The development of environmentally sustainable control strategies to fight insect pests is a key challenge nowadays. Pheromone-mediated mating disruption (MD) is based on the release of synthetic sex attractants into a crop, interfering with mate finding of a given pest species. However, a limited number of research items have been published on the optimization of MD strategies against the European grapevine moth, Lobesia botrana, as well as on the use of biodegradable dispensers to reduce waste production in vineyards, despite the high economic importance of this pest. Therefore, the present study evaluated the efficacy of the MD products Isonet® L TT and the biodegradable Isonet® L TT BIO, applied at various densities, in reducing L. botrana damage on grapevine in comparison to an untreated control and the reference MD product Isonet® L. Experiments were conducted in three different areas of grapevine cultivation, located in Central and Northern Italy, over three different years. Our MD approach allowed a reliable control of the three generations of L. botrana during the whole grape growing season, leading to a significant reduction in the infested flower clusters and bunches, as well as in the number of nests per flower cluster and bunch, if compared to the untreated control. The performances of Isonet® L TT BIO, Isonet® LTT, and Isonet® L did not differ in terms of infested flower clusters/bunches, as well as nests per flower cluster/bunch. This was confirmed in all experimental sites over 3 years of field experiments. Overall, the present research provides useful information for the optimization of MD programs against L. botrana, highlighting the interesting potential of biodegradable pheromone dispensers that can be easily applied at low densities in vineyards, reducing the use of chemical pesticides to control moth pests.