Investigating triazine-based modification of hyaluronan using statistical designs

• Published in: Carbohydrate polymers 2015-11, Vol.132, p.472-480
• Main Authors: Liang, Jue, Cheng, Lulu, Struckhoff, Jessica J., Ravi, Nathan
• Format: Article
• Language: English
• Subjects: Amidation; Cell Line; Cell Survival - drug effects; derivatization; Fractional-factorial design; Humans; Hyaluronan; hyaluronic acid; Hyaluronic Acid - chemistry; Hyaluronic Acid - toxicity; Magnetic Resonance Spectroscopy; molecular weight; Morpholines - chemistry; nuclear magnetic resonance spectroscopy; regression analysis; Response surface methodology; solvents; Sulfhydryl Compounds - chemistry; Triazine Coupling; Triazines - chemistry; Water - chemistry; water content
• ISSN: 0144-8617; 1879-1344
• DOI: 10.1016/j.carbpol.2015.06.067

•We used statistical designs to study triazine-based derivatization of hyaluronan.•We observed the triazine-method to be more efficient compared to carbodiimide-method.•We evaluated the effect of each factor on the derivatization using fractional factorial design.•We used response surface methodology to control the derivatization and optimize the reaction condition.•We determined the stability, purity and biocompatibility of derivatized hyaluronan. Hyaluronan (HA) and its derivatives have been extensively researched for many biomedical applications. To precisely tailor the property of HA by derivatizing it to a pre-determined extent is challenging, yet critical. In this paper, we used 2-chloro-4,6-dimethoxy-1,3,5-triazine (CDMT) and N-methylmorpholine (NMM) to derivatize HA via a triazine-based coupling reaction. Using a fractional factorial (FF) design, we observed that water content in the solvent, and molar ratios of CDMT and NaHCO3 to the carboxylate were the significant factors controlling the derivatization. We investigated how the effect of each factor changes as reaction conditions change. Moreover, by altering the amount of CDMT and NaHCO3, we developed a cubic regression model for precise control of the extent of derivatization using a response surface methodology (RSM) with a D-optimal design. No spurious peaks were detected by 1H NMR spectrum and only 10% decrease of molecular weight of the derivatized HA was determined by GPC. The HA with 6% modification was relatively biocompatible up to 15mg/mL.