Eugenol is a widely used fishery anesthetic. This study investigated the effects of various concentrations of eugenol on blood physiological and biochemical indexes, and muscle flavor, in crucian carp (Carassius auratus). To determine the appropriate concentration of eugenol anesthetic for use in crucian carp transportation and production operations, we evaluated seven anesthesia groups of 20, 30, 40, 50, 60, 70, and 80 mg/L and one control group (without eugenol) to determine the effects on blood physiological and biochemical indexes, and muscle flavor. The red blood cells and platelets of crucian carp decreased significantly (p < 0.05) with eugenol treatment. With increasing eugenol concentration, the white blood cells and hemoglobin did not change significantly, whereas lactate dehydrogenase, alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase increased significantly (p < 0.05). The content of phosphorus, magnesium, and sodium increased after anesthesia, whereas the content of potassium decreased with increasing eugenol concentration. After anesthesia, the content of albumin and total protein in the serum decreased with increasing eugenol concentration (p < 0.05); triglyceride first increased and subsequently decreased (p < 0.05); blood glucose content first increased and then decreased (p < 0.05); and no significant difference was observed in total cholesterol content (p > 0.05). No significant difference was observed in muscle glycogen and liver glycogen content after eugenol anesthesia (p > 0.05). The eugenol-based anesthesia test did not indicate major liver histomorphological effects, but the very small number of gill sheet edema cases observed requires further study. Analysis of electronic nose data indicated that eugenol treatment affected the flavor of the fish. The anesthesia concentration of 20–80 mg/L had some effect on the physiology and biochemistry of crucian carp, thus providing a reference for the application of eugenol in crucian carp transportation and experimental research.