TW201138758A - Method for dissolving hyaluronic acid and/or a salt thereof - Google Patents

Abstract

Disclosed is a method for dissolving hyaluronic acid and/or a salt thereof, in which a stirring tank provided with a stirring blade selected from a turbine-type serrated blade, a dispersion-type serrated blade, a dispersion-turbine-type serrated blade, an anchor-type serrated blade and a paddle-blade equipped serrated blade is used when dissolving hyaluronic acid and/or a salt thereof in an injectable solution selected from injection grade water, physiological saline, and phosphate buffered saline. The method for dissolving hyaluronic acid and/or a salt thereof enables large-scale production of injectable solutions of hyaluronic acid with a molecular weight of 1,500,000 to 4,000,000

Description

[Technical Field] The present invention relates to a hyaluronic acid and/or a salt thereof (hereinafter also collectively referred to as a uric acid) which is suitable for a pharmaceutical product from a liquid containing hyaluronic acid and/or a salt thereof. Note * The technique of ejaculation. - [Prior Art] Hyaluronic acid N-acetamidine-D-glucosamine and D_glucuronic acid-bonded disaccharide monoterpene are repeatedly linked to form a high molecular weight polysaccharide with a molecular weight of 50,000. Usually, the glucuronic acid is isolated and purified into sodium hyaluronate in the form of a sodium salt. It is known that sodium hyaluronate having a molecular weight of about 20,000 is more effective than the molecular weight of about 800,000 in the treatment of deformable knee joint disease, frozen shoulder, chronic rheumatoid arthritis, etc. Pharmacology and Treatment 'Vol.22, No.9, 289, (1994); Pharmacology and Treatment,

Vol.22, Νο·9, 319, (1994)). Further, it is known that it is an effect of preventing adhesion after surgery, and it is also effective as a pharmaceutical product in the field of dermatology and ophthalmology. The sodium hyaluronate produced by the microbial vinegar fermentation method is cultured using, for example, a certain species of Streptococcus, and the resulting culture solution is diluted and obtained in a powder form through various purification steps. * According to the microbial method, although sodium hyaluronate can be obtained in a high molecular weight form, there are various difficulties in the manufacture of a large amount of sodium hyaluronate injection. That is, it is difficult to efficiently dissolve the high molecular weight hyaluronic acid sodium in a short time, and since the viscosity of the solution is very high, it is difficult to handle and it is difficult to filter or sterilize due to the stability of heat or the like. Therefore, the method for mass-producing a high molecular weight sodium hyaluronate injection is still unclear. The inventors of the present invention conducted a repetitive study on a method for efficiently separating and removing impurities from a hyaluronic acid-containing solution to obtain a high-purity pharmaceutical grade hyaluronic acid when a large amount of a hyaluronic acid-containing solution is produced into an injection solution. It has been found that when hyaluronic acid is dissolved in an injection solution, hyaluronic acid does not sufficiently dissolve to cause agglomerates due to the use of a usual agitation tank under normal conditions, or causes a decrease in the molecular weight of hyaluronic acid when dissolved. The dissolution step becomes a bottleneck. SUMMARY OF THE INVENTION The present invention has been made in view of the above circumstances, and an object thereof is to provide a method for dissolving hyaluronic acid which can be preferably carried out in the case of generally producing an injection of hyaluronic acid. Further, an object of the present invention is to provide a method for dissolving hyaluronic acid which is capable of suppressing aggregation as much as possible and allowing a high molecular weight hyaluronic acid to be sufficiently dispersed in a solution for injection to obtain a clear solution. Further, an object of the present invention is to provide a method for dissolving hyaluronic acid which can dissolve a high molecular weight hyaluronic acid in a solution for injection without causing a decrease in the molecular weight as much as possible. According to a first aspect of the invention, the following method is provided. That is, (the method of dissolving hyaluronic acid and/or its salt, which uses a search tank having a selection/turning wheel type, a dispersion type, a dispersion type turbine type, an anchor type, and a half-wing with a paddle blade) It is selected from the group consisting of water for injection, physiological saline, and a solution for injecting 154809.doc 201138758, which dissolves hyaluronic acid and/or its salt; (2) dissolves hyaluronic acid and/or its salt as (1) The method wherein the shaft of the stirring wing is located at the center of the container or at an eccentric position; (3) The method for dissolving hyaluronic acid and/or a salt thereof as in (2), wherein the stirring blade is a one or more stirring tank (4) The method for dissolving hyaluronic acid and/or its salt according to (3), wherein a stirring tank having a stirring speed of 100 to 5000 rpm is used; (5) hyaluronic acid and/or its salt according to (4) a dissolution method, wherein the inner surface of the stirring tank and the pipeline is Teflon, a Teflon lining or a special fluorine coating; (6) the hyaluronic acid and/or any one of (1) to (5) a method for dissolving the salt thereof, wherein the average molecular weight of hyaluronic acid and/or its salt is from 150,000 to 4 million; (7) (1) The method for dissolving hyaluronic acid and/or a salt thereof according to any one of (6), wherein the hyaluronic acid and/or its salt is fermented with Streptococcus equi subsp. FM-100 or Streptococcus equi subsp. FM-3 00 ' According to this method, a high-quality hyaluronic acid solution which can be used in the production of an injection solution can be obtained. According to another aspect of the present invention, the following method is provided. That is, (8) a hyaluronic acid and/or Or a method for dissolving the salt thereof, which comprises using a stirring tank having a large-folded full-wheel type wing or a stirring wing of a dispersing turbine wing, in an injection solution selected from the group consisting of water for injection, physiological saline and buffered saline. Dissolving hyaluronic acid and/or its salt; 154809.doc 201138758 (9) The method for dissolving hyaluronic acid and/or its salt according to (8), wherein the stirring tank is substantially vertical cylindrical, and the stirring wing system is arranged to position the shaft (10) The method for dissolving hyaluronic acid and/or its salt according to (9), wherein the eccentric position of the shaft of the stirring wing divides the center line into a position of 1:2. (11) The hyaluronic acid of any one of (8) to (10) and/or The method for dissolving a salt, wherein the agitation tank is provided with a stirring blade having a ratio of agitating wing diameter/slot inner diameter of 〇3 to 〇5; (12) a dissolution method according to any one of (8) to (11), wherein (13) The method of dissolving any one of (8) to (12), wherein the stirring time is set to 45 minutes or more; (14) as in (8) to (13) The method according to any one of (13) wherein at least the inner surface of the agitation tank and the line connected to the search tank is made of stainless steel and subjected to electrolytic grinding; (15) as in (8) to (14) A method of dissolving, wherein the average molecular weight of hyaluronic acid and/or its salt is from 150,000 to 4 million; (16) The method of dissolving according to any one of (8) to (1), wherein hyaluronic acid and/or The dissolution method of any one of (8) to (16), wherein the hyaluronic acid and/or its salt system uses Streptococcus equi subsp. 100 or E. coli FM-300, manufactured by fermentation. According to this method, the high molecular weight hyaluronic acid can be sufficiently dispersed in the injectable solution while suppressing the aggregation or the decrease in the molecular weight as much as possible, and the injection can be produced on a large scale by 154809.doc 201138758. [Embodiment] Hereinafter, an aspect in which the present invention is carried out by Hx will be described. The hyaluronic acid used in the sputum includes a mixture of free form of uric acid, a salt of hyaluronic acid, or a salt of free hyaluronic acid and hyaluronic acid. Examples of the hyaluronic acid salt include a sodium salt, a potassium salt H chain salt, and the like. Further, the hyaluronic acid-containing liquid to be used in the present invention may be either an animal extractor or a manufacturer by a method, and it is preferred to use a yeast fermentation method. The hyaluronic acid by the acid fermentation method can be obtained by a known method using microorganisms such as bacteria having a hyaluronic acid-producing ability such as Streptococcus. Examples of the strains used in the brewing method include microorganisms which have been isolated from nature and which have the ability to produce this force, or Streptococcus equi subsp. FM 100 as described in Japanese Patent Laid-Open Publication No. The mutant of hyaluronic acid is stably produced in a high yield in a high yield, and it is preferred to use the mutant strain, which is described in Japanese Patent Application Laid-Open No. Hei No. No. No. 2,234,689. The hyaluronic acid-based high molecular weight solid (powder) which can be used in the present invention is produced by the above-mentioned method, and usually has an average molecular weight of i to 10,000. When the average molecular weight is less than 15 G, the effect as a pharmaceutical product is lowered, and on the other hand, it is difficult to obtain an average molecular weight of more than 40,000 by the above method. However, the dissolution method of the present invention can be used to dissolve low molecular weight hyaluronic acid. The solution for injection can be used as a step for dissolving hyaluronic acid. 154809.doc 201138758 Water for injection, physiological saline and buffered saline are used, and in particular, a buffer containing an acid, a base or a phosphate may be added. ? 11 Regulators, etc. are identified in the Pharmacopoeia of the Pharmacopoeia. The amount of the hyaluronic acid added in the step of the gluten is set so that the hyaluronic acid concentration is 0.75 to 1.25 w/v%. If the hyaluronic acid concentration is 0.75 w/v% or less, the hyaluronic acid solution has a low viscosity and is easy to manufacture. Also if it is 1_25 w/v. /. Above, it is difficult to prepare in large quantities due to the solubility of hyaluronic acid. Therefore, the hyaluronic acid concentration of the solution having a high viscosity is 0.75 to 1.25 w/v%, which is in accordance with the production conditions for the dissolution method of the present invention. Preferably, the valved airtight container is prefilled with dissolved hyaluronic acid and is supplied to the agitation tank through a feed passage through the valve. Preferably, the angle of the feed flow path is set to 50. The steep slope above is turned upside down, and when hyaluronic acid is introduced, the loss is reduced. The valve preferably uses a butterfly valve which does not cause the hyaluronic acid to contact the outside air by its conversion, and can be aseptically placed in the agitation tank. Further, as for the material of the valve-tight airtight container, it is preferable that the stainless steel or the inner surface thereof is coated with Teflon, or the inner surface is electrolytically ground, in terms of cleaning property, ease of handling, and the like. Processed by. The stirring tank for dissolving is formed by attaching a vertical axis type stirring device to a generally vertical cylindrical tank body, and the stirring device is provided with an axis that is disposed in the vertical direction of the groove body. The agitating wing is mounted substantially horizontally on the lower end of the shaft (and optionally in the middle portion), and a driving device mounted on the upper end of the shaft and disposed on the top cover portion of the trough body. 154809.doc 201138758: In the stirring and dissolving step in the stirring tank, since the solubility of hyaluronic acid in the solution for injection is poor, the solution is high in viscosity, and in most cases, the molecular weight is lowered in the dissolved material, there is In order to solve the ', * problem, the present inventors conducted a comparatively comparative study using various types of mixers in the above-mentioned agitation tank under various conditions. As a result, it was found that a specific type of mixer was preferably produced under specific conditions. That is, as the infusion device, a shattering machine having a wing type selected from the group consisting of a turbine type, a dispersion type, a dispersion type turbine type, a wrong type, and a slab toothed wing can be used. Comparative studies show that it is preferred to have a turbine type, a dispersion type, and a dispersion type turbine wing, and particularly preferably a large-depressed thirteen-wheel type wing and a dispersion turbine type wing. Moreover, it is understood that the position at which the agitating blade is disposed (the position of the shaft of the agitating blade) can be disposed substantially at the center of the groove body, but if the axial direction of the axial groove body of the agitating blade is eccentric outward In the case of setting, the dissolution rate of hyaluronic acid is accelerated, so it is preferable. For example, it can be seen that the eccentric position can be set to divide the groove diameter on the center line of the groove body into a position of 丨: 2, a position of 1:3, a position of 丨: 4, and a position of 丨: 5, etc. Among them, it is preferably divided into 1: 2 positions. Further, it is understood that the rotation speed of the stirring blade is usually 1 〇〇 5 5 rpm, for example, 800 to 2000 rpm, particularly for the larger large-wound full-wheel wing and the dispersed turbine wing in the stirring blade. In other words, it is preferably 15 rpm to 18 rpm. If the rotation speed is too small, the hyaluronic acid is poor in dispersibility in the solution for injection. On the other hand, even if you want to increase the rotation speed, the molecular weight of hyaluronic acid is too high, so that the stirring wing does not rotate v. In the case of dissolution, most cases 154809.doc 201138758 = effective is to warm while mixing, but The hyaluronic acid type 2 causes an abnormal physical property change such as a decrease in molecular weight due to heating. (4) It is not necessary to (4) It is considered that if the extension is granted - if the stirring time is prolonged, physical properties such as molecular weight decrease, etc., and the shape of the stirring blade, position, other operating conditions, etc. can be obtained. The speed is set to the above range, and the solution is dissolved in a short time under mild conditions without heating. Further, it was found that the size of the 'fourth wing' also has the best range. That is, the ratio of the throwing wing diameter (d)/slot inner diameter (D) is preferably set to 0.3 to 〇.5. The reason is that if the ratio is smaller than 〇_3, the mixing effect is insufficient. When the solubility and the squeegee are deteriorated, the other side is more than Q 5, and an abnormality such as a decrease in molecular weight is generated. Also, the same mixing time is short, (4) the mixing effect is insufficient, and the solubility: the dispersion is deteriorated. On the other hand, if it is too long, an abnormality such as a decrease in molecular weight may occur. The example is preferably from 45 minutes or more to about 100 minutes, and particularly preferably to about 60 minutes. Further, as described above, the stirring blade is appropriately designed. The type, the large J ° position, the rotation speed, etc., one section is sufficient, but it is not excluded to be set into a plurality of sections. In the dissolution operation, it is preferable to appropriately decompress the inside of the stirring tank, which is to remove hyaluronic acid and The bubble in the liquid is also effective for accelerating the dissolution rate. The hyaluronic acid solution is highly viscous, but for defoaming, it is preferred to use a vacuum decompression mechanism such as a vacuum fruit to reduce the pressure to 5 to 2 〇kpa abs. The temperature, or the simultaneous stirring of the solution, is more effective. 09.doc •10· 201138758 'The material of the inner surface of the mixing tank is the cleaning property of the inner surface after the combination. 'The corrosion resistance of the salt water, the solubility of β, etc. ), etc., the stainless steel, glass, and Teflon surface are stretched out and the uric acid solution is adhered to the surface of the material... 'It is preferably a special gas (registered trademark), Terry or Teflon (registered trademark) Coating [: 庐Μ 庐Μ 原因 : 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特 特Alternatively, it is also preferable to use stainless steel, in particular, § Cong said to replace the special (registered trademark). In this case, if the inner surface is subjected to electrolytic polishing, the performance equivalent to that of Teflon (registered trademark) can be obtained. After stirring and dissolving, 'the steps of sterilization, impurity filtration, defoaming, and filling are performed. If the steps are simply described, the sterilization of the hyaluronic acid solution is before the impurities are removed, or after filling in a small glass bottle or the like. And proceed. By filtering The filtration membrane used for filtration preferably has a pore diameter of 0.2 to 50 μm. When the pore diameter is smaller than the range, the sterilizing liquid obtained in the previous step is a liquid having a very high viscosity, so that it is difficult to pass the sterilizing liquid through the membrane. When the pore diameter is larger than the range, the impurity filtration may be incomplete, and the insoluble impurities which are visually discernible are mixed in the injection liquid, which is not preferable. The material of the ruthenium film can be self-assembled from polytetrafluoroethylene or polyester. , selected from Teflon (registered trademark), polypropylene, polyvinylidene fluoride and nylon, preferably polyvinylidene fluoride, polypropylene or nylon. As the shape of the filter membrane, it can be a flat membrane or a filter. Any one of the cartridge and the disposable filter is preferably a filter or a disposable filter in the case of a large amount of processing 154809.doc 201138758. As a specific example of the filter film which can be used in the present invention, it is manufactured by MiUip〇re.

Millipak or Durapore cartridge filters (Millidisk) and so on. The pH of the hyaluronic acid-containing solution is 2 to 1 Torr, and the temperature is 5 to i 〇〇 °c. The flow rate and pressure at the time of liquid permeation are set in consideration of the pressure resistance depending on the type of the filter. If pressure is applied, impurities will flow out from the filter, so care must be taken. In the cartridge filter 4, the flow rate is preferably 50 to 300 L/hr, and the treatment pressure is 0·01 to 0 5 Mpa. The concentration can also be adjusted by diluting the filtrate with a solution for injection. A filling machine is used in the filling step, and the filling machine includes a portion in which the hyaluronic acid solution is filled in the container, and a sealed portion is inserted into the filled container or the cereal is melted. As the container for the injectable solution to be filled, a usual ampule, a small glass bottle, a 歹2 7 7 1-3 type or prefilled syringes can be used. Hereinafter, the present invention will be described in detail by way of examples, but the invention is not limited to the description of the examples. [Examples] &lt;Example 1&gt; A sodium hyaluronate 1580 g having a molecular weight of 2.37 million obtained by a fermentation method was filled with 20 L with Streptococcus equi subsp. FM-100 (Microtechnology Research No. 9027). The butterfly valve is in an airtight container. A dispersing turbine type agitating blade was installed in a Teflon-coated stainless steel tank with a capacity of 200 L. The physiological saline (injection solution) containing 7.3 mM sodium phosphate buffer (pH 7.3) was 149 L. Put in the agitation tank. 154809.doc •12· 201138758 The airtight container filled with the above sodium hyaluronate is placed upside down on the raw material powder inlet of the feeding tank, the butterfly valve is opened, and the sodium hyaluronate is put into the stirring tank. 1 to 2 are plan views showing a dispersion turbine type agitating blade (dispersing agitating blade diameter 275 mm' including 12 blades and 6 blades of a folding blade blade). Stir at 1800 rpm for 50 minutes to completely dissolve the sodium hyaluronic acid. In order to remove the bubbles in the liquid, the pressure in the stirring tank was maintained at a vacuum of 15 kpa abs for 20 minutes, and after the bubbles were removed, the pressure was returned to normal pressure. The sodium hyaluronate concentration of the solution was measured by the carbazole sulfuric acid method, and the result was hydrazine. /(^ If the ultimate viscosity of the liquid is determined according to the fifteenth revision of the Japanese Pharmacopoeia, it is 33 8 dL/g, which is converted into a molecular weight of 2.37 million. The solution is continuously made by the KIDKOMAN company's kids c〇〇ker The sterilizer is continuously sterilized. The device consists of a double tube with an inner diameter of 23 mm 'with a built-in fixed mixer'. The volume of the heating section is 34 l, the volume of the valley is 0 · 6 L, and the volume of the cooling section is 2.6 L. The hot water of the outer tube of the heating unit was adjusted so that the temperature of the holding portion became 13 5 C, so that the stagnation time of the holding portion was 34 seconds, and the amount of pressure at the inlet of the heating unit was controlled. The outlet temperature is 40t or less, and the water in the outer tube of the cooling unit is adjusted. The pressure at the outlet of the cooling unit is 0.33 MPa, and the pressure regulating valve is used for control, and a filter film made of polyethylene dioxide having a pore diameter of 5 μm is used. After that, the cartridge filter manufactured by the company Miebopia was filtered, and the cooled sodium hyaluronate solution was filtered at a flow rate of 60 L/hr. The filtrate was stirred and mixed at 144 rpm for 30 minutes. Then, by including the separator type Filling pump filling part 'rubber stopper plug, rolling mechanism small 154809.doc -13- 201138758 glass bottle filling and sealing machine' fill this liquid in a small glass n oak county plug On the butyl rubber (made by Daxie Seiko Co., Ltd.). According to the fifteenth revision of the Pharmacopoeia, the general provisions of the preparation, the project of the injection, the quality test of the product was carried out to detect insoluble impurities, and the qualified rate was 99.9% or more. Use the dispersing turbine type agitating blades (wing diameter 243 _, including 12 blades and wing diameter 17 、, containing 4 blades) as shown in Figures 3 to 4 instead of the dispersing thirsty wheel type mixing shown in Figure 1-2. The operation was carried out under the same conditions, and the results were confirmed to be equally effective. <Example 2> In order to carry out (four) changes to the configuration of the (four) machine in the mixing tank, the best search wings and stirring bars #, and Dissolution test. In the dissolution test, 1280 g of sodium hyaluronic acid (limit viscosity 35.0 dL/g) was placed in a stirred tank of 149 L of the inner diameter of the physiological saline solution, and the mixer was operated to dissolve it. Based on the following benchmark pairs Evaluation of the dispersibility of uric acid sodium, solubility, and molecular weight reduction [Dispersibility] The behavior of hyaluronic acid was visually confirmed and evaluated according to the following criteria: 0: Hyaluronic acid dispersed in the entire tank △: Hyaluronic acid dispersion In the vicinity of the bottom of the tank and not dispersed in the entire tank X: Most of the hyaluronic acid is present at the bottom of the tank, and almost no dispersion is found. 154809.doc -14- 201138758 [Can be dissolved] The determination criterion of the following is a e, if all the criteria are satisfied, it is set to 〇 'If - one is not satisfied, it is set to Δ, and the case where all are not satisfied is set to χ. a: In the dissolution of hyaluronic acid, no condensate was found by visual standard (face 疙瘩) b: hyaluronic acid lysate was colorless and transparent c: no undissolved hyaluronic acid was found on the wall surface of the stirring tank by visual standard [molecular weight reduction Degree (Δ molecular weight)] The Δ molecular weight (= (pre-dissolution limit viscosity - post-dissolution limit viscosity) / pre-dissolution limit viscosity) was determined according to the following criteria. 〇: △ molecular weight $ 〇.〇3 △ : 〇.〇3&lt;Δ molecular weight $0.15 χ : 〇.15&lt;Δ molecular weight [test on the shape of the stirring blade] Preparation of large-blade turbine wing, two-stage folding blade turbine type The wing and the folded blade turbine wing and the paddle are used together, the dispersing turbine wing, the large agitating wing, and the dispersing agitating blade are used as the agitating blades, and are installed in the agitation tank, and the dissolution test is performed at a stirring speed of 18 rpm. The results are shown in Table 1 below. Here, the shape of each of the above-mentioned search wings used in the present embodiment and the mounting position in the stirring tank are shown in Figs. 5 to 10. Fig. 5 is a view showing a state in which a large-blade turbine-shaped wing is disposed in the scramble tank 4, (a) is a schematic side cross-sectional view, and (b) is a view of a bb arrow in the system (a), in which H = 754 mm, h=522 mm, D=550 mm, d=180

154809.doc S 201138758 mm (d/D=G·33). Further, in the test regarding the shape of the (four) wing, the large-depressed thirteen-wheel type wing 5 was attached to the eccentric position (four) mm of the stirring tank 4. This eccentric position corresponds to the eccentricity (1: 2) described later. Fig. 6 is a diagram showing a state in which a section of the turbine blade 6 is disposed in the agitation vessel 4 ((4) is a schematic side sectional view, (b) is a b_b arrow view in the system (4), and (4) is a cc arrow in the system (4) View, in the figure, H = 754 _, hl = 29 〇, 32 mm D - 550 mm, di (top) = 155 mm, ten (lower) mm (dl / D = 〇. 28 (top); d2 / D=〇33 (bottom)). In this test, the two-stage vane turbine wing 6 was also mounted at an eccentric position (e = 9 〇 mm). Fig. 7 is a view showing a state in which the hinged blade-shaped blade and the paddle blade 7 are disposed in the mixing tank 4, (a) is a schematic side sectional view, a b_b arrow view, and (c) is a system (4). C_c arrow view, in the figure, h=754 claws, = mm, h2 = 85 mm (paddle height), h3 = 154 mm, D=550 mm, d, (paddle position) 155 mm, d2 (turbine wing diameter) = 18 〇 mm (d丨 / D = 〇 28 (paddle); cb / D-0.33 (turbine wing)). In this test, the combined wings 7 were also mounted in an eccentric position (e = 90 mm). Fig. 8 is a view showing a state in which the dispersing turbine-shaped blade 8 is disposed in the agitation tank * (a) is a schematic side cross-sectional view, and (13) is a b-b arrow view in the hook, in the figure, H = 754 mm, h = 522 mm, D = 550 mm, d = 180 mm (d/D = 0.33). In this test, the dispersing turbine wing 8 was also mounted at an eccentric position (e = 90 mm). Figure 9 shows that The large agitating blade 9 is arranged in the state of the agitation tank*, (4) is a schematic side sectional view, and (b) is a view of the bb arrow in the system (4). In the figure, Η 677 mm 'h (wing south) = 450 mm, D = 550 mm, d = 290 mm 154809.doc • 16 - 201138758 (d/D = 0.53) Fig. 10 is a view showing a state in which the dispersion agitation blade 10 is disposed in the agitation vessel 4, and (a) is a schematic side section. Figure, (b) is a plan view of the dispersing agitating blade 10, in the figure, H = 677 mm, e = 137.5 mm, D = 550 mm, d = 250 mm (d / D = 0.53) o 154809.doc • 17- 201138758

[II

Homogenizer eccentricity 1800 0.45 &lt;1 X -9- Large mixing wing 1800 0.33 〇X i Dispersing turbine wing 1_.____________ Eccentricity (1 : 2) 1800 0.33 〇〇7 paddle (top) + folding blade turbine wing (lower Eccentricity (1 · 2) 1800 0.33 (upper), 0.28 (bottom) XX wins two-stage folding blade turbine wing eccentricity (1 : 2) 1800 0.33 &lt; plucking large hinged turbine wing 1 eccentric (1 : 2 ) 1800 0.33 〇〇7 Stirring wing shape Stirring wing position mixing speed [rpm] 搅拌 Stirring time [min] HA dispersibility dissolves molecular weight reduction 綦韹唼镅: VH 154809.doc -18- s 201138758 According to the results of Table 1 The large 苇 苇 , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , On the other hand, the second-stage folding field and the blade-winged wing are poor in the dispersibility of sodium hyaluronate, and the molecular weight is also reduced. The remaining folding leaves and the still-wing type of money are also used for dispersion or In the case of dissolution, the decrease in molecular weight is also seen in the case of 'good', especially in the large-scale search wings. [Position of the agitating wing] The agitating blade is attached to the eccentric work on the large-blade turbine wing and the dispersing turbine wing which are excellent in the results of the dissolution test in terms of the shape of the agitating blade: 2. Eccentricity 1: 3, eccentricity丨: Position 5 and investigate the effect of the position of the agitating wing during dissolution. The results are shown in Table 2 below. 154809.doc 19- 201138758 [(N&lt;] Dispersing turbine wing eccentricity (1: 5) 1800 0.33 § XX Dispersing turbine wing eccentricity (1: 3) 1800 0.33 &lt; 〇 分散 dispersion turbine wing eccentricity (1: 2 1800 0.33 折 large hinged turbine wing eccentric (1: 5) 1800 0.33 § 1-HXX Seven large hinged turbine wing eccentric (1: 3) 1800 0.33 宕Η &lt; X large hinged turbine wing eccentric (1 : 2) 1800 0.33 〇〇•δ- Stirring wing shape Stirring wing position mixing speed [rpm] Stirring wing diameter/slot inner diameter stirring time [min] HA dispersibility dissolves molecular weight decrease

綦 碜 碜 :: VH .20· 154809.doc s 201138758 According to the results of Table 2, in the case of either the large folding _ wheel wing and the dispersed wheel wheel type, the position of the eccentric 1: 2 The best installation location for the wing. [Stirring speed] The dispersing turbine wing that has been excellent in the test so far is installed in the eccentric 丨: 2 position with excellent performance, and the stirring speed is varied in the range of 1 〇〇〇卬 m to 2500 rm. The dissolution test was carried out. The results are shown in Table 3 below. [table 3]

According to the results of Table 3, 15 〇〇卬 (7) and a (8) rpm exhibited the same performance as the stirring rotation speed. However, considering the shorter dissolution time, 1800 rpm was the most excellent. [Agitating wing diameter/slot inner diameter] The size of the dispersion turbine wing which is excellent in the test so far is changed within a range of 0.H in the ratio of the agitating blade diameter/slot inner diameter, and the eccentricity 1 is excellent in performance. : 2 position, stirring speed 1800 rpm, the dissolution test I54809.doc 201138758 test. The results are shown in Table 4 below. [Table 4] Agitated wing shape dispersing turbine wing dispersing turbine wing dispersing turbine wing dispersing turbine wing agitating wing position eccentricity (1 : 2) Eccentricity (1 : 2) Eccentricity (1 * 2) Eccentricity (1 : 2 Stirring speed [rpm] 1800 1800 1800 1800 Stirring wing diameter/slot inner diameter 0.1 0.3 0.5 1 Stirring time [min] 120 60 60 60 HA dispersibility 〇〇〇〇 Dissolving Δ 〇〇〇 Molecular weight reduction small and small As can be seen from Table 4, the ratio of the stirring wing diameter/slot inner diameter was 0.3 and the same performance was exhibited by 0.5, but the ratio was 〇. The solubility was poor, and the ratio was one, and the molecular weight decreased greatly. [Stirring time] The dispersing turbine wing with a ratio of the agitating wing diameter/slot inner diameter of 0.33 is attached to the position of the eccentricity 1: 2, and the mixing time is changed within the range of 30 minutes to 120 minutes, and the dissolution test is performed. . The results are shown in Table 5 below. [Table 5] Agitated wing shape dispersing turbine wing dispersing turbine wing dispersing turbine wing dispersing turbine wing agitating wing position eccentricity (1 : 2) Eccentricity (1 : 2) Eccentricity (1 : 2) Eccentricity (1 : 2 Stirring speed [rpm] 1800 1800 1800 1800 Stirring wing diameter / groove inner diameter 0.33 0.33 0.33 0.33 Stirring time [min] 30 45 60 120 HA dispersibility 〇〇〇〇 Dissolving X 〇〇〇 Molecular weight reduction small and small 154809.doc •22· 201138758 According to Table 5, if the stirring time is 30 minutes, the solubility is insufficient. On the other hand, if the stirring time is 12 G minutes, the molecular weight is significantly lowered. In contrast to this, the application time is 45 minutes or 6 minutes, and all the bases are satisfied. [Simple Description of the Drawings] FIG. 1 is a plucking tank for performing the dissolution method of the present invention. A schematic plan view of the dispersed saddle (four) mixing wings. Figure 2 is a side elevational view of the dispersing turbine type agitating wing of Figure 1. Fig. 3 is a view similar to Fig. 1 showing an example of another dispersion type turbine agitating blade. Figure 4 is a side elevational view of the dispersing turbine type agitating wing of Figure 3. Fig. 5 is a schematic side sectional view showing a state in which a large-blade turbine-shaped wing is disposed in a stirring tank, and (b) is a view of a b-b arrow in (a). Fig. 6 is a cross-sectional view showing a state in which a two-stage folding blade type turbine blade is disposed in a stirring tank, (b) is a schematic side sectional view in (a), and (c) is a (b) system (a) The view of the ce arrow. Fig. 7 is a diagram showing the arrangement of the wing-shaped turbine wing and the paddle plate in the mixing tank, (a) is a schematic side sectional view, and (b) is a view of the bb arrow in the system (a). '(c) is the view of the cc arrow in (a). Fig. 8 is a view showing a state in which a dispersing turbine wing is disposed in a stirring tank, (a) is a schematic side sectional view, and (b) is a b-b arrow view in (a). Fig. 9 is a view showing a state in which a large agitating blade is disposed in a stirring tank, (a schematic side cross-sectional view of the 昀 system, and (b) a bb arrow view in (a). Fig. 10 shows a dispersing agitating blade. Diagram of the state of the agitation tank, (a) 154809.doc -23- 201138758 Surface diagram. It is a schematic side sectional view, (b) is a distributed mixing wing flat [Main component symbol description] 1, 10 Dispersing agitating wing 2 axis Hole 3 Folding blade 4 Stirring tank 5 Large vane turbine wing 6 Two-stage vane turbine wing 7 Folding turbine wing and paddle 8 Dispersing turbine wing 9 Large agitating wing b, c Arrow direction D Groove inner diameter d agitating wing diameter di paddle wheel thirteen wheel diameter e eccentric position h wing height h, ' h3 ' H 尚度 h2 paddle height with wings 154809.doc •24·

Claims (1)

201138758 VII. Patent application scope: 1 · A method for dissolving hyaluronic acid and/or its salt, which is a stirring tank equipped with a large-blade turbine wing or a dispersing turbine wing, selected from water for injection and saline And a buffer solution for injecting physiological saline, which dissolves hyaluronic acid and/or a salt thereof. 2. In the case of the method for dissolving hyaluronic acid and/or its salt, wherein the trough is a substantially vertical cylindrical shape, and the whipwing system is arranged to position the shaft at a position radially outward from the center of the groove. on. 3. The method for dissolving hyaluronic acid and/or a salt thereof according to claim 2, wherein the eccentric position of the shaft of the agitated wing is a position where the center line is divided into 1:2. 4. The method for dissolving hyaluronic acid and/or a salt thereof according to any one of claims 1 to 3, wherein the agitation tank has a ratio of agitating wing diameter/slot inner diameter of 〇.3 to 〇5. A method of dissolving any one of items 1 to 4, wherein the mixing wing is operated at a rotational speed of 1500 to 1800 rpm. 6. The dissolution method according to any one of items 1 to 5, wherein the stirring time is set to 45 minutes or longer. 7. The dissolution method according to any one of claims 1 to 6, wherein at least the inner surface of the agitation vessel and the line connected to the agitation vessel is made of stainless steel and electrolytically polished. The dissolution method according to any one of claims 7 to 7, wherein the average molecular weight of hyaluronic acid and/or its salt is from 1.5 to 4 million. The method of dissolving according to any one of claims 1 to 8, wherein the dissolution is carried out in such a manner that the concentration of hyaluronic acid and/or the salt thereof becomes 〇75 to 1.25 w/v%. The method of dissolving according to any one of claims 1 to 9, wherein the hyaluronic acid and/or its salt is produced by a fermentation method using Streptococcus equi subsp. FM-100 or Streptococcus equi subsp. FM-300. to make. 154809.doc