Pepsin

Low methoxyl pectin (LMP, degree of esterification 26%) from citrus peel was obtained from Ingredient Flavours Co., Ltd., Thailand. Hi-maize starch was procured from National Starch and Chemical Co., Ltd., Bangkok, Thailand. Bromelain from pineapple was purchased from Sisco Research Laboratories Pvt. Ltd., India. Pancreatin enzyme was purchased from Sigma-Aldrich, United States, and Pepsin from Acros, Denmark. All other chemicals used in the study were of analytical grade.

PVDF pellets were purchased from Sanowart Group Co., Ltd. (Zibo City, China). PEG with a molecular weight (MW) of 20 kDa, and microporous zeolite 96096 of the 13X type (crystalline, <10 µm particle size), comprising silica (SiO₂) and alumina (Al₂O₃) with metallic oxide, were purchased from Sigma Aldrich, Subang Jaya, Malaysia. N,N-dimethylformamide (DMF) was purchased from Fisher Scientific, Shah Alam, Malaysia. Egg albumin (EA) (MW = 44.3 kDa) was purchased from Sigma Aldrich, Malaysia, and bovine serum albumin (BSA) (MW = 66 kDa) and pepsin (MW = 35 kDa) were purchased from Acros Organics, Shah Alam, Malaysia.

The probiotic culture of Bifidobacterium animalis subsp. lactis BB-12 was obtained from Chr. Hansen (Hoersholm, Denmark). Sodium alginate, calcium chloride, xanthan gum, κ-carrageenan, pectin, inulin, glucose, and Tween 80 were provided by Sigma-Aldrich Chemie GmbH (Taufkirchen, Germany); glycerol was purchased from Lach-Ner (Brno, Czech Republic); and nanocrystalline cellulose (CNC) from CelluForce (Montreal, Canada). Olive pomace oil was kindly provided by MINERVA S.A. (Athens, Greece) and sweet whey with the following specifications: humidity 1%, fat content 1%, protein content 10.2%, lactose (hydrated) 75%, ash content 7.3%, was kindly provided by ION S.A. (Piraeus, Greece). Whole-fat milk was obtained from the local market. The materials for the microbiological analyses, such as MRS agar, Ringer’s solution, citric acid, and disodium phosphate, were acquired from Merck (Taufkirchen, Germany), whereas l-cysteine-HCl, neomycine sulfate, nalidixic acid, lithium chloride, and paromomycine sulfate by Thermo Fischer Scientific (MA, USA). Pepsin, pancreatin, and bile extract were obtained from Acros Organics (New Jersey, USA).

To perform the 11 experiments designed by the CCRD, S. Typhimurium cells recovered from BDL samples—MS cells (Section 2.2 and Section 2.3.1)—at each incubation condition of the CCRD runs were estimated by enumerating surviving cells (Table 1). S. Typhimurium inactivation was calculated as decimal logarithm reductions between the initial inoculum and the recovered cells—Log N₀/N.
The same 11 experimental conditions were used to assess the effect of each exposure condition in the BDL matrix on subsequent SGF S. Typhimurium survival (Table 1). After each experiment, 10 g of the BDL samples were homogenized using the stomacher with 90 mL of SGF and incubated at 37 ± 1 °C for 1 h. The SGF comprised 8.3 g proteose peptone (Sigma-Aldrich, Darmstadt, Germany); 2.05 g sodium chloride; 0.6 g potassium phosphate (Sigma-Aldrich, Darmstadt, Germany) 3.5 g D-glucose (Vetec Química Fina Ltd.a, RJ, Brazil), 0.11 g calcium chloride (Vetec Química Fina Ltd.a), 0.37 g potassium chloride (Tedia Company Inc, RJ, Brazil), 0.1 g lysozyme, and 13.3 mg pepsin (Thermo Fisher Scientific, MA, USA) per liter of distilled water [23 (link),24 (link),25 (link)]. S. Typhimurium enumerations were performed by plating an aliquot of the BDL-SGF cell suspension prior and after the 1 h incubation at 37 °C. S. Typhimurium inactivation was calculated by the logarithm difference from initial and final counts Log N₀/N.