Abstract: As a substitute of suture, bioadhesive attracts much attention in clinical application. However, the development of bioadhesive with excellent bonding performance for underwater usage remains a challenge. Herein, a non-aqueous bioadhesive with underwater bonding properties was manufactured based on hydrophobic hyperbranched polythioether (HBP). HBP with the surface containing abundant thiol groups was synthesized through a simple thiol-(meth)acrylate Michael addition polymerization of 2-(acryloyloxy)ethyl methacrylate (AA' monomer) and trimethylolpropane tris(3-mercaptopropionate) (B₃ monomer). HBP was a liquid material under room temperature, thus enabling the manufacturing of non-aqueous adhesives (HBP-x) by the direct addition of poly(ethylene glycol) diacrylate. The properties of HBP-x were studied using vial-tilt method, rheological test, lap-shear adhesion test, swelling test, degradation test and in vitro cytotoxicity test. Results showed that the HBP-adhesive could bond porcine skin with an underwater adhesive strength of up to 43 kPa. Moreover, the HBP-adhesive minimally swelled (7.2%—17.3%) and was degradable. In vitro cell experiments showed that the HBP-adhesive showed very low cytotoxicity to L929 cells, and allowed the proliferation of L929 cells on the surface of the adhesive. This non-aqueous HBP-adhesive based on thiol-acrylate Michael addition reaction displayed an underwater bonding ability and excellent cytocompatibility, holding great potential for clinical applications.