JP2004339356A - Method for refining cochineal coloring matter - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for refining a cochineal coloring matter having been used for coloring food or the like through efficiently removing allergenic proteins in view of the fact that one of such proteins, i.e. Coccus cacti L.( as raw material )-derived one contributing to allergy may be found in the cochineal coloring matter. <P>SOLUTION: The method for refining the cochineal coloring matter comprises adsorbing a cochineal extract liquid obtained by extracting the coloring matter from Coccus cacti L. to an adsorptive resin followed by washing the coloring matter with a hydrous alcohol adjusted to ≤pH2.5 to efficiently remove the allergenic protein contained in the extract liquid. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

【００３９】
上記実験結果より、比較例のように単に水で洗浄して得られた色素と、本発明の実施例であるｐＨ２．５以下の含水アルコールで洗浄した色素では、色素の収率の面では大きな差は生じていないが、含まれているタンパク質の含量に有意な差が生じていた。色素の収率及びタンパク質除去の効率の面から、硫酸によりｐＨ１．５に調整した１５〜１７．５％エタノール水により洗浄を行うことが好適であることが明らかとなった。
【００４０】
以上より、エンジムシから抽出されるコチニール色素抽出液に含まれる、エンジムシ由来のタンパク質を、本発明のような簡便な吸着処理を行うだけで、効率よくタンパク質を除去できることが可能となった。[0001]
BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a product orally-ingested products (foods, pharmaceuticals, etc.) and a product which comes into contact with the skin (cosmetics, etc.) by removing proteins and contaminants which may be allergens derived from the raw material, P. japonicum. The present invention relates to a cochineal pigment which is safe even if blended, a pigment preparation containing the pigment, and a purification method thereof.
[0002]
2. Description of the Related Art Cochineal pigments, which have been widely used as coloring agents for pharmaceuticals and foods, are parasitic on cactaceae, such as Nopalea coccinellifera, which are produced in desert areas in Mexico, Central America and South America. It is derived from the red pigment contained in the female body of the scale insect, Coccus cacti L .. For this reason, the cochineal colorant is prepared by extracting the dried product of the above-mentioned bark beetle as a raw material with water or alcohol.
[0003]
Recently, it has been reported that the cochineal pigment prepared from insects as described above contains contaminant proteins that are allergens and may cause allergies (Non-patented) Reference 1).
[0004]
In view of such a problem, the applicant of the present application has developed a method for removing a specific protein (protein having a molecular weight of 6,000 or more) derived from Japanese radish causing allergy from a subsequent research report, and disclosed the method (Patent Reference 1).
[0005]
[Patent Document 1] WO 02/22743 A1
[Non-Patent Document 1] Lizaso, M .; T. et al. , Ann. Allergy Asthma Immunol. , 84 (5), 549-552 (2000).
[0006]
However, in the above-mentioned prior art, a large number of steps are required to remove proteins, so that the production time is long and equipment is required.
[0007]
For this reason, there has been a need for a method capable of efficiently removing the protein serving as an allergen contained in the cochineal dye without using any special equipment and efficiently in terms of time.
[0008]
Means for Solving the Problems The inventors of the present invention have made intensive studies to solve the above-mentioned problems. As a result, the cochineal dye was adsorbed on the adsorbing resin, and the acid was strongly acidic, preferably pH 2.5 or less. More preferably, it was found that the protein could be effectively removed by washing with a hydrous alcohol having a pH of 2.0 or less.
[0009]
In the purification method disclosed in WO 02/22743 A1 by the present applicant, a step of using an adsorption resin as a means of removing proteins has been mentioned. However, such a step is different from that of the present invention in that a cochineal dye is used. It is adsorbed, desorbed immediately after washing with water, and no mention is made of washing with strongly acidic hydrous alcohol. From this, it is apparent that the present invention is a completely different purification method from the invention described in WO 02/22743 A1.
[0010]
BEST MODE FOR CARRYING OUT THE INVENTION
The present invention is a cochineal pigment characterized by being substantially free of proteins including those that become allergens derived from the worms used as raw materials.
[0011]
The cochineal dye is generally a red dye mainly containing an anthraquinone dye, carminic acid, and is prepared from the scale insect, Coccus cacti L., as described above.
[0012]
The cochineal dye of the present invention can be obtained by adsorbing a cochineal extract obtained by subjecting a cochlea extract to a cochineal extract obtained by extracting with a suitable solvent to an adsorption resin, washing with a strongly acidic aqueous alcohol and desorbing the cochineal extract. Can be.
[0013]
The proteolytic treatment can be usually performed using an enzyme. The enzyme used in the proteolytic treatment is not particularly limited as long as it has an action of decomposing proteins or peptides. For simplicity, a commercially available protease or peptidase may be used. Proteolytic treatment with an enzyme can be performed under conditions suitable for each enzyme used. For example, when the enzyme treatment under acidic conditions, the above cochineal extract 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. Therefore, it is preferable to adjust the pH to 3 to 5, preferably 3.5 to 4.5. Further, the temperature conditions for performing the protein decomposition treatment are not particularly limited, but the temperature can be generally in the range of 30 to 60 ° C.
[0014]
The protein-decomposed solution thus obtained is subjected to adsorption treatment after removing precipitates generated by filtration, co-precipitation or centrifugal separation, if necessary, as it is or after concentration.
[0015]
When the precipitate is filtered, for example, it can be performed using a filter aid such as diatomaceous earth or celite, and by using these filter aids, it is possible to remove fine and difficult to filter. it can.
[0016]
The adsorption treatment can be performed according to a conventional method, for example, an adsorption resin using a carrier such as activated carbon, silica gel or porous ceramic; styrene-based Duolite S-861 (trademark, Duolite, manufactured by USA Diamond Shamrock Co., Ltd.); The same applies hereinafter), Duolite S-862, Duolite S-863 or Duolite S-866; aromatic Sepabeads SP700 (trademark, manufactured by Mitsubishi Chemical Corporation, the same applies hereinafter), Sepabeads SP825; Diaion HP10 (trademark) Manufactured by Mitsubishi Chemical Corporation, hereinafter the same), Diaion HP20, Diaion HP21, Diaion HP40 and Diaion HP50; or Amberlite XAD-4 (trademark, manufactured by Organo, the same applies hereinafter), Amberlite XAD-7, Amberlite XAD-1180, Amberra It can be mentioned adsorption treatment using an adsorbent resin carrier such as preparative XAD-2000.
[0017]
The adsorption treatment is not limited, but is preferably performed under acidic conditions. Specifically, it is preferable that the above-mentioned inorganic acid or organic acid is added to the above-mentioned protein decomposition treatment liquid, which is a sample to be subjected to the adsorption treatment, to adjust the pH to about 2 to 4, preferably about pH 3.
[0018]
The protein degradation treatment solution containing the cochineal dye is subjected to an adsorption treatment, whereby the cochineal dye is adsorbed on a carrier such as the adsorption resin. Then, by washing with a strongly acidic aqueous alcohol, which is the production method according to the present invention, the contained proteins and proteins / peptides that have been degraded by the enzymatic treatment and reduced in molecular weight are removed. The hydrous alcohol adjusted to the strong acidity can be appropriately adjusted depending on the type of the adsorption resin to be used, but is preferably adjusted to pH 3 or less, preferably pH 2.5 or less.
[0019]
The strongly acidic hydrous alcohol used in the purification method of the present invention may be one containing 5 to 20% by weight, preferably 10 to 18% by weight of alcohol. Examples of the alcohol include lower alcohols having 1 to 4 carbon atoms such as methanol, ethanol, propanol, isopropyl alcohol and butanol. Preferably, it is ethanol. The alcohol concentration of the hydrated alcohol can be appropriately changed and used depending on the type and properties of the resin used as a carrier in the cochineal dye adsorption treatment. On the other hand, sulfuric acid, hydrochloric acid, etc. may be used without limitation as long as the pH becomes 2.5 or less, such as sulfuric acid, hydrochloric acid, etc., in order to make the pH of the hydrated alcohol strongly acidic, preferably pH 2.5 or less. it can.
[0020]
Next, the cochineal dye adsorbed on the carrier for adsorption is eluted from the carrier with a suitable solvent such as, for example, aqueous alcohol. In addition, before performing such elution treatment, it is preferable to wash with a suitable solvent such as water, for example, in which the cochineal dye does not desorb from the carrier for adsorption.
[0021]
In this case, the solvent for the elution treatment preferably has a pH of about 7 to 9, and the pH can be adjusted with, for example, trisodium citrate, sodium hydroxide, sodium carbonate, sodium hydrogen carbonate, or the like.
[0022]
Further, even if the liquid after the above-mentioned adsorption treatment is subjected to the adsorption treatment again, it can be subjected to various treatments such as an ion exchange treatment, a membrane treatment or an acid treatment.
[0023]
The ion exchange treatment is not particularly limited, and a treatment with a cation exchange resin or an anion exchange resin can be performed according to a conventional method using a conventional ion exchange resin.
[0024]
Further, the membrane treatment in the present invention broadly means a filtration treatment by a membrane, for example, a membrane filter (MF) filtration membrane, an ultra filter (UF) filtration membrane, a nano filter (NF) filtration membrane, a reverse osmosis. (Reverse osmosis (RO)) filtration treatment using a functional polymer membrane such as a filtration membrane and an electrodialysis membrane. As membrane treatment methods, in addition to the methods using these membranes, a dialysis method using a concentration gradient by an ion separation membrane, an electrodialysis method in which a voltage is applied using an ion exchange membrane as a diaphragm, and the like are known. ing.
[0025]
Further, the acid treatment can be carried out by adjusting the pH of the cochineal extract solution after proteolysis or the solution after the adsorption treatment to pH 1 to 6, preferably 1.5 to 4, and exposing it to acidic conditions. Specifically, it can be easily carried out by adding an acid to the treatment liquid. The acid is not particularly limited as long as it is an acid that can be generally used as a food additive, and any acid can be selected from the acids. The combination of acids and temperature conditions are not particularly limited.
[0026]
Further, as an optional treatment, a treatment in which carbon dioxide gas, ethylene, propane, or the like is brought into contact with the proteolysis-treated solution or the adsorption-treated solution of the cochineal extract in a closed system at a temperature and pressure higher than the critical point can be performed.
[0027]
Each of the above-mentioned treatments may be performed singly on the proteolysis-treated solution or the adsorption-treated solution of the cochineal extract, or may be performed by combining two or more kinds of treatments in any order. The processing may be repeatedly performed under the same or different conditions.
[0028]
Thus, the cochineal pigment obtained by the production method of the present invention is a food, drug, or medicine which is obtained by effectively removing the protein derived from the locust beetle, which is an allergen, and is orally ingested thereby. A cochineal pigment that does not induce allergic symptoms even when incorporated in quasi-drugs and skin-contact products (cosmetics) can be provided. Further, the cochineal dye obtained by the production method of the present invention hardly generates a precipitate over time, and has excellent storage stability.
[0029]
Incidentally, the cochineal dye mainly means carminic acid.In the category of the cochineal dye of the present invention, in addition to the carminic acid, a metal ion such as an aluminum ion or a calcium ion binds to carminic acid. (Including complexes and polymers thereof). Specific examples of carmine include a complex of carminic acid with an aluminum ion and a polymer thereof (aluminum lake) or a complex of carminic acid with a calcium ion and a polymer thereof (or calcium lake).
[0030]
Thus, the cochineal dye obtained from the present invention can be prepared as a dye composition by further mixing a food hygiene or pharmaceutically acceptable carrier or additive. Such pigment compositions are useful for coloring foods, including beverages, pharmaceuticals, quasi-drugs, or cosmetics, particularly for coloring orally ingestible or skin-contacting products (cosmetics). It can be suitably used as a pigment preparation to be used.
[0031]
The form of the dye preparation is not particularly limited. For example, water, an alcohol such as ethanol or another solvent (eg, propylene glycol, glycerin, etc.) as a carrier, and a cochineal dye dissolved or dispersed therein, or Dextrin, lactose, powdered starch syrup, etc. are used as carriers (excipients) and are mixed with cochineal pigments to form powder, granules, tablets or pills in a dry (solid) state. It can be prepared as a pigment preparation.
[0032]
The additives to be added to the pigment preparation are not particularly limited, but preservatives (such as sodium acetate and protamine) and stabilizers (eg, gum arabic, gellan gum, carrageenan, trehalose, water-soluble hemicellulose) which are usually used in pigments and pigment preparations , Sodium phosphate, sodium metaphosphate, etc.), emulsifier (lecithin, sucrose fatty acid ester, polyglycerin fatty acid ester, etc.) or antioxidant (tea extract, green coffee extract, sunflower seed extract, enzyme-treated rutin, rutin Food additives such as degradation products (quercetin), enzyme-treated isoquercitrin, tocopherols, ascorbic acids, and the like.
[0033]
The present invention will be specifically described below with reference to the following examples and comparative examples, but the present invention is not limited to these examples.
[0034]
Example 1
Purification of cochineal pigment To 35 kg of dried powder of Echinacea purpurea, 800 L of water adjusted to pH 7.5 with caustic soda was added, and the cochineal pigment was extracted while stirring at 90 to 100 ° C for 20 to 30 minutes. After the extraction, citric acid was added to the liquid (cochineal extract) that had been subjected to solid-liquid separation with a 60-mesh wire net, the cochineal extract was adjusted to pH 4 and cooled to about 50 ° C., and then 0.1% of protease was added. Then, the mixture was stirred at the above temperature for 3 hours. Next, after adjusting the pH of the obtained protein decomposition treatment solution to 3 with citric acid, a filter aid and diatomaceous earth were mixed, and the mixture was filtered through filter paper to obtain about 250 L of a protein decomposition solution. The cochineal proteolysis solution (color value: 7.81, pH 3.0, 256 g) was passed through an adsorbent resin Amberlite XAD-1180 (50 mL) filled in a resin tower to adsorb the cochineal dye, and then the resin was washed. Washing was performed with 200 mL of ethanol water having a concentration shown in the following table adjusted to pH 1.5 with sulfuric acid. Next, after washing with 200 mL of water, the cochineal dye was eluted using 40 mL of a 20% ethanol solution (pH 8.3) of 1.65% by weight of trisodium citrate, 40 mL of 20% ethanol, and ion-exchanged water. Obtained.
[0035]
The adsorption / desorption treatment was repeated, and the obtained eluate was concentrated under reduced pressure at 40 ° C., the dye yield was measured, and the protein content was measured by the semi-micro Kjeldahl method.
[0036]
As a comparative example, a comparative example using the same amount of ion-exchanged water as the cleaning solution after the adsorption was used.
[0037]
Here, the color value means the absorbance at the maximum absorption wavelength (around 495 nm) in the visible part of a solution (0.1 N hydrochloric acid aqueous solution) containing the subject cochineal dye, and the absorbance is measured at 10 w / v%. It is a numerical value converted into the absorbance of the solution.
[0038]
result

[0039]
From the above experimental results, the dye obtained by simply washing with water as in the comparative example, and the dye washed with aqueous alcohol having a pH of 2.5 or less, which is an example of the present invention, have a large dye yield. No difference was found, but a significant difference was found in the contained protein content. From the viewpoint of the yield of the dye and the efficiency of removing the protein, it was revealed that washing with 15 to 17.5% ethanol water adjusted to pH 1.5 with sulfuric acid was suitable.
[0040]
From the above, it became possible to efficiently remove proteins from the worms contained in the cochineal pigment extract extracted from the worms only by performing a simple adsorption treatment as in the present invention.

Claims (3)

In the step of purifying the cochineal dye, which is characterized by performing the adsorption resin treatment, after the adsorption treatment to the adsorption resin, the cochineal dye is desorbed by washing with a strongly acidic aqueous alcohol. A method for purifying a cochineal dye, which is a feature. The method for purifying a cochineal dye according to claim 1, wherein the pH of the hydroalcohol adjusted to a strong acidity is 2.5 or less. 3. The method for purifying a cochineal dye according to claim 1, wherein the aqueous alcohol is a 5 to 20% by weight ethanol solution.