JPS61282091A - Production of cacao butter substitute fat - Google Patents

Abstract

PURPOSE:To produce a cacao butter substitute fat, by cultivating a microorganisms suffering from growth inhibition by sterculic acid or a derivative thereof, accumulating a fat or oil rich in a specific component in the microbial cells and extracting and recovering the resultant fat or oil. CONSTITUTION:A microorganism, suffering from growth inhibition by sterculic acid or a derivative thereof, e.g. Candida humicola (FERM-P No.783), belonging to the genus Candida, Rhodotorula, Rhodosporidium or Cryptococcus, is cultivated. The cultivation is carried out in a culture medium containing nutrients required by the microorganism used under aerobic conditions to collect the wet microbial cells, from which the aimed fat or oil is recovered. When <=1ppm sterculic acid or a derivative is present in the culture medium, inhibition is defined as follows; the growth rate is suppressed to <=80% as compared with that in the absence thereof.

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Field of Application) The present invention relates to a method for producing a cocoa butter substitute using microbial cell oil.

(Prior Art) Currently, the raw material oils for cacao butter substitutes are southern vegetable oils such as shea butter and soybean oil, and these are used to produce cacao butter substitutes. In recent years, the importance of microbial cells themselves in terms of nutrition, nutritional value, and physiological effects has attracted attention.
Microbial cells have been developed as products for the food field.

*One example of this is the research involved in producing a cocoa butter substitute known as "hard putter" or "hard fat" using biological microorganisms (Japanese Patent Publication No. 5B-34114).

(Problem that the invention seeks to solve) However, the composition of fats and oils extracted and recovered from microbial cells is different from the composition of cocoa butter. For example, cocoa butter contains large amounts of 1,3-disaturated-2-unsaturated glycerides, including 1,3-distearo-2-olein (hereinafter referred to as SOS) 24.2%, 1-stearo-
2-oleo-3-baMmitin and 1-palmito-2-oleo-3-stearin (hereinafter referred to as pos) 32.8
%.

1.3-divalmitoe2-olein (hereinafter referred to as pop) is reported to be 12.1% (691% in total of S and U);
The composition of the fats and oils extracted and recovered from the bacterial cells of FO415@ is 5O86.4%, PO322,
2%, POP 19.3%C or moreSURU total 47.9%)
Therefore, one separation step is required. For this reason, the cost is high, and it is difficult to say that this method is necessarily sufficient from an economic standpoint as a method for producing a cocoa butter substitute fat.

Therefore, we obtained a microorganism with a high SIU content and a glyceride composition close to that of cocoa butter.

It is desired to develop an industrially advantageous method for producing a cocoa butter substitute.

(Means and effects for solving the problem) The present inventors have continued research on oil production by microorganisms, and have obtained microorganisms with unique characteristics using various methods. Among these, microbial oils whose growth is particularly inhibited by sterculic acid or its derivatives have higher S and U components than previously reported microbial oils, and their triglycephyte composition is similar to that of cacao tar. I found it.

The present invention provides a cocoa butter substitute, which is characterized by culturing microorganisms whose growth is inhibited by steruclic acid or its derivatives, accumulating fats and oils rich in S-like components in the microbial cells, and extracting and recovering the fats and oils. It is a manufacturing method.

Inhibition of growth of microorganisms according to the present invention refers to bacterial strains, medium composition,
Although it varies depending on factors such as concentration, it is defined as follows in the present invention. That is, in the presence of 1p or more of sterculic acid or its derivatives in the medium, the growth rate is suppressed to 80% or less compared to when no addition is made.

Microorganisms that can be used in the practice of the present invention are microorganisms whose growth is inhibited by sterculic acid or derivatives thereof;
Those obtained from nature can also be used as appropriate. For example, Canguida genus, Rhodoto71/ff
(Rhodotrula) genus, Rhodosboridium (
nhoaosportaxum), Endomyces (
Knaom7Qes) genus, Cryptococcus (0ry
ptooocous), Lipomyces (Ll, pomy
ces), genus Metostinichouia (mtsohnj-
kowia), Plchla, Saooharomyaes, Toruopsia, Trichosboa
Examples include microorganisms belonging to the genus Triohosporon. To give a specific example of the species name, Candida humicola (0andi+ia humicola)
Candida humicola mutant strains (0and1da humioδla cI3s 2
The bacterial cell oil of B22 KNK 286) has the following triglycephyte composition, POP 18.6%, 5O
P39.0%, SO814, 6%C or more S U total 72
．． 2%) and can be used as is as a cocoa butter substitute fat. This mutant strain was deposited with the Institute of Microbial Technology, Agency of Industrial Science and Technology, and was published as FIKEN Article No. 783 (FIRM BP).
-783).

In order to select microorganisms such as those mentioned above that have been collected from the natural world or have been artificially mutated, it is preferable to use, for example, the following method. i.e. 1 ppm to 1% of steruclic acid or methyl nucleucrate, preferably 5 ppm
Microorganisms were inoculated into a medium supplemented with ~t0 ppm and a medium not containing the substance, and when observed for 1 to 10 days, preferably 2 to 4 days, no growth was observed in the former medium, or no growth was observed in the former medium. However, microorganisms that grow well in the latter medium without additives are selected. A medium is a medium in which microorganisms normally grow.

For example, any medium for growing vines of the genus Candida (0and1da) may be used.

In order to obtain microorganisms having the above-mentioned properties by artificial mutation, for example, high-energy radiation treatments such as ultraviolet irradiation, CO60 treatment, and X-ray irradiation, or treatment with sodium sulfite, hydroxyamine, 4-methyl-N'-nitroso Guanidine, Acry7? A conventional mutagenesis treatment using a chemical inducer of a nitrogen mustard donor or the like can be used as appropriate.

Some of the bacteria collected in this way are also endowed with physiological and morphological variations such as auxotrophy for vitamins, amino acids, and other nutrients, and respiratory deficiencies; However, it goes without saying that if the production is inhibited by steruclic acid or its derivatives and the S and U components of the extracted fats and oils are high, it can be used in the present invention.

Culture may be carried out either by standing still or by shaking, but deep culture with aeration and agitation is generally advantageous. The medium used for culture is
It is usually a liquid medium that contains carbon sources, nitrogen sources, and other nutrient sources that the bacteria can use. For example, carbon sources include pentose (xylose, etc.), hexose (glucose, fructose, etc.), diflfAW4 (sucrose, maltose, etc.), but if they can be used by microorganisms, especially Not limited. In addition, highly assimilated industrial wastes such as sulfite valve waste liquid, waste molasses, and wood saccharification waste liquid may also be used, 1i! Sources include ammonium salts, nitrates, ammonium 71m salts, yeast extract, meat extract, and fishmeal.

Natural products such as corn steep liquor and soybean meal can be used. In addition, if necessary, metal salts such as iron,
Organic or inorganic salts such as magnesium, calcium, and manganese may be added, and if the improved bacteria used have auxotrophic properties, appropriate amounts of the necessary nutrients can be added as appropriate.

The various nutrient sources mentioned above are contained in the medium in an optimal formulation for the bacteria used. These conditions vary depending on the bacterial species, but especially for bacterial cell growth and oil accumulation, C
Since it is important, it should be decided after careful consideration.

Such a medium is usually pH 2 to 9, preferably pH 4.
~7, temperature 20°C ~ 40°C, preferably 25°C ~ 33°C
Maintain at °C and culture aerobically. During this time, an antifoaming agent that does not inhibit fermentation may be added, if necessary. In this way, fats and oils rich in 82U components are accumulated in the cultured bacterial cells.

To extract fats and oils from the bacterial cell growth obtained as described above, first, the culture solution is filtered or centrifuged to collect wet bacterial cells b6
Wet bacterial cells can be ground as they are using Colloidomi V, a ball mill, etc., and then extracted with a solvent such as hexane, or once frozen or dried, they can be ground using a gauge press, expeller, etc.
There are also methods of extraction using hexane, etc., or extraction using hexane, etc., or a combination of both. The waste bacteria after collecting oil and fats is a rich protein resource and can be used as feed.

On the other hand, the extracted crude fats and oils contain impurities such as hexanot and di-glycephyte, free fatty acids, and sterols, so they must be removed by purification. Refining may be carried out in accordance with the refining of ordinary fats and oils.

As described above, according to the present invention, it is possible to industrially advantageously produce from microorganisms a cocoa butter substitute fat with high s and tr contents and a glyceride composition similar to that of cocoa butter. This will be specifically clarified by examples.

(Example) The present invention will be explained more specifically using examples.

Examples Candida humicola
la) A malt extract agar slant culture of OB82822 (original strain) was scraped and suspended in sterile physiological saline to obtain a viable cell count of approximately 101 to 101 cells/11 t.
After mutating GV course 12.51/1.1 by NTG treatment. Yeast extract s, ysl/1, malt extract 5. ．． 7Sl/jl, polypeptone 6.25'Il
l.

Plate medium consisting of agar 251/11 A K Q, 2
After spreading the ml and culturing at 30°C for about 4B to 72 hours,
This was added to 3 ppm of methyl sterculate and inoculated onto optical plane medium B by the replica method (transfer).
After culturing for a period of time, colonies that do not appear in B are collected from A to obtain a mutant strain OB! 92822 NK-21
36 (FRtRM BP-783) was obtained.

Next, the thick and mutant strains were mixed with blackstrap molasses (35% sugar concentration) 1
42. BsI/l, urea a, 3f/l, 85% phosphate α
After sterilizing the medium consisting of 3 sl/1, each pure culture seed was inoculated and cultured with shaking at 30° C. for 4 days. After the culture was completed, the cells were collected by centrifugation, dried, and the yield of cells was measured. In addition, after the remaining wet bacterial cells were treated with sulfuric acid, crude oils and fats in the bacterial cells were extracted using Nisu V Aete V, and the fatty acid composition and triglyceride composition were measured. The results are described below.

Note that the mutant crude oil and fat has an acid value of 4Q, 4. Iodine value 38.3,
The melting point was 28.5°C. This mutant strain had a growth rate of 68% in the presence of Ste V ric acid Menal 4 pp+21 in the medium compared to that without the addition. No inhibition of growth was observed even when 5% methyl chlorate of Ste-V was added to the culture medium.

Thick KNK 286 (yI!XRMBP-783)
Fat content: 30.5% 29.8%Fatty acid composition e16:o 29.6% 25.8%01a
:o 12. OS 3.8 C1a:t 46.532.7 C18:2 11. q b,.

018:3 Trace Trace 020:O trace α7 Triglyceride composition: Thick OLL, (1-oleo-2-riluo-3-liruine and 1-liluo-2-liluo-3-olein) 1
．． 1% 0 (1(1-oleo-2-oleo-3-lyluine and 1-oleo-2-oleo-3-olein) 6,1%
000()liolein) 1a1%PI,
L(1-palmito-2-riluo-3-liruine and 1-
Riluo 2-Riluo 3-valmitin) 11%M
OI, (1-myristo-2-oleo-3-lyluine and 1-lino-V-o-2-oleo-3-myristin) 1.6%
POL (1-paV mido-2-oleo-3-liruine and 1-liluo-2-oleo-3-palmitine) 9.2%P
OO (1-palmito-2-oleo-3-olein and
-oleo-2-oleo-3-valmitine) 2
6.6% 800 (1-stearo-2-oleo-3-olein and 1-oleo-2-oleo-3-stearin),
10.8% PLP (1,3-palmito-2-riluine)
2.3% POP (1.S-palmito-2-olein)
14.0% 5op (t-stearo-2-oleo-3-valmitine and 1-palmito-2-oleo-
3-stearin) 14.7%5os(1.3
-stearo-2-olein) 2.6% KNK 286
(Fil: RM BP-783) ooz (1-oleo-2-oleo-3-liruine and 1-oleo-2-oleo-3-olein) 1.6% POL (1-palmito-2
S, O% POO (1-palmito-2-oleo-3-olein and 1-oleo-2-
oleo-3-valmitine) a2A800(
1-Stearo-2-oleo-3-olein and 1-oleo-2-oleo-3-stearin) 6.5% psp (1
・3-BaV Mido 2-Liluin) 3.3% POF (1 bow- Palmito 2-Olein) 18.6%
BOP (1-stearo-2-oleo-3-valmitin and 1-pal-1-2-oleo-3-stearin)
39.0% Boa (1,3-stearo-2-olein) 14.6% 80A (1-stearo-2-
oleo-3-elaidin and hi-fi 2-oleo-
3-Stearin) 1.2% PPP () Riba V Mitin)
α9% PPB (1-paV mido 2-palmito 3-stearin and 1-stearo-2
-palmito-3-stearin) 1. b%5sp
r t-stearo-2-stearo-3-valmitin and H-1-vaV mido-2-sdearo-3-stearin) 1.6
%pop, sop, sos 8. The sum of υ is 3 in thickness.
1.3%, whereas the mutant strain KHx 2a6 (yx
iMBP-783), it was 172.2%.

Mutant strain KNK 286 (FIRM BP-78x)
As shown in the cooling curve in Figure 1, the extracted fats and oils of
It is clear that its physical properties are similar to cocoa butter.

This KNK 286 (FIRM BP-783) extracted oil and fat 13%, cocoa butter 10%, cocoa mass 16%,
Mixture of 18% whole milk powder and 43% powdered sugar, 7% extracted oil and fat in KN (yvRMBP-7g3), medium melting point part of palm oil (dissolved in twice the amount of normal hexane as compared to the oil, -1S℃) and 6% cocoa butter (medium melting point portion), which was stirred for 60 minutes at 5°C and separated.
0%, cocoa mass 16%, whole milk powder 18% and powdered sugar 43%
% were used to prepare chocolate in a conventional manner. As a result, the allowable temperature difference during tempering was ±2.0°C for all formulation examples, and release from the mold was also good. In addition, in anti-V-muchist, temperature changes of 18 °C and 32 °C were carried out for 10 times in each 12-hour cycle.
When repeated several times, no plume was observed, indicating excellent plume resistance.

[Brief explanation of drawings]

FIG. 1 is a cooling carp of extracted oil and fat of Candida humicola OBS 2822 KNK-286 used in the present invention.

Claims (3)

[Claims] (1) Microorganisms whose growth is inhibited by steruclic acid or its derivatives are cultured in a medium, and S_2U (where S_2 is the hydroxyl group at the 1st and 3rd positions of glycerin is C_1_6, C_1_8, / or C_2_0 saturated fatty acid residue, U means a disaturated-monounsaturated triglyceride in which the hydroxyl group at the 2-position of the same glycerin is substituted with a C_1_4 to C_2_0 unsaturated fatty acid residue, respectively. A method for producing a cocoa butter substitute, which comprises collecting accumulated microbial cells and recovering fats and oils rich in S_2U components from the collected microbial cells. (2) The production method according to claim 1, wherein the microorganism is a microorganism belonging to the genus Candida. (3) The growth inhibition is inhibition in which the growth rate is suppressed to 80% or less in the presence of 1 ppm or more of sterculic acid or its derivative in the medium compared to the case without addition. Manufacturing method.