KR101613403B1 - Drug delivery system by Gelatin Sponge or Gelatin microsphere and Drug delivery system prepared thereby - Google Patents

Abstract

The present invention discloses a method for preparing gelatin sponge or gelatin microspheres containing phosphate buffered saline (PBS) or phosphate buffer and is directed to improve the conventional process having problems of difficulty in controlling shapes caused by the collapse of pores during a refreezing process and irregular and non-uniform shapes of embolic agent granules during pulverization. The present invention relates to a method for preparing a gelatin-based drug delivery system that allows higher stability and shape retainability by setting detailed conditions in a series of processes to obtain desired gelatin foam and an additional process suggesting a post-treatment process for the thus obtained gelatin foam, as well as a gelatin-based drug delivery system obtained by the method. The gelatin-based drug delivery system is obtained by adding phosphate buffered saline or phosphate buffer to crosslinked gelatin foam, drying the foam while maintaining pH in a range from neutral pH to alkaline pH, and inducing ionic bonding with a liver cancer-treating agent.

Description

TECHNICAL FIELD [0001] The present invention relates to a method for manufacturing a gelatin drug delivery system and a gelatin drug delivery system,

The present invention relates to a gelatin drug delivery system applicable to carotid artery embolization (ICA), and more particularly to a gelatin sponge or a gelatin sponge containing a phosphate buffer (PBS) or a phosphoric acid buffer (Phosphate Buffer) In order to improve the conventional processes in which the morphological control due to the breakage of the pore size caused in the re-freezing process and the morphology of the precursor granules in the pulverization become irregular and non-uniform in starting the manufacturing process of the microsphere (Gelatin microsphere) A series of processes for obtaining a gelatin foam and a post-treatment process of the obtained gelatin foam are presented, and a gelatin drug delivery system which is manufactured so as to be more stable and advantageous in shape preservation by detailed setting conditions due to additional processes And a gelatin drug carrier prepared thereby.

Carotid chemoembolization is a treatment to block blood vessels by administering anticancer drugs to find arteries supplying nutrients to liver tumors. Unlike other organs, the liver is double-fed with oxygen and nutrients by two blood vessels. One is the portal vein that goes around the small intestine and the large intestine. The other is the hepatic artery that comes directly from the aorta.

Normal liver tissue is mainly supplied with blood in the context, but tumor tissue is supplied with blood from the hepatic artery. Therefore, when only hepatic artery supplying the tumor is selected, administration of the anticancer drug and blocking of the blood vessel after the administration of the anticancer drug, But only the tumor can be selectively necrosed.

Therefore, carotid artery chemoembolization, which induces tumor necrosis by artificially blocking the blood vessels supplying the oxygen and nutrients necessary for cancer cells to grow, has recently been popularized. The hospitalization period required for a single operation is as short as several days, And it is recognized as an excellent method for the treatment of liver cancer in recent years because it is less difficult to return to the society and less restrictive than the other treatments and limited in the patient 's liver function and systemic condition.

On the other hand, in the "Gelatin sponge containing phosphoric acid buffer solution or phosphate buffer applied to carotid artery chemoembolization and method for manufacturing the same" filed by the applicant on July 27, 2015 and registered as December 1, Lt; / RTI >

A step of immersing a gelatin sponge in a phosphate buffer (PBS) or a phosphate buffer at 4 ° C to 37 ° C for 3 to 12 hours and a step of immersing the gelatin precipitated with a phosphate buffer solution (PBS) or a phosphate buffer A step of lyophilizing the sponge at a temperature of -10 ° C to -35 ° C for 30 minutes to 16 hours, a step of pulverizing the lyophilized gelatin sponge, a step of pulverizing the pulverized gelatin sponge at 10 to 100 μm, Mu] m, 300-500 [mu] m, and 500-700 [mu] m, respectively. The pulverized gelatin sponge is stored in a sterilized vial, a phosphate buffer solution or a phosphate buffer solution, Saline solution, distilled water, and an anticancer drug are added to each other to be ion-bonded to each other to be used as a therapeutic agent for liver cancer.

The above-described technical idea is advantageous in that the process is relatively simple and easy to manufacture. However, in the process of immersing a lyophilized gelatin sponge in a phosphate buffer solution or a phosphate buffer solution and re-freezing it, It is difficult to control the shape of the product, and it is difficult to control the shape of the product due to the irregular shape of the pre-granule granules in the process of crushing the gelatin sponge through the crusher or the uniform size of the surface being cut off There is a problem.

In the disclosed process, if the moisture content can not be controlled to an appropriate level during the lyophilization, a phosphoric acid buffer solution is formed into a powder and a disadvantage that it becomes clogged with a gelatin sponge is caused.

- Registration No. 10-0588555 (registered date 2006.06.02) - Registration No. 10-0601293 (Registered on July 07, 2006) - Registration No. 10-1172066 (Registration date 2012. 08. 01)

The present invention has been made in order to overcome the problems of the prior art described above, and it is an object of the present invention to provide a method for treating hepatocellular carcinoma cells using phosphate buffer (PBS) or phosphate buffered saline (PBS) It is an object to be solved to provide a gelatin drug delivery vehicle containing a phosphate buffer (phosphate buffer).

It is another object of the present invention to provide a process for producing a gelatin drug delivery system in which the gelatin drug delivery system is formed in a gelatin sponge or a gelatin microsphere form, It is a challenge.

In addition, a gelatin drug delivery system capable of more effectively healing various problems such as the development of irregular shape of color precursor granules caused by breakage or crushing of pore size caused by re-freeze drying in the process of manufacturing the above- It is another problem to provide a method for producing

In order to accomplish the above object, the present invention provides a gelatin drug delivery system and a method of manufacturing the same.

The method for preparing a gelatin drug delivery system proposed in the present invention is a method for manufacturing a gelatin drug delivery system in which a phosphate buffer solution or a phosphate buffer is added to a crosslinked gelatin foam and the pH of the gelatin is maintained in a range from neutral to alkaline, And the like.

For this purpose, the crosslinked gelatin foam was washed five times with acetone or distilled water, centrifuged by a centrifuge, and then added to a phosphate buffer solution or phosphate buffer solution to maintain a pH of 7 or more. After centrifugal dehydration, Thereby producing a gelatin sponge or a gelatin microsphere.

Meanwhile, in order to obtain a gelatin concentration of 1 to 6 (% w / v) proposed in the present invention, gelatin is soaked in 100 ml of water for 30 minutes, heated until the gelatin is dissolved, A gelatin solution forming step of adding 0.1 to 2 mg / ml of formaldehyde and curing the gelatin solution at a speed of 500 to 1000 rpm, 1200 to 1400 rpm and 1400 to 1500 rpm to form a gelatin foam A step of washing the obtained gelatin foam with acetone or distilled water followed by centrifugal separation using a centrifugal separator, and a step of adding a centrifuged gelatin foam to a phosphate buffer solution or a phosphate buffer solution, A gelatin sponge forming step of forming a gelatin sponge by drying the gelatin foam with a freeze dryer; The method comprising the steps of: pulverizing and sorting the particles into particles of 50 to 100 μm, 100 to 300 μm, 300 to 500 μm, and 500 to 700 μm, storing them in the form of a packaging container, and diluting the sterilized water to a therapeutic agent for liver cancer, do.

The gelatin microspheres are prepared by dissolving gelatin in water at 30 to 70 ° C. and filtering to obtain a gelatin solution having a gelatin content of 15 to 60%; and a step of putting the gelatin solution into a stabilizer having a mass ratio of 1 to 2% After stirring for 15 seconds at a volume ratio of oil of 1/1 to 1/10 and 200 to 800 r / min, a compound of a crosslinking aldehyde having a mass ratio of 2 to 50% at a temperature of 10 ° C was added and solidified for 1 to 2 hours. A gelatin foam processing step in which the gelatin foam is added to a phosphate buffer solution or a phosphate buffer solution to maintain a pH of 7 or more, washed five times, and centrifugally dehydrated; Gelatin foam was dissolved in a spray dryer to obtain gelatin microspheres of 50 to 150 μm, 150 to 350 μm, 350 to 560 μm, 560 to 710 μm, 710 to 1000 μm, and 1000 to 1500 μm, Sterilized water It characterized by consisting of; dilute ionic bond derived by the step of input.

According to the present invention, the present invention provides a gelatin drug delivery system capable of treating hepatocellular carcinoma as an embolization agent by binding an anticancer agent to a gelatin sponge or a gelatin microsphere, wherein the gelatin is delivered in the form of a gelatin sponge or a gelatin microsphere It is more effective to provide a gelatin drug delivery system that has been improved from a conventional state by providing a specific and differentiated process for producing a gelatin drug delivery system.

In particular, in the present invention, various improvements such as breakage of pore size and irregular shape of pre-formed granules caused by re-freeze drying in the process of manufacturing a gelatin drug delivery vehicle are performed by a predetermined addition process, The gelatin foam thus obtained was washed five times with acetone to distilled water, centrifuged by a centrifuge, and then added to a phosphate buffer solution or phosphate buffer solution to maintain a pH of 7 or more, followed by centrifugal dehydration to induce ionic bonding with the therapeutic agent for liver cancer And thus can be used more effectively for carotid chemoembolization.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a flow chart schematically showing a method of manufacturing a gelatin drug delivery system according to a preferred embodiment of the present invention. FIG.
2 is a graph showing measured values of doxorubicin absorbance by concentration using three wavelengths in the experimental example presented in the present invention.
FIG. 3 is a graph showing the adsorption capacity of doxorubicin of gelatin particles according to the type of hydration solution in the experimental example presented in the present invention.
FIG. 4 is a graph showing doxorubicin eluted in physiological saline according to the experimental example presented in the present invention. FIG.

Hereinafter, the present invention will be described with reference to the accompanying drawings, and the present invention will be described in detail with reference to preferred embodiments.

BRIEF DESCRIPTION OF THE DRAWINGS The advantages and features of the present invention, and how to accomplish them, will become apparent by reference to the embodiments described in detail below with reference to the accompanying drawings. However, it is to be understood that the present invention is not limited to the disclosed embodiments, but may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. It is intended that the disclosure of the present invention be limited only by the terms of the appended claims. And, throughout the specification, like reference numerals refer to like elements.

The present invention is disclosed in relation to the technical idea of a gelatin drug delivery vehicle applied to carotid artery embolization.

In particular, the present invention relates to a process for preparing a gelatin sponge or a gelatin microsphere containing phosphate buffered saline (PBS) or a phosphate buffer (phosphate buffer) It is proposed to solve some problems of the registered patent documents.

That is, it is difficult to control the morphology due to the breakage of the pore size caused by the re-freezing process and to obtain a predetermined gelatin foam in order to improve the conventional process in which the shape of the pre- The present invention relates to a method for producing a gelatin drug delivery system and a method for manufacturing the gelatin drug delivery system, which are manufactured so as to be more stable and more advantageous in shape preservation due to detailed setting conditions caused by additional steps, .

As shown schematically in FIG. 1, the gelatin drug delivery system proposed by the present invention is characterized in that a phosphate buffer solution or a phosphate buffer is added to a crosslinked gelatin foam and the hydrogen ion concentration is maintained in a range from neutral to alkaline And then inducing ionic bonding with the liver cancer therapeutic agent.

For example, the crosslinked gelatin foam is washed five times with acetone or distilled water, centrifuged by a centrifuge, and then added to a phosphate buffer solution or phosphate buffer solution to maintain a pH of 7 or more. After centrifugal dehydration, And is produced by gelatin sponge or gelatin microsphere by inducing ionic bonding with a therapeutic agent.

The gelatin sponge of the present invention is divided into a predetermined step and a gelatin solution forming step; A gelatin foam forming process; Gelatin foam processing; A gelatin sponge forming process; And an ionic bond induction process are sequentially performed.

In detail, in order to obtain a gelatin concentration of 1 to 6 (% w / v), the gelatin sponge is immersed in 100 ml of water for 30 minutes, heated to reduce the gelatin to dissolve, A gelatin solution forming step in which the gelatin solution is formed by adding formaldehyde at a concentration of 0.1 to 2 mg / ml and curing the gelatin solution at a speed of 500 to 1000 rpm, 1200 to 1400 rpm and 1400 to 1500 rpm, A centrifuging step of washing the obtained gelatin foam with acetone or distilled water and then centrifuging using a centrifugal separator; and adding a centrifuged gelatin foam to a phosphate buffer solution or a phosphate buffer solution, A gelatin sponge forming step of drying the gelatin foam by a freeze dryer to form a gelatin sponge; And an ionic bonding inducing step in which the ponge is pulverized and sorted into particles of 50-100um, 100-300um, 300-500um, and 500-700um, stored in a packaging container, and then diluted with sterilized water to the liver cancer treatment agent.

The gelatin foam processing process described above is given specific conditions as described above in order to eliminate the problems inherent in the conventional processes.

In the conventional patent document proposed by the present applicant, the freeze-dried gelatin sponge is immersed in a phosphate buffer solution or a phosphate buffer solution, and then subjected to a re-freeze drying process, which inevitably breaks the pore size of the sponge, It is difficult to crush the gelatin sponge through the crusher. In addition, in the process of crushing the gelatin sponge through the crusher, the shape of the precursor granules is irregular or the size of the surface is not uniform as in the case of cutting the gelatin. After the foam is cured, the remaining formaldehyde is washed with acetone or distilled water and then washed five times with phosphate buffer solution or phosphate buffer solution while maintaining the condition of hydrogen ion concentration of 7 or more. do.

In other words, unlike the conventional method, instead of using a ready-made gelatin sponge, a more detailed and specific process for forming a predetermined gelatin sponge is carried out, and after the processed gelatin foam is maintained at a proper hydrogen ion concentration, It is possible to escape from the various problems brought about by the time.

As will be described later, as a result of confirming through the present embodiment, the present invention is limited to the range from neutral to alkaline due to the characteristic that more anticancer agent binds when the pH is alkaline, that is, 11, but it is important that the pH is maintained at 7 or more.

The buffer solution according to the present invention plays a role of maintaining the pH of the solution so that the pH does not change abruptly. Here, the buffer solution may include buffers of Tris and glycine, PBS and PB as well as a mixture of two or more buffers It is most preferable to use PBS to PB buffer solution.

Most preferably, the pH of the Tris buffer constituting the buffer solution is pH 9 regardless of the molar concentration, and the pH is preferably 8 at the molar concentrations of 12.5 mM and 25 mM. The pH of the glycine buffer is preferably 9 at 25 mM, and the pH of the sodium phosphate buffer at 12.5 mM and 25 mM is preferably 8, and the pH at 7 is most preferred when the sodium phosphate buffer concentration is 25 mM. It is most preferable to remove NaCl from the PBS buffer, but it is not limited thereto.

The centrifugal dewatering as described above has an excellent effect of achieving an appropriate moisture content in order to improve the problem that when the water content is not adjusted to an appropriate level during lyophilization, the phosphate buffer solution is formed into a powder and is entangled with the gelatin sponge .

Meanwhile, the gelatin microspheres of the present invention are prepared by dividing the gelatin microspheres into predetermined steps, including a gelatin solution forming step; A gelatin foam forming process; Gelatin foam processing; Ion-coupling induction process.

Specifically, the gelatin microsphere is prepared by dissolving gelatin in water at 30 to 70 ° C and filtering to obtain 15 to 60% of a gelatin solution; and a gelatin solution in a mass ratio of 1 to 2 % Stabilizer, and the mixture was stirred at a volume ratio of water and oil of 1/1 to 1/10 and 200 to 800 r / min for 15 seconds. Then, a compound of a crosslinking aldehyde having a mass ratio of 2 to 50% Gelatin foam, which is obtained by coagulation for 2 hours, washing or centrifugation of the cross-linking agent, gelatin foam is added to phosphate buffer solution or phosphate buffer solution to maintain pH 7 or more, Gelatin foam processing step of dewatering gelatin foam by dissolving gelatin foam and securing gelatin microspheres of 50-100um, 100-300um, 300-500um, 500-700um, 700-1000um, 1000-1,500um by a spray dryer, After storage, Include; diluted with sterile water to the ion-binding cancer induction step of commitment.

Now, each embodiment for obtaining a gelatin sponge and a gelatin microsphere for realizing the gelatin drug delivery system proposed in the present invention will be described.

In Example 1, the following procedure was sequentially performed to obtain a gelatin sponge of the level required in the present invention.

1. Gelatin was dipped in 100 ml of water for 30 minutes to obtain a gelatin concentration of 1 to 6 (% w / v), heated to dissolve the gelatin, and then cooled down to a temperature of 40 ° C.

2. Formaldehyde at a concentration of 0.1 to 2 mg / ml was added to the gelatin solution, and the resulting gelatin solution was cured at a speed of 500-1000 rpm, 1200-1400 rpm, and 1400-1500 rpm to form a predetermined gelatin foam.

3. The obtained gelatin foam was washed with acetone or distilled water and centrifuged using a centrifuge.

4. Centrifuged gelatin foam was added to phosphate buffer solution or phosphate buffer solution to maintain pH 7 and above, washed five times, and centrifuged.

In this case, phosphate buffer (PBS) or phosphate buffer (Phosphate Buffer) at 4 ° C to 37 ° C was used and immersed for 3 to 12 hours.

5. The gelatin foam obtained in 4 was dried with a freeze dryer to form a gelatin sponge.

The gelatin sponge precipitated with ginseng salt buffer (PBS) or phosphate buffer (Phosphate buffer) was lyophilized for 30 minutes to 16 hours at a temperature of -10 ° C to -35 ° C to obtain a required level of gelatin sponge I went through the process.

6. After the dried gelatin sponge is pulverized into particles of 50-100um, 100-300um, 300-500um, 500-700um, and stored in a packaging container, the sterilized water is diluted into the liver cancer treatment agent, Binding.

For example, the gelatin sponge that has undergone pulverization and screening has been stored in a packaging container containing either a sterilized vial, a phosphate buffer solution, or a phosphate buffer solution, and then physiological saline, distilled water, Could be used as a treatment for liver cancer.

In Example 2, the following procedure was sequentially performed to obtain gelatin microspheres of the level required in the present invention.

1. A predetermined gelatin was dissolved in water at 30 to 70 占 폚 and filtered to obtain a 15 to 60% gelatin solution.

The gelatin solution obtained in 2.1 was put into a span 80 as a stabilizer having a mass ratio of 1 to 2% and stirred for 15 seconds at a volume ratio of water and oil of 1/1 to 1/10 and 200 to 800 r / min.

3. The stabilized gelatin solution was mixed with a crosslinking aldehyde compound having a mass ratio of 2 to 50% at a temperature of 10 ° C., and the gelatin solution was coagulated for 1 to 2 hours. The crosslinking agent was washed or centrifuged to remove the gelatin foam.

4. Gelatin foam was added to phosphate buffer solution or phosphate buffer solution to maintain pH 7 or higher, washed five times, and centrifuged.

In this case, phosphate buffer (PBS) or phosphate buffer (Phosphate Buffer) was used at a temperature in the range of 4 ° C to 37 ° C, and the immersion duration was 3 to 12 hours.

5. Dissolve the gelatin foam and store in gelatin microsphere of 50 ~ 100um, 100 ~ 300um, 300 ~ 500um, 500 ~ 700um, 700 ~ 1000um, After that, ionic bond was induced by adding sterilized water to liver cancer treatment agent by dilution.

The gelatin microsphere obtained by the above process is stored in a packaging container in a sterilized vial, a phosphate buffer solution, or a phosphate buffer solution. Then, physiological saline, distilled water and an anticancer drug are injected into the gelatin microsphere, And it could be used as one treatment for liver cancer.

The gelatin microsphere obtained in Example 2 was found to be a fine spherical color precursor. The interior of the gelatin microsphere was found to have a good absorption capacity in the form of a sponge, a uniform size, and a certain elasticity and swelling capacity.

(PBS) or phosphate buffer (PBS) at 4 ° C to 37 ° C to form a gelatin sponge containing a phosphate buffer (PBS) or a phosphate buffer (phosphate buffer) applied to carotid artery embolization as described above. (Phosphate Buffer) and immersed for 3 to 12 hours. The gelatin sponge is then lyophilized at a temperature of minus 10 ° C to minus 35 ° C for 30 minutes to 16 hours to adjust the moisture content to 10-20%.

In this process, gelatin sponge precipitated with phosphate buffer (PBS) or phosphate buffer (Phosphate Buffer) is lyophilized at -10 ℃ ~ -35 ℃ for 16 ~ 48 hours. The process of making the water present in the range of 0 to 10% can be carried out according to the work order.

Thereafter, the lyophilized gelatin sponge is pulverized, and the pulverized gelatin sponge is separated into a size of 50 to 100 μm, 100 to 300 μm, 300 to 500 μm, and 500 to 700 μm by means of a net or the like And then packaged in a sterilized vial bottle.

On the other hand, a gelatin sponge which has not been frozen is cut and immersed in a phosphate buffer (PBS) or a phosphate buffer (phosphate buffer) at 4 ° C to 37 ° C for 3 to 12 hours in a standardized state, . The gelatin sponge crushed by the standard is stored in a packaging container which is contained in a sterilized vial bottle, a phosphate buffer solution or a phosphate buffer solution. Then, physiological saline solution, distilled water and an anticancer drug are injected and combined with each other, Is used.

For example, a gelatin sponge in which a phosphate buffer solution (PBS) or a phosphate buffer has been precipitated can be dried by any one of dehydration drying, nitrogen drying, cold air drying, vacuum drying and natural drying methods in addition to the freeze drying method.

Experiment 1

For the present invention, doxorubicin for rectal use was Adriamycin AL DF (10 mg vial) manufactured by Ildong Pharma. A stock solution of 1 mg / ml was prepared by dissolving the drug in 10 ml of sterilized distilled water.

Using the characteristic of red color of doxorubicin in aqueous solution, it was confirmed whether it is easy to quantify doxorubicin by using spectrophotometer instead of HPLC which is difficult to manage quality.

 The absorbance of the doxorubicin solution was measured using various wavelengths to find the wavelength of the light source having a proportional correlation between the measured absorbance and the concentration of doxorubicin.

First, doxorubicin was diluted in distilled water and prepared at concentrations of 20 ug / ml, 40 ug / ml, 60 ug / ml, 80 ug / ml and 100 ug / ml. Absorbance values were measured at 420 nm, 450 nm and 510 nm.

The measurement results are shown in Table 1.

In all three wavelengths, the absorbance of each concentration of doxorubicin was directly proportional to the measured value. In this study, the experiment was conducted using the wavelength of 450 nm.

Absorbance by concentration of doxorubicin according to measurement wavelength Wave length
Doxorubicin (ug / ml) 420 nm 450 nm 510 nm 20 0.154 0.274 0.278 40 0.31 0.558 0.585 60 0.464 0.847 0.886 80 0.627 1.125 1.194 100 0.776 1.407 1.487

Experiment 2

Particles were hydrated in various solutions to investigate the conditions under which doxorubicin adsorbed on the gelatin microparticles. The solutions used here are 12 buffer solutions containing phosphate buffered saline (PBS), sterile water and normal saline.

Gelatin fine particle hydration solution No. Solution No. Solution One 200 mM Tris-HCl pH 9.8 8 100 mM citrate pH3 2 200 mM Tris-HCl pH8 9 100 mM citrate pH4 3 200 mM Tris-HCl pH 7.5 10 100 mM citrate pH 5 4 200 mM Tris-HCl pH 6.8 11 100 mM citrate pH 6 5 200 mM KCl pH2 D.W Sterilized water 6 200 mM glycine pH3 saline 0.85% NaCl 7 200 mM glycine pH 9.6 PBS Phosphate Buffered Saline pH 7.5

The hydration process of gelatin particles is summarized as follows. 10 mg of the gelatin particles were added to each of the 14 kinds of the solutions shown in Table 2 and stirred overnight at 4 캜. 1 ml of the supernatant was transferred to an e-tube and vortexed for 10 minutes. After centrifugation at 13,000 rpm for 2 minutes, supernatant was removed and 1 ml of doxorubicin diluted to 200 ug / ml was added and vortexed for 10 minutes. After centrifugation at 13,000 rpm for 2 minutes, the absorbance of the supernatant was measured at 450 nm.

The results are shown in the following Table 3 and FIG. 3. As the absorbance and the concentration of doxorubicin were lower, it was confirmed that the drug adsorbed well on the gelatin fine particles.

Experiment 3

The ability of the gelatin particles to adsorb doxorubicin according to the type of hydration solution (measured absorbance above supernatant and doxorubicin concentration) Solution Absorbance Conc. of doxorubicin (ug / ml) 0.2 mg / ml 2.762 195.0493 D.W 2.736 193.2183 saline 2.686 189.6972 PBS 0.841 59.76761 One 0.094 7.161972 2 0.519 37.09155 3 0.866 61.52817 4 1.238 87.72535 5 2.85 201.2465 6 2.842 200.6831 7 0.103 7.795775 8 2.768 195.4718 9 2.736 193.2183 10 2.522 178.1479 11 1.47 104.0634

The gel particles which were hydrated with distilled water and physiological saline had almost no adsorption of doxorubicin and a considerable amount of doxorubicin was adsorbed to gel particles which were hydrated in PBS. However, the remarkable fact is that regardless of the type of buffer, the adsorption of doxorubicin depends on the pH condition. In the case of Tris and glycine buffers, most of the drugs were adsorbed at pH 9.6 ~ 9.8, which is the strongest alkaline, and no adsorption of doxorubicin was observed in glycine and citrate buffers at pH 3 ~ 4.

These results indicate that the adsorption can be understood as a phenomenon in which the adsorption is more likely to occur in a high pH alkaline state and the acidic state becomes stronger, and the ionic bond is a chemical bond satisfying these conditions.

To confirm that the chemical binding of doxorubicin adsorbed on the gelatin microparticles is reversible, doxorubicin-adsorbed gel particles were reacted with physiological saline at neutral pH. Gel particles with red color were suspended in distilled water, and the supernatant was discarded by centrifugation at 13,000 rpm for 2 minutes. 0.5 ml of physiological saline was added to resuspend the gel particles, followed by vortexing for 10 minutes. The mixture was decanted again at 13,000 rpm for 2 minutes, and the absorbance of the supernatant was measured at 450 nm.

The measurement results are shown in Table 4.

Doxorubicin adsorbed on gel particles in PBS, Tris and citrate buffers was eluted in physiological saline solution. The degree of elution was confirmed to be the best separation of the adsorbed drug by the buffer solution with near neutral pH.

Taken together, these observations indicate that the adsorption rate of doxorubicin on gel particles increases with the alkalinity of the hydration buffer, and in contrast, the dissolution rate of the adsorbed drug increases when the buffer near neutral is used for hydration.

Doxorubicin eluted in physiological drinking water Solution Absorbance Conc. of doxorubicin (ug / ml) D.W 0.003 0.753521 saline 0.006 0.964789 PBS 0.663 47.23239 One 0.117 8.78169 2 0.421 30.19014 3 0.621 44.27465 4 0.777 55.26056 5 0.001 0.612676 6 0.004 0.823944 7 0.032 2.795775 8 0.003 0.753521 9 0.001 0.612676 10 0.007 1.035211 11 0.483 34.55634

Experiment 4

In order to investigate the ability of gelatin particles to adsorb doxorubicin according to the type and concentration of hydration solution, particles were hydrated in various solutions.

The solutions used were Tris buffer, Glycine buffer, sodium phosphate buffer, DW, 0.2 mg / ml, PBS pH 7.4, PBS pH 7.4 without KCl, PBS pH 7.4 without NaCl minus KCl and NaCl And 11 kinds of buffers, sterilized water and normal saline containing phosphate buffered saline (PBS) such as PBS pH 7.4.

Gelatin fine particle hydration solution No. Buffer 12.5 mM 25mM 50 mM 100 mM 200 mM One Tris-HCL pH 9 0.154 0.109 0.087 0.174 0.141 2 Tris-HCL pH8 0.208 0.141 0.331 0.526 0.552 3 Tris-HCL pH7 0.726 0.685 0.793 1.033 1.056 4 Tris-HCL pH6 2.062 1.864 1.614 1.774 1.816 5 Glycine pH9 0.109 0.057 0.505 0.394 0.206 6 Glycine pH8 1.812 1.063 0.505 0.394 0.206 7 Glycine pH7 2.358 2.360 2.274 2.178 1.836 8 Glycine pH6 2.36 2.364 2.302 2.386 2.404 9 Sodium phosphate buffer pH8 0.178 0.173 0.401 0.901 0.922 10 Sodium phosphate buffer pH7 0.523 0.159 0.686 1.232 1.191 11 Sodium phosphate buffer pH6 1.548 1.308 1.594 1.796 1.782 12 D.W 2.498 13 0.2 mg / ml 2.578 14 PBS pH 7.4 0.723 15 PBS (without KCl) pH 7.4 0.904 16 PBS (without NaCl) pH 7.4 0.399 17 PBS (without KCl and NaCl) pH 7.4 0.408

Tris buffer exhibited strong adsorption regardless of molarity at pH 9 and adsorption power at 12.5 mM and 25 mM when Tris buffer was pH 8. The higher the concentration, the lower the adsorption power. Glycine buffer showed the strongest adsorption power at pH 9 and 25 mM, and the higher the molar concentration, the stronger the adsorption power. Sodium phosphate buffer pH 8 showed the strongest adsorption power at 12.5 mM and 25 mM, and sodium phosphate buffer pH 7 showed the strongest adsorption at 25 mM. And the PBS buffer showed the best adsorption power when NaCl was removed.

PBS pH7.4 and KCl and NaCl minus adsorption power 10X 5X 2X 1X 0.5X PBS pH 7.4 1.716 1.492 1.025 0.723 0.583 PBS (KCl x,
NaCl x) pH 7.4 0.858 0.602 0.563 0.408 0.443

Adsorption power according to PBS pH PBS pH9.5 0.605 PBS pH 4.5 2.500 100 mM Tris-phosphate pH 11 0.191

Experiment 5

Hydration The particles were hydrated in solution to investigate the adsorption capacity of doxorubicin of gelatin particles according to the pH and concentration of sodium phosphate buffer. The solution used here is one kind of buffer, sterilized water and normal saline.

Gelatin fine particle hydration solution 1: sodium phosphate buffer pH8 2: sodium phosphate buffer pH7 3: sodium phosphate buffer pH6 12.5 mM 25mM 50 mM One 0.178 0.173 0.401 2 0.523 0.159 0.686 3 1.548 1.308 1.594 D.W 2.498 0.2 mg / ml 2.578 12.5 mM 25mM 50 mM One 0.192 0.283 0.467 2 0.476 0.510 0.784 3 1.480 1.456 1.414 D.W 2.458 0.2 mg / ml 2.526

The gelatin drug delivery system obtained by the above-described structure provides a gelatin drug delivery system capable of treating hepatocellular carcinoma as an embolization agent by binding an anticancer agent to a gelatin sponge or gelatin microsphere, wherein the gelatin drug delivery system is in the form of a gelatin sponge or gelatin microsphere By providing a specific and differentiated process for producing a stable gelatin drug delivery system.

In particular, in the present invention, various improvements such as breakage of pore size and irregular shape of pre-formed granules caused by re-freeze drying in the process of manufacturing a gelatin drug delivery vehicle are performed by a predetermined addition process, The gelatin foam thus obtained was washed five times with acetone to distilled water, centrifuged by a centrifuge, and then added to a phosphate buffer solution or phosphate buffer solution to maintain a pH of 7 or more, followed by centrifugal dehydration to induce ionic bonding with the therapeutic agent for liver cancer And thus can be used more effectively for carotid chemoembolization.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be clearly understood that the same is by way of illustration and example only and is not to be taken by way of limitation, will be. Therefore, it is intended that the present invention cover the modifications and variations of this invention provided they come within the scope of the appended claims and their equivalents.

Claims (5)

delete delete The crosslinked gelatin foam was washed five times with acetone or distilled water, centrifuged by centrifuge, and then added to phosphate buffer solution or phosphate buffer solution to maintain pH 7 or more. After centrifugal dehydration, Thereby producing a gelatin drug delivery system comprising a gelatin sponge or a gelatin microsphere,
The gelatin sponge,
In order to obtain a gelatin concentration of 1 to 6 (% w / v), gelatin is dipped in 100 ml of water for 30 minutes, heated to reduce the gelatin to dissolution, and then lowered to a temperature of 40 ° C;
Forming a gelatin foam by adding formaldehyde at a concentration of 0.1 to 2 mg / ml and allowing the gelatin solution to cure at a speed of 500-1000 rpm, 1200-1400 rpm, and 1400-1500 rpm to form a gelatin foam;
A centrifugal separation step of washing the obtained gelatin foam with acetone or distilled water and then centrifuging using a centrifuge;
A gelatin foam processing step in which the centrifuged gelatin foam is added to a phosphate buffer solution or a phosphate buffer solution to maintain a pH of 7 or more, washed five times, and centrifugally dehydrated;
A gelatin sponge forming process for drying a gelatin foam with a freeze dryer to form a gelatin sponge;
An ion binding induction process in which a gelatin sponge is pulverized and sorted into particles of 50 to 100 urn, 100 to 300 urn, 300 to 500 urn, and 500 to 700 urn and stored in a packaging container, and then sterilized water is diluted in the liver cancer treatment agent. The method of claim 1,
The method of claim 3,
The gelatin microspheres may be prepared by,
A step of forming a gelatin solution by dissolving gelatin in water at 30 to 70 캜 and filtering to obtain a gelatin solution of 15 to 60%;
The gelatin solution was added to a stabilizer having a mass ratio of 1 to 2% and stirred for 15 seconds at a volume ratio of water to oil of 1/1 to 1/10 and 200 to 800 r / min for 15 seconds. Then, Adding a compound of an aldehyde group and solidifying the mixture for 1 to 2 hours, washing or centrifuging the cross-linking agent to obtain a gelatin foam;
A gelatin foam processing step in which a gelatin foam is added to a phosphate buffer solution or a phosphate buffer solution to maintain a pH of 7 or more, washed five times, and centrifugally dehydrated;
Gelatin foam was dissolved and spray dried to secure gelatin microspheres of 50 ~ 100um, 100 ~ 300um, 300 ~ 500um, 500 ~ 700um, 700 ~ 1000um and 1000 ~ 1500um, And an ion-binding induction step of diluting the sterilized water and adding the diluted sterilized water to the sterilized water. delete

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