Formulation and characterization of gelatin-based hydrogels for the encapsulation of <em>Kluyveromyces lactis</em> and chitosan nanoparticles: biotech and biomedical applications

Abstract

A difficulty when orally administering microorganism-based probiotics is the significant loss of their bioactivity as they pass through the gastrointestinal (GI) tract. To overcome these issues, we propose to encapsulate the probiotic yeast Kluyveromyces lactis on chemically crosslinked gelatin hydrogels to protect the bioactive agents in different environments. Moreover, a challenge to obtain favorable results with therapeutic drugs and biomolecules is the inefficient cellular administration process. For this reason, we considered chitosan and gelatin nanoparticle loading systems to improve therapeutic efficacy. Also, we prepared hydrogels to encapsulate such nanoparticles by the chemical crosslinking of gelatin, an inexpensive and commercially available polymer. To explore changes in key physicochemical parameters and their impact on cell viability, we varied the concentration of the crosslinking agent (glutaraldehyde) and the gelatin. The synthesized hydrogels were characterized in morphological, physical-chemical, mechanical, thermal, and rheological properties. This comprehensive characterization allowed us to identify critical parameters to facilitate encapsulation and enhance system performance. Mainly due to pore size in the range of 5-10 µ m, sufficient rigidity (breaking forces of about 1 N), low brittleness and structural stability under swelling and relatively high shear conditions, we selected hydrogels with a high concentration of gelatin (7.5% (w/v)) and concentrations of the crosslinking agent of 3.0% and 5.0% (w/w) for cell and nanoparticles encapsulation. Yeasts and nanoparticles were encapsulated and subsequently tested in bioreactor operation and GI tract simulated media, thereby leading to cell viability levels that approached 95% and 50%, respectively. After testing, the hydrogels' firmness was only reduced to half of the initial value and maintained resistance to shear even under extreme pH conditions. These encouraging results indicate that the proposed encapsulates are suitable for overcoming most of the major issues of oral administration of drugs and probiotics and open the possibility to further explore additional biotech applications.