U.S. patent application number 09/831379 was filed with the patent office on 2003-01-02 for purified cochineal and method for its production.
Invention is credited to Ichi, Takahito, Koda, Takatoshi, Sakata, Makoto, Sato, Hiroyuki, Yukawa, Chiyoki.
Application Number | 20030003201 09/831379 |
Document ID | / |
Family ID | 18760333 |
Filed Date | 2003-01-02 |
United States Patent
Application |
20030003201 |
Kind Code |
A1 |
Ichi, Takahito ; et
al. |
January 2, 2003 |
Purified cochineal and method for its production
Abstract
The invention provides a cochineal color, a colorant product
containing the cochineal color and the method of its production,
wherein the cochineal color is free of allergenic proteins and
contaminants derived from the raw material Coccus cacti L. insect
and can be added with safety to products taken by mouth, such as
foods, pharmaceutical products, etc. The invention relates to a
cochineal color substantially free of proteins of molecular weight
not less than 6000. The invention also relates to a process for
producing a cochineal color which comprises subjecting a cochineal
extract solution to proteolysis and then to at least one treatment
selected from the group consisting of adsorption treatment, ion
exchange treatment, acid treatment, extraction treatment and
membrane treatment.
Inventors: |
Ichi, Takahito; (Osaka,
JP) ; Koda, Takatoshi; (Osaka, JP) ; Yukawa,
Chiyoki; (Osaka, JP) ; Sakata, Makoto; (Osaka,
JP) ; Sato, Hiroyuki; (Osaka, JP) |
Correspondence
Address: |
ARMSTRONG,WESTERMAN & HATTORI, LLP
1725 K STREET, NW.
SUITE 1000
WASHINGTON
DC
20006
US
|
Family ID: |
18760333 |
Appl. No.: |
09/831379 |
Filed: |
May 17, 2001 |
PCT Filed: |
March 23, 2001 |
PCT NO: |
PCT/JP01/02310 |
Current U.S.
Class: |
426/262 |
Current CPC
Class: |
C09B 61/00 20130101 |
Class at
Publication: |
426/262 |
International
Class: |
A23L 001/27 |
Foreign Application Data
Date |
Code |
Application Number |
Sep 11, 2000 |
JP |
2000-274596 |
Claims
1. A purrified cochineal color characterized by its being
substantially free of the allergen derived from the insect Coccus
cacti L.
2. A purrified cochineal color as claimed in claim 1 further
characterized in that it is substantially free of proteins having
molecular weights not less than 6000.
3. A color composition comprising the purrified cochineal color
defined in claim 1 and a carrier or additive which is food
sanitation-wise or pharmaceutically acceptable.
4. A color composition as claimed in claim 3 wherein the cochineal
color is at least one member selected from the group consisting of
carminic acid, carminic acid aluminum lake, carminic acid calcium
lake, and a polymer of said aluminum lake or calcium lake.
5. A colorant product for food, pharmaceutical, quasi-drug or
cosmetic use which comprises the color composition defined in claim
3.
6. A process for producing a cochineal color which comprises
subjecting a cochineal extract solution to proteolysis and removing
a fraction of molecular weight not less than 6000.
7. A process for producing a cochineal color which comprises
subjecting a cochineal extract solution to proteolysis and, then,
to at least one treatment selected from the group consisting of
adsorption treatment, ion exchange treatment, acid treatment and
membrane treatment.
8. A process for producing a cochineal color which comprises
subjecting a cochineal extract solution to proteolysis and
adsorption treatment and, then, to at least one treatment selected
from the group consisting of adsorption treatment, ion exchange
treatment, acid treatment and membrane treatment.
9. A process for producing a cochineal color as claimed in claim 6
wherein the cochineal extract solution is an extracted product
obtained by extracting Coccus cacti L. insect bodies with use of an
aqueous alcohol.
10. A process for producing a cochineal color as claimed in claim 6
wherein the cochineal extract solution is subjected to proteolysis
under an acidic condition.
11. A process for producing a cochineal color as claimed in claim 7
or 8 wherein the adsorption treatment is carried out under an
acidic condition.
12. A process for producing a cochineal color as claimed in claim 7
or 8 wherein the adsorption treatment comprises causing a cochineal
color to be adsorbed on an adsorbent under an acidic condition and,
then, desorbed with an aqueous alcohol under a condition of pH
7.about.9.
13. A process for producing a cochineal color as claimed in claim 7
or 8 wherein the acid treatment is conducted with an acid used as a
food additive.
14. A process for producing a cochineal color as claimed in claim 7
or 8 wherein the membrane treatment is at least one treatment
selected from the group consisting of reverse osmosis membrane
treatment, membrane filter treatment, ultrafiltration membrane
treatment and nanofiltration membrane treatment.
15. A process for producing a cochineal color as claimed in claim 7
or 8 wherein the membrane treatment is a treatment using a membrane
having a cut-off molecular weight of 2000.about.8000.
16. A process for producing a cochineal color as claimed in claim 7
or 8 wherein a fraction of molecular weight not less than 6000 is
removed by the membrane treatment.
Description
TECHNICAL FIELD
[0001] This invention relates to the cochineal color characterized
by its being so pure as to be substantially allergen-free and to a
coloring composition comprising the color. Furthermore, this
invention relates to a method of producing said cochineal color of
high purity and free of allergens.
BACKGROUND ART
[0002] Cochineal in broad use as a coloring agent, for example in
pharmaceutical products and foods, is derived from the red coloring
matter occurring in the female insect bodies of Coccus cacti L.
which grows on Nopalea coccinellifera and other plants of the
family Cactaceae cultivated in the desert areas of Mexico and
Central and South America. As such, cochineal is harvested from
said insect by extracting its dried bodies with water or
alcohol.
[0003] It has been reported, of late, that being an insect-derived
substance, cochineal contains allergenic impurity proteins and as
such may be an etiologic factor in allergic diseases (Ann Allergy
Asthma, Vol. 84(5), 549-552, 2000.).
[0004] Further, the use of cochineal colors produced by
conventional methods poses problems that the types and applications
of the products, and the usage concentration of the color are
limited due to the odors probably attributable to the above raw
material. Another problem is also pointed out that the conventional
cochineal colors tend to form a sediment with time since they
contain proteins and like contaminants derived from the above raw
material.
DISCLOSURE OF INVENTION
[0005] The inventors of this invention made an intensive
investigation to overcome the above allergenicity problem of the
conventional cochineal colors and have come to develop the
cochineal colors defined below.
[0006] This invention, therefore, is directed to the following
purified cochineal colors (1) and (2).
[0007] (1) A purified cochineal color characterized by its being
substantially free of the C. cacti L. insect-derived allergens.
[0008] (2) A purified cochineal color as defined in paragraph (1)
further characterized by its being substantially free of proteins
of molecular weight not less than 6000.
[0009] This invention is further directed to the following color
compositions and associated colorant product (3).about.(5).
[0010] (3) A color composition comprising the purified cochineal
color defined in paragraph (1) and a food sanitation-wise or
pharmaceutically acceptable carrier or additive.
[0011] (4) A color composition as defined in paragraph (3) wherein
the cochineal color is at least one member selected from the group
consisting of carminic acid, carminic acid aluminum lake, carminic
acid calcium lake, and a polymer of carminic acid aluminum lake or
carminic acid calcium lake.
[0012] (5) A colorant product for application to foods,
pharmaceutical preparations, quasi-drug preparations or cosmetic
products which comprises the color composition defined in paragraph
(3).
[0013] The invention is further directed to the following processes
(6).about.(16) for producing a cochineal color.
[0014] (6) A process for producing a cochineal color which
comprises subjecting a cochineal extract solution to proteolysis
and removing a fraction of molecular weight not less than 6000.
[0015] (7) A process for producing a cochineal color which
comprises subjecting the cochineal extract solution to proteolysis
and then to at least one treatment selected from the group
consisting of adsorption treatment, ion exchange treatment, acid
treatment and membrane treatment.
[0016] (8) A process for producing a cochineal color which
comprises subjecting the cochineal extract solution to proteolysis
and adsorption treatment and then to at least one treatment
selected from the group consisting of adsorption treatment, ion
exchange treatment, acid treatment and membrane treatment.
[0017] (9) A process for producing a cochineal color as defined in
paragraph (6) wherein the cochineal extract solution is an
extracted product obtained from the Coccus cactus L. insect with
use of an aqueous alcohol.
[0018] (10) A process for producing a cochineal color as defined in
paragraph (6) wherein the cochineal extract solution is subjected
to proteolysis under an acidic condition.
[0019] (11) A process for producing a cochineal color as defined in
paragraph (7) or (8) wherein the adsorption treatment is carried
out under an acidic condition.
[0020] (12) A process for producing a cochineal color as defined in
paragraph (7) or (8) wherein the adsorption treatment comprises
adsorbing cochineal color on an adsorbent under an acidic condition
and desorbing it with an aqueous alcohol under a condition of pH
7.about.9.
[0021] (13) A process for producing a cochineal color as defined in
paragraph (7) or (8) wherein the acid treatment is conducted with
an acid used as a food additive.
[0022] (14) A process for producing a cochineal color as defined in
paragraph (7) or (8) wherein the membrane treatment is at least one
treatment selected from the group consisting of reverse osmosis
membrane treatment, nanofiltration membrane treatment,
ultrafiltration membrane treatment and microfiltration membrane
treatment.
[0023] (15) A process for producing a cochineal color as defined in
paragraph (7) or (8) wherein the membrane treatment is at least one
treatment selected from the group consisting of reverse osmosis
(RO) membrane treatment, membrane filter (MF) treatment,
ultrafiltration (UF) membrane treatment and nanofiltration (NF)
membrane treatment.
[0024] (16) A process for producing a cochineal color as defined in
paragraph (7) or (8) wherein the membrane treatment is carried out
using a membrane having a cut-off molecular weight of
2000-8000.
[0025] (17) A process for producing a cochineal color as defined in
paragraph (7) or (8) wherein a fraction of molecular weight not
less than 6000 is removed by the membrane treatment.
BRIEF DESCRIPTION OF THE DRAWING
[0026] FIG. 1 is an SDS-polyacrylamide gel electrophoretogram
(SDS-PAGE) of the liquors in various production stages. The lanes,
from left to right, represent {circle over (1)} molecular weight
markers (16950 Da, 14410 Da, 10704 Da, 8167 Da, 6217 Da, 2512 Da),
{circle over (2)} cochineal extract solution, {circle over (3)}
protein hydrolysate, {circle over (4)} adsorption-treated solution,
{circle over (5)} MF filtrate, and {circle over (6)} NF
filtrate.
BEST MODE FOR CARRYING OUT THE INVENTION
[0027] This invention relates to a cochineal color derived from
Coccus cacti L. and characterized by its being substantially
allergen-free.
[0028] Cochineal is generally a red coloring matter composed
predominantly of carminic acid which is an anthraquinone color and,
as mentioned above, is produced by a process starting with Coccus
cacti L.
[0029] The cochineal color of the invention has been purified, for
implementing the above feature, to a degree substantially not
containing proteins of molecular weight not less than 6000.
[0030] The cochineal color of the invention as such can be produced
by a technology which comprises using a cochineal extract solution
obtained by extracting C. cacti insect bodies with a suitable
solvent, subjecting said cochineal extract solution to proteolysis
and, then, to a suitable purification treatment such as ion
exchange treatment and/or membrane treatment to remove a fraction
of molecular weight not less than 6000 from the cochineal
extract.
[0031] More particularly, a highly purified cochineal color
according to the invention can be produced and acquired by
subjecting the liquor after proteolysis of a cochineal extract
solution to any one of adsorption treatment, ion exchange
treatment, acid treatment or membrane treatment or an arbitrary
combination of any two or more of said treatments.
[0032] The cochineal extract solution for use in the present
invention can be obtained, by advantageously comminuting the dried
C. cactis L. insect bodies and extracting the powder with water, an
alcohol or an aqueous alcohol. The alcohol includes lower alcohols
containing 1.about.4 carbon atoms, such as methanol, ethanol,
propanol, isopropyl alcohol and butanol. The preferred is
ethanol.
[0033] The method of extraction may be one in routine use for
extraction. Thus, the extraction method includes but is not
restricted to a method which comprises soaking the dry powder of C.
cactis L. bodies in said solvent by a cold extraction technique or
a warm extraction technique, a method comprising extracting said
powder under warming and stirring and filtering the same, and a
percolation method. The preferred technique comprises immersing the
dry powder of C. cacti L. insect bodies in a solvent, preferably
prewarmed, for several minutes through a few hours, preferably for
tens of minutes. The pH of the extractant solvent is not
particularly restricted but is preferably neutral.about.alkaline,
specifically pH 6.about.8, preferably pH 7.about.8, more preferably
about pH 7.5. The basic compound to be used for alkalinizing the
extractant solvent can be selected from a broad range of compounds
which can be used in foods and is not particularly restricted
otherwise. As typical examples, sodium hydroxide, sodium carbonate,
sodium hydrogencarbonate and sodium citrate can be mentioned.
[0034] The cochineal extract solution obtained in the above manner
is filtered, coprecipitated or centrifuged to remove a solid
fraction where necessary and, then, subjected to proteolysis either
as it is or after concentration. The proteolysis can be generally
effected by means of an enzyme.
[0035] The enzyme for use in the proteolysis is not particularly
restricted insofar as it is capable of digesting protein or
peptides. Expediently, enzyme preparations commercially available
as proteases or peptidases can be employed. The enzymatic
proteolysis can be carried out under specific conditions suited to
the respective enzymes. For the enzymatic treatment under acidic
conditions, for instance, said cochineal extract solution is
preferably adjusted to pH 3.about.5, preferably pH 3.5.about.4.5,
using an inorganic acid such as hydrochloric acid, sulfuric acid or
phosphoric acid or an organic acid such as citric acid, malic acid,
tartaric acid, lactic acid, acetic acid or fumaric acid. The
temperature for this proteolysis is not particularly restricted but
may generally be within the range of 30.about.60.degree. C.
[0036] The protein hydrolysate thus obtained is subjected to
filtration, coprecipitation or centrifugation for removal of the
precipitate formed where necessary and, either directly or after
concentration, is further subjected to adsorption treatment, ion
exchange treatment, acid treatment or membrane treatment. The
preferred is membrane treatment.
[0037] The filtration procedure for removal of the precipitates
formed in the various stages in the process according to the
invention, inclusive of the precipitate mentioned just above, can
be carried out in the presence of a filter aid, such as
diatomaceous earth or celite, the use of which enables removal of
fine particles which are not easy to filter off.
[0038] The adsorption treatment can be carried out in the routine
manner. For example, it may be an adsorption treatment using such
an adsorbent as activated carbon, silica gel or a porous ceramic
material or an adsorption treatment using an adsorbent resin as
follows: a stirenic adsorbent resin such as Duolite S-861 (TM,
Diamond Shamrock, U.S.A.; the same applies below), Duolite S-862,
Duolite S863 and Duolite S-866; an aromatic adsorbent resin such as
Sepabeads SSP700 (TM, Mitsubishi Chemical; the same applies below),
Sepabeads SP825, Diaion HP10 (TM, Mitsubishi Chemical; the same
applies below), Diaion HP20, Diaion HP21, Diaion HP40 and Diaion
HP50, etc.; and Amberlite XAD-4 (TM, Organo; the same applies
below), Amberlite XAD-7 and Amberlite XAD-2000, etc., for
instance.
[0039] The adsorption treatment is carried out preferably, but not
essentially, under acidic conditions. To be specific, the sample to
be adsorbed is preferably adjusted to pH above 2.about.4, more
preferably pH about 3, with said inorganic or organic acid.
[0040] The cochineal color adsorbed on the adsorbent can be
recovered by desorbing (eluting) it with a suitable solvent such as
an aqueous alcohol. Preferably, prior to elution, the adsorbent is
washed with a suitable solvent, such as water, which does not cause
desorption of the cochineal color. The preferred aqueous alcohol
generally contains 10.about.60% by volume, more preferably
20.about.80% by volume, of an alcohol. The alcohol includes lower
alcohols of 1.about.4 carbon atoms, such as methanol, ethanol,
propanol, isopropyl alcohol and butanol. The preferred alcohol is
ethanol.
[0041] In this case, the desorbing solvent or eluent is preferably
neutral.about.alkaline, namely pH 7.about.9, preferably pH
8.about.9, and the adjustment to such a pH level can be made using
sodium citrate, sodium hydroxide, sodium carbonate, sodium
hydrogencarbonate or the like.
[0042] The liquor available on adsorption treatment of the
cochineal extract solution can be subjected to a further adsorption
treatment or to any of various other treatments such as ion
exchange treatment, membrane treatment and acid treatment.
[0043] This adsorption treatment may be the same treatment as
described above.
[0044] The ion exchange treatment is not particularly restricted
but includes the routine cation exchange or anion exchange
treatment using the conventional ion exchange resin. The cation
exchange resin, for instance, includes but is not restricted to
strongly cation exchange resins and weakly cation exchange resins,
such as Diaion SK1B (TM, Mitsubishi Chemical; the same applies
below), Diaion SK102, Diaion SK116, Diaion PK208, Diaion WK10 and
Diaion WK20, among others. The anion exchange resin is not
particularly restricted, either, but includes Diaion SA10A (TM,
Mitsubishi Chemical; the same applies below), Diaion SA12A, Diaion
SA20A, Diaion PA306, Diaion WA10, and Diaion WA20, among
others.
[0045] The membrane treatment in the context of the present
invention means a variety of filtration treatments using membranes,
thus including a treatment with a membrane filter (MF), a
functional high polymer membrane such as an ultrafiler (UF), a
nanofilter (NF), a reverse osmosis (RO) filter or an
electrodialysis membrane.
[0046] The known membrane treatment technology includes not only
the UF and RO membrane methods but also the dialysis treatment
utilizing the concentration gradient across an ion-selective
membrane and the electrodialysis treatment using an ion exchange
membrane with application of a voltage, among others. Industrially,
the NF membrane filtration method is preferred. The membrane
material for use in the membrane treatment method may be natural,
synthetic or semisynthetic and includes cellulose, cellulose
diacetate or triacetate, polyamides, polysulfones, polystirenes,
polyimides and polyacrylonitrile, among others.
[0047] The membrane treatment according to the invention includes a
treatment by which high molecular compounds are removed with a
membrane having a cut-off molecular weight of, for example,
10000.about.1000000 and a treatment by which low molecular
compounds are removed with a membrane having a cut-off molecular
weight of about 2000.about.8000, preferably about 6000, more
preferably about 3000.
[0048] The former technology specifically includes the UF membrane
technique using any of NTU-3150 membrane, NTU-3250 membrane,
NTU-3550 membrane and NTU-3800 UF membrane (all available from
Nitto Denko Corporation); Cefilt-UF (product of Nippon NGK
Insulators, Ltd.); AHP-2013 membrane, AHP-3013 membrane and
AHP-1010 membrane (all available from Asahi Chemical Industry),
among others. The later technology includes the reverse osmosis
membrane (NF membrane; cut-off mol. wt. ca 3000) technique using
any of NTR-7250 membrane, NTR-7410 membrane, NTR-7430 membrane and
NTR-7450 membrane (all from Nitto Denko Corporation); AIP-3013
membrane; ACP-3013 membrane, ACP-2013 membrane, AIP-2013 membrane
and AIO-1010 membrane (all available from Asahi Chemical Industry),
among others. These membranes can be used each independently or in
a suitable combination.
[0049] The acid treatment can be effected by adjusting the protein
hydrolysate of a cochineal extract solution or the liquor subjected
to the various treatments mentioned above to pH 1.about.6,
preferably pH 1.5.about.4, to thereby expose the liquor to an
acidic environment. The acid treatment can be expediently carried
out by adding an acid to said liquor. The acid to be used is not
particularly restricted insofar as it is in routine use as a food
additive and can be arbitrarily selected from among such acids. For
example, such organic acids as citric acid, acetic acid, malic
acid, lactic acid, etc. and such inorganic acids as sulfuric acid,
hydrochloric acid, phosphoric acid, nitric acid, etc. can be
employed. The preferred is a treatment using an inorganic acid
which is in routine use as a food additive.
[0050] The temperature condition for said acid treatment is not
particularly restricted but usually can be arbitrarily selected
from the range of 5.about.100.degree. C. For example, the range of
20.about.100.degree. C. or the range of 40.about.100.degree. C. may
be used. The duration of acid treatment is not particularly
restricted but usually can be arbitrarily selected from the range
of 1.about.300 minutes. Generally speaking, a shorter treatment
time suffices at a high treatment temperature and, in this sense,
the treatment time at 40.about.100.degree. C., for instance, may be
selected from the range of 5.about.60 minutes. In this treatment,
the treatment load liquor may optionally be agitated, there being
no particular rule to be complied with.
[0051] As an optional treatment, carbon dioxide gas, ethylene,
propane or the like may be contacted with the protein hydrolysate
of a cochineal extract solution or said liquor after said
adsorption treatment under supercritical temperature and pressure
conditions in a hermetically closed system.
[0052] Only one of the above treatments may be carried out or two
or more of them may be carried out in an optional combination in an
arbitrary order. Or one and the same treatment may be carried out
repeatedly under the same or varied conditions.
[0053] The preferred treatment method, though it is not
particularly restricted, comprises subjecting the cochineal extract
after proteolytic treatment to an adsorption treatment and
subjecting the liquor obtained by desorption from the adsorbent to
a membrane treatment. Furthermore, where necessary, an ion exchange
treatment may be carried out after said adsorption treatment. The
membrane treatment mentioned above is preferably an MF, UF or NF
membrane treatment, more preferably an MF or NF membrane treatment.
Furthermore, this membrane treatment is preferably carried out
using a membrane having a cut-off molecular weight of
2000.about.8000, preferably about 6000, more preferably about 3000,
to thereby remove high molecular compounds, preferably those having
molecular weights over 6000, more preferably over 3000.
[0054] The cochineal color of the present invention, thus obtained,
has been effectively deprived of various contaminants inclusive of
the C. cacti L. insect-derived proteins which are allergenic or
potentially allergenic. There can, thus, be provided a cochineal
color which, when formulated in foods, pharmaceuticals, quasi drugs
or cosmetic products which are, or are liable to be, taken by
mouth, does not induce allergic responses. Furthermore, the
cochineal color obtained by the above method of the invention
scarcely forms a precipitate with time, thus enjoying a good
shelf-life.
[0055] Incidentally, the term "cochineal color" means carminic acid
in the main but the cochineal color in the context of the invention
includes not only carminic acid but also carmine which is the
pigment (inclusive of its complexes and their polymers) formed as a
metal ion, such as aluminum or calcium ion, is complexed with
carminic acid. As the carmine, there can be mentioned such species
as the complex of carminic acid with aluminum ion, inclusive of its
polymer (aluminum lake), and the complex of carminic acid with
calcium ion, inclusive of its polymer (calcium lake).
[0056] The cochineal color of the invention, as obtained in the
above manner, can be formulated with a carrier and/or additive
which is acceptable from food sanitation points of view or
pharmaceutically acceptable and be provided in the form of a color
composition. This color composition can be used with advantage for
the colorant product to be added for the purpose of coloring foods
inclusive of drinks, pharmaceutical products, quasi drugs or
cosmetic products, particularly those products which are intended,
or are liable, to be taken by mouth.
[0057] The dosage form of the colorant product is not particularly
restricted but may be a solution or dispersion of the cochineal
color in a medium such as water, an alcohol, e.g. ethanol, or a
suitable other solvent (for example, propylene glycol, glycerol or
the like) or a dry (solid) product as prepared by formulating the
cochineal color in a carrier (excipient) such as dextrin, lactose,
dried starch syrup or the like and molding the mixture into
powders, granules, tablets or pills.
[0058] The additives which can be formulated in the colorant
product are not particularly restricted but include those food
additives which are generally used in colors and colorant products,
such as preservatives (sodium acetate, protamine, etc.),
stabilizers (gum arabic, gellan gum, carrageenin, trehalose,
water-soluble hemicellulose, sodium phosphate, sodium
metaphosphate, etc.), emulsifiers (lecithin, sucrose fatty acid
esters, polyglycerol fatty acid esters, etc.), antioxidant (tea
extract, raw coffee extract, sunflower seed extract, myrica
extract, rutin extract, rosemary extract, enzymatically treated
rutin, decomposition product of rutin (quercetin), enzymatically
treated isoquercitrin, tocopherols, ascorbic acid compounds,
etc.).
[0059] As shown in Example 3 given below, the cochineal color
significantly deodorized can be prepared by the process of the
invention. The process for producing a cochineal color of the
invention may be understood as a process for deodorizing a
cochineal color. Thus, the invention includes the following
modes.
[0060] (a). A process for deodorizing a cochineal color which
comprises subjecting a cochineal extract solution to proteolysis
and removing a fraction of molecular weight not less than 6000.
[0061] (b). A process for deodorizing a cochineal color which
comprises subjecting the cochineal extract solution to proteolysis
and then to at least one treatment selected from the group
consisting of adsorption treatment, ion exchange treatment, acid
treatment and membrane treatment.
[0062] (c). A process for deodorizing a cochineal color which
comprises subjecting the cochineal extract solution to proteolysis
and adsorption and then to at least one treatment selected from the
group consisting of adsorption treatment, ion exchange treatment,
acid treatment and membrane treatment.
[0063] (d). A process for deodorizing a cochineal color as defined
in paragraph (a) wherein the cochineal extract solution is an
extracted product obtained from the Coccus cactus L. insect with
use of an aqueous alcohol.
[0064] (e). A process for deodorizing a cochineal color as defined
in paragraph (a) wherein the cochineal extract solution is
subjected to proteolysis under an acidic condition.
[0065] (f). A process for deodorizing a cochineal color as defined
in paragraph (b) or (c) wherein the adsorption treatment is carried
out under an acidic condition.
[0066] (g). A process for deodorizing a cochineal color as defined
in paragraph (b) or (c) wherein the adsorption treatment comprises
adsorbing cochineal color on an adsorbent under an acidic condition
and desorbing it with an aqueous alcohol under a condition of pH
7.about.9.
[0067] (h). A process for deodorizing a cochineal color as defined
in paragraph (b) or (c) wherein the acid treatment is conducted
with an acid used as a food additive.
[0068] (i). A process for deodorizing a cochineal color as defined
in paragraph (b) or (c) wherein the membrane treatment is at least
one treatment selected from the group consisting of reverse osmosis
membrane treatment, membrane filter treatment, ultrafiltration
membrane treatment and nanofiltration membrane treatment.
[0069] (j). A process for deodorizing a cochineal color as defined
in paragraph (b) or (c) wherein the membrane treatment is carried
out using a membrane having a cut-off molecular weight of
2000.about.8000.
[0070] (k). A process for deodorizing a cochineal color as defined
in paragraph (b) or (c) wherein a fraction of molecular weight not
less than 6000 is removed by the membrane treatment.
EXAMPLES
[0071] The following examples illustrate the invention in further
detail. It should, however, be understood that the scope of the
invention is by no means defined by these examples.
Example 1
[0072] (1) Preparation of Cochineal Color and a Colorant
Product
[0073] To 35 kg of the dry powder of the insect Coccus cacti L. was
added 800 L of water adjusted to pH 7.5 with sodium hydroxide and
the cochineal color was extracted at 90.about.100.degree. C. with
stirring for 20.about.30 minutes. The extract solution was passed
through a 60-mesh metal sieve for solid-liquid separation and the
liquid phase (cochineal extract solution) was adjusted to pH 4 with
citric acid and cooled to about 50.degree. C. At this stage, 0.1%
of protease was added and the mixture was stirred at that
temperature for 3 hours. Then, the protein hydrolysate obtained was
adjusted to pH 3 with citric acid, mixed with the filter aid and
diatomaceous earth, and filtered through filter paper to recover
about 250 L of the protein hydrolysate. This cochineal protein
hydrolysate was run onto a column packed with the adsorbent resin
Amberlite XAD-7 (150 L) to adsorb cochineal color. After the resin
column was washed with 8 volumes of water, the cochineal color was
eluted out with 130 L of 50% aqueous solution of ethanol (pH 7.5)
to collect 82 L of eluate. Then, this adsorption-treated fluid was
treated with an MF membrane (a ceramic filter, 0.2 .mu.m; NGK
Insulators) at 2 kg/cm.sup.2 and 20.degree. C. to recover 82 L of
MF filtrate.
[0074] Further, a treatment using an NF membrane (CF30-S, cut-off
mol. wt.=6000; Nitto Denko) was carried out at 3 kg/cm.sup.2 and
20.degree. C. to give 80 L of NF filtrate. This NF filtrate was
concentrated under reduced pressure to give 20 kg of a
significantly purified color solution with a color value of
E.sup.10%.sub.1 cm=200. To 20 kg of this color solution were added
32.4 kg of water, 13 kg of ethyl alcohol and 600 g of citric acid
(crystals) to give 66 kg of a cochineal colorant product with a
color value of E.sup.10%.sub.1 cm=60.
[0075] The color value (E.sup.10%.sub.1 cm) mentioned above is a
value found by measuring the absorbance of a solution of the
objective cochineal color (0.1 N HCl) at the maximum absorption
wavelength (ca 410 nm) in the visible region of the spectrum and
converting the absorbance value to the absorbance of a 10 w/v %
solution.
[0076] (2) Analysis by SDS-Polyacrylamide Gel Electrophoresis
(SDS-PAGE)
[0077] If the treated solution obtained in each production stage
(cochineal extract solution, protein hydrolysate,
adsorption-treated solution, MF filtrate or NF filtrate) is
directly electrophoresed, a protein analysis can hardly be made
owing to the influence of pigments, salts, etc. occurring in the
cochineal color. Therefore, using a liquid chromatograph, the
protein alone was selectively separated from each treated solution
and analyzed by electrophoresis. Thus, 5 .mu.l of each solution
with a color value of E.sup.10%.sub.1 cm=80 was run onto a Symmetry
C18: ODS column (.phi. 0.46 mm.times.250 mm) under the conditions
of developer solvent: 70% MeOH, flow rate: 1 ml/min. and detection
wavelength: 280 nm to separate pigments and salts from the protein
and the protein fraction alone was harvested. This treatment was
repeated 10 times and the protein fractions were pooled,
concentrated to dryness, dissolved in 100 .mu.l of water to prepare
a sample and subjected to electrophoresis. The electrophoresis was
performed according to the Laemmli method (Nature, 227, 680 (1970))
and the gel was silver-stained in the routine manner. The sample
for electrophoresis was prepared by mixing 105 .mu.l of the above
sample with 10 .mu.l of 2-mercaptoethanol, 25 .mu.l of 0.5 M
Tris-HCl (pH 6.8), 40 .mu.l of 10 wt. % SDS and 20 .mu.l of 70 wt.
% glycerin and boiling the mixture for 3 minutes.
[0078] The conditions of SDS-PAGE were as follows.
[0079] Sample for assay: 10 .mu.l/lane
[0080] Buffer: Tris-glycine (pH 6.8), 0.1% SDS
[0081] The results of electrophoresis are shown in FIG. 1. In FIG.
1, the lanes from left to right represent {circle over (1)}
Molecular weight markers (16950 Da, 14410 Da, 10704 Da, 8167 Da,
6217 Da, 2512 Da), {circle over (2)} cochineal extract solution ,
{circle over (3)} protein hydrolysate, {circle over (4)}
adsorption-treated solution, {circle over (5)} MF filtrate, and
{circle over (6)} NF filtrate.
[0082] Recently, M. T. Lizaso et al. have reported that the
allergens occurring in the cochineal color are proteins with
molecular weights of 17000. 28000 and 50000 (Ann Allergy Asthma,
Vol. 84(5), 549-552 (2000)). While it is generally acknowledged
that proteins inducing allergic responses are comparatively large
proteins with molecular weights over 10000, it has also been
reported that a protein may become an allergen on coupling to
another component.
[0083] If only from the results shown in FIG. 1, it is apparent
that, in accordance with the production technology of the
invention, not only said proteins having molecular weights of
17000, 28000 and 50000 but also proteins having molecular weights
of 6000 and up, which are potential allergens, can be eliminated to
the extent not detectable by silver staining (below about 100 ppb)
any longer. Furthermore, by the production technology of the
invention, the sample after NF treatment is already so pure that
substantially no protein can be detected therein, indicating that
the cochineal color with an extremely high safety rating can be
obtained.
Example 2
[0084] To 24 kg of the cochineal colorant product with a color
value of E.sup.10%.sub.1 cm=60 as prepared in Example 1 was added
156 kg of water, and 8 kg of tartaric acid, 9.3 kg of burnt alum
and 0.6 kg of slaked lime (calcium hydroxide) were dissolved in the
resulting color liquor (pigment solution). Then, under stirring,
the temperature was raised to 80.degree. C. and held at this level
for 5 hours, whereby a water-insoluble cochineal aluminum lake
(carmine) was obtained. This aqueous suspension of carmine was
filtered with a filter press (Yabuta Co.) to recover a carmine
cake, which was then dried in vacuo at 50.degree. C. and crushed to
prepare a carmine powder.
[0085] Protein analysis of this carmine by the same method of
electrophoresis as in Example 1 showed no evidence of protein. It
was, therefore, clear that a carmine substantially free of proteins
as potential allergens can be prepared by the method described in
Example 1.
[0086] In this connection, according to the findings obtained by
the inventor of the present invention, when a carminic acid
aluminum lake or calcium lake (carmine) is caused to form a
polymer, there is the tendency that when a low molecular protein is
allowed to be present, a more neat lake can be obtained with an
increased intensity of red color. Therefore, a cochineal color
(carmine) with a higher intensity of redness and an increase market
value may be prepared by adding a protein of comparatively low
molecular weight which does not become an allergen to the above
allergen-free color solution.
Example 3
[0087] The cochineal colorant product with a color value of
E.sup.10%.sub.1 cm=60 prepared in Example 1 was examined for the
presence of an odor. More specifically, 5 g of the cochineal color
product was diluted with water to thereby prepare 1 L of a solution
with a color value of E.sup.10%.sub.1 cm=0.3. The resulting
solution was heated to the temperature of 50.degree. C. and then
evaluated for its odor based on the judgments made by a panel of 10
highly-trained specialists. As a comparative sample for the
evaluation were used a cochineal extract (pH 7.5) obtained from the
dry powder of the insect Coccus cacti L. by a water extraction
following the procedures of Example 1 (comparative color 1) and a
cochineal color prepared by further subjecting the cochineal
extract obtained by the water extraction to resin adsorption
treatment (comparative color 2). The comparative samples were used
for the evaluation of odors after diluted in the same manner as
described above to prepare a solution with a color value of
E.sup.10%.sub.1 cm=0.3 and then heated to the temperature of
50.degree. C. The results are shown in Table 1.
1 TABLE 1 Purified cochineal color Comparative Comparative
Evalutation product color 1 color 2 + + + 0 7 3 + + 0 3 6 + 0 0 1
.+-. 2 0 0 - 8 0 0 (Each number in Table 1 shows the number of
panelists who selected the grades according to the below
"Evaluation Criteria".)
[0088] <Evaluation Criteria>
[0089] +++: An odor derived from the insect Coccus cacti L. is
strongly perceived.
[0090] ++: An odor derived from the insect Coccus cacti L. is
perceived.
[0091] +: An odor derived from the insect Coccus cacti L. is
faintly perceived.
[0092] .+-.: An odor derived from the insect Coccus cacti L. is
hardly perceived.
[0093] -: An odor derived from the insect Coccus cacti L. is not
perceived.
[0094] As is apparent from the results, the cochineal color of the
invention is odorless or odorous in a degree hardly perceivable,
and the cochineal color significantly deodorized can be prepared by
the process of the invention.
Example 4
[0095] The cochineal colorant product with a color value of
E.sup.10%.sub.1 cm=60 prepared in Example 1 was examined for the
formation of sediment with time. Specifically, the cochineal
colorant product was preserved in dissolved state at the
temperature of 5.degree. C. for six months and then visually
observed for the sedimentation. As the result, the colorant product
of the invention formed no sediment and was stable for a long
period.
[0096] The result shows that the cochineal color significantly
inhibited from settling during a long-term storage can be prepared
by the process of the invention.
INDUSTRIAL APPLICABILITY
[0097] The cochineal color according to this invention is a
substantially allergen-free, safe color which has been highly
purified to remove the protein and other contaminants derived from
the raw material Coccus cacti L. insect. Therefore, this color and
the colorant product containing the color according to the
invention can be used without fear of risks for allergic responses
as the coloring matter for products which are, or are liable to be,
taken by mouth, such as beverages and other foods, pharmaceutical
products, quasi drugs and cosmetic products.
[0098] Further, the cochineal color of the invention is
significantly deodorized to a degree hardly perceivable, so that
the color can be used for coloring products such as foods,
cosmetics, etc. of which commercial values are influenced by their
odor.
[0099] Furthermore, the cochineal color according to the invention
is least liable to form a precipitate with time so that it can
evenly color various products, such as drinks, foods including
confectionery, etc. without the need for filtration in advance of
use.
* * * * *