the enzymes (lipases/amidases), the oxidant, the fatty acids
and the
ketones, the bleaching system can also contain phenolic and/or nonphenolic
compounds
with one or more benzene rings which are capable of improving "oxidation
transfer"
(redox cascade) and/or of scavenging the radicals which possibly could cause
polymerization of the lignin.
Besides the abovesaid preferred oxidant, H202, particularly preferred are air,
oxygen
(possibly in addition to H202), organic peroxides, percompounds such as sodium
perborate and/or sodium percarbonate, persulfates etc. (optionally together
with
activators such as TAED, nitrilamines, dicyandiamines etc.). Oxygen can also
be
generated in situ by H202 + catalase or the like, or H202, can be generated in
situ from
GOD + glucose or similar systems.
The efficacy of the enzyme component system (ECS) as the oxidation system of
the
invention in the modification, degradation or bleaching of lignin, lignin-
containing
materials or similar substances is often even enhanced when Mg2+ ions are
present
besides the said constituents. The Mg2+ ions can be derived, for example, from
a salt
such as MgS04. The Mg2+ concentration is in the range from 0.1 to 2 mg/g, and
2o preferably from 0.2 to 0.6 mg/g, of lignin-containing material.
In some cases, the efficacy of the enzyme component system (ECS) of the
invention can
be increased even further by adding to the system besides Mg2+ ions also
complexing
agents, for example ethylenediaminetetraacetic acid (EDTA),
diethylenetriaminepentaacetic acid (DTPA), hydroxyethylenediaminetriacetic
acid
(HEDTA), diethylenetriaminepentamethylenephosphonic acid (DTMPA), nitrilotri-
acetic acid (NTA), polyphosphoric acid (PPA) etc. The concentration of said
complexing agents is in the range from 0.2 to S mg/g, and preferably from 1 to
3 mg/g,
of the lignin-containing material.
Surprisingly, we have also found that for many wood pulps an acid wash (pH 2
to 6 and
3o preferably 2 to 5) or a Q step (pH 2 to 6 and preferably 2 to S) preceding
the ECS step
causes a marked decrease in kappa value compared to processing without this
special
pretreatment. In the Q step, chelators usually employed for this purpose (for
example,
CA 02335253 2000-12-15
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EDTA or DTPA) are preferably used at a concentration f rom 0. 1 to 1 % and
particularly from 0.1 to 0.5 %.
Reducing agents can be added at the same time, which with the oxidants present
give
rise to a certain redox potential. Suitable reducing agents are sodium
bisulfate, sodium
dithionite, ascorbic acid, thio compounds, mercapto compounds or glutathione
etc..
It is also possible to add to the system radical generators or radical
scavengers
(scavenging, for example, OH- or OOH- radicals). These can improve the
interaction
between the redox and the radical mediators.
Additional metal salt can also be added to the reaction solution. These are
important in
to that they interact with chelators as radical generators or redox centers.
In the reaction
solution, the salts form canons. Such ions are, among others, Fe2+, Fe3+,
Mn2+, Mn3+,
Mn4+, Cul+, Cu2+, Ca2+, Ti3+, Ce4+ and A13+
The chelates present in the solution can also serve as mimicking substances
for certain
oxido- reductases, such as the laccases (copper complexes) or for the lignin
or
manganese peroxidases (heme complexes). By mimicking substances are meant
substances simulating the prosthetic groups of (in the present case)
oxidoreductases and,
for example, capable of catalyzing oxidation reactions.
Moreover, NaOCI can be added to the reaction mixture. This compound can form
singlet oxygen by interacting with hydrogen peroxide.
2o Finally, it is also possible to use detergents. These include nonionic,
anionic, cationic
and amphoteric surfactants. Detergents improve the penetration of the enzymes
and
other components into the fibers.
It may also be necessary to add polysaccharides and/or proteins to the
reaction mixture.
Suitable polysaccharides are, in particular, glucans, mannans, dextrans,
levans, pectins,
alginates or vegetable gums, and suitable proteins are gelatins and albumins.
These
substances serve mainly as protective colloids for the enzymes.
Other proteins that can be added are proteases such as pepsin, bromelain,
papain etc.
These substances can, among other things, bring about the degradation of
extensin
(hydroxyproline-rich protein) present in wood, thus improving access to the
lignin.
3o Other suitable protective colloids are amino acids, monosaccharides,
oligosaccharides,
polyethylene glycol [PEG] types of a wide range of molecular weights,
polyethylene
oxides, polyethyleneimines and polydimethylsiloxanes.
CA 02335253 2000-12-15
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It is also possible to add to the enzyme component system of the invention
substances
capable of increasing the hydrophobicity of the reaction mixture, thus
bringing about
the swelling of the lignin and the fibers and which makes them more
susceptible to
attack. Such substances are, for example, glycols, such as propylene glycol
and ethylene
s glycol, glycol ethers such as ethylene glycol dimethyl ether etc., and
solvents, for
example, alcohols such as methanol, ethanol, butanol, amyl alcohol,
cyclohexanol,
benzyl alcohol and chlorohydrin, phenols such as phenol, methylphenols and
methoxyphenols, aldehydes such as formaldehyde and chloral, mercaptans such as
butyl
mercaptan, benzyl mercaptan and thioglycolic acid, organic acids such as
formic, acetic
1o and chloroacetic acid, amines such as ammonia and hydrazine, hydrotropic
solvents, for
example concentrated solutions of sodium benzoate, other substances such as
benzenes,
pyridines, dioxane, ethyl acetate, and other basic solvents such as OH-/ HZO
or
OH- /alcohol etc..
The process according to the invention can be used not only for the
delignification
15 (bleaching) of sulfate, sulfite, organosolv or other wood pulps or lignins,
but also for the
preparation of wood pulp in general, whether from wood or annual plants, when
it is
desired to carry out the defibrillation by the usual cooking (digestion)
process (possibly
combined with mechanical processing or pressure), namely by very gentle
digestion, up
to kappa numbers in the range from about 50 - 120 kappa.
2o In the bleaching as in the preparation of wood pulps, the treatment with
the enzyme
component system (ECS) of the invention can be applied once or several times,
either
before and/or after the washing and extraction of the treated material with
NaOH etc., or
without these intermediate steps, but also before and/or after pretreatment or
post-
treatment steps, such as acid washing, Q-steps, alkaline leaching or bleaching
steps such
25 as peroxide bleaching, 02-enhanced peroxide steps, pressurized peroxide
steps, Oz-
delignification, Cl2 -bleaching, CIOZ -bleaching, Cl2/CIOZ- bleaching, peracid
bleaching, peracid-enhanced OZ/peroxide bleaching, ozone bleaching, dioxirane
bleaching, reductive bleaching steps, other treatments such as swelling steps,
sulfonations, NO/NOZ treatments, nitrosylsulfuric acid treatment, enzyme
treatments,
3o for example treatments with hydrolases, such as cellulases and/or
hemicellulases (for
example, xylanase, mannase etc) and/or amylases and/or pectinases and/or
proteinases
CA 02335253 2000-12-15
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and/or lipases and/or amidases and/or oxidoreductases such as, for example,
laccases
and/or peroxidases etc., or several combined treatments.
This results in substantially further reduced kappa values and substantially
increased
brightness. Before the ECS treatment, it is also possible to insert an OZ step
or, as
already mentioned, carry out an acid wash or a Q-step (chelation step).
The invention will be further illustrated by way of the following examples:
EXAMPLE 1
Enzymatic Bleaching of OZ-delignified Softwood (Sulfate Pulp)
g, absolutely dry basis, of wood pulp (OZ-delignified softwood), pulp
consistency 30%
(about 17 g moist) was added to solutions prepared as follows:
A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
benzophenone and 2.5 mg of H20z (30%) with agitation. The pH was adjusted with
sulfuric acid and/or sodium hydroxide solution so that, after addition of the
wood pulp
and the enzyme, the pH was 7.5.
B) To 5 mL of tap water was added 5 mg of lipase from Humicola lanuginosa
(about
25,000 IU).
Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
pulp, the material was mixed in a dough mixer for 2 minutes. The material was
then
transferred to a reaction vessel preheated to 45 °C and was allowed to
incubate 1-4
2o hours under atmospheric pressure.
The material was washed over a nylon screen (30 p,m) and extracted for one
hour at
60 °C, 2% consistency and using 8% NaOH per gram of wood pulp. The
material was
again washed after which the kappa number was determined. For results see
Table 1.
EXAMPLE Ia
Enzymatic Bleaching of OZ-delignified Softwood (Sulfate Pulp)
CA 02335253 2000-12-15
-50-
g, absolutely dry basis, of wood pulp (Oz-delignified softwood), pulp
consistency 30%
(about 17 g moist) was added to solutions prepared as follows:
A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
acetone and
2.5 mg of H202 (30%) with agitation. The pH was adjusted with sulfuric acid
and/or
5 sodium hydroxide solution so that, after addition of the wood pulp and the
enzyme, the
pH was 7.5.
B) To 5 mL of tap water was added 5 mg of lipase from Humicola lanuginosa
(about
25,000 IU).
Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
1o pulp, the material was mixed in a dough mixer for 2 minutes. The material
was then
transferred to a reaction vessel preheated to 45 °C and was allowed to
incubate 1-4
hours at atmospheric pressure.
The material was washed over a nylon screen (30 pm) and extracted for one hour
at
60 °C, 2% pulp consistency and using 8% NaOH per gram of wood pulp. The
material
was again washed, after which the kappa number was determined. For results see
Table
1.
EXAMPLE Ib (+ H202, with Nitrilamine as Activator)
Enzymatic Bleaching of 02-delignified Softwood (Sulfate Pulp)
5 g, absolutely dry basis, of wood pulp (Oz-delignified softwood), pulp
consistency 30%
(about 17 g moist) was added to solutions prepared as follows:
A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
acetone, 2.5
mg of H202 (30%) and 0.5 mg of nitrilamine with agitation. The pH was adjusted
with
sulfuric acid and/or sodium hydroxide solution so that, after addition of the
wood pulp
and the enzyme, the pH was 7.5.
2s B) To 5 mL of tap water was added S mg of lipase from Humicola lanuginosa
(about
25,000 IU).
CA 02335253 2000-12-15
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Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
pulp, the material was mixed in a dough mixer for 2 minutes. The material was
then
transferred to a reaction vessel preheated to 45 °C and was allowed to
incubate 1-4
hours at atmospheric pressure.
The material was washed over a nylon screen (30 pm) and extracted for one hour
at 60
°C, 2 % pulp consistency and using 8% NaOH per gram of wood pulp. The
material was
again washed, after which the kappa number was determined. For results see
Table 1.
EXAMPLE 2
1o Enzymatic Bleaching of Oi-delignified Hardwood (Sulfate Pulp)
5 g, absolutely dry basis, of wood pulp (OZ-delignified hardwood), pulp
consistency
30% (about 17 g moist) was added to solutions prepared as follows:
A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
benzophenone and 2.5 mg of H202 (30%) with agitation. The pH was adjusted with
sulfuric acid and/or sodium hydroxide solution so that, after addition of the
wood pulp
and the enzyme, the pH was 7.5.
B) To 5 mL of tap water was added 5 mg of lipase from Humicola lanuginosa
(about
25,000 ICT).
Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
2o pulp, the material was mixed in a dough mixer for 2 minutes. The material
was then
transferred to a reaction vessel preheated to 45 °C and was allowed to
incubate 1-4
hours at atmospheric pressure.
The material was washed over a nylon screen (30 pm) and extracted for one hour
at 60
°C, 2% pulp consistency and using 8% NaOH per gram of wood pulp. The
material was
again washed after which the kappa number was determined. For results see
Table 1.
CA 02335253 2000-12-15
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EXAMPLE 2a
g, absolutely dry basis, of wood pulp (OZ-delignified hardwood), pulp
consistency
30% (about 17 g moist) was added to solutions prepared as follows:
A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
acetone and
5 2.5 mg of HZOZ (30%) with agitation, The pH was adjusted with sulfuric acid
and/or
sodium hydroxide solution so that, after addition of the wood pulp and the
enzyme, the
pH was 7.5.
B) To 5 mL of tap water was added 5 mg of lipase from Humicola lanuginosa
(about
25,000 IU).
1o Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
pulp, the material was mixed in a dough mixer for 2 minutes. The material was
then
transferred to a reaction vessel preheated to 45 °C and was allowed to
incubate 1-4
hours at atmospheric pressure.
The material was washed over a nylon screen (30 pm) and extracted for one hour
at 60
°C, 2% pulp consistency and using 8% NaOH per gram of wood pulp. The
material was
again washed, after which the kappa number was determined. For results see
Table 1.
EXAMPLE 3
Enzymatic Bleaching of OZ-delignified Softwood (Sulfate Pulp)
5 g, absolutely dry basis, of wood pulp (OZ-delignified softwood), pulp
consistency 30%
(about 17 g moist) was added to solutions prepared as follows.
A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
benzophenone and 2.5 mg of HZOZ (30%) with agitation. The pH was adjusted with
sulfuric acid and/or sodium hydroxide solution so that, after addition of the
wood pulp
and the enzyme, the pH was 7.5.
CA 02335253 2000-12-15
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B) To 5 mL of tap water was added 200 IU of amidase from Pseudomonas
aeruginosa
(Sigma A 6691) (1 IU = conversion of 1 mole of acetamide and hydroxylamine to
acetohydroxamic acid and NH3 per minute at pH 7.2 and 37 °C).
Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
pulp, the material was mixed in a dough mixer for 2 minutes. The material was
then
transferred to a reaction vessel preheated to 45 °C and was allowed to
incubate 1-4
hours at atmospheric pressure.
The material was washed over a nylon screen (30 Vim) and extracted for one
hour at 60
°C, 2% pulp consistency and using 8% NaOH per gram of wood pulp. The
material was
1o again washed, after which the kappa number was determined. For results see
Table 1.
TABLE 1
Wood pulp % Delignification % Delignification
(before extraction) (after extraction)
a) Softwood
(untreated) ---- 5. 8%
b) Softwood
(lipase-treated) 19% 17.5% 32.0% 31%* 35%**
c) Hardwood
(untreated) ----- 6. 5
d) Hardwood
(lipase-treated) 21% 18%* 33% 28%*
e) Softwood
(amidase-treated) 15. 5% 23%
f) Comparative example:
laccase + HOBT 5 kg/ton
of wood pulp, other conditions
as in WO 96/18770
(pulp a/b) 17.5% 22%
*Underligned values were obtained with acetone as the ketone
**Value obtained with added nitrilamine.
CA 02335253 2000-12-15
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II) Use in Enzymatic Wastewater Treatment, for Example of Grinder Wastewater
in the Paper Industry
Because in this application the polymerization of lignin or lignin
constituents contained
in the wastewater is desired rather than lignin degradation, the enzyme
component
system (ECS) of the invention is used with a small amount of added
polymerization
catalyst.
One component of the enzyme component system (ECS) of the invention used is an
enzyme, preferably lipase from Aspergillus spec., at a concentration from 0.05
to 50 mg
per liter of wastewater and preferably from 0.05 to 10 mg of enzyme per liter
of
1o wastewater (corresponding to about 250 to 50,000 IU per liter of
wastewater) (1 IU
hydrolyzes 1 g equivalent of triglyceride fatty acid in 1 hour at pH 7.7 and
37 °C).
The treatment of the grinder wastewater with the enzyme component system of
the
invention is preferably carried out in the presence of oxygen or air at
atmospheric
pressure or slight positive oxygen pressure and at a pH from 2 to 11 and
preferably from
15 3 to 6, at a temperature from 20 to 95 °C and preferably from 40 to
95 °C.
Another component is the oxidant, preferably H202, which is used at a
concentration
from 0.05 to 200 mg per liter of wastewater (100% basis) and preferably from
0.05 to
50 mg per liter of wastewater.
Another component consists of one or more fatty acids, preferably C6 to C26
and
2o particularly C6 to C16, fatty acids, and more particularly tetradecanoic or
dodecanoic
acid at a concentration from 0.05 to 200 mg per liter, and preferably at a
concentration
from 0.05 to 10 mg per liter, of wastewater.
Another component is a ketone, preferably, for example, benzophenone at a
concentration from 0.05 to 200 mg per liter of wastewater and preferably at a
25 concentration from 0.05 to 50 mg per liter of wastewater.
Moreover, to increase the efficiency of the process and to use less
precipitant (mostly
sodium aluminate/aluminum sulfate) which represents the main cost factor, a
polymerization catalyst is used, mostly a phenolic substance or a polycyclic
compound
CA 02335253 2000-12-15
- 55 -
with several oxidizable hydroxyl groups, in our case preferably, for example,
purpurogallin.
These substances are used at a concentration from 0.005 to 200 mg per liter of
wastewater and preferably at a concentration from 0.005 to 50 mg per liter of
s wastewater.
The invention will be further illustrated by way of the following examples:
EXAMPLE 4
190 mL of grinder wastewater was adjusted to pH 6, its temperature was
adjusted to 45
°C in an appropriate jacketed reaction vessel and to it were added the
following
to solutions:
1) Enzyme solution: lipase (Aspergillus spec.): 1 mg in 0.1 mL of water
2) Fatty acid solution: 1 mg of dodecanoic acid in 1 mL of water
3) Ketone solution: 1 mg of 2,2',4,4'-tetrahydroxybenzophenone in 1 mL of
water
15 4) Polymerization catalyst: 0.1 mg of purpurogallin in 0.1 mL of water.
The reaction was initiated by addition of solution 5) (oxidant: H202), namely
of a
solution of 3.3 mg of H20z (30%) in 0.1 mL of water, and the volume was
adjusted to
200 mL with preheated water. The reaction was allowed to proceed for 1 to 4
hours and
preferably for 2 hours. The wastewater was then either only filtered or
filtered and
2o precipitated with 0.2%/0.2% or 0.5%/0.5% aluminum sulfate solution/sodium
aluminate
solution, 10 wt % in each case and based on the initial value without
treatment. The
lignin which in untreated grinder wastewater is usually present in an amount
from 600
to 900 mg per liter was determined quantitatively by photometry at 280 nm. The
drop in
lignin is a measure of COD reduction and of the efficiency of the system.
25 The results are collected in Table 2.
EXAMPLE 5 (without polymerization catalyst)
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190 mL of grinder wastewater was adjusted to pH 6, its temperature was
adjusted to 45
°C in an appropriate jacketed reaction vessel and to it were added the
following
solutions:
1 ) Enzyme solution: lipase (Aspergillus spec.): 1 mg in 0.1 mL of water
2) Fatty acid solution: 1 mg of dodecanoic acid in 1 mL of water
3) Ketone solution: 1 mg of 2,2',4,4'-tetrahydroxybenzophenone in 1 mL of
water
The reaction was initiated by addition of solution 4) (oxidant- HzOz), namely
a solution
of 3.3 mg of HzOz (30%) in 0.1 mL of water, and the volume was adjusted to 200
mL
with preheated water. The reaction was allowed to proceed for 1 to 4 hours and
1o preferably for 2 hours. The wastewater was then either only filtered or
filtered and
precipitated with 0.2%/0.2% or 0.5%/0.5% aluminum sulfate solution/sodium
aluminate
solution, 10 wt % in each case based on the initial value without treatment.
The lignin
which in untreated grinder wastewater is usually present in an amount from 600
to 900
mg per liter was determined quantitatively by photometry at 280 nm. The drop
in lignin
is a measure of COD reduction and of the efficieney of the system.
The results are collected in Table 2.
25
35
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TABLE 2
Treatment Residual Lignin After 2 Hours
(0 Value = 600 mg/liter)
COD (mg/L)
No treatment
(filtered) 800
No treatment
(filtered/precipitated*) 720
With treatment (lipase)
(filtered/precipitated),
(with polymerization cat.)** 100
Comparative treatment:
with 25,000 IU of laccase
per liter of wastewater
(filtered/precipitated*)
(with polym. catalyst)** 170
2o With treatment (lipase)
(filtered/precipitated*)
(without polym. catalyst) 220
With treatment
(filtered/precipitated*)
with polym. catalyst
(Humicola lipase) 160
(For the test with polymerization
and/or modification of lignin, see below)
* Only the precipitations at 0.5%/0.5% are shown.
* * Polymerization catalyst
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>~ Use in the Preparation of Lignin Solutions or Gels, Corresponding
Binders/Adhesives and of Wood-Based Composites
In this application, too, the polymerization of the lignin or lignin-
containing materials is
desired and not lignin degradation. Hence, the enzyme component system (ECS)
of the
invention is used with a small amount of added polymerization catalyst.
Because it was found that the polymerization of lignin, for example in
groundwood pulp
wastewater (grinder wastewater) is also a good system for evaluating general
polymerization properties for the use of the enzyme component system (ECS) in
the
1o preparation of lignin solutions or gels, of the corresponding
binders/adhesives and of
wood-based composites, tests were carried out with the same experimental
formulations
as for the wastewater.
In this regard, it is known from the cited patents WO 94/01488, WO 93/25622,
WO
93/23477 and DE 3 037 992 C2 that, for example, in the production of particle
board the
15 binder made by polymerization and dissolution of lignin is applied by
spraying it onto
the wood fiber material in an amount of about 40 to 100 g per kg of said
material which
is then subjected to pressing at a pressure of about 20-40 kg / cm2 for about
2-4 min
and at a temperature rising from about 3 5 to 190 °C within about 20
seconds. The
pressures and temperatures used for pressing can, of course, also be
substantially lower,
2o and subsequent curing of the binder/wood fiber mixture by continuing enzyme-
catalyzed reactions may be desired.
To evaluate the polymerization properties of ECS for this application, we used
as the
model system, as stated hereinabove, the above-described system for removing
lignin
from grinder wastewater.
25 As one component of the enzyme component system (ECS) of the invention was
used
an enzyme, preferably lipase from Humicola lanuginosa, at a concentration from
0.05 to
50 mg per liter of wastewater and preferably from 0.05 to 10 mg of enzyme per
liter of
wastewater (corresponding to about 250 to 50,000 IU per liter of wastewater)
(1 IU
hydrolyzes 1 p equivalent of triglyceride fatty acid in 1 hour at pH 7.7 and
37 °C).
3o The treatment of the grinder wastewater with the enzyme component system of
the
invention is preferably carried out in the presence of oxygen or air at
atmospheric
CA 02335253 2000-12-15
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pressure or slight positive oxygen pressure and at a pH from 2 to 11 and
preferably from
3 to 6, at a temperature from 20 to 95 °C and, preferably from 40 to 95
°C.
Another component is the oxidant, preferably H20z which is used at a
concentration
from 0.05 to 200 mg per liter of wastewater (100% basis) and preferably from
0.05 to
50 mg/liter of wastewater.
Another component consists of one or more fatty acids, preferably a C6 to C26,
particularly C6 to C16 fatty acid and more particularly tetradecanoic or
dodecanoic acid
at a concentration from 0.05 to 200 mg/liter of wastewater and preferably at a
concentration from 0.05 to 50 mg/liter of wastewater.
1o Another component is a ketone, preferably, for example, benzophenone at a
concentration from 0.05 to 200 mg/liter of wastewater and preferably at a
concentration
from 0.05 to 50 mg/liter of wastewater.
Moreover, as already stated in the foregoing, to increase the eWciency of the
process a
polymerization catalyst is used, mostly a phenolic substance or a polycyclic
compound
with several oxidizable hydroxyl groups, in our case preferably, for example,
purpurogallin.
These substances are used at a concentration from 0.005 to 200 mg per liter of
wastewater and preferably at a concentration from 0.005 to 50 mg per liter of
wastewater.
EXAMPLE 6
190 mL of grinder wastewater was adjusted to pH 8.5, preheated to a
temperature of 45
°C in an appropriate jacketed reaction vessel and to it were added the
following
solutions:
1) Enzyme solution: lipase (Humicola lanuginosa): 1 mg in 0.1 mL of water
2) Fatty acid solution: 1 mg of dodecanoic acid in 1 mL of water
3) Ketone solution: 1 mg of 2,2',4,4'-tetrahydroxybenzophenone in 1 mL of
water
4) Polymerization catalyst: 0.1 mg of purpurogallin in 0.1 mL of water.
The reaction was initiated by addition of solution 5) (oxidant: H20z), namely
a solution
of 3.3 mg of H202 (30%) in 0. 1 mL of water, and the volume was adjusted to
200 mL
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with preheated water. The reaction was allowed to proceed for 1 to 4 hours and
preferably for 2 hours. The wastewater was then either only filtered or
filtered and
precipitated with 0.2%/0.2% or 0.5%/0.5% aluminum sulfate solution/sodium
aluminate
solution, 10 wt % in each case, based on the initial value without treatment.
The lignin
which in untreated grinder wastewater is usually present in an amount from 600
to 900
mg per liter was determined quantitatively by photometry at 280 nm. The drop
in lignin
is a measure of COD reduction and of the efficiency of the system.
The results are collected in Table 2.
I~ Use as Enzymatic Deinking System
In this application, no lignin degradation is desired but, rather, a swelling
effect on the
lignin-containing fibers so as to bring about the detachment of adhering
printing ink
particles, an effect similar to that of sodium hydroxide solution in
conventional
chemical deinking.
To the enzyme component system (ECS) are added besides the usual components
such
as lipase, oxidant, fatty acid and ketone, also a number of phenolic
substances which
serve as polymerization catalysts in wastewater treatment and in lignin
2o polymerization/modification applications. Here, we found, surprisingly,
that these
substances cause a shift in the pH for the optimum enzyme activity thus
improving
performance.
Also surprisingly, we found that the addition of a reducing agent, preferably
dithionite
or bisulfite increases the efficiency of ink detachment.
One component of the enzyme component system (ECS) of the invention used is an
enzyme, preferably lipase from Humicola lanuginosa, at a concentration from 5
to 500
mg per kg of air-dried waste paper, and preferably from 5 to 100 mg of enzyme
per kg
of waste paper (corresponding to about 25,000 to 500,000 IU per kg of waste
paper)
(1 IU hydrolyzes 1 ~ equivalent of triglyceride fatty acid in 1 hour at PH 7.7
and 37
°C). The treatment of the waste paper with the enzyme component system
of the
invention for the purpose of removing printing ink particles is preferably
carried out in
the presence of oxygen or air at atmospheric pressure or slightly positive
oxygen
CA 02335253 2000-12-15
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pressure (maximum 2 bar) and at a pH from 7 to 11 and preferably 7 to 9, at a
temperature from 20 to 95 °C and preferably from 40 to 95 °C.
Another component is the oxidant, preferably H202, which is used at a
concentration
from 5 to 5000 mg per kg of waste paper (100% basis) and preferably from 5 to
1000
mg per kg of waste paper.
Another component consists of one or more fatty acids, preferably C6 to C26,
and
particularly C6 to C16, fatty acids, and more particularly tetradecanoic or
dodecanoic
acid at a concentration from 5 to 2000 mg per kg of waste paper and preferably
at a
concentration from 5 to 500 mg per kg of waste paper.
to Another component is a ketone, preferably, for example, benzophenone at a
concentration from 5 to 2000 mg per kg of waste paper and preferably at a
concentration from 5 to 500 mg per kg of waste paper.
Moreover, to increase the efficiency of the process, the abovesaid compounds
are used,
for example phenolic substances or polycyclic compounds with several
oxidizable
hydroxyl groups and preferably, for example, bisphenol A. These substances are
employed at a concentration from 1 to 2000 mg per kg of waste paper and
preferably at
a concentration from 1 to 500 mg per kg of waste paper.
In addition, a reducing agent is used, preferably Na-dithionate or Na-
bisulfite, at a
concentration from 0. 1 to 1000 mg per kg of waste paper and preferably at a
2o concentration f rom 0. 1 to 200 mg per kg of waste paper.
To collect the printing ink particles, commercial detergents are used as
collectors,
preferably Incopur brands, for example Incopur RSGA, at a concentration from 1
to
5000 mg per kg of waste paper, and preferably from 1 to 1000 mg per kg of
waste
paper.
To enhance the detaching effect on many waste paper compositions, additional
enzymes
can be added, for example cellulases and/or hemicellulases (for example,
xylanase
and/or mannase etc.) and/or pectinases and/or oxidoreductases.
The invention will be further illustrated by way of the following examples:
EXAMPLE 7
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About 10 kg of water (preheated to about 45 °C) was added to the pulper
of a Lamort
laboratory deinking apparatus, and the pH was adjusted with sodium hydroxide
solution
(and/or sulfuric acid) so that after the addition of 1.5 kg of air-dried waste
paper (50%
newspapers, 50% magazines) which had been cut into about 2 x 3 cm pieces and
after
the addition of the other system constituents the pH was 8.0 to 8.5.
These system constituents were (per kg of air-dried waste paper):
a) 500,000 IU of lipase from Humicola lanuginosa per 100 mL of tap water
b) 0.1 g of dodecanoic acid per 100 mL of tap water
c) 0.1 g of benzophenone per 100 mL of tap water
1o d) 0.1 g of bisphenol A per 20 mL of 0. 1 molar NaOH
e) 0.02 g of Na bisulfite per 10 mL of tap water
fJ 0.5 g of Incopur RSGA per 100 mL of tap water
g) 1 g of H202 (30%) per 100 mL of tap water (added at the end).
1s The pulper was started after the addition of system constituents a) to g)
and during the
addition of the waste paper. The total quantity of water was then adjusted to
15 kg with
approximately 45 °C tap water. The pulping process was allowed to
proceed for 10
minutes. For further reaction, the fiber suspension was transferred to a
heated vessel
where it was allowed to incubate at about 40-45 °C for 15 to 45
minutes.
2o In a Voith flotation cell, 100 g of pulp, absolutely dry basis, (after this
incubation) was
adjusted to a total volume of about 20 L with tap water (45 °C) and
subjected to
flotation for 10 to 20 minutes. The accepted stock was discharged, test sheets
were
prepared from it in a commercial sheet former, dried and the ISO brightness
were
determined. Table 3 shows the results.
2s
(ISO = International Standardization Organization)
EXAMPLE 8
The method was the same as in Example 7, but instead of the enzyme component
system (ECS) of the invention only water was used. Table 3 shows the results.
CA 02335253 2000-12-15
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EXAMPLE 9
About 1 kg of water (preheated to about 45 °C) was added to a dough
mixer, and the pH
was adjusted with sodium hydroxide solution (and/or sulfuric acid) so that
after the
addition of 150 g of air-dried waste paper (50% newspapers, 50% magazines)
which
had been cut into about 2 x 3 cm pieces and after the addition of the other
system
constituents the pH was 8.0 to 8.5.
These system constituents were (per 100 g of air-dried waste paper):
1o a) 5000 IU of amidase from Pseudomonas aeruginosa (Sigma A 6691) per 100 mL
of
tap water
(1 IU = conversion of 1 pmole of acetamide and hydroxylamine to
acetohydroxamic
acid and NH3 per minute at pH 7.2 and 37 °C)
b) 0.01 g of dodecanoic acid per 100 mL of tap water
c) 0.01 g of benzophenone per 100 mL of tap water
d) 0,01 g of bisphenol A per 20 mL of 0.1 molar NaOH
e) 0.002 g of Na bisulflte per 10 mL of tap water
f) 0.05 g of Incopur RSGA per 100 mL of tap water
g) 0.1 g of H202 (30%) per 100 mL of tap water (added at the end).
The dough mixer was started after the addition of system constituents a) to g)
and
during the addition of the waste paper. The total quantity of water was then
adjusted to
1.5 kg with tap water (45 °C). The pulping process was allowed to
proceed for 10
minutes.
For further reaction, the fiber suspension was transferred to a heated vessel
where it was
allowed to incubate at about 40-45 °C for 15 to 45 minutes.
In a Voith flotation cell, 100 g of pulp, absolutely dry basis, (after this
incubation) was
adjusted to a total volume of about 20 L with tap water (45 °C) and
subjected to
flotation for I O to 20 minutes. The accepted stock was discharged, test
sheets were
3o prepared from it in a commercial sheet former, dried and the ISO brightness
and were
determined. Table 3 shows the results.
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EXAMPLE 10 (Chemical System)
About 10 kg of water (preheated to about 45 °C) was added to the pulper
of a Lamort
laboratory deinking apparatus, and 1.5 kg of air-dried waste paper (50%
newspapers,
50% magazines), cut into approximately 2 x 3 cm pieces, was added after the
addition
of the following chemicals (based on air- dried pulp):
1) 0.8 wt % of soap (DR 3, Henkel)
l0 2) 3.5% of water glass
3) 2% of sodium hydroxide (100%)
4) 1 % H202 ( 100%)
The pulper was started during waste paper addition. The total quantity of
water was then
adjusted to 15 kg with about 45 °C tap water. The pulping process was
allowed to
proceed f or 10 minutes.
For further reaction, the fiber suspension was transferred to a heated vessel
where it was
allowed to incubate at about 40-45 °C for 15 to 45 minutes,
In a Voith flotation cell, 100 g of pulp, absolutely dry basis, (after this
incubation) was
2o adjusted to a total volume of about 20 L with tap water (45 °C) and
subjected to
flotation for 10 to 20 minutes. The accepted stock was discharged, test sheets
were
prepared from it in a commercial sheet former, dried and the ISO brightness
were
determined'. Table 3 shows the results.
30
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TABLE 3
System ISO Brightness,
Water only 51
Chemical system 61
ECS/lipase 58.5
ECS/amidase 57
Comparative system:
lactase (800,000 IU/kg of
waste paper + bisphenol A +
Na bisulfite (0.1 or 0.02 g/kg
of waste paper), for other con-
ditions see WO 91/14820;
WO 92/20857 55.5
V) Use as Oxidation System in Organic Synthesis
From the multiplicity of possible uses of the enzyme component system (ECS) of
the
invention, such as in hydroxylation reactions, oxidation of unsaturated
aliphatics,
Baeyer-Villiger oxidations, oxidation of heterocycles, carbon-carbon
dehydrogenations
and other oxidation reactions, by way of the following examples, the oxidation
of
aicohols to aldehydes and of aromatic methyl groups to aldehydes are
described.
It is known from the literature that these reactions can be carried out with
the
oxidoreductase lactase and a mediator such as ABTS (2,2'-azino-bis(3-
ethylbenzothiazolin-6-sulfonic acid), see T. Rosenau et al., Synthetic
Communications
26 (2), 315-320 (1996) and A Potthast et al., J. Org. Chem. (60), pp. 4320-
4321 (1995).
The main advantage of the process of the invention over these processes is its
lower cost
and better performance, particularly based on the cost.
One of the components of the enzyme component system (ECS) of the invention is
an
enzyme, preferably lipase from, for example, Humicola lanuginosa, used at a
concentration of 0.05 to 5 mg per 10 mmoles of substrate, preferably from 0.05
to 3 mg
per 10 mmoles of substrate (which corresponds to about 250 to 15,000 IU) (1 IU
hydrolyzes 1 ~ equivalent of a triglyceride fatty acid in 1 hour at pH 7.7 and
37 °C).
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Preferably, the oxidation reaction with the enzyme component system of the
invention
is carried out in the presence of oxygen or air at atmospheric pressure or a
slightly
positive OZ pressure and at pH from 2 to 1 l, preferably at pH 3-9, at a
temperature of 20
to 95 °C, preferably 40-95 °C and a 5 to 100 mmolar, preferably
5 to SO mmolar,
substrate concentration.
Another component is the oxidant, preferably H202 (100%), which is used at a
concentration from 0.05 to 100 mg per 10 mmoles of substrate and preferably
from 0.05
to 30 mg per 10 mmoles of substrate.
Another component consists of one or more fatty acids, preferably C6 to C26,
and
1o particularly Cs to C16 fatty acids, and more particularly tetradecanoic or
dodecanoic acid
at a concentration from 0.05 to 100 mg per 10 mmoles of substrate and
preferably at a
concentration f rom 0.05 to 30 mg per 10 mmoles of substrate.
Another component is a ketone, preferably, for example, benzophenone at a
concentration from 0.05 to 100 mg per 10 mmoles of substrate and preferably at
a
concentration from 0.05 to 30 mg per 10 mmoles of substrate.
The invention will be further illustrated by way of the following examples:
EXAMPLE 11 (Oxidation of Benzyl Alcohols to Aldehydes)
2o The following components were added to 50 mL of 0.1 molar pH 4.5 acetate
buffer in a
250-mL reaction flask:
1) p-methoxybenzyl alcohol in 30 mL of tetrahydrofuran [THF] (20-molar in
total
volume)
2) 2 mg of lipase from Humicola lanuginosa
3) 5 mg of dodecanoic acid
4) 25 mg of benzophenone
The reaction was started by addition of 12.5 mg of H202 (30%) and was allowed
to
proceed for 12 to 24 hours. Then, 0.5 mL of the reaction solution was removed,
3o extracted with CHZCl2, and the p-methoxybenzaldehyde content was determined
by GC
or GC-MS. The results are shown in Table 4
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EXAMPLE 12 (Oxidation of Aromatic Methyl Groups to Aldehydes)
The following components were added to 50 mL of 0.1 molar pH 4.5 acetate
buffer in a
250-mL reaction flask:
1) toluene in 30 mL of THF (20-molar in total volume)
2) 2 mg of lipase from Humicola lanuginosa
3) 5 mg of dodecanoic acid
4) 25 mg of benzophenone
The reaction was started by addition of 12.5 mg of H202 (30%) and was allowed
to
proceed for 12 to 24 hours. Then, 0.5 mL of the reaction solution was removed,
extracted with CHZC12, and the benzaldehyde content was determined by GC or GC-
MS. The results are shown in Table 4.
TABLE 4
Substrate Oxidized Substrate Conversion,
P-Methoxybenzyl
alcohol p-methoxybenzaldehyde 98
(lipase)
p-Methoxybenzyl
alcohol p-methoxybenzaldehyde 90
(ABTS/laccase)
Toluene
(lipase) benzaldehyde 98
Toluene
(ABTS/laccase) benzaldehyde 92
VI) Use in Coal Liquefaction
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Recently, the use of white rotting fungi was studied in the liquefaction of
lignite and
anthracite, and the general feasibility thereof was confirmed. Patent
applications WO
94/29510 and WO 96/18770 have also disclosed the general feasibility of using
fixngus-
free systems based on oxido- reductases and special mediators.
Surprisingly, we were able to confirm for the enzyme component system (ECS) of
the
invention that it, too, can be used for "liquefying" the lignin-like
tridimensional network
of polycyclic aromatic ring systems of lignite and anthracite.
One of the components of the enzyme component system (ECS) of the invention is
an
enzyme, preferably lipase from, for example, Humicola lanuginosa, used at a
1o concentration of 0.05 to 20 mg per gram of ground lignite, absolutely dry
basis,
preferably from 0.05 to 10 mg per gram of coal (corresponding to about 250 to
50,000
IU) (1 IU hydrolyzes 1 ~ equivalent of a triglyceride fatty acid in 1 hour at
pH 7.7 and
37 °C).
Preferably, the coal treatment with the enzyme component system of the
invention is
~5 carried out in the presence of oxygen or air at atmospheric pressure or at
a slightly
positive 02 pressure and at pH from 2 to 1 I, preferably at pH 3-9, at a
temperature of 20
to 95 °C, preferably 40-95 °C, and a coal consistency of 0.5 to
40%.
An unusual and surprising finding concerning the use of enzymes is that when
the
enzyme component system of the invention is employed, the consistency of the
material
2o can be increased and the performance is markedly improved. For economic
reasons, the
process according to the invention is carried out at a coal consistency from 4
to 35% and
particularly from 4 to 15%.
Another component is the oxidant, preferably H202, which is used at a
concentration
from 0.05 to I 00 mg per gram of coal ( 100% basis) and preferably from 0.05
to 50 mg
25 per gram of coal.
Another component consists of one or more fatty acids, preferably C6 to C26,
particularly C6 to Cis fatty acids, and more particularly tetradecanoic or
dodecanoic aeid
at a concentration from 0.05 to 100 mg per gram of coal and preferably at a
concentration from 0.05 to SO mg per gram of coal.
3o Another component is a ketone, preferably, for example, benzophenone at a
concentration from 0.05 to 100 mg per gram of coal and preferably at a
concentration
from 0.05 to 50 mg per gram of coal.
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-
The invention will be further illustrated by way of the following example:
EXAMPLE 13
Enzymatic Coal Liquefaction
5 g of lignite or anthracite, absolutely dry basis, (particle size about 200
to SOO,u) was
added to solutions prepared es follows:
1o A) To 20 mL of tap water were added 5 mg of tetradecanoic acid, 25 mg of
benzophenone and 12.5 mg of H202 (30%) per gram of coal with agitation. The pH
was
adjusted with sulfuric acid and/or sodium hydroxide solution so that, after
addition of
the coal and the enzyme, the pH was 8.
B) To 5 mL of tap water was added 10 mg of lipase from Humicola lanuginosa
(about
50,OOOIU).
Solutions A and B were combined and diluted to 45 mL. After addition of the
coal, the
material was mixed for 2 minutes. The material was then transferred to a
reaction vessel
preheated to 45 °C and allowed to incubate 1-4 hours.
The resulting coal of modified consistency was then removed from the reaction
flask.
VII) Use as Bleaching Agent in Detergents
In using the enzyme component System (ECS) of the invention as a bleaching
agent in
detergents, one of the components is an enzyme, preferably lipase from, for
example,
Humicola lanuginosa, used at a concentration from 0.05 to 20 mg per 100 mL of
washing solution and preferably from 0.05 to l0 mg of enzyme per 100 mL of
washing
solution (which corresponds to about 250 to 100,000 IU per 100 mL of washing
solution) (1 IU hydrolyzes 1 p equivalent of a triglyceride fatty acid in 1
hour at PH 7.7
and 3 7 °C).
3o Preferably, the bleaching with the enzyme component system of the invention
is carried
out in the presence of oxygen or air at atmospheric pressure or at a slightly
positive OZ
CA 02335253 2000-12-15
pressure and at pH from 2 to 12, preferably at pH 3-10, at a temperature of 20
to 95 °C
and preferably 30-95 ° C.
Another component is the oxidant, preferably H202, which is used at a
concentration
from 0.05 to 50 mg per 100 mL of washing solution (100% basis) and preferably
from
0.05 to 20 mg per 100 mL of washing solution.
Another component consists of one or more fatty acids, preferably a C6 to C26,
particularly a C6 to C16 fatty acid and more particularly tetradecanoic or
dodecanoic
acid, at a concentration from 0.05 to 50 mg per 100 mL, of washing solution
and
preferably at a concentration from 0.05 to 20 mg per 100 mL of washing
solution.
1o Another component is a ketone, preferably, for example, benzophenone at a
concentration from 0.05 to 50 mg per 100 mL of washing solution and preferably
at a
concentration from 0.05 to 20 mg per 100 mL of washing solution.
Other Components
The bleaching system can also contain phenolic and/or nonphenolic compounds
with
one or more benzene rings. The following oxidants besides those mentioned
hereinabove are particularly preferred: air, oxygen, H202, organic peroxides,
sodium
perborate and/or sodium percarbonate. Oxygen can also be generated in situ by
H202+
2o catalase or the like, or HZOa can be generated in situ by GOD + glucose or
similar
systems.
Also preferred are multicomponent bleaching systems containing cation-
generating
metals salts. The cations Fe Z+ Fe3+ Mn2+ Mn3+ Mn4+ Cu'+, Cu2+, Ti3+, Ce4+, M
2+ and
> > > > > g
A13+ are preferably used.
The bleaching system can also contain polysaccharides and/or proteins.
Suitable
polysaccharides are, glucans, mannans, dextrans, levans, pectins, alginates,
vegetable
gums and/or polysaccharides formed by fungi or produced in mixed cultures with
yeasts. Suitable proteins are gelatins and albumins, among others. Also
suitable are
monosaccharides, oligosachharides, amino acids, PEG, polyethylene oxides,
3o polyethyleneimines and polydimethylsiloxanes.
CA 02335253 2000-12-15
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Use of the Multicomponent System
The multicomponent system can be used in combination with surface-active
detergent
constituents or detergent additives, which in themselves are known.
s The invention will be further illustrated by way of the following examples:
EXAMPLE 14
Effect of ECS on Tea-Stained Standard Cotton Fabrics
A (5x5 cm) piece of fabric was allowed to incubate in 100 mL of washing
solution (in a
300-mL Erlenmeyer flask) at 40 °C for 40 min with reciprocating shaking
(120 rpm).
Before the beginning of incubation, the washing solution was subjected to a 10-
min
temperature equilibration period.
1s The washing solution was prepared with standard tap water (STW) at
14° dH'. The
following system component doses were used: 2.5 mg of lipase from Humicola
lanuginosa/100 mL, 2.5 mg of tetradecanoic acid/100 mL, 12.5 mg of
benzophenone/100 mL and 6.5 mg of H202 (30%). After decanting the washing
liquor",
cold water was added 3x in the form of a strong water jet and then decanted.
2o The results are shown in Table 5.
EXAMPLE 15
Effect of ECS (Amidase) on Tea-Stained Standard Cotton Fabrics
A (5x5 cm) piece of fabric was allowed to incubate in 100 mL of washing
solution (in a
300-mL Erlenmeyer flask) at 40 °C for 40 min with reciprocating shaking
(120 rpm).
Before the beginning of incubation, the washing solution was subjected to a 10-
min
temperature equilibration period.
3o The washing solution was prepared with standard tap water (STW) at
14° dH. The
following doses were used: 1000 IU of amidase/ 100 mL (1 IU = conversion of 1
gmole
of acetamide and hydroxylamine to acetohydroxamic acid per minute at pH 7.2
and 37 °
CA 02335253 2000-12-15
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C), 2.5 mg of tetradecanoic acid/1 00 mL, 12.5 mg of benzophenone/100 mL and
6.5
mg of H202 (30%). After decanting the "washing liquor", cold water was added 3
x in
the form of a strong water jet and then decanted.
The results are shown in Table 5.
(dH = one degree of German water hardness = 10 mg of Ca0/L)
TABLE 5
1o pH Whiteness Brightness
STW zero value 4.5 2.55 2.3
Heavy-duty detergent 10.1 8.9 6.15
STW + ECS (lipase) 8.5 7.5 7,2
STW + ECS (amidase) 8 6.9 6.3
Liquid detergent +
ECS (lipase) 8.5 8.5 8.0
Comparative test:
Liquid detergent + laccase
+ HOBT (conditions
as in
PCTBP 96/02658;
PCT/EP 94101967 5 6.5 6.0
VIII) Use in the Bleaching/Decolorizing of Textiles
One of the components of the enzyme component system (ECS) of the invention
used in
the bleaching/decolorizing of textile fabrics is an enzyme, preferably lipase
from, for
3o example, Humicola lanuginosa, employed at a concentration of 0.05 to 10 mg
per gram
of denim (corresponding to about 250 to 25,000 IU per gram of denim) (1 IU
hydrolyzes 1 p equivalent of a triglyceride fatty acid in 1 hour at pH 7.7 and
37 °C).
Preferably, the bleaching/decolorizing with the enzyme component system of the
invention is carried out in the presence of oxygen or air at atmospheric
pressure or at a
slightly positive OZ pressure and at pH from 2 to 11, preferably at pH 3-9, at
a
temperature of 20 to 95 °C, preferably 40-95 °C and a fabric
density of 0.5 to 40%
[fabric density = ratio of weight of fabric to weight of solution]
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An unusual and surprising finding concerning the use of enzymes is that when
the
enzyme component system of the invention is used, the fabric density can be
increased
and the performance markedly improved. For economic reasons, the process
according
to the invention is carried out at a fabric density from 4 to 35% and
particularly from 4
to 15%.
Another component is the oxidant, preferably H202, which is used at a
concentration
from 0.05 to 20 mg per gram of denim (100% basis) and preferably from 0.05 to
10 mg
per gram of denim.
Another component consists of one or more fatty acids, preferably C6 to C26
and
1o particularly C6 to Ci6 fatty acids, and more particularly tetradecanoic or
dodecanoic acid
at a concentration from 0.05 to 20 mg/g of denim and preferably at a
concentration from
0.05 to 10 mg per gram of denim.
Another component is a ketone, preferably, for example, benzophenone, used at
a
concentration from 0.05 to 20 mg/g of denim and preferably at a concentration
from
0.05 to 10 mg/g of denim.
The invention will be further illustrated by way of the following examples:
EXAMPLE 16
2o Bleaching with ECS + Lipase
1 g of denim fabric was placed in a 200-mL Erlenmeyer flask (fabric density
2%). The
pH of the solution (tap water), the volume of which was 50 mL after the
addition of all
components, was preadjusted to pH 6 with 0.5 N HZS04.
1 mg of lipase from Humicola lanuginosa, 0.5 mg of tetradecanoic acid, 1 mg of
benzophenone and 2.5 mg of H202 (30%) were added per gram of denim. The
experiment was carried out in a shaking water bath (200 rpm) at 45 °C
and a reaction
time of 45 minutes. The piece of fabric was washed with tap water and dried in
air. The
brightness was then determined with an Elrepho instrument. The values obtained
are
3o given in Table 6.
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EXAMPLE 17
Bleaching with ECS + Amidase
1 g of denim fabric was placed in a 200-mL Erlenmeyer flask (fabric density
2%). The
pH of the solution (tap water), the volume of which was 50 mi, after the
addition of all
components, was preadjusted to pH 6 with 0.5 N HZS04.
200 I(J of amidase from Pseudomonas aeruginosa (Sigma A 6691), 0.5 mg of
tetradecanoic acid, 1 mg of benzophenone and 2.5 mg of H202 (30%) were added
per
1o gram of denim. The experiment was carried out in a shaking water bath (200
rpm) at 45
°C and a reaction time of 45 minutes. The piece of fabric was washed
with tap water and
dried in air. The brightness was then determined with an Elrepho instrument.
The values
obtained are given in Table 6.
TABLE 6
System pH ISO Brightness
Untreated specimen -- 4.5
Laccase + violuric acid
(comparative system) 3 .5 13 . S
ECS system + lipase 6.0 16.9
ECS system + amidase 6.0 14.5
Hypochlorite 4.5 n.d.
Addition of Other Substances to the Enzyme Component System (ECS)
3o For all applications, the components of the enzymatic oxidation systems
with enzyme
action-enhancing compounds disclosed in DE 198 21 263.1, DE 198 20 947.9 and
PCTlDE 98/01313 can be added to the enzyme component system (ECS) of the
invention, such systems containing the following:
a) At least one oxidation catalyst, preferably enzymes such as oxidoreductases
of
classes 1.1.1. to 1.97 according to the International Enzyme Nomenclature:
Committee
of the International Union of Biochemistry and Molecular Biology (Enzyme
CA 02335253 2000-12-15
- 75 -
Nomenclature, Academic Press, Inc., 1992, pp. 24-154) among which the
following are
particularly preferred: cellobiose: oxygen-1-oxidoreductase (cellobiose
reductase)
1.1.3.25, cellobiose: quinone-1-oxidoreductase 1.1.5.1, bilirubin oxidase
1.3.3.5, cyto-
chrome oxidase 1.9.3, oxygenases, lipoxygenases 1.13, 1.14, superoxide
dismutase
1.15.11, ferrioxidase, for example ceruloplasmin 1. 16.3. 1, especially
preferred being
the enzymes of class 1. 10 which act on related compounds. They catalyze the
oxidation
of biphenols and ascorbates. Suitable acceptors are NAD+, NADP+ (1.10.1),
cytochrome
(1.10.2), oxygen (1.10.3) or others (1.10.99). Among these, particularly
preferred as
acceptors are the enzymes of class 1. 10.3 with oxygen (02) as acceptor.
to Particularly preferred among the enzymes of this class are catechol oxidase
(tyrosinase)
(1.10.3.1), L- ascorbate oxidase (1.10.3.3), O-aminophenol oxidase (1.10.3.4)
and
laccase (benzenediol: oxygen oxidoreductase) (1.10.3.2), the laccases
(benzenediol:
oxygen oxidoreductase) (1.10.3.2) being particularly preferred.
Other particularly preferred enzymes are those of group 1.11 which act on a
peroxide as
acceptor. Only subclass (1.11.1) contains peroxidases. Especially preferred
here are
cytochrome C peroxidases (1.11.1.5), catalase (1.11.1.6), peroxidase
(1.11.1.7), iodide
peroxidase, (1.11.1.8), glutathione peroxidase (1.11.1.9), chloride peroxidase
(1.11.1.10), L-ascorbate peroxidase (1.11.1.11), phospholipid hydroperoxide
glutathione
2o peroxidase (1.11.1.12), manganese peraxidase (1.11.1.13) and diarylpropane
peroxidase
(ligninase, lignin peroxidase) (1. 11. 1. 14).
b) At least one suitable oxidant,
c) At least one mediator selected from the group consisting of hydroxylamines,
hydroxylamine derivatives, hydroxamic acids, hydroxamic acid derivatives,
aliphatic,
cycloaliphatic, heterocyclic or aromatic compounds containing at least one N-
hydroxy,
oxime, N-oxy or N,N'-dioxy function and/or at least one mediator from the
group of
amides, such as, for example, hydrazides or 1,2,4-triazolidin- 3,5-diones
(urazoles)
3o and/or at least one mediator from the group of imides such as, for example,
the
hydantoins, and/or at least one mediator from the group of oxocarbons.
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Moreover, it is possible to use at least one mediation enhancer selected from
the group
consisting of carbonyl compounds, aliphatic ethers, phenol ethers or olefins
(alkenes),
and/or at least one mediation enhancer selected from the group consisting of
the
abovesaid mediators of the NO-, NOH- or HRN-OH type and/or amides such as the
hydrazides or urazoles and/or the imides such as the hydantoins and/or the
oxocarbons.
It is also possible to use at least one mediation enhancer selected from the
group
consisting of cation radical-generating substances of the phenothiazine and/or
phenoxazine type and/or of the (R = N-N = R) type* (for example, ARTS) or of
aryl-
1o substituted alcohols (nonphenols) such as, for example, veratryl alcohol
and/or phenol
derivatives, such as p-hydroxycinnamic acid, 2,4-dichlorophenol, p-
hydroxybenzenesulfonate, vanillin (4-hydroxy-3-methoxybenzaldehyde), p-
hydroxybenzoic acid, 5- amino-2-hydroxybenzoic acid (S-aminosalycilic acid)
and/orWurster-type radical cation compounds [see Angewandte Chemie 91, pp. 982-
997 (1979); Chem. Unserer Zeit 12, pp. 89-98 (1978); Rompp Chemie Lexikon
[Rompp
Chemical Encyclopedial, 9th edition, 1995) and/or radical anions, for example
semiquinones formed by enzymatic oxidation of hydroquinones.
It is essential for improving the performance of the enzyme/mediator systems
with the
aid of comediators that the mediator/comediator ratio be from 5000 : 1 to 1 :
1, a ratio
2o from 500 : 1 to 1 : 1 being particularly preferred. When several mediators
and
comediators are used at the same time, the ratio of these mediator or
comediator
concentrations depends on the particular combinations employed.
* N means nitrogen, R denotes groups.
These enzymatic oxidation systems according to the invention contain at least
one
oxidant. Suitable oxidants are, for example, air, oxygen, ozone, peroxides,
such as H202,
organic peroxides, per acids such as peracetic, performic, persulfuric,
pernitric,
3o metachloroperoxybenzoic and perchloric acid, per compounds such as
perborates,
percarbonates or.persulfates, or oxygen species and the radicals thereof such
as the OH,
CA 02335253 2000-12-15
OOH and OH+ radical, superoxide (Oz -), dioxygenyl cation Oz+, singlet oxygen,
ozonide (03-), dioxiranes, dioxitanes or Fremy radicals.
Said preferred mediator/mediation enhancer substances of formulas I to XXII
and the
said other mediation-enhancing compounds are represented in Appendix IV and
Appendix IVa.
In the following, by way of an example of enzymatic pulp bleaching, the
improvement
in performance is presented that can be achieved for many types of pulps by
use of a
1o combination of the enzyme component system (ECS) of the invention and the
above-
described oxidation systems with enzyme action-enhancing compounds.
EXAMPLE 18 (Enzyme: lipase/laccase)
15 Enzymatic Bleaching of Softwood (Sulfate Pulp)
g, absolutely dry basis, of wood pulp (02-delignified softwood), pulp
consistency 30%
(about I7 g moist) was added to solutions prepared as follows:
2o A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
benzophenone, 2.5 mg of H202 (30%), 37 moles of violuric acid + 0.37 pmole of
4-
tert.butylurazole with agitation. The pH was adjusted with 0.5 mole/L sulfuric
acid
solution so that, after addition of the wood pulp and the enzyme, the pH was
5.
25 B) To 5 mL of lap water was added 5 mg of lipase from Humicola lanuginosa
(about
25,000 IU), and to this mixture was added an amount of laccase from Trametes
versicolor sufficient to give an activity of IS U (1 U = conversion of 1 pmole
of
ABTS/min/mL of enzyme) per gram of wood pulp.
Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
3o pulp, the material was mixed in a dough mixer for 2 minutes. The material
was then
transferred to a reaction vessel preheated to 45 °C and allowed to
incubate I-4 hours at
atmospheric pressure.
CA 02335253 2000-12-15
_ ')g _
The pulp was washed over a nylon screen (30pm) and extracted for one hour at
60 °C,
2% pulp consistency and using 8% NaOH per gram of wood pulp. The pulp was
again
washed, after which the kappa number was determined. For results see Table 7.
EXAMPLE 19 (Enzyme: lipase/peroxidase)
Enzymatic Bleaching of Softwood (Sulfate Pulp)
5 g, absolutely dry basis, of wood pulp (OZ-delignified softwood), pulp
consistency 30%
(about 17 g moist) was added to solutions prepared as follows:
A) To 20 mL of tap water were added 1 mg of tetradecanoic acid, 5 mg of
benzophenone, 2.5 mg of H202 (30%), 37 pmoles of violuric acid + 0.37 pmole of
4-
~ s tert.butylurazole with agitation. The pH was adjusted with 0.5 mole/L
sulfuric acid
solution so that, after addition of the wood pulp and the enzyme, the pH was
7.
B) To 5 mL of tap water was added 5 mg of lipase from Humicola lanuginosa
(about
25,000 IU) and 0.1 mg of peroxidase (horseradish) per gram of wood pulp.
2o Solutions A and B were combined and diluted to 33 mL. After addition of the
wood
pulp, the material was mixed in a dough mixer for 2 minutes. The material was
then
transferred to a reaction vessel preheated to 45 °C and allowed to
incubate 1-4 hours at
atmospheric pressure.
The pulp was washed over a nylon screen (30 pm) and extracted for one hour at
60 °C,
25 2% pulp consistency and using 8% NaOH per gram of wood pulp. The pulp was
again
washed, after which the kappa number was determined. For results see Table 7.
35
CA 02335253 2000-12-15
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TABLE 7
SYSTEM % DELIG. % DELIG.
(lipase/lacc.) (lipase/peroxid.)
ECS + lipase system
(+ laccase + violuric acid) +* 44 ---
ECS + lipase system
to (+ peroxidase + violuric acid) +* --- 38
Laccase (+ violuric acid) +* 27 --
Peroxidase (+ violuric acid) +* --- 28
* Mediation enhancer 4-tert.butylurazole
25
35
45
55
CA 02335253 2000-12-15
APPENDIX I
System Component
2 of the Enzyme
Component System
(ECS) of the Invention
s The fatty acids
used in the process
of the invention
as a source of
peracid are, for
example:
1) Saturated fatty
acids
Butanoic acid (butyric acid)
toPentanoic acid (valeric acid)
Hexanoic acid (caproic acid)
Heptanoic acid (enanthic acid)
Octanoic acid (caprylic acid)
Nonanoic acid (pelargonic acid)
1 Decanoie acid (capric acid)
s
Undecanoic acid
Dodecanoic acid (lauric acid)
Tridecanoic acid
Tetradecanoic acid(myristic acid)
2oPentadecanoic acid
Hexadecanoic acid (palmitic acid)
Heptadecanoic acid
Octadecanoic acid (stearic acid)
Nonadecanoic acid
2sEicosanoic acid (arachic acid)
Heneicosanoic acid
Docosanoic acid (behenic acid)
Tricosanoic acid
Tetracosanoic acid(lignoceric acid)
3oPentacosanoic acid
Hexacosanoic acid (cerotic acid)
Octacosanoic acid
Triacontanoic acid(melissic acid)
CA 02335253 2000-12-15
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2) Unsaturated fatty acids
10-Undecenoic acid
9-cis-Dodecenoic acid (lauroleic acid)
9-cis-Tetradecenoic acid (myristoleic
acid)
9-cis-Hexadecenoic acid (paimitoleic
acid)
6-cis-Octadecenoic aeid (petroselic acid)
6-trans-Octadecenoic acid (petroselaidic
acid)
9-cis-Octadecenoic acid (oleic acid)
l0 9-trans-Octadecenoic acid (elaidic acid)
9-cis, 12 cis-Octadecadienoic acid (linoleic acid)
9-trans, 1 2-trans-Octadecadienoic acid(linolaidic acid)
9-cis, 12-cis,15-cis-Octadecatrienoic (linolenic acid)
acid
9-trans, 11 -trans, 1 3-trans- Octadecatrienoic(a,-eleostearic
acid acid)
9-cis, 11 -trans, 1 3-trans-Octadecatrienoic(f3-eleostearic
acid acid)
9-cis-Icosenic acid (gadoleic acid)
Icosa-5,8,11,14-tetraenoic acid (arachidic acid)
13-cis-Docosenoic acid (erucic acid)
13-trans-Docosenoic acid (brassidic acid)
4,8,12,15,19-Docosapentaenoic acid (clupanodonic
acid)
3) Polyunsaturated fatty acids
9,12-Octadecadienoic acid (linoleic acid)
9,12,1 5-Octadecatrienoic acid (linolenic acid)
5,9,1 2-Octadecatrienoic acid
9,11,13-Octadecatrienoic acid (eleostearic acid)
9,11,13,15-Octadecatetraenoic acid(parinaric acid)
5,11,14-Icosatrienoic acid
5, 8,11,1 4-Icosatetraenoic acid (arachidic acid)
4, 8,12,1 S,1 8-Icosapentaenoie
acid
CA 02335253 2000-12-15
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4,8,12,15,19-Docosapentaenoic acid (clupanodonic acid)
4,8,12,15,18,21 -Tetracosahexaenoic acid (nisinic acid)
Particularly preferred are tetradecanoic acid (myristic acid) and dodecanoic
acid (lauric
s acid).
APPENDIX II
System Component 4 (Ketones) of the Enzyme Component System (ECS) of the
l0 Invention
Particularly preferred are carbonyl compounds of general formula I:
O
15 R~RZ
I
The Rl and RZ groups can be equal or different and denote aliphatic or
aromatic groups.
Moreover, the R' and RZ groups can form a ring containing besides carbon also
2o heteroatoms such as nitrogen, oxygen and sulfur.
Particularly preterred are 1,2-diketones (formula II), 1,3-diketones (formula
III),
polyketones (polyketides) and the tautomeric enols (formula IV):
O
3~ R4 O O O OH
R II R~R6 ~ R~~Rs
O
II III IV
CA 02335253 2000-12-15
- 83 -
wherein the R3 to R6 groups, once again, can be equal or different and denote
aliphatic
or aromatic groups. Moreover, groups R3 and R4 and groups RS and R6, together,
can
form a ring containing besides carbon also heteroatoms such as nitrogen,
oxygen or
sulfur. The possibility of tautomerization or formation of a resonance hybrid
is
particularly important in this case.
Besides general carbonyl compounds, particularly preferred are ketones, such
as, in
general hydroxyketones, a,,13-unsaturated ketones, oxycarboxylic acids,
quinones and
halogenated ketones.
Particularly preferred among these are the following:
Acetone, methyl ethyl ketone, diethyl ketone, methyl n-butyl ketone, methyl
isobutyl
ketone, cyclohexanone, cyclopentanone, 2-methylcyclohexanone, 3-
methylcyclohexanone, 4-methylcyclohexanone, dihydroxyacetone, diacetyl
monohydrazone, diacetyl dihydrazone, acetophenone, p-hydroxyacetophenone,
1 -phenyl-3-butanone, 3-pentanone, 4-heptanone, 2-nonanone, cycloheptanone,
cyclooctanone, cyclodecanone, cyclododecanone, dimethyl ketone, ethyl propyl
ketone,
methyl amyl ketone, acetylacetone, pinacoline, methyl isopropyl ketone, methyl
2o isoamyl ketone, ethyl amyl ketone diisopropyl ketone, diisobutyl ketone,
methyl vinyl
ketone, methyl isopropenyl ketone, mesityl oxide, isophorone, hydroxyacetone,
methoxyacetone, 2,3-pentanedione, 2,3-hexanedione, phenylacetone,
propiophenone,
benzophenone, benzoin, benzil, 4,4'-dimethoxybenzil, 4'-methoxyacetophenone,
3'- methoxyacetophenone, O-ethylbenzoin, (2-methoxyphenyl)acetone, (4-
methoxyphenyl)acetone, methoxy-2-propanone, glyoxylic acid, benzyl glyoxylate,
benzylacetone, methyl benzyl ketone, methylcyclohexyl ketone, 2-decanone,
dicyclohexyl ketone, 3,3-dimethyl-2-butanone, methyl isobutyl ketone, methyl
isopropyl ketone,2-methyl-3-heptanone, 5-methyl-3-heptanone, 6-methyl-5-hepten-
2-
one, 5-methyl-2-hexanone, 3-nonanone, 5-nonanone, 2-octanone, 3-octanone, 2-
3o undecanone, 1,3- dichloroacetone, 1-hydroxy-2-butanone, 3-hydroxy-2-
butanone, 4-
hydroxy-4-methyl-2-pentanone, 2-(1S)- adamanantone, anthrone,
bicyclo(3.2.0)hept-2-
en-6-one, cis-bieyclo(3.3.0)octan-3,7-dione, (1S)- (-)-camphor, p-chloranil,
CA 02335253 2000-12-15
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cyclobutanone, 1,3-cyclohexanedione, 1,4-cyclohexanedione monoethylene ketal,
dibenzosuberone, ethyl 4-oxocyclohexanecarboxylate, 9-fluorenone, 1,3-
indandione,
methyl- cyclohexanone, phenylcyclohexanone, 4-propylcyclohexanone, 1,2,3,4-
tetrahydro-1-naphthalenone, 1,2,3,4-tetrahydro-2-naphthalenone, 3,3,5-
trimethylcyclo-
hexanone, 3-acetoxy-2-cyclohexen-1-one, benzylideneacetone, (R)-(-)-carvone,
(S)-(-)carvone, curcumin, 2-cyclohexen-I-one, 2,3-diphenyl-2- cyclopropen-1-
one, 2-
hydroxy-3-methyl-2-cyclopentene-1-one, isophorone, a,-ionone, f3-ionone, 3-
methoxy-2-
cyclohexen-1-one, 3-methyl-2-cyclopenten-1-one, 3-methyl-3-penten-2-one, (R)-
(+)-
pulegone, tetraphenyl-2,4-cyclopentadien-1-one, 2,6,6-trimethyl-2-cyclohexen-
1,4-
~o dione, 2-acetylbenzoic acid, 1-acetylnaphthalene, 2-acetylnaphthalene,
3'-aminoacetophenone, 4'-aminoacetophenone, 4'-cyclohexylacetophenone, 3',4'-
diacetoxyacetophenone, diacetylbenzene, 2',4'-dihydroxyacetophenone, 2',5'-
dihydroxyacetophenone, 2',6'-dihydroxyacetophenone, 3,4-dimethoxyacetophenone,
2'-hydroxyacetophenone, 4'-hydroxyacetophenone, 3'-methoxyacetophenone, 4'-
15 methoxyacetophenone, 2'-methylacetophenone, 4'-methylacetophenone,
2'-nitroacetophenone, 3'-nitroacetophenone, 4'-phenylacetophenone, 3,'4',5'-
trimethoxy-
acetophenone, 4'-aminopropiophenone, benzoylacetone, benzoylpropionic acid,
benzylideneacetophenone, cyclohexyl phenyl ketone, desoxybenzoin, 4',4'-
dimethoxybenzil, 1,3-diphenyl-1,3-propanedione, ethylbenzoyl acetate, ethyl
2o phenylglyoxylate, 4'- hydroxypropiophenone, 1,3-indandione, 1-indanone,
isopropyl
phenyl ketone, 6-methoxy-1,2,3,4- tetrahydronaphthalen-1-one, methylphenyl
glyoxylate, phenylglyoxylonitrile, 1-phenyl-1,2-propanedione 2-oxime,
valerophenone,
2-acetyl-y-butyrolactone, 2-acetylpyrrole, l -benzylpiperidin-4-one,
dehydroacetic acid,
3,4-dihydro-4,4-dimethyl-2H-pyran-2-one, 1,4-dihydro-4-pyridinone, N-eth-
25 oxycarbonyl-4-piperidinone, 2-methyl furyl ketone, 5-hydroxy-2-
hydroxymethyl-4H-
pyran-4-one, 3-hydroxy-2-methyl-4-pyranone, 3-indolyl methyl ketone, isatin, 1-
methyl-4-piperidinone, methyl 2-pyridyl ketone, methyl 3-pyridyl ketone,
methyl 4-
pyridyl ketone, methyl 2-thienyl ketone, phenyl 2-pyridyl ketone, phenyl 4-
pyridyl
ketone, tetrahydrofuran-2,4-dione, tetrahydro-4H-pyran-4-one, 2,2,6,6-
tetramethyl-4-
3o piperidone, xanthone, acenaphthene quinone, pyruvic acid, (1 R)-(-)-camphor
quinone,
(1S)-(+)-camphor quinone, 3,5-ditert.butyl-o-benzoquinone, 1,2-dihydroxy-3,4-
cyclobutendione, ethyl (2-amino-4-thiazolyl)glyoxylate, ethyl pyruvate, 2,3-
CA 02335253 2000-12-15
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hexanedione, 3,4- hexanedione, 3-methyl-2-oxobutyric acid, 3-methyl-2-
oxovaleric
acid, 4-methyl-2-oxovaleric acid, 2- oxobutyric acid, 2,3-pentandione, 9,10-
phenanthrene quinone, acetoacetanilide,2-acetyl-y-butyrolactone, 2-acetylcyclo-
pentanone, allyl acetoacetate, benzoylacetone, tert.butyl acetoacetate, 1,3-
cyclopentanedione, diethyl 3-oxoglutarate, dimethyl acetylsuccinate, dimethyl
3-
oxoglutarate, 1,3-diphenyl-1,3-propanedione, ethyl acetoacetate, ethyl
benzoylacetate,
ethyl butyrylacetate, ethyl 2-oxocyclohexanecarboxylate, ethyl 2-
phenylacetoacetate,
methyl acetoacetate, 2-methyl-1,3- cyclohexanedione, 2-methyl-1,3-
cyclopentanedione,
methyl isobutyrylacetate, methyl 3-oxopentanoate, methyl pivaloylacetate, 3-
oxoglutaric acid, tetrahydrofuran-2,4-dione, 2,2,6,6-tetramethyl-3,5-
heptanedione, 3-benzoylpropionic acid, 1,4-cyclohexanedione, dimethyl
acetylsuccinate, ethyl levulinate, 2-aminoanthraquinone, anthraquinone, p-
benzoquinone, 1,4-dihydroxyanthraquinone, 1,8-dihydroxyanthraquinone,
2-ethylanthraquinone, methyl-p-benzoquinone, 1,4-naphthoquinone, tetramethyl-p-
benzoquinone, 2,2-dimethyl-1,3-dioxan-4,6-dione, 2-benzoylbenzoic acid, 3-
benzoyl-
propionic acid, 5,6-dimethoxyphthataldehydic acid, levulinic acid, methyl
trans-4-oxo-
2-pentenoate, phthalaldehydic acid, terephthalaidehydic acid, dibutyl maleate,
dibutyl
succinate, dibutyl phthalate, dicyclohexyl phthalate, diethyl
acetamidomalonate, diethyl
adipate, diethyl benzylmalonate, diethyl butylmalonate, diethylethoxymethylene-
2o malonate, diethyl ethylmalonate, diethyl fumarate, diethyl glutarate,
diethyl
isopropylidenemalonate, diethyl maleate, diethyl malonate, diethyl
methylmalonate,
diethyl oxalate, diethyl 3-oxoglutarate, diethyl phenylmatonate, diethyl
phthalate,
diethyl pimelate, diethyl sebacate, diethyl suberate, diethyl succinate,
diisobutyl
phthalate, dimethyl acetylene- dicarboxylate, dimethyl acetylsuccinate,
dimethyl
adipate, dimethyl 2-aminoterephthalate, dimethyl fumarate, dimethyl
glutaconate,
dimethyl glutarate, dimethyl isophthalate, dimethyl malonate, dimethylmethoxy-
malonate, dimethyl methylenesuccinate, dimethyl oxalate, dimethyl 3-
oxoglutarate,
dimethyl phthalate, dimethyl succinate, dimethyl terephthalate, ethylene
glycol
diacetate, ethylene glycol dimethacrylate, monoethyl fumarate, monomethyl
malonate,
3o monoethyl adipate, monomethyl phthalate, monomethyl pimelate, monomethyl
terephthalate, 1,2-propylene glycol diacetate, triethyl methanetricarboxylate,
trimethyl
1,2,3-propanetricarboxylate, 3-acetoxy-2-cyclohexen-1-one, allyl acetoacetate,
allyl
CA 02335253 2000-12-15
-86-
cyanoacetate, benzyl acetoacetate, tert.butyl acetoacetate, butyl
cyanoacetate,
chlorogenic acid hemihydrate, coumarin-3-carboxylic acid, diethyl ethoxy-
carbonylmethanephosphonate, dodecyl gallate, dodecyl 3,4,5-trihydroxybenzoate,
(2,3-
epoxypropyl) methacrylate, (2-ethoxyethyl) acetate, ethyl
acetamidocyanoacetate, ethyl
2-aminobenzoate, ethyl 3-aminopyrazol-4-carboxylate, ethyl benzoxylacetate,
ethyl
butyrylacetate, ethyl cyanoacetate, ethyl 2-cyano-3-ethoxyacrylate, ethyl
cyanoformate,
ethyl 2-cyanopropionate, ethyl 3,3-diethoxypropionate, ethyl 1,3-dithian-2-
carboxylate,
ethyl 2-ethoxyacetate, ethyl 2-furancarboxylate, ethyl levulinate, ethyl
mandelate, ethyl
gallate, ethyl 2-methyllactate, ethyl 4- nitrocinnamate, ethyl oxamate, ethyl
2-
oxocyclohexanecarboxylate, ethyl 4-oxocyclohexane- carboxylate, ethyl 5-
oxohexanoate, ethyl 2-phenylacetoacetate, ethyl 4-piperidinecarboxylate,
ethyl 2-pyridinecarboxylate, ethyl 3-pyridinecarboxylate, ethyl 4-
pyridinecarboxylate,
ethyl thioglycolate, ethyl 3,4,5-trihydroxybenzoate, 2-hydroxyethyl
methacrylate, 2-
hydroxypropyl methacrylate, 3-indole acetate, 2-methoxyethyl acetate, 1-
methoxy-2-
propyl acetate, methyl 2- aminobenzoate, methyl 3-aminocrotonate, methyl
cyanoacetate, methyl 4-cyanobenzoate, methyl 4- formylbenzoate, methyl 2-
furancarboxylate, methyl isobutyrylacetate, methyl methoxyacetate, methyl 2-
methoxybenzoate, methyl 3-oxopentanoate, methyl phenylglyoxylate, methyl
phenyl-
sulfinylacetate, methyl pivatoylacetate, methyl 3-pyridinecarboxylate, 5-
2o nitrofurfurylidene diacetate, propyl gallate, propyl 3,4,5-
trihydroxybenzoate, methyl 3-
methylthiopropionate, acetamide, acetanilide, benzamide, benzanilide, N,N-
diethylacetamide, N,N-dimethylformamide, N,N-diethyl-3-methyl- benzamide,
diethyltoluamide, N,N-dimethylacetamide, N,N-diphenylacetamide, N-
methylformamide, N-methylformanilide, N-acetylthiourea, adipic acid diamide, 2-
aminobenzamide, 4-aminobenzamide, succinic acid diamide, malonic acid diamide,
N,N'-methylene diacrylamide, oxalic acid diamide, pyrazine-2-carboxamide,
pyridine-4-carboxamide, N,N,N',N'-tetramethylsuccinic acid diamide,
N,N,N',N'-tetramethylglutaric acid diamide, acetoacetanilide, benzohydroxamic
acid,
cyanoacetamide, 2-ethoxybenzamide, diethyl acetamidomalonate, ethyl
3o acetamidocyanoacetate, ethyl oxamate, hippuric acid Na salt, N-(hydroxy-
methyl)acrylamide, L-(-)-lactamide, 2'-nitroacetanilide, 3'- nitroacetanilide,
4'-
nitroacetanilide, paracetamol, piperine, salicylanilide, 2-acetyl-~y-
butyrolactone, y-
CA 02335253 2000-12-15
-87-
butyrolactone, s-caprolactone, dihydrocoumarin, 4-hydroxycoumarin, 2-(SH)-
furanone,
2,5-dihydro-5-methoxy-2-furanone, phthalide, tetrahydrofuran-2,4-dione, 2,2,6-
trimethyl-1,3-dioxin-4-one, y- valerolactone, 4-amino-1,3-dimethyluracil,
barbituric
acid, O-benzyloxycarbonyl-N-hydroxysuccinimide, succinimide, 3,6-dimethyl-
piperazin-2,5-dione, 5,5-diphenylhydantoin, ethyl 1,3-dioxoisoindoline-2-
carboxylate,
9-fluorenylmethylsuccinimidyl carbonate, hydantoin, maleimide, 3-methyl-1-
phenyl-2-
pyrazolin-5-one, 1-methyl-2-pyrrolidone, methyluracil, 6-methyluracil,
oxindole,
phenytoin, 1-(2H)-phthalazinone, phthalimide, 2,5-piperazinedione, 2-
piperidinone, 2-
pyrrolidone, rhodanine, saccharin, 1,2,3,6-tetrahydrophthalimide, 1,2,3,4-
tetrahydro-
l0 6,7-dimethoxyquinazolin-2,4-dione, 1,5,5-trimethyl-hydantoin, 1-vinyl-2-
pyrrolidone,
ditert.butyl dicarbonate, diethyl carbonate, dimethyl carbonate, dimethyl
dicarbonate,
diphenyl carbonate, 4,5-diphenyl-1,3-dioxol-2-one, 4,6- diphenylthieno-(3,4-d)-
1,3-
dioxo[-2-one 5,5-dioxide, ethylene carbonate, magnesium methoxide methyl
carbonate,
monomethyl carbonate Na salt, propenyl carbonate, N-allylurea,
azodicarbonamide, N-
benzylurea, biuret, 1,1'-carbonyldiimidazol, N,N-dimethylurea, N-ethylurea, N-
formylurea, urea, N-methylurea, N-phenylurea, 4-phenylsemicarbazide,
tetramethylurea, semicarbazide hydrochloride, diethyl azodicarboxylate, methyl
carbamate, 1-(4-methoxyphenyl)-2-(2- methoxyphenoxy)ethanone and 1-(4-
methoxyphenyl)-2-(2-methoxyphenoxy)ethanol.
Also preferred are anhydrides, such as the following:
Benzoic anhydride, benzene-1,2,4,5-tetracarboxylic acid-1,2,4,5-dianhydride,
3,3',4,4'-
benzophenonetetracarboxylic anhydride, succinic anhydride, butyric anhydride,
crotonic
anhydride, cis-1,2-cyclo- hexanedicarboxylic anhydride, ditert.butyl
dicarbonate,
dimethyl dicarbonate, dodecenylsuccinic anhydride, Epicon B 4400, acetic
anhydride,
glutaric anhydride, hexanoic anhydride, isatoic anhydride, isobutyric
anhydride,
isovaleric anhydride, malefic anhydride, 1,8-naphthalenedicarboxylic
anhydride, 3-
nitrophthalic anhydride, 5-norbornene-2,3-dicarboxylic anhydride, phthalic
anhydride,
2-phenylbutyric anhydride, pivalic anhydride, propionic anhydride, cis-1,2,3,6-
tetrahydrophthalic anhydride and valeric anhydride.
CA 02335253 2000-12-15
_$g_
Particularly preferred are benzophenones such as the following:
Benzophenone, 4-aminobenzophenone, 2-amino-5-chlorobenzophenone,
benzophenone-2-carboxylic acid, (S)-(-)-2-(N-benzopropyl)aminobenzophenone,
4,4'-
bis(dimethylamino)benzophenone, 4,4'- bis(diethylamino)benzophenone, 3,4-
dimethoxybenzophenone, 4,4'-dihydroxybenzophenone, 2,4- dihydroxybenzophenone,
4-hydroxybenzophenone, 2-hydroxy-4-methoxybenzophenone, 4-methoxybenzo-
phenone, 4,4'-dimethoxybenzophenone, 2,2',4,4'-tetrahydroxybenzophenone and 2-
chlorobenzophenone.
APPENDIX III
Polymerization Catalysts: Phenolic Compounds, Phenol Derivatives or Other
Phenolic Polycyclic Compounds with a Number of Oxidizable Hydroxyl Groups
Preferably, such polymerization catalysts are, for example, the following:
alizarin, 5-amino-2-hydroxybenzoic acid, 3-aminophenol, pyrocatechol, 2,2-
bis(4-
hydroxyphenyl)- propane, bis(4-hydroxyphenyl)methane, quinalizarin, 4-chloro-1-
naphthol, coniferyl alcohol, 2,4-di- aminophenol dihydrochloride, 3,5-dichloro-
4-
hydroxyaniline, 1,4-dihydroxyanthraquinone, 2,2-di- hydroxybiphenyl, 4,4-
dihydroxybiphenyl, 2,3-dihydroxynaphthalene, 2,6-diisopropylphenol, 3,5-di-
methoxy-
4-hydroxybenzhydrazine, 2,5-ditert.butylhydroquinone, 2,6-ditert.butyl-4-
methylphenol, 4-hydroxybiphenyl, 2-hydroxydiphenylmethane, 2-(2-hydroxy-
phenyl)benzothiazole, 5-indanol, 2-iso- propoxyphenol, 4-isopropyl-3-
methylphenol, 5-
isopropyl-2-methylphenol, 4-isopropylphenol, lauryl gallate, 2-naphthol, 4-
nonylphenol,
3-(pentadecyl)phenol, 2-propylphenol, 4-propylphenol, purpurine, pyrogallol, 4-
(1,1,3,3-tetramethylbutyl)phenol, 1,2,4-trihydroxybenzene, 2,4,6-
trimethylphenol,
2,3,5-trimethylphenol, 2,3,6-trimethylphenol, 3,4,5-trimethylphenol, 6,7-
dihydroxy-4-
methyl coumarin, 2-(2-hydroxyethoxy)benzaldehyde, 1 -naphthol,
nordihydroguaiaretic
acid, octyl gallate, silibinin, 3,4,6-trihydroxyben-zoate-octylester, 2,4,6-
tritert.butyl-
phenol, 2,4-ditert.butylphenol, 2,6-dichlorophenol, indophenol, ethoxyquin, 1-
3o aminoanthraquinone, 2-amino-5-chlorobenzophenone, 4-aminodi- phenylamine, 7-
CA 02335253 2000-12-15
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amino-4-hydroxy-2-naphthalenesulfonic acid, 2-(4-aminophenyl)-6-
ethylbenzothiazole,
benzanthrone, trioctyl trimellitate, trans-chalcone, bis(4-aminophenyl)amine
sulfate,
2,2'- ethylidenebis (4,6-ditert.butylphenol), 2,2-bis(2,6-dibromo-4-(2-hydroxy-
ethoxyphenyl)propane, bis(3,5-ditert.butyl-4-hydroxyphenyl)methane, 2,2-
bis(3,5-
dichloro-4-hydroxyphenyl)propane, Bismarck Brown Y, 1-bromophthalein, 4-
butylaniline, 2-tert.butyl-5-methylphenol, 1-chloro-anthraquinone, 2-chloro-
anthraquinone, triallyl 1,3,5-benzenetricarboxylate, l,1,1-tris(hydroxy-
methyl)propane,
tri-methacrylate, pentaerythrityl triacrylate, 1,2,4-trivinylcyclohexane,
trans,cis-
cyclododeca-1 5,9-tri- ene, pentaerythritol tetrabenzoate, 4,4'-methylene-
bis(2,6-
to ditert.butylphenol), 4,4'-isopropylidene-bis(2,6-dichlorophenol), 4,4'-
isopropylidene-
bis(2,6-dibromophenol), 4,4'-isopropylidene-bis[2-(2,6-
dibromophenoxy)ethanol, 2,2'-
ethylidene-bis(4,6-ditert.butylphenol), 3-tert.butyl-4-hydroxy-5-methylphenol,
5-
tert.butyl-4-hydroxy-2-methylphenol, syringaldazine, 4,4'-
dimethoxytriphenylmethane
and di-sec.butylphenol.
Also particularly preferred are compounds with several hydroxyl groups, such
as:
ellagic acid, gallic acid, gallein, gallangin, myoinositol, morin, nitranilic
acid,
phenolphthalein, purpurin, purpurogallin, quinizarin, chrysazin, quercitin,
quinhydrone,
chloranilic acid, carmine, rhodizonie acid, croconic acid, meilitic acid,
hematoxylin, 9-
phenyl-2,3,7-trihydroxy-6-fluorene, 9-methyl-2,3,7- trihydroxy-6-fluorene,
2o tetrahydroxy-p-benzoquinone, 2,2',4,4'-tetrahydroxybenzophenone, Pyragallol
Red, 1-
nitrophloroglucinol, 1,4-dihydroxyanthraquinone, 5,8-dihydroxy-1,4-
naphthoquinone,
hexa- oxocyclohexane octahydrate, 5,7-dihydroxyflavanone, 3',4'-
dihydroxyflavanone,
glyoxal hydrate, 1,3,5-tris(2-hydroxyethyl)isocyanuric acid, quinalizarin and
2,4,5-
trihydroxybenzamine.
30
CA 02335253 2000-12-15
APPENDIX IV
Appendix IV shows the formulas of mediators/mediation enhancers (NO-, NOH- and
HNR-OH compounds) which according to the invention can be added to the enzyme
component system (ECS) together with oxidoreductases, such as, for example:
Hydroxylamines (linear or cyclic, aliphatic or aromatic, heterocyclic) of
general
formula I)
1 2
R,N,R
i
OH
(I)
such as compounds of general formula II
R
~X
1s N
R ~s
R
is (II)
2o such as compounds of general formula III:
R5
6
R ~ ~ X
N
R
R8 R (III)
such as compounds of general formula IV:
CA 02335253 2000-12-15
-91-
R5
6
R
X
N
R i
R$ O ~ R 16 (IV)
such as compounds, namely derivatives, of 1-hydroxybenzotriazole
and the tautomeric benzotriazole 1-oxide, as well as the esters and salts
thereof, particularly compounds of formula V
R5
6
R / N
~N
R N,
R$ OH
such as, for example, the following compounds:
1 -hydroxybenzotriazole
1 -hydroxybenzotriazole, sodium salt
1 -hydroxybenzotriazole, potassium salt
1 -hydroxybenzotriazole, lithium salt
1 -hydroxybenzotriazole, ammonium salt
1 -hydroxybenzotriazole, caicium salt
1 -hydroxybenzotriazole, magnesium salt
1 -hydroxybenzotriazole-6-sulfonic acid, monosodium salt
1 -methoxy-1 H-benzotriazole
1 -acetoxy- 1 H-benzotriazole
1 -hydroxy-(4,5-f)-dioxolo-1 H-benzotriazole
1 -hydroxy-6-methyl-3H-benzotriazole
1-hydroxy-6-nitro-1 H-benzotriazole
CA 02335253 2000-12-15
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1 -hydroxy-5,6-dimethyl-1 H-benzotriazole
1 -hydroxy-6-methoxy-1 H-benzotriazole
1 -hydroxy-5,6-dimethoxy-1 H-benzotriazole
1 -hydroxy-1 H-benzotriazole-6-carboxylic acid
1,5-dihydroxy-1 H-benzotriazole
1 -hydroxy-1 H-benzotriazole-6-sulfonic acid hydrazide
1 -hydroxy-1 H-benzotriazole-6-carboxamide
1 -hydroxy-5-methoxy-1 H-benzotriazole
6-amino-1 -hydroxy-1 H-benzotriazole
6-amino-5-methoxy-1 H-benzotriazole
6-chloro- 1 -hydroxy- 1 H-benzotriazole
6-acetamido-1 -hydroxy-1 H-benzotriazole
1 -hydroxy- 1 H-benzotriazole-6-carboxylic acid ethyl ester
1 -hydroxy-4-nitro-1 H-benzotriazole
4-chloro-1 -hydroxy-1 H-benzotriazole
1 -hydroxy-6-tert. butyl- 1 H-benzotriazole
6-cyclohexyl-1 -hydroxy-1 H-benzotriazole
6-isopropyl-1 -hydroxy-1 H-benzotriazole
1 -hydroxy-6-phenyl-1 H-benzotriazole
3-methyl-3H-benzotriazole 1 -oxide
2-phenyl-ZH-benzotriazole 1 -oxide
such as compounds of general formula A (cyclic N-hydroxy compounds):
X Y
I I
- C - N -- C -
I
OH
(A)
CA 02335253 2000-12-15
-93-
Such as compounds of general formula VI, VII, VIII or IX:
R3 X X
R4 / I 'R
N-OH N-OH
$R \ ~ gR
R6 Y Y
VI VII
OH
I
Y_~ _N_ ~.X
X 13
R9 ~ ls~
/ \
\N - OH
11R
\ / Rl
y
Y R, 6 R15
VIII IX
for example, compounds such as:
N-hydroxyphthalimide and optionally substituted N-hydroxyphthalimide
derivatives, N-
hydroxymaleimide and optionally substituted N-hydroxymaleimide derivatives, N-
CA 02335253 2000-12-15
-94-
hydroxynaphthalimide and optionally substituted N-hydroxynaphthalimide
derivatives,
N-hydroxysuccinimide and optionally substituted N-hydroxysuccinimide
derivatives,
such as, for example:
N-hydroxyphthalimide, N-hydroxybenzene-1,2,4-tricarboximide,
N,N'-dihydroxypyromellitic acid diimide,
N,N'-dihydroxybenzophenone-3,3',4,4'-tetracarboxylic acid diimide,
for example, (formula VII):
1o N-hydroxymaleimide,
pyridine-2,3-dicarboxylic acid N-hydroximide
for example (formula VIII):
N-1-hydroxysuccinimide, N-1-hydroxytartarimide,
N-hydroxy-5-norbornene-2,3-dicarboximide,
exo-N-hydroxy-7-oxabicyclo[2.2.1 ]-5-heptene-2,3-dicarboximide,
N-hydroxy-cis-cyclohexane-1,2-dicarboximide,
N-hydroxy-cis-4-cyclohexene-1,2-dicarboximide
for example (formula IX):
N-hydroxynaphthalimide sodium salt
for example (six-membered ring of formula A):
X Y
I I
_- C N _ C ___.
I
OH
(A)
CA 02335253 2000-12-15
-95-
N-hydroxyglutarimide
such as compounds of general formula X or XI (oximes):
OH
N
OH X ~ ~ X
N ~ ~~/
3R~~R4 ,N N
S R I ~ R6
X X X
X XI
for example (formula X):
dimethyl 2-hydroxyiminomalonate
for example (formula XI):
1-methylvioluric acid, 1,3-dimethylvioluric acid, thiovioluric acid, alloxane
4,5-
dioxime, alloxane 5- oxime hydrate (violuric acid) and/or the esters or salts
thereof,
such as compounds from the class of N-aryl-N-hydroxyamides of general formula
XII, XIII and XIV, XIVa, XIVb, XIVc, XIVd and XIVe:
OH
A-N-B XH
CA 02335253 2000-12-15
-yf)-
OH OH
I
A-N-C-N-A XII1
OH
I
N
s D C XIV
OH O
Art-N-CI -Rs
XIVa
OH O OH O
Arl N-CI (R6)p CI-N-Arl
XIVb
OH O
Ar~N-CI
~(CR3R4)q
2o XIVc
CA 02335253 2000-12-15
9'j
OH O
Arl-N-IS-Rs
I I
O
XIVd
OH O
~t-N-p-Rs
Rs
XIVe
1o such as compounds of general formula XV, XVI and XVII (nitroxyl
radicals/nitroxides)
O' I I
I ,N\ ~R3 R3w iNw ,R3
Ar~N~Ar Ar R3iC~R3 R3~R3 R3 R3
XV XVI XVII
Is
CA 02335253 2000-12-15
-98-
such as compounds of general formula XVII a and XVII b (nitroxyl radicals):
2
R2 R R2 R2 R2 RZ R2 R2
R2 R2
R~ R~ R1 ~R~
R~ N R1 R~ N~R1
O~ O~
XVIIa XVIIb
1o Such as compounds of general formula XVIIIa (amides) and XVllI b
(hydrazides):
R
x
R~ ~N~ ~R
I
R R
XVBIa XVIII b
such as compounds of general formula XVIII c (cyclic hydrazides):
X\N~R
G
~X~N~
R
XVIII c
CA 02335253 2000-12-15
-99-
such as urazoles (formula XVIII d) and phthalhydrazides (formula XVIII e):
0
0
/ 'NH R ~ ~ NH
HN NH ~ NH
O O
XVnI d XVIII a
such as compounds of general formula XIX (imides):
R
I
R N R
O O
XIX
such as compounds of general formula XIXa (imides):
R R
I I
R~N~N~R
IYO IYO
XIX a
CA 02335253 2000-12-15
-loo-
such as compounds of general formula XIXb (cyclic imides):
R
I
G~N
>O
N
O I
R
XIX b
such as compounds of general formula XIXc (hydantoin derivatives):
H
~N
>O
N
O H
XIX C
I5 such as compounds of general formula XX, such as a,-hydroxycarbonyl
compounds
of general formula XXa, a-dicarbonyl compounds of general formula XXb,
I3-hydroxycarbonyl compounds of general formula XXc and I3-dicarbonyl
compounds of general formula XXd:
O O
XX a
CA 02335253 2000-12-15
- 1~1 -
O
R2 R II R4 O OH
R
R R
OH
XX b XX c XX d
such as compounds of general formula XXI (linear compounds with double
bonds/enols):
O OH
s~ / R1o
R
OH
XXI
1s such as compounds of general structure XXII (cyclic compounds, groups not
OH,
derivatives of squaric acid, OH group derivatized):
11
O R
//
R12
m
20 XXII
CA 02335253 2000-12-15
-102-
such as, for example:
deltic acid, squaric acid, croconic acid and rhodizonic acid
*The formula descriptions (groups/R ... are given in patent application DE 197
19
857Ø
io APPENDIX IVa
Appendix IVa shows compounds which can be added to the enzyme component system
(ECS) of the invention as mediation enhancers, primarily as additives together
with
mediators and oxidoreductases:
Aliphatic ethers and aryl-substituted alcohols, such as:
2,3-dimethoxybenzyl alcohol, 3,4-dimethoxybenzyl alcohol, 2,4-dimethoxybenzyl
alcohol, 2,6-dimethoxybenzyl alcohol, homovanillic alcohol, ethylene glycol
2o monophenyl ether, 2-hydroxybenzyl alcohol, 4-hydroxybenzyl alcohol, 4-
hydroxy-3-
methoxybenzyl alcohol, 2-methoxybenzyl alcohol, 2,5-dimethoxybenzyl alcohol,
3,4-
dimethoxybenzylamine, 2,4-dimethoxybenzylamine hydrochloride, veratryl
alcohol,
coniferyl alcohol.
olefins (alkenes), for example:
2-allylphenol, 2-allyl-6-methylphenol, allylbenzene, 3,4-
dimethoxypropenylbenzene, p-
methoxystyrene, 1-allylimidazole, 1-vinylimidazole, styrene, stilbene, allyl
phenyl
ether, benzyl cinnamate, methyl cinnamate, 2,4,6-triallyloxy-1,3,5-triazine,
CA 02335253 2000-12-15
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1,2,4-trivinylcyclohexane, 4-allyl-1,2-dimethoxybenzene, 4-tert. benzoic acid
vinyl
ester, squalene, benzoin allyl ether, cyclohexene, dihydropyran and N-
benzylcinnamanilide.
Phenol ethers, such as:
2,3-dimethoxybenzyl alcohol, 3,4-dimethoxybenzyl alcohol,
2,4-dimethoxybenzyl alcohol, 2,6-dimethoxybenzyl alcohol, homovanillic
aicohol, 4-
hydroxybenzyl alcohol, 4-hydroxy-3-methoxybenzyl alcohol, 2-methoxybenzyl
alcohol,
2,5-dimethoxybenzyl aicohol, 3,4-dimethoxybenzylamine, 2,4-
dimethoxybenzylamine
1o hydrochloride, veratryl alcohol, coniferyl alcohol, veratrol, anisole.
Carbonyl compounds, such as:
4-aminobenzophenone, 4-acetylbiphenylbenzophenone, benzil, benzophenone
15 hydrazone, 3,4-dimethoxybenzaldehyde, 3,4-dimethoxybenzoic acid, 3,4-
dimethoxybenzophenone, 4-dimethylaminobenzaldehyde, 4-acetylbiphenyl
hydrazone,
benzophenone 4-carboxylic acid, benzoylacetone, bis(4,4'-
dimethylamino)benzophenone, benzoin, benzoin oxime, N-benzoyl-N-
phenylhydroxylamine, 2-amino-S-chlorobenzophenone, 3-hydroxy-4-
2o methoxybenzaldehyde, 4-methoxybenzaldehyde, anthraquinone-2-sulfonic acid,
4-
methylaminobenzaldehyde, benzaldehyde, benzophenone-2-carboxylic acid,
3,3',4,4'-benzophenonetetracarboxylic dianhydride, (S)-(-)-2-(N-benzylpropyl)-
aminobenzophenone, benzylphenylacetanilide, N-benzylbenzanilide, 4,4'-
bis(dimethylamino)thiobenzophenone, 4,4'-bis(diacetylamino)benzophenone,
25 2-chlorobenzophenone, 4,4'-dihydroxybenzophenone, 2,4-
dihydroxybenzophenone, 3,5-
dimethoxy-4-hydroxybenzaldehyde hydrazine, hydroxybenzophenone, 2-hydroxy-4-
methoxybenzophenone, 4-methoxybenzophenone, 3,4-dihydroxybenzophenone, p-
anisic acid, p-anisaldehyde, 3,4-dihydroxybenzaldehyde, 3,4-dihydroxybenzoic
acid,
30 2,5-dimethoxy-4-hydroxybenzaldehyde, 3,5-dimethoxy-4-hydroxybenzoic acid, 4-
hydroxybenzaldehyde, salicylaldehyde, vanillin, vanillic acid.
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APPENDIX 5
Possible Oxidation Reactions of the Enzyme Component System
1) Hydroxylation reactions
a) Synthesis
of alcohols
b) Hydroxylationof steroids
c) Hydroxylationof terpenes
d) Hydroxylationof benzenes
e) Hydroxylationof alkanes
1o Hydroxylationof aromatic compounds
f)
g) Hydroxylationof double bonds
h) Hydroxylationof nonactivated methyl
groups
i) Dihydroxylation of aromatie compounds
2) Oxidation of unsaturated aliphatics
a) Preparation of epoxides
b) Preparation of compounds by
epoxidation
c) Preparation of arene oxides
d) Preparation of phenols
2o Preparation of cis-dihydrodiols
e)
3) Baeyer-Villiger oxidations
a) Baeyer-Villiger conversion of steroids
4) Oxidation of heterocycles
a) Transformation of organic sulfides
b) Oxidation of sulfur compounds
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c) Oxidation of nitrogen compounds (formation of N-oxides etc.)
d) Oxidation of other heteroatoms
5) Carbon-carbon dehydrogenation
a) Dehydrogenation of steroids
6) Other oxidation reactions
a) Oxidation of alcohols and aldehydes
b) Oxidation of aromatic methyl groups to aldehydes
c) Oxidative coupling of phenols
1o d) Oxidative degradation of alkyl chains (l3-oxidation etc.)
e) Formation of peroxides or percompounds
f) Initiation of free-radical induced chain reactions.
20
30
