A Biomaterial-Based Approach to Trypsin Sensing: Design and Optimization of Gelatin–Casein Films

Real-time postoperative monitoring systems have tremendous potential to detect postoperative complications faster before patients become systemically ill. This study investigates the potential of gelatin-casein blend films as a biodegradable, implantable biomaterial platform for trypsin detection, which is a potential biomarker for an anastomotic leak from the duodenum or proximal jejunum. Although implantability has not been verified in this case, the implantability of gelatin and casein-based biomaterials is substantiated by their demonstrated cytocompatibility as evidenced below and established utility in medical applications, as evidenced by recent advancements in biomaterials research. We systematically evaluated nine gelatin-casein blends, ranging from 0:100 to 100:0 ratios to optimize their enzymatic degradability and response, establishing a material platform for future biosensing applications. Optimal refers to the sensor's ability to rapidly detect physiological concentrations of trypsin in the body while generating the maximum detectable response. Characterization of the films was performed using Fourier-Transform Infrared Spectroscopy (FTIR), UV-visible Spectroscopy (UV-vis), and Quartz Crystal Microbalance (QCM). The films' responses to trypsin were analyzed through limit of detection, initial reaction rates, and absorbance shifts. For statistical analysis, a flexible exponential decay model, was employed to assess the significance of the results. Our findings reveal that the 75:25 casein:gelatin blend exhibits superior performance, with the lowest limit of detection (7.81 × 10^-11 M), highest initial reaction rate (6.936 ΔHz/s by QCM, -0.095 AU/min by UV-vis), and most significant absorbance shift (-2.208 AU after 10 min). This optimal blend demonstrates a 10-fold improvement in detection limit compared to pure gelatin films and a 5-fold enhancement over pure casein films. The remarkable sensitivity, rapid response, and significant signal change of the 75:25 casein:gelatin blend make it a promising candidate biomaterial platform for an implantable trypsin sensor.