Objective  To compare the advantages and disadvantages of three methods for determining the molecular weight of sodium hyaluronate in sodium hyaluronate eye drops: viscosity method, size exclusion chromatography-refractive index detector (SEC-RI), and size exclusion chromatography-refractive index detector-multi-angle laser light scattering (SEC-RI-MALLS).

Methods The weight-average molecular weight (Mw) of sodium hyaluronate in sodium hyaluronate eye drops was determined using these three methods, and the molecular weight distribution [Mw/number-average molecular weight (Mn)] was determined using SEC-RI and SEC-RI-MALLS methods.

Results  The Mw of three batches of purified sodium hyaluronate ophthalmic solution determined by SEC-RI method, viscosity method and SEC-RI-MALLS method were 365,359-367,508, 918,593-938,397 and 987,300-1,003,000 g/mol, respectively.

Conclusion  The Mw of sodium hyaluronate determined by SEC-RI method is relatively small, while the results obtained by viscosity method and SEC-RI-MALLS method are closer. The SEC-RI-MALLS method does not rely on reference standards, its operation is simple and fast, and it can also reflect the parameters such as Mw and Mw/Mn of sodium hyaluronate, which is suitable for the detection of sodium hyaluronate in sodium hyaluronate eye drops.

1.Gomes JAP, Santo RM. The impact of dry eye disease treatment on patient satisfaction and quality of life: a review[J]. Ocul Surf, 2019, 17(1): 9-19. DOI: 10.1016/j.jtos.2018.11.003.

2.Pinto-Fraga J, de la Rosa AL, Arauzo FB, et al. Efficacy and safety of 0.2% hyaluronic acid in the management of dry eye disease[J]. Eye Contact Lens, 2017, 43(1): 57-63. DOI: 10.1097/ICL.0000000000000236.

3.Becker LC, Bergfeld WF, Belsito DV, et al. Final report of the safety assessment of hyaluronic acid, potassium hyaluronate, and sodium hyaluronate[J]. Int J Toxicol, 2009, 28 (4 Suppl): 5-67. DOI: 10.1177/1091581809337738.

4.Cyphert JM, Trempus CS, Garantziotis S. Size matters: molecular weight specificity of hyaluronan effects in cell biology[J]. Int J Cell Biol, 2015, 2015: 563818. DOI: 10.1155/2015/563818.

5.Marcellin E, Steen JA, Nielsen LK. Insight into hyaluronic acidmolecular weight control[J]. Appl Microbiol Biotechnol, 2014, 98(16): 6947-6956. DOI: 10.1007/s00253-014-5853-x.

6.JP18-XVIII[S]. 2021: 1710-1711.

7.EP11.0[S]. 2023: 3993-3994.

8.国家食品药品监督管理局. 国家药品标准WS1-(X072)2011Z 玻璃酸钠[S]. 2011.

9.袁俊杰, 袁华. 黏度法测聚合物分子量实验的一些细节问题[J]. 当代化工, 2024, 53(10): 2505-2510. [Yuan JJ, Yuan  H. Some detailed issues in the experiment of measuring polymer molecular weight by viscosity method[J]. Contemporary Chemical Industry, 2024, 53(10): 2505-2510.] DOI: 10.13840/j.cnki.cn21-1457/tq.2024.10.024.

10.宋奇奇, 刘颖, 原琳, 等. 黏度法测定聚合物相对分子质量的改进[J]. 山东化工, 2019, 48(7): 160-161, 163. [Song QQ, Liu Y, Yuan L, et al. Improvement of determination of polymer molecular weight by viscosity method[J]. Shandong Chemical Industry, 2019, 48(7): 160-161, 163.] DOI: 10.19319/j.cnki.issn.1008-021x.2019.07.068.

11.中国药典2020年版. 二部[S]. 2020: 201-203.

12.邹剑, 杨蕾, 林涛, 等. SEC-RI-MALLS测定甘露聚糖肽分子量与分子量分布[J]. 药物分析杂志, 2024, 44(2): 256-263. [Zou J, Yang L, Lin T, et al. Determination of molecular weight and distribution of mannatide by means of SEC-RI-MALLS[J]. Chinese Journal of Pharmaceutical Analysis, 2024, 44(2): 256-263.] DOI: 10.16155/j.0254-1793.2024.02.08.

13.刘菁, 吴燕, 李华龙, 等. SEC-RI-MALLS联用技术测定甲基纤维素的分子量及分子量分布[J]. 药物分析杂志, 2019, 39(6): 1058-1062. [Liu J, Wu Y, Li HL, et al. Establishment of determination method for the molecular weight and molecular weight distribution of methyl cellulose by SEC-RI-MALLS[J]. Chinese Journal of Pharmaceutical Analysis, 2019, 39(6): 1058-1062.] DOI: 10.16155/j.0254-1793.2019.06.13.

14.中国药典检验标准操作规范2019年版[S]. 2010: 114.

15.中国药典2020年版. 四部[S]. 2020: 480.

16.何素婷, 张卫东, 吕晶, 等. 3种方法测定玻璃酸钠的分子量比较[J]. 中国药师, 2015, 18(6): 929-931. [He ST, Zhang WD, Lyu J, et al. Comparison of three methods for determining molecular weight of sodium hyaluronate[J]. Chinese pharmacist, 2015, 18(6): 929-931.] DOI: 10.3969/j.issn.1008-049X.2015.06.015.

17.宋玉娟, 李颖颖, 范慧红. 玻璃酸钠及制剂分子质量与分子质量分布测定方法研究[J]. 药物分析杂志, 2014, 34(8): 1444-1448. [Song YJ, Li YY, Fan HH. Determination of molecular weight and molecular weight distribution of sodium hyaluronate and its preparations[J]. Chinese Journal of Pharmaceutical Analysis, 2014, 34(8): 1444-1448.] DOI: 10.16155/j.0254-1793.2014.08.033.

18.程广文, 范晓东, 周志勇, 等. 凝胶渗透色谱-多角度激光散射测定聚乙二醇分子量[J]. 高分子材料科学与工程, 2008, 24(10): 127-130. [Cheng GW, Fan XD, Zhou ZY, et al. Determination of molecular weight of polyether glycol by size exclusion chromatography-multi angle laser light scattering[J]. Polymer Materials Science & Engineering, 2008, 24(10): 127-130.] DOI: 10.16865/j.cnki.1000-7555.2008.10.033.

19.王悦, 宋玉娟, 刘倩, 等. 国产羟乙基淀粉130/0.4注射液质量评价及质量研究[J]. 中国药物警戒, 2022, 19(10): 1067-1072, 1090. [Wang Y, Song YJ, Liu Q, et al. Quality evaluation and quality research of domestic hydroxyethyl starch 130/0.4 injection[J]. Chinese Journal of Pharmacovigilance, 2022, 19(10): 1067-1072, 1090.] DOI: 10.19803/j.1672-8629.20220224.

20.Becker A, Köhler W, Müller B. A scanning michelson interferometer for the measurement of the concentration and temperature derivative of the refractive index of liquids[J]. Ber Bunsenges Phys Chem, 1995, 99: 600-608. DOI: 10.1002/bbpc.19950990403.

21.EP11.0 [S]. 2022: 1017.

22.WHO Expert Committee on Biological Standardization. Recommendations to assure the quality, safety and efficacy of group Amenin gococcal conjugate vaccines[S]. 2011, 962: 115-171.