Prebiotic Potential of Cellulase-Xylanase Treated Guava By-Products
We applied commercial fungi-origin cellulase and xylanase separately or in combination on two types of guava puree by-products and studied the prebiotic activity of the ethanolic extract (EEC) of the treated sample. We also added the untreated and treated whole guava by-products as a prebiotic source into UHT fresh milk containing a yogurt starter culture to investigate the practical application of these guava by-products. Two types of guava purée by-products: refiner (the seed-rich fraction), and decanter (the pulp-rich fraction) were washed twice to ensure most of the sugar were removed (1), and then treated with 150 U xylanase or cellulase separately or in combination at a ratio of 1:2 (xylanase to cellulase) to investigate the prebiotic potential. The enzymatic treatments were labeled as 150 XY (xylanase); 150 CE (cellulase), 150 CX (combined cellulase-xylanase), and CT (control containing buffer without enzyme, termed as untreated hereafter). The reducing sugar amount liberated into the effluent reflects the extent of hydrolysis followed the order 150 XY < 150CE < 150CX (2). The ethanolic extract (EEC) derived from the untreated and enzymatic-treated by-products contained mainly rhamnose and xylose and were found the most in the combined enzymatic-treated samples. (3) The total soluble carbohydrates content of the EEC followed the order of CT < 150XY < 150CE < 150CX (4). Enzymatic treatments also led to the increment of the EEC's prebiotic activity score tested on Lactobacillus brevis, Lactobacillus rhamnosus, and Lactobacillus plantarum. (5) The incorporation of the whole guava by-products into yogurt starter culture-containing UHT fresh milk to form yogurt has caused a significant increment of the log CFU/mL (6), decrease in pH value (7), and enhancement in hardness, stickiness, and adhesiveness (7), with the most prominent effect exerted by the combined enzymatic-treated guava by-products. Thus, combined cellulase-xylanase treatment can be a good option to valorize guava purée by-products as prebiotic sources for food applications. 1. Reducing sugar content in 1st washing and 2nd washing of the guava by-products. 2. Reducing sugar content in the effluent after enzymatic treatments. 3. Different sugar amounts in the ethanolic extract (EEC) of differently treated guava by-products. 4. Total soluble carbohydrate content in the ethanolic extract (EEC) of differently treated guava by-products. 5. Data of prebiotic activity score of the ethanolic extract (EEC) 6. Total plate count of yogurt incorporated with differently treated guava by-products 7. Textural properties and pH of yogurt incorporated with differently treated guava by-products
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1. The washed solution was analyzed using 3.5-dinitrosalicylic acid (DNS) assay based on Miller (1959) 2. The effluent after enzymatic treatment was analyzed using the Nelson Somogyi method (Nelson, 1944; Gusakov et al. 2011) 3.high-performance liquid chromatography (HPLC) system equipped with a Shimadzu LC-20AD pump, a refractive index detector (RID-10A), and a manual injector was used together with an NH2 Purosphere® STAR column (Merck, United States) operated at 30℃. The mobile phase used was 80% acetonitrile, while the flow rate was 0.8 mL/min and 0.2 mL/min for Pump A and B. We injected 10 μL filtered and suitably diluted sample into the HPLC column to generate a chromatogram for the sugar profile analysis. 4. The total soluble carbohydrate content determination using phenol-sulphuric method (Dubios, 1956) 5. The prebiotic activity score assay was carried out based on Lim et al. (2018), the following formula was applied: Prebiotic activity score = [ ((Probiotic O.D.on prebiotic at 24 h - Probiotic O.D.on prebiotic at 0 h))/((Probiotic O.D.on glucose at 24 h - Probiotic O.D.on glucose at 0 h)) ] – [ ((Enteric O.D.on prebiotic at 24 h - Enteric O.D.on prebiotic at 0 h))/((Enteric O.D.on glucose at 24 h - Enteric O.D.on glucose at 0 h)) ] 6. A serially diluted yogurt sample was transferred aseptically to a 9 cm-diameter petri dish and added with sterile Luria-Bertani (LB) agar, mixed well and allowed to solidify. After incubated at 37℃ for 24 h, visible colonies between 30 and 300 in each sample plate were enumerated, and the average was expressed as log CFU/mL. 7. The pH of the formed yogurt was measured at a ratio of 1: 1 (yogurt: deionized water) using a calibrated pH meter. Textural attributes of yogurt were measured by using a TA.XTplus Texture Analyzer (Stable Micro Systems, United Kingdom) equipped with a load cell of 5 kg and a 25 mm-diameter cylindrical probe, with a speed setting of 20 mm/s, by the software Texture Exponent Programs. The hardness (mm), stickiness (g), stringiness (mm) and adhesiveness (g.s) were analyzed from the texture profile generated.