WO2017177668A1 - 一种地榆皂苷ii聚合物胶束及其制备方法 - Google Patents

一种地榆皂苷ii聚合物胶束及其制备方法 Download PDF

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WO2017177668A1
WO2017177668A1 PCT/CN2016/104876 CN2016104876W WO2017177668A1 WO 2017177668 A1 WO2017177668 A1 WO 2017177668A1 CN 2016104876 W CN2016104876 W CN 2016104876W WO 2017177668 A1 WO2017177668 A1 WO 2017177668A1
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saponin
mpeg
pla
polymer micelle
parts
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PCT/CN2016/104876
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French (fr)
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杨世林
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四川英路维特医药科技有限公司
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Priority to US16/093,118 priority Critical patent/US10493029B2/en
Publication of WO2017177668A1 publication Critical patent/WO2017177668A1/zh

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
    • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
    • A61K31/704—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin attached to a condensed carbocyclic ring system, e.g. sennosides, thiocolchicosides, escin, daunorubicin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/34—Macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyesters, polyamino acids, polysiloxanes, polyphosphazines, copolymers of polyalkylene glycol or poloxamers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/19—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles lyophilised, i.e. freeze-dried, solutions or dispersions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants

Definitions

  • the invention relates to a saponin II polymer micelle and a preparation method thereof, and belongs to the field of medicine.
  • Myelosuppression is a clinically common hematopoietic system disease. Radiation therapy and/or chemotherapy of various systemic neoplastic diseases, radiation damage caused by ionizing radiation, viral hepatitis, parvovirus infection or drugs can cause bone marrow suppression. Myelosuppression mainly manifests as damage to the bone marrow microenvironment, hematopoietic stem cells, hematopoietic growth factors, etc., and the granulocyte, red, and megakaryocyte system is inhibited by one, two or three lines; among them, granulocyte deficiency causes serious infection, red blood cells Significant reduction can cause severe anemia, and a significant drop in platelets causes severe bleeding and even death. Because myelosuppression is a serious threat to the health of patients, especially the quality of life of cancer patients undergoing radiotherapy and chemotherapy, many research groups are currently working to find effective drugs to prevent bone marrow suppression.
  • Earthworm saponin II chemical name: 3-O- ⁇ -L-arabinose-19 ⁇ -hydroxyisoxa-12-carboxylic acid (ziyu-glycoside II), is a mantle from the genus Rosaceae
  • CN101119740A discloses the use of saponin II in the preparation of a medicament for increasing red blood cells and hemoglobin.
  • the saponin II has low solubility in water and low oral gastrointestinal absorption rate, resulting in low oral bioavailability of the drug, which greatly limits its clinical application.
  • polymer micelles can solubilize poorly soluble drugs, and have the characteristics of stable structure, excellent tissue permeability, long residence time in the body, and effective drug reaching the target.
  • An excellent carrier for poorly soluble drugs, An excellent carrier.
  • carrier materials for preparing polymer micelles are generally used, and amphiphilic block copolymers are usually used, and hydrophilic segments may be selected from polyethylene glycol, polyoxyethylene, povidone, chitosan, etc., hydrophobic segments.
  • the materials mainly include polypropylene, polystyrene, polyamino acid, polylactic acid, spermine or short-chain phospholipids. It is the main problem in designing polymer micelles to select suitable polymer materials according to the main pharmacological properties, to prepare micelles with the required quality of the preparation, and to effectively release the drugs therefrom and to exert pharmacodynamic effects in the living body.
  • the object of the present invention is to provide a saponin II polymer micelle and a preparation method thereof.
  • the present invention provides a polymer micelle which is prepared by containing a raw material of the following weight ratio: 20 parts of saponin II and 100 parts to 5000 parts of mPEG-PLA-Phe (Boc).
  • it is prepared from the following raw materials by weight ratio: 20 parts of saponin II and 400 parts to 5000 parts of mPEG-PLA-Phe (Boc).
  • it is prepared from the following raw materials by weight ratio: 20 parts of saponin II and 4000 parts of mPEG-PLA-Phe (Boc).
  • the polymer micelle further comprises 0-5 parts of sugar, and the sugar is selected from one or more of glucose, sucrose, trehalose, fructose, mannitol or lactose.
  • the mPEG-PLA-Phe (Boc) has a number average molecular weight of 1000 to 5000, and a PLA number average molecular weight of 1000 to 50000.
  • the mPEG-PLA-Phe (Boc) has a mPEG number average molecular weight of 2,000 and a PLA number average molecular weight of 4,000.
  • the invention provides a preparation method of the polymer micelle, comprising the steps of: taking each weight ratio of saponin II, mPEG-PLA-Phe (Boc), and dissolving in 12-125 times the volume of ethanol. In the middle, the ethanol is removed by evaporation to obtain a polymer micelle.
  • the preparation method further comprises the steps of: taking 0 to 5 parts of sugar, and dissolving the polymer micelle together in water to make the concentration of the saponin II is 0.01 mg/mL to 20 mg/mL. Filtered with a 0.22 um microporous membrane, sterilized, and lyophilized to obtain a polymer micelle lyophilized powder.
  • the present invention provides the use of the polymeric micelles in the manufacture of a medicament for the treatment and/or prevention of myelosuppression.
  • the medicament is a medicament for treating and/or preventing bone marrow suppression caused by a chemical substance.
  • the drug is a drug that increases the amount of one or more of peripheral blood leukocytes (WBC), neutral granules (NEUT), red blood cells (RBC), platelets (PLT), or hemoglobin (HGB).
  • WBC peripheral blood leukocytes
  • NUT neutral granules
  • RBC red blood cells
  • PHT platelets
  • HGB hemoglobin
  • the present invention provides a method of treating and/or preventing myelosuppression using the polymeric micelles described.
  • the invention provides a saponin II polymer micelle and a preparation method thereof.
  • the quality evaluation experiment shows that the preparation is only obtained when the mPEG-PLA-Phe (Boc) of the present invention is used as a carrier material.
  • the quality of the mantle saponin II polymer micelles is the best, and the use of other materials leads to a decrease in the quality of the preparation.
  • the saponin II micelles of the present invention can significantly increase the number of WBC, RBC, PLT, NEUT and HGB in the peripheral blood, and the drug efficacy is obviously superior to that of the saponin II drug, indicating that the present invention
  • the invented saponin II polymer micelle has a good therapeutic and/or preventive effect on myelosuppression, and can improve the bioavailability of the poorly soluble drug saponin II.
  • the raw materials and equipment used in the specific embodiments of the present invention are known products and are obtained by purchasing commercially available products.
  • the above-mentioned saponin II micelle freeze-dried powder has good resolubility, and the particle size distribution changes little before and after reconstitution. Its main drug content is 0.4mg/mL, encapsulation efficiency is >99%, particle size is ⁇ 100nm, PDI ⁇ 0.3.
  • Sample preparation Take the saponin micelle solution and dilute the appropriate multiples with methanol, and inject the high-performance liquid chromatograph to measure the particle size and distribution.
  • Sample preparation Take the saponin micelle solution and dilute the appropriate multiples with ultrapure water, and inject the particle size analyzer to measure the particle size and distribution.
  • amphiphilic block copolymers composed of various hydrophilic and hydrophobic segments were investigated according to the properties of the saponins II.
  • the five materials with good polymer micelles are listed below.
  • mPEG-PLA-Phe(Boc) a methoxypolyethylene glycol-polylactic acid block copolymer terminated with tert-butoxyphenylalanine, the hydrophilic chain of which is a mPEG having a number average molecular weight of 1000-5000
  • the parent ester chain is a PLA having a number average molecular weight of 1000 to 5000, and the structure is as in Formula I:
  • mPEG-PLA is a methoxy polyethylene glycol-polylactic acid block copolymer
  • the hydrophilic chain is mPEG having a number average molecular weight of 1000-5000
  • the lipophilic chain is a PLA having a number average molecular weight of 1000-5000
  • Poloxamer F68 poloxamer is a polyoxyethylene polyoxypropylene ether block copolymer.
  • Solutol HS15 is polyethylene glycol-12-hydroxystearate.
  • Poloxamer F68, Solutol HS15, and polyoxyethylene castor oil EL35 were purchased from BASF, Germany.
  • the polymer micelles prepared by the carrier material mPEG-PLA-Phe(Boc) of the invention have the highest content of saponin II (up to 0.391 mg/mL), the average particle size is small (87 nm), and the dispersion index ( PDI) was only 0.16, indicating that the particle size of the resulting micelles was very uniform.
  • the other two materials, mPEG-PLA and Poloxamer F68 resulted in a significant decrease in drug content and particle size uniformity, and the micelles obtained with mPEG-PLA had larger average particle size and were not suitable for injection.
  • the use of Solutol HS15 and polyoxyethylene castor oil EL35 also has the problem of a decrease in the drug content.
  • the ceramide saponin II polymer micelle can be prepared; wherein, mPEG2000-PLA4000-Phe (Boc) is used.
  • the content of saponin II in polymer micelles was the highest (up to 0.391 mg/mL), the average particle size was small (87 nm), and the dispersion index (PDI) was only 0.16, indicating that the quality of the obtained micelles was the best.
  • the saponin II and MPEG-PLA-Phe (Boc) were weighed according to the mass ratio of 1:3-1:250 (the mass of the fixed saponin II was 20 mg, and the mass of MPEG-PLA-Phe (Boc) varied with the ratio) Dissolve in 50ml of ethanol, remove the solvent by rotary evaporation at 55 °C, add 50ml of water for injection to dissolve the film, and filter the obtained micelle solution through a 0.22 ⁇ m sterile membrane to determine the content and particle size distribution of saponin II in the micelle. table 3.
  • the carrier material MPEG-PLA-Phe (Boc) dosage is 5 ⁇ 250 times of the saponin II
  • the polymer micelles have better quality: the drug content is not less than 0.01mg / mL, the average particle size is below 90nm, The PDI is lower than 0.2; among them, the dosage of MPEG-PLA-Phe(Boc) is in the range of 20-250 times of the mass of the saponin II, and the quality of the preparation can be further optimized, mainly in the high drug content, which can reach 0.4 mg/ About mL, wherein the mass ratio of saponin II:mPEG-PLA-Phe(Boc) is 1:200, the micelle quality is the best: the drug content is the highest and the particle size distribution is the most uniform.
  • mice All animals were fed ad libitum for 1 week and were randomly divided into: blank group; model group; saponin II micelle sample 1 to 4 (using the micelles prepared in Examples 1-4, respectively), the dose was 2.5 mg. ⁇ kg-1, prepared before use; saponin II group: saponin II original drug powder, dissolved in 10% DMSO-physiological saline, the dose is 2.5mg ⁇ kg-1, prepared before use.
  • the other groups of mice were intraperitoneally injected with cyclophosphamide physiological saline solution at a dose of 120 mg ⁇ kg-1, and the blank group was intraperitoneally injected with an equal volume of normal saline.
  • Each experimental group was given the corresponding drug by dose and administration from the first day of the experiment.
  • the blank group and the model group were injected with the same volume of normal saline in the tail vein for 6 consecutive days.
  • WBC Peripheral blood leukocytes
  • NUT neutrophils
  • RBC red blood cells
  • PHT platelets
  • HGB hemoglobin

Abstract

一种地榆皂苷II聚合物胶束，其由下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)100份～5000份。质量评价实验表明，仅有在使用mPEG-PLA-Phe(Boc)作为载体材料时，制备得到的地榆皂苷II聚合物胶束质量最佳，采用其他材料则会导致制剂质量的下降。药效实验中，与模型组比较，该地榆皂苷II胶束能显著升高外周血WBC、RBC、PLT、NEUT和HGB数量，且药效明显优于地榆皂苷II原药，表明该地榆皂苷II聚合物胶束对骨髓抑制有较好的治疗和/或预防作用，能提高难溶性药物地榆皂苷II的生物利用度。

Description

一种地榆皂苷II聚合物胶束及其制备方法 技术领域
本发明涉及一种地榆皂苷II聚合物胶束及其制备方法，属于医药领域。
背景技术
骨髓抑制是临床上常见的造血系统疾病，各系统肿瘤性疾病的放射治疗和/或化学治疗、电离辐射引起的放射损伤、病毒性肝炎、微小病毒感染或药物等因素均可引起骨髓抑制。骨髓抑制主要表现为骨髓微环境、造血干细胞、造血细胞生长因子等的损伤，粒、红、巨核细胞系统一系、二系或三系细胞受抑制；其中，粒细胞缺乏会引起严重感染，红细胞明显减少会引起严重贫血，血小板明显下降引起严重出血，甚至导致死亡。由于骨髓抑制严重威胁着患者的生命健康，尤其对进行放化疗的肿瘤患者的生活质量带来不利影响，因此目前许多研究小组都致力于寻找有效的药物来防治骨髓抑制。
地榆皂苷Ⅱ，化学名：3-O-α-L-阿拉伯糖基-19α-羟基乌索-12烯-28-羧酸(ziyu-glycoside II)，是从蔷薇科地榆属植物地榆(Sanguisorba officinalis L.)或长叶地榆[S.officinalis L.var.longifolia(Bertol.)Yu et Li]的根中提取得到的一种有效成分。CN101119740A公开了地榆皂苷Ⅱ在制备升高红细胞和血红蛋白的药物中的用途。然而，地榆皂苷II在水中溶解度低，口服胃肠吸收率小，造成该药物口服生物利用度低，大大限制了在临床上的应用。
聚合物胶束作为药用载体可使难溶药物增溶，且具有结构稳定、优良的组织渗透性、体内滞留时间长、能使药物有效地到达靶点等特点，对难溶性药物来说是一种优良的载体。目前，用于制备聚合物胶束的载体材料种类繁多，通常采用两亲性嵌段共聚物，亲水段可选用聚乙二醇、聚氧乙烯、聚维酮、壳聚糖等，疏水段材料主要有聚丙烯、聚苯乙烯、聚氨基酸、聚乳酸、精胺或短链磷脂等等。根据主药理化性质选择合适的聚合物材料，从而制备得到制剂质量符合要求的胶束，并使药物能够从中有效地释放进而在生物体内发挥药效，是设计聚合物胶束面临的主要难题之一。
因此，开发一种制剂质量较好且药效明确的地榆皂苷Ⅱ聚合物胶束，用于治疗和/或预防骨髓抑制，成为了一个亟待解决的问题。
发明内容
本发明的目的在于提供一种地榆皂苷II聚合物胶束及其制备方法。
本发明提供了一种聚合物胶束，它是包含下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)100份～5000份。
进一步的，它是由下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)100份～5000份。
优选的，它是由下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)400份～5000份。
进一步优选的，它是由下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)4000份。
其中，所述的聚合物胶束还包含糖0～5份，所述的糖选自葡萄糖、蔗糖、海藻糖、果糖、甘露醇或乳糖中的一种或几种。
其中，所述的mPEG-PLA-Phe(Boc)中mPEG数均分子量在1000～5000之间，PLA数均分子量在1000～50000之间。
进一步的，所述的mPEG-PLA-Phe(Boc)中mPEG数均分子量为2000，PLA数均分子量为4000。
本发明提供了一种所述聚合物胶束的制备方法，包括如下步骤：取各重量配比的地榆皂苷II、mPEG-PLA-Phe(Boc)，溶解于12～125倍体积量的乙醇中，再将乙醇蒸发除去，即得聚合物胶束。
进一步的，所述的制备方法还包括如下步骤：取0～5份糖，与所述的聚合物胶束一并溶解于水中，使地榆皂苷II的浓度为0.01mg/mL～20mg/mL，用0.22um微孔滤膜过滤，灭菌，冷冻干燥，即得聚合物胶束冻干粉。
本发明提供了所述聚合物胶束在制备治疗和/或预防骨髓抑制的药物中的用途。
进一步的，所述的药物是治疗和/或预防化学物质所致骨髓抑制的药物。
更进一步的，所述的药物是升高外周血白细胞(WBC)、中性粒细(NEUT)、红细胞(RBC)、血小板(PLT)或血红蛋白(HGB)中一种或几种的数量的药物。
本发明提供了一种治疗和/或预防骨髓抑制的方法，它是使用所述的聚合物胶束。
本发明提供了一种地榆皂苷II聚合物胶束及其制备方法。质量评价实验表明，仅有在使用本发明mPEG-PLA-Phe(Boc)作为载体材料时，制备得到的 地榆皂苷II聚合物胶束质量最佳，采用其他材料则会导致制剂质量的下降。药效实验中，与模型组比较，本发明地榆皂苷II胶束能显著升高外周血WBC、RBC、PLT、NEUT和HGB数量，且药效明显优于地榆皂苷II原药，表明本发明地榆皂苷II聚合物胶束对骨髓抑制有较好的治疗和/或预防作用，能提高难溶性药物地榆皂苷II的生物利用度。
显然，根据本发明的上述内容，按照本领域的普通技术知识和惯用手段，在不脱离本发明上述基本技术思想前提下，还可以做出其它多种形式的修改、替换或变更。
以下通过实施例形式的具体实施方式，对本发明的上述内容再作进一步的详细说明。但不应将此理解为本发明上述主题的范围仅限于以下的实例。凡基于本发明上述内容所实现的技术均属于本发明的范围。
具体实施方式
本发明具体实施方式中使用的原料、设备均为已知产品，通过购买市售产品获得。
实施例1本发明聚合物胶束的制备
取20mg地榆皂苷II、100mg mPEG1000-PLA1000-Phe(Boc)，溶解于15ml乙醇，55℃旋转蒸发去除溶剂。
加入50ml注射用糖水(含5mg葡萄糖)溶解药膜，所得胶束溶液经0.22μm除菌膜过滤后冻干，得地榆皂苷II胶束冻干粉。
实施例2本发明聚合物胶束的制备
取20mg地榆皂苷II、400mg mPEG2000-PLA4000-Phe(Boc)，溶解于15ml乙醇，55℃旋转蒸发去除溶剂。
加入50ml注射用糖水(含5mg蔗糖)溶解药膜，所得胶束溶液经0.22μm除菌膜过滤后冻干，得地榆皂苷II胶束冻干粉。
上述地榆皂苷II胶束冻干粉复溶性好，复溶前后粒径分布变化较小。其主药含量达0.4mg/mL，包封率>99％，粒径<100nm，PDI<0.3。
实施例3本发明聚合物胶束的制备
取20mg地榆皂苷II、2000mg mPEG5000-PLA3000-Phe(Boc)，溶解于50ml乙醇，55℃旋转蒸发去除溶剂。
加入50ml注射用糖水(含5mg海藻糖)溶解药膜，所得胶束溶液经0.22μm除菌膜过滤后冻干，得地榆皂苷II胶束冻干粉。
实施例4本发明聚合物胶束的制备
取20mg地榆皂苷II、4000mg mPEG2000-PLA5000-Phe(Boc)，溶解于50ml乙醇，55℃旋转蒸发去除溶剂。
加入50ml注射用糖水(含5mg果糖)溶解药膜，所得胶束溶液经0.22μm除菌膜过滤后冻干，得地榆皂苷II胶束冻干粉。
以下通过实验例证明本发明的有益效果。
含量测定：
仪器：高效液相色谱仪Waters e2695
色谱条件：色谱柱：十八烷基键合硅胶(4.6×250mm，5μm)
检测波长：203nm
流动相：甲醇：0.1％甲酸＝80:20
流速：1mL/min
柱温：25℃
进样量：20μL
样品配制：取地榆皂苷胶束溶液以甲醇稀释适当倍数，注入高效液相色谱仪中测量粒径及分布。
粒径及分布测定：
仪器：马尔文ZS90
样品配制：取地榆皂苷胶束溶液以超纯水稀释适当倍数，注入粒度仪中测量粒径及分布。
实验例1采用不同载体材料制备的聚合物胶束的质量评价
本研究前期根据地榆皂苷II的性质考察了多种亲水段、疏水段材料组成的两亲性嵌段共聚物，以下列出所得聚合物胶束质量较好的五种材料。
1、mPEG-PLA-Phe(Boc)：为叔丁氧基苯丙氨酸封端的甲氧基聚乙二醇-聚乳酸嵌段共聚物，其亲水链为数均分子量为1000～5000的mPEG，亲酯链为数均分子量为1000～5000的PLA，结构如式I：
Figure PCTCN2016104876-appb-000001
该高分子聚合物合成方法见文献：[1]陈雪冬，杨世林，冯育林，刘珂等，牛蒡子苷元mPEG-PDLLA聚合物胶束的制备及表征[J].医药，2015，18(8)：153；[2]陈雪冬，一种新型载多西他赛mPEG-PDLLA-Phe(Boc)胶束的制备及评价，江西中医药大学硕士学位论文，2016.6。
2、mPEG-PLA：为甲氧基聚乙二醇-聚乳酸嵌段共聚物，其亲水链为数均分子量为1000～5000的mPEG，亲酯链为数均分子量为1000～5000的PLA，结构如式II：
Figure PCTCN2016104876-appb-000002
该高分子聚合物合成方法见上述文献。
3、Poloxamer F68即泊洛沙姆，为聚氧乙烯聚氧丙烯醚嵌段共聚物。
4、Solutol HS15为聚乙二醇-12-羟基硬脂酸酯。
5、聚氧乙烯蓖麻油EL35。
Poloxamer F68，Solutol HS15，聚氧乙烯蓖麻油EL35均购于德国BASF公司。
实验方法
1、分别称取20mg地榆皂苷II及400mg的不同高分子材料mPEG2000-PLA4000-Phe(Boc)，mPEG2000-PLA4000，Poloxamer F68，Solutol HS15及聚氧乙烯蓖麻油EL35溶解于15ml乙醇，55℃旋转蒸发去除溶剂后加入50ml注射用水溶解药膜，所得胶束溶液经0.22μm除菌膜过滤后，测定胶束中地榆皂苷II含量及粒径分布，结果见表1。
表1采用不同载体材料制备的聚合物胶束的质量评价
Figure PCTCN2016104876-appb-000003
实验结果：采用本发明载体材料mPEG-PLA-Phe(Boc)制备的聚合物胶束中地榆皂苷II含量最高(达到0.391mg/mL)，平均粒径较小(87nm)，且分散指数(PDI)仅0.16，表明所得胶束的粒径非常均匀。另两种材料mPEG-PLA、Poloxamer F68会导致药物含量及粒径均匀度的明显降低，而且采用mPEG-PLA得到的胶束平均粒径较大，不适于注射。使用Solutol HS15、聚氧乙烯蓖麻油EL35同样存在药物含量下降的问题。
以上实验结果表明：仅有在使用本发明mPEG-PLA-Phe(Boc)作为载体材料时，制备得到的地榆皂苷II聚合物胶束质量最佳，采用其他材料则会导致制剂质量的下降。
2、分别称取20mg地榆皂苷II及400mg的不同数均分子量的mPEG-PLA-Phe(Boc)溶解于15ml乙醇，55℃旋转蒸发去除溶剂后加入50ml注射用水溶解药膜，所得胶束溶液经0.22μm除菌膜过滤后，测定胶束中地榆皂苷II含量及粒径分布，结果见表2。
表2采用不同分子量mPEG-PLA-Phe(Boc)制备的聚合物胶束的质量评价
Figure PCTCN2016104876-appb-000004
实验结果：采用本发明数均分子量范围内的mPEG-PLA-Phe(Boc)作为载体材料，均能制备得到地榆皂苷II聚合物胶束；其中，采用mPEG2000-PLA4000-Phe(Boc)制备的聚合物胶束中地榆皂苷II含量最高(达到0.391mg/mL)，平均粒径较小(87nm)，且分散指数(PDI)仅0.16，表明所得胶束的质量最佳。
实验例2根据不同载体材料用量制备的聚合物胶束的质量评价
按照质量比1:3-1:250的比例称取地榆皂苷II及MPEG-PLA-Phe(Boc)(固定地榆皂苷II质量为20mg，MPEG-PLA-Phe(Boc)质量随比例变化)，溶解于50ml乙醇，55℃旋转蒸发去除溶剂后加入50ml注射用水溶解药膜，所得胶束溶液经0.22μm除菌膜过滤后，测定胶束中地榆皂苷II含量及粒径分布，结果见表3。
表3根据不同载体材料用量制备的聚合物胶束的质量评价
Figure PCTCN2016104876-appb-000005
实验结果：载体材料MPEG-PLA-Phe(Boc)用量为地榆皂苷II 5～250倍时所得聚合物胶束质量较好：药物含量不低于0.01mg/mL，平均粒径在90nm以下，PDI均低于0.2；其中，MPEG-PLA-Phe(Boc)用量在地榆皂苷II质量20～250倍范围内，制剂质量可得到进一步优化，主要表现在药物含量较高，可达到0.4mg/mL左右；其中，地榆皂苷II：mPEG-PLA-Phe(Boc)质量比为1：200时胶束质量最佳：药物含量最高、粒径分布最为均匀。
实验例3本发明地榆皂苷II聚合物胶束的药效实验
1、实验材料、试剂、仪器
受试药物地榆皂苷II胶束、地榆皂苷II原粉。
工具药物环磷酰胺。
实验动物KM-小鼠：18.5～22.5g。
实验仪器：全自动血球分析仪；BS-600L电子天平：规格：600g/0.1g，上海友声衡器有限公司。
2、统计方法
用SPSS 17.0软件进行统计分析。数据以均数±标准差
Figure PCTCN2016104876-appb-000006
表示，组间采用单因素方差分析，方差齐者组间进行LSD检验，方差不齐者进行Tamhane’s T2检验。
3、实验方法
3.1实验动物分组及模型制备
所有动物适应性喂养1周后按体重随机分为：空白组；模型组；地榆皂苷II胶束样品①～④组(分别使用实施例1～4制备的胶束)，给药剂量2.5mg·kg-1，临用前配制；地榆皂苷II组：地榆皂苷II原药粉末，用10％DMSO-生理盐水溶解，给药剂量2.5mg·kg-1，临用前配制。实验第1天，除空白组外，其余各组小鼠按120mg·kg-1剂量腹腔注射环磷酰胺生理盐水溶液，空白组小鼠腹腔注射等体积生理盐水。
3.2给药
各实验组自实验第1天开始按剂量、给药方式给予相应药物，空白组和模型组小鼠尾静脉注射等体积生理盐水，连续6天。
3.3标本采集
实验第7天，各实验组小鼠眼眶取血，用装有EDTA抗凝剂的0.5mlEP管收集待测。
3.4检测指标及方法
采用全自动血球计数仪对各实验组小鼠外周血白细胞(WBC)、中性粒细胞(NEUT)红细胞(RBC)、血小板(PLT)、血红蛋白(HGB)进行计数。
4、实验结果
表4各实验组小鼠外周血WBC、RBC和PLT数量
Figure PCTCN2016104876-appb-000007
注：与模型组比较，*P<0.05，**P<0.01；注：与地榆皂苷II组比较，△P<0.05，△△P<0.01。
实验结果：与模型组比较，本发明地榆皂苷II胶束样品实验组①～④小鼠外周血WBC、RBC、PLT数量均有显著升高(P<0.05)，地榆皂苷II组则没有显著性差异；与地榆皂苷II组比较，本发明地榆皂苷II胶束样品实验组①～④小鼠外周血WBC、RBC、PLT数量均有显著升高(P<0.05)。
表5各实验组小鼠外周血NEUT、HGB计数
Figure PCTCN2016104876-appb-000008
注：与模型组比较，*P<0.05，**P<0.01；与地榆皂苷II组比较，△P<0.05，△△P<0.01。
实验结果：与模型组比较，本发明地榆皂苷II胶束样品实验组①～④小鼠外周血NEUT和HGB数量均有显著升高(P<0.05)，地榆皂苷II组无显著 性差异；与地榆皂苷II组比较，本发明地榆皂苷II胶束样品实验组①～④小鼠外周血NEUT和HGB数量均有显著升高(P<0.05)。
以上实验结果表明：本发明地榆皂苷II聚合物胶束对骨髓抑制有较好的治疗和/或预防作用，药效明显优于直接用地榆皂苷II原药给药。

Claims (11)

  1. 一种聚合物胶束，其特征是：它是包含下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)100份～5000份。
  2. 如权利要求1所述的聚合物胶束，其特征是：它是由下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)100份～5000份。
  3. 如权利要求2所述的聚合物胶束，其特征是：它是由下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)400份～5000份。
  4. 如权利要求3所述的聚合物胶束，其特征是：它是由下述重量配比的原辅料制备而成：地榆皂苷II 20份、mPEG-PLA-Phe(Boc)4000份。
  5. 如权利要求1所述的聚合物胶束，其特征是：它还包含糖0～5份，其中，所述的糖选自葡萄糖、蔗糖、海藻糖、果糖、甘露醇或乳糖中的一种或几种。
  6. 如权利要求1～5任意一项所述的聚合物胶束，其特征是：所述的mPEG-PLA-Phe(Boc)中mPEG数均分子量在1000～5000之间，PLA数均分子量在1000～50000之间。
  7. 如权利要求6所述的聚合物胶束，其特征是：所述的mPEG-PLA-Phe(Boc)中mPEG数均分子量为2000，PLA数均分子量为4000。
  8. 一种权利要求1～7任意一项所述聚合物胶束的制备方法，其特征是：包括如下步骤：取各重量配比的地榆皂苷II、mPEG-PLA-Phe(Boc)，溶解于12～125倍体积量的乙醇中，再将乙醇蒸发除去，即得聚合物胶束。
  9. 如权利要求8所述的制备方法，其特征是：还包括如下步骤：取0～5份糖，与所述的聚合物胶束一并溶解于水中，使地榆皂苷II的浓度为0.01mg/mL～20mg/mL，用0.22um微孔滤膜过滤，灭菌，冷冻干燥，即得聚合物胶束冻干粉。
  10. 权利要求1～7任意一项所述聚合物胶束在制备治疗和/或预防骨髓抑制的药物中的用途。
  11. 一种治疗和/或预防骨髓抑制的方法，其特征是：它是使用权利要求1～7任意一项所述的聚合物胶束。
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