Enzymes are very important components which are vital for the existence of every cellular life. There is significant interest in the use of structurally stable and catalytically active enzymes in pharmaceutical, food, fine chemicals industries, and in various industrial processes as catalysts. Stem bromelain (BM) is a proteolytic enzyme which is widely used in chemical, medical, and pharmaceutical fields. However, harsh process conditions are the main barriers to the effective use of this enzyme in different applications. To overcome these drawbacks, biocompatible biobased ionic liquids (ILs), composed of the choline cation (an essential nutrient) and different anions are used. The ILs namely choline chloride [Ch](+)[Cl](-), choline acetate [Ch](+)[Ac](-), choline dihydrogen phosphate [Ch](+)[Dhp](-), choline bitartrate [Ch](+)[Bit](-), choline iodide [Ch](+)[I](-), and choline hydroxide [Ch](+)[OH](-) are chosen for the current work. Therefore, in the present study, structural stability and activity of BM have been evaluated in the presence of choline-based ILs using various biophysical techniques at different concentrations. The present work demonstrated that [Ch](+)[OH](-) is the strongest destabilizer, whereas [Ch](+)[Cl](-) is the best stabilizer for the native structure of BM among all studied ILs. This work revealed the suitability of some choline-based ILs as potential media for sustained stability and activity of BM.