2.3. Samples Preparation

The crude juices were prepared as described previously [15]. The stem and peels (pineapple by-products) were reduced to a crude juice, using a juice machine (model: HR1922/21 1200 W, Philips) that splits the press cake (solid parts) from the crude juice (liquid parts). The crude juice was centrifuged 7370× g at 4 °C for 10 min, and the remaining supernatant was centrifuged to remove the remaining pulp.

The enzyme from pineapple fruit is a well-known enzyme in the market, associated with its proteolytic activity and when present in crude juices, influences the prebiotic potential associated, and for that reason an enzymatic extraction was performed to remove the enzymatic fraction from the final product. For that, a green chemistry separation methodology was applied to the remain supernatant as described by Campos et al. [16] to remove bromelain.

The biological activities—prebiotic and antioxidant were assessed for the resulting supernatant (PBLF, without the enzymatic fraction).

The resulting supernatant from enzyme extraction was frozen at −20 °C, with a cryoprotectant Maltodextrin (MAL 2%; w/v) and applied to a freeze-drying process through a vacuum pressure of 0.1 torr in the freeze drier (Model FT33, Armfield, UK), and the freezing temperature in the chamber was −46 °C, while the samples chamber temperature was 15 °C.