We investigate the influence of Kondo effect, namely, the nonperturbative effect induced by heavy impurities, on the chiral separation effect (CSE) in quark matter. We employ a simple effective model incorporating the Kondo condensate made of a light quark and a heavy quark, and compute the response function of axial current to the magnetic field in static limit and dynamical limit. As a result, we find that the Kondo effect catalyzes the CSE in both the limits, and particularly the CSE in dynamical limit can be enhanced by a factor of approximately three. Our findings clearly show that the presence of heavy impurities in quark matter can play an important role in the transport phenomena of light quarks induced by a magnetic field.