Diamine oxidase (DAO) from Pisum sativum is an enzyme that catalyzes the degradation of biogenic amines (BA) present in wine, producing harmless aldehydes and hydrogen peroxide (H2O2). H2O2 promotes a rapid inactivation of the immobilized enzyme. At first glance, co-immobilization of DAO and catalase (CAT) could improve the elimination of the released hydrogen peroxide. Two different co-immobilized derivatives were prepared: (a) both enzymes co-localized and homogeneously distributed across the whole structure of a porous support, and (b) both enzymes we de-localized inside the porous support: DAO immobilized on the outer part of the porous support and catalase immobilized in the inner part. Co-localized derivatives were seven-fold more effective than de-localized ones for the elimination of hydrogen peroxide inside the porous support. In addition to that, the degradation of putrescine by DAO was three-fold more rapid when using both co-localized enzymes. The optimal co-localized derivative (containing 1.25 mg of DAO plus 25 mg of CAT per g of support) promoted the instantaneous elimination of 91% H2O2 released inside the porous support during putrescine oxidation. This optimal derivative preserves 92% of activity after three reaction cycles and DAO immobilized without catalase only preserves 41% of activity. Co-localization seems to be the key strategy to immobilize two sequential enzymes. When enzymes are immobilized in close proximity to each other in a co-localized pattern, the generation of byproducts as H2O2 is strongly reduced.