# Toxicity Evaluation of Sulfobetainized Branched Polyethyleneimine via Antibacterial and Biocompatibility Assays

## TL;DR

This paper evaluates the toxicity and antibacterial properties of modified polyethyleneimine (PEI) after a chemical process called sulfobetainization.

## Contribution

The study introduces a novel sulfobetainization method to improve PEI's biocompatibility while retaining or adjusting its antibacterial properties.

## Key findings

- Sulfobetainized PEI showed no toxicity to fibroblast cells even at high concentrations.
- Multiple sulfobetainization steps increased thermal stability and zwitterionic content of PEI.
- Antimicrobial properties of PEI can be modulated through sulfobetainization and pH adjustments.

## Abstract

Branched polyethyleneimine (PEI), possessing different types of amines—e.g., primary, secondary, and tertiary—in the polymer chains are well known for their antibacterial properties and DNA condensing ability, affording substantial advantages in many biomedical uses, including gene therapy. However, because of PEI’s toxicity, depending on the molecular weight, its widespread biomedical use is hindered. Therefore, in this study, PEIs with different molecular weights—i.e., 600, 1200, and 1800 g/mol—were modified with 1,3-propane sultone, undergoing a sulfobetainization reaction in a single step to attain a zwitterionic structure: sulfobetainized PEI (b-PEI). The sulfobetainization reaction was carried out twice to increase the zwitterionic repeating unit on PEI chains. The increasing number of SO3− groups on the PEI chains was confirmed by the increased peak intensities around 1160 and 1035 cm−1 on the FT-IR spectrum, which are assigned to symmetric and asymmetric S=O peaks. The elemental analysis results for first- and second- betainization PEIs, abbreviated as b1-PEI and b2-PEI, respectively, were revealedthe increased wt% of S confirming the successful multiple-sulfobetainization of the PEI chains. Thermal stability analyses of PEIs and their corresponding multiple-sulfobetainized forms showed that multiple-sulfobetainization reactions increased the thermal stability of bare PEI chains. PEIs with lower molecular weights exhibited more antimicrobial properties. As PEI is sulfobetainated, its antimicrobial properties can be further adjusted via sulfobetainization (once or twice), or by adjusting the corresponding solution pH, or by protonating them with different acids with different counter anions. The cell toxicity of PEI on L929 fibroblast cells was slightly increased by increasing the molecular weight of the PEI, but all forms of sulfobetainized PEIs were found to be safe (no toxicity), even at 1000 µg/mL concentrations.