Abstract
Hyaluronic acid and its acrylate derivatives are important intermediates for various pharmaceutical, biomedical, and cosmetic applications due to their biocompatibility and viscoelasticity properties. However, these polymers are inherently difficult to characterize due to their significant heterogeneity regarding molar mass and chemical composition (degree of substitution, DS). The present study describes the development of a comprehensive online two-dimensional liquid chromatography (2D-LC) approach to characterize hyaluronic acid and its acrylate derivatives (DS ranging from 0.4 to 3.1) in terms of molar mass and degree of substitution. In the first dimension of the 2D-LC method, separation according to chemical composition/DS was achieved by using a stepwise solvent gradient and a reversed phase C8 column. Fractions from the first dimension were automatically transferred to the second dimension comprising size exclusion chromatographic separation of the fractions according to molar mass. It was found that the hyaluronic acid derivatives were broadly distributed with regard to both chemical composition and molar mass. Fractions with different degrees of substitution were identified, and their molar mass distributions were determined. The study proved that comprehensive 2D-LC is a powerful approach to reveal the complex nature of hyaluronic acid and its derivatives.
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Liu L, Liu Y, Li J, Du G, Chen J. Microbial production of hyaluronic acid: current state, challenges, and perspectives. Microb Cell Factories. 2011;10:99–108.
Chong BF, Blank LM, Mclaughlin R, Nielsen LK. Microbial hyaluronic acid production. Appl Microbiol Biotechnol. 2005;66:341–51.
Necas J, Bartosikova L, Brauner P, Kolar J. Hyaluronic acid (hyaluronan): a review. Vet Med (Praha). 2008;53:397–411.
Sze JH, Brownlie JC, Love CA. Biotechnological production of hyaluronic acid: a mini review. 3 Biotech. 2016;6:1–9.
Burdick JA, Prestwich GD. Hyaluronic acid hydrogels for biomedical applications. Adv Mater. 2011;23:41–56.
Schanté CE, Zuber G, Herlin C, Vandamme TF. Chemical modifications of hyaluronic acid for the synthesis of derivatives for a broad range of biomedical applications. Carbohydr Polym. 2011;85:469–89.
Liu J, Willför S, Xu C. A review of bioactive plant polysaccharides: biological activities, functionalization, and biomedical applications. Bioact Carbohydrates Diet Fibre. 2015;5:31–61.
Zong A, Cao H, Wang F. Anticancer polysaccharides from natural resources: a review of recent research. Carbohydr Polym. 2012;90:1395–410.
Alkrad JA, Merstani Y, Neubert RHH. New approaches for quantifying hyaluronic acid in pharmaceutical semisolid formulations using HPLC and CZE. J Pharm Biomed Anal. 2002;30:913–9.
Čožíková D, Šílová T, Moravcová V, Šmejkalová D, Pepeliaev S, Velebný V, et al. Preparation and extensive characterization of hyaluronan with narrow molecular weight distribution. Carbohydr Polym. 2017;160:134–42.
Oudshoorn MHM, Rissmann R, Bouwstra JA, Hennink WE. Synthesis of methacrylated hyaluronic acid with tailored degree of substitution. Polymer. 2007;48:1915–20.
Finelli I, Chiessi E, Galesso D, Renier D, Paradossi G. Gel-like structure of a hexadecyl derivative of hyaluronic acid for the treatment of osteoarthritis. Macromol Biosci. 2009;9:646–53.
Buffa R, Šedová P, Basarabová I, Bobula T, Procházková P, Vágnerová H, et al. A new unsaturated derivative of hyaluronic acid—synthesis, analysis and applications. Carbohydr Polym. 2017;163:247–53.
Pravata L, Braud C, Boustta M, El Ghzaoui A, Tømmeraas K, Guillaumie F, et al. New amphiphilic lactic acid oligomer–hyaluronan conjugates: synthesis and physicochemical characterization. Biomacromolecules. 2008;9:340–8.
Šedová P, Buffa R, Kettou S, Huerta-Angeles G, Hermannová M, Leierová V, et al. Preparation of hyaluronan polyaldehyde—a precursor of biopolymer conjugates. Carbohydr Res. 2013;371:8–15.
Palumbo FS, Pitarresi G, Mandracchia D, Tripodo G, Giammona G. New graft copolymers of hyaluronic acid and polylactic acid: synthesis and characterization. Carbohydr Polym. 2006;66:379–85.
Vasi A-M, Popa MI, Butnaru M, Dodi G, Verestiuc L. Chemical functionalization of hyaluronic acid for drug delivery applications. Mater Sci Eng C. 2014;38:177–85.
Pouyani T, Prestwich GD. Functionalized derivatives of hyaluronic acid oligosaccharides: drug carriers and novel biomaterials. Bioconjug Chem. 1994;5:339–47.
Eenschooten C, Guillaumie F, Kontogeorgis GM, Stenby EH, Schwach-Abdellaoui K. Preparation and structural characterisation of novel and versatile amphiphilic octenyl succinic anhydride–modified hyaluronic acid derivatives. Carbohydr Polym. 2010;79:597–605.
Zawko SA, Truong Q, Schmidt CE. Drug-binding hydrogels of hyaluronic acid functionalized with β-cyclodextrin. J Biomed Mater Res Part A. 2008;87A:1044–52.
Botha C, Viktor Z, Moire C, Farcet C, Brothier F, Pfukwa H, et al. Separation of hydrophobically modified hyaluronic acid according to the degree of substitution by gradient elution high performance liquid chromatography. Anal Bioanal Chem. 2018;410:4259–73.
Botha C, Kuntz J-F, Moire C, Farcet C, Pfukwa H, Pasch H. Molar mass analysis of hydrophobically modified hyaluronic acid by SEC-MALLS: facing the challenges of amphiphilic biomacromolecules. Macromol Chem Phys 2018;https://doi.org/10.1002/macp.201800233.
Pasch H. Hyphenated separation techniques for complex polymers. Polym Chem. 2013;4:2628–50.
Pasch H, Trathnigg B. HPLC of polymers. Berlin: Springer; 1997. p. 223.
Radke W, Falkenhagen J. Chapter 5. Liquid interaction chromatography of polymers. In: Fanali S, Haddad PR, Poole CF, Schoenmakers P, Lloyd D, editors. Liquid chromatography. Amsterdam: Elsevier; 2013. p. 93–129.
Raust J-A, Brüll A, Moire C, Farcet C, Pasch H. Two-dimensional chromatography of complex polymers: 6. Method development for (meth)acrylate-based copolymers. J Chromatogr A. 2008;1203:207–16.
Radke W. Polymer separations by liquid interaction chromatography: principles – prospects – limitations. J Chromatogr A. 2014;1335:62–79.
Maiko K, Hehn M, Hiller W, Pasch H. Comprehensive two-dimensional liquid chromatography of stereoregular poly(methyl methacrylates) for tacticity and molar mass analysis. Anal Chem. 2013;85:9793–8.
Schoenmakers P, Aarnoutse P. Multi-dimensional separations of polymers. Anal Chem. 2014;86:6172–9.
Shakun M, Heinze T, Radke W. Characterization of sodium carboxymethyl cellulose by comprehensive two-dimensional liquid chromatography. Carbohydr Polym. 2015;130:77–86.
Ghareeb HO, Radke W. Characterization of cellulose acetates according to DS and molar mass using two-dimensional chromatography. Carbohydr Polym. 2013;98:1430–7.
Stern R, Kogan G, Jedrzejas MJ, Šoltés L. The many ways to cleave hyaluronan. Biotechnol Adv. 2007;25:537–57.
Acknowledgements
The preparation of the HA and HAM samples by L’Oréal Laboratories is acknowledged.
Funding
The authors received financial support from the L’Oréal Research and Innovation (Aulnay-sous-Bois, France). ZV received funding for her MSc project from the NRF.
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Viktor, Z., Farcet, C., Moire, C. et al. Comprehensive two-dimensional liquid chromatography for the characterization of acrylate-modified hyaluronic acid. Anal Bioanal Chem 411, 3321–3330 (2019). https://doi.org/10.1007/s00216-019-01799-x
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DOI: https://doi.org/10.1007/s00216-019-01799-x