Effect of Al content on the stacking fault energy (SFE) was investigated in the austenitic Fe-25Mn-(1.16-9.77)Al-0.68C (at%) alloys by X-ray diffraction line profile analysis and thermodynamic estimation, and was discussed on the basis of anomaly in shear modulus caused by the antiferromagnetic transition. The experimental results show that the stacking fault probability decreases with increasing Al content, the observed SFE increases linearly when Al content is lower than 6.27 at%, and markedly as it is more than 6.27 at%. The thermodynamic estimation indicates that the non-magnetic component of SFE increases faster than the observed one with increasing Al content in the antiferromagnetic state, and both are almost equal in the paramagnetic state. The magnetic order increases SFE in the antiferromagnetic state, and the magnetic component of SFE depends on the average magnetic moment and Neel temperature. The increases in the localized magnetic moment and the decrease in the Neel temperature are caused by the addition of Al atoms into the austenitic Fe-Mn alloys and are accompanied by the anomaly in shear modulus, which affects SFE in the antiferromagnetic state. The anomalous drop in shear modulus leads to the inconsistency for the variations of the observed SFE and non-magnetic component with Al content in the antiferromagnetic state.