Vxr 300 mhz

The synthesis of CPTEG and CPH diacids was performed as previously described (56 (link)). Melt polycondensation was used to synthesize 20:80 CPTEG : CPH copolymer as described previously (56 (link)). Polymer purity, copolymer composition, and molecular weight were evaluated using ¹H nuclear magnetic resonance spectroscopy (VXR 300 MHz, Varian, Palo Alto, CA).
Before NP synthesis, BRSV post-fusion F protein (GenScript, Piscataway, NJ) and BRSV G protein (KanPro, Lawrence, KS) were separately dialyzed with the use of 10k MWCO Spin-X UF Concentrators (Corning, Corning, NY) and lyophilized for 48 h. A double emulsion nanoprecipitation technique was used to synthesize polyanhydride particles as previously described (57 (link)). Briefly, 20:80 CPTEG : CPH copolymer, post-fusion F protein (1.5 wt.%), G protein (1.5 wt.%) and CpG ODN 2007 (1 wt.%) (InvivoGen, San Diego, CA) were dissolved in methylene chloride and sonicated at 30Hz for 30s using a probe sonicator. Following sonication, the solution was quickly poured into a bath of chilled pentane (-4°C) at a solvent to non-solvent ratio of 1:200 and particles were collected by vacuum filtration.

All reagents from commercial suppliers were used without further purification. All solvents were freshly distilled before use from appropriate drying agents. All other reagents were recrystallized or distilled when necessary. Reactions were performed under a dry nitrogen atmosphere. Analytical thin layer chromatography (TLC) was performed with silica gel 60 F₂₅₄ plates. Visualization was accomplished by UV light. Column chromatography was carried out using silica gel 60 (230–400 mesh ASTM). Melting points were determined with an Electrothermal IA9100 Digital Melting Point Apparatus without correction. NMR spectra were obtained on Bruker Avance 400 MHz and Varian VXR 300 MHz spectrometers and recorded at 25 °C. Chemical shifts for ¹H NMR spectra are reported in ppm downfield from TMS, chemical shifts for ¹³C NMR spectra are recorded in ppm relative to internal deuterated chloroform (δ = 77.2 ppm for ¹³C), and chemical shifts for ¹⁹F NMR are reported in ppm downfield from fluorotrichloromethane (CFCl₃). Coupling constants (J) are reported in Hertz. The terms m, s, d, t, and q refer to multiplet, singlet, doublet, triplet, and quartet, respectively. ¹³C NMR and ¹⁹F NMR were broadband decoupled from hydrogen nuclei. High-resolution mass spectra (HRMS) were measured by EI method with an Agilent LC-Q-TOF-MS 6520 spectrometer.