Potential harmful effects of spent coffee grounds (SCGs)-derived biochar on earthworms (Lumbricus terrestris) were investigated through two complementary experiments, which assessed the avoidance response of earthworms to biochar-amended soils (experiment 1), and the response of oxidative stress biomarkers and digestive enzymes (experiment 2). The main results were: 1) the highest dose of biochar (5% w/w) caused a significant avoidance response of earthworms (75% individuals avoided these treated soils after 48 h); 2) signs of oxidative stress were early detected in earthworms exposed to biochar (1 and 5% w/w) as indicated by the integrated biological response index; 3) earthworms exposed to biochar-amended soils for 30 d experienced a significant increase of digestive enzyme activities measured in both the gastrointestinal tissue and the luminal content; 4) interaction between earthworms and biochar led to a higher soil extracellular enzyme activities in the 1% biochar treatment than that of control and 5% biochar treatments. These findings suggest that the joint application of SCG-biochar and L. terrestris is a workable approach for improving soil quality in terms of soil biochemical promotion, although earthworms may develop some physiological mechanisms of biochar tolerance (antioxidant defenses).