A conditioner containing lugworm autolysate was prepared, and its feasibility as a new conditioner was investigated. The treatment of hair with the conditioner, before and after dyeing showed positive effects on the thickness, weight, and amino acid contents of the hair. These properties were significantly enhanced, and similar results were obtained by hair surface examination by using scanning electron microscopy and tensile strength measurements. The surface of the cuticles in the experimental group was smoother than that of the control group, and the arrangement of the hair scales was maintained even after shampooing for 20 times. The hair tensile strength of the experimental group was significantly higher than that of the control group. The significant treatment effect of the conditioner containing lugworm autolysate could be attributed to the synergistic effect of adhesive strength and effective film formation properties of lugworm autolysate.