JP5424268B2 - Method for producing high protein content organic matter, high protein content organic matter, feed production method, and feed - Google Patents

Description

  The present invention relates to a method for producing a high protein-containing organic substance and feed by removing phorbol ester from an organic substance containing phorbol ester.

  Patent Document 1 and Patent Document 2 describe processing methods for the purpose of removing the environmental pollutants and toxic substances contained in feed or feed material raw materials, which are intended for processing and production of animal feed or feed material. What is disclosed is given as a representative example.

  The technique described in Patent Document 1 is a method of adding a fatty acid ester, a fatty acid amide, a free fatty acid or a hydrocarbon, which is a volatile working fluid, to a fat or oil containing an environmental pollutant or a toxic component, By stripping with a volatile working fluid, environmental pollutants or toxic components are separated from the fat or oil along with the volatile working fluid. Note that the stripping process is performed by blowing vapor or gas into a liquid containing a specific substance to be removed, mixing a highly volatile liquid and then volatilizing it, or putting the whole liquid under vacuum conditions. In this processing method, the specific substance is removed from the liquid by moving the specific substance into a vapor, gas phase or volatile fluid phase, or volatilizing the specific substance itself.

  The technique described in Patent Document 2 is intended to remove phytic acid contained in grain as feed or food. If an animal ingests feed or food containing high concentrations of phytic acid, it can interfere with normal intestinal absorption of nutritionally important trace metals, resulting in a series of deficiency disorders. For this reason, it is required to remove phytic acid from grains containing phytic acid as described above. This technology inoculates cereals such as soybean koji containing phytic acid with koji molds and produces koji, thereby using phytate degrading enzymes such as phytase and phosphatase that are produced during the growth of koji molds. Is removed.

Japanese Patent No. 3905538 Japanese Patent Laid-Open No. 8-214822

  However, in the technique disclosed in Patent Document 1, a volatile working fluid containing environmental pollutants or toxic components remains after the stripping process. Since it is not easy to separate environmental pollutants or toxic components from this volatile working fluid, it is difficult to reuse the volatile working fluid once it has undergone the stripping process. Therefore, it is required to use a new volatile working fluid for each stripping process, and the volatile working fluid that has undergone the stripping process contains environmental pollutants or toxic components. There is a problem that it is required to dispose of safely, and the running cost related to the processing becomes high.

  In addition, the stripping process used in this technology is the environment where the temperature and pressure in the tank and the supply rate of the volatile working fluid are not accurately controlled in the stripping tank that actually performs the stripping process. The removal rate of pollutants and toxic components does not increase. For this reason, in order to realize these, expensive control devices and equipment are inevitably required, and there is a problem that the initial cost at the time of installation of the equipment is increased.

  Furthermore, the stripping process used in this technology increases the removal rate if the environmental pollutants and toxic components in the processing object and the volatile working fluid are not sufficiently mixed and contacted with each other in the stripping tank. I can't. For this reason, the object to be treated is necessarily limited to a liquid substance such as fat or oil. Therefore, it has the big problem that it is difficult to apply to solid processing objects such as soybean meal and other squeezed rice cakes.

  Moreover, although the technique disclosed in Patent Document 2 can be applied to solid processing objects such as soybean meal and other squeezed rice cakes, those that can be removed are included in the processing object. Limited to phytic acid. For this reason, there exists a problem that it cannot apply, when the other toxic component which cannot be decomposed | disassembled with the phytate decomposing enzyme called the phytase and phosphatase produced in the process in which a koji mold is propagated is contained.

Here, Jatropha curcas L. seeds contain a high content of 30-40% oil, but this oil contains phorbol esters that have been reported as carcinogenic promoters. Therefore, it is not suitable for food. For this reason, in recent years, it has attracted attention as a promising renewable energy resource that does not cause competition with food applications.
In addition, when a large amount of seeds is squeezed, a large amount of seed pomace is inevitably generated. The protein content of this pomace is about 60%, and the protein content of soybean pomace, which is the main feed ingredient ( Jatropha seed pomace has the potential to be used as a feed ingredient superior to soybean pomace.
However, since phorbol esters are also contained in this Jatropha seed pomace, it is actually difficult to use it as a feed material, and it can be used as a fertilizer with low added value or discarded without using it. The current situation is that there is no other way.

  Even if it is going to apply the technique indicated by patent documents 1 in order to remove the phorbol ester which is the toxic component from the organic substance containing phorbol ester which has been reported as a carcinogenic promoter like Jatropha seed pomace, Since Jatropha seed pomace is a solid organic substance, it is physically difficult to sufficiently remove the phorbol ester by the stripping treatment used in this technique. In addition, even when trying to apply the technique disclosed in Patent Document 2, gonococcus cannot be sufficiently removed because of its low ability to decompose phorbol esters.

  As described above, the purpose of sufficiently removing phorbol esters, which are toxic components, from organic substances containing phorbol esters that have been reported as carcinogenic promoters such as Jatropha seed pomace has already been disclosed. However, it has been required to develop a new technology that can achieve the above-mentioned purpose.

Therefore, the present inventors have intensively studied and obtained phorbol ester from an organic substance containing phorbol ester at a low cost by utilizing microorganisms that are easily available while having high degradability for phorbol ester. The present invention has been completed by finding that it can be decomposed and removed with a high throughput.
That is, the present invention relates to a method for producing a high protein-containing organic substance that can be suitably used as a livestock feed by removing phorbol ester from an organic substance containing phorbol ester using Bacillus sp. It aims at providing a feed manufacturing method and feed.

In order to achieve the above object, the method for producing a high protein-containing organic material of the present invention comprises mixing and fermenting an organic material containing a phorbol ester and a bacterium belonging to the genus Bacillus to decompose the phorbol ester in the organic material. There is a way.

  Moreover, the high protein content organic substance of this invention is set as the structure which mixed and fermented the organic substance containing a phorbol ester, and Bacillus genus bacteria, and decomposed | disassembled the phorbol ester in an organic substance.

  Moreover, the manufacturing method of the feed of this invention is as a method of decomposing | disassembling the phorbol ester in an organic substance by mixing and fermenting the organic substance containing a phorbol ester, and a Bacillus genus microbe.

  Moreover, the feed of this invention is set as the structure containing the fermented product obtained by mixing and fermenting the organic substance containing a phorbol ester, and Bacillus genus bacteria, and decomposing | disassembling a phorbol ester.

According to the present invention, from an organic substance containing phorbol ester that has been reported as a carcinogenic promoter such as Jatropha seed pomace, waste liquid containing phorbol ester using an expensive volatile reagent or containing a toxic substance that is difficult to process. Can be removed without introducing an expensive control device or control equipment.
In addition, phorbol esters are decomposed and removed more efficiently than when other microorganisms are used by fermenting organic substances containing phorbol esters under relatively mild conditions using inexpensive and readily available Bacillus spp. can do.
By these, the initial cost related to the treatment, the production method of the high protein-containing organic substance, which can decompose and remove phorbol ester with excellent efficiency while keeping the running cost low, the high protein-containing organic substance, the production method of the feed, and The feed can be provided.

It is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of 1st embodiment of this invention. It is a figure explaining the productivity of the oil of Jatropha. It is a figure explaining the generation | occurrence | production amount of the oil squeezed slag of Jatropha. It is a figure explaining the predominance as a feed raw material of a Jatropha seed nuclear oil extraction pomace. It is a figure which shows the phorbol ester decomposition rate in organic substance. It is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of 2nd embodiment of this invention. It is a figure which shows the change of phorbol ester content in organic substance. It is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of 3rd embodiment of this invention. It is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of 4th embodiment of this invention. It is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of 5th embodiment of this invention. It is a figure which shows the change of phorbol ester content in the organic substance in an Example and a comparative example. It is a graph which shows the change of phorbol ester content in the organic substance in an Example and a comparative example. It is a graph which shows the figure which shows the breeding test result of the chicken by the feed of this invention, and an average body weight. It is the figure and graph which show the feed intake of the chicken by the feed of this invention.

  Hereinafter, preferred embodiments of a method for producing a high protein content organic material, a high protein content organic material, a feed production method, and a feed according to the present invention will be described with reference to the drawings.

[First embodiment]
First, the structure of 1st embodiment of this invention is demonstrated with reference to FIGS. Drawing 1 is a figure showing a process of a manufacturing method of high protein content organic matter and feed of this embodiment. FIGS. 2 to 5 are diagrams for explaining the productivity of Jatropha oil, diagrams for explaining the amount of jatropha oil squeezed, diagrams showing the phorbol ester decomposition rate in organic matter, and Jatropha seed kernel oil squeezed It is a figure explaining the predominance as feed materials.

As shown in FIG. 1, in the method for producing a high protein-containing organic substance and feed according to this embodiment, first, an organic substance containing a phorbol ester to be treated is mixed with a bacterium belonging to the genus Bacillus in a stirring step. Stir well until distribution.
Next, a fermentation process is performed in which the mixture obtained by the stirring process is transferred to a temperature-controlled fermentation chamber or fermentation vessel and fermented for a predetermined period.
After the fermentation process is finished, the mixture after treatment taken out from the fermentation chamber or the fermentation vessel is a high protein-containing organic substance in which the phorbol ester is decomposed by the action of Bacillus. In addition, components such as vitamins and minerals in the mixture after treatment are increased by the secondary action of Bacillus sp.

Here, the high protein-containing organic substance means an organic substance having a high protein content. Specifically, an organic substance having a protein content of 40 to 65% or more can be referred to as a high protein-containing organic substance.
The high protein-containing organic material thus obtained can be blended in livestock feed or used as it is to obtain a feed. When blended and used in livestock feed, the blending ratio is not particularly limited, but as described later in the examples, for example, even if 10% by weight of high protein-containing organic matter is added to the feed, chickens can be grown without any particular problems. It has been confirmed that this is possible. The livestock refers to mammals and birds other than humans such as dogs, cats, pigs, cows, horses, sheep, chickens, etc., and are used as those raised by humans.

Examples of the organic substance containing a phorbol ester used in the method for producing a high protein-containing organic substance of the present embodiment include squeezed rice cakes after squeezing seeds of Jatropha curcas L. Thus, the squeezed rice cake after the oil extracted from the inner seed kernel (kernel) can be used.

  Fig. 2 compares the annual production of oil per unit cultivated area for various representative oil crops cultivated around the world. According to the figure, the oil production of palm is prominently large, followed by Jatropha. However, the area where palm can be cultivated is limited to the tropical region where precipitation is abundant and relatively fertile, and palm oil can be used as food. For this reason, it has become difficult to obtain a global consensus in recent years when palm is used in large quantities for fuel and industrial applications, and it is difficult to expand production using palm oil as a renewable energy resource.

  On the other hand, while Jatropha has the second highest oil production after palm, Jatropha oil contains phorbol ester which has been reported as a carcinogenic promoter and therefore cannot be edible. For this reason, it does not cause competition with food use unlike palm oil. Jatropha can be cultivated not only in the rainy tropical areas where palm can be grown, but also in land where there is little precipitation and it is dry and where food crops are not grown. Attention has been paid.

FIG. 3 compares the amount of Jatropha seeds produced from per unit area of Jatropha cultivated land with the amount of oil generated by squeezing the seeds and the amount of squeezed wrinkles. As shown in the figure, when using Jatropha as a raw material, about 1.5 tons of oil per unit cultivated area can be produced per year, but at the same time, unit cultivated land equivalent to more than twice the amount of oil generated As much as 3.5 tons of oil squeezed per year is incidentally generated.
Thus, squeezed pomace that is produced in large quantities at the same time as producing oil contains phorbol ester that has been reported as a carcinogenic promoter similar to oil. Therefore, as it is, it cannot be used as animal feed materials, and its use is limited to fertilizers with low added value and solid fuels with low unit prices, and the entire renewable resources obtained by growing Jatropha can be used. It was difficult to make effective use.

  In such a situation, in this embodiment, the phorbol ester can be decomposed and removed by using the squeezed rice cake after squeezing the seeds of Jatropha as an organic substance containing the phorbol ester. It is possible to increase the value of the squeezed rice cake from which the slag has been removed as an animal feed material and put it on the market. As a result, it is possible to greatly improve the profitability of the business when making Jatropha cultivation as a business, and to supply lower-cost oil to the market. In addition, it is possible to more effectively utilize renewable biomass resources produced by growing Jatropha as a plant.

  Next, referring to FIG. 4, the squeezed rice cake obtained by squeezing out the seed kernel (kernel) that has been unshelled from Jatropha seeds is treated in the method for producing a high protein-containing organic material of this embodiment. The advantage in the case of using as an organic substance containing the target phorbol ester will be described. The figure shows the result of comparison of important composition items as feed raw materials for post-pressing pomace of Jatropha seed kernels and post-soy pruning soybeans which is a typical feed raw material.

  From this figure, it can be seen that the protein content, which is the most important composition as a feed raw material, is about 45% for soybeans and 60% or more for Jatropha. As for the lipid content and the ash content, it is shown that Jatropha is not much different from soybean, and that the fiber content is less than Jatropha than soybean. From these, as long as the phorbol ester, which has been reported as a carcinogenic promoter, can be removed, high-protein and low-fiber Jatropha seeds after nuclear extraction can be a better feed ingredient than soybean pomace. Recognize.

  In addition, in the manufacturing method of the high protein content organic substance of this embodiment, the organic substance of the object which decomposes | disassembles a phorbol ester by mixing and fermenting a Bacillus-genus microbe is not limited to Jatropha. The technical idea of the present embodiment can be applied in the same manner as long as it is a high protein-containing organic substance containing phorbol ester, and by suitably decomposing the phorbol ester with a Bacillus genus, a high protein-containing organic substance is suitably produced. Is possible.

The Bacillus bacteria used in the method for producing a high protein content organic matter present embodiment, for example, Bacillus coagulans (Bacillus coagulans), Bacillus smithii (Bacillus smithii), Bacillus subtilis ssp subtilis (Bacillus subtilis subsp. Subtilis ), Bacillus licheniformis , Bacillus cereus , Bacillus subtilis var. Natto, etc., but is not limited to this. Various Bacillus bacteria in the genus can be preferably used.

  FIG. 5 shows the result of comparative measurement of the phorbol ester degradation rate in organic matter for each bacterium. As experimental conditions, 1% of the weight of the organic substance containing phorbol ester was mixed, and then the optimum culture temperature of each bacterium (Bacillus subtilis varietus natto) and 37 ° C for yeast, The koji mold was fermented at 30 ° C. for 3 weeks. And it was compared between each microbe what percentage of the amount of phorbol ester contained in the organic substance before a process was decomposed | disassembled by HPLC analysis. As a result, it is shown that Bacillus spp. Have the best resolution.

  As described above, according to the method for producing a high protein-containing organic substance and feed according to the present embodiment, an organic substance containing a phorbol ester is used by using a Bacillus sp. Therefore, it is possible to remove the phorbol ester that has been reported as a carcinogenic promoter at a high decomposition removal rate and low cost.

In other words, in the process of decomposing and removing phorbol esters, expensive control devices and equipment are introduced without using expensive volatile reagents or generating waste liquid containing toxic substances that are difficult to process. Processing can be performed without any problem.
In addition, phorbol esters are decomposed and removed more efficiently than when other microorganisms are used by fermenting organic substances containing phorbol esters under relatively mild conditions using inexpensive and readily available Bacillus spp. can do. As a result, it is possible to achieve a high phorbol ester removal rate while keeping the initial cost and running cost related to the processing low.
Furthermore, the secondary action of Bacillus can increase components such as vitamins and minerals in the organic matter after treatment. Especially when the organic matter after treatment is used for animal feed raw materials, etc. It is also possible to increase the nutrients inside.

In addition, as an organic substance containing phorbol ester, by using the pomace after squeezing the seeds of Jatropha, according to the present embodiment, the squeezed squeeze that is generated in a larger amount than oil as a by-product of oil production by Jatropha cultivation The phorbol ester in the straw can be decomposed and removed, and the resulting oil-drawn squeezed meal can be increased in value as an animal feed material and put on the market.
Furthermore, as an organic substance containing phorbol ester, by using the squeezed rice cake (jatropha seed squeezed squeezed rice cake) after squeezing out the seed core of Jatropha and extracting the inner seed core, The concentration of nutrients can be significantly increased, and a feed material superior to soybean pomace can be obtained. For this reason, it becomes possible to raise the value of the processed oil squeezed culm as an animal feed material to the market. As a result, it is possible to contribute to further improvement of the profitability of the business when making Jatropha cultivation as a business, and thereby the effect of stabilizing the market price of Jatropha oil as a renewable energy resource at a lower price level can be expected.

[Second Embodiment]
Next, the structure of 2nd embodiment of this invention is demonstrated with reference to FIG.6 and FIG.7. FIG. 6 is a diagram showing steps of a method for producing a high protein-containing organic material and feed according to the present embodiment. FIG. 7 is a diagram showing changes in the phorbol ester content in organic materials.
As shown in FIG. 6, the method for producing a high protein-containing organic substance and feed according to the present embodiment includes (A1) a mixing step, (A2) a high-temperature and high-pressure step, (A3) a stirring step, and (A4) a fermentation step. be able to. About another point, it can be set as the thing similar to 1st embodiment, and after squeezing the seed of Jatropha curcas L. of the Euphorbiaceae family as an organic substance containing a phorbol ester also in this embodiment. Squeezed rice cake or a squeezed rice cake after squeezing out the seed kernels extracted from the seed kernel (kernel) can be used.

(A1) Mixing step First, water is mixed with an organic substance containing phorbol ester. At this time, as a mixing ratio, it is preferable to mix 0.5-3 mass parts of water with respect to 4 mass parts of organic substance containing phorbol ester. This is because the efficiency of fermentation increases when the mixing ratio of water is set in this way. From such a viewpoint, it is more preferable that the mixing ratio of water is 2 to 3 parts by mass.

(A2) High-temperature and high-pressure process Next, a mixture of an organic substance containing phorbol ester and water is sterilized at high temperature and high pressure. This kills microorganisms that can inhibit fermentation by Bacillus. This can be done in a general manner by autoclaving.
Various kinds of microorganisms are included in the organic substances to be treated that are usually used, and some of them may include microorganisms that inhibit the action of degrading phorbol esters by Bacillus sp. Therefore, high temperature and high pressure sterilization is performed to kill these inhibitory microorganisms.

(A3) Stirring step Next, the sterilized water added with Bacillus spp. Is added to the sterilized mixed solution described above and stirred sufficiently until a uniform distribution is obtained. At this time, it is preferable to add 0.004-0.2 mass part of Bacillus-genus bacteria with respect to 0.5-1 mass part of sterilized water. This is because uniform fermentation can be realized if the mixing ratio of the genus Bacillus is as described above. Moreover, from such a viewpoint, it is more preferable that the mixing ratio of the genus Bacillus is 0.04 to 0.12 parts by mass.

(A4) Fermentation process Next, the sterilized water which added the Bacillus-genus microbe is added, and the liquid mixture stirred is fermented on sealing conditions. From the viewpoint of efficient fermentation, the temperature condition is preferably 30 to 50 ° C, more preferably 37 to 50 ° C. The fermentation time is preferably 2 to 4 weeks.
After the fermentation process is finished, the mixture after treatment taken out from the fermentation chamber or the fermentation vessel is a high protein-containing organic substance in which the phorbol ester is decomposed by the action of Bacillus. In addition, components such as vitamins and minerals in the mixture after treatment are increased by the secondary action of Bacillus sp.

  Here, the reason why the fermentation time is set to 2 to 4 weeks will be described with reference to FIG. As shown in the figure, the degradation rate of phorbol ester is about 50% at one week after the start of the fermentation process, but about 80% after 2 weeks, and about 95% after 3 weeks. It is about 99% after 4 weeks. In this way, the longer the fermentation time, the higher the degradation rate of the phorbol ester, but if the treatment time is prolonged, the cost for maintaining the state increases, so the selection of the fermentation time that balances the degradation rate and cost is possible. Desired. As a result of the above examinations, it was found that the most efficient fermentation time is 2 to 4 weeks.

According to the method for producing a high protein-containing organic substance and feed according to this embodiment, phorbol ester can be decomposed and removed from organic substances containing phorbol ester at low cost and with high processing capacity, and is suitable for livestock feed. It becomes possible to produce a high protein-containing organic substance that can be used, and a feed using the same.
In addition, the fermentation time of the organic matter to be treated by the Bacillus genus can be optimized, and the factor that inhibits the phorbol ester decomposition action by the Bacillus genus can be removed. For this reason, the maximum amount of phorbol ester decomposing action can be obtained with a minimum amount of Bacillus sp. Input, and the cost required for removing the phorbol ester in the organic matter can be further reduced.

[Third embodiment]
Next, the configuration of the third embodiment of the present invention will be described with reference to FIG. The figure is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of this embodiment. This embodiment is different from the second embodiment in that the high protein-containing organic matter obtained in any of the embodiments of the present invention is not used as a Bacillus genus per se, but as an inoculum for fermentation. About another point, it can be set as the thing similar to 2nd embodiment.
That is, the (A1) mixing step and the (A2) high-temperature and high-pressure step in the method for producing a high protein-containing organic matter and feed according to this embodiment can be the same as those in the second embodiment.

  Unlike the second embodiment, the (A3) stirring step in the method for producing a high protein-containing organic matter and feed according to this embodiment is different from that of the second embodiment in that a solution obtained by adding Bacillus to sterile water is not added to the mixture. The high protein-containing organic substance obtained by the production method of the embodiment is added to sterilized water, and this is added to the sterilized mixed liquid obtained by the high-temperature and high-pressure step and stirred. In this way, it is not necessary to prepare a new Bacillus genus every time, and it is possible to reduce the cost required for producing a high protein-containing organic substance.

At this time, it is preferable to add 0.02-1 mass part of high protein containing organic substance with respect to 0.5-1 mass part of sterilized water. This is because a high fermentation efficiency can be realized at a low cost if the mixing ratio of the high protein-containing organic substance is set in this way. From such a viewpoint, the mixing ratio of the high protein-containing organic substance is more preferably 0.2 to 0.4 parts by mass.
And the (A4) fermentation process in the manufacturing method of the high protein content organic substance and feed of this embodiment can be made the same as that of 2nd embodiment.

  According to the method for producing a high protein-containing organic substance and feed according to this embodiment, it is not necessary to prepare a new Bacillus bacterium each time. For this reason, the total amount of Bacillus necessary for decomposing phorbol ester can be further reduced as compared with the case of the second embodiment, and as a result, the cost required for the production of high protein-containing organic matter and feed is further increased. It can be kept low.

[Fourth embodiment]
Next, the manufacturing method of the high protein content organic substance of 4th embodiment is demonstrated with reference to FIG. The figure is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of this embodiment.
In the present embodiment, an organic substance containing phorbol ester is mixed with a Bacillus bacterium and pre-cultured in advance, and the pre-culture product is added to the organic substance containing phorbol ester and subjected to main fermentation, whereby a high protein-containing organic substance is obtained. It differs from the second embodiment in that it is manufactured. About another point, it can be set as the thing similar to 2nd embodiment.

<Pre-culture>
(B1) First mixing step First, water is mixed with an organic substance containing a phorbol ester. At this time, it is preferable to mix 0.5-1.5 mass parts of water with respect to 2 mass parts of organic substance containing phorbol ester. This is because the efficiency of fermentation increases when the mixing ratio of water is set in this way. From such a viewpoint, the mixing ratio of water is more preferably 1 to 1.5 parts by mass.

(B2) First High-Temperature High-Pressure Step Next, similarly to the high-temperature high-pressure step in the second embodiment, a mixture of an organic substance containing phorbol ester and water is sterilized at high temperature and high pressure.

(B3) 1st stirring process Next, what added the Bacillus genus microbe to sterilized water is added to the above-mentioned sterilized liquid mixture, and is stirred. At this time, it is preferable to add 0.002-0.1 mass part of Bacillus genus bacteria with respect to 0.5 mass part of sterilized water. This is because uniform fermentation can be realized if the mixing ratio of the genus Bacillus is as described above. From such a viewpoint, it is more preferable that the mixing ratio of the genus Bacillus is 0.02 to 0.06 parts by mass.

(B4) Pre-culture process Next, the liquid mixture obtained by the 1st stirring process is fermented on airtight conditions. From the viewpoint of efficient fermentation, the temperature condition is preferably 30 to 50 ° C, more preferably 37 to 50 ° C. The fermentation time is preferably 1 to 7 days.

<Main fermentation>
(B5) Second mixing step Next, water is mixed with an organic substance containing phorbol ester. At this time, it is preferable to mix 2-4 mass parts of water with respect to 5 mass parts of organic substance containing phorbol ester. This is because the efficiency of fermentation increases when the mixing ratio of water is set in this way. From such a viewpoint, it is more preferable that the mixing ratio of water is 3 to 4 parts by mass.

(B6) Second High Temperature and High Pressure Step Next, similarly to the first high temperature and high pressure step, a mixture of an organic substance containing phorbol ester and water is sterilized at high temperature and high pressure.

(B7) Second stirring step Next, the preculture product obtained by preculture is added to sterilized water. And the sterilized water which added this preculture product is added and stirred to the liquid mixture sterilized by the 2nd high temperature / high pressure process.
At this time, it is preferable to add 1 to 4 parts by mass of the preculture product with respect to 1 part by mass of sterilized water. This is because if the mixing ratio of the precultured product is set in this way, high fermentation efficiency can be realized at low cost. From such a viewpoint, it is more preferable that the mixing ratio of the precultured product is 2 to 4 parts by mass.

(B8) Main fermentation process Next, the mixed liquid which added and stirred the Bacillus genus bacteria-containing water is fermented on airtight conditions. From the viewpoint of efficient fermentation, the temperature condition is preferably 30 to 50 ° C, more preferably 37 to 50 ° C. The fermentation time is preferably 2 to 4 weeks as described above in the second embodiment.
After the fermentation process is finished, the mixture after treatment taken out from the fermentation chamber or the fermentation vessel is a high protein-containing organic substance in which the phorbol ester is decomposed by the action of Bacillus. In addition, components such as vitamins and minerals in the mixture after treatment are increased by the secondary action of Bacillus sp.

As described above, according to the method for producing a high protein-containing organic material and feed according to the present embodiment, Bacillus spp. Can be efficiently grown by preculture, and the obtained preculture product is converted into an organic material containing phorbol ester. Since it can be added and fermented, the fermentation action can be promoted.
Thereby, it becomes possible to decompose | disassemble phorbol ester in organic substance more efficiently.

[Fifth embodiment]
Next, the manufacturing method of the high protein content organic substance of 5th embodiment is demonstrated with reference to FIG. The figure is a figure which shows the process of the manufacturing method of the high protein containing organic substance and feed of this embodiment.
In the present embodiment, an organic substance containing phorbol ester is mixed with a Bacillus bacterium and pre-cultured in advance, and the pre-culture product is added to the organic substance containing phorbol ester and subjected to main fermentation, whereby a high protein-containing organic substance is obtained. To manufacture. The fourth embodiment is different from the fourth embodiment in that the first and second mixing steps are not provided, and the mixing ratio of the bacterial cells and the water is different. About another point, it can be set as the thing similar to 4th embodiment.

<Pre-culture>
(C1) First High Temperature and High Pressure Step First, 10 parts by mass of an organic substance containing phorbol ester is sterilized at high temperature and high pressure. This kills microorganisms that can inhibit fermentation by Bacillus. This can be done in a general manner by autoclaving.

(C2) 1st stirring process Next, what added the Bacillus-genus microbe to sterilized water is added to the above-mentioned sterilized organic substance, and is stirred. At this time, it is preferable to add 0.01 to 0.6 parts by mass of Bacillus bacteria to 5 to 10 parts by mass of sterilized water. This is because uniform fermentation can be realized if the mixing ratio of the genus Bacillus is as described above. From such a viewpoint, it is more preferable that the mixing ratio of the genus Bacillus is 0.1 to 0.6 parts by mass.

(C3) Pre-culture process Next, the liquid mixture obtained by the 1st stirring process is fermented on sealing conditions. From the viewpoint of efficient fermentation, the temperature condition is preferably 30 to 50 ° C, more preferably 37 to 50 ° C. The fermentation time is preferably 1 to 7 days.

<Main fermentation>
(C4) Second High Temperature and High Pressure Step Next, as in the first high temperature and high pressure step, 100 parts by mass of organic matter containing phorbol ester is sterilized at high temperature and high pressure.

(C5) Second stirring step Next, the preculture product obtained by preculture is added to sterilized water. And the sterilized water which added this preculture product is added to the organic substance sterilized by the 2nd high temperature / high pressure process, and is stirred.
At this time, it is preferable to add 5-20 mass parts of precultured products with respect to 50-100 mass parts of sterilized water. This is because if the mixing ratio of the precultured product is set in this way, high fermentation efficiency can be realized at low cost. From such a viewpoint, it is more preferable that the mixing ratio of the precultured product is 10 to 20 parts by mass.

(C6) Main fermentation process Next, the mixed liquid which added and stirred the Bacillus genus bacteria-containing water is fermented on airtight conditions. From the viewpoint of efficient fermentation, the temperature condition is preferably 30 to 50 ° C, more preferably 37 to 50 ° C. The fermentation time is preferably 2 to 4 weeks as described above in the second embodiment.
After the fermentation process is finished, the mixture after treatment taken out from the fermentation chamber or the fermentation vessel is a high protein-containing organic substance in which the phorbol ester is decomposed by the action of Bacillus. In addition, components such as vitamins and minerals in the mixture after treatment are increased by the secondary action of Bacillus sp.

In the method for producing a high protein-containing organic material and feed according to this embodiment, unlike the fourth embodiment, water is added not to an organic material containing a phorbol ester but to a Bacillus bacterium. In this way, by increasing the amount of water that dissolves the bacteria as much as possible, in the organic matter to which the water in which the bacteria are dissolved is added, the bacteria are more uniformly dispersed over a wider area, and the fermentation becomes more uniform throughout the organic matter.
For this reason, it is anticipated that the fermentation efficiency of the whole organic substance will become high, and it becomes possible to decompose | disassemble the phorbol ester in organic substance more efficiently.

  Hereinafter, with respect to Examples and Comparative Examples of the method for producing high protein-containing organic matter and feed according to the present invention, and evaluation regarding the usefulness of the high protein-containing organic matter and feed thus obtained, refer to FIGS. explain. 11 and 12 are diagrams and graphs showing changes in the phorbol ester content in the organic matter in Examples and Comparative Examples, respectively. FIG. 13: is a figure which shows the breeding test result of the chicken by the feed of this invention, and a graph which shows average body weight transition. FIG. 14 is a diagram and a graph showing the feed intake of chickens by the feed of the present invention.

Example 1
Prior to the steps of the method for producing high protein-containing organic matter and feed, oil was extracted from Jatropha by the following steps to obtain a Jatropha residue.
First, 24 kg of Jatropha seeds were separated into a kernel part and a seed coat part using a molting machine, and only the kernel part was collected. The amount of kernel part collected was about 14.4 kg. Next, using a pulverizer, the kernel part was pulverized so as to have a diameter of about 2 mm.

  Next, using an electric oil press (Part No. S100-200, manufactured by San Seiki Co., Ltd.), the pulverized kernel portion is extracted, the oil is extracted, and the kernel wrinkle discharged from the oil press (= Jatropha residue) Were collected and allowed to cool naturally. The collected Jatropha residue was about 7.2 kg.

Next, pre-culture and main fermentation were performed according to the following steps to obtain a high protein-containing organic material in which phorbol ester was decomposed.
First, 10 g of Jatropha residue was sterilized with an autoclave at 120 ° C. for 15 minutes.
Next, Bacillus coagulans ( Bacillus coagulans , NBRC12583 strain, obtained from the National Institute of Technology and Evaluation Biological Genetic Resources Division) is used as a Bacillus bacterium, and 10 g of sterilized water containing 0.6 g of the cells is added. The mixture was added to the sterilized Jatropha residue and stirred well until the bacterial distribution became uniform, and then precultured at 37 ° C. for 3 days.

  Stirring was carried out by horizontally tilting and rotating a beaker containing a mixed solution to which Bacillus was added. This is because, when stirring with a spatula or the like, the solid organic matter collapses into a mud and the ventilation becomes worse. The stirring method is the same in the following examples and comparative examples.

Next, 100 g of Jatropha residue was sterilized by autoclaving at 120 ° C. for 15 minutes, and 50 g of sterilized water added with 10 g of a culture solution (culture product) obtained by pre-culture was added to the sterilized Jatropha residue to Stir well until the distribution is uniform.
Then, 25 g of water was added every 8 days, and the mixture was stirred and subjected to main fermentation at 50 ° C. for 24 days. As a fermentation product, fermented Jatropha, which is a high protein-containing organic substance in which phorbol ester was decomposed, was obtained.
Finally, 10% by weight of this high protein-containing organic substance was added to a chicken test feed (WYNMOORE Chickbooster, Breeders Business Group (Philippines)) to produce the feed of this example.

(Example 2)
Under the same conditions as in Example 1, the high protein-containing organic matter and feed of this example were produced.

(Examples 3 and 4)
In the same manner as in Example 1, except that Bacillus smithi ( Bacillus smithii , NBRC15311 strain, obtained from the National Institute of Technology and Evaluation, Biological Genetic Resource Department) was used as the Bacillus genus. Protein-containing organic matter and feed were produced.

(Examples 5 and 6)
As Bacillus, Bacillus subtilis ssp subtilis (Bacillus subtilis subsp. Subtilis, NBRC13719 strain, obtained by the National Institute of Technology and Evaluation Biological Resource Center) except using, in the same manner as in Example 1 Thus, a high protein-containing organic substance and feed of this example were produced.

(Examples 7 and 8)
In the same manner as in Example 1, except that Bacillus licheniformis ( Bacillus licheniformis , NBRC12200 strain, obtained from the National Institute of Technology and Evaluation Biological Genetic Resources Division) was used as the Bacillus genus. Protein-containing organic matter and feed were produced.

Example 9
In the same manner as in Example 1, except that Bacillus cereus ( Bacillus cereus , NBRC15305 strain, obtained from the Biological Genetic Resource Department, National Institute of Technology and Evaluation) was used as the Bacillus genus. Protein-containing organic matter and feed were produced.

(Examples 10 and 11)
Example except that Bacillus subtilis var. Natto (obtained from Miyagino Natto Factory, 4-29 Ginkgo-cho, Miyagino-ku, Sendai-shi, Miyagi) was used as the Bacillus genus. In the same manner as in Example 1, the high protein-containing organic matter and feed of this example were produced.

(Comparative Example 1)
First, 10 g of the Jatropha residue obtained in Example 1 was sterilized at 120 ° C. for 15 minutes by an autoclave.
Next, a bacteria added to the mixture, Lactobacillus Deruburikki ssp Deruburikki (Lactobacillus delbrueckii subsp. Delbrueckii, NBRC3202 strain obtained by the National Institute of Technology and Evaluation Biological Resource Center) was used to the bacteria 10 g of sterilized water to which 0.6 g of body was added was added to the sterilized mixed solution and stirred well until a uniform distribution was obtained, and then pre-cultured at 30 ° C. for 3 days.

Next, 100 g of Jatropha residue was sterilized by autoclaving at 120 ° C. for 15 minutes, and 50 g of sterilized water to which 10 g of a culture solution (culture product) obtained by pre-culture was added was added to the sterilized Jatropha residue, Stir well until the distribution is uniform.
And it fermented for 24 days at 30 degreeC, and fermented Jatropha which is a high protein content organic substance by which phorbol ester was decomposed | disassembled was obtained as this fermentation product.
In Comparative Example 1, addition of water every 8 days and stirring are not performed, and the fermentation temperature is 30 ° C. This is because in the bacterial species of Comparative Example 1, these conditions are suitable for fermentation.
Finally, 10% by weight of this high protein-containing organic substance was added to a chicken test feed (WYNMOORE Chickbooster, Breeders Business Group (Philippines)) to produce a feed for this comparative example.

(Evaluation)
<1. Degradation rate of phorbol ester>
The phorbol ester content (PE content) of the high protein-containing organic matter in Examples and Comparative Examples was measured every 8 days from the start date of the main fermentation to the 24th day, and the average degradation rate of the phorbol ester was calculated. . The results are shown in FIGS.

Here, in each of the examples and comparative examples, the organic substance containing phorbol ester was obtained by unshelling the seeds of Jatropha curcas L. and removing the internal seed kernel (kernel). Used. In addition, “bacterial species” in FIG. 11 indicates the species name of the bacterium used for decomposing the phorbol ester in the organic matter. The “strain” in Examples 10 and 11 indicates that it was obtained from the Miyagino Natto Factory (4-29 Ginkgo-cho, Miyagino-ku, Sendai City, Miyagi Prefecture) in Examples 10 and 11. For other examples, NBRC (NITE Biological It is obtained from the Resource Center) and its strain number. In “PE content (mg / g)”, the content of phorbol ester in the mixture in the fermentation process is shown every 8 days from the start of the main fermentation.

(1) Measuring method of PE content Content of the phorbol ester in the mixture in a fermentation process was measured as follows.
First, 20 ml of dichloromethane (Dichloromethane) was added to 1.5 g of a sample to be measured, pulverized with a homogenizer for 2 minutes, and centrifuged to collect the residue. To this, 20 ml of dichloromethane (Dichloromethane) was added, vigorously vibrated for 1 minute, and filtered through a centrifuge five times. All filtrates were collected and dried under a stream of nitrogen gas.

Next, this was dissolved in 5 ml of tetrahydrofuran, filtered through a 0.2 μm filter, and 20 μl was injected into HPLC (high performance liquid chromatography). WATERS (registered trademark) 2690 (manufactured by Waters Corporation) was used for HPLC, and WATERS 996 (manufactured by Waters Corporation) was used for the detector. HPLC was performed under the following conditions.
-HPLC column: symmetry C18, 3.5 μm, 100 × 4.6 mm
Solvent: A 1.75 ml o-phosphoric acid (85%) in 1 L distilled water, B Acetonitrile (HPLC grade), C Tetrahydrofuran (HPLC grade)
・ Flow rate: 1 ml / min ・ Process: (1) Solution A 60%, solution B 40% (15 minutes), (2) Solution A 25%, solution B up to 75% (15 minutes), (3) Solution B 100% (10 minutes), (4) C solution 100% (10 minutes)

(2) Degradation rate of phorbol ester As shown in FIG. 11, the PE content in Example 1-11 was 2.98 mg / g on the start day of the main fermentation. On the eyes and 24th day, the average values are 1.17 mg / g, 0.25 mg / g, and 0.14 mg / g, respectively.
Therefore, the phorbol ester decomposition rates on the 8th, 16th, and 24th days are 60.8%, 91.7%, and 95.4%, respectively.

On the other hand, the PE content in Comparative Example 1 was 2.98 mg / g on the start day of the main fermentation, but 2.08 mg / g, 1 day on the 8th day, 16th day, and 24th day, respectively. 0.78 mg / g and 1.66 mg / g.
Therefore, the phorbol ester decomposition rates on the 8th, 16th, and 24th days are 30.9%, 40.3%, and 44.3%, respectively.

From this result, it was revealed that phorbol ester can be efficiently decomposed by using a Bacillus genus as a bacterium for decomposing phorbol ester from an organic substance containing phorbol ester.
On the other hand, when Lactobacillus delbricki subspices delblick is used as a bacterium for degrading phorbol ester from organic matter containing phorbol ester, phorbol ester should be sufficiently removed. I couldn't.

<2. Chicken breeding test>
Using the feeds obtained in Examples and Comparative Examples, chicken chick growth tests were conducted at Palawan Agribusiness Development Foundation Inc. (Philippines) in the following manner.
In order to make sure that the chicken chicks on the 4th day after hatching are divided into groups of 3 chicks, divided into gauges A, B, and C, and fed to the feeds obtained in the examples, and that they grow sufficiently. The test was conducted.

  For Gauge A, a high protein-containing organic substance (processed meal) obtained by decomposing phorbol ester by Bacillus Smithy of Example 3 was used as a test feed for chicken WYNMOORE Chickbooster, manufactured by Breeders Business Group (Philippines). ) Was added to the feed obtained by adding 10%. Further, the high protein-containing organic substance obtained by decomposing phorbol ester by Bacillus subtilis subsp. Subtilis subtilis of Example 6 is used for gauge B, and the Bacillus subtilis varietas of example 11 is used for gauge C. A feed obtained by adding 10% of the high protein-containing organic substance obtained by decomposing phorbol ester with natto to the same test feed for chickens as gauge A was fed.

And it reared until 14th after hatching, and measured the body weight of each chicken at the start and every 7th day. All gauges were fed ceaselessly until the end of the test, and drinking water was free drinking.
The chicken chicks used were 9 female chicks with no history of vaccine hatched from broiler breeder eggs, which were preliminarily raised for 4 days. The change in body weight of each chicken is shown in FIG. 13, and the amount of feed intake is shown in FIG.

As shown in FIG. 13, it can be seen that chickens of any gauge are growing smoothly. Moreover, as shown in FIG. 14, feed intake is appropriately performed in any gauge.
Therefore, the high protein content organic matter obtained by decomposing phorbol ester in the organic matter using Bacillus sp. By the method for producing high protein content organic matter and feed of the present invention can be suitably used as feed. It became clear.

The present invention is not limited to the above-described embodiments and examples, and it goes without saying that various modifications can be made within the scope of the present invention.
For example, although Jatropha is used in the above embodiment, the present invention can be applied to other organic substances containing phorbol ester. Moreover, although said evaluation was performed about the chicken chick, the high protein content organic substance manufactured by this invention can also be used as a feed of a pig, a cow, a horse, and other livestock.

The present invention can be suitably used for producing livestock feed such as chickens.

Claims (11)

A method for producing a high protein-containing organic substance, comprising mixing an organic substance containing a phorbol ester and a Bacillus bacterium (excluding Bacillus natto) and fermenting, and decomposing the phorbol ester in the organic substance. . After mixing 0.5 to 3 parts by mass of water with 4 parts by mass of organic matter containing phorbol ester and sterilizing at high temperature and high pressure, Bacillus (excluding Bacillus natto) 0.004 to 0.2 parts by mass What is added to 0.5-1 mass part of sterilized water is added, and it ferments for 2 to 4 weeks at 30-50 degreeC. The manufacturing method of the high protein content organic substance of Claim 1 characterized by the above-mentioned. After mixing 0.5-3 mass parts of water with 4 mass parts of organic substance containing phorbol ester and sterilizing at high temperature and high pressure, the production method according to claim 1 or 2 is added to 0.5-1 mass part of water. What added 0.02-1 mass part of the obtained high protein content organic substance is added, and it ferments at 30-50 degreeC for 2 to 4 weeks. The manufacturing method of the high protein content organic substance characterized by the above-mentioned. Organic matter containing phorbol ester and Bacillus genus (excluding natto) are mixed and pre-cultured, then organic matter containing phorbol ester and Bacillus genus (excluding natto) are mixed, and this mixture The method for producing a high protein-containing organic material according to claim 1, wherein a precultured product is added to the product and fermented to decompose phorbol ester in the organic material. After mixing 2 parts by weight of an organic substance containing phorbol ester and 0.5 to 1.5 parts by weight of water and sterilizing at high temperature and high pressure, 0.5 parts by weight of sterilized water is added to the genus Bacillus (excluding Bacillus natto). Add 0.002 to 0.1 parts by mass, pre-culture at 30 to 50 ° C. for 1 to 7 days,
After mixing 5 parts by mass of organic substance containing phorbol ester and 2-4 parts by mass of water and sterilizing at high temperature and high pressure, 1 to 4 parts by mass of the precultured product obtained by pre-culture is added to 1 part by mass of sterilized water. The method for producing a high protein-containing organic material according to claim 4, wherein a main fermentation is performed at 30 to 50 ° C. for 2 to 4 weeks. After 10 parts by mass of an organic substance containing phorbol ester is sterilized at high temperature and high pressure, 5 to 10 parts by mass of sterilized water and 0.01 to 0.6 parts by mass of Bacillus (excluding Bacillus natto) are added. Pre-culture at 30-50 ° C. for 1-7 days,
After 100 parts by mass of an organic substance containing phorbol ester is sterilized at high temperature and high pressure, 50 to 100 parts by mass of sterilized water plus 5 to 20 parts by mass of a preculture product obtained by preculture is added, and 30 to 50 The method for producing a high protein-containing organic material according to claim 4, wherein the main fermentation is carried out at 2 ° C for 2 to 4 weeks. After squeezing after squeezing the seeds of Jatropha curcas L. as an organic substance containing the phorbol ester, or squeezing out the seed nuclei from the shells of the seeds A method for producing an organic substance with a high protein content according to any one of claims 1 to 6, wherein: A high protein-containing organic substance, wherein an organic substance containing a phorbol ester and a Bacillus genus bacterium (excluding Bacillus natto) are mixed and fermented to decompose the phorbol ester in the organic substance. After squeezing after squeezing the seeds of Jatropha curcas L. as an organic substance containing the phorbol ester, or squeezing out the seed nuclei from the shells of the seeds The high protein-containing organic substance according to claim 8, wherein: A method for producing a feed comprising mixing an organic substance containing a phorbol ester and a Bacillus bacterium (excluding Bacillus natto) and fermenting to decompose the phorbol ester in the organic substance. It contains a fermented product obtained by decomposing the phorbol ester by mixing and fermenting an organic substance containing a phorbol ester and a Bacillus genus (excluding Bacillus natto). feed.

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