Avance 3 500 mhz high field nmr spectrometer

Alg-MA was synthesized utilizing an anhydrous reaction to control the degree of methacrylation (DOM).^{28 -29 (link)} Sodium alginate was rendered soluble in anhydrous DMSO through an ion exchange with DTAB. Aqueous solutions of sodium alginate (1%, w/v) and DTAB (2%, w/v) were prepared and slowly mixed while stirring at 1000 rotations per minute (rpm). The precipitate was washed in DI water and lyophilized. A 1% (w/v) alginate-DTA/DMSO solution was reacted with MA in the presence of a catalyst, DMAP, for 24 hours at room temperature. The solution was hydrolyzed through extensive dialysis in 0.2 M sodium phosphate dibasic salt solution followed by further dialysis in DI water. Alginate methacrylation was confirmed using ¹H-NMR spectroscopy, (Bruker AVANCE III 500 MHz high-field NMR spectrometer) by the presence of methacrylate (6.25, 5.75 ppm) and alginate methyl resonances (2.0 ppm). A 1% (w/v) polymer solution in D₂O was analyzed at room temperature, spinning at 20 Hz for 16 scans.^{28 , 30 (link)} The DOM was quantified by peak integration and calculation of the ratio between of the methyl protons at 2.0 ppm and the newly formed methylene protons of methacrylate at 5.75 ppm and 6.25 ppm.^{30 (link)-31 (link)}

Methacrylated alginate (AMA) was synthesized as previously described.^{24 (link), 56 (link)} Briefly, a 2% (w/v) solution of Manugel® GMB (MW≈ 170–240 kDa, FMC Biopolymer) in deionized (DI) water was mixed with a 20-fold molar excess of methacrylic anhydride (Sigma-Aldrich). The solution was maintained at pH 8 using 5 N sodium hydroxide (Fisher) for 24 hours. Purification was performed via dialysis (MWCO=6–8 kDa) against DI water for 5-days, and lyophilized to yield a dry product. A 1% (w/v) AMA was prepared in deuterium oxide (D₂O, Acros Organics) and subsequently analyzed using ¹H-NMR spectroscopy (Bruker AVANCE III 500 MHz high-field NMR spectrometer), for 64 scans at 20 Hz. The degree of methacrylation (DOM) was established through integration of the peaks associated with methacrylate (6.24, 5.78 ppm) and alginate methyl resonances (1.96 ppm), and calculation of the ratios between.^{29 (link), 56 (link), 58}

Alg-MA was synthesized using an aqueous-based reaction. One percent (w/v) sodium alginate (Manugel, Mw ≈ 200 kDa, FMC Biopolymer) solution, in deionized (DI) water, was reacted with a 20-molar excess of methacrylic anhydride (Sigma Aldrich) for 12 hours.[12 (link), 15 (link), 30 –32 (link)] The reaction product was treated with sodium hydroxide and dialyzed in DI water for 3 days. The degree of methacrylation (DOM), or ratio of methacrylate groups per repeat unit of alginate, was determined using ¹H-NMR spectroscopy (Bruker AVANCE III 500 MHz high-field NMR spectrometer). Briefly, Alg-MA was dissolved in deuterium oxide (D₂O) at 1% (w/v) solution and was analyzed at room temperature, spinning at 20 Hz for 16 scans. The DOM was determined by calculating ratios of methylene (6.1, 5.6 ppm) and alginate methyl (1.9 ppm) peak integrations.[15 (link)] Alg-MA material with a single DOM was used for subsequent rheological and mechanical analysis, and modification via oxidation.