WO2020113812A1 - Nadh-containing biological high-molecular-weight nanosphere, preparation method for same, and applications thereof - Google Patents

Abstract

An NADH-containing biological high-molecular-weight nanosphere, comprising a biological high-molecular-weight carrier and NADH dispersed on the biological high-molecular-weight carrier. The biological high-molecular-weight carrier is a high-molecular-weight fiber polymer, a fiber frame thereof constitutes a three-dimensional interpenetrating network structure, thus offering protection for the NADH, solving the problem of same being prone to decomposition upon exposure to light or oxygen, extending the preservation time of the active component of NADH, and reducing the difficulty of preservation. Disclosed is a preparation process for the biological high-molecular-weight nanosphere. A biological high-molecular-weight material of a suitable chain length acquired via physical modification allows spherical particles to be formed easily, constitutes a honeycomb-shaped three-dimensional interpenetrating network structure, and favors NADH micromolecules to be loaded on the inner and outer surfaces of the network structure. The biological high-molecular-weight nanosphere is applicable in preparing preparations such as tablets, granules, capsules or soft capsules to serve as a human body healthcare or pet therapeutic medicament/functional food product.

Description

Biopolymer nanospheres containing NADH, preparation method and application thereof Technical field

The invention relates to the technical field of health care products, in particular to a biopolymer nanosphere containing NADH and a preparation method and application thereof.

Background technique

The reduced state of nicotinamide adenine dinucleotide (NADH) is a proton-transporting coenzyme, also known as reduced coenzyme I. NADH serves as an electron donor and has oxidation during hundreds of thousands of metabolic reactions in living cells. The reduction activity plays an important role in electron transfer during glycolysis, citric acid cycle and photosynthesis. In vivo, NADH is involved in vitamin derivation and the production of adenosine triphosphate (ATP), and is a coenzyme of more than 250 dehydrogenases that have been identified, and has the role of maintaining cell growth, differentiation and energy metabolism.

NADH is considered to have beneficial effects on the body and intelligence without side effects. It can be used as a drug to improve health and quality of life. Clinical studies have shown that parenteral administration of NADH has a positive effect on the treatment of Parkinson's disease and major depression. NADH can promote the cognitive ability and exercise ability of patients with Parkinson's disease and Alzheimer's disease, and at the same time relieve fatigue, drowsiness, pigmented fatigue syndrome, and enhance the vitality of users.

In the middle of the last century, NADH has successfully treated various neurological diseases such as Parkinson's disease, Alzheimer's disease and delayed dementia by infusion, but due to the high sensitivity of NADH liquid (very sensitive to light and oxygen), As a result, its chemical properties are unstable (requiring ready-to-use), which has not been widely used. In addition, NADH is decomposed immediately after contact with gastric acid, which makes oral administration impossible.

Summary of the invention

In view of this, the present invention provides a NADH-containing biopolymer nanosphere with good stability, easy storage, oral administration, and mass production.

In addition, it is also necessary to provide a method for preparing the above-mentioned NADH-containing biopolymer nanospheres and the application of the above-mentioned NADH-containing biopolymer nanospheres.

The invention provides a biopolymer nanosphere containing NADH, which includes a biopolymer carrier and NADH dispersed on the biopolymer carrier.

The invention also provides a method for preparing biopolymer nanospheres containing NADH, including the following steps:

Mixing and grinding the biopolymer raw material and the auxiliary materials to physically modify the biopolymer raw material to obtain a biopolymer carrier;

Grind NADH raw material and sieve to obtain NADH;

The NADH and the biopolymer carrier are mixed and stirred to obtain the NADH-containing biopolymer nanosphere, wherein NADH is dispersed on the biopolymer carrier.

Preferably, the auxiliary material includes at least one of 0.05-30 parts of sodium alginate and 0.05-30 parts of xanthan gum.

The invention also provides an application of the NADH-containing biopolymer nanospheres in the preparation of drugs for preventing and treating sub-health and tumors and functional foods.

The invention also provides a pharmaceutical preparation comprising the NADH-containing biopolymer nanospheres and pharmaceutically acceptable auxiliary materials;

Preferably, the pharmaceutical preparation is one of tablets, capsules, granules, injections, tinctures, suppositories, patches, pills, syrups, mixtures, powders, lotions, films and dripping pills;

Preferably, the pharmaceutical preparation is a capsule, and the capsule includes hard capsules and soft capsules.

The invention also provides a method for preparing the pharmaceutical preparation, which includes the following steps:

Providing the NADH-containing biopolymer nanospheres, adding granulation auxiliary materials to the biopolymer nanospheres and stirring;

Screening to obtain granular wet material with uniform particle size;

Drying the wet material of particles to obtain particles of biopolymer nanospheres containing NADH;

Preferably, the method further includes tabletting the NADH-containing biopolymer nanosphere particles into tablets;

Preferably, the method further includes filling the NADH-containing biopolymer nanosphere particles into a capsule shell to make a capsule;

Preferably, the method further comprises mixing the NADH-containing biopolymer nanosphere particles with edible oil and filling the soft capsule shell to make a soft capsule;

Preferably, the granulation auxiliary material is at least one of microcrystalline cellulose, polyvinylpyrrolidone, sodium bicarbonate, magnesium stearate, calcium polyphosphate, wetting agent and binder.

The invention also provides a functional food comprising the NADH-containing biopolymer nanospheres and food additives.

The advantages of the present invention: The NADH-containing biopolymer nanospheres provided by the present invention extend the storage time of NADH active ingredients, reduce storage difficulty, improve stability, and maintain biological activity.

BRIEF DESCRIPTION

FIG. 1 is a preparation flow chart of NADH-containing biopolymer nanospheres in a preferred embodiment of the present invention.

Fig. 2 is a preparation flow chart of a pharmaceutical preparation in a preferred embodiment of the present invention.

FIG. 3 is an XRD test chart of NADH-containing biopolymer nanospheres under different NADH contents in a preferred embodiment of the present invention.

4A and 4B are scanning electron microscope images of NADH-containing biopolymer nanospheres in Example 1 of the present invention.

5 is a graph showing the release rate of the active ingredient after soaking in the simulated gastric acid solution and the simulated colon solution after the NADH-containing biopolymer nanospheres are formed into tablets in Example 2 of the present invention.

6 is a graph showing the release rate of the active ingredient after the NADH-containing biopolymer nanospheres in Example 2 of the present invention are formed into tablets and soaked in a simulated colon solution.

detailed description

The technical solutions in the embodiments of the present invention will be described clearly and completely in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, but not all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by a person of ordinary skill in the art without making creative efforts fall within the protection scope of the present invention.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the technical field of the present invention. The terminology used in the description of the present invention herein is for the purpose of describing specific embodiments, and is not intended to limit the present invention.

A preferred embodiment of the present invention provides a NADH-containing biopolymer nanosphere, which includes a biopolymer carrier and NADH dispersed on the biopolymer carrier.

The biopolymer carrier has a three-dimensional network structure, and the average particle size of the biopolymer carrier is 200-1000 nm. The biopolymer carrier includes at least one of chitosan (CS) and konjac glucomannan (KGM). KGM is a natural polymer soluble dietary fiber. The fiber has a honeycomb three-dimensional interpenetrating network structure. The size of the mesh is tens to thousands of nanometers, NADH enters through the mesh and is loaded on the surface of the KGM fiber network. Preferably, the konjac glucomannan is at least one of konjac glucomannan, quaternized konjac glucomannan, carboxymethyl konjak glucomannan and deacetylated konjac glucomannan. In terms of parts by weight, the biopolymer nanospheres include 1-20 parts of the NADH and 20-80 parts of the biopolymer carrier.

Preferably, the biopolymer nanosphere further includes an auxiliary material, and the auxiliary material includes at least one of 0.05-30 parts of sodium alginate and 0.05-30 parts of xanthan gum in parts by weight. Sodium alginate is a natural polysaccharide. It has the ability to concentrate solutions, form gels and form films. It can form a film structure on the outer surface of the biopolymer carrier. It has a waterproof and anti-oxidation effect and avoids NADH from being decomposed by light or oxygen. problem. The average particle diameter of the biopolymer nanosphere is 500-1000 nm.

Please refer to FIG. 1, a preferred embodiment of the present invention further provides a method for preparing the NADH-containing biopolymer nanospheres, including the following steps:

S11. Mix and grind the biopolymer raw material and the auxiliary materials to physically modify the biopolymer raw material to obtain a biopolymer carrier;

After mixing and grinding the biopolymer raw material and the auxiliary material, the molecular chain length and molecular weight of the biopolymer raw material are reduced, forming a honeycomb three-dimensional interpenetrating network structure and forming a cluster with an average particle size of 200-1000 nm Nanosphere.

S12. Grind NADH raw material and sieve to obtain NADH;

The average particle diameter of the obtained NADH is 10 to 100 nm.

S13. Mixing and agitating the NADH with the biopolymer carrier to obtain the NADH-containing biopolymer nanosphere, wherein NADH is dispersed on the biopolymer carrier.

It can be understood that, in the present invention, the order of addition between the NADH and the biopolymer carrier is in no particular order, and can be added in any order of addition.

Adjust the addition amount of NADH in step S13, and detect the NADH-containing biopolymer nanospheres with different NADH contents (0%, 8%, and 10%) by X-ray diffraction (XRD) to obtain XRD as shown in FIG. 3 Diffraction pattern, as can be seen from Fig. 3, the biopolymer carrier (blank microspheres) without NADH loading exhibited broad diffraction peaks of polysaccharide cellulose (diffraction peaks at 2θ angles of 18° and 22.5°), After loading NADH, several new diffraction peaks appeared, which are characteristic diffraction peaks of NADH, and the intensity of XRD diffraction peak of nanospheres containing 10% NADH is higher than that of nanospheres containing 8% NADH, which further proves that NADH has succeeded Loaded on biopolymer carrier.

Further, the NADH raw material in step S12 is prepared by the following process:

S121. Extract the crude extract of nicotinamide riboside adenylyl transferase or its pure enzyme;

S122. A crude extract of immobilized recombinant nicotinamide riboside adenylyl transferase or its pure enzyme;

S123. The NADH raw material is prepared by catalyzing with immobilized recombinant nicotinamide riboside adenylyl transferase, using nicotinamide nucleotide and adenosine triphosphate (ATP) as substrates.

The specific steps of extracting the crude extract of nicotinamide riboside adenylyl transferase or its pure enzyme in S121 are as follows:

The plasmid pRSET containing the nicotinamide riboside adenylyl transferase gene was transformed into competent bacterial cells E. coli HB101 and cultured on a Luriabroth (LB) plate (containing 100 mg/L kanamycin) at 37°C for 24 hours. Inoculate a single clone in 5 ml of LB liquid medium (containing 100 mg/L kanamycin) and incubate at 30°C for 20-24 hours. The cells were collected by centrifugation and suspended in 1 mL of 100 mM Tris hydrochloric acid buffer (pH=7.5). Then, the bacterial cells were lysed with ultrasonic waves, centrifuged (17°C, 10800 g, 10 min) and the supernatant was collected as crude protein extract (or crude extract). The crude nicotinamide riboside adenylyl transferase crude protein was heat-treated at 70°C for 10 min, centrifuged (10°C, 17800 g, 10 min), and the supernatant was collected as the partially purified protein.

The specific steps of immobilizing nicotinamide riboside adenylyl transferase in S122 are as follows: take the crudely extracted protein of nicotinamide riboside adenylyl transferase or partially purified protein, and wash with enzyme buffer (0.02M TrisHCl/0.001M EDTA, pH=7.0 solution) diluted to protein content 5～10mg/mL. Mix the enzyme diluent and PB solution (2.0mol/L potassium dihydrogen phosphate, pH=7.5) in equal volumes, add the epoxy-type immobilized enzyme carrier LX3000 (10 mg enzyme/g carrier) on a shaker (speed 100rpm ) Reaction at 25 ℃ for 20h. After the reaction is completed, filter with a filter bag and wash with enzyme buffer 5 to 6 times to obtain immobilized nicotinamide riboside adenylyl transferase.

The specific steps for preparing the NADH raw material with immobilized nicotinamide riboside adenylyl transferase in S123 are as follows: preparing a substrate solution: containing 5 mM nicotinamide nucleotide, 10 mM disodium adenosine triphosphate (ATP), 100 mM Tris hydrochloric acid The buffer and MgCl _{2 at a} final concentration of 10 mM were adjusted to pH 7.5. Take 1 mL of the substrate solution, then add 0.05 g of immobilized nicotinamide riboside adenylyl transferase, and perform the reaction at 37°C for 2 to 20 hours. Centrifuge (17,800 g, 15 min at 10°C) and collect the supernatant. The content of nicotinamide adenine dinucleotide in the resulting supernatant was determined by high pressure liquid chromatography (HPLC). The results show that the conversion rate of nicotinamide nucleotides to nicotinamide adenine dinucleotide exceeds 80%.

The invention also provides an application of the NADH-containing biopolymer nanospheres in the preparation of drugs for preventing and treating sub-health and tumors and functional foods.

The invention also provides a pharmaceutical preparation comprising the NADH-containing biopolymer nanospheres and pharmaceutically acceptable excipients. Preferably, the pharmaceutical preparation is one of tablets, capsules, granules, injections, tinctures, suppositories, patches, pills, syrups, mixtures, powders, lotions, films, and pills. Preferably, the pharmaceutical preparation is a capsule, and the capsule includes hard capsules and soft capsules. The pharmaceutically acceptable auxiliary materials include granulation auxiliary materials. Preferably, the granulation auxiliary material is at least one of microcrystalline cellulose, polyvinylpyrrolidone, sodium bicarbonate, magnesium stearate, calcium polyphosphate, wetting agent and binder.

Preferably, a suitable daily dosage of NADH oral dosage form is 5 to 500 mg. Preferably, a suitable daily dosage of NADH oral dosage form is 25-100 mg.

Referring to FIG. 2, a preferred embodiment of the present invention also provides a method for preparing the pharmaceutical preparation, which includes the following steps:

S21. Provide the NADH-containing biopolymer nanospheres, add granulation auxiliary materials to the biopolymer nanospheres and stir;

S22, sieve to prepare granule wet material with uniform particle size;

S23. Dry the wet material of the particles to obtain particles of biopolymer nanospheres containing NADH.

Preferably, the method further includes tabletting the NADH-containing biopolymer nanosphere particles into tablets;

Preferably, the method further includes filling the NADH-containing biopolymer nanosphere particles into a capsule shell to make a capsule;

Preferably, the method further comprises mixing the NADH-containing biopolymer nanosphere particles with edible oil and filling the soft capsule shell to make a soft capsule;

Preferably, the granulation auxiliary material is at least one of microcrystalline cellulose, polyvinylpyrrolidone, sodium bicarbonate, magnesium stearate, calcium polyphosphate, wetting agent and binder.

The wetting agent includes one or two of 70% volumetric ethanol and water, and the binder includes pregelatinized starch with a mass concentration of 5-20%, and a starch slurry with a mass concentration of 10-15% And at least one of hypromellose solutions with a mass concentration of 10%. The drying condition is vacuum drying at 10-50°C for 0.5-24 hours.

It can be understood that, in the present invention, the order of addition between the NADH-containing biopolymer nanospheres and the granulation auxiliary materials is in no particular order, and they can be added in any order of addition. In the present invention, the preparation of the NADH-containing biopolymer nanospheres can be performed simultaneously with the preparation of the pharmaceutical preparation, that is, when the NADH is mixed with the biopolymer carrier, the granulation can be added The accessories are stirred together.

The invention also provides a functional food comprising the NADH-containing biopolymer nanospheres and food additives. The food additives include pectin, fumaric acid, polydextrose, maltose, phospholipid, citric acid, hydroxypropyl starch, lactic acid, sorbitol, milk powder, maltodextrin, honey, corn starch, corn oil, sesame oil , Sucrose, vitamin C, vitamin E, xylitol and gelatin one or more.

The present invention will be specifically described below through examples.

Example 1

Preparation of biopolymer nanospheres containing NADH:

S11, kneading konjac glucomannan (KGM) and sodium alginate with a colloid mill to obtain a biopolymer carrier.

S12. Grinding NADH raw material and sieving to obtain NADH, the average particle diameter of the NADH is 10-100 nm.

S13. The NADH obtained in step S12 is mixed with the biopolymer carrier and stirred evenly to obtain a biopolymer nanosphere containing NADH. The biopolymer nanospheres include 20 parts of biopolymer carrier and 1 part of NADH in parts by weight. The material of the biopolymer carrier is konjac glucomannan (KGM). The NADH-containing biopolymer nanospheres further include an auxiliary material, and the auxiliary material includes 0.05 parts of sodium alginate in parts by weight.

The above-mentioned biopolymer nanospheres are used to prepare a pharmaceutical preparation, which is a granule.

Preparation of granules for pharmaceutical preparations:

S21. Add granulation auxiliary materials to the biopolymer nanospheres containing NADH and stir. In terms of parts by weight, the granulation auxiliary materials include 10 parts of wetting agent and 5 parts of binder. The wetting agent is Ethanol with a volume concentration of 70%, and the binder is a starch slurry with a mass concentration of 10%;

S22. Wet granulation with a medium-sized steel sieve. Weigh the weighed material on a set of sieve screens with a sieve pore size decreasing from the inside to the outside. The material is left on each sieve surface according to the particle size to obtain granulated wet material. ;

S23. Dry the wet material at a drying temperature of 40°C for a period of 3 hours to obtain particles of biopolymer nanospheres containing NADH.

Example 2

The difference from Example 1 is that the biopolymer nanospheres include 80 parts of biopolymer carrier and 20 parts of NADH, and the material of the biopolymer carrier is quaternized konjac glucomannan (QKGM) and The mixture of carboxymethyl konjac glucomannan (CKGM) has a mass ratio of 1:1. The auxiliary material includes 30 parts of sodium alginate.

The quaternized konjac glucomannan (QKGM) and carboxymethyl konjac glucomannan (CKGM) are prepared from konjac glucomannan (KGM) through quaternization and carboxymethylation treatment, respectively. The quaternized konjac glucomannan (QKGM) and carboxymethyl konjac glucomannan (CKGM) can also be stirred with a concentration of 0.1 to 1 mol/L HCl or 0.1 to 1 mol/L NaOH for 2 to 10 hours The molecular weight of konjac glucomannan facilitates subsequent physical modification.

The pharmaceutical preparation is a tablet, and the tablet is prepared by the following method:

The granulation method described in Example 1 is used for granulation, and then the prepared granules are compressed with a tablet machine. By adjusting the parameters and pressure of the compression mold, the tablets with the target diameter, thickness and hardness can be prepared. Agent. The granulation method of this example is different from that in Example 1 in that granulation auxiliary materials include 40 parts of binder, 30 parts of magnesium stearate and 30 parts of microcrystalline cellulose, wherein The binder is a mixture of pregelatinized starch with a mass concentration of 5% and hypromellose solution with a mass concentration of 10%.

Example 3

Different from Example 1: the biopolymer nanospheres include 45 parts of biopolymer carrier and 5 parts of NADH, and the material of the biopolymer carrier is deacetylated konjac glucomannan (da-KGM) . The auxiliary material includes 15 parts of sodium alginate.

The deacetylated konjac glucomannan (da-KGM) is prepared by deacetylation of KGM. Before step S11, the method further includes mixing the deacetylated konjac glucomannan (da-KGM) with a concentration of 0.1 to 1 mol/L HCl or 0.1 to 1 mol/L NaOH for 2 to 10 hours to reduce the konjac glucomannan The molecular weight of sugar is convenient for subsequent physical modification.

The pharmaceutical preparation is a capsule, and the capsule is prepared by the following method:

The granulation method described in Example 1 is used for granulation, and after granulation, the prepared granules are filled into the shell of a commercially available edible hard capsule by a quantitative filling method to obtain a capsule preparation. The granulation method of this example is different from that of Example 1 in that granulation auxiliary materials include 20 parts of binder, 0.5 parts of magnesium stearate, 0.5 parts of microcrystalline cellulose, 1 Parts of polyvinylpyrrolidone, 2 parts of sodium bicarbonate and 20 parts of wetting agent. Wherein, the wetting agent is ethanol with a volume concentration of 70%, and the binder is a starch slurry with a mass concentration of 10%.

Example 4

The difference from Example 1 is that the biopolymer nanospheres include 60 parts of biopolymer carrier and 5.2 parts of NADH. The auxiliary material includes 30 parts of xanthan gum.

Before step S11, the method further includes stirring KGM with 0.1-1 mol/L HCl or 0.1-1 mol/L NaOH together for 2-10 hours to reduce the molecular weight of the konjac glucomannan to facilitate subsequent physical modification.

The pharmaceutical preparation is a soft capsule, and the soft capsule is prepared by the following method:

The granulation method described in Example 1 is used for granulation. After granulation, the prepared granules are mixed with an appropriate amount of edible oil and biosurfactant, and then encapsulated in a commercially available soft capsule shell to obtain a soft capsule preparation The edible oil may be olive oil, sesame oil, soybean oil, corn oil, camellia oil, grape seed oil, etc. The granulation method of this embodiment is different from that of Embodiment 1 in that granulation auxiliary materials are different. In this embodiment, the granulation auxiliary materials include 16 parts of binder, 2 parts of magnesium stearate, and 1 part of poly Calcium phosphate and 15 parts of wetting agent, wherein the wetting agent is ethanol with a volume concentration of 70%, and the binder is hypromellose solution with a mass concentration of 10%.

Example 5

Different from Example 1, the biopolymer nanospheres include 40 parts of biopolymer carrier and 12 parts of NADH. The auxiliary material includes 0.05 parts of xanthan gum and 13 parts of sodium alginate.

Before step S11, the method further includes stirring KGM with 0.1-1 mol/L HCl or 0.1-1 mol/L NaOH together for 2-10 hours to reduce the molecular weight of the konjac glucomannan to facilitate subsequent physical modification.

In this embodiment, the pharmaceutical preparation is a granule. The granule is prepared by the method described in Example 1. The difference is that the granulation auxiliary material includes 5 parts of binder and 12 parts of microcrystalline cellulose. And 15 parts of a wetting agent, wherein the wetting agent is a mixture of ethanol and water with a volume concentration of 70%, and the binder is pregelatinized starch with a mass concentration of 15%.

Example 6

The difference from Example 1 is that the biopolymer nanosphere includes 70 parts of biopolymer carrier and 15 parts of NADH, and the material of the biopolymer carrier is chitosan (CS). The auxiliary material includes 15 parts of xanthan gum and 24 parts of sodium alginate.

The biopolymer nanospheres containing NADH described in this example can be prepared by the preparation method of Example 1. The difference is that, before step S11, the chitosan is further subjected to quaternization treatment for chemical modification to reduce the shell The molecular weight of glycan facilitates subsequent physical modification.

In this embodiment, the pharmaceutical preparation is a tablet, and the tablet is prepared by the following method:

The granulation method described in Example 1 is used for granulation, and then the prepared granules are compressed with a tablet machine. By adjusting the parameters and pressure of the compression mold, the tablets with the target diameter, thickness and hardness can be prepared. Agent. The granulation auxiliary material includes 16 parts of binder, 8 parts of microcrystalline cellulose, 10 parts of polyvinylpyrrolidone, 20 parts of sodium bicarbonate, and 13 parts of wetting agent, wherein the wetting agent is A mixture of ethanol and water with a volume concentration of 70%, and the binder is a starch slurry with a mass concentration of 10%.

Example 7

The difference from Example 1 is that the biopolymer nanospheres include 55 parts of biopolymer carrier and 8 parts of NADH, and the material of the biopolymer carrier is chitosan (CS). The auxiliary material includes 20 parts of xanthan gum and 5 parts of sodium alginate. In this embodiment, the material of the biopolymer carrier is chitosan (CS), and CS is a natural polymer material. The material has a honeycomb three-dimensional interpenetrating network structure with a mesh size of tens to thousands Nanometer, NADH enters through the mesh and is loaded on the surface of the chitosan network.

In this embodiment, the pharmaceutical preparation is a granule. The granule is prepared by the method described in Example 1. The difference is that: granulation auxiliary materials are different. The granulation auxiliary materials include 8 parts of binder and 6 parts. Of microcrystalline cellulose, 4 parts of polyvinylpyrrolidone, 5 parts of sodium bicarbonate, 10 parts of magnesium stearate, 10 parts of calcium polyphosphate and 15 parts of wetting agent, wherein the wetting agent is A mixture of ethanol and water at a volume concentration of 70%, and the binder is a mixture of starch slurry at a mass concentration of 10% and pregelatinized starch at a mass concentration of 15%.

Table 1 NADH-containing biopolymer nanospheres prepared in Examples 1-7 of the present invention and their specific processing conditions

Scanning electron microscopy tests were performed on the biopolymer nanospheres containing NADH prepared in Example 1, and the test results are shown in FIGS. 4A and 4B. As can be seen from FIGS. 4A and 4B, the biopolymer nanospheres have a honeycomb three-dimensional interpenetrating network structure.

The pregelatinized starch in the granulation adjuvant in Example 2 not only has good disintegration and adhesion, but also significantly improves the hardness, disintegration and surface brightness of the tablet, and more importantly, it improves the dissolution rate. The granulation difficulty is reduced, and the granulation and compressibility of the granules are improved. The compressed tablets have high hardness, low brittleness and smooth surface. Hydroxypropyl methylcellulose is a derivative of a mixed ether of hydroxypropyl and methoxycellulose. The replacement group in the molecule is an ether. It is used in tablets, mainly as a binder and a disintegrant. Disintegration improved and dissolution increased. Microcrystalline cellulose has good fluidity and compressibility, and has both adhesive, lubricating, and disintegration-assisting properties. It has no interaction with drugs, and can make the tablet shape smooth, beautiful, and easy to disintegrate.

The tablets prepared in Example 2 were immersed in simulated gastric acid solution SGF pH=2 for 2 hours, and then simulated colonic solution SCF pH=6.8 for 6 hours, and tested for NADH at 2h, 4h, 6h and 8h respectively The release rate and test results are shown in Table 2 and Figure 5.

Table 2 The release rate of the active ingredient after soaking in the simulated gastric acid solution and the simulated colon solution after the NADH-containing biopolymer nanospheres in Example 2 are made into tablets

Time h	Release rate%
0	0
2	1.12
4	20.72
6	50.24
8	90.33
remaining	0.30
Total	90.63

The tablets prepared in Example 2 were immersed in the simulated colon solution SCF pH=6.8 for 8 hours, and the release rates of NADH at 2h, 4h, 6h and 8h were tested respectively. The test results are shown in Table 3 and FIG. 6.

Table 3 The release rate of the active ingredient after soaking in the simulated colon solution after the NADH-containing biopolymer nanospheres in Example 2 are made into tablets

Time h	Release rate%
0	0
2	39.57
4	66.85
6	85.5
8	93.27
remaining	2.10
Total	95.37

The prepared preparation can release NADH active ingredients within 8 to 10 hours after oral administration. It stays in the gastric juice and releases the active ingredients 2 hours before the preparation, and stays in the small intestine and releases the active ingredients after 6 to 8 hours. The above test results show that, under the immersion of the simulated gastric acid solution and the simulated colon solution, the active ingredient NADH in the prepared tablet can be slowly released into the solution and is a long-acting medicament.

In Example 3, the microcrystalline cellulose in the excipient serves as a filler and a disintegrant, and polyvinylpyrrolidone plays a role in protecting and dispersing the drug, which is helpful for drug release.

Compared with the prior art, the above technical solution of the present invention has the following advantages:

(1) The biopolymer nanosphere of the present invention includes a biopolymer carrier and NADH dispersed on the biopolymer carrier. The biopolymer carrier is a polymer fiber polymer, and its fiber skeleton forms a three-dimensional interpenetrating network structure through random arrangement, cross arrangement, and crimp arrangement, so that NADH is protected and its exposure to light or The problem of easy decomposition after oxygen prolongs the preservation time of NADH active ingredients and reduces the difficulty of preservation.

(2) In the biopolymer nanospheres of the present invention, most of the NADH is dispersed inside the three-dimensional interpenetrating network of the biopolymer carrier, and a small part is loaded on the outer surface of the biopolymer carrier. This three-dimensional interpenetrating network structure biopolymer carrier has the functions of biological activity and loading NADH active ingredients, so that the NADH loaded between the network structures is protected, and the problem that NADH is rapidly decomposed by stomach acid and cannot be fully utilized is solved. , Is a polymer carrier with intestinal sustained-release effect, and provides an ideal drug carrier for NADH to treat diseases such as Parkinson's disease, Alzheimer's disease, depression and cancer. By slowly releasing the active ingredients of NADH and activating DNA repair enzymes to repair DNA, it can further prevent the occurrence of cancer.

(3) The biopolymer carrier of the present invention is chitosan and/or konjac glucomannan. Both chitosan and konjac glucomannan can form a three-dimensional interpenetrating fiber network, which is easy to load NADH in the network Be protected. In addition, chitosan is a natural polymer material with good biofunctionality and compatibility, safety and microbial degradation, and because of its positive charge, it also has a bacteriostatic effect; konjac glucomannan Sugar is a natural high-molecular soluble dietary fiber with high viscosity, high water absorption and fast expansion. Due to its special glycosidic bond structure, it is also immunogenic.

(4) The process for preparing the biopolymer nanospheres according to the present invention first prepares a physically modified biopolymer carrier, and mixes and grinds the biopolymer raw materials with xanthan gum and/or sodium alginate to Long biopolymer chains are cut into molecular chains of moderate length. The physically modified biopolymer material has a moderate chain length, and it is easier to form spherical particles to form a honeycomb three-dimensional interpenetrating network structure, which is beneficial to the loading of NADH small molecules on the inner and outer surfaces of the network structure. In addition, the physically modified biopolymer carrier has a membrane-like structure on its outer surface, which has the function of waterproofing and oxidation resistance, solves the problem of NADH decomposing by light and oxygen, and prolongs the preservation time of NADH; by grinding NADH raw materials After sieving, NADH is ground to a particle size of tens of nanometers, which makes it easier for NADH to enter the three-dimensional interpenetrating network structure of biopolymer materials and be protected.

(5) The biopolymer nanospheres described in the present invention can be applied to the preparation of tablets, granules, capsules or soft capsules. First, by preparing biopolymer nanospheres containing NADH, the NADH is protected from the network by biopolymer carriers In the structure, it is re-made into a pharmaceutical dosage form, which prevents the problem that NADH in the dosage form is decomposed by gastric juice and cannot be absorbed by the human body. In the above dosage forms, soft capsules disperse NADH-containing biopolymer nanospheres in the oil phase, which isolates the contact of NADH with air and water, further solves the problem of easy decomposition of NADH, and has a longer shelf life.

The above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to the preferred embodiments, those of ordinary skill in the art should understand that the technical solutions of the present invention can be modified or equivalently replaced Without departing from the spirit and essence of the technical solution of the present invention.

Claims (11)

1. A biopolymer nanosphere containing NADH is characterized in that it includes a biopolymer carrier and NADH dispersed on the biopolymer carrier.
2. The biopolymer nanosphere containing NADH according to claim 1, wherein the biopolymer carrier has a three-dimensional network structure, and the average particle size of the biopolymer carrier is 200-1000 nm.
3. The NADH-containing biopolymer nanosphere according to claim 1, wherein the biopolymer carrier includes at least one of chitosan and konjac glucomannan;

   Preferably, the konjac glucomannan is at least one of konjac glucomannan, quaternized konjac glucomannan, carboxymethyl konjak glucomannan and deacetylated konjac glucomannan.
4. The biopolymer nanospheres containing NADH according to claim 1, characterized in that the biopolymer nanospheres include 1-20 parts of the NADH and 20-80 parts of the organism in parts by weight Polymer carrier.
5. The biopolymer nanosphere containing NADH according to claim 1, characterized in that the biopolymer nanosphere further comprises an auxiliary material, and the auxiliary material includes 0.05 to 30 parts of sodium alginate and 0.05 in parts by weight. ~30 parts of at least one of xanthan gum.
6. The NADH-containing biopolymer nanosphere according to claim 1, wherein the average particle diameter of the biopolymer nanosphere is 500-1000 nm.
7. A method for preparing NADH-containing biopolymer nanospheres according to any one of claims 1-6, characterized in that it includes the following steps:

   Mixing and grinding the biopolymer raw material and the auxiliary materials to physically modify the biopolymer raw material to obtain a biopolymer carrier;

   Grind NADH raw material and sieve to obtain NADH;

   Mixing and stirring the NADH with the biopolymer carrier to obtain the NADH-containing biopolymer nanosphere, wherein NADH is dispersed on the biopolymer carrier;

   Preferably, the auxiliary material includes at least one of 0.05-30 parts of sodium alginate and 0.05-30 parts of xanthan gum in terms of parts by weight.
8. An application of the biopolymer nanospheres containing NADH according to any one of claims 1 to 6 in the preparation of drugs for preventing and treating sub-health and tumors and functional foods.
9. A pharmaceutical preparation comprising the NADH-containing biopolymer nanosphere according to any one of claims 1-6 and pharmaceutically acceptable excipients;

   Preferably, the pharmaceutical preparation is one of tablets, capsules, granules, injections, tinctures, suppositories, patches, pills, syrups, mixtures, powders, lotions, films and dripping pills;

   Preferably, the pharmaceutical preparation is a capsule, and the capsule includes hard capsules and soft capsules.
10. A method for preparing the pharmaceutical preparation according to claim 9, characterized in that it comprises the following steps:

    Providing the NADH-containing biopolymer nanospheres, adding granulation auxiliary materials to the biopolymer nanospheres and stirring;

    Screening to obtain granular wet material with uniform particle size;

    Drying the wet material of particles to obtain particles of biopolymer nanospheres containing NADH;

    Preferably, the method further includes tabletting the NADH-containing biopolymer nanosphere particles into tablets;

    Preferably, the method further includes filling the NADH-containing biopolymer nanosphere particles into a capsule shell to make a capsule;

    Preferably, the method further comprises mixing the NADH-containing biopolymer nanosphere particles with edible oil and filling the soft capsule shell to make a soft capsule;

    Preferably, the granulation auxiliary material is at least one of microcrystalline cellulose, polyvinylpyrrolidone, sodium bicarbonate, magnesium stearate, calcium polyphosphate, wetting agent and binder.
11. A functional food, comprising the biopolymer nanospheres containing NADH according to any one of claims 1-6 and food additives.

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