U.S. patent application number 09/872922 was filed with the patent office on 2001-11-22 for enzyme containing granule.
This patent application is currently assigned to Novozymes A/S. Invention is credited to Markussen, Erik Kjaer.
Application Number | 20010044403 09/872922 |
Document ID | / |
Family ID | 27220980 |
Filed Date | 2001-11-22 |
United States Patent
Application |
20010044403 |
Kind Code |
A1 |
Markussen, Erik Kjaer |
November 22, 2001 |
Enzyme containing granule
Abstract
This invention relates to an enzyme containing granular
composition comprising: a) an enzyme containing core and b) a
protective substantially continuous layer or coating encapsulating
the core comprising at least 60% of a water soluble compound,
having a molecular weight below 500 grams per mole, a pH below 11
and a constant humidity at 20.degree. C. of more than 81%. The
invention provides an improved stability of enzymes upon
storage.
Inventors: |
Markussen, Erik Kjaer;
(Vaerlose, DK) |
Correspondence
Address: |
NOVOZYMES NORTH AMERICA, INC.
C/O NOVO NORDISK OF NORTH AMERICA, INC.
405 LEXINGTON AVENUE, SUITE 6400
NEW YORK
NY
10174
US
|
Assignee: |
Novozymes A/S
Bagsvaerd
DK
|
Family ID: |
27220980 |
Appl. No.: |
09/872922 |
Filed: |
June 1, 2001 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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09872922 |
Jun 1, 2001 |
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09344877 |
Jun 25, 1999 |
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6268329 |
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60092003 |
Jul 8, 1998 |
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Current U.S.
Class: |
510/392 ;
510/396; 510/446 |
Current CPC
Class: |
C11D 3/0042 20130101;
C12N 11/14 20130101; C11D 3/0084 20130101; C11D 3/39 20130101; C12N
9/96 20130101; C11D 3/38672 20130101; C11D 7/16 20130101; C12N 9/98
20130101; C12N 11/04 20130101; C11D 7/10 20130101 |
Class at
Publication: |
510/392 ;
510/396; 510/446 |
International
Class: |
C11D 003/00; C11D
017/06 |
Foreign Application Data
Date |
Code |
Application Number |
Jun 30, 1999 |
DK |
PA 1998 00876 |
Claims
1. An enzyme containing granule comprising: a) an enzyme containing
core and b) a protective substantially continuous layer or coating
encapsulating the core comprising at least 60% of a water soluble
compound, having a molecular weight below 500 grams per mole, a pH
below 11 and a constant humidity at 20.degree. C. of more than
81%.
2. The granule according to claim 1, wherein said water soluble
compound has a molecular weight between 30-500 g/mole.
3. The granule according to claim 2, wherein said water soluble
compound has a molecular weight between 75-400 g/mole.
4. The granule according to any preceding claim, wherein said water
soluble compound has a solubility of at least 0.1 gram per 100 gram
water.
5. The granule according to any preceding claim, wherein said water
soluble compound has a solubility of at least 10 gram per 100 gram
water.
6. The granule according to any preceding claim, wherein said water
soluble compound has constant humidity at 20.degree. C. of more
than 90%.
7. The granule according to any preceding claim, wherein said core
is a non absorbing core.
8. The granule according to claim 1, wherein said salt is selected
from the group consisting of alkali or earth alkali metal ion salt
of sulfate, sulfite, phosphate, phosphonate, nitrate, chloride,
carbonate and simple organic acids.
9. The granule according to claim 1, wherein said salt is selected
from the group consisting of Na.sub.2HPO.sub.4, Na.sub.3PO4,
(NH.sub.4)H.sub.2PO.sub.4, KH.sub.2PO.sub.4, Na.sub.2SO.sub.4,
K.sub.2SO.sub.4, KHSO.sub.4, ZnSO.sub.4 and sodium citrate.
10. The granule according to any preceding claim, wherein said
coating further comprises one or more protective agents capable of
inactivating components hostile to the enzyme entering the granule
from a surrounding matrix.
11. The granule according to claim 10, wherein said hostile
component is a detergent bleach component.
12. The granule according to claim 11, wherein said protective
agent is selected from the group consisting of reducing agents,
antioxidants and salts of transition metals.
13. The granule according to claim 12, wherein said reducing agent
is a salt of thiosulfate.
14. The granule according to any preceding claim, wherein said
coating constitutes 1-75% w/w of the coated granule.
15. The granule according any preceding claim, wherein said enzyme
containing core comprises a particulate carrier and an enzyme
containing layer.
16. The granule according to claim 15, wherein additional enzyme is
adsorbed into the carrier.
17. The granule according to claim 16, wherein said absorbed enzyme
and/or enzyme containing layer comprises a protective reducing
agent.
18. The granule according to any preceding claim, wherein said
enzyme is selected from the group consisting of oxidoreductases (EC
1.-.-.-), transferases (EC 2.-.-.-), hydrolases (EC 3.-.-.-),
lyases (EC 4.-.-.-), isomerases (EC 5.-.-.-) and ligases (EC
6.-.-.-).
19. The granule according to any preceding claim further comprising
an additional coating.
20. The granule according to claim 19, wherein said additional
coating is a lubricant.
21. A method for producing the granule of claims 1-20 comprising:
a) mixing an enzyme containing core material with a liquid medium
comprising said water soluble compound and, b) removing volatile
components of the liquid medium from the mixture, so as to deposit
the nonvolatile components of the liquid medium as solid coating
layer on the core material.
22. The method according to claim 21 wherein the granule is
obtained by a fluid bed process comprising: a) fluidising an enzyme
containing core material in a fluid bed apparatus, b) introducing a
liquid medium comprising the water soluble compound of the
invention by atomization into the fluid bed, so as to deposit the
nonvolatile components of the liquid medium as solid coating layer
on the core material and, c) removing volatile components of the
liquid medium from the coated core material.
23. A detergent composition comprising the enzyme containing
granular composition of any of the claims 1-20.
24. A cleaning method comprising contacting an object with an
aqueous solution comprising the particulate composition of any of
the claims 1-20.
Description
1. CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation application of U.S.
application Ser. No. 09/344,877, filed Jun. 25, 1999, and claims,
under 35 U.S.C. 119, priority of Danish application no. PA 1998
00876, filed Jun. 30, 1999, and benefit of U.S. provisional
application No. 60/092,003, filed Jul. 8, 1998. Priority from U.S.
application Ser. No. 09/344,877, filed on Jun. 25, 1999, is claimed
under 35 U.S.C. 120, the contents of which are fully incorporated
herein by reference
FIELD OF THE INVENTION
[0002] The present invention relates to an enzyme containing
granule protecting the enzyme from inactivation when the granules
are stored, i.e. improving the storage stability of the enzyme. The
invention further relates to a process for producing such a granule
and to the use of the granule in a number of industrial
applications such as incorporation of the granule in a detergent
composition.
BACKGROUND OF THE INVENTION
[0003] The industrial use of enzymes, notably enzymes of microbial
origin, has become increasingly common. Enzymes are used in
numerous industries, including, for example, the starch-processing
industry and the detergent industry.
[0004] It is well known that enzymes upon storage is liable to be
degraded or inactivated by components (such as oxygen or bleach
components) from the surrounding matrix (such as a detergent),
which are capable of oxidizing or otherwise inactivating the
enzyme. Further it is well known that a low formation of
enzyme-containing dust of granules is desired.
[0005] Since the introduction of enzymes into the detergent
industry, a lot of effort has been devoted to improving the
formulation of enzyme products by applying granulation and coating
of the enzyme so as to both protect the enzyme from degradation as
well as control enzyme dust formation.
[0006] Many granulates are composed of a core particle upon which
an enzyme containing layer is added. The core may also in itself
contain enzyme. To apply the desired properties of this
construction, e.g. color, dust formation, solubility rate, size,
enzyme stability, physical strength etc. the core-enzyme
construction is usually added additional layers of coatings
providing such properties.
[0007] Some coating layers described in the art are complex
multi-component compositions such as in:
[0008] WO 90/09440 which discloses an enzyme containing granulate
1) one coating (or shell) comprising a binder (e.g. kaolin), a
filler (e.g. inorganic salts), granulating agents (e.g. cellulose
fibers providing physical strength), and an enzyme and 2) a second
dust suppressing coating (mono-, di- or triglyceride).
[0009] DE 4322229 which discloses an enzyme containing granule with
a coating comprising an inorganic pigment, an alcohol, an
emulsifier, a pigment dispersant and water.
[0010] JP 61162185 which discloses a process for production of an
enzyme containing granule comprising coating a core with a solution
containing enzyme(s), sodium sulfate and optionally binders and
coating agents.
[0011] Other coating layers apply polymers or even macroscopic
particles to gain improved properties of the granulate such as: WO
97/23606 which discloses an enzyme containing granule comprising an
outer coating of polyvinyl pyrrolidone, PVA or PEG.
[0012] WO 96/38527 which discloses an enzyme containing granulated
substance with a coating comprising water insoluble particles and a
binder.
[0013] U.S. Pat. No. 5,324,649 which discloses an enzyme containing
granule comprising an outer coating of polyvinyl alcohol or a
copolymer.
[0014] WO 93/07263 which discloses an enzyme containing granule
comprising an outer coating of vinyl (co)polymer.
[0015] WO 92/12645 which discloses an enzyme containing T-granulate
coated with high melting fat or wax.
[0016] WO 89/08694 which discloses an enzyme containing granulate
with a coating comprising a mono- or diglyceride of a fatty
acid.
[0017] DD 263790 which discloses a protease containing granule with
a coating of skim milk and/or maltodextrin.
[0018] WO 87/07292 which discloses an enzyme containing granulate
with a coating containing a copolymers of acrylic acid, and/or an
filler and/or a plasticiser.
[0019] EP 193829 and U.S. Pat. No. 4,689,297 which discloses a
process for production of an enzyme containing particle comprising
coating the particle with a macromolecular, film forming water
soluble or water dispersible coating agent.
[0020] JP 58179492 which discloses an enzyme supporting particle
with a coating of modified cellulose.
[0021] WO 89/08695 which discloses an enzyme containing particles
with a coating containing clay.
[0022] Still other prior art disclosures apply non aqueous liquids
as coatings to gain improved properties of the granulate such
as:
[0023] WO 96/16151 which discloses a enzyme containing granule with
a coating of a non-aqueous liquid.
[0024] Some prior art disclosures mention use of agents in a
coating layers providing a special functions upon dissolution of
the granulates such as:
[0025] DE 4344215 which discloses an enzyme containing granule with
a coating containing an inorganic Ag-corrosion inhibitor.
[0026] EP 206417 which discloses an enzyme containing granule with
a coating containing an alkaline buffer salt having a pH of 7-11.
The buffer salt may constitute 50-100% of the coating.
[0027] WO 93/07263 discloses an enzyme containing granule which
contains a scavenger layer, preferably ammonium sulfate.
[0028] EP-415652-A2 as well as U.S. Pat. No. 5,093,021 describes
enzyme granules coated with unsoluble and highly alkaline alkali
metal silicates optionally in combination with alkali metal
carbonates.
[0029] Use of low molecular water soluble compounds in coatings in
moderate amounts have been disclosed within the art of formulating
stabile enzyme compositions usually as filler material. However, it
has not been acknowledged that the amount and hygroscopicity of
such compounds has essential impact on the stability of an enzyme
in an enzyme granule coated with such compounds.
SUMMARY OF THE INVENTION
[0030] We have in our search for enzyme formulation with improved
enzyme stability surprisingly found, that an enzyme containing
granule coated or encapsulated by a continuous layer of a simple,
predominantly water soluble and low cost material with a high
constant humidity significantly increases the storage stability of
the enzyme, especially at high humidity conditions.
[0031] The present invention provides thus in a first aspect an
enzyme containing granule comprising:
[0032] a) an enzyme containing core and
[0033] b) a protective substantially continuous layer or coating
encapsulating the core comprising at least 60% of a water soluble
compound, having a molecular weight below 500 grams per mole, a pH
below 11 and a constant humidity at 20.degree. C. of more than
81%.
[0034] In accordance with the first aspect a second aspect of the
invention is a method for producing said enzyme containing granule
comprising coating said enzyme containing core with said coating
material.
[0035] In accordance with the previous aspects further aspects of
the invention relates to applications of the enzyme containing
granules, e.g. incorporation of the granule in a detergent or an
animal feed composition or a baking composition and a cleaning
method comprising contacting an object with an aqueous solution of
the enzyme containing granule.
DESCRIPTION OF DRAWING
[0036] FIG. 1 shows a cross section of an example of a granule
according to the invention depicting different granule layers.
DETAILED DESCRIPTION OF THE INVENTION
[0037] Definitions
[0038] The term "% RH" is used throughout the text, and in the
context of the invention the term is to be understood as the
relative humidity of air. 100% RH is air saturated with water
moisture at a fixed temperature and % RH thus reflects the percent
moisture saturation of the air.
[0039] The term "constant humidity" (in the context of the
invention sometimes abbreviated as CH) of a compound or substance
is to be understood as the % RH of atmospheric air in equilibrium
with a saturated aqueous solution of said compound in contact with
the solid phase of said compound, all confined within a closed
space at a given temperature. This definition is in accordance with
"Handbook of chemistry and physics" CRC Press, Inc., Cleveland,
USA, 58th edition, p E46, 1977-1978. Accordingly CH.sub.20.degree.
C.=50% for a compound means that air with a 50% humidity will be in
equilibrium with a saturated aqueous solution of the compound at
20.degree. C. Accordingly the term constant humidity is a measure
of the hygroscopic properties of a compound.
[0040] The term "pH" of a compound in the context of the invention
is to be understood as the pH of a 10% w/w aqueous solution of the
said compound.
[0041] The term "water soluble compound" in the context of the
invention is to be understood as a compound for which at least 0.1
grams of the compound may dissolved in 100 g of water at 20.degree.
C., preferably at least 0.5 g per 100 g water, e.g. at least 1 g
per 100 g water.
[0042] The Protective Coating
[0043] Without being bound to this theory we believe that a coating
with a high constant humidity value has at least one important
functional property: The coating inhibits moisture from entering
the enzyme containing core material. The coating is thus a barrier
between the potentially harmful matrix which surrounds the enzyme
containing granules (e.g. a detergent and/or air). For moisture
and/or harmful components carried by the moisture to reach the
enzyme within a granule, the moisture must pass through the coating
(i.e. the moisture must be absorbed on the outside and liberated on
the inside surface of the coating) before coming in contact with
and inactivating the enzyme. Coatings with a high constant humidity
(CH) thus provides a better protection of the enzyme than coatings
with a lower constant humidity, i.e. the high humidity constant
coating will inhibit moisture from entering the granule at a
broader ranger of humidity conditions (% RH) of the surrounding
matrix. We believe that if as an example the % RH is higher than
the CH of the coating material the coating will absorb moisture
from the surrounding matrix and allow the moisture to be
transported inside the granule.
[0044] For coatings allowing absorption of moisture from the
surrounding matrix, the rate of moisture absorption, and thus the
damaging affects on the enzyme is believed to be further increased
if the core material also readily absorbs the incoming moisture,
which is the case for many known core materials. The moisture
absorption process may thus be accelerated by swelling or expansion
of the core materials forming cracks, holes or disruptions in the
coating, thus making moisture access to the enzyme in the core even
easier. Choosing a coating with a constant humidity value higher
than the expected % RH of the surrounding air thus effectively
reduces this process.
[0045] Further a coating with a high constant humidity value is
believed to inhibit entry of microorganisms thus reducing the
possibility of microbial growth within the granule.
[0046] As mentioned vide supra a suitable coating according to the
invention comprises at least 60% w/w of a water soluble compound,
having a molecular weight below 500 grams per mole, a pH below 11
and a constant humidity of more than 81% w/w. Water soluble
compounds having a molecular weight below 500 g/mole are usually
much cheaper and easier to handle with respect to coating processes
than high molecular substances such as polymers. Preferred water
soluble compounds should have a molar weight in the range of 30-500
g/mole preferably 75-400, e.g. 100-300 g/mole.
[0047] Further a water soluble compound benefits release and/or
dissolution of the enzyme upon introduction of the granule in an
aqueous medium (e.g. a cleaning or washing liquor) much more than
insoluble salts such as calcium carbonate and minerals or inorganic
compounds such as kaolin and/or titandioxide. Accordingly a
preferred water soluble compound is an inorganic salt, e.g. salts
of sulfate, sulfite, phosphate, phosphonate, nitrate, chloride or
carbonate or salts of simple organic acids (less than 10 carbon
atoms e.g. 6 or less carbon atoms) such as citrate, malonate or
acetate. Preferred cations in these salt are alkali or earth alkali
metal ions, although the ammonium ion or metal ions of the first
transition series, e.g. Zinc may also be used. Especially alkali-
or earth alkali metal salts of sulfate, sulfite, phosphate,
phosphonate, nitrate, chloride or carbonate or salts of simple
organic acids such as citrate, malonate or acetate are
preferred.
[0048] A suitable solubility should be that at least 0.1 grams of
the salt in 100 g of water at 20.degree. C., preferably at least
0.5 g per 100 g water, e.g. at least 1 g per 100 g water. In a most
preferred embodiment of the invention the solubility of the water
soluble compound is at least 10 grams or at least 20 grams of
compound per 100 gram water at 20.degree. C. A high solubility is
very advantageous as it benefits the coating process as it lowers
the amount of water needed to be evaporated after delivering the
coating on the core. Further it is important that the compound is
dissolved in an aqueous phase before coating, because if a core is
coated by applying the water soluble compound in dry form as a
particulate powder or as a slurry on the core, these particles will
form channels or openings in the coating allowing access of
moisture to the core.
[0049] The water soluble compound should also have a moderate pH in
aqueous solution as extreme pH values of coating solution may
corrode equipment as well as being potentially dangerous to work
with. Thus the water soluble compound may be a slightly alkaline or
slightly acidic compound. Accordingly the pH of the water soluble
compound should be below 11, preferably below 10, e.g. below 9,
below 8 or even below 7, when measured as a 10% w/w aqueous
solution of the water soluble compound. Some soluble salts like
sodium carbonate has a very high pH (above 11) and may not be
suitable as a coating material in the context of this invention.
Also bicarbonate salts may be unsuitable as they although they have
a lower pH tend to form carbon dioxide gas in solution which may
interfere with the coating process.
[0050] Specific examples of suitable water soluble compounds of the
invention are Na.sub.2HPO.sub.4 (CH.sub.20.degree. C.=95%),
Na.sub.3PO.sub.4 (CH.sub.25.degree. C.=92%),
(NH.sub.4)H.sub.2PO.sub.4 (CH.sub.20.degree. C.=93.1%),
KH.sub.2PO.sub.4 (CH.sub.20.degree. C.=92%),
Na.sub.2SO.sub.4(CH.sub.20.degree. C.=93%),
K.sub.2SO.sub.4(CH.sub.25.degree. C.=99%), KHSO.sub.4
(CH.sub.20.degree. C.=86%), ZnSO.sub.4 (CH.sub.20.degree. C.=90%)
and sodium citrate (CH.sub.25.degree. C.=86%). Sodium sulfate and
sodium citrate are the most preferred water soluble compounds as
they are cheap chemicals.
[0051] The coating comprises as said at least 60% w/w, e.g. 65% w/w
or 70% w/w of the water soluble compound, which preferably may be
at least 75% w/w, e.g. at least 80% w/w, at least 85% w/w, e.g. at
least 90% w/w or at least 95% w/w. The coating may even essentially
consist of the water soluble compound. With due respect for
maintaining a desired suitable constant humidity value for the
total coating material minor amounts of other feasible compounds
may be present in the coating such as conventional coating
materials. Examples of are, inter alia, described in the paragraph
"additional coatings" given below. Other examples of conventional
coating materials may be found in references such as U.S. Pat. No.
4,106,991, EP 170360, EP 304332, EP 304331, EP 458849, EP 458845,
WO 97/39116, WO 92/12645A, WO 89/08695, WO 89/08694, WO 87/07292,
WO 91/06638, WO 92/13030, WO 93/07260, WO 93/07263, WO 96/38527, WO
96/16151, WO 97/23606, U.S. Pat. No. 5,324,649, U.S. Pat. No.
4,689,297, EP 206417, EP 193829, DE 4344215, DE 4322229 A, DD
263790, JP 61162185 A and/or JP 58179492.
[0052] In a particular embodiment the coating may comprise minor
amounts of a protective agent capable of reacting with a component
capable of inactivating (being hostile to) the enzyme entering the
granule from the surrounding matrix, i.e. before the component come
into contact and inactivate the enzyme. The protective agent may
thus e.g. be capable of neutralizing, reducing or otherwise
reacting with the hostile component rendering it harmless to the
enzyme. Typical components capable of inactivating the enzyme are
oxidants such as perborates, percarbonates, organic peracids and
the like.
[0053] Protective agents may fall into several categories: alkaline
or neutral materials, reducing agents, antioxidants and/or salts of
first transition series metal ions. Each of these may be used in
conjunction with other protective agents of the same or different
categories. Examples of alkaline protective agents are alkali metal
silicates, -carbonates or bicarbonates which provide a chemical
scavenging effect by actively neutralizing e.g. oxidants. Examples
of reducing protective agents are salts of sulfite, thiosulfite or
thiosulfate, while examples of antioxidants are methionine,
butylated hydroxytoluene (BHT) or butylated hydroxyanisol (BHA).
Most preferred agents are salts of thiosulfates, e.g. sodium
thiosulfate. The amounts of protective agent in the coating may be
1-40% w/w of the coating, preferably 5-30%, e.g. 10-20%.
[0054] In accordance with the concept of the invention the coating
does not contain any enzyme, as the purpose of the coating is to
protect enzymes encapsulated by the coating.
[0055] The protective effect of the coating depends on the
thickness of the coating and the amount of coating relative to the
rest of the enzyme containing granule. An increased coating
thickness provides a better protection of the enzyme, but at the
same time result in increased manufacturing costs as well as a risk
of poorer granule properties with respect to enzyme solubility rate
upon introduction of the enzyme containing granule in an aqueous
medium. For effective protection the coating thickness also must be
adjusted to the size of the core, e.g. to obtain a desired size of
the finishes granule. Depending on the size of the core material
the coating may be applied in 1-75% w/w of the weight of the coated
granule to obtain a desired size of the coated granule. For small
sizes of core material the coating may be applied in 50-75% w/w or
15-50% of the coated granule. Usually coatings constituting 2-20%
w/w, preferably 3-10% w/w, e.g. 6% of the coated granule is however
preferred.
[0056] The coating should encapsulate the enzyme containing core by
forming a substantially continuous layer. The layer or coating is
preferably homogenous in thickness and by substantially continuous
is meant that the core surface should have few or none uncoated
areas.
[0057] The Core
[0058] The core contains the enzyme(s). Besides of the enzyme(s)
the core may be constructed in any way or of any material which
provides the desired functional properties of the core material,
e.g. the core may consist of materials which allows readily release
of the enzyme(s) upon introduction to an aqueous medium. In one
preferred embodiment the core is constructed of a particulate
carrier (I) with the enzyme absorbed and/or an enzyme containing
layer (II) applied on the carrier surface, optionally comprising an
enzyme protecting reducing agent. There may even be additional
coating within the core material providing desired functional
properties of the core material. Another preferred core is the so
called T-granulate wherein enzyme and granulation material is mixed
to form granules incorporating the enzyme distributed throughout
the core such as described in U.S. Pat. No. 4,106,991 e.g. Example
1. Any conventional methods and non-enzyme materials may be used to
prepare the core. Examples of known conventional cores and
materials is, inter alia, described in, U.S. Pat. No. 4,106,991 (in
particular), EP 170360, EP 304332, EP 304331, EP 458849, EP 458845,
WO 97/39116, WO 92/12645, WO 89/08695, WO 89/08694, WO 87/07292, WO
91/06638, WO 92/13030, WO 93/07260, WO 93/07263, WO 96/38527, WO
96/16151, WO 97/23606, U.S. Pat. No. 5,324,649, U.S. Pat. No.
4,689,297, EP 206417, EP 193829, DE 4344215, DE 4322229 A, DD
263790, JP 61162185 A, JP 58179492.
[0059] As a particularly preferred embodiment of the invention the
core may be prepared by applying an enzyme layer onto a "placebo"
carrier (enzyme-free carrier) according to the methodology
described in U.S. Pat. No. 4,106,991. Optionally additional enzyme
may be absorbed into the surface of the carrier.
[0060] In a particular embodiment of the invention the enzyme
containing core may also comprise a protective agent as described
for the coating, vide supra, preferably mixed with the enzyme in
suitable amounts such as 0.1-1% w/w of the coated granule,
preferably 0.1-0.5% w/w, e.g. 0.33% w/w.
[0061] As described, supra, the core may through the coating absorb
moisture from the surrounding environment, a process which may
cause the core to swell resulting in crack formation in the coating
and further moisture absorbance. The core may even in at high
relative humidity dissolve and become fluid. Accordingly in order
to provide further stabilization of the enzyme the core should
preferably be a non absorbing core, i.e. the should only be able of
absorbing less moisture than 20% w/w of it own dry weight,
preferably less than 10% w/w, e.g. less than 8% w/w or less than 5%
w/w, measured at 75% RH at 20.degree. C.
[0062] Enzymes
[0063] The enzyme in the context of the present invention may be
any enzyme or combination of different enzymes, which benefits from
being granulated and thus be protected against a hostile
environment in order to be applicable for a specific use.
Accordingly, when reference is made to "an enzyme" this will in
general be understood include combinations of one or more
enzymes.
[0064] It is to be understood that enzyme variants (produced, for
example, by recombinant techniques) are included within the meaning
of the term "enzyme". Examples of such enzyme variants are
disclosed, e.g., in EP 251,446 (Genencor), WO 91/00345 (Novo
Nordisk A/S), EP 525,610 (Solvay) and WO 94/02618 (Gist-Brocades
NV).
[0065] The enzyme classification employed in the present
specification with claims is in accordance with Recommendations
(1992) of the Nomenclature Committee of the International Union of
Biochemistry and Molecular Biology, Academic Press, Inc., 1992.
[0066] Accordingly the types of enzymes which may appropriately be
incorporated in granules of the invention include oxidoreductases
(EC 1.-.-.-), transferases (EC 2.-.-.-), hydrolases (EC 3.-.-.-),
lyases (EC 4.-.-.-), isomerases (EC 5.-.-.-) and ligases (EC
6.-.-.-).
[0067] Preferred oxidoreductases in the context of the invention
are peroxidases (EC 1.11.1), laccases (EC 1.10.3.2) and glucose
oxidases (EC 1.1.3.4)], while preferred transferases are
transferases in any of the following sub-classes:
[0068] a) Transferases transferring one-carbon groups (EC 2.1);
[0069] b) transferases transferring aldehyde or ketone residues (EC
2.2); acyltransferases (EC 2.3);
[0070] c) glycosyltransferases (EC 2.4);
[0071] d) transferases transferring alkyl or aryl groups, other
that methyl groups (EC 2.5); and
[0072] e) transferases transferring nitrogenous groups (EC
2.6).
[0073] A most preferred type of transferase in the context of the
invention is a transglutaminase (protein-glutamine
.gamma.-glutamyltransferase; EC 2.3.2.13). Further examples of
suitable transglutaminases are described in WO 96/06931 (Novo
Nordisk A/S).
[0074] Preferred hydrolases in the context of the invention are:
Carboxylic ester hydrolases (EC 3.1.1.-) such as lipases (EC
3.1.1.3); phytases (EC 3.1.3.-), e.g. 3-phytases (EC 3.1.3.8) and
6-phytases (EC 3.1.3.26); glycosidases (EC 3.2, which fall within a
group denoted herein as "carbohydrases"), such as .alpha.-amylases
(EC 3.2.1.1); peptidases (EC 3.4, also known as proteases); and
other carbonyl hydrolases].
[0075] In the present context, the term "carbohydrase" is used to
denote not only enzymes capable of breaking down carbohydrate
chains (e.g. starches) of especially five- and six-membered ring
structures (i.e. glycosidases, EC 3.2), but also enzymes capable of
isomerizing carbohydrates, e.g. six-membered ring structures such
as D-glucose to five-membered ring structures such as
D-fructose.
[0076] Carbohydrases of relevance include the following (EC numbers
in parentheses): .alpha.-amylases (3.2.1.1), .beta.-amylases
(3.2.1.2), glucan 1,4-.alpha.-glucosidases (3.2.1.3), cellulases
(3.2.1.4), endo-1,3(4)-.beta.-glucanases (3.2.1.6),
endo-1,4-.beta.-xylanases (3.2.1.8), dextranases (3.2.1.11),
chitinases (3.2.1.14), polygalacturonases (3.2.1.15), lysozymes
(3.2.1.17), .beta.-glucosidases (3.2.1.21), .alpha.-galactosidases
(3.2.1.22), .beta.-galactosidases (3.2.1.23),
amylo-1,6-glucosidases (3.2.1.33), xylan 1,4-.beta.-xylosidases
(3.2.1.37), glucan endo-1,3-.beta.-D-glucosidases (3.2.1.39),
.alpha.-dextrin endo-1,6-.alpha.-glucosidases (3.2.1.41), sucrose
.alpha.-glucosidases (3.2.1.48), glucan endo-1,3-.alpha.-glucosid-
ases (3.2.1.59), glucan 1,4-.beta.-glucosidases (3.2.1.74), glucan
endo-1,6-.beta.-glucosidases (3.2.1.75), arabinan
endo-1,5-.alpha.-L-arab- inosidases (3.2.1.99), lactases
(3.2.1.108), chitosanases (3.2.1.132) and xylose isomerases
(5.3.1.5).
[0077] Examples of commercially available oxidoreductases (EC
1.-.-.-) include Gluzyme.TM. (enzyme available from Novo Nordisk
A/S). Further oxidoreductases are available from other
suppliers.
[0078] Examples of commercially available proteases (peptidases)
include Esperase.TM., Alcalase.TM., Neutrase.TM., Durazym.TM.,
Savinase.TM., Kannase, Pyrase.TM., Pancreatic Trypsin NOVO (PTN),
Bio-Feed.TM. Pro and Clear-Lens.TM. Pro (all available from Novo
Nordisk A/S, Bagsvaerd, Denmark).
[0079] Other commercially available proteases include Maxatase.TM.,
Maxacal.TM., Maxapem.TM., Opticlean.TM. and Purafect.TM. (available
from Genencor International Inc. or Gist-Brocades).
[0080] Examples of commercially available lipases include
Lipolase.TM., Lipolase.TM. Ultra, LipoPrime, Lipozyme.TM.,
Palatase.TM., Novozym.TM. 435 and Lecitase.TM. (all available from
Novo Nordisk A/S).
[0081] Other commercially available lipases include Lumafast.TM.
(Pseudomonas mendocina lipase from Genencor International Inc.);
Lipomax.TM. (Ps. pseudoalcaligenes lipase from
Gist-brocades/Genencor Int. Inc.; and Bacillus sp. lipase from
Solvay enzymes. Further lipases are available from other
suppliers.
[0082] Examples of commercially available carbohydrases include
Alpha-Gal.TM., Bio-Feed.TM. Alpha, Bio-Feed.TM. Beta, Bio-Feed.TM.
Plus, Bio-Feed.TM. Plus, Novozyme.TM. 188, Celluclast.TM.,
Cellusoft.TM., Ceremyl.TM., Citrozym.TM., Denimax.TM., Dezyme.TM.,
Dextrozyme.TM., Finizym.TM., Fungamyl.TM., Gamanase.TM.,
Glucanex.TM., Lactozym.TM., Maltogenase.TM., Pentopan.TM.,
Pectinex.TM., Promozyme.TM., Pulpzyme.TM., Novamyl.TM.,
Termamyl.TM., AMG.TM. (Amyloglucosidase Novo), Maltogenase.TM.,
SweetZyme.TM. and Aquazym.TM. (all available from Novo Nordisk
A/S). Further carbohydrases are available from other suppliers.
[0083] The amount of enzyme to be incorporated in a granule of the
invention will depend on the intended use of the granulate. For
many applications, the enzyme content will be as high as possible
or practicable.
[0084] The content of enzyme (calculated as pure enzyme protein) in
a granule of the invention will typically be in the range of from
about 0.5% to 20% by weight of the core.
[0085] When, for example, a protease (or peptidase) is incorporated
in granules according to the invention, the enzyme activity
(proteolytic activity) of the finished granules will typically be
in the range of 1-50 KiloNovoProteaseUnits per gram. Likewise, in
the case of, for example, .alpha.-amylases, an activity of 10-500
KiloNovoUnits per gram will be typical, whilst for lipases, an
activity in the range of 50-400 KiloLipolaseUnits per gram will
normally be suitable. All units are known to the art.
[0086] Additional Coatings
[0087] The granules of the present invention may comprise one, two
or more additional coating layers on the inside or outside surface
of the protective coating according to the invention.
[0088] The additional coating layers may perform any of a number of
functions in the granule, depending on the intended use of the
granule. Thus, for example, an additional coating may achieve one
or more of the following effects:
[0089] (i) further reduction of the dust-formation tendency of a
granule without the additional coating according to the
invention;
[0090] (ii) further protection of enzyme(s) in the granule against
oxidation by bleaching substances/systems (e.g. perborates,
percarbonates, organic peracids and the like);
[0091] (iii) dissolution at a desired rate upon introduction of the
granule into a liquid medium (such as an aqueous medium);
[0092] (iv) provide a better physical strength of the granule.
[0093] Any additional conventional coating(s) of desired properties
may be applied and examples of conventional coating materials and
coating methods is, inter alia, described in U.S. Pat. No.
4,106,991, EP 170360, EP 304332, EP 304331, EP 458849, EP 458845,
WO 97/39116, WO 92/12645, WO 89/08695, WO 89/08694, WO 87/07292, WO
91/06638, WO 92/13030, WO 93/07260, WO 93/07263, WO 96/38527, WO
96/16151, WO 97/23606, U.S. Pat. No. 5,324,649, U.S. Pat. No.
4,689,297, EP 206417, EP 193829, DE 4344215, DE 4322229 A, DD
263790, JP 61162185 A, JP 58179492.
[0094] In appropriate embodiments of granules according to the
present invention, the additional coating layer may be composed as
described in U.S. Pat. No. 4,106,991 [e.g. with a waxy material
such as polyethylene glycol (PEG), optionally followed by powdering
with a whitener such as titanium dioxide].
[0095] Additional coating layers may further comprise one or more
of the following: anti-oxidants, chlorine scavengers, plasticizers,
pigments, lubricants (such as surfactants or antistatic agents)
additional enzymes and fragrances.
[0096] Plasticizers useful in coating layers in the context of the
present invention include, for example: polyols such as sugars,
sugar alcohols, or polyethylene glycols (PEGs) having a molecular
weight less than 1000; urea, phthalate esters such as dibutyl or
dimethyl phthalate; and water.
[0097] Suitable pigments include, but are not limited to, finely
divided whiteners, such as titanium dioxide or kaolin, coloured
pigments, water soluble colorants, as well as combinations of one
or more pigments and water soluble colorants.
[0098] As used in the present context, the term "lubricant" refers
to any agent which reduces surface friction, lubricates the surface
of the granule, decreases tendency to build-up of static
electricity, and/or reduces friability of the granules. Lubricants
can also play a related role in improving the coating process, by
reducing the tackiness of binders in the coating. Thus, lubricants
can serve as anti-agglomeration agents and wetting agents.
[0099] Examples of suitable lubricants are polyethylene glycols
(PEGs) and ethoxylated fatty alcohols.
[0100] In an especially preferred embodiment of the invention, only
a lubricant is applied as additional coating. The composition of 1)
an enzyme containing core, 2) a coating and 3) and an additional
lubricant coating has shown particularly good properties with
respect to enzyme stability.
[0101] Coating Methods
[0102] The invention also relates to a method for
manufacture/production of the enzyme containing granule described
herein. Conventional coating methods may be used to apply the
coating according to the invention as described by the references
stated in the previous section (above).
[0103] A method for production of the enzyme containing granule may
comprise the following steps:
[0104] a) mixing an enzyme containing core material with a liquid
medium comprising the water soluble compound of the invention
and,
[0105] b) removing volatile components of the liquid medium from
the mixture, so as to deposit the nonvolatile components of the
liquid medium as solid coating layer on the core material.
[0106] In a preferred embodiment of the invention the enzyme
containing granule is produced by a fluid bed process
comprising:
[0107] a) fluidising an enzyme containing core material in a fluid
bed apparatus,
[0108] b) introducing a liquid medium comprising the water soluble
compound of the invention by atomization of the liquid medium into
the fluid bed, so as to deposit nonvolatile components of the
liquid medium as a solid coating layer on the core material
and,
[0109] c) removing volatile components of the liquid medium from
the coated core material.
[0110] In a further preferred embodiment of the invention the core
material is prepared by a method comprising:
[0111] a) Preparing a particulate carrier material,
[0112] b) introducing a liquid medium comprising an enzyme by
atomization of the liquid medium into the fluid bed, so as to
deposit nonvolatile components including the enzyme of the liquid
medium as an enzyme containing layer on the carrier, and
[0113] c) removing volatile components of the liquid medium from
the core material.
[0114] The particulate carrier material may in a preferred
embodiment comprise a binder (such as Glucidex.TM. 21D, from
Roquette Freres), a fibre material (such as cellulose fibres) and a
filler (such as finely ground sodium sulfate and/or kaolin). The
particulate carrier may as well be prepared/granulated and dried as
described in Example 1 in U.S. Pat. No. 4,106,991. Following
granulation the dry particulate carrier may suitably be sieved, and
fractionated after size to obtain a uniform carrier size. Preferred
carrier sizes measured as the diameter of the carrier are between
0.1-2 mm, e.g. 0.3-1.0 mm.
[0115] As a further preferred embodiment additional enzyme may be
absorbed on the particulate carrier prior to applying the enzyme
layer (II) This absorption may be achieved by:
[0116] a) absorbing the enzyme(s) into the surface of the carrier
by contacting the particulate carrier with a liquid comprising the
enzyme in a mixer,
[0117] b) mixing the composition by means of mixing blades, and
[0118] c) drying the enzyme loaded carrier by fluidising it in a
fluid bed apparatus,
[0119] Conventional mixing equipment can satisfactorily be used to
mix the particulate carrier with the enzyme-containing liquid
medium. The mixing equipment can be a batch mixer or a continuous
mixer, such as a convective mixer [see, e.g., Harnby et al., Mixing
in the Process Industries, pp. 39-53 (ISBN 0-408-11574-2)].
Non-convective mixing equipment, e.g. rotating drum mixers or
so-called pan-granulators, may also be employed.
[0120] Drying of enzyme-loaded particulate carrier, application of
the enzyme containing layer (II) the coating (III) and any
additional coatings may be performed in any type of fluidising
equipment (such as in a fluid-bed apparatus or other form of
fluidizing equipment, such as a Huttlin-type fluidizer). For a
description of suitable fluid-bed equipment, see, e.g., Harnby et
al. , Mixing in the Process Industries, pp. 54-77 (ISBN
0-408-11574-2).
[0121] Applications of the Enzyme Containing Granule
[0122] The enzyme containing granule according to the invention is
useful where ever enzymes are to be stored alone or to be
incorporated in another dry product, and an improved enzyme
stability is needed to enable good storage properties (improved
shelf life) of the granule. Especially at relatively humid
conditions, i.e. under an atmosphere with a % RH of more than 55%
RH, preferably more than 60% RH, e.g. more than 70% RH. Especially
at conditions with more than 75% RH, more than 85% RH or more than
95% RH the invention is useful. The granule is also particularly
useful in dry products comprising oxidative compounds such as
peroxides or superoxides, e.g. bleach (e.g. perborates or
percarbonates) or other reactive components, which in case of
contact with the enzyme is able of inactivating the enzyme. Thus
the invention provides a detergent composition comprising the
granule of the invention. The enzyme containing granule is further
useful for cleaning an object (e.g. textile of cotton or other
natural or synthetic fabrics) by contacting the object with an
aqueous solution of the enzyme containing granule. Finally the
enzyme containing granule is useful in products such as animal
feed/fodder or bakers flour.
[0123] Detergent Disclosure
[0124] A detergent composition of the invention comprises the
enzyme containing granule of the invention and a surfactant.
Additionally, it may optionally comprise a builder, another enzyme,
a suds suppresser, a softening agent, a dye-transfer inhibiting
agent and other components conventionally used in detergents such
as soil-suspending agents, soil-releasing agents, optical
brighteners, abrasives, bactericides, tarnish inhibitors, coloring
agents, and/or encapsulated or non-encapsulated perfumes.
[0125] The detergent composition according to the invention can be
in bars or granular forms. The pH (measured in aqueous solution at
use concentration) will usually be neutral or alkaline, e.g. in the
range of 7-11.
[0126] An enzyme contained in the granule of the invention
incorporated in the detergent composition, is normally incorporated
in the detergent composition at a level from 0.00001% to 2% of
enzyme protein by weight of the composition, preferably at a level
from 0.0001% to 1% of enzyme protein by weight of the composition,
more preferably at a level from 0.001% to 0.5% of enzyme protein by
weight of the composition, even more preferably at a level from
0.01% to 0.2% of enzyme protein by weight of the composition.
[0127] Surfactant System
[0128] The surfactant system may comprise nonionic, anionic,
cationic, ampholytic, and/or zwitterionic surfactants. The
surfactant system preferably consists of anionic surfactant or a
combination of anionic and nonionic surfactant, e.g. 50-100% of
anionic surfactant and 0-50% nonionic. The laundry detergent
compositions may also contain cationic, ampholytic, zwitterionic,
and semi-polar surfactants, as well as the nonionic and/or anionic
surfactants other than those already described herein.
[0129] The surfactant is typically present at a level from 0.1% to
60% by weight. Some examples of surfactants are described
below.
[0130] a) Nonionic Surfactant:
[0131] The surfactant may comprise polyalkylene oxide (e.g.
polyethylene oxide) condensates of alkyl phenols. The alkyl group
may contain from about 6 to about 14 carbon atoms, in a straight
chain or branched-chain. The ethylene oxide may be present in an
amount equal to from about 2 to about 25 moles per mole of alkyl
phenol.
[0132] The surfactant may also comprise condensation products of
primary and secondary aliphatic alcohols with about 1 to about 25
moles of ethylene oxide. The alkyl chain of the aliphatic alcohol
can either be straight or branched, and generally contains from
about 8 to about 22 carbon atoms.
[0133] Further, the nonionic surfactant may comprise polyethylene
oxide condensates of alkyl phenols, condensation products of
primary and secondary aliphatic alcohols with from about 1 to about
25 moles of ethylene oxide, alkylpolysaccharides, and mixtures
hereof. Most preferred are C8-C14 alkyl phenol ethoxylates having
from 3 to 15 ethoxy groups and C8-C18 alcohol ethoxylates
(preferably C10 avg.) having from 2 to 10 ethoxy groups, and
mixtures thereof.
[0134] b) Anionic Surfactants:
[0135] Suitable anionic surfactants include the alkyl sulfate
surfactants which are water soluble salts or acids of the formula
ROSO3M wherein R preferably is a C10-C24 hydrocarbyl, preferably an
alkyl or hydroxyalkyl having a C10-C20 alkyl component, more
preferably a C12-C18 alkyl or hydroxyalkyl, and M is H or a cation,
e.g., an alkali metal cation (e.g. sodium, potassium, lithium), or
ammonium or substituted ammonium.
[0136] Other anionic surfactants include salts (including, for
example, sodium, potassium, ammonium, and substituted ammonium
salts such as mono- di- and triethanolamine salts) of soap, C8-C22
primary or secondary alkanesulfonates, C8-C24 olefinsulfonates,
sulfonated polycarboxylic acids prepared by sulfonation of the
pyrolyzed product of alkaline earth metal citrates.
[0137] Alkylbenzene sulfonates are suitacble, especially linear
(straight-chain) alkyl benzene sulfonates (LAS) wherein the alkyl
group preferably contains from 10 to 18 carbon atoms. The laundry
detergent compositions typically comprise from about 1% to about
40%, preferably from about 3% to about 20% by weight of such
anionic surfactants.
[0138] Builder System
[0139] The compositions according to the present invention may
further comprise a builder system. Any conventional builder system
is suitable for use herein including aluminosilicate materials,
silicates, polycarboxylates and fatty acids, materials such as
ethylenediamine tetraacetate (EDTA), metal ion sequestrants such as
aminopolyphosphonates. Phosphate builders can also be used
herein.
[0140] Suitable builders can be an inorganic ion exchange material,
commonly an inorganic hydrated aluminosilicate material, more
particularly a hydrated synthetic zeolite such as hydrated zeolite
A, X, B, HS or MAP. Detergency builder salts are normally included
in amounts of from 5% to 80% by weight of the composition.
Preferred levels of builder for liquid detergents are from 5% to
30%.
[0141] Bleaching Agents
[0142] The detergent composition may also comprise a bleaching
agents, e.g. an oxygen bleach or a halogen bleach. The oxygen
bleach may be a hydrogen peroxide releasing agent such as a
perborate (e.g. PB1 or PB4) or a percarbonate, or it may e.g. be a
percarboxylic acid. The particle size of a bleaching agent may be
400-800 microns. When present, oxygen bleaching compounds will
typically be present at levels of from about 1% to about 25%.
[0143] The hydrogen peroxide releasing agent can be used in
combination with bleach activators such as
tetra-acetylethylenediamine (TAED), nonanoyloxybenzene-sulfonate
(NOBS), 3,5-trimethyl-hexsanoloxybenzene-sul- fonate (ISONOBS) or
pentaacetylglucose (PAG).
[0144] The halogen bleach may be, e.g. a hypohalite bleaching
agent, for example, trichloro-isocyanuric acid and the sodium and
to potassium salt of dichloroisocyanurates and N-chloro and N-bromo
alkane sulfonamides. Such materials are normally added at 0.5-10%
by weight of the finished product, preferably 1-5% by weight.
[0145] Granular detergent compositions according to the present
invention can also be in "compact form", i.e. they may have a
relatively higher density than conventional granular detergents,
i.e. form 550 to 950 g/l.
[0146] The compositions of the invention may for example, be
formulated as hand and machine laundry detergent compositions
including laundry additive compositions and compositions suitable
for use in the pretreatment of stained fabrics, rinse added fabric
softener compositions, and compositions for use in general
household hard surface cleaning operations and dishwashing
operations.
[0147] More specifically, the enzyme containing granules of the
invention may be incorporated in the detergent compositions
described in WO 97/04079, WO 97/07202, WO 97/41212, and PCT/DK
97/00345.
MATERIALS AND METHODS
[0148] Preparation of Carrier Material
[0149] Portions of carrier material was prepared by spraying 3.0 kg
of fibrous cellulose (Arbocel.TM. FTC200), 3.0 kg of kaolin
(Speswhite.TM., English China Clay) and 20.5 kg of finely ground
sodium sulfate with 9.5 kg of a 21% w/w aqueous solution of
carbohydrate binder (Glucidex.TM. 21D, from Roquette Freres). This
mixtures was granulated and dried as described in Example 1 in U.S.
Pat. No. 4,106,991. The dry granulated carrier material was sieved,
and the fraction between 0.3 and 1.0 mm was separated and used for
further processing.
[0150] Absorbing Enzyme on the Carrier Material
[0151] The granulated and fractionated carrier material was
transferred to a Lodiger mixer equipped with a multiple chopper
head, and sprayed with a liquid Savinase.TM. enzyme concentrate
solution approximately 33 KNPU/g containing 0.4% w/w g of a
nonionic surfactant (Softanol 50). The enzyme containing solution
was sprayet onto the carrier at dosage of 0.15 kg solution per kg
carrier material. The nonionic surfactant was added to the enzyme
concentrate to enhance the absorption of the enzyme onto the
carrier. The Savinase.TM. concentrate was applied using a pressure
nozzle submerged in the carrier and spraying directly into the
chopper. The carrier and the Savinase.TM. concentrate was mixed by
means of mixing blades and the mixing blades and the chopper were
operated continuously during the spraying. The resulting product
was transferred to a Glatt WSG 15 fluid-bed apparatus (Glatt,
Germany) with an air-inlet temperature of 62.degree. C., and dried
for 30 minutes, or until the product temperature exceeded
50.degree. C., and then sieved on a 1.2 mm mesh screen, leaving
only 0.8% w/w of residual, oversized particles on the sieve.
[0152] Measurement of Enzyme Stability
[0153] For each type of enzyme containing granule samples of the
granules are mixed with a commercial detergent. One or of these
samples are immediately sealed in glass jars and stored below
-18.degree. C. These samples are reference samples and the enzyme
herein is by definition 100% stabile. Other samples are placed in
climate controlled cupboards and stored for a preset period of time
in open glass jars at different temperatures and humidities (% RH).
When a preset storage period is finished samples are removed from
the "climate static" conditions, and the glass jars immediately
sealed and cooled to below -18.degree. C. to stop any progressing
inactivation of the enzyme. When all samples has been stored for
the preset period of time, all samples including the reference
samples are analyzed the day in an appropriate enzyme activity
assay and the test results of the samples are calculated as percent
of the reference sample test results.
[0154] Enzyme Assay
[0155] The enzyme assay used herein is a protease activity assay,
and the unit for protease activity herein is Kilo Novo Protease
Units per gram of sample (KNPU/g). The activity is determined
relatively to an enzyme standard (Savinase.TM.) of known activity.
The enzyme standard is standardized by measuring for a given amount
of enzyme the formation rate (.mu.mol/minute) of free amino groups
liberated from digestion of di-methyl-casein (DMC) in solution by
the enzyme. The formation rate is monitored by recording the linear
development of absorbance at 420 nm of the simultaneous reaction
between the formed free amino groups and added
2,4,6-tri-nitro-benzene-sulfonic acid (TNBS). The digestion of DMC
and the color reaction is carried out at 50.degree. C. in a pH 8.3
boric acid buffer with a 9 min. reaction time followed by a 3 min.
measuring time. A folder AF 220/1 is available upon request to Novo
Nordisk A/S, Denmark, which folder is hereby included by
reference.
[0156] For samples of the enzyme containing detergents and the
standardized enzyme standard a modified assay was used, wherein the
reaction was carried out at 40.degree. C. in a pH 8.3 boric acid
buffer containing 3.1 g/L boric acid (Merck), 11.18 g/L potassium
chloride (Merck), 1.5 mL/L 15% BriJ 35 (Merck) and 20 g/L sodium
sulfite bleach scavenger.
[0157] For measuring Endolase activity (a cellulase) any
conventional endo cellulase viscosity reduction method may be used
(such as the method described in B1087a-GB available upon request
from Novo Nordisk A/S--Denmark). Endolase digests CMC
(carboxy-methyl-cellulose) in solution thereby reducing the
viscosity of the solution and the viscosity reduction is related to
the activity of the endolase.
[0158] For measuring alfa-amylase activity (e.g. Natalase.RTM.) any
conventional alfa-amylase assay may be used (such as the method
described in AF318/1-GB available upon request from Novo Nordisk
A/S--Denmark). Alfa-amylases cleaves 1-6 alfa bonds between
adjacent glucose units. By using e.g. a
2-chlor-4-nitrophenyl-b-D-maltoheptaosid substrate and alfa- and
beta-glucosidase enzymes the substrae may be completely digested
into monosaccharides and 2-chlor-4-nitrophenol, which form a
detectable colour. Kits for performing these assays are
commercially available.
[0159] Measurement of Constant Humidity Values for Coating
Compounds
[0160] The constant humidity of water soluble compound coating
materials were measured by preparing a saturated aqueous solution
with excess of solid phase of the compound in an open beaker. After
equilibrium has been reached and no more of the compound dissolves
in the liquid phase, and solid particles of the compound are still
visible, the beaker is placed in a sealed thermostated humidity
measuring device (e.g. a Novasina apparatus), which measures the %
RH at the chosen temperature of the atmosphere over the saturated
solution.
EXAMPLES
[0161] The invention disclosed herein is illustrated by the
non-limiting examples given below
Example 1
[0162] A sample of the dry enzyme-loaded granular carrier was
transferred to a Glatt WSG5 conventional fluid bed apparatus. Using
a conventional top spray coating technique with an air inlet
temperature of 70.degree. C., air outlet temperature of 42.degree.
C. and with air volume of 600 m3/h the following steps were carried
out in sequence:
[0163] a) an enzyme containing layer was applied onto the carrier
by spraying an enzyme containing aqueous solution containing
Savinase.TM., PVP/VA co-polymer (Luviscol VA64) and titaniumdioxide
(TiO.sub.2) onto the carrier at a spraying rate for the enzyme
solution of 100 g/min. Approximately 210 g Savinase.TM. concentrate
(30 KNPU/g), 2.34 g of PVP/VA co-polymer (Luviscol VA64) and 2.5 g
of titaniumdioxide (TiO.sub.2) were applied per kg carrier,
[0164] b) a 80 g sodium citrate per kg carrier coating layer of was
applied by spraying a 37% w/w aqueous solution of sodium citrate
onto the product of a), at a spraying rate for the coating solution
of 100 g/min,
[0165] c) an additional coating layer of 50 g titaniumdioxide
(Kronos 2044), 50 g kaolin (ECC Supreme), 55 g Glascol LS27 (46%
suspension from Allied Colloids Ltd.--GB), 30 g PEG 4000 and 1.67 g
Softanol 50 per kg carrier was applied by spraying an aqueous
solution of these components onto the product of b), at a spraying
rate for the coating solution of 100 g/min, and
[0166] d) a final coating was applied by spraying a solution of 7.5
g PEG 4000 per kg carrier onto the product of c), at a spraying
rate for the coating solution of 100 g/min.
[0167] The finished enzyme containing granule was dried for 5
minutes and then cooled to 30.degree. C., where after it was
removed from the fluid bed and sifted between 300 and 1200
.mu.m.
[0168] As a reference two different types of granules were
prepared. The first type of granule was prepared by repeating steps
a)-d) with the exception that the coating described in b) was
replaced by a 80 g ammonium sulfate per kg carrier coating layer by
spraying a 37% w/w aqueous solution of ammonium sulfate onto the
product of a), at a spraying rate for the coating solution of 100
g/min. The second type of granule was also prepared by repeating
steps a)-d), but with the exception that the coating described in
b) was replaced by a 80 g sodium formiate per kg carrier coating
layer by spraying a 37% w/w aqueous solution of sodium formiate
onto the product of a), at a spraying rate for the coating solution
of 100 g/min.
Example 2
[0169] The storage stability of the enzyme containing granulates
from Example 1 were tested in one powder detergent (A) containing
perborate bleach and TAED and one powder detergent (B) containing
percarbonate bleach. 100 mg samples of the enzyme containing
granulates and 10.0 g samples of the detergents were mixed to
constitute test samples. The samples were incubated in open jars at
air conditions 35.degree. C. and 55% RH in a thermostated and
humidity controlled cupboard. Identical samples were taken out from
the cupboard after 2 and 4 weeks and analyzed for enzyme
(Savinase.TM.) activity together with identical reference samples,
which had been stored in sealed jars below -18.degree. C. The
results are given in Example 1, wherein the enzyme activities of
the samples are given as a percentage of the corresponding
reference samples.
1 TABLE 1 Residual activity Residual activity Coating in Detergent
A in Detergent B compound 2 weeks 4 weeks 2 weeks 4 weeks Invention
Sodium 76% 51% 82% 69% granule citrate Reference Ammonium 56% 39%
70% 59% granule sulfate Reference Sodium 35% 21% 32% 21% granule
formiate
[0170] Constant humidities at 25.degree. C.: Ammonium sulfate=79;
sodium formiate=56 and sodium citrate=86. The results show that the
coating with highest constant humidity provides the best storage
stability of the enzyme.
Example 3
[0171] The hygroscopicity of the samples from Example 1 were tested
including a reference sample which was identical to the granulates
of Examples 1 to 3 except that it did not contain any salt coating
of ammonium sulfate, sodium citrate or sodium formiate. All samples
were incubated for 1 week in open jars at 3 different conditions:
55% RH and 35.degree. C., 60% RH and 30.degree. C. and 74% RH and
37.degree. C. Subsequently the water absorption of the samples were
determined by weighing the samples before and after the incubation.
Further after the incubation the samples were examined under a
microscope. The results are given in Table 2, which shows the water
absorption of the samples at different humidities. These results
clearly shows big difference in how much water is absorbed in
identical granules with different coatings, indicating that sodium
citrate has the lowest water absorbing properties (the highest
constant humidity value). For comparison a sample of a granulate
with-a different core material and a coating of ammonium sulfate is
included (Purafect G). This result shows that if the coating allows
moisture to pass, the core may absorb huge amounts of water. Table
2 further lists the results of the microscope examination, which
showed that the coating of some of the samples were damaged (cracks
had developed) which was most clearly observed when the water
absorption of the coating (absorption of the sample minus the
absorption of the reference) had reached a level of approximately
50% of the salt layer weight (which constitutes 6% of total
formula).
2TABLE 2 Coating damage Coating % water absorption observed Sample
(III) 55% RH 60% RH 70% RH at Core no salt 1.1 1.7 2.8 No material
coating damage reference Ammonium 1.4 2.2 6.1 70% RH granule
sulfate invention Sodium 1.2 1.9 3.4 No granule citrate damage
Reference Sodium 4.8 7.3 12.3 60% RH granule formiate Purafect G
Commercial 0.9 1.9 23.2 Liquefies product at 70% RH
[0172] Constant humidities at 25.degree. C.: Ammonium sulfate=79;
sodium formiate=56 and sodium citrate=86.
Example 4
[0173] An enzyme containing granule coated with sodium citrate was
produced as in Example 1 with the exception that the pH of the
sodium citrate was adjusted to 7.5 by adding citric acid to the
coating solution prior to the coating process.
Example 5
[0174] An enzyme containing granule was produced as in Example 4
with the exception that the coating of pure sodium citrate was
replaced by a mixture of sodium citrate and sodium thiosulfate in
the weight ratio 9:1. Thus a coating consisting of 72 g sodium
citrate and 8 g thiosulfate per kg carrier was applied.
Example 6
[0175] An enzyme containing granule was produced as in Example 4
with the exception that the coating of pure sodium citrate was
replaced by a mixture of sodium citrate and sodium thiosulfate in
the weight ratio 4:1. Thus a coating consisting of 64 g sodium
citrate and 16 g thiosulfate per kg carrier was applied.
Example 7
[0176] The storage stability of the enzyme containing granules from
Examples 4-6 as well as the ammonium sulfate reference granule of
Example 1 were tested in one powder detergent (A) containing
perborate bleach and TAED and one powder detergent (B) containing
percarbonate bleach. 100 mg samples of the enzyme containing
granulates (Purafect 50 mg) and 10.0 g samples of the detergents
were mixed to constitute test samples. The samples were incubated
in open jars at air conditions 35.degree. C. and 55% RH in a
thermostated and humidity controlled cupboard. Identical samples
were taken out from the cupboard after 2 and 4 weeks and analyzed
for enzyme (Savinase.TM.) activity together with identical
reference samples, which had been stored in sealed jars below
-18.degree. C. The results are given in Example 3, wherein the
enzyme activities of the samples are given as a percentage of the
corresponding reference samples.
3 TABLE 3 Residual activity Residual activity Coating in Detergent
A in Detergent B compound 2 weeks 4 weeks 2 weeks 4 weeks Reference
ammonium 53% 36% 73% 61% granule of sulfate (Ex. 1) Invention
Sodium 80% 51% 74% 68% granule citrate pH 7.5 Invention 90% Sodium
96% 78% 90% 92% granule citrate, 10% thiosulfate Invention 80%
Sodium 103% 89% 95% 92% granule citrate, 20% thiosulfate Purafect G
Commercial 89% 79% 90% 81% product
[0177] Constant humidities at 25.degree. C.: Ammonium sulfate=79
and sodium citrate=86.
Example 8
[0178] An enzyme containing granule was produced as in Example 1
with the exception that i) the enzyme solution used for both
absorption of enzyme onto the carrier and for applying the enzyme
layer contained sodium thiosulfate in an amount corresponding to
0.33% w/w of the final coated granulate and ii) the sodium citrate
coating was replaced by a 80 g per kg carrier sodium sulfate
coating by applying a 45.degree. C., 28.6% w/w aqueous sodium
sulfate solution.
[0179] Further a reference granule was prepared identical to the
first granule in this example with the exception that the sodium
sulfate coating was replaced with a 80 g per kg carrier coating of
ammonium sulfate.
Example 9
[0180] The storage stability of the enzyme containing granules from
Example 8 were tested along with a commercial product in one powder
detergent (A) containing sodium perborate bleach and TAED, one
powder detergent (B) containing sodium percarbonate bleach and one
powder detergent (C) without bleach. 100 mg samples (50 mg Purafect
G) of the enzyme containing granulates and 10.0 g samples of the
detergents were mixed to constitute test samples. The bleach
containing samples were incubated in open jars at air conditions
35.degree. C. and 55% RH, while the samples without bleach were
incubated in open jars at air conditions 37.degree. C. and 70% RH.
All samples were incubated in thermostated and humidity controlled
cupboards. Identical samples were taken out from the cupboard after
2 and 4 weeks and analyzed for enzyme (Savinase.TM.) activity
together with identical reference samples, which had been stored in
sealed jars below -18.degree. C. The results are given in Table 4,
wherein the enzyme activities of the samples are given as a
percentage of the corresponding reference samples.
4 TABLE 4 Residual activity Residual activity Residual Coating in
Detergent A in Detergent B activity com- 2 4 2 4 in Deter- pound
weeks weeks weeks weeks gent C Reference ammo- 68% 49% 87% 78% 74%
53% granule nium sulfate Invention Sodium 90% 72% 98% 100% 93% 92%
granule sulfate Purafect G com- 92% 83% 83% 47% mercial product
[0181] Constant humidities at 25.degree. C.: Ammonium sulfate=79;
sodium sulfate=93.
[0182] The results show that coating materials with higher constant
humidity results in improved stability.
Example 10
[0183] An enzyme containing granule coated with sodium sulfate was
produced as in Example 8 with the exception that a different ultra
filtrated Savinase.TM. concentrate solution (app 41 KNPU/g) was
used throughout the process.
Example 11
[0184] A sample of dry enzyme-loaded granular carrier was produced
as described vide supra with the exception that the enzyme solution
used for absorbing enzyme onto the carrier was added 5.33 g of
sodium thiosulfate per kg finished dry carrier. The enzyme and
thiosulfate loaded carrier was transferred to a Glatt WSG5
conventional fluid bed apparatus. Using a conventional top spray
coating technique with an air inlet temperature of 70.degree. C.,
air outlet temperature of 42.degree. C. and with air volume of 600
m3/h the following steps were carried out in sequence:
[0185] a) an enzyme containing layer was applied onto the carrier
by spraying an enzyme containing aqueous solution containing
Savinase.TM., PVP/VA co-polymer (Luviscol VA64), titaniumdioxide
(TiO.sub.2) and sodium thiosulfate onto the carrier at a spraying
rate of 100 g/min. Approximately 87 g Savinase.TM. concentrate (41
KNPU/g), 2.67 g of PVP/VA co-polymer (Luviscol VA64), 1.67 g of
titaniumdioxide (TiO.sub.2) and 1.67 g sodium thiosulfate were
applied per kg carrier,
[0186] b) a 267 g sodium sulfate per kg carrier coating layer of
sodium sulfate was applied by spraying a 28.5% w/w aqueous solution
of sodium sulfate at approximately 45.degree. C. onto the product
of a), at a spraying rate for the coating solution of 100
g/min,
[0187] c) a final lubricant coating was applied by spraying a
solution of 7.33 g PEG 4000 per kg carrier onto the product of c),
at a spraying rate of 100 g/min.
[0188] The finished enzyme containing granulate was dried for 5
minutes and then cooled to 30.degree. C., where after it was
removed from the fluid bed and sifted between 300 and 1200
.mu.m
Example 12
[0189] The storage stability of the enzyme containing granulates
from Examples 13 and 14 were tested along with two commercial
products in one powder detergent (A) containing perborate bleach
and TAED, one powder detergent (B) containing percarbonate bleach
and one powder detergent (C) without bleach. 150 mg samples of the
enzyme containing granulates (75 mg Purafect E and G) and 10.0 g
samples of the detergents were mixed to constitute test samples.
The bleach containing samples were incubated in open jars at air
conditions 35.degree. C. and 55% RH, while the samples without
bleach were incubated in open jars at air conditions 37.degree. C.
and 70% RH. All samples were incubated in thermostated and humidity
controlled cupboards. Identical samples were taken out from the
cupboard after 2 and 4 weeks and analyzed for enzyme (Savinase.TM.)
activity together with identical reference samples, which had been
stored in sealed jars below -18.degree. C. The results are given in
Table 5, wherein the enzyme activities of the samples are given as
a percentage of the corresponding reference samples.
5 TABLE 5 Residual activity Residual activity Residual Coating in
Detergent A in Detergent B activity com- 2 4 2 4 in Deter- pound
weeks weeks weeks weeks gent C Invention Sodium 82% 55% 85% 77% 94%
80% granule sulfate (Ex. 10) Invention Sodium 98% 98% 96% 96% 99%
95% granule sulfate (Ex. 11) Purafect E com- 88% 65% 51% 41% 65%
17% mercial product Purafect G com- 95% 75% 73% 62% -- -- mercial
product
[0190] The results given in Table 5 shows that application of a
thick coating of with a high constant humidity leads to superior
stability even if only a lubricant is applied as an additional
coating.
Example 13
[0191] 20 kg of uncoated Savinase.RTM. (protease enzyme) containing
granulate (type TX) was produced as described in U.S. Pat. No.
4,106,991 Example 1 with the following exceptions:
[0192] 1) sodium sulfate was used in stead of sodium chloride as
filler material
[0193] 2) the enzyme concentrate was an aqueous suspension of
crystalline enzyme containing also a binder (Glucidex) and 0.4% w/w
methionine as an antioxidant
[0194] The uncoated enzyme granule was transferred to a 50 liter
Lodige mixer and sprayed with 2.0 kg of a solution/suspension
consisting of 25% sodium sulfate, 12% dextrine, 7% TiO2 and 56% of
water. The granular mass was during the spraying treated with the
chopper as described in U.S. Pat. No. 4,106,991.
[0195] The mixer treated granulate was subsequently transferred to
a Glatt WSG5 fluid bed and dried. 15 kg of the dried granulate was
thereafter successively coated in three steps with parameters as
described in Example 1.
[0196] a) the enzyme containing granulate was in a first step
coated with 11.0 kg of a solution/suspension consisting of 27.1%
sodium sulfate 3.9% TiO2, 1.0% dextrin and water ad 100%. The salt
solution was during the spraying kept at 45-50.degree. C. to avoid
crystallisation of the sodium sulfate. The coated the enzyme
containing granule with a coating containing 85% w/w sodium
sulfate.
[0197] b) an additional coating was applied as a dust suppressing
film by spraying with 3.5 kg of a solution consisting of 6.3%
methylhydroxypropyl cellulose (Aqualon 8MP5C) 6.3% PEG 4000 and
water ad 100%
[0198] c) the granulate was in a final step sprayed with 0.46 kg of
a 24% PEG 4000/water solution.
Example 14
[0199] A comparative base granulate was prepared as described in
Example 13 without the mixer treatment and without fluid bed
coatings and was conventionally coated as described in US 106,991
Example 22 by applying a solution of 7% PEG 4000 and 12.5% of a 1:1
TiO2/Kaolin mixture.
Example 15
[0200] The storage stability of granulates of Examples 13 and 14
was tested in a perborate containing (A) and a percarbonate
containing (B) detergent as described in Example 4.
6 TABLE 6 Residual Residual activity in activity in detergent A
detergent B Coating 2 4 2 4 compound weeks week weeks weeks Example
13 Sodium 98 92 103 99 granulate sulfate Example 14 comparative 69
43 63 48 granulate granule Purafect G Commercial 90 73 97 83
product
Example 16
[0201] 15 kg of dry Savinase enzyme absorbed on a granular carrier
was transferred to a Glatt WSG conventional fluid bed apparatus.
Using a conventional top spray coating technique with an air inlet
temperature of 70.degree. C. and an air outlet of 42.degree. C. and
with an air volume air volume of 600 m.sup.3/h were the following
steps carried out in sequence:
[0202] a) an enzyme containing layer was applied onto the carrier
by spraying granular carrier with an enzyme containing solution
consisting of 2.07 kg of a liquid Savinase concentrate (82% dry
matter, 24 KNPU/g), 50 g Glucidex 21D and 54 g TiO2 and with a
spraying rate of 100 g/min.
[0203] b) The salt layer was applied in a second step by spraying
with 14.7 kg of a solution consisting of 27% sodium sulfate, 3.9%
TiO2, 1,0% Glucidex 21D and water ad 100%. The temperature of the
solution was kept at 45-50.degree. C. to avoid crystallisation of
the salt.
[0204] c) The salt coated granulate was in a next step coated with
a dust suppressing film by spraying with 3.4 kg of a solution
consisting of 6.3% methylhydroxypropyl cellulose, 6.3% PEG 4000 and
water ad 100%.
[0205] d) the granulate was in a final step sprayed with 0.46 kg of
a 24% PEG 4000/water solution.
[0206] All enzyme concentrate used for this preparation had an
addition of sodium thiosulfate to the concentrate corresponding to
0.3% w/w of the final granulate.
Example 17
[0207] This example was prepared as Example 16 with the exception
15that the solution for the coating (b) was reduced to 11,0 kg.
Example 18
[0208] This example was prepared as Example 16 with the exception
that the solution for the coating (b) was reduced to 7.35 kg.
Example 19
[0209] This example was produced according to Example 16 step (a),
i.e. without the salt coating and the dust suppressing film.
Example 20
[0210] The storage stability of granulates of Examples 16-19 was
tested and compared to references in three detergents as described
in Example 9.
7 TABLE 7 Residual Residual Residual activity (%) activity (%)
activity (%) in detergent in detergent in detergent A B C Coating 2
4 2 4 2 4 compound weeks week weeks weeks weeks week Ex- Sodium 99
99 98 98 94 82 ample sulfate 16 Ex- Sodium 100 96 94 97 92 71 ample
sulfate 17 (75% relative to Example 16) Ex- Sodium 97 94 96 97 93
76 ample sulfate 18 (50% relative to Example 16) Ex- Com- 60 44 55
43 79 49 ample parative 19 granule without sodium sulfate Ex- Com-
69 43 63 48 88 62 ample parative 14 granule Purafect Com- 90 73 97
83 89 47 G mercial product
Example 21
[0211] An Endolase (cellulase enzyme) containing granulate was
produced as described in U.S. Pat. No. 4,106,991 Example 1 with the
following exceptions:
[0212] a) The filler was sodium sulfate
[0213] b) The liquid enzyme concentrate was used as the granulating
liquid.
[0214] c) The granulate had furthermore an addition of 10% w/w of a
carbohydrate binder and 0.5% w/w of sodium thiosulfate.
[0215] The granulate was conventionally coated as described in US
106,991 Example 22 by applying a solution of 7.2% w/w PEG 4000 and
13,0% w/w of a 1:1 mixture of TiO2 and kaolin.
Example 22
[0216] An Endolase (cellulase enzyme) containing granulate was
produced as described in Example 21. This base granulate was coated
according to the steps (b)-(d) in Example 16 with the exception
that 10.6 kg of the salt solution was applied in step (b).
Example 23
[0217] The storage stability of granulates of Examples 21 and 22
was tested in a sodium perborate containing detergent (A) with
conditions as described in Example 9.
8 TABLE 8 Residual activity (%) in detergent A Coating 1 3 5
compound week weeks weeks Example 21 Comparative 97 28 23 granule
Example 22 Sodium 100 70 sulfate
Example 24
[0218] A Natalase.RTM. (an amylase enzyme) containing granulate was
produced as described in U.S. Pat. No. 4,106,991 Example 1 with the
following exceptions:
[0219] 1) sodium sulfate was used in stead of sodium chloride as
filler material
[0220] 2) the enzyme concentrate was an aqueous suspension of
crystalline enzyme which was also used as the granulating liquid
containing also a binder (Glucidex).
[0221] 3) The granulate had furthermore an addition of 0.4% w/w
sodium thiosulfate (calculated as % of the uncoated granulate).
[0222] The granulate was conventionally coated as described in U.S.
Pat. No. 106,991 Example 22 by applying a solution of 7% of PEG
4000 and 12.5% of a 1:1 mixture of TiO2 and kaolin.
Example 25
[0223] A Natalase.RTM. containing granulate was produced as
described in Example 24. 1300 g of the Natalase containing base
granulate was transferred to a UniGlatt fluid bed where coated with
a salt coating by spraying it with a 50.degree. C. salt solution at
consisting of:
[0224] 234 g of sodium sulfate
[0225] 9 g of Glucidex 21D
[0226] 25 g of TiO2
[0227] 585 g of water
[0228] The spraying conditions were air inlet temperature
70.degree. C. and air outlet temperature 42.degree. C. The
granulate was after finishing the spraying further dried in the
fluid bed for 5 minutes
Example 26
[0229] The storage stability of granulates of Examples 24 and 25
was tested in the percarbonate containing detergent (B) with
conditions as described in Example 9.
9 TABLE 9 Residual activity in detergent B Coating 1 2 4 compound
week weeks weeks Example 24 Comparative 82 79 69 granule Example 25
Sodium 98 97 94 sulfate
Example 27
[0230] A non-coated Savinase.RTM. containing granulate (granulate
A) was produced as described in U.S. Pat. No. 4,106,991 Example 1
wit the following exceptions:
[0231] 1) sodium sulfate was used in stead of sodium chloride as
filler material
[0232] 2) the enzyme concentrate was an aqueous suspension of
crystalline enzyme containing also a carbohydrate binder (Glucidex)
and methionine as an antioxidant
[0233] The non-coated granules was coated with a salt layer using a
fluid bed according to the following procedure:
[0234] a) 15 kg of the uncoated granules were fluidized in a Glatt
WSG-5 fluid bed using 550 m.sup.3 air per hour. The air inlet
temperature was 70.degree. C.
[0235] b) A salt solution of 2.0 kg Na.sub.2SO.sub.4 dissolved in
5.0 kg water at 50.degree. C. was prepared. 2.1 kg of this solution
was sprayed onto the fluidized granules at a rate of 100 grams
solution per minute. During spraying of the liquid the product
temperature was approximately 42.degree. C. After adding the
solution the water was allowed to evaporate from the coated
granules (until the product temperature raised quickly in the fluid
bed). A sample of 2.0 kg (granulate Al) of the coated granules was
taken out and the coating process was repeated to add further
coating to the granules remaining in the fluid bed by spraying the
remaining granules with an additional 1.75 kg of the salt solution.
A sample of 2.0 kg of the additionally coated granules (granulate
A2) was taken out and the coating process was repeated once more to
add even further coating to the granules remaining in the fluid bed
by spraying the remaining granules with the remaining 3.15 kg of
the salt solution (granulate A3).
Example 28
[0236] The enzyme stability in the granules were tested in a model
powder detergent containing Sodium Perborate at 35.degree. C. and
55% relative humidity in open boxes according to Example 9.
10TABLE 10 Residual Savinase activity in % 2 4 Granulate weeks
weeks Uncoated Savinase .RTM. 47 36 granules A1 (4.0%
Na.sub.2SO.sub.4) 68 42 A2 (7.8% Na.sub.2SO.sub.4) 77 55 A3 (15.8%
Na.sub.2SO.sub.4) 92 75
[0237] The % salt was calculated as % w/w salt of the uncoated
cello granules under the condition that all water was evaporated.
From Example 10 it was concluded that the enzyme stability in the
Savinase.RTM. granules were significantly improved by the salt
coating and that increasing the amount of salt improves the
stability.
* * * * *