CN101862447A - A kind of nattokinase composition and preparation method thereof - Google Patents

Abstract

The invention relates to a natto kinase composition and a preparation method thereof. The natto kinase composition comprises natto kinase, film forming materials and film forming agents. The preparation method comprises the following steps: a. mixing the film forming materials and solvents according to a weight ratio of 1/(3 to 19) for preparing film forming liquid; b. adding the natto kinase into the film forming liquid according to a weight ratio of (0.5 to 2)/1, and sufficiently and uniformly mixing the mixture; c. adding the film forming agents accounting for 0.5 to 2 times of the amount into the mixture of the step c, sufficiently and uniformly mixing the mixture, and preparing the mixture into soft materials; and d. taking the soft materials to be made into natto kinase composition grains through the work procedures of granulation, drying at 40 to 60 DEG C and grain pelletization. The invention has the advantages that: 1. the preparation process is simple and can be easily implemented, the automation can be realized, the production efficiency is high, the process production equipment investment is low, and the manufacture cost is low; and 2. the simple process is utilized for improving the stability, and the natto kinase composition can be easily absorbed by human bodies.

Description

A kind of nattokinase composition and preparation method thereof

Technical field:

The invention relates to a nattokinase composition and a preparation method thereof.

Background technique:

In modern social life, thrombus in the human body often causes serious cardiovascular diseases such as vascular embolism, cerebral thrombosis, stroke, acute myocardial infarction, etc., especially causing great harm to the health of middle-aged and elderly people. There are thrombus all over the world. With 15 million patients, the potential thrombolytic market is $2 billion. Nattokinase, as a new type of safe bioactive substance for the prevention and treatment of human thrombotic cardiovascular disease, has been extensively studied since it was discovered by Dr. Sumi Yoko in 1987. According to a large number of scientific studies and clinical trials, nattokinase has a strong effect of dissolving fibrin, can reduce blood viscosity, improve blood circulation, maintain the normal shape and function of blood cells and other physiological functions. Therefore, nattokinase preparations are mainly used for the prevention and treatment of cerebral thrombosis, stroke, and Alzheimer's disease. The development of oral preparations of nattokinase has broad market prospects and far-reaching social significance as the prevention and treatment of thrombotic diseases.

The enzyme activity of nattokinase remains relatively stable in the range of 25-45°C, and the enzyme inactivates rapidly as the temperature rises. When the temperature exceeds 60°C, the enzyme activity will be completely lost within 2 hours. Nattokinase is relatively stable in the natural pH state: the activity is relatively stable in the range of pH 4-12, the activity is the most stable in the range of pH 7-9, and when the pH is lower than 4, it will rapidly denature and inactivate.

The traditional method of preparing enteric-coated tablets, that is, plain tablets with enteric coating, is not suitable for nattokinase. The reason is that this method involves mechanical tablet pressing, and the tablet will be compressed inside the material. Local high temperature is generated, which will cause the loss of nattokinase activity. And enteric-coated capsule is owing to have easy rupture, leakage, various shortcomings such as slow disintegration, directly filling nattokinase with enteric-coated capsule is also limited in use.

Utilize microcapsule preparation technology (as described in patent publication number: CN1766096A), mix nattokinase and pharmaceutical auxiliary material, make soft material, make ball core, spray into coating liquid coating, pack into capsule. The test results of drug release test showed that this method of preparing enteric-coated microcapsules was beneficial to reduce the loss of activity of nattokinase in vivo. But there are certain shortcomings in this preparation method: (1) due to the low temperature (30°C) during coating, it is easy to cause the adhesion of the pellets, which affects the molding of the microcapsule preparation. (2) After coating, the microcapsule still needs a film healing process in a low-temperature (30°C) oven. Due to the low temperature, the film healing time is long, 8 hours, which greatly limits the production.

There is also technology (as described in Patent Publication No.: CN1714675), the mixture of nattokinase and other auxiliary materials except the core auxiliary material is microencapsulated and fixed on the core auxiliary material by airflow spray drying. The microencapsulated nattokinase pharmaceutical composition prepared by air spray drying technology can be directly taken orally. The active ingredient nattokinase is not decomposed and inactivated by pepsin in the stomach of mammals, which improves the stability and effectiveness of the drug. However, the disadvantage of using airflow spray drying technology is that the dryer has a large volume and low heat transfer coefficient, resulting in low thermal efficiency and high power consumption. At present, the thermal efficiency of domestic industrial-scale spray dryers is generally 30% to 50%. The thermal efficiency of foreign spray dryers with waste heat recovery can reach 70%. It makes economic sense. Therefore, the preparation of the nattokinase microcapsule pharmaceutical composition using airflow spray drying technology not only puts forward higher requirements on the production process, but also is a very large limitation on the economic cost.

Invention content:

In order to solve the technical problem, the invention provides a nattokinase composition and a preparation method thereof.

The technical scheme that the present invention solves its technical problem is: a kind of nattokinase composition, comprises nattokinase, film-forming material, film-forming agent, and described film-forming material is made of cellulose esters, acrylic resins, One or more of polyvinyl alcohol phthalate, methacrylic acid copolymer, shellac, and corn gluten are mixed with a soluble solvent.

Its preparation method step is as follows: a, mix film-forming material and solvent by the weight ratio of 1: (3～19), prepare film-forming liquid; B, press (0.5～2): 1 weight ratio adds nattokinase In the film-forming solution, mix well; c. Add 0.5 to 2 times the amount of film-forming agent to the mixture in step b, mix well, and prepare a soft material; d. The soft material is granulated and heated at 40-60°C The nattokinase composition granules are prepared through drying and granulation processes.

Further explanation: the film-forming agent is formed by mixing one or two of water and organic alcohols.

In order that the nattokinase composition is more beneficial to health, further: the nattokinase composition is also added with a diluent.

Further: the diluent is formed by mixing one or more of starches, sugars, celluloses, inorganic salts, and sugar alcohols.

In order to take the different habits of customers, further: the nattokinase composition granules prepared in the step d are converted into dosage forms.

Compared with the prior art, the present invention has the following salient features: 1. The preparation process is simple and easy, can realize automation, high production efficiency, less investment in process production equipment, and lower manufacturing cost compared with the prior art. 2. The acid-resistant stability of nattokinase in the prepared composition is effectively improved by using a simple process, and it is easy to be absorbed by the human body.

Detailed ways:

A kind of nattokinase composition, comprises nattokinase, film-forming material, film-forming agent, and described film-forming material is made of cellulose esters, acrylic resins, polyvinyl alcohol phthalate (PVAP), methacrylic acid One or more of copolymer, shellac, and zein are mixed with a soluble solvent, and the film-forming agent is mixed with one or two of water and organic alcohols, and for the combination of nattokinase The food is more beneficial to health, and the nattokinase composition is also added with a diluent, and the components of the diluent are mixed by one or more of starches, sugars, celluloses, inorganic salts, and sugar alcohols made.

Its specific formula and dosage are as follows:

Formula Amount (%)

             

Nattokinase 10～100000 units

Film-forming solution 8～20

Film former 8～20

Thinner 0～60

The preparation method steps of described nattokinase composition are as follows: a, mix film-forming material and solvent by the weight ratio of 1: (3～19), prepare film-forming liquid; b, press (0.5～2): 1 weight ratio Ratio: Add nattokinase into the film-forming solution and mix well; c. Add 0.5 to 2 times the amount of film-forming agent to the mixture in step b, mix well and prepare a soft material; d. The nattokinase composition granules are prepared through the processes of granulation, drying at 40-60° C. and granulation.

In order to take according to the different habits of customers, further: transform the nattokinase composition granules prepared in the step d into dosage forms, such as capsules, tablets, pills, granules, powders, etc.

Customers can effectively prevent and treat cerebral thrombosis, cerebral apoplexy, senile dementia, etc. by taking the nattokinase composition of the present invention. The detection data and results of the nattokinase composition of the present invention are given below:

1. The formula of nattokinase composition is as follows: (the activity of nattokinase in the composition is 1,000FU)

ethanol 0 35 15 25 5

2. Detection index of nattokinase composition:

Acid resistance investigation of nattokinase activity in nattokinase composition:

1. Preparation of artificial gastric juice and phosphate buffer

(1) Artificial gastric juice: add 0.9ml of concentrated hydrochloric acid to a 100ml volumetric flask, add distilled water to make the volume to 100ml, and obtain artificial gastric juice of pH 1.2.

(2) PH7.8 phosphate buffer:

Solution A: Disodium hydrogen phosphate 71.8g, dissolved in 1000ml water

Solution B: 2.76g sodium dihydrogen phosphate, dissolved in 100ml water

Take 997.35ml of A solution and 92.65ml of B solution and mix them.

2. Acid damage

Take by weighing 10 parts of nattokinase composition samples (above-mentioned sample numbers 1, 2, 3, 4, 5, 2 parts for each numbered sample), add to a 25ml volumetric flask, add 10ml of artificial gastric juice and shake evenly, place in a water bath Place at 37°C for 2 hours, shaking every 10 minutes.

3. Neutralization

Add 10ml of 0.2MPH7.8 phosphate buffer, shake to disperse the composition evenly, centrifuge at 3000 rpm for 3 minutes, and take the supernatant for testing.

4. Determination of activity retention rate by fibrin plate method

(1) Preparation of fibrin plate:

Weigh fibrinogen, and use 30ml 0.1MPH7.8PBS solution to make a concentration of 0.2%-0.5%; weigh thrombin, dissolve it with 1ml0.1MPH7.8PHS solution, and make a concentration of 0.2%-0.5%. Weigh the agar powder, add 80ml of 0.1MPH7.8PHS solution to make the concentration 1.0%-2.0%, heat to boil, cool to about 50°C, add fibrin stock solution and bovine thrombin solution to the agar solution, shake gently Pour it evenly into a plate, remove the air bubbles with a medicine spoon, cool at room temperature, and punch holes with a puncher;

(2) Use a microsampler to pipette a little enzyme solution of each sample and add it to the well of the fibrin plate;

(3) Incubate the fibrin plate with the sample enzyme solution in a 37°C oven for 4 hours;

(4) Measure the diameter of the dissolution circle with a vernier caliper, and calculate the size of the enzyme activity.

Table 1 Nattokinase composition sample acid resistance investigation data table

Sample serial number Sampling activity (FU) Loading concentration (FU/ml) Loading amount (ul) Undamaged sample dissolution ring diameter (cm) Acid damage sample dissolution ring diameter (cm) Undamaged sample dissolution area (cm ² ) Acid-damaged sample dissolution zone area (cm ² ) Sample activity retention rate (%) 01 1600 40 40 1.833 1.085 2.6375 0.926 35.11 02 1600 40 40 1.797 0.989 2.536 0.7685 30.39 03 1600 40 40 1.808 1.017 2.566 0.812 31.64 04 1600 40 40 1.785 1.018 2.501 0.822 32.92 05 1600 40 40 1.793 0.937 2.525 0.691 27.31 Listed product 1 1600 40 40 1.836 0.227 2.646 0.040 1.53 Listed product 2 1600 40 40 1.789 0.767 2.512 0.462 18.37 Listed products 3 1600 40 40 1.805 0.552 2.558 0.239 9.36

in conclusion:

The results of the acid resistance investigation of 5 batches of samples show that the activity of the nattokinase composition prepared by the method is improved under acidic conditions. The 5 batches of nattokinase compositions prepared by this method have obvious advantages in acid resistance (sample activity retention rate) compared with 3 products on the market.

Nattokinase composition (sample number 01) release check:

Method: small cup method, speed: 100rpm, temperature: 37°C, release medium: 0.1mol/LHCl in 100ml for 2 hours, sampled at 20min, 40min, 60min, 80min, 100min, 120min, and added to pH7.8 borax buffer And as a sample solution, measure the enzyme activity; after 120 minutes of sampling, add 100ml of pH7.8 borax buffer solution to the cup, continue to release for 45 minutes, take a sample, and measure the enzyme activity. The release medium was degassed before use.

Enzyme activity assay method:

(1) Take a test tube and add 1.4ml of borax buffer and 0.4ml of fibrinogen, and place it in a water bath at 37±0.2°C for 5 minutes.

(2) Add 0.1ml bovine thrombin solution to the above test tube, shake and mix well, and let stand in a water bath at 37±0.2°C for 10 minutes.

(3) Reaction group——Step 2 is completed. At the precise 10th minute, start to add 0.1ml of the sample solution that has just been diluted, and fully oscillate and mix for 5 seconds. Place it in a water bath at 37±0.2°C for 1 hour, and After 20 minutes and 40 minutes, shake and mix again.

Control group——step 2 is completed. At the precise 10th minute, start to add 2ml of methyl trichloroacetate, shake and mix well, then add 0.1ml of the sample solution that has just been diluted, and shake and mix well, in 37±0.2℃ water Let stand in the bath for 20 minutes.

(4) The part of the reaction group is completed in step 3. At the precise first hour, start to add 2ml of methyl trichloroacetate, shake and mix well, and let it stand in a water bath at 37±0.2°C for 20 minutes.

(5) Centrifuge the reactant in the test tube at 12,000 rpm for 10 minutes.

(6) Take the supernatants of the reaction group and the control group to measure the absorbance at UV _{275 nm} , and record the absorbance Ar of the reaction group and the absorbance Ac of the control group.

(7) Calculate the enzyme activity according to the absorbance.

The experimental results are as follows:

Table 2: Nattokinase sample solution enzyme activity assay data table within 2 hours of release experiment

From the test results, it can be seen that the intragastric release of nattokinase in the nattokinase composition prepared by this method is less than 90%, ensuring that more than 10% of the nattokinase in the composition enters the intestine for release, and after the nattokinase is absorbed through the intestine exert physiological effects.

Claims (6)

1. a nattokinase composition, is characterized in that: described nattokinase composition comprises nattokinase, film-forming material, film-forming agent, and described film-forming material is made of cellulose esters, acrylic resins, One or more of polyvinyl alcohol phthalate, methacrylic acid copolymer, shellac, and corn gluten are mixed with a soluble solvent. 2. The nattokinase composition according to claim 1, characterized in that: the film-forming agent is formed by mixing one or both of water and organic alcohols. 3. The nattokinase composition according to claim 1, characterized in that: the nattokinase composition is also added with a diluent. 4. the nattokinase composition according to claim 3 is characterized in that: said diluent is mixed by one or more in starches, sugars, celluloses, inorganic salts, sugar alcohols become. 5. a kind of preparation method of nattokinase composition according to claim 1 is characterized in that: described preparation method step is as follows: a, mix film-forming material and solvent by the weight ratio of 1: (3～19) , prepare the film-forming solution; b, add nattokinase into the film-forming solution in a weight ratio of (0.5-2): 1, and mix well; c, add 0.5-2 times the amount of forming The film preparation is fully mixed and prepared into a soft material; d, the soft material is granulated, dried at 40-60° C. and granulated to prepare the nattokinase composition granules. 6. the preparation method of preparing nattokinase composition according to claim 5 is characterized in that: the nattokinase composition granule that described step d is made carries out dosage form conversion.