Extraction method and content detection method of polyvinyl alcohol in polyester particles

Abstract

The invention discloses an extraction method and a content detection method of polyvinyl alcohol in polyester particles. The method for extracting the polyvinyl alcohol in the polyester particles comprises the following steps: (1) dissolving the polyester particles in an organic solvent, mixing with water, extracting, centrifuging, and collecting a water phase and an organic phase; the volume ratio of the organic solvent to the water is (1.8-10): 1; (2) mixing the organic phase with water, extracting and centrifuging, and collecting a water phase; the volume ratio of the organic phase to the water is (1.8-10): 1; and (3) mixing the water phases collected in the steps. And repeating the operation for more than two times in the step (2). The polyvinyl alcohol extraction method can ensure that the recovery rate reaches 85% or above, and the polyvinyl alcohol residue detection method is higher in accuracy and simple in process, and the detection cost is reduced.

Description

Technical Field

[0001] The invention relates to the field of quantitative detection of polyvinyl alcohol, and in particular to a method for extracting polyvinyl alcohol in polyester particles and a method for detecting its content. Background Art

[0002] Polyester particles are common injectable filling products in the medical beauty field. For example, polycaprolactone particles or polylactic acid particles. When these materials degrade in the body, they can stimulate an immune response in tissues, promote fibroblast proliferation and myofibroblast differentiation, and then synthesize a large amount of collagen, thereby restoring subcutaneous volume and repairing wrinkles.

[0003] At present, the emulsified solvent volatilization method is often used to prepare microparticles. Specifically, the polyester material is dissolved in an organic solvent to obtain an organic phase, and the organic phase is added dropwise to an aqueous phase containing polyvinyl alcohol, and then the organic solvent is removed and solidified to obtain polyester microparticles. There will be trace amounts of polyvinyl alcohol residues in the polyester microparticles obtained by this method. Polyvinyl alcohol cannot be degraded in the body, and will accumulate for a long time after local injection, and is not easy to be excreted from the body. Therefore, it is necessary to strictly control the amount of polyvinyl alcohol residues in polyester microparticles. It has been reported that the content of polyvinyl alcohol in polyester microparticles is tested by ultraviolet-visible spectrophotometry. Specifically, polyvinyl alcohol is first separated from polyester microparticles to reduce the influence of materials such as polyester in the sample to be tested on the accuracy of the test results. The extracted polyvinyl alcohol sample is colored with an iodine reagent in a boric acid medium, and then the absorbance value of the test system is tested to calculate the amount of polyvinyl alcohol residues. However, the separation effect of the existing polyvinyl alcohol separation method is not ideal, and the extraction rate of polyvinyl alcohol is low, resulting in inaccurate test results.

[0004] Therefore, there is an urgent need in the art to develop a method that can accurately test the amount of polyvinyl alcohol residue in polyester microparticles. Summary of the invention

[0005] The technical problem to be solved by the present application is to overcome the defects of inaccurate detection results when detecting the residual polyvinyl alcohol content in polyester microparticles in the prior art, and to provide a method for extracting polyvinyl alcohol in polyester microparticles and a method for detecting the content. The polyvinyl alcohol extraction method of the present application can ensure a recovery rate of more than 85%, and the polyvinyl alcohol residual detection method has higher accuracy and a simple process.

[0006] This application solves the above technical problems through the following technical solutions.

[0007] The present application provides a method for extracting polyvinyl alcohol from polyester microparticles, comprising the following steps:

[0008] (1) dissolving polyester microparticles in an organic solvent, mixing with water, extracting, centrifuging, and collecting an aqueous phase and an organic phase; the volume ratio of the organic solvent to the water is (1.8-10):1;

[0009] (2) taking the organic phase, mixing it with water, extracting it, centrifuging it, and collecting the aqueous phase; the volume ratio of the organic phase to the water is (1.8-10):1;

[0010] (3) Mix the water phases collected in the above steps;

[0011] Repeat step (2) two or more times.

[0012] In step (1) of some embodiments, the mass percentage of the polyvinyl alcohol in the polyester particles is less than 0.5%.

[0013] In step (1) of some embodiments, the polyester is selected from at least one of polyglycolide, polylactic acid, polycaprolactone, polyhydroxyalkanoate, glycolide-lactide copolymer, caprolactone-lactide copolymer, polycaprolactone-polyethylene glycol copolymer and polylactic acid-polyethylene glycol copolymer. Wherein, the polylactic acid-polyethylene glycol copolymer is selected from polylactic acid-polyethylene glycol-polylactic acid triblock copolymer and / or polylactic acid-polyethylene glycol diblock copolymer. The number average molecular weight of the polylactic acid-polyethylene glycol-polylactic acid triblock copolymer is 40,000 to 130,000, wherein the number average molecular weight of PEG is 100 to 800.

[0014] In step (1) of some embodiments, the preparation method of the polyester microparticles is selected from at least one of a solvent volatilization method, a phase separation method, a spray drying method, a surface deposition method and a grinding method.

[0015] In step (1) of some embodiments, the dissolution can be performed under ultrasonic conditions according to conventional techniques in the art.

[0016] In step (1) of some embodiments, the mass volume ratio of the polyester particles to the organic solvent is 0.005 to 0.5 g / mL, for example, 0.005 g / mL, 0.01 g / mL, 0.015 g / mL, 0.02 g / mL, 0.025 g / mL, 0.03 g / mL, 0.035 g / mL, 0.0375 g / mL, 0.04 g / mL, 0.045 g / mL, 0.05 g / mL, 0.0 55g / mL, 0.06g / mL, 0.065g / mL, 0.07g / mL, 0.075g / mL, 0.08g / mL, 0.085g / mL, 0.09g / mL, 0.095g / mL, 0.1g / mL, 0.15g / mL, 0.2g / mL, 0.25g / mL, 0.3g / mL, 0.35g / mL, 0.4g / mL, 0.45g / mL or 0.5g / mL.

[0017] In step (1) of some embodiments, the organic solvent is a halogenated alkane solvent, preferably a C2-C6 halogenated alkane, such as dichloromethane.

[0018] In step (1) of some embodiments, the volume ratio of the organic solvent to the water is (1.8-5):1, for example, 2:1, 2.5:1, 3:1, 3.5:1, 4:1, 4.5:1, 5:1, 5.5:1, 6:1, 6.5:1, 7:1, 7.5:1, 8:1, 8.5:1, 9:1, 9.5:1 or 10:1.

[0019] In step (1) and step (2) of some embodiments, the centrifugal speed is independently 800-30000 rpm, preferably 1000-15000 rpm, for example 1000 rpm, 2000 rpm, 3000 rpm, 4000 rpm, 5000 rpm, 6000 rpm, 7000 rpm, 8000 rpm, 9000 rpm, 10000 rpm, 11000 rpm, 12000 rpm , 13000rpm, 14000rpm, 15000rpm, 16000rpm, 17000rpm, 18000rpm, 19000rpm, 20000rpm, 21000rpm, 22000rpm, 23000rpm, 24000rpm, 25000rpm, 26000rpm, 27000rpm, 28000rpm, 29000rpm or 30000rpm.

[0020] In steps (1) and (2) of some embodiments, the centrifugation time is independently 1 to 15 min, preferably 1 to 10 min, for example 1 min, 2 min, 3 min, 4 min, 5 min, 6 min, 7 min, 8 min, 9 min, 10 min, 11 min, 12 min, 13 min, 14 min or 15 min.

[0021] In step (2) of some embodiments, the volume ratio of the organic phase to the water is (1.8-5):1, for example, 2:1, 2.5:1, 3:1, 3.5:1, 4:1, 4.5:1, 5:1, 5.5:1, 6:1, 6.5:1, 7:1, 7.5:1, 8:1, 8.5:1, 9:1, 9.5:1 or 10:1.

[0022] In some embodiments, step (2) is repeated 2 to 5 times.

[0023] The present application also provides a method for quantitatively detecting polyvinyl alcohol in polyester microparticles, comprising the following steps:

[0024] (1) obtaining an aqueous phase by the method for extracting polyvinyl alcohol from polyester microparticles as described above, heating the aqueous phase, and fixing the aqueous phase to obtain a solution A;

[0025] (2) taking part of the solution A and mixing it with a boric acid solution, and then mixing it with an iodine-potassium iodide reagent to obtain a solution B;

[0026] (3) The absorbance value of the solution B is measured by ultraviolet-visible spectrophotometry, and the content of the polyvinyl alcohol is calculated by comparing with a standard curve.

[0027] In step (1), the heating may be a boiling water bath heating conventionally used in the art.

[0028] In step (1), the heating time is 5 to 40 min, for example, 5 min, 10 min, 15 min, 20 min, 25 min, 30 min, 35 min or 40 min.

[0029] In step (1), after the volume is fixed, the concentration of the polyester microparticles is 2 to 30 mg / mL, preferably 4 to 16 mg / mL, for example 3 mg / mL, 5 mg / mL, 7 mg / mL, 9 mg / mL, 11 mg / mL, 13 mg / mL, 15 mg / mL, 17 mg / mL, 19 mg / mL, 21 mg / mL, 23 mg / mL, 25 mg / mL, 27 mg / mL, 29 mg / mL or 30 mg / mL, based on the mass of the polyester microparticles in the aqueous phase per unit volume.

[0030] In the boric acid solution of step (2), the mass volume ratio of boric acid to solvent is 0.015 to 0.05 g / mL, for example 0.03.1 g / mL.

[0031] In step (2), the volume ratio of the solution A to the boric acid solution is 1:(0.5-3), for example 1:1.5.

[0032] In step (2), the concentration of potassium iodide is 0.01 to 0.05 g / mL, for example 0.025 g / mL, based on the mass of potassium iodide per unit volume of iodine-potassium iodide reagent.

[0033] In step (2), the concentration of iodine is 0.005 to 0.03 g / mL, for example 0.0127 g / mL, based on the mass of iodine per unit volume of iodine-potassium iodide reagent.

[0034] In step (2), the volume ratio of the solution A to the iodine-potassium iodide reagent is 1:(0.05-0.3), for example, 1:0.1.

[0035] In step (2), after the operation of mixing with the iodine-potassium iodide reagent, the operation of fixing the volume may be further included.

[0036] In step (3), when the UV-visible spectrophotometer is used for testing, the absorption wavelength is 650 to 720 nm, for example, 690 nm.

[0037] On the basis of conforming to the common sense in the art, the above-mentioned preferred conditions can be arbitrarily combined to obtain the preferred embodiments of the present application.

[0038] The reagents and raw materials used in this application are commercially available.

[0039] The positive progress of the present application is that the present application improves the extraction process of polyvinyl alcohol in polyester microparticles, increases the recovery rate of polyvinyl alcohol, and thereby improves the accuracy of the detection method for polyvinyl alcohol residues in polyester microspheres, simplifies the extraction process, reduces the detection cost, and the detection method is specific, and other components in the sample to be tested have no effect on the test results, and there will be no false positive problems. DETAILED DESCRIPTION

[0040] The present application is further described below by way of examples, but the present application is not limited to the scope of the examples. The experimental methods without specific conditions in the following examples are selected according to conventional methods and conditions, or according to the product instructions.

[0041] In the following examples and comparative examples, the preparation method of the boric acid solution includes the following steps: weigh 6.4 g of boric acid, add 200 mL of water, and dissolve and mix.

[0042] In the following examples and comparative examples, the preparation method of the iodine-potassium iodide reagent used includes the following steps: weigh 2.5 g of potassium iodide, place it in a 100 mL volumetric flask, add an appropriate amount of water to dissolve it, add 1.27 g of iodine, shake to dissolve it, dilute it to the scale with water, and shake well.

[0043] In the following examples and comparative examples, the preparation method of polyester microspheres comprises the following steps:

[0044] ① Preparation of aqueous phase: Add an appropriate amount of purified water to a container, slowly add 30g of polyvinyl alcohol 1788, stir until completely dissolved, and dilute to 1L to obtain the aqueous phase;

[0045] ② Preparation of organic phase: add an appropriate amount of dichloromethane to a container, weigh 20 g of PLA-PEG-PLA block copolymer (number average molecular weight is 68375, wherein the number average molecular weight of polyethylene glycol is 400); add it to the container, stir to completely dissolve it and adjust the volume to 100 mL to obtain an organic phase;

[0046] ③ Membrane emulsification: slowly pour the above organic phase into the aqueous phase, and use a membrane emulsifier to emulsify the mixed liquid. The pore size of the membrane tube is 50 μm to obtain an emulsion;

[0047] ④ Solidification: Stir the emulsified emulsion and blow nitrogen to volatilize the dichloromethane until the dichloromethane is completely evaporated;

[0048] ⑤ Collection: After the dichloromethane is completely evaporated, the suspension is centrifuged to collect the solid components, which are then dried to obtain polyester microspheres.

[0049] Plotting a standard curve

[0050] Preparation of reference solution: Take 40 mg of polyvinyl alcohol, weigh accurately, place in a 250 mL volumetric flask, add appropriate amount of 90°C water, sonicate to dissolve, cool to room temperature, dilute to scale with water, and shake well.

[0051] Preparation of linear series solutions: measure 0.4mL, 0.7mL, 1mL, 1.3mL, and 1.6mL of the above-prepared reference solution respectively and place them in 10mL volumetric flasks, accurately add 3mL of boric acid solution to each volumetric flask, accurately add 0.2mL of iodine-potassium iodide test solution to each volumetric flask, dilute to 10mL with water, shake well, and let stand for 10 minutes.

[0052] Preparation of blank solution: Measure 5 mL of water and place it in a 10 mL volumetric flask, accurately add 3 mL of boric acid solution, then accurately add 0.2 mL of iodine-potassium iodide test solution, dilute with water to 10 mL, shake well, and let stand for 10 min;

[0053] Determination method: UV-visible spectrophotometry was used, with blank solution as background, and the absorbance of the linear series solution prepared above was measured at a wavelength of 690 nm. The results are shown in Table 1. The polyvinyl alcohol standard working curve was drawn with the polyvinyl alcohol concentration as the abscissa and the absorbance at 692 nm as the ordinate.

[0054] As a result, the linear regression equation of polyvinyl alcohol is y=0.0198x+0.0047, and the correlation coefficient R 2 is 0.9995.

[0055] Table 1

[0056] Polyvinyl alcohol concentration (μg / mL) Absorbance value 6.48 0.134 11.35 0.232 16.21 0.323 21.07 0.418 25.94 0.523

[0057] Example 1

[0058] (A) Extraction of polyvinyl alcohol:

[0059] Step (1): 0.4 g of the polyester microsphere sample containing polyvinyl alcohol prepared above was taken, accurately weighed, placed in a 15 mL centrifuge tube, 8 mL of dichloromethane was accurately added, ultrasonicated to dissolve, and 4 mL of water was added, shaken thoroughly to mix evenly, fully extracted, centrifuged after extraction (10000 rpm, 5 min), and the aqueous phase and the organic phase were collected;

[0060] Step (2): Mix the organic phase with 4 mL of water, shake thoroughly to make the mixture uniform, extract thoroughly, centrifuge after extraction (10000 rpm, 5 min), and collect the aqueous phase; repeat this step 4 times;

[0061] Step (3): Mix the 20 mL of water phase collected in the above steps.

[0062] (B) Quantitative detection of polyvinyl alcohol:

[0063] Preparation of test solution: Place the 20mL supernatant collected above in an evaporating dish and heat in a boiling water bath for 20min. Wash with appropriate amount of hot water and transfer completely to a 25mL volumetric flask, cool, dilute to scale with water, and shake well. Accurately measure 2mL and place in a 10mL volumetric flask, accurately add 3mL of boric acid solution and 3mL of water, then accurately add 0.2mL of iodine-potassium iodide test solution, dilute to 25mL with water, shake well, and let stand for 10min.

[0064] Preparation of blank solution: Measure 5 mL of water and place it in a 10 mL volumetric flask, accurately add 3 mL of boric acid solution, then accurately add 0.2 mL of iodine-potassium iodide test solution, dilute with water to 10 mL, shake well, and let stand for 10 min;

[0065] The absorbance of the test solution was measured at 690 nm using a UV-visible spectrophotometer with a blank solution as the background. The content of polyvinyl alcohol was calculated using the absorbance according to the standard curve method.

[0066] Examples 2 to 5, Comparative Examples 1 to 2

[0067] The only difference compared with Example 1 is that during the polyvinyl alcohol extraction, the number of repetitions of step (2), the centrifugal speed, the centrifugal time, and the volume ratio of dichloromethane to water are different, and other condition parameters remain unchanged, see Table 2 below.

[0068] Table 2

[0069]

[0070] Comparative Example 3

[0071] Compared with Example 1, the only difference is that in the polyvinyl alcohol extraction process, no centrifugation is performed after extraction, and the layers are separated by standing. Other condition parameters are the same as in Example 1.

[0072] The results showed that the system could not demulsify during extraction, and the separation effect between the organic phase and the aqueous phase was poor and could not be detected.

[0073] Effect Example 1 Recovery Verification

[0074] Verify the recovery rate of the above-mentioned embodiment and comparative example detection method.

[0075] Test sample extract: The method is the same as step (A) in the above embodiment or comparative example, except that the amount of polyester microsphere sample in step (1) is adjusted to 0.3 g, and other conditions and parameters remain unchanged to obtain a polyvinyl alcohol extract;

[0076] Test sample spiked extract: Compared with the test sample extract, in step (1), before adding water, 0.5 mL of polyvinyl alcohol stock solution (concentration of 4 mg / mL) was added to the system, and the amount of water added was adjusted to 3.5 mL, while other parameters remained unchanged.

[0077] The method of the above embodiment or comparative example step (B) was used to detect the absorbance value of the corresponding test sample extract, and the mass of polyvinyl alcohol in the test sample extract was calculated as the background value; the absorbance value of the test sample spiked extract was detected, and the mass of polyvinyl alcohol in the test sample spiked extract was calculated as the detection value; the recovery rate was calculated using the following formula, and the results are shown in Table 3.

[0078] Recovery rate = (detection value - background value) / added amount × 100%; wherein the added amount is the mass of polyvinyl alcohol in the polyvinyl alcohol stock solution added to the spiked extract of the test sample.

[0079] Table 3

[0080] Recovery rate Example 1 97.6％ Example 2 87.0％ Example 3 101.7％ Example 4 96.0％ Example 5 89.7％ Comparative Example 1 73.2％ Comparative Example 2 67.5％

[0081] The results show that the number of extractions and the volume ratio of organic solvent to water have a great influence on the recovery rate. When the sample to be tested is extracted and detected using the method described in the embodiment, the recovery rate is higher, that is, the detection accuracy is higher.

[0082] Effect Example 2 Specificity Verification

[0083] The polyester microsphere sample containing polyvinyl alcohol in the above Example 1 was replaced with an equal amount of PLA-PEG-PLA block copolymer (raw material for preparing polyester microspheres), and other parameters were the same as in Example 1. No absorption peak was detected under the wavelength of 690 nm. It can be seen that the water phase does not contain impurities that affect the accuracy of the detection results, and the detection method has specificity.

[0084] Finally, it should be noted that in this application, the terms "comprises", "includes" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or apparatus that includes a series of elements includes not only those elements, but also other elements that are not explicitly listed, or also includes elements that are inherent to such process, method, article or apparatus.

[0085] Although the present application has been disclosed above through the description of the specific embodiments of the present application, it should be understood that those skilled in the art can design various modifications, improvements or equivalents to the present application within the spirit and scope of the attached scheme. These modifications, improvements or equivalents should also be considered to be included in the scope of protection claimed in the present application.

Claims

1. A method for extracting polyvinyl alcohol from polyester microparticles, characterized in that: The extraction method comprises the following steps: (1) dissolving polyester microparticles in an organic solvent, mixing with water, extracting, centrifuging, and collecting an aqueous phase and an organic phase; the volume ratio of the organic solvent to the water is (1.8-10):1; (2) taking the organic phase, mixing it with water, extracting it, centrifuging it, and collecting the aqueous phase; the volume ratio of the organic phase to the water is (1.8-10):1; (3) Mix the water phases collected in the above steps; Repeat step (2) two or more times.

2. The extraction method according to claim 1, characterized in that The extraction method satisfies at least one of the following conditions (A) to (K): (A) In step (1), the mass percentage of the polyvinyl alcohol in the polyester microparticles is less than 0.5%; (B) in step (1), the polyester is selected from at least one of polyglycolide, polylactic acid, polycaprolactone, polyhydroxyalkanoate, glycolide-lactide copolymer, caprolactone-lactide copolymer, polycaprolactone-polyethylene glycol copolymer and polylactic acid-polyethylene glycol copolymer; (C) In step (1), the polyester microparticles are prepared by at least one method selected from the group consisting of solvent volatilization, phase separation, spray drying, surface deposition and grinding; (D) In ​​step (1), the dissolution is carried out under ultrasonic conditions; (E) In step (1), the mass volume ratio of the polyester microparticles to the organic solvent is 0.005 to 0.5 g / mL; (F) In step (1), the organic solvent is a halogenated alkane solvent; (G) In step (1), the volume ratio of the organic solvent to the water is (1.8-5):1; (H) In step (1) and step (2), the centrifugal speed is independently 800 to 30,000 rpm; (I) In step (1) and step (2), the centrifugation time is independently 1 to 15 min; (J) In step (2), the volume ratio of the organic phase to the water is (1.8-5):1; (K) Repeat step (2) 2 to 5 times.

3. The extraction method according to claim 2, characterized in that The extraction method satisfies at least one of the following conditions (A) to (C): (A) In step (1), the organic solvent is a C2-C6 halogenated alkane; (B) In step (1) and step (2), the centrifugal speed is independently 1000 to 15000 rpm; (C) In step (1) and step (2), the centrifugation time is independently 1 to 10 minutes.

4. The extraction method according to claim 3, characterized in that In step (1), the organic solvent is dichloromethane.

5. A method for quantitatively detecting polyvinyl alcohol in polyester microparticles, characterized in that: The steps include: (1) obtaining an aqueous phase by the method for extracting polyvinyl alcohol from polyester microparticles as described above, heating the aqueous phase, and fixing the aqueous phase to obtain a solution A; (2) taking part of the solution A and mixing it with a boric acid solution, and then mixing it with an iodine-potassium iodide reagent to obtain a solution B; (3) The absorbance value of the solution B is measured by ultraviolet-visible spectrophotometry, and the content of the polyvinyl alcohol is calculated by comparing with a standard curve.

6. The method for quantitatively detecting polyvinyl alcohol in polyester microparticles according to claim 5, characterized in that: The quantitative detection method satisfies at least one of the following conditions (A) to (C): (A) In step (1), the heating is performed in a boiling water bath; (B) In step (1), the heating time is 5 to 40 minutes; (C) In step (1), after the volume is fixed, the concentration of the polyester microparticles is 2 to 30 mg / mL, preferably 4 to 16 mg / mL, based on the mass of the polyester microparticles per unit volume of the aqueous phase.

7. The method for quantitatively detecting polyvinyl alcohol in polyester microparticles according to claim 5 or 6, characterized in that: The quantitative detection method satisfies at least one of the following conditions (A) to (G): (A) In the boric acid solution of step (2), the mass volume ratio of boric acid to solvent is 0.015 to 0.05 g / mL; (B) In step (2), the volume ratio of the solution A to the boric acid solution is 1:(0.5-3); (C) In step (2), the concentration of potassium iodide is 0.01 to 0.05 g / mL, based on the mass of potassium iodide per unit volume of iodine-potassium iodide reagent; (D) In ​​step (2), the concentration of iodine is 0.005 to 0.03 g / mL, based on the mass of iodine per unit volume of iodine-potassium iodide reagent; (E) In step (2), the volume ratio of the solution A to the iodine-potassium iodide reagent is 1:(0.05-0.3); (F) in step (2), after the operation of mixing with the iodine-potassium iodide reagent, further comprising an operation of fixing the volume; (G) In step (3), when the UV-visible spectrophotometer is used for testing, the absorption wavelength is 650-720 nm.

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