A LIQUID ENZYME FORMULATION AND A PROCESS FOR ITS PREPARATION
FIELD OF THE INVENTION
[0001] The present invention relates to a liquid enzyme formulation,
particularly to a liquid and stable formulation comprising a crosslinking enzyme
and/or an enzyme modifying milk proteins. Particularly the present invention
relates a liquid and stable transglutaminase formulation. In addition, the pre
sent invention relates to a method for preparing a liquid enzyme formulation.
BACKGROUND OF THE INVENTION
[0002] Enzyme formulations comprising a crosslinking enzyme
and/or an enzyme modifying milk proteins, such as laccase, tyrosinase, perox
idase, sulfhydryl oxidase or glucose oxidase are commercially available both in
powder and liquid formulations. However, transglutaminase and protein glutaminase
products are currently in the market only in powder form. The use of
a powdered enzyme product is not totally accepted due to the dust formation in
all production plants. The health hazards resulting from the dusting have in
curred concerns among the workers, especially.
[0003] A transglutaminase derived from Streptoverticillium mobaraense
strain and a process for its preparation has been disclosed in the Euro
pean patent No. 0 379 606 B 1 . Further, a method for the production of a
transglutaminase using a gene isolated from Streptomyces lydicus strain is
disclosed in the European Patent No. 0 777 726 B 1 .
[0004] One of the problems associated with formulating an enzyme,
such as a transglutaminase, in liquid form, is the lack of stability of the formula
tion. Further, one of the disadvantages associated with the present liquid e n
zyme formulations is that they contain at least one preservative.
BRIEF DESCRIPTION OF THE INVENTION
[0005] An object of the present invention is thus to provide a liquid
enzyme formulation comprising transglutaminase and/or another milk protein
crosslinking and/or modifying enzyme such as laccase, tyrosinase, peroxidase,
sulfhydryl oxidase, glucose oxidase or protein glutaminase, which is stable and
can be stored for a time period required from a commercial formulation in a
room temperature or in temperatures of a refrigerator and/or a freezer.
[0006] Another object of the present invention is to provide a liquid
formulation comprising a transglutaminase, which is stable and can be stored
for a time period required from a commercial formulation in a room tempera
ture or in temperatures of a refrigerator and/or a freezer.
[0007] A further object of the present invention is to provide a meth
od for the preparation of a stable, liquid enzyme formulation comprising trans
glutaminase and/or another milk protein crosslinking and/or modifying enzyme.
[0008] An even further object of the present invention is to provide a
method for the preparation of a stable, liquid formulation comprising a transglu
taminase.
[0009] The objects of the invention are achieved by the formulations
and methods set forth in the independent claims. Preferred embodiments of
the invention are described in the dependent claims.
[0010] Other objects, details and advantages of the present inven
tion will become apparent from the following detailed description and examples.
DETAILED DESCRIPTION OF THE INVENTION
[001 1] Milk protein crosslinking and/or modifying enzymes such as
transglutaminase, laccase, tyrosinase, peroxidase, sulfhydryl oxidase and pro
tein glutaminase catalyze milk protein modifications. There seems to be syner
gism within the action of these enzymes and further with the action of glucose
oxidase. Without being bound by any theory glucose oxidase and/or peroxi
dase seem to catalyze reactions wherein oxygen is released through hydrogen
peroxide formation. The oxygen can then catalyze (oxidate) crosslinking of ty
rosinase.
[0012] Milk protein crosslinking and/or modifying enzymes such as
transglutaminase, laccase, tyrosinase, peroxidase, sulfhydryl oxidase and pro
tein glutaminase optionally together with glucose oxidase are used in the man
ufacture of processed fish, meat and egg products, pastes and pates, fruits,
berries and vegetables, soy products, cereal products, bread and bakery prod
ucts.
[0013] Following milk protein crosslinking and/or modifying enzymes
are all relevant in food prosessing in dairy or other food categories.
[0014] Transglutaminases are a family of enzymes (EC 2.3.2.1 3)
that catalyze the generation of covalent linkages between the glutamine and
lysine amino acid residues present in the protein molecules. When linkages
are formed, ammonia is released.
[0015] Laccases (EC 1.1 0.3.2) derived from fungi and bacteria,
such as, fungus Trametes hirsute, catalyze the crosslinking between carbohy
drates and proteins (oxidation of aromatic compounds and cysteine) with a p
plications in food processing for reduction of allergenicity, for example.
[0016] Tyrosinases (EC 1.14.1 8.1 ) are enzymes which catalyzes
the oxidation of phenols such as tyrosine, with applications in food processing
for reduction of allergenicity, for example.
[0017] Peroxidases (EC 1.1 1.1 .7) are a family of enzymes that cata
lyze the oxidation of aromatic compounds with applications in food processing
for reduction of allergenicity, for example.
[0018] Sulfhydryl oxidase (EC 1.8.3.3) catalyzes the formation of d i
sulfide bonds, oxidation of glutathione.
[0019] Protein glutaminase catalyzes the deamidation of protein
bound glutamine, and glutamine is converted to glutamic acid.
[0020] Glucose oxidase catalyzes the formation of protein crosslinks
and oxidative gelation of pentosans.
[0021] Milk protein crosslinking and/or modifying enzymes such as
transglutaminase, laccase, tyrosinase, peroxidase, sulfhydryl oxidase and pro
tein glutaminase are used in dairy industry to stabilize the structure of milkbased
products. In addition to dairy industry, these enzymes are used in the
manufacture of processed fish, meat and egg products, pastes and pates,
fruits, berries and vegetables, soy products, cereal products, bread and bakery
products. Accordingly, a liquid enzyme formulation is suited for the manufac
ture of dairy, processed fish, meat, egg, pastes and pates, fruits, berries and
vegetables, soy, cereal, bread and bakery products, for example. The use of a
liquid enzyme preparation would be more practical and advantageous than the
use of powder formulation, especially in the industrial scale manufacturing.
[0022] Transglutaminases are most active in the pH range from 5.2
to 8. When a liquidized transglutaminase is stored at pH 5.2 or at pH 8, the
enzyme loses its activity quickly. After storing a transglutaminase at pH 5.2 for
7 days at room temperature or in the temperature of a refrigerator, only half of
the activity is left.
[0023] The invention is based on the finding that when a transglu
taminase, tyrosinase or protein glutaminase is stored in a suspension of a
polyol, such as glycerol or sorbitol, and water in the pH range from 4.4 to 5.1 ,
its activity is remained moderately during storage at room temperature and
excellently during storage in the temperatures of a refrigerator and/or a
freezer. In addition, the invention is based on the finding that when a
transglutaminase together with a protein glutaminase is stored in a suspen
sion of glycerol and water at pH 4.6, the activity of the enzymes remained
moderately during storage at room temperature and excellently during sto r
age in the temperatures of a refrigerator and/or a freezer. Further, the inven
tion is based on the finding that when a transglutaminase together with a pro
tein glutaminase and tyrosinase is stored in a suspension of glycerol and wa
ter at pH 4.6, the activity of the enzymes remained moderately during storage
at room temperature and excellently during storage in the temperatures of a
refrigerator and/or a freezer. In addition, the liquid preparations of the present
invention are also microbiologically stable during the storage at room temper
ature and in the temperatures of a refrigerator and/or a freezer. Further, it
was unexpectedly found that liquid enzyme formulation of the present inven
tion maintained its enzyme activity and microbiological purity (no microbial
growth) without any preservatives in the formulation in the temperatures of a
refrigerator and/or a freezer.
[0024] In one embodiment, the liquid enzyme formulation in polyolwater
suspension has pH value within the range from 4.4 to 5.1 . In another
embodiment of the present invention, the pH of the enzyme formulation is with
in the range from 4.4 to 4.8. In one embodiment of the present invention, the
pH of the enzyme formulation is 4.4. In another embodiment of the present
invention, the pH of the enzyme formulation is 4.6. In a further embodiment of
the present invention, the pH of the enzyme formulation is 4.8. In an even further
embodiment of the present invention, the pH of the enzyme formulation is 5.1 .
[0025] Polyols, such as glycerol, sorbitol, xylitol and/or mannitol can
be used in the liquid enzyme formulation of the present invention. Mixtures of
the polyols, such as mixtures of glycerol and the other polyols are also operable.
[0026] The suspension of a polyol and water or a mixture of two or
more polyols and water suitable for formulating the enzyme preparation of the
present invention may contain the polyol(s), such as glycerol, sorbitol, xylitol
and/or mannitol from 25% up to 100%, preferably from 50% up to 100%
(w/w%). In one embodiment of the present invention, the enzyme is dissolved
into 25% polyol/75% water (w/w) suspension. In another embodiment of the
present invention, the enzyme is dissolved into 50% polyol/ 50% water (w/w)
suspension. In a further embodiment of the present invention, the enzyme is
dissolved into 75% polyol/25% water (w/w) suspension.
[0027] The suspension of glycerol and water suitable for formulating
the enzyme preparation of the present invention may contain glycerol from
25% up to 100%, preferably from 50% up to 100%. In one embodiment of the
present invention, the enzyme is dissolved into 25% glycerol/75% water sus
pension. In another embodiment of the present invention, the enzyme is d is
solved into 50% glycerol/ 50% water suspension. In a further embodiment of
the present invention, the enzyme is dissolved into 75% glycerol/25% water
suspension. Further, the suspension of sorbitol and water suitable for formulat
ing the enzyme preparation of the present invention may contain sorbitol from
25% up to 100%, preferably from 50% up to 100%. In one embodiment of the
present invention, the enzyme is dissolved into 25% sorbitol/75% water sus
pension. In another embodiment of the present invention, the enzyme is d is
solved into 50% sorbitol/50% water suspension. In a further embodiment of the
present invention, the enzyme is dissolved into 75% sorbitol/25% water sus
pension.
[0028] The pH of the suspension may be adjusted to the desired
range with an acid approved for food use, such as, lactic acid, GDL (gluconodeltalactone),
citric acid, acetic acid, oxalic acid, malic acid, pantothenic acid,
propionic acid and/or hydrochloric acid, or any mixtures/combinations thereof
in the form of acid or salt. In one embodiment of the present invention, lactic
acid is used for the pH adjustment.
[0029] The liquid enzyme preparation of the present invention may
optionally contain also a preservative such as Na-benzoate. In one embodi
ment, the liquid enzyme preparation of the present invention does not contain
any additional preservatives i.e., the formulation is free from preservatives. In
another embodiment, the liquid enzyme preparation of the present invention
contains an additional preservative. In a further embodiment, the liquid enzyme
preparation of the present invention contains Na-benzoate as a preservative.
In an even further embodiment, the liquid enzyme preparation of the present
invention contains Na-benzoate, in an amount of 0.1 to 1%, preferably in an
amount of 0.7% as a preservative.
[0030] The liquid enzyme preparation of the present invention main
tains its activity in room temperature for about two weeks, in a refrigerator for
about 1.5 to six months, at least and in freezer from a minimum of 5 months up
to 24 months.
[0031] In one embodiment of the invention, the liquid formulation
comprises one milk protein crosslinking and/or modifying enzyme. In another
embodiment of the invention, the liquid formulation comprises two or more milk
protein crosslinking and/or modifying enzymes. In one embodiment, the milk
protein crosslinking and/or modifying enzyme in the formulation of the present
invention is transglutaminase. In another embodiment, the milk protein crosslinking
and/or modifying enzyme in the formulation of the present invention is
tyrosinase. In another embodiment, the milk protein crosslinking and/or modify
ing enzyme in the formulation of the present invention is protein glutaminase.
In another embodiment, the milk protein crosslinking and/or modifying e n
zymes in the formulation of the present invention are transglutaminase and
protein glutaminase. In another embodiment, the milk protein crosslinking
and/or modifying enzymes in the formulation of the present invention are
transglutaminase and tyrosinase. In another embodiment, the milk protein
crosslinking and/or modifying enzymes in the formulation of the present inven
tion are transglutaminase and laccase. In a further embodiment, the milk pro
tein crosslinking and/or modifying enzymes in the formulation of the present
invention are transglutaminase, protein glutaminase and laccase. In an even
further embodiment, the milk protein crosslinking abd/or modifying enzymes in
the formulation of the present invention are transglutaminase, protein glutami
nase and tyrosinase.
[0032] The present invention relates also to a method for preparing
a liquid enzyme formulation wherein a milk protein crosslinking and/or modify
ing enzyme is added to polyol-water suspension having pH value within the
range from 4.4 to 5.1 . In one embodiment of the present invention, the pH of
the polyol-water suspension is adjusted to a value within the range from 4.4 to
5.1 . In one embodiment of the present invention, the pH of the polyol-water
suspension is adjusted to a value within the range from 4.4 to 4.8. In one em
bodiment of the present invention, the pH of the enzyme formulation is adjust
ed to pH 4.4. In another embodiment of the present invention, the pH of the
polyol-water suspension is adjusted to pH 4.6. In a further embodiment of the
present invention, the pH of the polyol-water suspension is adjusted to pH 4.8.
In an even further embodiment of the present invention, the pH of the enzyme
formulation is adjusted to pH 5.1 .
[0033] The suspension of polyol and water suitable for the method
of the present invention may contain polyol from 25% up to 100% (w-%). In
one embodiment of the present invention, the enzyme is dissolved into 25%
polyol-water suspension. In another embodiment of the present invention, the
enzyme is dissolved into 50% polyol-water suspension. In a further embodi
ment of the present invention, the enzyme is dissolved into 75% polyol-water
suspension. In one embodiment of the invention, the polyol is glycerol. In a n
other embodiment of the invention, the polyol is sorbitol.
[0034] In the present method, the pH of the suspension may be ad
justed to the desired range with an acid approved for food use (food grade,
GRAS), such as, lactic acid, GDL, citric acid, acetic acid, oxalic acid, malic acid,
pantothenic acid, propionic acid and/or hydrochloric acid or any mix
tures/combinations thereof in the form of acid or salt. In one embodiment, lactic
acid is used in the method of the present invention for the pH adjustment.
[0035] In the method of the present invention, also a preservative,
such as Na-benzoate, may optionally be included in the liquid enzyme formula
tion. In one embodiment, the method of the present invention does not com
prise addition of a preservative. In another embodiment, the method of the
present invention comprises an addition of a preservative. In a further embod
iment, the method of the present invention comprises addition of Na-benzoate
as a preservative. In an even further embodiment, the method of the present
invention comprises addition of Na-benzoate, in an amount of 0.1 to 1%, pref
erably in an amount of 0.7% as a preservative.
[0036] In one embodiment, the method of the present invention
comprises the following steps:
a) pH of a polyol-water suspension is adjusted with food grade acid(
s) to a value within the range from 4.4 to 5.1
b) a milk protein crosslinking and/or modifying enzyme is added to
the suspension
c) optionally a preservative is added.
[0037] In another embodiment, the method of the present invention
comprises the following steps:
a) a milk protein crosslinking and/or modifying enzyme is added to a
polyol-water suspension
b) pH of the suspension is adjusted with food grade acid(s) to a val
ue within the range from 4.4 to 5.1
c) optionally a preservative is added.
[0038] In one embodiment of the present invention, one milk protein
crosslinking and/or modifying enzyme is added to the suspension. In another
embodiment of the present invention, two or more milk protein crosslinking
and/or modifying enzymes are added to the suspension. In one embodiment,
the milk protein crosslinking and/or modifying enzyme is transglutaminase. In
another embodiment, the milk protein crosslinking and/or modifying enzyme is
tyrosinase. In another embodiment, the milk protein crosslinking and/or modify
ing enzyme is protein glutaminase. In another embodiment, the milk protein
crosslinking and/or modifying enzymes are transglutaminase and protein glu
taminase. In another embodiment, the milk protein crosslinking and/or modify
ing enzymes are transglutaminase and tyrosinase. In another embodiment, the
milk protein crosslinking and/or modifying enzymes are transglutaminase and
laccase. In a further embodiment, the milk protein crosslinking and/or modify
ing enzymes are transglutaminase, protein glutaminase and laccase. In an
even further embodiment, the milk protein crosslinking and/or modifying e n
zymes are transglutaminase, protein glutaminase and tyrosinase.
[0039] The invention will be described in more detail by means of
the following examples. The examples are not to be construed to limit the
claims in any manner whatsoever.
EXAMPLES
COMPARATIVE EXAMPLE 1
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol -water
suspension at pH 5.2 at 4°C
[0040] The transglutaminase derived from Streptoverticillium mobaraense-
strain having activity of 16.300 nkat/g (Activa® TG, Ajinomoto), was
dissolved in activity of 274 nkat g into 50% glycerol-water (w/w) suspension
which pH was adjusted to 5.2 with lactic acid. The enzyme activity was moni
tored for 7 days. Additionally, the microbiological purity of the preparation was
monitored.
[0041] On day 7, only 50% of the activity of the transglutaminase
was left. No microbial growth was detected during the seven day preservation
test.
COMPARATIVE EXAMPLE 2
Transglutaminase preparation Activa® TG (Ajinomoto) in water at pH 5.2
at 4°C
[0042] The transglutaminase Activa® TG (Ajinomoto) was dissolved
in activity of 274 nkat/g into water having pH 5.2 adjusted with lactic acid. The
suspension contained also 0.7% Na-benzoate as a preservative.
[0043] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0044] On day 50, only 43% of the activity of the transglutaminase
was left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 1
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol -water
suspension at pH 4.6 at 4°C
[0045] The transglutaminase Activa® TG (Ajinomoto) was dissolved
in activity of 274 nkat/g into 50% glycerol -water (w/w) suspension having pH
4.6 adjusted with lactic acid.
[0046] The enzyme activity was monitored for 7 days. Additionally,
the microbiological purity of the preparation was monitored.
[0047] On day 7, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the seven day preservation test.
EXAMPLE 2
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol -water
suspension at pH 4.6 at 4°C
[0048] The transglutaminase Activa® TG (Ajinomoto) was dissolved
in activity of 326 nkat/g into 50% glycerol -water (w/w) suspension having pH
4.6 adjusted with lactic acid. The suspension contained also 0.7% Nabenzoate
as a preservative.
[0049] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0050] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 3
Transglutaminase preparation Activa® TG-YG (Ajinomoto) in glycerolwater
suspension at pH 4.6 at 4°C
[0051] The liquid transglutaminase formulation was prepared by
dissolving Ajinomoto's transglutaminase preparation Activa® TG-YG, which
contains a transglutaminase derived from Streptoverticillium mobaraensestrain
and glutathione into 50% glycerol -water (w/w) suspension having pH 4.6
adjusted with lactic acid.
[0052] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0053] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 4
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol -water
suspension at pH 4.4 at 4°C
[0054] The liquid transglutaminase preparation was prepare by d is
solving transglutaminase Activa® TG (Ajinomoto) in activity of 274 nkat/g into
50% glycerol -water (w/w) suspension which pH was adjusted with lactic acid to
pH 4.4.
[0055] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0056] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 5
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol -water
suspension at pH 4.8 at 4°C
[0057] The liquid transglutaminase preparation was prepare by d is
solving transglutaminase Activa® TG (Ajinomoto) in activity of 274 nkat/g into
50% glycerol-water (w/w) suspension which pH was adjusted with lactic acid to
pH 4.8.
[0058] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0059] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 6
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol-water
suspension at pH 5.1 at 4°C
[0060] The liquid transglutaminase preparation was prepare by d is
solving transglutaminase Activa® TG (Ajinomoto) in activity of 274 nkat/g into
50% glycerol-water (w/w) suspension which pH was adjusted with lactic acid to
pH 5.1 at 4°C.
[0061] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0062] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 7
Transglutaminase preparation Saprona TG (Yiming Biological Products
Co, China) in glycerol-water suspension at pH 4.6 at 4°C
[0063] The liquid transglutaminase preparation was prepare by d is
solving transglutaminase Yiming Saprona TG derived from Streptoverticillium
mobaraense-strain into 50% glycerol-water (w/w) suspension which pH was
adjusted with lactic acid to pH 4.6.
[0064] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0065] On day 50, 100% of the activity of the transglutaminase was left.
No microbial growth was detected during the 50 day preservation test.
EXAMPLE 8
Transglutaminase preparation Reactyn CL 1000 TG (Campus SpA, Italy)
in glycerol-water suspension at pH 4.6 at 4°C
[0066] The liquid transglutaminase preparation was prepare by d is
solving transglutaminase Campus Reactyn CL 1000 TG into 50% glycerolwater
(w/w) suspension which pH was adjusted with lactic acid to pH 4.6.
[0067] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0068] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 9
Transglutaminase preparation TG-PG (Ajinomoto) in glycerol-water sus¬
pension at pH 4.6 at 4°C
[0069] The liquid preparation was prepare by dissolving into 50%
glycerol-water (w/w) suspension having pH 4.6 adjusted with lactic acid, an
enzyme preparation TG-PG (Ajinomoto). The preparation TG-PG (Ajinomoto)
contains a transglutaminase derived from Streptoverticillium mobaraensestrain
and a protein glutaminase derived from Chryseobacterium proteolyticum.
[0070] The transglutaminase activity of the liquid preparation was
100 U/g and the protein glutaminase activity of the liquid preparation was 100
U/g.
[0071] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0072] On day 50, 100% of the activity of the transglutaminase and
100% of the activity of protein glutaminase were left. No microbial growth was
detected during the 50 day preservation test.
EXAMPLE 10
Transglutaminase preparation Activa® TG (Ajinomoto) in 75% glycer¬
ol/25% water suspension at pH 4.6 at 4°C
[0073] The transglutaminase Activa® TG (Ajinomoto) was dissolved
in activity of 326 nkat/g into 75% glycerol/25%water (w/w) suspension having
pH 4.6 adjusted with lactic acid.
[0074] The enzyme activity was monitored for 50 days at the tem
perature of 4°C. Additionally, the microbiological purity of the preparation was
monitored.
[0075] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the preservation test.
EXAMPLE 11
Transglutaminase preparation Activa® TG (Ajinomoto) in 25% glycer¬
ol/75% water suspension at pH 4.6 at 4°C
[0076] The transglutaminase Activa® TG (Ajinomoto) was dissolved
in activity of 326 nkat/g into 25% glycerol/75%water (w/w) suspension having
pH 4.6 adjusted with lactic acid.
[0077] The enzyme activity was monitored for 50 days at the tem
perature of 4°C. Additionally, the microbiological purity of the preparation was
monitored.
[0078] On day 50, 72% of the activity of the transglutaminase was
left. No microbial growth was detected during the preservation test.
EXAMPLE 12
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol -water
suspensions at pH 4.4 - 4.8 at 22°C
[0079] The liquid transglutaminase preparations were prepare by
dissolving transglutaminase Activa® TG (Ajinomoto) in activity of 2789 nkat/g
into 50% glycerol-water (w/w) suspension which pH was adjusted with lactic
acid to pH 4.4 - 4.8.
[0080] The enzyme activity of the preparations was monitored for 13
weeks at the temperature of 22°C. Additionally, the microbiological purity of the
preparations was monitored.
[0081] After two weeks storage, the activity of the enzyme in the
preparations started decreasing. No microbial growth was detected during the
preservation test.
EXAMPLE 13
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol-water
suspensions at pH 4.4 - 4.8 at 4°C
[0082] The liquid transglutaminase preparations were prepare by
dissolving transglutaminase Activa® TG (Ajinomoto) in activity of 2789 nkat/g
into 50% glycerol-water (w/w) suspension which pH was adjusted with lactic
acid to pH 4.4 - 4.8.
[0083] The enzyme activity of the preparations was monitored for 26
weeks at the temperature of 4°C. Additionally, the microbiological purity of the
preparations was monitored.
[0084] After 26 weeks storage, 89% of the activity of the enzyme
was left. No microbial growth was detected during the preservation test.
EXAMPLE 14
Transglutaminase preparation Activa® TG (Ajinomoto) in glycerol-water
suspensions at pH 4.4 - 4.8 at -20°C
[0085] The liquid transglutaminase preparations were prepare by
dissolving transglutaminase Activa® TG (Ajinomoto) in activity of 2789 nkat/g
into 50% glycerol-water (w/w) suspension which pH was adjusted with lactic
acid to pH 4.4 - 4.8.
[0086] The enzyme activity of the preparations was monitored for 26
weeks at the temperature of -20°C. Additionally, the microbiological purity of
the preparations was monitored.
[0087] After 26 weeks storage, 97% of the activity of the enzyme
was left. No microbial growth was detected during the preservation test.
EXAMPLE 15
Tyrosinase preparation in glycerol-water suspension at pH 4.6 at 4°C
[0088] The tyrosinase enzyme was dissolved in activity of 100 U/g in
to 50% glycerol-water (w/w) suspension which pH was adjusted with lactic acid
to pH 4.6.
[0089] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0090] After 50 days storage, 97% of the activity of the tyrosinase
was left. No microbial growth was detected during the preservation test.
EXAMPLE 16
Protein glutaminase preparation in glycerol-water suspension at pH 4.6
at 4°C
[0091] The protein glutaminase was dissolved in activity of 100 U/g in
to 50% glycerol-water (w/w) suspension which pH was adjusted with lactic acid to
pH 4.6.
[0092] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0093] After 50 days storage, 96% of the activity of the protein glu
taminase was left. No microbial growth was detected during the preservation
test.
EXAMPLE 17
Transglutaminase preparation Activa® TG (Ajinomoto) in sorbitol-water
suspension at pH 4.6 at 4°C
[0094] The liquid transglutaminase preparation was prepared by d is
solving transglutaminase Activa® TG (Ajinomoto) in activity of 274 nkat/g into
50% sorbitol-water (w/w) suspension which pH was adjusted with lactic acid to
pH 4.6.
[0095] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0096] On day 50, 100% of the activity of the transglutaminase was
left. No microbial growth was detected during the 50 day preservation test.
EXAMPLE 18
Liquid enzyme preparation containing TG-PG (Ajinomoto) and tyrosinase
in glycerol-water suspension at pH 4.6 at 4°C
[0097] The liquid preparation was prepare by dissolving into 50%
glycerol-water (w/w) suspension having pH 4.6 adjusted with lactic acid, an
enzyme preparation TG-PG (Ajinomoto) and tyrosinase. The transglutaminase
activity of the liquid preparation was 100 U/g, the protein glutaminase activity
of the liquid preparation was 100 U/g and tyrosinase activity of the liquid prepa
ration was 100 U/g.
[0098] The enzyme activity was monitored for 50 days. Additionally,
the microbiological purity of the preparation was monitored.
[0099] On day 50, 100% of the activity of the transglutaminase,
100% of the activity of protein glutaminase and 98% of the activity of tyrosi
nase were left. No microbial growth was detected during preservation test.
[0100] It will be obvious to a person skilled in the art that, as the
technology advances, the inventive concept can be implemented in various
ways. The invention and its embodiments are not limited to the examples de
scribed above but may vary within the scope of the claims.
Claims
1. A liquid enzyme formulation c h a r a c t e r i z e d in that it com
prises at least one milk protein crosslinking and/or modifying enzyme in polyolwater
suspension comprising from 25% to 100% (w/w) polyol and having pH
value within the range from 4.4 to 5.1 .
2 . The formulation according to claim 1, c h a r a c t e r i z e d in that
the polyol-water suspension comprises from 50% to 75% polyol.
3 . The formulation according to claim 1 or claim 2, c h a r a c t e r
i z e d in that the polyol is glycerol or sorbitol.
4 . The formulation according to any one of claims 1 to 3, c h a r -
a c t e r i z e d in that the pH is 4.6.
5 . The formulation according to any one of claims 1 to 4, c h a
a c t e r i z e d in that the formulation comprises transglutaminase, tyrosinase
or protein glutaminase.
6 . The formulation according to any one of claims 1 to 4 c h a r a c -
t e r i z e d in that the formulation comprises transglutaminase and protein glu
taminase.
7 . The formulation according to claim 5 or claim 6 c h a r a c t e r
i z e d in that the formulation comprises also laccase and/or tyrosinase.
8 . The formulation according to any one of claims 1 to 7 c h a r a c -
t e r i z e d in that the formulation is free from preservatives.
9 . A method for preparing a liquid enzyme formulation c h a a c
t e i z e d in that at least one milk protein crosslinking and/or modifying e n
zyme is added to polyol-water suspension comprising from 25% to 100% (w/w)
polyol and having pH value within the range from 4.4 to 5.1 .
10 . The method according to claim 9 c h a r a c t e r i z e d in that
the method comprises the following steps:
a) pH of the polyol-water suspension is adjusted with food grade a c
id to a value within the range from 4.4 to 5.1 ,
b) at least one milk protein crosslinking and/or modifying enzyme is
added to the suspension,
c) optionally a preservative is added.
11. The method according to claim 9 c h a r a c t e r i z e d in that
the method comprises the following steps:
a) at least one milk protein crosslinking and/or modifying enzyme is
added to a polyol-water suspension comprising from 25% to 100% polyol,
b) pH of the suspension is adjusted with food grade acid(s) to a val
ue within the range from 4.4 to 5.1
c) optionally a preservative is added.
12 . The method according to any one of claims 9 to 11, c h a r a c
t e i z e d in that the polyol-water suspension comprises from 50% to 75%
polyol.
13 . The method according to any one of claims 9 to c h a r a c
t e i z e d in that the pH of the polyol-water suspension is 4.6.
14. The method according to any one of claims 9 to 13, c h a r a c
t e i z e d in that the polyol is glycerol or sorbitol.
15 . The method according to any one of claims 9 to 14 c h a r a c -
t e r i z e d in that the enzyme is transglutaminase, tyrosinase or protein glu
taminase.
16 . The method according to any one of claims 9 to 14 c h a r a c -
t e r i z e d in that the enzymes are transglutaminase and protein glutaminase.
17 . The method according to claim 15 o 16 c h a r a c t e r i z e d in
that also laccase and/or tyrosinase is added to the suspension.
18 . The method according to any one of claims 9 to c h a r a c
t e i z e d in that the method does not comprise addition of a preservative.