发明领域field of invention
本发明涉及基于透明质酸的产品领域。The present invention relates to the field of products based on hyaluronic acid.
现有技术current technology
下文通常所述的透明质酸，连同它的盐称为透明质素HA，是负电荷直链多糖，由n个二糖单元(-4GlcUAβ1-3GlcNAcβ1-)的重复组成，其中D-葡糖醛酸(GlcUA)和N-乙酰基-D-葡糖胺(GlcNAc)是用交替的β-1,3和β-1,4糖苷键连接的。The hyaluronic acid generally described below, together with its salts called hyaluronan HA, is a negatively charged linear polysaccharide composed of repeats of n disaccharide units (-4GlcUAβ1-3GlcNAcβ1-), in which D-glucuronide The acid (GlcUA) and N-acetyl-D-glucosamine (GlcNAc) are linked by alternating β-1,3 and β-1,4 glycosidic linkages.
HA是高水溶性多糖，并且HA溶液表现出非牛顿型粘弹性性能。这些性质取决于分子量(由于HA是线性聚合物，因此，分子量取决于链的长度)、浓度、pH和离子强度。HA is a highly water-soluble polysaccharide, and HA solutions exhibit non-Newtonian viscoelastic properties. These properties depend on molecular weight (since HA is a linear polymer, molecular weight depends on chain length), concentration, pH and ionic strength.
由于它的生物学性质和功能，HA具有很高的附加值(它的商业价值大大超过了其它天然多糖)，应用范围从医药方面到药用化妆品和营养食品。它的粘弹性连同完全没有毒性或免疫原性(HA的结构在其存在的所有活的有机体中通常是一样的)产生多种且广泛的应用。Due to its biological properties and functions, HA has high added value (its commercial value greatly exceeds that of other natural polysaccharides), and its applications range from pharmaceuticals to cosmeceuticals and nutraceuticals. Its viscoelasticity coupled with the complete absence of toxicity or immunogenicity (the structure of HA is generally the same in all living organisms in which it exists) yields diverse and wide-ranging applications.
在很多这些应用中，性能取决于HA的分子量。对于这一点，当开发HA的制备方法和应用策略时，HA的平均分子量和多分散性指数Mw/Mn(其测量分子量分布曲线的宽度，其中Mn是数量-平均分子量，定义为样品中所有聚合物分子的总重量除以分子总数，并且Mw是重量-平均分子量，其考虑存在的分子的变化质量)必须是考虑的金标准。In many of these applications, performance depends on the molecular weight of the HA. For this, when developing HA preparation methods and application strategies, the average molecular weight and polydispersity index Mw/Mn of HA (which measures the width of the molecular weight distribution curve, where Mn is the number-average molecular weight, defined as the Dividing the total weight of the molecule by the total number of molecules, and Mw being the weight-average molecular weight, which takes into account the varying masses of molecules present) must be the gold standard for consideration.
特别地，目前与其分子量相关的HA的多种生物学响应意味着在文中必须使用低分子量HA(L-HA)和高分子量HA(H-HA)。In particular, the present multiple biological responses of HA in relation to its molecular weight mean that low molecular weight HA (L-HA) and high molecular weight HA (H-HA) must be used in this context.
发明描述Description of the invention
本发明描述了L-HA与H-HA之间的协同杂化复合物(cooperative hybridcomplexes)(命名为简称L/H-HA)、它们的特性、其制备方法以及其在医药、化妆品和食品领域的用途。弱作用力(例如氢键或疏水相互作用)能在分子间产生非常稳定的相互作用，如果这些是协同类型。当在分子间可能形成多重键时，产生协同性，并且很弱，随后它们随机断裂，但是由于存在完整的邻位键，多重键能立即重新形成，所述的邻位键在可用于重新形成的距离处维持键的结构组成。The present invention describes cooperative hybrid complexes (cooperative hybrid complexes) between L-HA and H-HA (named L/H-HA for short), their properties, their preparation methods and their applications in the fields of medicine, cosmetics and food the use of. Weak forces (such as hydrogen bonds or hydrophobic interactions) can produce very stable interactions between molecules, if these are of the cooperative type. Cooperativity arises when multiple bonds may form between molecules, and are weak, then they break randomly, but multiple bonds can be reformed immediately due to the presence of intact ortho bonds, which are available for reformation The structural composition of the bond is maintained at a distance of .
溶液中HA分子的特征在于基于形成疏水键和链间氢键的相互作用的协同现象，并且这些相互作用的协同性取决于链的长度，并且因此取决于链的分子量。H-HA的长链提供它们之间稳定的相互作用，其涉及溶液中存在的所有分子，其产生三维网络，然而L-HA分子提供不太稳定的相互作用，导致聚集的体系，其没有同时涉及存在的所有分子，其替代聚簇中的相互作用。溶液中H-HA和L-HA的这种不同方式的聚集引起流变学性能的很大差异，例如HA溶液的粘度，对于很多应用(特别是在医药领域中)，这是非常重要的性质。HA molecules in solution are characterized by synergistic phenomena based on interactions that form hydrophobic bonds and interchain hydrogen bonds, and the cooperativity of these interactions depends on the length of the chains, and thus on the molecular weight of the chains. The long chains of H-HA provide stable interactions between them, which involve all molecules present in solution, which create a three-dimensional network, whereas L-HA molecules provide less stable interactions, leading to aggregated systems, which do not simultaneously All molecules present are involved, which replace the interactions in the cluster. This different way of aggregation of H-HA and L-HA in solution causes large differences in rheological properties, such as the viscosity of HA solutions, which is a very important property for many applications, especially in the field of medicine .
事实上，作为分子量函数的HA溶液的粘度快速下降实际上取决于分子间相互作用的这种不同能力，因此，在相同浓度下，分子量大于1·106Da的H-HA溶液比分子量在5·103至5·105Da之间的L-HA溶液具有更高数量级的粘度。由于H-HA长链之间相互作用的强的协同性，当L-HA溶于H-HA的粘稠溶液中时，短期内没有观察到产生的溶液的粘度的显著差异，这表明两种分子群体独立表现，并且L/H-HA协同杂化聚集体的形成是热力学不利的过程。只有经过一段时间(数天至数周)，才能观察到粘度缓慢但持续下降，这不能归因于水解过程，并且如果L-HA的分子量小于105Da，这些效应更明显。这些溶液理化性质(特别是粘度)的不断变化使得它们不适合实际应用，其需要不变的流变学特征。In fact, the rapid decrease in the viscosity of HA solution as a function of molecular weight actually depends on this different ability of intermolecular interaction, therefore, at the same concentration, the H-HA solution with molecular weight greater than 1·10 L-HA solutions between • 103 and 5 • 105 Da have a higher order of magnitude viscosity. Due to the strong synergistic nature of the interactions between the long chains of H-HA, when L-HA was dissolved in a viscous solution of H-HA, no significant difference in the viscosity of the resulting solution was observed in the short term, suggesting that the two Molecular populations behave independently, and the formation of L/H-HA cooperative hybrid aggregates is a thermodynamically unfavorable process. Only over a period of time (days to weeks) is a slow but continuous decrease in viscosity observed, which cannot be attributed to a hydrolysis process, and these effects are more pronounced if the molecular weight of the L-HA is less than 105 Da. The constant variation of the physicochemical properties of these solutions (especially viscosity) makes them unsuitable for practical applications, which require constant rheological characteristics.
因此，鉴于以上描述，很清楚目前还没有获得低分子量透明质酸和高分子量透明质酸之间的协同杂化复合物，所述的复合物具有使得它们能用于所需目的的性质。Therefore, in view of the above description, it is clear that no synergistic hybrid complexes between low molecular weight hyaluronic acid and high molecular weight hyaluronic acid have been obtained so far, said complexes possessing properties that would allow them to be used for the desired purpose.
现在，令人惊讶地发现通过将含有H-HA和L-HA的水溶液一起经过合适配置的热循环来产生稳定的L/H-HA协同杂化物是可能的。Now, it has been surprisingly found that it is possible to generate stable L/H-HA synergistic hybrids by subjecting aqueous solutions containing H-HA and L-HA together through suitably configured thermal cycling.
本发明的稳定的L/H-HA协同杂化物溶液的特征在于粘度不随时间变化并且显著低于热循环之前。The stable L/H-HA synergistic hybrid solutions of the present invention are characterized by a viscosity that does not change over time and is significantly lower than before thermal cycling.
这些性能不能仅仅归因于HA的热解聚过程。These properties cannot be solely attributed to the thermal depolymerization process of HA.
存在四个精密决定L/H-HA复合物形成和它们流变学性质的参数：There are four parameters that precisely determine the formation of L/H-HA complexes and their rheological properties:
a)在相同溶液中同时存在两种类型的HA(L-HA和H-HA)；a) There are two types of HA (L-HA and H-HA) in the same solution;
b)在形成L/H-HA杂化物体系的过程中所用的两种HA的分子量；b) the molecular weights of the two HAs used in the formation of the L/H-HA hybrid system;
c)所用的两种HA的相对比例；c) the relative proportions of the two HAs used;
d)溶液所暴露的热循环的分布。d) Distribution of thermal cycles to which the solution is exposed.
在相同溶液中同时存在两种类型HA(L-HA和H-HA)是一个必要条件，因为当进行热循环，溶液达到足够高温度时，产生的能量条件能同时破裂H-HA链间的所有相互作用以及L-HA链间的所有相互作用，并且在这些条件中不再存在必要条件，因为溶液中分子间产生的弱相互作用是协同型，并且聚合物链以独立的实体表现。接着，当溶液在热处理循环范围内冷却时，链间相互作用开始逐渐重新形成，在这种情况下其在溶液中存在的所有HA分子间(高分子量和低分子量)随机产生，引起杂化物体系，随着弱分子间键数量的增加，杂化物体系是稳定的，它们的协同性意味着不同分子量的聚合物链间产生的相互作用方式不随时间变化。确认这种机制有效性实际上是，当两种溶液(一种L-HA和一种H-HA)分别进行热循环，然后冷却后将它们以相同浓度在溶液中一起混合时，我们没有观察到由杂化物系形成引起的粘度的显著且快速降低，只有在热循环过程中同时存在两种分子时才能形成杂化物体系。The simultaneous presence of two types of HA (L-HA and H-HA) in the same solution is a necessary condition, because when thermal cycling is performed and the solution reaches a sufficiently high temperature, the energy conditions generated can simultaneously break the bonds between the H-HA chains. All interactions, and all interactions between L-HA chains, and in these conditions no longer exist as a necessary condition, because the weak interactions generated between molecules in solution are of the cooperative type, and the polymer chains behave as independent entities. Next, when the solution is cooled within the range of the heat treatment cycle, interchain interactions begin to gradually reform, in which case they are generated randomly among all HA molecules (both high and low molecular weight) present in the solution, giving rise to the hybrid system , as the number of weak intermolecular bonds increases, the hybrid system is stable, and their cooperativity means that the interaction mode between polymer chains of different molecular weights does not change with time. Confirming the validity of this mechanism is actually that when two solutions (one L-HA and one H-HA) were thermally cycled separately and then mixed together in solution at the same concentration after cooling, we did not observe Due to the dramatic and rapid decrease in viscosity caused by the formation of hybrid systems, hybrid systems can only be formed when both molecules are present simultaneously during thermal cycling.
在构建L/H-HA杂化物体系中所用的HA的分子量精密地决定了它们的流变学特性；所用的L-HA和H-HA之间分子量差异越大，相同浓度时，相比H-HA的粘度，杂化物体系的粘度降低越多。The molecular weight of HA used in the construction of L/H-HA hybrid systems precisely determines their rheological properties; the greater the difference in molecular weight between the used L-HA and H-HA, the greater the - the viscosity of the HA, the more the viscosity of the hybrid system decreases.
如果L-HA的分子量在1·104至1·106Da之间并且H-HA的分子量是以公式MWH-HA≥MWL-HA/0.9给定时，可以获得具有粘度降低特征的协同杂化L/H-HA复合物。If the molecular weight of L-HA is between 1·104 and1· 106 Da and the molecular weight of H-HA is given by the formula MWH-HA ≥ MWL-HA /0.9, a synergistic characteristic of viscosity reduction can be obtained Hybrid L/H-HA complex.
L-HA和H-HA的相对比例(决定杂化物的化学计量)相比它们之间杂化的种类引起它们流变学性质的修饰；由于复合物的形成引起的粘度降低随着所用的L-HA/H-HA化学计量比的增加而增加。通常所述的比值在0.1至10之间，优选0.5至2。The relative proportions of L-HA and H-HA (determining the stoichiometry of the hybrids) compared to the type of hybridization between them cause a modification of their rheological properties; the viscosity decrease due to complex formation increases with the amount of L-HA used HA/H-HA stoichiometric ratio increases. Usually said ratio is between 0.1 and 10, preferably 0.5 to 2.
本发明的复合物的粘度通常比单独含有用于形成复合物的H-HA透明质酸的溶液的粘度小1.1至200倍。The viscosity of the complex of the present invention is generally 1.1 to 200 times smaller than that of a solution containing H-HA hyaluronic acid used to form the complex alone.
引起由含有L-HA和H-HA的溶液开始形成协同杂化L/H-HA体系的热分布发现，首先将溶液加热至温度为80至160℃、优选100至120℃，然后快速冷却至室温。从而获得的L/H-HA杂化物体系随时间稳定，证明能维持它们的流变学特征。The thermal profile leading to the formation of a synergistic hybrid L/H-HA system starting from a solution containing L-HA and H-HA found that the solution was first heated to a temperature of 80 to 160°C, preferably 100 to 120°C, and then rapidly cooled to room temperature. The L/H-HA hybrid systems thus obtained are stable over time, demonstrating the ability to maintain their rheological characteristics.
如已经提及的，本发明的L/H-HA杂化复合物溶液可以易于通过将所需分子量的H-HA和L-HA水溶液混合并且将产生的溶液进行上面所示的热循环而获得，优选L-HA溶液的浓度为0.01至50％w/w，而H-HA溶液的浓度为0.01至10％w/w。As already mentioned, the L/H-HA hybrid complex solutions of the present invention can be readily obtained by mixing aqueous solutions of H-HA and L-HA of the desired molecular weight and subjecting the resulting solution to thermal cycling as indicated above , preferably the L-HA solution has a concentration of 0.01 to 50% w/w and the H-HA solution has a concentration of 0.01 to 10% w/w.
固态协同杂化L/H-HA复合物可以从含有它们的溶液以多种方式获得：Solid-state cooperative hybrid L/H-HA complexes can be obtained from solutions containing them in several ways:
a)沉淀含有它们的溶液，加入水可混溶的有机溶剂，例如低分子量醇、丙酮等；a) Precipitate the solution containing them, adding water-miscible organic solvents, such as low molecular weight alcohols, acetone, etc.;
b)溶剂蒸发；b) solvent evaporation;
c)喷雾干燥；c) spray drying;
d)冻干。d) lyophilization.
此外，类似于上述那些具有低值动态粘度特征的协同杂化复合物可以通过高温热处理含有低分子量多糖(例如软骨素、硫酸软骨素、糊精、环糊精、葡聚糖)的H-HA水溶液而获得。In addition, synergistic hybrid complexes similar to those above characterized by low values of dynamic viscosity can be obtained by high-temperature heat treatment of H-HA containing low-molecular-weight polysaccharides (e.g., chondroitin, chondroitin sulfate, dextrin, cyclodextrin, dextran) obtained from an aqueous solution.
由于它们流变学特性，协同杂化L/H-HA复合物在某些生物医学应用中具有很多益处，例如：通过皮内注射HA生物再生皮肤；用于消退与关节炎性障碍相关的病理学状况的粘度补充技术；膀胱内治疗膀胱炎；治疗阴道炎性疾病；治疗肺泡疾病；治疗口腔疾病。Due to their rheological properties, synergistic hybrid L/H-HA complexes have many benefits in certain biomedical applications, such as: bioregeneration of skin by intradermal injection of HA; for regression of diseases associated with arthritic disorders; Viscosity replenishment techniques for physiologic conditions; intravesical treatment of cystitis; treatment of vaginal inflammatory diseases; treatment of alveolar diseases; treatment of oral diseases.
与使用协同杂化L/H-HA复合物相关的最重要的优势是它们的低粘度，在医学实践中其允许使用更高浓度的溶液，但在用小口径针和导管注射或喷雾时仍能充分流动。The most important advantage associated with the use of synergistic hybrid L/H-HA complexes is their low viscosity, which allows the use of more concentrated solutions in medical practice, but remains stable when injected or sprayed with small-bore needles and catheters. able to flow adequately.
一旦接触生物环境，协同杂化L/H-HA复合物作为体系表现出缓慢释放L-HA和H-HA，因为微环境的化学复杂性(特征在于溶液中存在其它种类和细胞结构的巨大表面)容许逐渐分解分子间相互作用，其表现为所述的复合物使得本文从头开始可获得L-HA和H-HA，它们是在体内具有不同作用的分子种类，L-HA通过与细胞表面存在的受体相互作用传递信号，H-HA作为细胞外基质的重要组成。Upon exposure to the biological environment, the synergistic hybrid L/H-HA complex as a system exhibits slow release of L-HA and H-HA due to the chemical complexity of the microenvironment (characterized by the presence of other species and the large surface area of cellular structures in solution). ) allows a gradual breakdown of intermolecular interactions, which manifest themselves as complexes that make available here ab initio L-HA and H-HA, which are molecular species with different roles in the body, L-HA exists by interacting with the cell surface The receptor interaction transmits signals, and H-HA serves as an important component of the extracellular matrix.
下面给出了非限制性实施例，描述了协同杂化L/H-HA复合物的制备、特性和用途。Non-limiting examples are given below describing the preparation, properties and use of the synergistic hybrid L/H-HA complexes.
实施例1-在不同温度下制备协同杂化L/H-HA复合物Example 1 - Preparation of Synergistic Hybrid L/H-HA Complex at Different Temperatures
H-HA(MW 1.4·106Da；Mw/Mn 1.5)和L-HA(MW 3.3·104Da；Mw/Mn 1.8)的两种水溶液是以2％w/v在蒸馏水中制备的，其用于制备表1中给出的多种溶液。Two aqueous solutions of H-HA (MW 1.4·106 Da; Mw/Mn 1.5) and L-HA (MW 3.3·104 Da; Mw/Mn 1.8) were prepared at 2% w/v in distilled water, It was used to prepare various solutions given in Table 1.
将同时含有L-HA和H-HA的这些溶液在高压釜中进行热循环，其设定为：a)加热相在10分钟内由25℃升至最高温度；b)维持该温度特定一段时间(10分钟或40分钟)；c)在10分钟内冷却溶液至25℃。These solutions, containing both L-HA and H-HA, were thermally cycled in an autoclave with the settings: a) the heating phase was ramped from 25°C to a maximum temperature within 10 minutes; b) this temperature was maintained for a specified period of time (10 minutes or 40 minutes); c) Cool the solution to 25° C. within 10 minutes.
测定MW和多分散性指数Mw/Mn，使用装有多检测器的尺寸排阻色谱系统，所述的多检测器包括四桥粘度计、折射计、直角光散射检测器(RALS)和低角光散射检测器(LALS)，专利权归American group Viscotek(www.viscotek.com)所有。用LALS测量的信号与分子量和浓度成比例，用粘度检测器测量的信号与样品浓度和固有粘度成比例，而折射计提供浓度的测量。Viscotek装置不仅能测量HA的分子量，而且能评价存在的分子群体中分子量的不均一度，通过多分散性指数Mw/Mn描述，通过Viscotek装置自动计算，并且定义为平均分子量(Mw＝ΣimiMi/Σimi，其中mi是分子量Mi的聚合物的质量，并且Σimi是聚合物的总质量，设定mi＝niMi，表达式还可以为Mw＝ΣiniMi2/ΣiniMi)和重量-平均分子量(Mn＝ΣiniMi/Σini，其中niMi是分子量Mi的聚合物的质量，并且Σini是存在的聚合物的总摩尔数)的比。动态粘度η的测量是在Anton Paar Physica MCR 301流变仪上进行的，使用具有同轴圆筒的几何图。η是在25℃下以恒定剪切率(γ’＝2s-1)测定的，其在聚合物溶液的牛顿粘度范围内(相对于γ’，η是常数，并且仅取决于溶液中聚合物的构象)。The MW and polydispersity index Mw/Mn were determined using a size exclusion chromatography system equipped with multiple detectors including a four-bridge viscometer, a refractometer, a right-angle light scattering detector (RALS) and a low-angle Light Scattering Detector (LALS), patented by American group Viscotek (www.viscotek.com). The signal measured with LALS is proportional to molecular weight and concentration, the signal measured with viscometer is proportional to sample concentration and intrinsic viscosity, and the refractometer provides a measurement of concentration. The Viscotek device can not only measure the molecular weight of HA, but also evaluate the heterogeneity of the molecular weight in the existing molecular population, described by the polydispersity index Mw/Mn, automatically calculated by the Viscotek device, and defined as the average molecular weight (Mw=Σi mi Mi /Σi mi , wherein mi is the quality of the polymer of molecular weight Mi , and Σi mi is the total mass of the polymer, setting mi =ni Mi , the expression can also be Mw = Σi ni Mi2 /Σi ni Mi ) and weight-average molecular weight (Mn = Σi ni Mi /Σi ni , where ni Mi is the mass of the polymer with molecular weight Mi , and Σ ini is the ratio of the total moles of polymer present) . The measurement of dynamic viscosity η was performed on an Anton Paar Physica MCR 301 rheometer using a geometry with coaxial cylinders. η is measured at a constant shear rate (γ' = 2s-1 ) at 25°C in the range of Newtonian viscosities of polymer solutions (with respect to γ', η is a constant and depends only on the polymer in solution conformation).
表1Table 1
*2％w/v的两种溶液首先在高温下处理，然后以体积比1:1混合。*The two solutions at 2% w/v were first treated at high temperature and then mixed in a volume ratio of 1:1.
表1-测量浓度为1％w/v的L-HA(MW 3.3·104Da；Mw/Mn 1.8)和H-HA(MW 1.4·106Da；Mw/Mn 1.5)的溶液的动态粘度和浓度为1％w/v和L-HA/H-HA比例为1:1w/w的相应稳定的协同L/H-HA复合物的动态粘度。高压釜中热处理循环设定为加热相在10分钟内由25℃至Tmax，在Tmax维持特定时间，并且冷却相在10分钟内由Tmax至25℃。在热处理后立即测量η。Table 1 - Measurement of dynamic viscosities of solutions of L-HA (MW 3.3·104 Da; Mw/Mn 1.8) and H-HA (MW 1.4·106 Da; Mw/Mn 1.5) at a concentration of 1% w/v and the dynamic viscosities of the corresponding stable synergistic L/H-HA complexes at a concentration of 1% w/v and an L-HA/H-HA ratio of 1:1 w/w. The heat treatment cycle in the autoclave was set as a heating phase from 25°C toTmax within 10 minutes, maintained atTmax for a specified time, and a cooling phase fromTmax to 25°C within 10 minutes. η was measured immediately after heat treatment.
表1中的数据表明：a)在所述的条件下加热H-HA溶液导致长聚合物链的轻微水解(120℃，10分钟，MW 9.51·105Da；110℃，10分钟，MW1.04·106Da；100℃，10分钟，MW 1.20·106Da；100℃，40分钟，MW 9.10·105Da)并且η成比例降低；b)在所述的条件下加热L-HA溶液导致聚合物链的轻微水解(120℃，10分钟，MW 2.96·104Da；110℃，10分钟，MW 3.12·104Da；100℃，10分钟，MW 3.25·104Da；100℃，40分钟，MW 2.88·104Da)并且η成比例降低，加热后其不再可测；c)在L-HA和H-HA溶液中简单混合，混合后立即导致η的轻微降低，因为对于非常低分子量(3.3·104Da)的L-HA，甚至在室温下，基于短链和长链之间的氢键的部分相互作用开始活化；d)加热同时含有H-HA和L-HA的溶液导致氢键的定量破裂，导致长链间和短链间先前存在的协同性条件的丧失；e)在随后的冷却相中，如果溶液中存在短链和长链，在短链和长链间能随机恢复具有氢桥的协同相互作用，引起通过协同相互作用稳定的杂化物体系；f)分别加热H-HA和L-HA溶液，冷却相后它们的混合不能形成协同杂化物，但性能类似于字母c)下描述的性能；g)没有与其它分子或表面的相互作用的L/H-HA杂化物在室温下保持稳定，因为即使氢键随机打开，随链存在的多重此类相互作用也能维持结构元件在适合的距离内引起键合用于重新形成；h)H-HA+L-HA混合物经历的温度越高或暴露的时间越长，协同杂化物形成越有效。The data in Table 1 show that: a) Heating the H-HA solution under the conditions described resulted in slight hydrolysis of the long polymer chains (120 °C,10 min, MW 9.51·105 Da; 110 °C, 10 min, MW 1. 04·106 Da; 100°C, 10 min, MW 1.20·106 Da; 100°C, 40 min, MW 9.10·105 Da) and η decreases proportionally; b) heating L-HA under the conditions described The solution causes slight hydrolysis of the polymer chains (120°C,10 min, MW 2.96·104 Da; 110°C,10 min, MW 3.12·104 Da; 100°C, 10 min, MW 3.25·104 Da; 100°C , 40 min, MW 2.88·104 Da) and η decreases proportionally, it is no longer measurable after heating; c) simple mixing in L-HA and H-HA solutions immediately after mixing results in a slight decrease in η because For very low molecular weight (3.3·104 Da) L-HA, even at room temperature, partial interactions based on hydrogen bonds between short and long chains start to activate; d) heating containing both H-HA and L- A solution of HA leads to a quantitative breakdown of hydrogen bonds, leading to the loss of the pre-existing cooperativity conditions between long and short chains; e) in the subsequent cooling phase, if short and long chains are present in the solution, between short and short chains The synergistic interactions with hydrogen bridges can be randomly restored between long chains, resulting in a hybrid system that is stable through synergistic interactions; f) H-HA and L-HA solutions are heated separately, and their mixing after cooling phases cannot form synergistic hybrids, But the properties are similar to those described under letter c); g) L/H-HA hybrids without interactions with other molecules or surfaces remain stable at room temperature because multiple Class interactions can also maintain structural elements within a suitable distance to cause bonding for reformation; h) The higher the temperature experienced or the longer the exposure time of the H-HA+L-HA mixture, the more efficient the cooperative hybrid formation.
实施例2-用不同组合物制备协同杂化L/H-HA复合物Example 2 - Preparation of Synergistic Hybrid L/H-HA Complexes with Different Compositions
不同组合物的协同杂化L/H-HA复合物是通过将H-HA(MW 1.4·106Da；Mw/Mn 1.5)和L-HA(MW 3.3·104Da；Mw/Mn 1.8)溶于100mL水中制备的，如表2所示。将产生的溶液在高压釜中进行下列热循环：10分钟内从25℃至120℃，120℃10分钟，10分钟内从120℃至25℃。如实施例1所描述的那样测定样品的动态粘度，L-HA、H-HA和L/H-HA的MW和多分散性指数Mw/Mn。表2中的数据表明L/H-HA协同复合物的粘度取决于L-HA/H-HA比例：比例越高，粘度越低。Synergistic hybrid L/H-HA complexes of different compositions were obtained by combining H-HA (MW 1.4·106 Da; Mw/Mn 1.5) and L-HA (MW 3.3·104 Da; Mw/Mn 1.8) Prepared by dissolving in 100mL water, as shown in Table 2. The resulting solution was subjected to the following thermal cycles in an autoclave: 25°C to 120°C in 10 minutes, 120°C for 10 minutes, 120°C to 25°C in 10 minutes. The dynamic viscosity, MW and polydispersity index Mw/Mn of the L-HA, H-HA and L/H-HA of the samples were determined as described in Example 1. The data in Table 2 show that the viscosity of the L/H-HA synergistic complex depends on the L-HA/H-HA ratio: the higher the ratio, the lower the viscosity.
表2Table 2
*混合后立即测量η。*Measurement of η immediately after mixing.
表2-测量不同L-HA/H-HA比例的协同杂化L/H-HA复合物的动态粘度浓度η。H-HA(MW 1.4·106Da；Mw/Mn 1.5)的浓度维持在1％w/v不变，而L-HA(MW 3.3·104Da；Mw/Mn1.8)的浓度从0变化至1.5w/v。高压釜中热处理循环设定为10分钟加热相由25℃至Tmax，在Tmax维持特定时间，并且冷却相在10分钟内由Tmax至25℃。在热处理后立即测量η。Table 2 - Measurement of dynamic viscosity concentration η of synergistic hybrid L/H-HA complexes with different L-HA/H-HA ratios. The concentration of H-HA (MW 1.4·106 Da; Mw/Mn 1.5) was maintained at 1% w/v, while the concentration of L-HA (MW 3.3·104 Da; Mw/Mn 1.8) was changed from 0 Change to 1.5w/v. The heat treatment cycle in the autoclave was set as a heating phase from 25°C toTmax in 10 minutes, a hold atTmax for a specified time, and a cooling phase fromTmax to 25°C in 10 minutes. η was measured immediately after heat treatment.
实施例3-使用不同分子量的L-HA制备协同杂化L/H-HA复合物Example 3 - Preparation of Synergistic Hybrid L/H-HA Complex Using L-HA of Different Molecular Weights
H-HA(MW 1.4·106Da；Mw/Mn 1.5)、L-HA(MW 3.3·104Da；Mw/Mn 1.8)和L-HA(MW2.2·105Da；Mw/Mn 1.7)的水溶液是以2％w/v在蒸馏水中制备的，所述的溶液用于制备表3中给出的多种溶液。将产生的溶液在高压釜中进行下列热循环：10分钟内从25℃至120℃，120℃10分钟，10分钟内从120℃至25℃。如实施例1所描述的那样测定样品的动态粘度η，L-HA、H-HA和L/H-HA的MW和多分散性指数Mw/Mn。H-HA (MW 1.4·106 Da; Mw/Mn 1.5), L-HA (MW 3.3·104 Da; Mw/Mn 1.8) and L-HA (MW2.2·105 Da; Mw/Mn 1.7 ) in water was prepared at 2% w/v in distilled water and used to prepare various solutions given in Table 3. The resulting solution was subjected to the following thermal cycles in an autoclave: 25°C to 120°C in 10 minutes, 120°C for 10 minutes, 120°C to 25°C in 10 minutes. The dynamic viscosity η of the samples, the MW and the polydispersity index Mw/Mn of L-HA, H-HA and L/H-HA were determined as described in Example 1.
表3中的数据表明协同杂化物中L-HA的MW越低，η降低得越多，所有其它参数是相同的。比较使用MW 3.3·104Da或MW 2.20·105Da的L-HA的L/H-HA杂化复合物的η值，ηH-HA/ηL/H-HA的比值增加约50倍。The data in Table 3 indicate that the lower the MW of the L-HA in the synergistic hybrids, the more η decreases, all other parameters being equal. Comparing the η values of L/H-HA hybrid complexes using L-HA of MW 3.3·104 Da or MW 2.20·105 Da, the ratio of ηH-HA /ηL/H-HA increased about 50 times .
表3table 3
表3-测量由不同MW的L-HA构建的且L-HA/H-HA比例为1w/w的协同杂化L/H-HA复合物的动态粘度η。H-HA(MW 1.4·106Da；Mw/Mn 1.5)、L-HA(MW 3.3·104Da；Mw/Mn 1.8)和L-HA(MW 2.2·105Da；Mw/Mn 1.7)的水溶液是以2％w/v在蒸馏水中制备的，所述的溶液用于制备表中给出的多种溶液。将产生的溶液在高压釜中进行下列热循环：10分钟内从25℃至120℃，120℃10分钟，10分钟内从120℃至25℃。Table 3 - Measurement of dynamic viscosity η of synergistic hybrid L/H-HA complexes constructed from L-HA of different MW and L-HA/H-HA ratio 1 w/w. H-HA (MW 1.4·106 Da; Mw/Mn 1.5), L-HA (MW 3.3·104 Da; Mw/Mn 1.8) and L-HA (MW 2.2·105 Da; Mw/Mn 1.7) Aqueous solutions of ® were prepared at 2% w/v in distilled water, and said solutions were used to prepare the various solutions given in the table. The resulting solution was subjected to the following thermal cycles in an autoclave: 25°C to 120°C in 10 minutes, 120°C for 10 minutes, 120°C to 25°C in 10 minutes.
实施例4-经历和未经历热循环形成的协同杂化L/H-HA复合物的动力学分析Example 4 - Kinetic analysis of cooperative hybrid L/H-HA complexes formed with and without thermal cycling
H-HA(MW 1.4·106Da；Mw/Mn 1.5)、L-HA(MW 3.3·104Da；Mw/Mn 1.8)和L-HA(MW2.2·105Da；Mw/Mn 1.7)的水溶液是以2％w/v在蒸馏水中制备的，所述的溶液用于制备表4中给出的多种溶液。将一半产生的溶液维持在该温度，并且另一半先在高压釜中进行下列热循环：10分钟内从25℃至120℃，120℃10分钟，10分钟内从120℃至25℃，然后保持在室温。随时间测量两个系列样品的动态粘度η。如实施例1所描述的那样测定L-HA、H-HA和L/H-HA的MW、多分散性指数Mw/Mn和样品的动态粘度η。H-HA (MW 1.4·106 Da; Mw/Mn 1.5), L-HA (MW 3.3·104 Da; Mw/Mn 1.8) and L-HA (MW2.2·105 Da; Mw/Mn 1.7 ) in water was prepared at 2% w/v in distilled water and used to prepare various solutions given in Table 4. Half of the resulting solution was maintained at this temperature, and the other half was first subjected to the following thermal cycles in the autoclave: 25°C to 120°C in 10 minutes, 120°C for 10 minutes, 120°C to 25°C in 10 minutes, then hold At room temperature. The dynamic viscosity η of two series of samples was measured over time. The MW, the polydispersity index Mw/Mn and the dynamic viscosity η of the samples were determined as described in Example 1 for L-HA, H-HA and L/H-HA.
表4的数据表明：a)当L-HA的MW是104Da数量级时，开始形成L/H-HA协同复合物，即使是缓慢的，甚至在室温，因为L-HA短链间存在的较低的协同性使得这些链竞争H-HA长链间存在的协同相互作用，从而引起形成杂化物体系；b)对于这个原因，在室温下通过混合H-HA和MW为104Da数量级的L-HA获得的溶液表现出动态粘度随时间而变化；c)相反，热处理在数分钟内产生协同杂化复合物，一旦它们达到平衡条件，所述的复合物的动态粘度不随时间而变化；d)当L-HA的MW是105Da数量级时，没有热处理，简单混合两种溶液不会随时间而显著改变它们的动态粘度，因为L-HA链间预先存在很强的协同性，其阻止它们与H-HA链相互作用。The data in Table 4 show that: a) When the MW of L-HA is on the order of 104 Da, the L/H-HA synergistic complex starts to form, even slowly, even at room temperature, because of the The lower cooperativity allows these chains to compete for the cooperative interactions that exist between the long chains of H-HA, thereby causing the formation of hybrid systems; b) for this reason, at room temperature by mixing H-HA and MW of the order of 104 Da The solutions obtained with L-HA exhibit a dynamic viscosity change with time; c) in contrast, heat treatment produces synergistic hybrid complexes within minutes, said complexes having a dynamic viscosity that does not change with time once they have reached equilibrium conditions; d) When the MW of L-HA is on the order of 105 Da, without heat treatment, simply mixing the two solutions will not significantly change their dynamic viscosities over time, because there is a strong cooperativity between the L-HA chains, which prevent them from interacting with the H-HA chains.
表4Table 4
表4-不同MW的L-HA构建的且L-HA/H-HA比例为1w/w的协同杂化L/H-HA复合物的动态粘度η动力学。H-HA(MW 1.4·106Da；Mw/Mn 1.5)、L-HA(MW 3.3·104Da；Mw/Mn 1.8)和L-HA(MW 2.2·105Da；Mw/Mn 1.7)的水溶液是以2％w/v在蒸馏水中制备的，所述的溶液用于制备表中给出的多种溶液。将一半产生的溶液维持在该温度，并且另一半先在高压釜中进行下列热循环：10分钟内从25℃至120℃，120℃10分钟，10分钟内从120℃至25℃，然后保持在室温。Table 4 - Dynamic viscosity η kinetics of synergistic hybrid L/H-HA complexes constructed with L-HA of different MW and L-HA/H-HA ratio 1 w/w. H-HA (MW 1.4·106 Da; Mw/Mn 1.5), L-HA (MW 3.3·104 Da; Mw/Mn 1.8) and L-HA (MW 2.2·105 Da; Mw/Mn 1.7) Aqueous solutions of ® were prepared at 2% w/v in distilled water, and said solutions were used to prepare the various solutions given in the table. Half of the resulting solution was maintained at this temperature, and the other half was first subjected to the following thermal cycles in the autoclave: 25°C to 120°C in 10 minutes, 120°C for 10 minutes, 120°C to 25°C in 10 minutes, then hold At room temperature.
实施例5-通过从包含它们的溶液中沉淀来制备固态协同杂化L/H-HA复合物Example 5 - Preparation of solid-state cooperative hybrid L/H-HA complexes by precipitation from solutions containing them
将如实施例1所描述的那样获得的协同杂化L/H-HA复合物的水溶液(热循环设定为暴露于Tmax为120℃10分钟)用2倍体积的无水乙醇处理，缓慢加入并且搅拌。获得白色粉状沉淀，其快速沉淀，并且可以在真空下加热干燥。该处理导致形成白色干燥的粉末，相对于理论值，产率为99％。如果以浓度为1％w/w溶于水中，粉末状的协同杂化L/H-HA复合物得到具有与最初沉淀的溶液相同动态粘度η值的溶液。The aqueous solution of the synergistic hybrid L/H-HA complex obtained as described in Example 1 (the thermal cycle was set to expose to aTmax of 120 °C for 10 min) was treated with 2 times the volume of absolute ethanol, slowly Add and stir. A white powdery precipitate is obtained which precipitates rapidly and can be dried with heat under vacuum. The treatment resulted in the formation of a white dry powder in a yield of 99% relative to theory. If dissolved in water at a concentration of 1% w/w, the powdered synergistic hybrid L/H-HA complex gave a solution with the same dynamic viscosity η value as the initially precipitated solution.
实施例6-通过冻干包含它们的溶液来制备固态协同杂化L/H-HA复合物Example 6 - Preparation of solid-state cooperative hybrid L/H-HA complexes by lyophilization of solutions containing them
将如实施例1所描述的那样获得的协同杂化L/H-HA复合物的水溶液(热循环设定为暴露于Tmax为120℃10分钟)冻干。获得海绵状团块，通过机械处理其易于转变成白色粉末。冻干的粉末的产率与理论值相同。如果以浓度为1％w/w溶于水中，粉末状冻干的协同杂化L/H-HA复合物得到具有与最初沉淀的溶液相同动态粘度η值的溶液。An aqueous solution of the synergistic hybrid L/H-HA complex obtained as described in Example 1 (thermal cycle set to expose to aTmax of 120° C. for 10 minutes) was lyophilized. A spongy mass was obtained which was easily transformed into a white powder by mechanical treatment. The yield of the lyophilized powder was the same as theoretical. If dissolved in water at a concentration of 1% w/w, the powdered lyophilized synergistic hybrid L/H-HA complex gave a solution with the same dynamic viscosity η value as the initially precipitated solution.
实施例7-制备协同杂化C/H-HA和CS/H-HA复合物Example 7 - Preparation of Cooperative Hybrid C/H-HA and CS/H-HA Complexes
H-HA(MW 1.4·106Da；Mw/Mn 1.5)、软骨素(C；MW 6.6·104Da；Mw/Mn 1.4)和硫酸软骨素(CS；MW 3.8·104Da；Mw/Mn 1.4)的水溶液是以2％w/v在蒸馏水中制备的，所述的溶液用于制备表5中给出的多种溶液。将产生的溶液在高压釜中进行下列热循环：10分钟内从25℃至120℃，120℃10分钟，10分钟内从120℃至25℃。如实施例1所描述的那样测定样品的动态粘度η，L-HA、H-HA和L/H-HA的MW和多分散性指数Mw/Mn。H-HA (MW 1.4·106 Da; Mw/Mn 1.5), chondroitin (C; MW 6.6·104 Da; Mw/Mn 1.4) and chondroitin sulfate (CS; MW 3.8·104 Da; Mw/ Aqueous solutions of Mn 1.4) were prepared at 2% w/v in distilled water and used to prepare the various solutions given in Table 5. The resulting solution was subjected to the following thermal cycles in an autoclave: 25°C to 120°C in 10 minutes, 120°C for 10 minutes, 120°C to 25°C in 10 minutes. The dynamic viscosity η of the samples, the MW and the polydispersity index Mw/Mn of L-HA, H-HA and L/H-HA were determined as described in Example 1.
表5的数据表明：a)MW数量级为104Da的软骨素和硫酸软骨素通过热处理引起形成稳定的协同杂化C/H-HA和CS/H-HA复合物，其特征在于低动态粘度η值；b)C和CS溶液与H-HA溶液简单混合在动态粘度η中没有产生显著变化；c)C/H-HA复合物的ηpre/ηpost比值约为CS/H-HA复合物的两倍。The data in Table 5 show that: a) chondroitin and chondroitin sulfate with MW on the order of 104 Da induced by heat treatment form stable synergistic hybrid C/H-HA and CS/H-HA complexes characterized by low dynamic viscosities η values; b) simple mixing of C and CS solutions with H-HA solutions produced no significant change in dynamic viscosity η; c) ηpre /ηpost ratios for C/H-HA complexes were approximately twice as much.
表5table 5
表5-测量协同杂化C/H-HA和CS/H-HA复合物的动态粘度η。H-HA(MW 1.4·106Da；Mw/Mn 1.5)、C(MW 6.6·104Da；Mw/Mn 1.4)和CS(MW 3.8·104Da；Mw/Mn 1.4)的水溶液是以2％w/v在蒸馏水中制备的，所述的溶液用于制备表中给出的多种溶液。将产生的溶液在高压釜中进行下列热循环：10分钟内从25至120℃，120℃10分钟，10分钟内从120至25℃。Table 5 - Measurement of dynamic viscosity η of synergistic hybrid C/H-HA and CS/H-HA composites. Aqueous solutions of H-HA (MW 1.4·106 Da; Mw/Mn 1.5), C (MW 6.6·104 Da; Mw/Mn 1.4) and CS (MW 3.8·104 Da; Mw/Mn 1.4) were obtained by 2% w/v in distilled water was used to prepare the various solutions given in the table. The resulting solution was subjected to the following thermal cycles in an autoclave: 25 to 120°C in 10 minutes, 120°C for 10 minutes, 120 to 25°C in 10 minutes.
实施例8-协同杂化L/H-HA复合物在生物再生领域中的用途Example 8 - Use of Synergistic Hybrid L/H-HA Complex in the Field of Biological Regeneration
将如实施例1所描述的那样获得的4g协同杂化复合物溶于100mL盐水中，在120℃下加热10分钟，然后如实施例5所描述的通过冻干干燥复合物。所用的透明质酸是注射用药物级的，并且所有操作是在保证无菌和无致热原溶液的条件下进行的。将含有40mg/mL L/H-HA复合物的溶液引入安装30号针的1mL注射器中。面部生物再生治疗是在10名被告知的志愿者上进行，所述的志愿者面部存在明显的皮肤老化体征。实验设计设定为每位个体进行相同的生物再生治疗，在面部右侧皮下微注射本发明的制剂(1mL)并且在左侧注射已经上市的早期产品(1mL)。仪器客观化获得的结果表明稳定的协同L/H-HA复合物治疗在治疗的质量和持续时间上均有优势。4 g of the synergistic hybrid complex obtained as described in Example 1 were dissolved in 100 mL of saline, heated at 120° C. for 10 minutes, and then the complex was dried by lyophilization as described in Example 5. The hyaluronic acid used is of pharmaceutical grade for injection, and all operations are carried out under conditions that ensure sterile and pyrogen-free solutions. Introduce the solution containing 40 mg/mL L/H-HA complex into a 1 mL syringe fitted with a 30-gauge needle. The facial biorejuvenation treatment was performed on 10 informed volunteers with visible signs of skin aging on the face. The experimental design was set such that each individual underwent the same bioregenerative treatment, microinjected the formulation of the present invention (1 mL) subcutaneously on the right side of the face and injected an earlier product already on the market (1 mL) on the left side. The results obtained with instrumental objectification showed the advantages of stable synergistic L/H-HA complex therapy both in quality and duration of treatment.
实施例9-协同杂化L/H-HA复合物在粘度补充领域中的用途Example 9 - Use of a synergistic hybrid L/H-HA complex in the field of viscosity supplementation
将如实施例1所描述的那样获得的4g协同杂化复合物溶于100mL盐水中，在120℃下加热10分钟，然后如实施例5所描述的通过冻干干燥复合物。所用的透明质酸是注射用药物级的，并且所有操作是在保证无菌和无致热原溶液的条件下进行的。将含有40mg/mL L/H-HA复合物的溶液引入安装30号针的1mL注射器中。粘度补充治疗是在5名被告知的志愿者上进行的，所述的志愿者患有双侧膝盖障碍，治疗适应证是透明质酸渗入关节中。实验设计设定为每位个体接受相同的粘度补充治疗，在右侧关节用本发明的制剂(1mL)并且在左侧关节用已经上市的早期产品(1mL)。仪器客观化获得的结果表明稳定的协同L/H-HA复合物治疗在疼痛快速减少和病理学病症消退功效上均有优势。4 g of the synergistic hybrid complex obtained as described in Example 1 were dissolved in 100 mL of saline, heated at 120° C. for 10 minutes, and then the complex was dried by lyophilization as described in Example 5. The hyaluronic acid used is of pharmaceutical grade for injection, and all operations are carried out under conditions that ensure sterile and pyrogen-free solutions. Introduce the solution containing 40 mg/mL L/H-HA complex into a 1 mL syringe fitted with a 30-gauge needle. Viscosupplementation treatment was carried out on 5 informed volunteers with bilateral knee disturbances, the indication for treatment being the infiltration of hyaluronic acid into the joints. The experimental design was set up so that each individual received the same viscosity supplementation treatment with the formulation of the invention (1 mL) in the right joint and with the already marketed earlier product (1 mL) in the left joint. The results obtained by instrumental objectification show the advantages of stable synergistic L/H-HA complex therapy in terms of rapid pain reduction and resolution of pathological conditions.
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| ITMI2010A001635AIT1402382B1 (en) | 2010-09-09 | 2010-09-09 | HYBRID COOPERATIVE COMPLEX HYALURONIC ACID |
| ITMI2010A001635 | 2010-09-09 | ||
| PCT/EP2011/065633WO2012032151A2 (en) | 2010-09-09 | 2011-09-09 | Hybrid cooperative complexes of hyaluronic acid |
| Publication Number | Publication Date |
|---|---|
| CN103119068A CN103119068A (en) | 2013-05-22 |
| CN103119068Btrue CN103119068B (en) | 2016-11-30 |
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5442053A (en)* | 1982-09-28 | 1995-08-15 | Fidia, S.P.A. | Salts and mixtures of hyaluronic acid with pharmaceutically active substances, pharmaceutical compositions containing the same and methods for administration of such compositions |
| CN1585645A (en)* | 2001-11-13 | 2005-02-23 | 艾尔科公司 | Use of a mixture of sodium hyaluronate and chondroitin sulfate for the treatment of osteoarthritis |
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5442053A (en)* | 1982-09-28 | 1995-08-15 | Fidia, S.P.A. | Salts and mixtures of hyaluronic acid with pharmaceutically active substances, pharmaceutical compositions containing the same and methods for administration of such compositions |
| CN1585645A (en)* | 2001-11-13 | 2005-02-23 | 艾尔科公司 | Use of a mixture of sodium hyaluronate and chondroitin sulfate for the treatment of osteoarthritis |
| Publication | Publication Date | Title |
|---|---|---|
| CA2810742C (en) | Hybrid cooperative complexes of hyaluronic acid | |
| Deng et al. | Enhanced gelation of chitosan/β-sodium glycerophosphate thermosensitive hydrogel with sodium bicarbonate and biocompatibility evaluated | |
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