CN100579575C - Peptide protein drug sublingual instant nano drug film and its three-dimensional printing preparation method - Google Patents

Abstract

The invention relates to a sublingual instant nano drug film of polypeptide protein drugs and its three-dimensional printing preparation method. Formed by conjunctival fluid bonding, the central area of the middle layer is formed by bonding the binder of the microemulsion system containing polypeptide protein drugs and soybean phospholipids, and the peripheral area is formed by bonding the same adhesive liquid as the top and bottom layers; preparation: prepared by nanotechnology For the microemulsion system of polypeptide protein drugs, the microemulsion system is positioned and printed in the center of the membrane by three-dimensional printing technology, and the peripheral area is sprayed with a binder containing a membrane penetration enhancer through another print head to form the drug film. The drug film can be quickly dissolved under the tongue to release the microemulsion particles containing the drug, and under the action of the penetration enhancer, it can pass through the sublingual mucosa and enter the circulatory system. The preparation process of the whole process is simple, highly automated and reproducible.

Description

Peptide protein drug sublingual instant nano drug film and its three-dimensional printing preparation method

technical field

The invention belongs to the field of drug delivery systems for polypeptide protein drugs, in particular to a sublingual instant nano drug film for polypeptide protein drugs and a three-dimensional printing preparation method thereof.

Background technique

The three-dimensional printing (Three Dimensional Print, 3DP) forming technology first proposed by MIT Sachs et al. (US patent, NO.5204055, 1993) is based on the concept of "layer-by-layer printing, layer-by-layer superposition" to prepare a special shape. or objects with complex internal structures. This technology uses powder as the material, the processing process is very flexible, the forming speed is fast, the operating cost is low and the reliability is high. It is one of the most viable new technologies in the rapid prototyping industry. The key equipment of this technology - 3D printer is generally composed of computer terminal, powder processing system (including powder feeding, layering and recycling), nozzle and adhesive supply device, precision platform and mobile device.

3DP forming technology has a high degree of processing flexibility that has never been seen in traditional manufacturing. It does not require various tools in traditional powder processing and is not limited by any geometric shape. Since the spraying position, spraying frequency and spraying speed can be controlled at will; different materials can be sprayed by different nozzles; spraying substances can be solutions, suspensions, emulsions and molten substances, etc., so 3DP forming technology can easily control local materials Composition, microstructure and surface properties. At the same time, because many traditional processing processes are unified into a process of repeated bonding on one machine, it is easy to design and research, and there is no problem of scale in the transformation process to industrial production, which can save a lot of time and money, and truly embodies Advantages of rapid prototyping technology. Compared with other rapid prototyping technologies, 3DP forming technology has its unique advantages: compared with laser selective sintering, equipment manufacturing costs and process operating costs are much lower; compared with fused deposition, it can be operated at room temperature and runs more efficiently. Convenient and reliable; the method of spray bonding avoids the use of laser or heating and melting, and will not affect the activity of active ingredients. For this reason, from the moment the 3D printing technology appeared, it has started various application research in the field of pharmacy.

For example, Wu et al [J.control.Release, 1996, 40 (1): 77-87] first used 3DP forming technology to carry out research on the preparation of implanted drug delivery systems; Katsta [J.control.Release, 2000 (66): 1 -9] and Rowe[J control.Release, 2000 (66): 11-17] etc. used 3DP forming technology to carry out the preparation research of oral sustained and controlled release drug delivery system; WO2000/29202 discloses a Oral-containing fast disintegrating tablets prepared by the technology; US2003/0198677A1 discloses a zero-order sustained and controlled release drug delivery system prepared by 3DP forming technology. Drug concentration gradient distribution with high center and low circumference; Yu et al. [J Pharm Sci, 2007(96): 2446-2456] used 3DP forming technology to prepare an oral drug that obtains zero-order controlled release effect through the gradient distribution of rent-release materials. medicine system. The drugs used in all kinds of relevant literatures are based on dyes as models, or use small molecule synthetic chemical drugs as research objects. At present, the applicant has not found that 3D printing technology is specially applied to polypeptide protein drug delivery systems. There are no research documents and patents, and there is no report on the preparation of oral drug delivery system for polypeptide protein drugs by integrating the advantages of 3D printing technology and nanotechnology.

Contents of the invention

The purpose of the present invention is to provide a sublingual instant nano drug film of polypeptide protein drugs and its three-dimensional printing preparation method, the use of three-dimensional printing technology can accurately control the dosage and the position of the drug in the preparation, and the good membrane penetration performance of the phospholipid nano system , to prepare a nano drug film that can be rapidly dissolved under the tongue. The whole process is simple, highly automated and reproducible.

The sublingual instant nano drug film of polypeptide protein drugs of the present invention is a membrane formed by bonding multi-layer powder, the top layer and the bottom layer of the membrane are formed by bonding liquid containing penetration enhancer, and the center The area is formed by bonding the binder of the microemulsion system containing polypeptide protein drugs and soybean phospholipids, and the surrounding area is formed by bonding the same adhesive liquid as the top and bottom layers.

The penetration enhancer is a penetration enhancer that can open the mucosal junction without damaging the mucosa, such as azone, sodium lauryl sulfate, etc.;

The binding liquid containing penetration enhancer is ethanol aqueous solution containing 0～8% penetration enhancer as binding agent, preferably the ethanol aqueous solution containing 4% penetration enhancer, the ethanol content in its ethanol aqueous solution is 60～100%, Preferably 90%;

The content of polypeptide protein drugs in the microemulsion system is 5-20%, preferably 10%;

The binder containing the microemulsion system can be suitable for the spraying system used by the three-dimensional printer.

The three-dimensional printing preparation method of the sublingual instant nano drug film of the polypeptide protein drug of the present invention comprises the following steps:

(1) Prepare the microemulsion system of polypeptide protein drugs by using nanotechnology, prepare layer powder and prepare powder forming binders for the peripheral areas of the top layer, bottom layer and middle layer;

(2) The powder feeding device of the system first transports the solid powder to the platform, and then rolls the layer by laying rods, and then the nozzles on the 3D printing system run on the fast and slow double tracks of the X-Y plane, selectively in different Binder is sprayed on the area to bond the powder together to form a two-dimensional layered sheet;

(2) On the Z-axis, the powder bed is driven down by the piston to determine the height (that is, the thickness of the powder layer), and a new layer of powder layer and bonded printing are performed, and so on, until the three-dimensional object to be processed is sprayed and formed, and the process is carried out. Appropriate post-processing (such as drying, powder removal, appropriate compression, etc.) can obtain a three-dimensional solid product.

The layer-laying powder is that each layer is covered with the same mixed powder, and the powder composition is Kollidon 25 and lactose, and the weight ratio is 60-90:10-40, especially 75:25.

The sublingual instant nano drug film of polypeptide protein drugs provided by the present invention, due to the porosity and the hydrophilicity of the auxiliary materials, can make the film dissolve rapidly within a few seconds without drinking water, and release it in order releases the penetration enhancer in the peripheral areas of the top, bottom, and middle layers, and then releases the microemulsion particles in the center of the middle layer. Under the action of the penetration enhancer, the microemulsion particles can quickly pass through the sublingual mucosa and enter the blood circulation system, avoiding the degradation of polypeptide and protein drugs by the first-pass effect of the gastrointestinal tract and liver.

The preparation method of the present invention uses computer-aided (CAD) to design sublingual instant nano drug film, provides a drug delivery system model containing material information, and directly controls the operation by outputting instructions from the computer terminal through the drug delivery system model and the forming machine data exchange interface program Preparation; preparation is carried out by "printing layer by layer, layer by layer superposition". All layers are covered with the same mixed powder, and different layers and different areas of each layer can be bonded and formed by spraying different binders with multi-nozzle 3D printers.

The present invention provides a determination and optimization scheme for the process parameters of the sustained and controlled release drug delivery system prepared by three-dimensional printing forming technology: through the "drop test" (dropping the binder onto the corresponding powder outside the three-dimensional printing system), strip test ( Use a 3D printing system to spray lines and strips on the corresponding powder bed, etc., observe and compare the bonding effect, bonding drying speed, deformation and shrinkage after bonding, the number of defects in the bonding strip, and the strength of the strip with a microscope , Determine and optimize the interval time between layers, the thickness of the powder layer, the spraying rate (spraying drop volume × spraying frequency), the number of spraying times and other forming preparation process parameters.

The drug film of the present invention can quickly dissolve under the tongue, release the microemulsion particles containing the drug, and under the action of the penetration enhancer, pass through the sublingual mucosa and enter the circulatory system. The whole process is simple, highly automated, and heavy Good performance.

Description of drawings

Figure 1 is a schematic diagram of the three-dimensional printing technology process;

Fig. 2 is the structural diagram of multi-layer nano drug film;

Fig. 3 is microemulsion particle diameter laser granulometer measurement figure;

Fig. 4 is the experimental diagram of the in vitro porcine tongue mucosa drug administration of the nano-medicine film;

Fig. 5 is the cumulative penetration of insulin through the sublingual mucosa of the drug film, the microemulsion and the control solution.

Detailed ways

Below in conjunction with specific embodiment, further illustrate the present invention. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. In addition, it should be understood that after reading the teachings of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the appended claims of the present application.

Example 1

(1) Preparation of insulin microemulsion

Insulin (titer 24.5IU·mL ^-1 ) was dissolved in pH 7.4 phosphate buffer solution to prepare 3.0 mg·mL ^-1 insulin solution. Soybean lecithin was dissolved in propylene glycol under the condition of ultrasound at a ratio of 1:3. The insulin solution was added dropwise to the propylene glycol solution of soybean lecithin at a ratio of 1:1 with stirring, and the 1.5 mg·mL ^-1 insulin microemulsion was prepared by proper ultrasonic treatment, which was used as the printing solution for the drug-loading area in the middle of the drug film.

Detection of microemulsion particle size: the particle size and distribution of microemulsion are measured by laser scattering method. Take 1.2mL samples and place them in a laser particle size analyzer (Nano ZS90 laser particle size analyzer, Malvern Instruments Co., Ltd., UK) to measure particle size and distribution. The particle size analyzer working conditions: the light source is He-Ne, the wavelength is 633nm, and the measurement temperature is 25°C. The microemulsion particle size laser particle size analyzer measurement chart is shown in Figure 3.

(2) 3D printing powder preparation

Pass Kollidon 25 and lactose through a 200-mesh sieve, mix the powder with a particle size of less than 74 μm evenly, and use it as a layer powder; weigh 2 grams of the penetration enhancer Azone and 15 grams of polyvinylpyrrolidone PVP 30, dissolve in 100 mL In 60% ethanol aqueous solution, the peripheral areas of the top layer, the bottom layer and the middle layer are formulated as powder forming adhesives.

The raw material composition and content (by weight percentage) of intermediate mixing powder are as follows:

Kollidon25 10 servings

Lactose 90 servings

(3) Determine the 3D printing forming parameters

Top and bottom surface spray forming parameters:

Layer interval time 2min

Powder layer thickness 100μm

Spraying rate [spraying drop volume (droplet quantity × droplet size) × spraying frequency] 4nL × 1200Hz

Spraying times 2 times

Parameters of intermediate medicated mixed powder spraying:

Layer interval time 4min

Powder layer thickness 100μm

Spray rate (spray drop volume × spray frequency) 4nL × 1200Hz

Spraying times 1 time

(4) Preparation of insulin nano-drug film

The operation preparation is directly controlled by the computer terminal output command, as shown in Figure 1. First spread a layer of mixed powder with a thickness of 100 μm, spray 2 times of binder containing penetration enhancer to form the bottom surface of the drug film, and then the piston rod drives the powder bed of the workbench to descend as a whole to prepare a new layer of powder.

The mixed powder of the middle three layers remains unchanged, the thickness of the layer is 100 μm, the microemulsion containing insulin is used as the binder, and the binder containing the penetration enhancer is sprayed on the outermost periphery to form.

Subsequently, another layer of mixed powder with a thickness of 100 μm was spread, and the binder containing the penetration enhancer was sprayed twice to form the bottom surface of the drug film. Finally, the obtained drug film is dried and powder-removed, and the structure diagram of the drug film is shown in FIG. 2 .

(5) Analysis of drug content in nano-drug film

A reversed-phase high performance liquid chromatography method was established to detect the content of insulin in the nano-drug film. The chromatographic column is HypersilODS ₂ (250mm×4.6mm, id5μm); Chromatographic conditions: mobile phase is 0.1mol L ^-1 sodium dihydrogen phosphate: 0.1mol L ^-1 sodium sulfate: acetonitrile=36:36:28, using Phosphoric acid was used to adjust the pH to 3.0; the flow rate was 1.0mL·min ^-1 ; the ultraviolet detection wavelength was 214nm; the injection volume was 20 μL, and the column temperature was 30°C. Use 0.01M HCl solution containing 0.15% TritonX-100 to dilute to the volume, shake well, filter through a 0.45 μm microporous membrane, and inject samples for analysis. Excipients do not affect drug determination.

After absorbance measurement, the content of insulin in the drug film was calculated according to the regression equation C=1.67322×10 ^-4 A+0.05702 (linear range: 5-200 μg·mL ^-1 ), and the result was 2.61±0.07 mg (n=6). The content difference between films is 2.68%, and the potency is 63.95±1.75IU/tablet.

(6) In vitro instant dissolving performance of nanomedicine film

According to the Chinese Pharmacopoeia 2005 edition appendix disintegration test method, the instant disintegration time limit of the drug film was determined. Since the minimum unit of the LD-2D disintegration time limit tester (Shanghai Huanghai Drug Inspection Instrument Factory) is minutes, a stopwatch is used for timing. The dissolving time of 6 fast-dissolving films in 25°C water is determined to be 28.7±3.6s. Put 4 pieces of drug film into 100mL water and stir until completely dispersed, the formed uniform dispersion can pass through a sieve with a caliber of 480μm, and the fineness of the dispersion meets the requirements.

(7) The drug delivery performance of the nano-drug film through the porcine tongue mucosa

Fresh pork tongue was taken from adult healthy pigs just after slaughter, and the sublingual mucosal tissue was removed, washed with normal saline, and cut into a certain size for later use. As shown in Figure 4, the sublingual mucosa of the fresh pork tongue is tightly stretched between the receiving pool and the drug supply pool of the diffusion cell. The surface of the mucosa faces the drug supply pool, and the inner layer of the mucosa faces the receiving pool. In the constant temperature (37±1)°C transdermal diffusion tester, 5 mL each of insulin nanofilm, insulin microemulsion and insulin control solution were added to the administration pool, and the receiving solution was 5 mL of 0.9% normal saline. Take 1 mL of the receiving solution regularly at 10, 20, 30, 60, 90, 120, and 180 min, and supplement with normal saline at the same temperature and volume. Insulin was detected at a wavelength of 214 nm.

The cumulative penetration of insulin was calculated by the regression equation C=1.67322×10 ^-4 A+0.05702, and plotted against the sampling time. The results are shown in FIG. 5 . Due to the synergistic effect of the penetration enhancer azone and phospholipids, the drug film has good transmembrane drug delivery performance, and the insulin permeation amount reaches (5.843±0.322) IU·mL ^-1 in 180 minutes, which is better than the permeation amount of microemulsion and insulin solution ( 5.591±0.240) IU·mL ^-1 and (2.457±0.181) IU·mL ^-1 . Especially in the first 10 minutes, the insulin permeation amount of the drug film reached (1.352±0.235) IU·mL ^-1 , far greater than the permeation amount of microemulsion and insulin solution (0.743±0.110) IU·mL ^-1 and (0.640±0.090 ) IU·mL ⁻¹ .

Claims (7)

1. A sublingual instant nano drug film of polypeptide protein drugs, characterized in that: the drug film is prepared by three-dimensional printing method, formed by bonding multilayer powder, the top and bottom layers of the film are made of The binder containing the penetration enhancer is bonded and formed, the central area of the middle layer is formed by bonding the adhesive of the microemulsion system containing polypeptide protein drugs and soybean phospholipids, and the surrounding area is formed by bonding the same adhesive as the top and bottom layers; The polypeptide protein drug is insulin; the penetration enhancer is azone; the binder containing the penetration enhancer is an ethanol aqueous solution of azone and polyvinylpyrrolidone; the powder has a weight ratio of 10:90 Composition of Kollidon 25 and lactose; The specific steps of the described three-dimensional printing method are as follows: (1) Using nanotechnology to prepare the microemulsion system of polypeptide protein drugs, preparing layer powder and preparing powder-forming binders for the surrounding areas of the top layer, bottom layer and middle layer; (2) The powder feeding device of the system first transports the solid powder to the platform, and then rolls the layer by laying rods, and then the nozzles on the 3D printing system run on the fast and slow double tracks of the X-Y plane, selectively in different Spray the binder in the area to bond the powder together to form a two-dimensional layered sheet; (3) On the Z-axis, the piston drives the powder bed to descend as a whole to determine the height, and a new layer of powder layering and bonding printing are performed, and so on, until the three-dimensional objects to be processed are sprayed and formed, and dried, powder removed, and properly compressed , that is, a three-dimensional solid product is obtained. 2. The sublingual instant nanomedicine film of polypeptide and protein drugs according to claim 1, characterized in that: the binder containing penetration enhancer is 60-100% ethanol aqueous solution containing 0-8% penetration enhancer. 3. The sublingual instant nano-drug film of polypeptide and protein drugs according to claim 2, characterized in that: the binder containing penetration enhancer is 90% ethanol aqueous solution containing 4% penetration enhancer. 4 . The sublingual instant nano drug film of polypeptide and protein drugs according to claim 1 , characterized in that: the content of polypeptide and protein drugs in the microemulsion system is 5-20%. 5 . The sublingual instant nano drug film of polypeptide and protein drugs according to claim 4 , characterized in that: the content of polypeptide and protein drugs in the microemulsion system is 10%. 6. the preparation method of a kind of polypeptide protein drug sublingual instant nano drug film according to claim 1, comprises the following steps: (1) Using nanotechnology to prepare the microemulsion system of polypeptide protein drugs, preparing layer powder and preparing powder-forming binders for the surrounding areas of the top layer, bottom layer and middle layer; (2) The powder feeding device of the system first transports the solid powder to the platform, and then rolls the layer by laying rods, and then the nozzles on the 3D printing system run on the fast and slow double tracks of the X-Y plane, selectively in different Spray the binder in the area to bond the powder together to form a two-dimensional layered sheet; (3) On the Z-axis, the piston drives the powder bed to descend as a whole to determine the height, and a new layer of powder layering and bonding printing are performed, and so on, until the three-dimensional objects to be processed are sprayed and formed, and dried, powder removed, and properly compressed , that is, a three-dimensional solid product is obtained. 7. The preparation method according to claim 6, characterized in that: the powder layer is that each layer is evenly covered with the same mixed powder layer, and the powder composition is Kollidon 25 and lactose, and the weight ratio is 10:90.

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