To investigate the effect of oxygen content on the quality of pearl gentian grouper during waterless keeping alive, the changes of physiological indicators and muscle quality of pearl gentian grouper before waterless keeping alive, during keeping alive (0,3, 6,9, 12h) and after revival for 12h under different oxygen volume fraction (60%, 80%, 99%) in package were analyzed, and the quality prediction model was established. The results showed that when the oxygen volume fraction was 99%, the plasma glucose and cortisol mass concentrations of pearl gentian grouper were significantly increased with the prolongation of keeping alive time (P<0.05), and the maximum values were (0.86±0.01) mg/mL and (3034.21±14.84) ng/L, respectively. In addition, muscle springiness decreased from 0.79±0.07 to 0.50±0.01, and cohesiveness declined from 0.61±0.01 to 0.51±0.00. When the oxygen volume fraction was 60%, the activities of alanine aminotransferase, aspartate aminotransferase and the mass concentration of creatinine in plasma were increased steadily during the keeping alive period. The mass ratio of glycogen in muscle decreased from (1.47±0.02) mg/g to (0.71±0.02) mg/g, resulting in the accumulation of a large amount of lactic acid. In contrast, grouper under 80% oxygen volume fraction could more actively adapt to low temperature and anhydrous environment because of a relatively low stress level. Finally, the quality prediction model was established with plasma glucose, muscle glycogen, springiness and cohesiveness as key quality indexes for the health quality of grouper, and fitting degree of which was good. This study could provide reference for the excavation of health quality indicators during the actual transportation process of grouper.