KR20180120634A - Delayed release enzyme preparations and the preparation method for the same - Google Patents

Abstract

The present invention relates to an enteric digestive enzyme composition using a delay-type emission system, and to a manufacturing method thereof, in which hydroxypropyl cellulose phthalate, as a delay-type emission polymer, is coated on the enzyme composition manufactured by solid fermentation and the core of granules of hardening oil to improve digestion rate of livestock feeds and increase biological availability.

Description

FIELD OF THE INVENTION [0001] The present invention relates to a digestive enzyme for enteral use,

The present invention relates to the field of animal feed, and more particularly, to an animal-derived digestive enzyme containing an enzyme as an active ingredient, and to a method for producing the same.

Enzymes are natural compounds and have no toxicity. They are widely used as feed additives and help increase animal productivity. Adding β-glucanase to the feed can improve the broiler's growth rate and feed efficiency because it allows the barley starch and protein to be fully utilized in the small intestine. In addition, the addition of xylanase to weaned diets has an effect of increasing building and nitrogen utilization as compared with the control group.

In addition to the general availability of these enzymes, amylase and protease, the digestive enzymes used to increase productivity, are extremely unstable in the production of both enzymes and require external supply do. In the case of piglets, the digestive enzyme system changes rapidly around the reason. When digestive organs are not completely developed, the digestive enzymes cause an increase in digestibility. In particular, feeding of digestive enzymes through feed during this period can help minimize the digestibility of piglets. However, the enzymes supplied through the feed are affected by the stomach acid during the passage through the stomach, so that the effect of enzyme addition is not maximized. In addition, the pellet processing method for increasing the availability of feed ingredients may lead to the depolymerization of the enzyme only at the temperature at which the pellet is produced, and development of a technique for increasing the efficiency of the digestive enzyme is required.

As described above, when the enzyme is simply added to the feed after the development and production of the enzyme, the enzyme is exposed to stomach acid after ingestion, which causes destabilization at low pH, mixing with simple feed, and inactivation during the molding process. And improve the efficiency of the enzyme to maximize its effect.

In general, in the field of pharmacy, a delayed-release dosage form utilizing enteric coating or the like is actively utilized as a means for absorbing drugs or exhibiting its drug efficacy at a small intestine or below . The nature of the drug is that the drug is degraded or inactivated by gastric acid with a low pH, or when the main absorption or efficacy of the drug is in the small intestine or large intestine, it is effectively utilized as a means for delivering the drug to a desired site.

For example, digestive enzymes, especially pancreatin, are digestive enzymes with the activity of amylase, protease, and lipase, isolated from the pancreas of mammals such as cattle and pigs. So that the digestive activity can be promoted in the upper part of the small intestine by being activated only in neutral to weakly alkaline pH which is similar pH condition. However, due to pancreatic insufficiency, pancreatin ingested from the outside of the patient must pass through the stomach, so that it is irreversibly inactivated by gastric acid and pepsin. Therefore, an orally administered enzyme requires a suitable protective barrier to prevent it from being inactivated in the stomach, and enteric coating is optimal.

In the past, complex enzyme digestion enzymes including pancreatin were coated with sugar coating or enteric coating from the top to the components that need to express the drug efficacy, so that sufficient effect was not obtained. However, recently, only pancreatin It has been designed and developed into a form that can be used in combination with other enzymes after intestinal coating, and is on the market as a pharmaceutical product.

There are several types of enteric coating machines that can be used depending on the target point of drug release. Shellac begins to dissolve slowly in slightly acidic conditions and is solubilized at pH 7.0 and above and at the same time has the advantage of protecting the destabilization of the drug due to moisture absorption. Cellulose acetate phthalate (CAP) dissolves at pH 6.0 or higher And has a property showing somewhat hygroscopicity. If necessary, the entire duodenum may be targeted by applying hydroxypropylmethyl cellulose phthalate (HPMCP-50) or HPMCP-55, which can rapidly induce drug release from the upper part of the duodenum. Methacrylic acid copolymers, especially Eudragit S, are also widely used for the purpose of drug release in the large intestine.

On the other hand, in order to more efficiently apply to the human body, the enteric coating agent having the precise conditions as described above is used. However, in view of economical efficiency and the like, alginic acid, ethylcellulose, methylcellulose Agar, carrageenan, pectin, and guar gum), chitin or chitin acid or an artificially processed analog thereof, soy protein or wheat protein or an artificially processed analog thereof, chitin or chitin acid or an artificially processed analog thereof, agar, carrageenan, Gums and the like, which can be used. However, when compared with enteric coating agents that respond quickly to pH changes, they are low in drug release efficiency.

Korean Patent Application No. 10-1999-0005822 discloses a method for producing enteric-coated granules by first coating pancreatin with seed at a low temperature and then performing secondary coating if necessary.

Korean Patent Application No. 10-2005-7009145 discloses a granular composition containing an enzyme and a hardening oil in the core of saccharides.

Korean Patent Application No. 10-1988-0700451 describes a formulation in which an enzyme is microgranulated to encapsulate with a water-soluble mechanical barrier and an enteric coated layer is applied thereto.

Korean Patent Application No. 10-2005-0067227 discloses a preparation in which a binding liquid containing pancreatin is injected into a spherical pellet and the enteric coating polymer is injected thereinto.

Patent Application No. 10-1999-0005822 Patent Application No. 10-2005-7009145 Patent Application No. 10-1988-0700451 Patent Application No. 10-2005-0067227

When the enzyme is simply added to the feed after the enzyme is developed and produced, the enzyme is exposed to the stomach acid and becomes unstable at a low pH, or is inactivated due to mixing with a simple feed, or heat during the pelleting process And some of them have been pointed out, and we are trying to maximize the efficiency by improving the efficiency of the enzyme.

If the manufacturing cost is not taken into consideration, it can be sufficiently implemented in the conventional technology, but it is very difficult to implement an appropriate technology that can be manufactured at a low cost considering mass production and consumption of livestock feed. In fact, the pelletizing process to produce the core for the acid-resistant coating is not only low in yield but also takes a long time to manufacture, and the amount of raw materials that can be added in a small amount in one production is limited. Accordingly, in consideration of mass production, it is necessary to introduce a granulation process capable of quickly and mass-producing particles used as a core in coating. However, since the formed granules are irregular in shape and the surface is not smooth, the thickness of the coating layer after coating on the surface of the granules is not constant, and the coating amount should be greatly increased. To solve this problem, we intend to develop an intestinal digestive enzyme which can be manufactured at low cost and has high acid resistance.

As a means for solving the problem to be solved, in the present invention, in an enteric digestive enzyme agent comprising an enzyme agent as an active ingredient, the enteric digestive enzyme agent is one in which a coating layer of hydroxypropylcellulose phthalate is formed on the surface of granules containing an enzyme agent and hardening oil A digestive enzymatic agent for enteral purposes is provided. Also, there is provided a method for producing an enteric digestive enzymatic agent comprising an enzyme as an active ingredient, comprising the steps of: preparing an association product in an environment in which an enzyme and a hardened oil are not present in a solvent, And a step of forming a coating layer of methyl cellulose phthalate.

It is stable to the heat generated in the manufacturing process, increases the productivity by reducing the coating process, reduces the production cost due to low cost raw material and simple process, and is highly applicable to the industry. In the present invention, the intestinal digestive enzymes can be manufactured at a low cost, can be produced at low cost, have high acid resistance, and exhibit excellent results in actual specification tests, considering mass production and consumption of livestock feeds .

Therefore, the present invention can simultaneously improve efficiency and economical efficiency for application of enzymes as animal feeds and animal pharmaceuticals.

The present invention relates to an intestinal digestive enzyme utilizing a delayed-release system, and is made by applying a long-acting coagulant to a core composed of an enzymatic agent prepared by solid fermentation and a hardening oil.

The enzyme of the present invention may include conventional enzymes that can be used in the art, and examples thereof include amylase, beta-glucanase, cellulase, pectinase, phytase, protease, Lipase, and the like.

Particularly, in order to achieve the above-mentioned object, the active ingredient of the present invention is prepared by inoculating aspergillus oryzae as an enzyme produced by solid-state fermentation (SSF) And can be evaluated on the basis of protease as an indicator substance for evaluating the activity of an enzyme in an enzyme agent after fermentation.

The enzymes used in the present invention can be used in an amount of 5,000 U / g or more based on the protease, preferably 10,000 U / g. In addition, it is preferable that the protease is used in an amount of 30% (w / w) or more per unit production unit in the final formulation. Since the enzyme itself is relatively inexpensive, it is preferable to use a minimum amount of additives and the manufacturing process can be shortened.

On the other hand, a method of imparting acid resistance by coating an enzyme with an enzyme can be variously applied depending on the manufacturing method and the polymer used. First of all, according to the manufacturing method, the effect may be greatly varied depending on the method of applying the polymer to the enzyme. The granule-forming method of producing the enzyme-polymer granule so that the polymer protects the enzyme by mixing and bonding the polymer and the enzyme, Coating method and a coating-coating method which is a method of once coating an enteric core with an enzyme as an enteric polymer and then coating the enteric polymer again after preparing the pellet using a binder. In the latter case, since the inert core itself is spherical, the spherical shape is maintained even after the coating of the enzyme, so that the coating film is dense and constant after the application of the enteric polymer, and the acid resistance is high. However, when a large amount of an enzyme is required to be coated, the process of applying the enzyme is consumed for a long time, so that it is not suitable as a method for implementing the present invention. In the case of the granule-coating method, since the granules are produced by coalescence without using an additional coating agent using an appropriate binder, a large amount of endogenous enzyme can be produced within a short time, which is suitable for the present invention.

Examples of usable binders include cellulosic binders, synthetic binders, saccharides, animal-derived binders such as gelatin, and the like. However, the present invention uses hydrogenated oil which is inexpensive and does not require a post-association drying step. The hydrogenated oil is a white solid artificial fat made by adding hydrogen to a liquid oil such as fish oil or soybean oil. In the present invention, palm oil-based hydrogenated oil is used. Particularly, in the present invention, granules after the granulation process were irregular in shape and smooth on the surface, but showed a certain level of acid resistance even when a small amount of polymer was applied.

The curing oil used in the present invention has a melting point of 60 to 80 degrees and is considered to be in a liquid state upon heating and becomes a solid state upon cooling so that the drying process after the combination is not necessary and the cost is lower than that of a pharmaceutical binder And the present invention is applied to the present invention. Surprisingly, the use of the hydrogenated oil as a binder showed superior acid resistance in the case of the same amount of the enteric polymer applied as compared with the water-soluble binder, and the enzyme could be easily released in the neutral medium. In general, when the coating is performed on the surface of the granules, the amount of coating should be very high due to the properties of the core. However, in the present invention, the coating amount can be reduced by introducing the hardening oil into the granules.

The curing oil used in the present invention may be used in an amount of 30% (w / w) to 100% (w / w) based on the weight of the enzyme active ingredient per unit manufacturing unit, preferably 60% w / w) is appropriate.

Hydroxypropylmethylcellulose phthalate is most preferable as a delayed release polymer in view of being able to simultaneously improve efficiency and economy for application of enzymes as animal feeds and animal drugs.

Although intestinal coagulants that do not change quickly due to pH changes are inexpensive and may be suitable for the present invention, they are relatively low in drug release efficiency. Indeed, sodium alginate and shellac were used to evaluate the acid resistance.

The hydroxypropylmethylcellulose phthalate used in the present invention may be used in an amount of 30% (w / w) to 100% (w / w) based on the weight of the enzyme, which is an active ingredient per unit manufacturing unit, and preferably 50% ) To 85% (w / w) is appropriate.

A specific method for producing the intestinal digestive enzymes of the present invention is as follows.

The enzymes are added to a coalescing machine such as a planetary mixer or a high speed mixer, and the mixture is fed with pre-dissolved hydrogenated oil while stirring. At this time, the curing oil in contact with the enzyme agent is cooled by the enzyme agent and changes to a solid component immediately or in a short time. It is passed through an oscillator equipped with 10mesh ~ 20mesh sieve. In addition to the oscillator, it is also possible to formulate the body with a perforated body such as a power-mill or a co-mill.

The sizing material thus prepared is put into a partitioned fluidized bed coater and is sprayed with a coating solution in which hydroxypropylmethylcellulose phthalate is dissolved in a solvent. At this time, a plasticizer such as triethyl citrate, dibutyl sebacate and an anti-sticking agent such as talc, silicates, and metal stearate may be added to the coating solution.

The solvent used in the coating solution may be any solvent which can completely dissolve the polymer such as ethanol and methylene chloride and is volatile. In addition, since the solvent used is volatilized during the process, it can be recovered from the collecting concentrator and reused, so that it is economically advantageous.

The time spent in the fluidized bed coating process can vary depending on the type of solvent, concentration of the bound solution, process temperature, injection speed, etc., but varies from 30 minutes to 8 hours depending on the process variables. Considering that the product temperature during the process is in the range of 60 to 70 ° C., there is no significant decrease in the activity when heat is applied for 5 hours.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the following examples are for illustrative purposes only and are not intended to limit the scope of the present invention.

Example 1

Add 600 g of the enzyme to a planetary mixer (Whirlpool, USA) and stir at the first stage (about 60 rpm) of the control knob. 400 g of the hardening oil melted in an oven at 70 ° C is added in advance. Allowed to stand at room temperature to solidify the allied material, and passed through an oscillator equipped with a 14 mesh stainless steel body to produce a sizing product. 500 g of hydroxypropylmethylcellulose phthalate is dissolved in 8 L of ethanol and methylene chloride (1: 1), and 30 g of triethyl citrate is dissolved therein to prepare a coating solution. The slurry was put into a fluidized bed coater (DPL 1/3) and flowed at an inlet air temperature of 60 ° C and an inlet air velocity of 25. When the product temperature reached 30 ° C, a feed rate of 25 rpm, an atomizing pressure of 1.5 bar, Lt; RTI ID = 0.0 > 32 C. < / RTI >

Comparative Example 1

500 g of the enzyme is added to a planetary mixer (Whirlpool, USA) and stirred at the first stage (about 60 rpm) of the control knob. 50 g of molasses is added thereto. This was dried in an oven at 60 ° C and passed through an oscillator equipped with a 14 mesh stainless steel body to prepare a sizing product. 500 g of hydroxypropylmethylcellulose phthalate is dissolved in 8 L of ethanol and methylene chloride (1: 1), and 30 g of triethyl citrate is dissolved therein to prepare a coating solution. The slurry was put into a fluidized bed coater (DPL 1/3) and flowed at an inlet air temperature of 60 ° C and an inlet air velocity of 25. When the product temperature reached 30 ° C, a feed rate of 25 rpm, an atomizing pressure of 1.5 bar, Lt; RTI ID = 0.0 > 32 C. < / RTI >

Comparative Example 2

Add 600 g of the enzyme to a planetary mixer (Whirlpool, USA) and stir at the first stage (about 60 rpm) of the control knob. 400 g of the hardening oil melted in an oven at 70 ° C is added in advance. Allowed to stand at room temperature to solidify the allied material, and passed through an oscillator equipped with a 14 mesh stainless steel body to produce a sizing product. 500 g of shellac is dissolved in 2 L of ethanol to prepare a coating solution. The slurry was put into a fluidized bed coater (DPL 1/3) and flowed at an inlet air temperature of 60 ° C and an inlet air velocity of 25. When the product temperature reached 30 ° C, the flow rate was 25 rpm, the atomizing pressure was 2.0 bar, Lt; RTI ID = 0.0 > 32 C. < / RTI >

Experimental Example 1

The eluate (acidic activity) was recovered by reacting the above examples, comparative examples and uncoated enzymes at pH 1.2 for 2 hours using the pharmacodynamic dissolution assay method 2 (paddle method) The remaining residue was reacted again at pH 6.8 for 2 hours, and the protease enzyme activity of the extract (transition activity) was measured to confirm the acid resistance. Separately, the reaction was carried out at pH 6.8 alone for 2 hours under the above elution test conditions to confirm the activity (neutral activity) in the intestinal digestive enzymes. The acid resistance can be determined by the following formula, and the results are shown in Table 1.

Protease Enzyme potency Extract Enzyme Example 1 Comparative Example 1 Comparative Example 2 Neutral activity 16029 17776 17595 5883 Acidic activity 2179 1218 1958 715 Transition active 112 10950 2595 426 Acid resistance (U / g) 2291 12168 4553 1141 Acid resistance% 14.3 68.5 25.9 19.4

 In the case of neutral activity, that is, activity of the enzyme, the activity of the uncoated enzyme was similar to that of Example 1 and Comparative Example 1, so that there was no problem in preparation and formulation compatibility. In Comparative Example 2 using shellac, Respectively. It is considered that this phenomenon is caused by the fact that the release of the enzyme is not smooth due to the low solubility of the shellac itself in the aqueous solution rather than the problem of the preparation and mixing suitability. As for the acidic activity, the enzyme, When the activity was lost under acidic conditions after all of the enzymes were released, activity of 2179 U / g was shown. In the case of Example 1 and Comparative Example 2, this value is lower than that, which effectively suppresses the release of the enzyme in the acidic medium . On the other hand, Comparative Example 1 shows an approximate value of the value of the enzyme, indicating that the enzyme released a considerable amount of the enzyme in the acidic medium. In addition, when the transition activity measuring the activity of the remaining enzyme released from the acidic medium was observed, it can be seen that the uncoated enzyme released all of the enzyme from the acidic medium. In Example 1, about 70% of the enzyme was dissolved in the neutral medium It can be seen that it emits. In the case of the comparative example, since less than 30% of the enzyme is released from the neutral medium due to decomposition or delay of release, it is not suitable for the purpose of the present invention. From the above results, it is possible to confirm the superiority of the present invention by using a process of preparing granules by using hardening oil and coating the surface with hydroxypropylmethylcellulose.

Experimental Example 2

Specification Test

The piglets produced from each sow were randomly assigned to each treatment according to body weight, with a mean age of 28 days and an average weight at placement of 8.47 kg (standard deviation 0.828). The number of housewives was 6, and 12 repetitions were performed. The treatments were divided into a control group without the addition of the enzyme, a 0.1% uncoated enzyme, a 0.05% additive in Example 1, and a 0.1% additive in Example 1. The temperature of the experimental pigs was 32 ° C for the first week, and the temperature was gradually lowered to 28 ° C (at the end of the test) at about 21 ° C and the relative humidity was 50 to 60%. The floor of the pig house was a concrete floor equipped with thermal insulation and heating device, and two pairs of feed diversion per pound of water and a water nipple device were provided. Feed intake and water intake were unlimited. Feed intake was calculated by subtracting the residual amount and the amount of fat from the total amount fed for 28 days. Body weight was measured after the placement and body weight was measured at the end. Dietary digestibility was measured by adding 0.3% Cr ₂ O ₃ and 1.5% Celite-545 to the diets. The diets were fed for a period of one week, Collected in a plastic bag, and stored in a freezer (-55 ° C). The stored feces were pooled and melted after the experiment, and then mixed with a kneader. Then, a certain amount of the feces was collected and rapidly frozen (-55 ° C), followed by lyophilization and pulverization. Celite was analyzed by the method of Prabucki et al. (1975) (HCl-Insoluble Ash). Chromic oxide was applied by Fenton and Fenton (1979). The digestibility of nutrients was derived from the method developed by Schneider and Flatt (1975). The digestibility of each nutrient was calculated from the mean values derived from the two indicators. As a result, the daily gain was significantly (p = 0.04) higher than that of the control (p <0.05). The feed conversion ratios were also significantly improved (p <0.01) by adding the enzymes. This is shown in Table 2.

Control Enzyme
0.1% Example  One
0.05% Example  One
0.10% Standard error P value Daily weight gain, g 347 373 374 383 8.4 0.04 Daily dietary intake, g 520 524 515 523 13.1 0.96 Feed rate 1.50 1.41 1.38 1.37 0.021 <0.01

Claims (9)

An enteric digestive enzymic agent comprising an enzyme as an active ingredient,
Wherein the intestinal digestive enzymes are formed by coating a coating layer of hydroxypropylcellulose phthalate on the surface of granules containing an enzyme agent and a hardening oil. 2. The enteric digestive enzyme according to claim 1, wherein the enzyme is an enzyme produced by solid-state fermentation (SSF). The digestive enzyme according to claim 1 or 2, wherein the hardened oil is prepared by adding hydrogen to a liquid oil of an animal or a plant and has a melting point of 60 to 80 ° C. A method for producing an enteric digestive enzymic agent comprising an enzyme as an active ingredient,
A method for producing an endo-digestive enzyme, comprising the steps of: preparing an association product in an environment in which an enzyme and a hardening oil are not present, preparing a granule by the preparation, and forming a coating layer of hydroxypropylmethylcellulose phthalate on the surface of the granule Gt; [Claim 5] The method according to claim 4, wherein the enzyme is an enzyme produced by solid-state fermentation (SSF). The method according to claim 4, wherein the enzyme is 30 parts by weight or more per unit of production. The method according to claim 4, wherein the hydrogenated oil is prepared by adding hydrogen to the liquid oil of an animal or a plant and has a melting point of 60 to 80 ° C. 5. The method according to claim 4, wherein the hardening oil is 30 to 100 parts by weight based on the weight of the enzyme, which is an active ingredient. The method according to claim 4, wherein the hydroxypropylmethylcellulose phthalate is from 30 to 100 parts by weight based on the weight of the enzyme, which is an active ingredient per unit manufacturing unit.