The synthesis of vanillin has meanwhile a tradition of more than 150 years, with recently a clear trend towards methods for biobased and possibly natural-quality vanillin. Large waste-streams of lignin are available for the production of lignin-oil, which consist of substituted aromats with unsaturated sidechains (monolignols). Aromatic DiOxygenases (ADOs) activate molecular oxygen and produce two species of carbonyl compounds from a monolignol. We call them ‘ozonylases’, because these enzymes may be a substitution for chemical ozonolysis – a rather dangerous process on scale due to the formation of explosive intermediates. We identified and expressed 13 novel ozonylases. Among the tested enzymes, we found a highly effective enzyme that cleaves isoeugenol, 4-vinylguaiacol and most importantly coniferyl alcohol to vanillin. Herein we present selected characteristics of this enzyme and show gram scale whole cell biotransformations with quantitative conversion of 50 mM isoeugenol to vanillin (Scheme 1). We also show promising enzymatic cascade reactions from eugenol and ferulic acid to vanillin.