WO2016060411A1

WO2016060411A1 - Method for producing organic mineral and nucleic acid composite

Info

Publication number: WO2016060411A1
Application number: PCT/KR2015/010677
Authority: WO
Inventors: 조향현; 김희경; 김영호
Original assignee: 조향현
Priority date: 2014-10-13
Filing date: 2015-10-08
Publication date: 2016-04-21
Also published as: KR20160043480A; KR101676327B1

Abstract

The present invention relates to a method for producing an organic mineral and nucleic acid composite, comprising the step of: mixing a nucleic acid monomer containing a purine base, or a powdery salt thereof, with a water-soluble powdery mineral having a negative enthalpy of solution and then reacting the resulting mixture by mixing same with water; or mixing a nucleic acid monomer containing a purine base, or a powdery salt thereof, with water, dissolving same, and then reacting the resulting aqueous solution by mixing same with a water-soluble powdery mineral having a negative enthalpy of solution. The organic mineral and nucleic acid composite produced using the method according to the present invention replaces inorganic minerals in a wide range of fields such as food, pharmaceuticals, feed, or fertilizers and, at the same time, has superior bioavailability compared with the inorganic minerals and thus can achieve the effect of sufficient mineral supply even by using a small amount thereof and prevent the occurrence of discoloration, off-flavors, and foul smells caused by using an excessive amount of the inorganic minerals.

Description

Method for preparing organic passivated mineral and nucleic acid complex

The present invention relates to a method for preparing an organic mineral source. In particular, the present invention relates to a method for preparing an organosibilized mineral and a nucleic acid complex from a nucleic acid.

Normal growth of flora and fauna requires a certain amount of minerals. Animals, including humans, also require minerals such as calcium, zinc, magnesium, potassium, iron, copper, selenium, chromium, molybdenum and iodine, and these minerals form a skeleton, although their proportion is very small in animal tissues, and osmotic pressure in the body Its role is to maintain the acid-base equilibrium of body fluids, and to participate as an activator of the enzyme system or as a component of the enzyme itself.

However, since minerals cannot be synthesized in the body, they must be supplied from the outside, and are mainly supplied in the form of inorganic minerals such as hydrochloride, sulfate, nitrate, phosphate and carbonate, but inorganic minerals have very low bioavailability.

Organic minerals, on the other hand, have high bioavailability, but face limitations in their widespread use as mineral sources for food, medicine, feed, and fertilizers due to high price or sanitary issues.

Korean Patent Publication No. 10-0513011 manufactures a soluble calcium-nucleic acid complex which combines calcium and macromolecule nucleic acid, but there may be a hygiene problem using the polymer nucleic acid extracted from salmon testis, and the manufacturing method is complicated. In addition, the production cost is high, but the production yield is low, there is a limit that it is uncertain whether the organic passivation of minerals other than calcium is possible.

In addition, Korean Patent Publication No. 10-1166546 manufactures an organo-calculated calcium-enriched whey protein by combining calcium with unsterilized whey powder, but it is not easy to secure unsterilized whey powder as a raw material, and production yield is high. There was a limit to the low, high heating costs involved in the heating process, and uncertainty about the organic passivation of minerals other than calcium.

The present invention is to provide a method for producing an organic mineral source of high bioavailability by chelation of minerals to nucleic acids.

The present invention comprises the steps of reacting a purine base-containing nucleic acid monomer or a salt powder thereof, and a water-soluble mineral powder negative soluble enthalpy, by mixing water to the mixture; Or mixing and dissolving a purine base-containing nucleic acid monomer or a salt powder thereof and water, followed by mixing and reacting a water-soluble mineral powder having a negative dissolution enthalpy in the aqueous solution. to provide.

The purine base-containing nucleic acid monomer has a purine base containing a hydroxyl group (-OH) at position 6 of the purine ring, combined with a water-soluble mineral powder having a dissolution enthalpy of negative to form an insoluble precipitate, and the yield of production of the organic passivated mineral and nucleic acid complex. It is preferable at this point. For example, the purine base-containing nucleic acid monomer may be any one or more nucleic acid monomers selected from inosinic acid, guanylic acid and xanthyl acid, preferably inosinic acid or guanylic acid.

Most preferably it is the sodium salt of inosine acid or the sodium salt of guanic acid.

The water-soluble mineral powder of which the dissolution enthalpy is negative can be selected by a person skilled in the art according to the type of the organic passivated mineral to be targeted. Water-soluble minerals with negative enthalpy of dissolution do not ionize when mixed with water, especially when mixed with a small amount of water, causing an exothermic reaction and forming chelate bonds when a substrate for organic passivation is present.

The soluble enthalpy of the water-soluble mineral powder may be mixed with a plurality of different minerals sequentially or simultaneously with a purine base-containing nucleic acid monomer or salt powder thereof.

The water-soluble mineral powder having a negative enthalpy of dissolution is well known to those skilled in the art. For example, calcium chloride, calcium carbonate, calcium lactate, or the like may be used for calcium, and preferably dissolution enthalpy is large calcium chloride.

The water is not particularly limited as long as it is water used in food, medicine, feed or fertilizer or additives thereof, but preferably deionized water is used to reduce the influence of other metal salts.

Mixing of the purine base-containing nucleic acid monomer or a salt powder thereof, a water-soluble mineral powder having a negative enthalpy, and water, and a mixture of the purine base-containing nucleic acid monomer or a salt powder thereof and a water-soluble mineral powder having a negative enthalpy thereof The ratio may be from 1: 5 to 95: 1 weight ratio, preferably from 1: 2 to 10: 1 weight ratio, more preferably from 1: 1 to 5: 1 weight ratio, and the chelated minerals of the organic passivated mineral and the nucleic acid complex Can be adjusted according to the target content of.

Regardless of the mixing order of the purine base-containing nucleic acid monomer or a salt powder thereof, a water-soluble mineral powder having a negative dissolution enthalpy, and water, a mixing ratio of the water-soluble mineral powder having a negative dissolution enthalpy and water is 1:50 to 2: 1 weight ratio. , Preferably from 1:10 to 1: 1 weight ratio. The amount of water may be appropriately adjusted for the reaction, but too much may be undesirable for chelation of minerals through exothermic reaction, and too little may limit the uniform hydration of a mixture of minerals and nucleic acids.

The reaction by the addition of water may be carried out at 10 to 100 ℃ for 30 minutes to 5 days. In particular, even if a separate heating in the reaction process is not advantageous to increase the production yield or increase the chelated mineral content, the reaction time need not be particularly limited, the time enough to complete the reaction, for example more than 1 hour , More than 2 hours, more than 4 hours, more than 6 hours, more than 12 hours can be adjusted, the maximum reaction time can be adjusted to 3 days, 2 days, 1 day.

*

When the chloride is used as the water-soluble mineral powder, the content of chlorine ions increases as the amount of the mineral chelated increases. This increased chlorine ions can be simply removed by a process of washing with water. For example, it is possible to lower the concentration of chlorine ions to harmless levels through 1 to 5 repeated washings with 0.5 to 10 times the volume of water used to dissolve calcium chloride, preferably 1 to 2 times the water. have. The washing may be performed by centrifugation to remove the supernatant after adding an appropriate amount of water, for example, centrifugation for 1 to 60 minutes at 500 to 50,000 rpm, and this centrifugation is performed at room temperature without setting temperature conditions. It may be, but may be carried out by setting the temperature to 25 ° C or less, preferably 15 ° C or less.

The organic passivated mineral and nucleic acid complex of the present invention is convenient to use by drying in a suitable form, such as hot air drying, spray drying, lyophilization and the like in powder form.

The present invention also provides an organic passivated mineral and nucleic acid complex which is prepared by the above method and is a chelate compound in which the mineral and the nucleic acid are chelate bound.

The mineral may be any one selected from calcium, zinc, magnesium, potassium, iron, copper, selenium, chromium, molybdenum and iodine or two or more complex minerals.

The present invention provides a method for preparing a highly bioavailable organic passivated mineral and nucleic acid complex by binding a mineral to a purine base-containing nucleic acid monomer. The organic passivated minerals and nucleic acid complexes prepared by the method of the present invention replace inorganic minerals in a wide range of fields such as food, medicine, feed or fertilizers, and have excellent bioavailability compared to inorganic minerals, thus providing sufficient mineral supply effect even in small amounts. To achieve this, it is possible to prevent the occurrence of discoloration, already odor, due to the use of excess mineral minerals.

In Experimental Example 1 of FIG. 1, A is a photograph after completion of the organic passivation reaction, and B is centrifuged for 20 minutes at 25 to 3,000 rpm for the organic passivation reaction, followed by mixing and suspending two times the weight of purified water with respect to the centrifuge precipitate. After centrifuged 20 times at 25 ℃ 3,000 rpm at 25 ℃ is a photograph after five times.

FIG. 2 is a photograph of the supernatant obtained after centrifugation of the organic passivated reactant at 25 ° C. for 20 minutes at 3,000 rpm for 24 hours at room temperature.

Figure 3 is a photograph of the organo-calcified calcium and nucleic acid complex obtained in order to evaluate the production yield by performing the washing process three times in B of Figure 1, 1 is Ca-IMP, 2 is Ca-GMP, 3 is Ca- AMP, 4 is Ca-CMP and 5 is Ca-MIX.

Figure 4 is a photograph of the inorganic mineral powder before mixing, IMP and GMP powder used as a substrate, and the organic passivated mineral and nucleic acid complex prepared in Experimental Example 3, 1: CaCl2, 2: FeSO4, 3 :: ZnSO4, 4: CuSO4 5: GMP, 6: Ca-GMP, 7: Fe-GMP, 8: Zn-GMP, 9: Cu-GMP 10: IMP, 11: Ca-IMP, 12: Fe-IMP, 13: Zn-IMP, 14 is Cu-IMP.

5 to 9 are the photographs of the upper part A after the end of the organic passivation reaction in Experimental Example 3, the lower part B is centrifuged at 3,000 rpm for 20 minutes at 25 ° C. for the organic passivation reaction. After washing and mixing twice the weight of purified water and centrifuging for 20 minutes at 25 ℃ at 3,000 rpm, it is a photograph after five times, 1 is a photograph of mineral powder, 2 is a reactant of mineral and GMP, 3 is a mineral and IMP 5 is a calcium chloride powder as a mineral, FIG. 6 is a ferrous sulfate powder as a mineral, FIG. 7 is a sodium sulfate selenite powder, and FIG. 8 is a copper sulfate powder as a mineral. Fig. 9 shows the use of zinc sulfate powder as a mineral.

10 is a photograph after completion of the organic passivation reaction in Experimental Example 4, the bottom B is centrifuged at 3,000 rpm for 20 minutes at 25 ° C. for the organic passivation reaction, and then twice the weight of purified water with respect to the centrifugation precipitate. Is a photograph after five times of mixing and suspending the centrifugation at 3,000 rpm for 20 minutes at 25 ℃, 1: Ca-IMP, 2: CaFe-IMP, 3: CaCu-IMP, 4: CaSe-IMP, 5: CaCr-IMP, 6: CaMn-IMP, 7: CaMo-IMP, 8: CaZn-IMP.

11 is a photograph of each organic passivated mineral and nucleic acid complex prepared in Experimental Example 4.

12 is a photograph after the end of the organic passivation reaction in Experimental Example 5, the bottom (after CF) is a photograph after five times the washing process centrifuged for 20 minutes at 3,000 rpm.

13 is a photograph of each of the organic passivated mineral and nucleic acid complexes prepared in Experimental Example 5, left A is M (7) -GHAC, and right B is M (7) -IHAC.

14 is a photograph after centrifugation at 25 ° C. and 4,000 rpm for 20 minutes after completion of the organic passivation reaction in Experimental Example 7; A is a photograph from Preparation Example 7.1-1 to Preparation Example 7.1-8, and B is Preparation Example 7.2-1. To manufacture example 7.2-8.

Mixing of the purine base-containing nucleic acid monomer or a salt powder thereof, a water-soluble mineral powder having a negative enthalpy, and water, and a mixture of the purine base-containing nucleic acid monomer or a salt powder thereof and a water-soluble mineral powder having a negative enthalpy thereof The ratio may be from 1: 5 to 95: 1 weight ratio.

Regardless of the mixing order of the purine base-containing nucleic acid monomer or a salt powder thereof, a water-soluble mineral powder having a negative dissolution enthalpy, and water, a mixing ratio of the water-soluble mineral powder having a negative dissolution enthalpy and water is 1:50 to 2: 1 weight ratio. Can be.

The reaction by the addition of water may be carried out at 10 to 100 ℃ for 30 minutes to 5 days.

The purine base-containing nucleic acid monomer may be any one or more nucleic acid monomers selected from inosinic acid, guanylic acid and xanthyl acid.

Hereinafter, the present invention will be described in detail by Examples, Experimental Examples and Preparation Examples. However, the following Examples, Experimental Examples and Preparation Examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following Examples, Experimental Examples, and Preparation Examples.

Experimental Example 1: Confirmation of the preparation possibility of organo-calcified calcium and nucleic acid complex according to the nucleic acid type

1) Sample preparation method

5'-cytidyl, a nucleic acid containing a purine base, 5'-inosinoate disodium (IMP), 5'-diso guanylate (GMP) and 5'-disodenylate (AMP), a pyridine base Disodium acid (CMP) and 5 g of nucleic acid (MIX) powders mixed in the same amount were each collected 5 g, and 10 g of purified water was mixed with stirring at 150 ° C. for 90 minutes at 25 ° C. to prepare an aqueous nucleic acid solution. .

5 g of calcium chloride powder [CaCl ₂ · 2H ₂ O (Junsei, Japan)] was mixed with each of the aqueous nucleic acid solutions while stirring at 25 ° C. for 150 minutes at 150 rpm, and allowed to react for 24 hours without additional heating.

2) Experiment result

After the completion of the organic passivation reaction, the organic passivation reaction was centrifuged at 25 ° C. at 3,000 rpm for 20 minutes, and then compared to before and after centrifugation to compare the evaluation of properties (precipitation, dissolution, coagulation, adhesion, and gelation). ++), severe (++), initial phenomenon occurs (+), no change (-) was confirmed by the results are shown in Table 1.

Table 1

Figure 1 A is a photograph after the end of the organic passivation reaction, B is centrifuged for 20 minutes at 25 ℃ to 3,000 rpm the organic passivation reaction, the centrifuged precipitate was mixed and suspended in 25 times the weight of purified water to 25 ℃ After washing five times centrifuged at 3,000 rpm for 20 minutes at 5 times. Figure 2 is the supernatant obtained after centrifugation for 20 minutes at 25 ℃ to 3,000 rpm the organic passivation reaction in B of Figure 1 at room temperature It is photograph when 24 hours passed.

When calcium was added to single or four kinds of mixed nucleic acids, the solubility appeared to be very good at the beginning, but after a certain time, the nucleic acid showed an exothermic reaction due to the organic passivation reaction with calcium, especially IMP and GMP. In the case of adding calcium chloride powder, the exothermic reaction proceeded greatly about 30 minutes, and the temperature of the aqueous solution rose to 60 ° C. or more.

According to the organic passivation reaction, IMP and GMP treatment groups were similarly insoluble with precipitation, flotation, adhesion and coagulation, and most of the AMP, CMP and MIX treatment groups showed solubility. This initial tendency did not show a significant difference even after 1 day, except that the CMP treatment group had a gradual organic passivation reaction, resulting in gelation.

The nucleic acids used in the above experiments were similar in their basic structure, but the AMP and MIX treatment groups inducing liquid phase after induction of the organic passivation reaction, the CMP treatment group having gelation properties, and calcium and chelating reactions It was divided into IMP and GMP treated groups with insoluble conditions.

The production yield was centrifuged to the organic passivated reactant, mixed and suspended with about twice the amount of purified water, and centrifuged for 20 minutes at 25 rpm at 3,000 rpm for 5 minutes. It was confirmed in% relative to the nucleic acid weight of the substrate used.

The production yield was 69.6% for Ca-IMP, 74.1% for Ca-GMP, 81.48% for Ca-AMP, 1.2% for Ca-CMP and 1.6% for Ca-Mix treatment group. In terms of production yield, the most effective organic passivation reaction was the AMP treatment group, followed by IMP followed by the GMP treatment group.

The organic passivation efficiency of the prepared organic passivated mineral and nucleic acid complex is shown in Table 2 by performing ICP analysis, it was identified by dividing into a total of 15 species including the target mineral compared to the original substrate.

TABLE 2

It was evaluated that there was little chelating effect of calcium in the precipitates of AMP and CMP treatment group. This was confirmed that calcium was separated into the supernatant, and Ca-IMP, which was insoluble in organic organase, was 81,569ppm, Ca-GMP was detected 84,498 ppm.

As a result of investigating the calcium content in the precipitate of Ca-Mix treated group which showed the liquefaction pattern, 43,393ppm was detected, and it was estimated to have chelating efficacy of half level of Ca-IMP or Ca-GMP, which was partially CMP and AMP. The reaction of chelating calcium minerals proceeded at the same time, and it was also estimated that the sediment and the supernatant were separated into the supernatant upon separation.

When 5 g of calcium chloride powder [CaCl ₂ · 2H ₂ O] was added to 5 g of IMP and GMP powder among nucleic acids, it was evaluated to exhibit organic passivation efficiency in the range of 50,000 to 70,000 ppm. It was also expected to be freely adjustable.

Experimental Example 2: Confirmation of preparation possibility of organo-calcified calcium and nucleic acid complexes according to the mixing order

1) Sample preparation method

Using the 5'-inosinonate disodium (IMP), which was excellent in organic passivation efficiency and high in production yield in Experimental Example 1, to determine whether the organic calcium passivation and nucleic acid complexes were generated according to the mixing order of IMP powder, calcium chloride powder and water. In order to perform the following experiment.

Preparation Example 2-1 is a mixture of 5 g of calcium chloride powder [CaCl ₂ · 2H ₂ O] and 10 g of purified water, and Preparation Example 2-2 is a mixture of 5 g of 5'-diisosinate (IMP) powder and 10 g of purified water, In Preparation Example 2-3, the calcium chloride aqueous solution of Preparation Example 2-1 and the IMP aqueous solution of Preparation Example 2-2 were mixed, and Preparation Example 2-4 was obtained by mixing 5 g of IMP powder with 15 g of the calcium chloride aqueous solution of Preparation Example 2-1. Preparation Example 2-5 was a mixture of 5 g of calcium chloride powder and 5 g of IMP powder and mixed with 10 g of purified water, and Preparation Example 2-6 was mixed with 5 g of calcium chloride powder and 5 g of IMP powder and mixed with 10 g of purified water. Heated at < RTI ID = 0.0 >

In Preparation Examples 2-1 to 2-6, the reaction was performed at room temperature without additional heating for 24 hours after the mixing or heat treatment.

2) Experiment result

Properties evaluation (precipitation, dissolution, coagulation, adhesion, gelation phenomenon), production yield and chelated calcium content in the precipitate were confirmed in the same manner as Experimental Example 1, Preparation Example 2-3 30 minutes after mixing the aqueous solution and aqueous solution After the temperature, and the rest of the preparation examples are shown in Table 3 to determine the exothermic reaction by measuring the temperature of the aqueous solution after 30 minutes after mixing the powder raw material and water.

TABLE 3

In Preparation Example 2-1 and Preparation Example 2-5, the exothermic reaction proceeded severely, so that the temperature of the aqueous solution rose to 55 ° C. or higher, but in Preparation Example 2-2, Preparation Example 2-3, and Preparation Example 2-4, No exothermic reaction was induced. In particular, when calcium chloride aqueous solution was prepared first, as in Preparation Example 2-3 or Preparation Example 2-4, and then mixed with IMP aqueous solution or IMP powder, no exothermic reaction was induced and insoluble organocalcified and nucleic acid complexes were not formed. .

When heated separately at 80 ° C. for 30 minutes as in Preparation Example 2-6, insoluble organocalcified and nucleic acid complexes were formed in the same manner as in Preparation Example 2-5, but there was no difference in production yield or calcium content. Since the preparation of the organic passivated mineral and nucleic acid complex was determined to react without the heat treatment process in consideration of the ease of operation and the reduction of energy costs.

Insoluble precipitate due to the combination of IMP or GMP and calcium in the nucleic acid from the results of Experimental Example 1 and Experimental Example 2 was found to be organic passivation by chelate bond rather than ionic bond, phosphorus (P) or phosphate group between the four nucleic acids Since the content of was not different, the amine group (-NH ₂ ) commonly possessed by IMP or GMP or the purine base including hydroxyl group (-OH) at position 6 of the purine ring plays a major role in chelating bond. It was expected to.

From the above results, it was found that using IMP or GMP as a substrate is appropriate in consideration of the production yield and the degree of chelate binding when inducing the organic passivation reaction of calcium with the nucleic acid as a substrate.

Experimental Example 3: Confirmation of the preparation possibility of the organic passivated mineral and nucleic acid complex according to the introduction of a single mineral

1) Sample preparation method

In Experimental Example 1, 5'-inosinoate disodium (IMP) or 5'-sodium guanylate (GMP) powder, which had excellent organic passivation efficiency and high production yield, was used as a substrate.

In addition to the calcium chloride powder of Experimental Example 1 as an inorganic mineral, zinc sulfate [ZnSO ₄ · 7H ₂ O (Bixsol, Korea)] powder, ferrous sulfate [FeSO ₄ · 7H ₂ O (Yakuri Pure Chemical)] powder, copper sulfate [CuSO ₄ · 5H ₂ O (Yakuri Pure Chemical)] powder and sodium selenite [Na ₂ SeO ₃ (XinXianShi Qiyuan Food Additive Co.)] powder were used.

2 parts by weight of the inorganic mineral was first powder-mixed to 10 parts by weight of the IMP or GMP powder, and 10 parts by weight of purified water was mixed with the mixed powder and reacted at room temperature with stirring at 500 rpm for 24 hours.

Centrifuging the organic passivated reactant reacted for 24 hours at 25 ° C. at 3,000 rpm for 20 minutes to obtain a precipitate, followed by mixing and stirring two times the weight of the precipitated purified water and centrifuging under the same conditions as above. Repeated 5 times to remove the unreacted nucleic acid and minerals, and then lyophilized to prepare an organic passivated mineral and nucleic acid complex. The prepared organic passivated mineral and nucleic acid complexes are abbreviated as '(mineral used)-(nucleic acid used)'.

As a control, IMP and GMP dissolved in purified water at 20% by weight were used, respectively.

2) Experiment result

Properties evaluation (precipitation, dissolution, coagulation, adhesion, gelation phenomenon), production yield and chelated mineral content in the precipitate were confirmed in the same manner as in Experiment 1, and the organic passivation efficiency was subjected to ICP analysis, to determine the target mineral compared to the original substrate The total chelated mineral content was identified as 'PS +' containing phosphorus (P) and sulfur (S) and 'PS-' except phosphorus and sulfur.

Table 4

4 is a photograph of the inorganic mineral powder before mixing, IMP and GMP powder used as a substrate, and the prepared organic passivated mineral and nucleic acid complex, 1: CaCl2, 2: FeSO4, 3: ZnSO4, 4: CuSO4, 5: GMP , 6: Ca-GMP, 7: Fe-GMP, 8: Zn-GMP, 9: Cu-GMP 10: IMP, 11: Ca-IMP, 12: Fe-IMP, 13: Zn-IMP, 14: Cu- IMP.

5 to 9, the upper part A is a photograph after the completion of the organic passivation reaction, and the lower part B is centrifuged at 3,000 rpm for 20 minutes at 25 ° C. for the organic passivation reaction, and then purified water of twice the weight of the centrifugation precipitate. Is a photograph after five times of mixing and suspending, centrifuging at 3,000 rpm for 20 minutes at 25 ℃, 1 is a photograph of mineral powder, 2 is a reactant of mineral and GMP, 3 is a reactant of mineral and IMP, FIG. 5 shows the use of calcium chloride powder as a mineral, FIG. 6 uses ferrous sulfate powder as a mineral, FIG. 7 uses sodium selenite powder, and FIG. 8 uses copper sulfate powder as a mineral. 9 uses zinc sulfate powder as a mineral.

First, 20% GMP and 20% IMP used as a control group were excellent in solubility in purified water and showed colorless water solubility.

In Experimental Examples 1 and 2, the organocalcified calcium and nucleic acid complexes formed insoluble precipitates by calcium and nucleic acid chelate bonds. In this experiment, calcium, iron, copper, and zinc were used as the substrates when GMP and IMP were used. All formed precipitates, but only selenium did not show precipitation.

In terms of production yield, Cu-GMP treated with copper sulfate compared to the weight of the substrate nucleic acid used for the first time was 104%, whereas Cu-IMP treated group showed about 60%, indicating a difference in yield according to the substrate. It was. The rest of calcium, iron and zinc showed high production efficiency ranging from 95% to 133% regardless of substrate.

The organic passivation efficiency of the prepared organic passivated mineral and nucleic acid complex is shown in Table 5 by performing ICP analysis, and identified by dividing into a total of 15 species including the target mineral compared to the original substrate.

Table 5

In Table 5, the content of phosphorus uniquely possessed by the GMP and IMP controls was about 50,000 ppm, the calcium content was 36 ppm of GMP, no IMP was detected, iron was 2.82 ppm, and IMP was 5.92 ppm. Was detected, zinc, copper and selenium were not detected.

In the case of Ca-GMP, 41,000 ppm of phosphorus was detected, and the calcium content was increased to 62,000 ppm after the organic passivation reaction. The Ca-IMP content was 50,000 ppm and the calcium content was 58,000 ppm after the organic passivation reaction.

Iron content in Fe-GMP was 36,720ppm, Fe-IMP was 42,430ppm, and phosphorus was detected in the range of 47,000-48,000ppm.

The zinc content in Zn-GMP and Zn-IMP was about 11,000 ppm, with 49,000 to 54,000 ppm of phosphorus detected.

The copper content in Cu-GMP and Cu-IMP both exhibited a chelating ability of over 100,000 ppm, with phosphorus concentrations of 48,000-51,000 ppm.

From the above results, it was confirmed that even when the nucleic acid powder and the mineral powder were mixed in a 5: 1 weight ratio, the organic passivated mineral and the nucleic acid complex having increased mineral content were formed in a high yield.

Experimental Example 4: Confirmation of preparation of organic passivated mineral and nucleic acid complex by simultaneous introduction of two minerals

1) Sample preparation method

In Experimental Example 1, 5'-inosinoate disodium (IMP) powder having excellent organic passivation efficiency and high production yield was used as a substrate.

In addition to the calcium mineral powder, zinc sulfate powder, ferrous sulfate powder, copper sulfate powder, and sodium selenite powder of Experimental Example 3, four kinds of mineral powders were additionally used as inorganic minerals. The four minerals are magnesium sulfate [MgSO ₄ ] powder, potassium chloride [KCl (large crystallization)] powder, chromium chloride [CrCl ₃ · 6H ₂ O (large crystallization)] powder and sodium molybdate [MoNa ₂ O ₄ ( Sigma-Aldrich)] powder was used.

10 parts by weight of calcium chloride powder is mixed with 20 parts by weight of IMP powder, or 10 parts by weight of calcium chloride powder and 10 parts by weight of the other mineral powder are mixed with 20 parts by weight of IMP powder, and 30 parts by weight of purified water is mixed with the mixed powder. The reaction was carried out for 24 hours while stirring at 500 rpm at room temperature.

After the reaction, the washing method, the preparation method of the organic passivated mineral and the nucleic acid complex, or the abbreviated description method are the same as in Experimental Example 3.

2) Experiment result

Table 6

TABLE 7

10 is a photograph after completion of the organic passivation reaction, and the bottom B shows centrifugation of the organic passivation reaction at 25 ° C. at 3,000 rpm for 20 minutes, followed by mixing and suspending twice the weight of purified water with respect to the centrifugation precipitate. After centrifuged for 5 minutes at 25 ° C. at 3,000 rpm for 20 minutes, the photographs were taken five times, including: 1: Ca-IMP, 2: CaFe-IMP, 3: CaCu-IMP, 4: CaSe-IMP, 5: CaCr- IMP, 6: CaMn-IMP, 7: CaMo-IMP, 8: CaZn-IMP.

11 is a photograph of each of the prepared organic passivated mineral and nucleic acid complexes.

When calcium chloride powder and other mineral powder were mixed with the IMP powder and reacted, it was found that the two kinds of minerals chelate each to form an organic passivated mineral and a nucleic acid complex. In addition, the production yield was significantly increased from 39% to 213% than using calcium chloride powder alone.

CaCu-IMP was slightly lower in the calcium content based on Ca-IMP at 78,870 ppm (0.96 times) and 76,574 ppm (0.93 times) for CaMn-IMP, but the calcium content was increased by the other two complex mineral reactions. Increased by 1.2 to 1.75 times, it was found that the organic passivation efficiency is increased through mutual promotion rather than competition of chelate bonds due to the complex use of minerals.

Compared with Ca-IMP where no minerals other than calcium were used, iron in CaFe-IMP was 69,390 ppm (Ca-IMP 19 ppm), and in CaCu-IMP, copper was 99,450 ppm (Ca-IMP ND) and CaSe-IMP. Selenium is 279,000 ppm (Ca-IMP 23 ppm), CaCr-IMP is chromium 68,550 ppm (Ca-IMP 1.72 ppm), and CaMn-IMP is 85,500 ppm (Ca-IMP 3.23 ppm) and magnesium is CaMg-IMP At 36,230 ppm (Ca-IMP 73 ppm) and CaZn-IMP for zinc, 91,724 ppm (Ca-IMP 8 ppm), when calcium is used as a catalyst, all of the different minerals show very high organic passivation efficiency at the same time. Was evaluated.

In particular, IMP powder alone and organolated selenium and nucleic acid complex formation was impossible, but it was confirmed that the organic passivation of selenium also proceeded when the mixture of calcium chloride powder and sodium selenite powder was reacted.

Therefore, this result is an organic passivation method capable of simultaneously increasing the organic passivation efficiency of heterogeneous minerals, including calcium, with increased production yield when using calcium as a catalyst.

Experimental Example 5: Confirmation of preparation of organic passivated mineral and nucleic acid complexes by simultaneous introduction of seven minerals

1) Sample preparation method

In Experimental Example 1, 5'-inosinoate disodium (IMP) powder or 5'-sodium guanylate (GMP) powder, which had excellent organic passivation efficiency and high production yield, was used as a substrate.

As inorganic minerals, calcium chloride powder, zinc sulfate powder, ferrous sulfate powder, copper sulfate powder, sodium selenite powder, magnesium sulfate powder, and potassium chloride powder of Experimental Example 4 were used.

1 part by weight of each of the seven mineral powders was mixed with 10 parts by weight of IMP powder or GMP powder, and then 100 parts by weight of purified water was mixed with the mixed powder and reacted at room temperature with stirring at 500 rpm for 24 hours. The reaction of IMP as a substrate was abbreviated as 'M (7) -IHAC' and the reaction of GMP as a substrate was abbreviated as 'M (7) -GHAC'.

2) Experiment result

Properties evaluation (precipitation, dissolution, coagulation, adhesion, gelation phenomenon), production yield and chelated mineral content in the precipitate were confirmed in the same manner as in Experiment 1, and the organic passivation efficiency was subjected to ICP analysis to target target minerals compared to the substrate The total chelated mineral content was identified as 'PS +' containing phosphorus (P) and sulfur (S) and 'PS-' except phosphorus and sulfur.

Table 8

Table 9

The upper part (after reaction) of FIG. 12 is a photograph after completion | finish of an organic passivation reaction, and the lower part (after CF) is a photograph after performing the washing | cleaning process 5 times centrifuged at 3,000 rpm for 20 minutes.

FIG. 13 is a photograph of each of the prepared organic passivated mineral and nucleic acid complexes. The left A is M (7) -GHAC and the right B is M (7) -IHAC.

When the seven minerals were simultaneously treated with IMP or GMP, M (7) -GHAC showed 158% of M (7) -IHAC and 162% of M (7) -IHAC.

Comparing the total organic passivation amount and the detection efficiency with respect to the addition of the seven inorganic minerals, M (7) -GHAC contains about 14.1% of phosphorus (P) and sulfur (S). : 73.7%), and 15 total minerals excluding phosphorus and sulfur were 16.1% (68.8% of organic passivation efficiency compared to mineral input). In the case of M (7) -IHAC, the range of about 23.2% (organic passivation efficiency of mineral input: 59.6%) and 10.5% ((organic passivation efficiency of mineral input: 52.9%) was similarly high regardless of nucleic acid type. Organic passivation efficiency was shown.

The organic passivation efficiency of each mineral was compared with that of minerals, and it was found that calcium is 67.7 ~ 74%, iron is 99.8 ~ 100%, zinc is 76 ~ 97.9%, copper is 97.7 ~ 99.1%, and selenium is 73.6. Organic passivation efficiencies of ~ 91.1%, 7.79 ~ 24.6% for potassium, and 22 ~ 56.6% for magnesium, but IMP-based substrates were somewhat lower.

Experimental Example 6: Evaluation of External Resistance of Organostabilized Minerals and Nucleic Acid Complexes

1) Experiment Method

Four kinds of Ca-GMP, Fe-GMP, Zn-GMP and Cu-GMP, which are organic passivated mineral and nucleic acid complexes prepared in Experimental Example 4, and four kinds of Ca-IMP, Fe-IMP, Zn-IMP and Cu-IMP A total of eight species were evaluated for acid resistance and alkali resistance.

4 ml of each prepared pH solution was added to a 5 ml volume of Eppendorf tube, and 0.5 g of each of the prepared organic passivated mineral and nucleic acid complexes was added thereto, followed by sufficient dispersion and sonocation. After fermentation, the change of appearance was evaluated by sedimentation, dissolution, elution and gelation. The most severe case was +++, the severe case was ++, the initial phenomenon occurred at +, and there was no change. The cases were evaluated by dividing by.

After the evaluation of the properties, centrifugal (3,000rpm, 20 minutes, 25) treatment, only the supernatant was passed through a 0.25 μm filtration filter to compare the amount of minerals present in the supernatant through ICP analysis. Confirmed.

2) Experiment result

Table 10 shows the results of pH stability experiments of the organic passivated mineral and nucleic acid complexes.

Table 10

All test groups were evaluated to be stable within the pH 2-10 range. In case of Cu-GMP, the dissolution and gelation of minerals occurred severely while dissolving in the range of pH 1, which is strongly acidic, but this phenomenon did not occur when the pH was neutral and neutral.

We examined whether there was another test zone that caused the change of properties related to mineral separation and elution under acidic conditions. Among them, Fe-GMP and Fe-IMP were included. The phenomenon became more severe.

Table 11 shows the results of the long-term storage stability of the pH through the analysis of the content of the minerals eluted in the supernatant after the evaluation of the properties of the organic passivated calcium and nucleic acid complex.

Table 11

Ca-HAC materials, namely Ca-IMP (Ca-IHAC) and Ca-GMP (Ca-GHAC), were evaluated to have safety over the entire pH range. In other words, when comparing total minerals including calcium, Ca-GHAC showed 90-231ppm in total sum including P and S and 41-190ppm minerals minus phosphorus (P) and sulfur (S). In the case of using IMP as a substrate, only slight values of 71-114 ppm and 54-106 ppm were confirmed.

* The results of the long-term storage stability test results of the pH of the minerals eluted in the supernatant after evaluation of the properties of the organic iron iron and nucleic acid complexes are shown in Table 12.

Table 12

Fe-HAC material showed slightly higher mineral distribution compared to Ca-HAC material, but it was evaluated to have safety in the whole range. In other words, compared to only iron, Fe-GMP was investigated to be 2ppm at pH 7, 1ppm Fe-IMP was estimated to have little elution effect in neutral conditions, the amount of elution is increasing toward acidic and basic conditions Although evaluated, the levels of minerals contained in the sediment retained chelate bonds of more than 30,000 ppm of Fe-GMP (Fe-GHAC) and 42,000 ppm of Fe-IMP (Fe-IHAC). It was found to have basicity.

Table 13 shows the pH long-term storage stability test results through the analysis of the content of the minerals eluted in the supernatant after the evaluation of the properties of the organic passivated copper and nucleic acid complex.

Table 13

When comparing the safety of Cu-HAC material with only copper, Cu-GHAC was found to be 20ppm at pH 7 and 1.27ppm of Fe-IHAC, which showed little dissolution effect in neutral conditions. As the conditions were increased, the elution rate was estimated to increase within the range of 1,000-3,000 ppm, and the safety condition was evaluated to be the pH range of 4-10.

The levels of minerals contained in the sediments preserved the organic efficiency of more than 100,000 ppm in both Cu-GHAC and Fe-IHAC and their elution values for 10 days were evaluated as having acid and base resistance.

After evaluating the properties of the organo-zinc and nucleic acid complexes, the results of the long-term storage stability test results of the pH of the minerals eluted in the supernatant are shown in Table 14.

Table 14

When Zn-HAC materials were compared with zinc only, Zn-GHAC was found to have 25ppm at pH 7 and 366ppm at Zn-IHAC.

Investigation of the resistance to acid and base resistance showed that Zn-GHAC materials had excellent base resistance as the elution amount decreased to 3 ppm as the base condition rather than acid resistance. However, Zn-IHAC materials were found to have both acid and base resistance at the same time, showing a little difference than other materials.

Although the levels of minerals contained in the sediments preserved organic efficiency of more than 10,000 ppm in both Zn-GHAC and Fe-IHAC and the results of elution values for 10 days, they were slightly higher. It was assessed to possess.

Experimental Example 7: Confirmation of preparation of organic passivated mineral and nucleic acid complex of the polymer nucleic acid polymer

1) Confirmation of yeast extract characteristics

end. Molecular Weight

S. cereviase isolated from extract of freeze-dried sample and C.utilis extracted from OKK110426 extract (JP KOHJIN Co., product name: AROMILD) of each were prepared, as cross pyojunche FPLC analysis powder GMP (1g) + powder CMP (1g) + Powder After mixing AMP (1g) + powder IMP (1g), FPLC analysis was performed on the composition obtained by FPLC analysis of 50g mass up of purified water and 1% (w / w) dilution of NANA, whose molecular weight is 309. A NANA test zone was used.

The FPLC system (AKATA, Sweden) was measured in a Superdex Pep 10 / 300GL column using UPC900 + D920 + CU950, phosphate buffer, flow rate 0.5ml / min, pressure 0.84PSI and the results are shown in Table 15.

Table 15

Based on NANA (MW 309), a standard that knows the molecular weight, based on nucleic acids (5 mixtures) contained in yeast extract, the molecular weight detection time was analyzed by FPLC analysis. When the molecular weight was 309, it was 42.44 min. Detected in.

On this basis, if the S.cereviase CKK110426 in the case of 90% in 41.83min. The Aromild was 90% detected by the 42.08min..

Considering that the molecular weight of NANA is 309 and considering that the molecular weight of the nucleic acid is 392.17 (IMP) to 407.20 (GMP), it can be seen that 69% of the yeast extract is a polymer having a molecular weight of 1,000 Da or more when calculated in the formula.

I. Nucleic acid content

The content of nucleic acid in yeast extracts was 10.5%, GMP 10.5%, CMP 7.2% and

UMP

8,2% in Aromild (KOHJIN, Japan) compared to 19 standard nucleic acids. Was not detected, and the total nucleic acid was 36.4%. Nucleic acids in the S. cereviase CKK110426 extract were assayed and found to contain 6.57% CMP, 8.21% ppm for UMP, 4.89% for GMP, 2.74% for IMP, and 0.66% for AMP. Was%.

All. Mineral content

Mineral content in yeast extract is shown in Table 16 by the IPC analysis.

Table 16

2) Sample preparation method

end. Preparation of Samples by Non-thermal Treatment

A) Manufacturing Example 7.1-1

An experiment was conducted to determine whether yeast extract, an organic passivation substrate, forms precipitates upon dissolution. For this, mixed purified water with 6.66 g of yeast extract was mixed, but stirred after mass-up at a rate of 30 g (150rpm, 1 hour, 25 C) was dissolved.

B) Preparation example 7.1-2

In order to investigate the insoluble precipitate formation and the reaction pattern related to the organic passivation reaction according to the mixing reaction of inorganic iron powder with GMP, the organic passivation substrate, 0.040g of ferrous sulfate powder of Experimental Example 3 and 6.66g of GMP powder were mixed. After that, the mixture was mixed with 32 g of purified water and stirred (150 rpm, 1 hour, 25 ° C.).

C) Preparation Examples 7.1-3 to 8

As a result of increasing the amount of inorganic minerals added as a yeast extract ( S.cereviase CKK110426) substrate, the characteristics change and yield patterns of the organic passivation reaction were investigated.

Preparation Example 7.1-3 was prepared by mixing 6.66 g of yeast extract with respect to ferrous sulfate powder 0.040 g (pure Fe content: 0.008 g), and adding 28.6 g of purified water to the organic solvent through stirring (150 rpm, 1 hour). Induced the passivation reaction.

Production Example 7.1-4 was carried out in the same manner as in Production Example 7.1-3 except that 0.080 g (pure Fe content: 0.016 g) was added to the ferrous sulfate powder.

Production Example 7.1-5 was carried out in the same manner as in Production Example 7.1-3 except that 0.12 g of ferrous sulfate powder (pure Fe content: 0.024 g) was added.

Production Example 7.1-6 was carried out in the same manner as in Production Example 7.1-3 except that 0.16 g (pure Fe content: 0.032 g) was added to the ferrous sulfate powder.

Production Example 7.1-7 was carried out in the same manner as in Production Example 7.1-3 except that 0.20 g (pure Fe content: Fe 0.04 g) was added to the ferrous sulfate powder.

Production Example 7.1-8 was carried out in the same manner as in Production Example 7.1-3 except that 0.30 g (pure Fe content: Fe 0.06 g) was added to the ferrous sulfate powder.

I. Preparation of Samples by Heat Treatment

Preparation Examples 7.2-1 to 7.2-8 are carried out in the same manner as Preparation Examples 7.1-1 to 7.1-8, but after the final mixing process, only a step of heating with stirring at 150 rpm for 30 minutes at 80 ° C. is further added. It was.

3) Experiment result

Appearance evaluation (precipitation, dissolution, coagulation, adhesion, gelation phenomenon), production yield and chelated mineral content in the precipitate were confirmed in the same manner as in Experiment 1.

Table 17

Table 18

14 is a photograph after centrifugation at 25 ° C. and 4,000 rpm for 20 minutes after completion of the organic passivation reaction. A is a photograph from Preparation Example 7.1-1 to Preparation Example 7.1-8, and B is Preparation Example 7.2-1 to Preparation Example 7.2. It is a photograph up to -8.

In Preparation Example 1-1, in which only a single yeast extract substrate containing high nucleic acid was dissolved in purified water, the solubility was excellent, but the organic passivation reaction with the ferrous sulfate powder added separately was not induced, because it was combined with Mine contained in the yeast. .

In the case of inducing the organic passivation reaction of Fe using the GMP powder of Preparation Example 7.1-2, the yield was 131% compared to the substrate, and the physical property change related to the convenience of on-site production was observed. The precipitation effect was also evaluated as excellent.

Concentration gradients of ferrous sulfate powder from low concentration (one-fold basis) to high concentration (7.5-fold) were applied to the yeast extract of the same amount compared to the nucleic acid GMP substrate, and the yield of organic passivation was 11.9% when low concentration was added. The pattern was gradually increased, and production yield of 25,8% was obtained in Preparation Example 8, which was added up to 7,5 times. From the above results, it was found that the production yield when using the yeast extract is significantly lower than that of the GMP powder as a substrate, which is not suitable as a substrate for organic passivation.

Even when heat treatment conditions were added in Preparation Examples 7.2-1 to 7.2-8, no increase in production yield was observed in the yeast extract or in the GMP powder substrate.

A pattern similar to the example was shown.

Claims

Mixing a purine base-containing nucleic acid monomer or a salt powder thereof, and a water-soluble mineral powder having a negative enthalpy, and then reacting the mixture by mixing water; Or mixing and dissolving a purine base-containing nucleic acid monomer or a salt powder thereof and water, followed by mixing and reacting a water-soluble mineral powder having a negative dissolution enthalpy in the aqueous solution.
The purine base-containing nucleic acid monomer or salt powder thereof, and the enthalpy thereof are negative, regardless of the mixing order of the purine base-containing nucleic acid monomer or salt powder thereof, the water-soluble mineral powder having a negative dissolution enthalpy, and water. The mixing ratio of the phosphorus water-soluble mineral powder is 1: 5 to 95: 1 weight ratio method for producing an organic passivated mineral and nucleic acid complex.
The mixing ratio of water-soluble mineral powder and water of claim 1, wherein the purine base-containing nucleic acid monomer or a salt powder thereof, a water-soluble mineral powder having negative dissolution enthalpy, and water are mixed in an order of 1: Process for producing an organic passivated mineral and nucleic acid complex, characterized in that 50 to 2: 1 by weight.
The method of claim 1, wherein the reaction by adding water is carried out at 10 to 100 ° C. for 30 minutes to 5 days.
The method according to claim 1, wherein the purine base-containing nucleic acid monomer is at least one nucleic acid monomer selected from inosinic acid, guanylic acid and xanthyl acid.
An organic passivated mineral and nucleic acid complex prepared by the method of any one of claims 1 to 5, wherein the mineral and nucleic acid are chelate-bonded chelating compounds.
According to claim 6, The mineral is an organic passivated mineral and nucleic acid, characterized in that any one selected from calcium, zinc, magnesium, potassium, iron, copper, selenium, chromium, molybdenum and iodine or two or more complex minerals Complex.