JPH0829246B2 - Microcapsule manufacturing method - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION (A) Field of Industrial Application The present invention relates to a method for producing microcapsules having yeast as a microcapsule film.

(B) Prior Art A microcapsule is a microcapsule that contains a liquid, a solid, or a gas as fine particles having a size of 1 μm to several hundreds of μm and is uniformly covered with a thin film. Also, microcapsules of colored dyes, pharmaceuticals, agricultural chemicals, fragrances, feeds, etc. are industrially commercialized.

The most common of these is its application to pressure-sensitive copying paper. That is, each of the upper paper coated with microcapsules containing a hydrophobic liquid in which a colorless electron-donating dye is dissolved on the back surface of the support and the lower paper coated with a colorless electron-accepting developer on the surface of another support When the writing pressure is applied, the microcapsules will be destroyed and the inclusions will be released, and the coloring agent and developer will come into contact and a chemical reaction will cause the coloring substance to reach the surface of the lower paper. It is formed, and this is what is obtained as a copy image.

In this way, a microcapsule can contain a substance with a certain property by forming a thin film on the outside of the substance at the same time, and the encapsulated substance can be taken out if the film is destroyed when necessary. Is.

Conventionally known methods for producing microcapsules include: (1) Coacervation method using gelatin (US Pat. Nos. 2,800,457 and 2,800,458 etc.) (2) Forming a film from the outer phase (aqueous phase) In situ method (Japanese Patent Publication Nos. 36-9168 and 47-23165, JP-A-48-57892)
No. 51-9079, No. 54-25277, etc.) (3) An interfacial polymerization method utilizing a film-forming reaction between an inner phase and an outer phase is known as a powerful method.

Further, in US Pat. No. 4,001,480, the lipid content is 40 to 6
A method of encapsulating the lipid-soluble substance in 0% fungi has been introduced.

Furthermore, in JP-A-58-107189, as a method for increasing the lipid content of growing microorganisms, the lipid content recovered from the medium is
Select from organic substances (eg, aliphatic alcohols, esters, aromatic hydrocarbons, hydrogenated aromatic hydrocarbons) for increasing lipid content in growing microorganisms of 10 wt% or more (eg, oily yeast, brewer's yeast, etc.) A microbial capsule consisting of enclosing a liquid).

(C) Problems to be Solved by the Invention In the encapsulation method described above, microcapsules having a dense film excellent in protective power of the inclusion are obtained and widely applied industrially, It is also true that it has a number of problems regarding manufacturing. That is, in the coacervation method (1), the pH, temperature, and time operations involved in the reaction are complicated. There is a problem that the encapsulation process requires a long time. (2) in situ
The method (3) and the interfacial polymerization method (3) have a drawback that they are not suitable for encapsulation of an unstable substance or a substance that is easily thermally denatured because a highly reactive coating substrate is reacted at a relatively high temperature. Have Although an encapsulation method using microorganisms has also been reported, this encapsulation method has a mild intake condition for inclusions and can be relatively easily operated, but it is included in a certain amount of membrane material (cell amount). The amount of inclusions is extremely small, and it is more difficult than the conventional microencapsulation method to include more liquid in less bacterial cells.

However, when a hydrophobic liquid having a capacity higher than the capacity of the added cells is added to the yeast dispersion liquid, free inclusions, that is, non-capsules are formed, and the function as microcapsules is remarkably reduced.

It is an object of the present invention to provide an encapsulation method using a microorganism, and more particularly, a method using a yeast to allow a larger amount of hydrophobic liquid to be contained in the yeast cell.

(D) Means for Solving the Problems The present invention relates to a method for producing a microcapsule in which yeast cells are used as a microcapsule film and more hydrophobic liquid is contained in the cells, and the following 4 It consists of a process of stages.

1) Preparation process of yeast dispersion liquid 2) Addition process of alcohols to yeast dispersion liquid 3) Process of preparing hydrophobic liquid to be incorporated and adding it to yeast dispersion liquid 4) Capsule accompanied by heating and stirring In the present invention, in order to solve the problems of the conventional microencapsulation method using microorganisms, the addition process of alcohols described in 2) above is incorporated.

More specifically, in a method for obtaining a hydrophobic liquid microcapsule having yeast as a film by adding a hydrophobic liquid to be incorporated in a yeast dispersion liquid, in the microencapsulation process, the number of carbon atoms is 1 to 3 The addition of monohydric alcohols makes it possible to include a large amount of hydrophobic liquid in yeast cells.

In the process of the above four steps, it does not matter which of the operations 2) and 3) is performed first or simultaneously.

That is, the following three types can be considered when proceeding the processes of 2) and 3).

A) After adding alcohols to the yeast dispersion, the prepared hydrophobic liquid is added.

B) After adding the prepared hydrophobic liquid to the yeast dispersion liquid, alcohols are added.

C) Add a mixture of alcohol and hydrophobic liquid to the yeast dispersion.

However, the effects of the present invention are similarly exhibited by any of the methods, and the addition method is not limited. Also, depending on the type of yeast, by maintaining an appropriate nutrient source and environment,
Some produce ethyl alcohol (so-called alcohol fermentation). Usually 1% to 10% of yeast dispersion.
It is known to produce (w / v) ethyl alcohol, and the resulting ethyl alcohol can be used as the alcohol in the present invention without any problem.

The amount of alcohols added is preferably 0.01 to 4 parts, and particularly preferably 0.1 to 2 parts, relative to 1 part of yeast (dry weight part).

Sufficient effect does not appear if the amount added is less than this range,
On the other hand, if the concentration is higher than this, on the contrary, the phenomenon that the amount of the hydrophobic liquid contained in the yeast is decreased is seen, which is an unfavorable effect.

As one of constituents for producing microcapsules using microorganisms in JP-A-58-107189, alcohols having 4 to 15 carbon atoms is referred to as an organic substance for increasing lipid content. Although there is a description used, alcohols described in these publications are used as a hydrophobic liquid contained in microcapsules and do not have a water-soluble property. It is clearly distinguished from the water-soluble polar solvent in the invention.

That is, the alcohol used in the present invention is added to impart a catalytic effect in the microencapsulation process, and a part thereof may not penetrate into the yeast cell during the encapsulation process. However, it cannot be essentially a hydrophobic liquid contained in microcapsules, and its purpose is completely different.

Moreover, when a polyhydric alcohol such as ethylene glycol or glycerin was used, no catalytic effect was observed in the microencapsulation process as described in the present invention.

Furthermore, when ketones such as acetone and methyl ethyl ketone are used, similar catalytic effects are observed even though they are inferior to alcohols, but generally because ketones are highly effective in extracting lipids from yeast cells. , Immediately after the addition of ketones, or over time, yeast flocculates, resulting in poor dispersion stability of the obtained microcapsule particles and causing severe surface roughness when smeared on a support such as paper. there were.

Yeast used in the present invention is a general term for microorganisms that grow by budding or division, but as a classification, it is roughly divided into spore-producing yeast that reproduces sexually and non-spore-forming yeast that does not, and both fungal phyla. Belong to.

The former belongs to Ascomycota, Protozoa, Endomycetaceae, and the latter belongs to Incomplete Mycota, Cryptococcales Cryptococcaceae.

Furthermore, spore yeast (Endomycetaceae) is classified into the following subfamilies and genera.

I Eremascoideae [Eremascoideae] Eremascus [[Eremascus]] II Endomycoideae [Endomycoideae] Endomyces [[Endomyces]] Schizosaccharomyces [[Schizosaccharomyces]] III Saccharomycoideae ] A Endomycopseae [Endomycopseae] Endomycopsis [[Endomycopsis]] B Saccharomyceteae [Saccharomyces] [Saccharomyces] a. Saccharomyces [[Saccharomyces]] b. Chigosaccharomyces subgenus [(Zygosaccharomyce
s]] Toruruspora [[Torulaspora]] Pichia [[Pichia]] a. Pichia [[Pichia]] b. ] Schwaniomyces genus [[Schwaniomyces]] C Nazosonieae [Nadsonieae] Saccharomycodes [[Saccharomycodes]] Nadsonia [[Nadsonia]] IV Nematosporoideae [Nematosporoideae] Monosporella [[Monosporella]] ] [Nematospora] [[Coccidiascus]] Sporeless yeast (Cryptococciaceae) is classified into the subfamily and the genus shown below.

I Cryptococcoideae [Cryptococideae] Cryptococcus [[Cryptococcus]] Torulopsis [[Torulopsis]] Pichirosporum [[Pityrosporum]] [Plettanomyces] [[Brettanomyces]] Candida ] Trigonopsis [[Trigonopsis]] II Trichosporoideae [Trichosporon] [[Trichosporon]] III Rhodotoruloideae Rhodotorula [Rhodotorula] More specifically, Saccharomyces spp. Cereviche 〔〔saccharomyces cerev
iceae]] Saccharomyces rouxi
i]] Saccharomyces Carlsbergensis [[saccharomy
ces carlsbergensis]) Saccharomyces Uvaru
Endomyces Vernalis of the genus Endomyces [[Endomyces.Vernalis]] Lipomyces lipofer of the genus Lipomyces [[lypomyces.lipofer]] Lipomyces starkei [[lypomyces.starkeyi]]
s]] Candida utills] of the genus Candida ([Candida tropicalli]
s]] Candida lypolytica [[Candida lypolytica]] Candida flaveri!

The components constituting the yeast cells are roughly divided into glucan mainly constituting the cell wall, water-insoluble component mainly composed of mannan and phospholipid component mainly constituting the cell membrane. Water-soluble or polar solvent-soluble enzymes and proteins. It is divided into components, and these are distributed differently depending on the type of yeast, but the yeast used in the present invention has any composition. In addition, yeast has no proliferative function, that is, whether it is alive or dead, it has no effect on the effect of the present invention. There are various shapes of yeast such as oval, spherical, lemon-shaped, columnar, and oval depending on the type of yeast, but shapes such as circular, oval, and oval are preferable. The particle size is preferably 5 to 20 μm.

The yeast dispersion liquid may be a yeast dispersion liquid prepared by dispersing a commercially available yeast strain (commercially available as baker's yeast in a semi-dehydrated state) in a solvent such as water, or a nutrient source such as a carbon source or a nitrogen source. What was obtained by growing yeast in the medium containing the yeast may be used as it is as the yeast dispersion liquid. If necessary, pH adjustment or addition of a preservative is also performed.

The yeast concentration (dry solid content concentration) in the yeast dispersion liquid is not particularly limited, but is preferably 10 to 20% (W / W). If it is less than this range, the production efficiency will be poor, and if it exceeds 20%, the viscosity of the dispersion liquid will be rapidly increased, which will hinder uniform stirring, which is not preferable.

When the hydrophobic liquid used in the present invention is applied as a microcapsule for carbonless paper, it has a good solubility of an electron-donating colorless dye b. It is nearly colorless and odorless c. Wide temperature range However, it is required that the liquid be stable and the like, but a hydrophobic liquid can be easily encapsulated.

Specifically, paraffin oil, cottonseed oil, soybean oil, corn oil, olive oil, castor oil, fish oil, chlorinated paraffin,
Chlorinated diphenyl, dibutyl phthalate, dioctyl phthalate, dibutyl maleate, O-dichlorobenzene, alkylated naphthalenes such as diisopropyl naphthalene, 1-phenyl-1-xylyl ethane, and the like,
If necessary, dyes, fragrances, pharmaceuticals and the like are dissolved or dispersed in these hydrophobic liquids, but the hydrophobic liquids that are immiscible with the water-soluble liquid can be used alone. The addition of the hydrophobic liquid to the yeast dispersion may be performed as it is, but in order to allow the yeast to exist in a more uniform state, it is dispersed in an aqueous solution containing a suitable emulsifier, and the emulsified state After that, it is preferable to add.

The temperature in the encapsulation step is not particularly limited, but is preferably 20 ° C to 70 ° C. The time is required for 1 hour or more, but can be appropriately changed depending on the amount of the hydrophobic liquid contained and the encapsulation temperature.

(E) Examples The present invention will be described in more detail with reference to Examples. The yeast weights shown in Examples and Comparative Examples are all parts by weight in a dry and dehydrated state (both intracellular and extracellular).

Example-1 As an emulsifier, as a hydrophobic liquid in 20 parts of a 0.5% Tween 80 (Kao Atlas nonionic surfactant) aqueous solution,
3- (N-methylcyclohexylamino-6-methyl-
Hisol SASN containing 1.1 parts of 7-anilinofluoran (black color dye manufactured by Shin Nisso Chemical Co., Ltd., trade name PSD-150)
22 parts of -296 (hydrophobic liquid with high boiling point, manufactured by Nippon Petrochemical Co., Ltd.) was added with vigorous stirring to give an average liquid particle size of 8 μm to obtain a hydrophobic liquid emulsion. Next, 100 parts of a dispersion liquid containing 10 parts of commercially available baker's yeast (produced by Oriental Yeast Co., Ltd., Saccharomyces cerevisiae) [Saccharomyces cerevisiae] (cell concentration 10%), ethyl alcohol 15 parts and the above hydrophobic After adding 42 parts of the liquid dispersion, the mixture was continuously shaken in a rotary shaker at a temperature of 50 ° C. and a stirring speed of 200 rpm for 3 hours, so that the hydrophobic liquid was completely contained in the yeast. Microcapsulation was completed.When this microcapsule dispersion was applied directly to 40g / m ² of high quality paper with bar coating at an application amount of about 5g / m ^2, an excellent carbon-free top paper for carbon paper was obtained. It was

Example-2 16 parts of Hisol SAS N-296 containing 0.5 part of crystal violet lactone (blue color-developing, electron-donating colorless dye) as a hydrophobic liquid is added to 20 parts of an aqueous solution containing the same surfactant as in Example 1. Was added with vigorous stirring to an average particle size of 5 μm to obtain a hydrophobic liquid emulsion. Next, after adding 10 parts of isopropyl alcohol and 36 parts of the above-mentioned hydrophobic liquid dispersion to 100 parts of the same yeast dispersion as in Example 1,
Shaking was continued for 3 hours in a rotary shaker under conditions of a temperature of 50 ° C. and a stirring speed of 200 rpm. As a result, the hydrophobic liquid was completely contained in the yeast and the microencapsulation was completed. When this microcapsule was smeared on a paper in the same manner as in Example 1, an upper paper for carbonless paper with good color development was obtained.

Example-3 13 parts of corn oil containing 0.4 parts of apple flavor (manufactured by Takasago International Corporation) and 5 parts of n-propyl alcohol were mixed to obtain a hydrophobic liquid contained in microcapsules.
Dry baker's yeast as yeast (Oriental Yeast Co., Ltd.)
(Manufactured and killed yeast) 18 parts of the above hydrophobic liquid was added to 100 parts of the yeast dispersion containing 10 parts, and the temperature was kept at 40 ° C in a rotary shaker.
When the shaking was continued for 4 hours under the conditions of the temperature of 200 ° C. and the rotation speed of 200 rpm, all the hydrophobic liquid was contained in the yeast and the perfume microcapsules were obtained.

This microcapsule dispersion was applied directly to 80 g / m ² high-quality paper as a bar coat at a coating amount of about 7 g / m ² , and when pressure was applied to the coated part, paper that released an aroma only when the microcapsules were destroyed It was obtained and could be used for applications such as sample books for perfumes.

Example 4 One platinum loop of a Candida lypolytica [[Candida lypolytica]] (preserved strain transferred from the IFO-0717 Foundation for Fermentation Research Institute) was inoculated into a medium for yeast having the composition ratio described below to inoculate the bacteria. What was grown to a body concentration of 100 g / was used as a yeast dispersion liquid. (This strain does not undergo alcohol fermentation.) Next, 30 parts of the hydrophobic liquid dispersion described in Example 1 was added, and further 15 parts of methyl alcohol was added to 100 parts of the yeast dispersion. Then, shaking was continued for 4 hours in a rotary shaker at a temperature of 25 ° C. and a rotation speed of 200 rpm to complete the microencapsulation. As a result, the hydrophobic liquid was all contained in the yeast, and microcapsules for carbonless paper with good color development were obtained.

Medium composition Glucose 100 parts Yeast extract 5 parts Polypeptone 5 parts Water 1 pH5.6 Example-5 Saccharomyces was added to a yeast medium having the composition ratio described below.
Ubarum [(Saccharomyces-Uvarum]] (IFO-0290
((Beer yeast))) was inoculated with 1 platinum loop and the cell concentration was 10% (w /
The cells were grown until they reached w) to obtain a yeast dispersion liquid. In addition, the concentration of ethyl alcohol in the yeast dispersion liquid produced by alcohol fermentation was 5.2% (w / w) based on the yeast dispersion liquid, as measured by gas chromatography.

As a hydrophobic liquid, 20 parts of corn oil containing 1 part of vitamin-A (accelephthol) was vigorously stirred in 15 parts of 2.0% gelatin aqueous solution (Miyagi Chemical Co., Ltd., acid-treated gelatin YGK) to make the hydrophobic liquid. A dispersion of was prepared and added to 100 parts of the above yeast dispersion. (10 yeasts in this dispersion
Parts, ethyl alcohol contains 5.2 parts. ) Thereafter, shaking was continued for 4 hours in a rotary shaker under the condition of a temperature of 40 ° C. and a rotation speed of 200 rpm to complete the microencapsulation.

As a result, the vitamin-A solution was all contained in the yeast, and vitamin-A-containing microcapsules were obtained.

Comparative Example-1 Microencapsulation was attempted in the same manner as in Example-1, except that ethyl alcohol was not added to the yeast dispersion liquid in Example-1.

Example-6 Microencapsulation was attempted in the same manner as in Example-1, except that 0.5 part of ethyl alcohol was added to the yeast dispersion liquid in Example-1.

Comparative Example-2 Microencapsulation was attempted in the same manner as in Example-1, except that 45 parts of ethyl alcohol was added to the yeast dispersion liquid in Example-1.

Comparative Example-3 Microencapsulation was attempted in the same manner as in Example-1, except that 15 parts of isobutyl alcohol was added to the yeast dispersion liquid in Example-1.

Comparative Example-4 Microencapsulation was attempted in the same manner as in Example-1, except that 15 parts of isoamyl alcohol was added to the yeast dispersion liquid in Example-1.

Comparative Example-5 Microencapsulation was attempted in the same manner as in Example-1, except that the same amount of ethylene glycol was added to the yeast dispersion liquid instead of ethyl alcohol in Example-1.

Comparative Example-6 In Example-1, microencapsulation was tried in the same manner as in Example-1, except that the same amount of acetone was added to the yeast dispersion liquid instead of ethyl alcohol. As a result, vigorous yeast flocculation occurred when acetone was added, and the surface became rough even when applied to paper and dried.

In the above Examples and Comparative Examples, the "encapsulation amount" was used as an index to judge how much hydrophobic liquid was contained in the unit yeast.

The amount of encapsulation indicates how many g of the hydrophobic liquid was contained in 1 g of the yeast.

 The measuring method was according to the following procedure.

(1) Collect microcapsule slurry to a dry weight of 1 g.

(2) 10000r to separate uncapsulated hydrophobic liquid
Centrifugation is performed 3 times for 10 minutes at pm to remove the upper uncapsule and wash the microcapsule.

(3) Add 30 ml of concentrated hydrochloric acid / methanol (5 / 95v / v) solution as an extraction solvent to the dispersion liquid containing only microcapsules,
Disperse well and shake for 10 minutes at 70 ℃ to extract colorless dye.

(4) After removing the capsule film (membrane material residue) remaining in the extraction completed liquid with filter paper (Toyo Filter Paper Co., Ltd. No. 5C),
The amount of the hydrophobic liquid contained in the microcapsules is calculated by colorimetrically determining the filtrate at a wavelength of 600 nm.
Calculate the amount of hydrophobic liquid for 10 g of yeast and use it as the encapsulation amount.

In addition, in the method for quantifying vitamin A of Example-5, microcapsule slurry was centrifuged to collect unencapsulated vitamin A, and a chloroform solution of antimony trichloride was added to the uncapsulated vitamin A. Vitamin A
Was extracted, and the amount of vitamin A in the microcapsules was calculated by performing colorimetric determination at a wavelength of 325 nm.

The method for quantifying the perfume contained in the microcapsules of Example-3 was as follows. After extracting the inclusions from the microcapsules with the above-mentioned hydrochloric acid + methanol solution, the corn oil in the extract was quantified by a gas chromatography method. And the amount of encapsulation.

Table 1 shows a summary of the contents of Examples 1 to 6 and Comparative Examples 1 to 6 and the encapsulation amount.

The "amount of polar solvent in cells" in Table 1 indicates the weight of polar solvent present in 1 g of yeast after the completion of microencapsulation and was measured by the following procedure.

(1) Collect microcapsule slurry to a dry weight of 1 g.

(2) In order to remove the polar solvent present in the microcapsule dispersion medium, centrifugation is performed 3 times at 10,000 rpm for 10 minutes to wash the microcapsules.

(3) The above-mentioned hydrochloric acid + methanol solution was added as an extraction solvent to the microcapsule dispersion liquid, and after well dispersed,
Extract the polar solvent at 70 ° C for 10 minutes. At this time, a reflux tube for cooling is attached to prevent evaporation of the polar solvent.

(4) After removing the capsule film remaining in the extraction-completed liquid with paper, the polar solvent in the liquid was quantified and calculated by the gas chromatography method to obtain the intracellular polar solvent amount.

(F) Effect of the invention From the results of the examples, it is clear that the presence of a specific alcohol in the production of microcapsules using a microorganism dramatically improves the encapsulation amount. In particular, when the microcapsules according to the present invention are applied as pressure-sensitive copying paper, it is possible to reduce the total coating amount with a capsule having a higher encapsulation amount, and thus improve the drying property during the smearing process, It also brings about a great effect on the improvement of color printability. Food,
The same applies to microcapsules of pharmaceuticals, and the main target components when used as a powder after being smeared on paper or after undergoing some dehydration / drying process are:
Obviously, since it is an encapsulated substance, it is obvious that the higher the proportion of the internal phase substance in the unit microcapsule is, the more preferable it is in consideration of the production efficiency and the like.

As described above, it is clear that the method for producing microcapsules according to the present invention is an excellent method that has never been seen in the past, and can be very useful industrially.

Continuation of the front page (51) Int.Cl. ⁶ Identification code Office reference number FI Technical display location C12N 1/16 Z 8828-4B C12P 1/02 Z 7417-4B

Claims (2)

[Claims] 1. A method for producing a microcapsule comprising a yeast cell containing a hydrophobic liquid, wherein a monohydric alcohol having 1 to 3 carbon atoms is added to the yeast dispersion liquid. Yeast added (dry weight part)
The method for producing microcapsules is characterized by 0.01 to 4 parts per 1 part. 2. The method for producing microcapsules according to claim 1, wherein the alcohol is ethyl alcohol.