The Role of Crosslinker Content of Positively Charged NIPAM Nanogels on the In Vivo Toxicity in Zebrafish

Abstract

Polymeric nanogels as drug delivery systems offer great advantages, such as high encapsulation capacity and easily tailored formulations; however, data on biocompatibility are still limited. We synthesized N-isopropylacrylamide nanogels, with crosslinker content between 5 and 20 mol%, functionalized with different positively charged co-monomers, and investigated the in vivo toxicity in zebrafish. Our results show that the chemical structure of the basic unit impacts the toxicity profile depending on the degree of ionization and hydrogen bonding capability. When the degree of crosslinking of the polymer was altered, from 5 mol% to 20 mol%, the distribution of the positively charged monomer 2-tert-butylaminoethyl methacrylate was significantly altered, leading to higher surface charges for the more rigid nanogels (20 mol% crosslinker), which resulted in >80% survival rate (48 h, up to 0.5 mg/mL), while the more flexible polymers (5 mol% crosslinker) led to 0% survival rate (48 h, up to 0.5 mg/mL). These data show the importance of tailoring both chemical composition and rigidity of the formulation to minimize toxicity and demonstrate that using surface charge data to guide the design of nanogels for drug delivery may be insufficient.

Sustainable and Surfactant-Free Synthesis of Negatively Charged Acrylamide Nanogels for Biomedical Applications

“…DLS measurements were carried out to analyze the particle sizes with NG solutions in phosphate buffer (10 mM pH 7.4) forming stable colloidal solutions. The measurements (Figure 3a) showed that NGs synthesized in DMSO resulted in particles with hydrodynamic diameter (Dh) less than 10 nm (Figure S2), for both neutral and negatively charged particles, which is consistent with previously reported data 50 . Polymerization in DMSO with CMt = 1% (w/w) is affected by the solvent limiting the reactivity of Aam 48 , stabilizing the growing polymeric chains and leading to smaller particles.…”

Impact of Protein Corona Formation on the Thermoresponsive Behavior of Acrylamide-Based Nanogels

“…The incorporation of positively charged comonomers with pH-responsive behavior, such as tert -butylaminoethyl-methacrylate, was not evaluated in this work due to the observed toxicity of the resulting nanogels in vivo. 38 All nanogels were synthesized with high total monomer conversions (>90% as determined by 1 H NMR, Figure S1 ) and chemical yields (>70%) ( Table 1 ), providing evidence of consistency between the initial formulation and the composition of the isolated polymers. Higher monomer conversions compared to the chemical yields result from the loss of low-molecular-weight chains during the purification step via dialysis (molecular-weight cutoff 3.5 kDa), which also ensures the elimination of any residual monomers that could cause cytotoxicity.…”

“…Negatively charged nanogels were obtained by incorporating 2.5 molar % of ProAM, previously employed by our group to obtain nanogels with dual thermo- and pH-responsive properties. The incorporation of positively charged comonomers with pH-responsive behavior, such as tert -butylaminoethyl-methacrylate, was not evaluated in this work due to the observed toxicity of the resulting nanogels in vivo . All nanogels were synthesized with high total monomer conversions (>90% as determined by 1 H NMR, Figure S1) and chemical yields (>70%) (Table ), providing evidence of consistency between the initial formulation and the composition of the isolated polymers.…”

“…We previously reported the development of thermoresponsive nanogels as drug delivery vectors, demonstrating the role of chemical composition and synthetic methodologies on their thermoresponsive behavior, both in buffer solutions and at the air/water interface. , Additionally, our studies on the environmental exposure of nanogels to zebrafish larvae showed the high toxicity of positively charged formulations of N -isopropylacrylamide (NIPAM)-based nanogels, which prompted us to focus on negatively charged nanogels for this study.…”