Designing Zwitterionic Peptide with Superhydrophilicity through Carboxyl–Amino Spacing Adjustment for Robust Antifouling Electrochemical Biosensing in Complex Biofluids

The performance of electrochemical biosensors is severely compromised by biofouling in complex biofluids, underscoring the necessity to explore high-performance antifouling materials to construct robust nonfouling electrochemical biosensors. Here, the antifouling zwitterionic peptide CPPPP(D-Dap)(D-Dap)(D-Dap)(D-Dap) (CP(DDap)) was designed by adjusting the distance between the adjacent amino and carboxyl groups of the peptide, and it was further employed to fabricate antifouling electrochemical biosensors. Compared with traditional zwitterionic peptides, CP(DDap) exhibited not only superhydrophilicity but also higher structural rigidity and a smaller dipole moment, enabling it to achieve superior antifouling performance in complex biological media, while its unnatural characteristics endowed it with robust stability against hydrolysis by natural proteinase in biofluids. The CP(DDap)-based biosensor demonstrated high sensitivity for cortisol detection in serum with a remarkably low detection limit of 3.5 pg·mL^-1, and it displayed superior antifouling performance in real serum over 3 weeks. Notably, the assay results obtained by this biosensor in a series of clinical samples were consistent with those obtained using a commercial cortisol assay kit. The strategy of adjusting amino-carboxyl spacing in zwitterionic peptides explored in this study provides a new approach for designing highly efficient antifouling materials, thereby advancing the development of robust biosensors and bioelectronics tailored for practical applications in complex biofluids.