Design and characterization of electrospun fibers for controlled delivery of antimicrobial peptides and proteins
Authors:   , Kielholz Tobias 
Published: November 2024

Widespread antimicrobial resistance in bacteria such as Staphylococcus aureus and Pseudomonas aeruginosa leads to life-threatening infections and is a major challenge in clinical practice. Unfortunately, common antibiotic drugs are prone to induce antimicrobial resistance through various mechanisms like enzymatic cleavage and thus represent inadequate treatment options. Innovative therapeutics are therefore urgently sought to address the increasing number of infections complicated by resistant strains. Promising approaches in this context involve in particular antimicrobial peptides and bacteriophages. These innovative antimicrobials exert more complex or multiple modes of action, overcoming typical mechanisms that drive antibiotic resistance in bacteria. However, related to the local therapy of infected wounds, established drug delivery systems, such as semi-solid dosage forms, have disadvantages in terms of mechanical stress during application and in providing favorable conditions for optimal wound healing. Here, polymer fiber based wound dressings have proven to exhibit the desired properties for controlled drug delivery to eradicate bacteria while creating a beneficial microenvironment that allows the wound to heal. Such dressings can be produced by electrospinning, a versatile technique that generates nonwoven fiber mats of micro- to nanoscale fibers with a high surface-to-volume ratio by applying high voltage to a polymeric solution flux. In addition, the properties of the resulting fiber mats can be tuned by the choice of polymers, and advanced electrospinning setups can be employed to achieve optimal protection of the bioactive cargo. However, characterizing such electrospun wound dressings loaded with innovative antimicrobial agents is not trivial and is hampered by the extremely small and intersecting fiber mat structure. Characterization is further complicated because such antimicrobials are not structured with simple chemical moieties like small molecules, requiring even more sophisticated fiber analysis.
A comprehensive analytical toolset for in-depth characterization of complex electrospun fiber mats is pursued in the first section, with the additional objective of creating a coherent concept of complementary techniques as a basis for rational electrospun fiber development. In the second and third chapter, the aim was to develop electrospun wound dressing formulations that allow the bioactive encapsulation of innovative antimicrobial agents such as antimicrobial peptides and bacteriophages. In the second chapter, the aim was to develop an electrospun wound dressing formulation that allows the bioactive encapsulation of different bacteriophage types with the intention of rapid release for the treatment of wounds infected with the critical strains of bacteria Staphylococcus aureus and Pseudomonas aeruginosa. The objective of the last paragraph was to use a systematic approach for the preparation of electrospun wound dressings with prolonged release and enhanced aqueous solubility of the antimicrobial peptide tyrothricin and to characterize the antimicrobial performance against two prominent wound pathogens.
Electrospinning has emerged as a key technique for the fabrication of polymer-based fiber mats, which have shown tremendous potential as a platform for various pharmaceutical applications when encapsulating an active pharmaceutical ingredient. In this context, reproducible fiber properties are critical for realizing rational formulation development. Therefore, a thorough fiber characterization is necessary to guarantee, for example, the desired drug release kinetics. However, until now, there has been no standardized testing strategy for dosage forms such as tablets as we know it from the regulation by the International Pharmacopoeia, and the associated fiber mat analysis of such complex structures is challenging and further hampered by the introduction of innovative drugs such as (poly )peptides. To this end, two fiber mats with contrasting physico-chemical properties were electrospun, and a range of analytical techniques were evaluated for their respective capabilities and limitations.
Since adequate fiber formation during electrospinning is strongly dependent on the polymer solution used, rheological studies have been performed to systematically assess the optimal polymer concentration that yields flawless electrospun fibers. After electrospinning, the resulting products were examined using different complementary visualization methods. Stereomicroscopy was a suitable method to quickly determine whether fibers or particles were obtained. Higher magnification is required to detect defects in fiber morphology, such as drug crystals or inhomogeneities, and was achieved with scanning electron microscopy. The corresponding micrographs were further evaluated using an image analysis software to obtain the diameter distribution of the fibers. If multiple fiber types are evident, confocal Raman imaging can be used for complementary chemically selective elucidation. Attenuated total reflection infrared spectroscopy offers faster qualitative compositional characterization compared to the Raman approach, but without spatial resolution, spectral superposition remained a challenge to interpretation.