Ethanol (99%), acetone (≥99%), sulfuric acid (96%), oxygen peroxide (50%), and Nessler’s reagent were obtained from VWR chemicals (Leuven, Belgium). Celite, sodium hydroxide (1N solution), and hydrochloric acid (≥32%) were acquired from Sigma-Aldrich (Steinheim, Germany). MES hydrate (>99%) and ammonium sulfate were provided by Alfa Aesar (Kandel, Germany). TRIS (>99.8%) was obtained from Acros Organics (Geel, Belgium). Petroleum ether (60‒80 °C) was supplied by LAB-SCAN analytical sciences (Gliwice, Poland). Amylase thermostable (3000 U/mL), protease (9 tyrosine equivalent units/mg), and amyloglucosidase (3260 U/mL), suitable for AOAC International total dietary fiber and starch analytical procedures, were obtained from Megazyme (Te Huissen, The Netherlands). Ultrapure water was prepared in a Simplicity UV water purification system (Millipore, Molsheim, France).
Protease
Manufactured by Megazyme
Sourced in Ireland
Protease is a type of enzyme that catalyzes the breakdown or hydrolysis of proteins into smaller peptides or amino acids. It plays a crucial role in various biological processes and industrial applications.
Automatically generated - may contain errors
Lab products found in correlation
Amyloglucosidase, by Megazyme (4 mentions)
Endo arabinanase, by Megazyme (2 mentions)
α amylase, by Megazyme (2 mentions)
Sulfuric acid, by Avantor (1 mentions)
Acetone, by Avantor (1 mentions)
Ammonium sulfate, by Thermo Fisher Scientific (1 mentions)
Celite, by Merck Group (1 mentions)
Ethanol, by Avantor (1 mentions)
Nessler s reagent, by Avantor (1 mentions)
Simplicity uv water purification system, by Merck Group (1 mentions)
Hydrochloric acid (hcl), by Merck Group (1 mentions)
Cyanidin chloride, by Carl Roth (1 mentions)
Quercetin dihydrate, by Carl Roth (1 mentions)
Quercetin 3 o glucoside, by Carl Roth (1 mentions)
Cyanidin 3 o glucoside, by Carl Roth (1 mentions)
α amylase, by Novozymes (1 mentions)
Amyloglucosidase, by Novozymes (1 mentions)
Thermostable α amylase, by Merck Group (1 mentions)
Isoferulic acid, by Merck Group (1 mentions)
Luteolin 7 o glucoside, by Merck Group (1 mentions)
5 protocols using protease
1
Enzymatic Starch and Fiber Analysis
2
Chokeberry Pomace Powder Extrusion Trials
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Extrusion trials were performed using commercial chokeberry (Aronia melanocarpa) pomace powder (CPP) (Aronia Original Naturprodukte GmbH, Dresden, Germany) [20 (link)]. Chemicals and reagents of analytical purity grade were obtained from Merck KGaA (Darmstadt, Germany), unless stated otherwise. Amyloglucosidase (E-AMGDFPD, from Aspergillus niger, 36,000 U/g), α-amylase (E-PANAA, from pig pancreas 100,000 U/g), protease (E-BSPRPD from Bacillus licheniformis 9000 U/g), Celite 545 and endo-arabinanase (EC 3.2.1.99, from A. niger, 9 U/mg) were from Megazyme (Bray, Ireland) and Amberlite FPA53 and Ambersep 200 from Rohm and Haas Europe (Frankfurt, Germany). Thermostable α-amylase (Thermamyl 120 L, EC 3.2.1.1, from B. licheniformis, 120 KNU/g), protease (Alcalase 2.5 L, EC 3.4.21.62, from B. licheniformis, 2.5 AU/g), and Amyloglucosidase (AMG 300 L, EC 3.2.1.3, from A. niger, 300 AGU/g) were a gift from Novozymes, (Bagsværd, Denmark). Rotihydroquant C5 and D used for Karl Fischer titration as well as cyanidin-3-O-glucoside (≥96%), cyanidin chloride (≥97%), 5-caffeoylquinic acid (≥97%), quercetin-3-O-glucoside (≥99%), and quercetin dihydrate (≥99%) used as PP standards were obtained from Carl Roth GmbH & Co. KG (Karlsruhe, Germany). Ultrapure water was used for all experiments.
3
Enzymatic Hydrolysis of Polysaccharides
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If not stated otherwise, all chemicals used were of “p.a.” grade or better and were purchased from VWR (Radnor, PA, United States), Sigma-Aldrich (Schnelldorf, Germany), or Carl Roth (Karlsruhe, Germany). Thermostable α-amylase (EC 3.2.1.1, from Bacillus licheniformis, 20,000–60,000 U/mL) was purchased from Sigma. Protease (EC 3.4.21.14, subtilisin A from B. licheniformis, 300 U/mL), amyloglucosidase (EC 3.2.1.3, from Aspergillus niger, 200 U/mL), endo-arabinanase (EC 3.2.1.99, from A. niger, 9 U/mg), and endo-galactanase (EC 3.2.1.89, from A. niger, 408 U/mg) were purchased from Megazyme (Bray, Ireland).
4
Phenolic Standards Quantification Protocol
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Phenolic standards chlorogenic acid, caffeic acid, trans-cinnamic acid, gallic acid, ferulic acid, isoferulic acid, rutin, protocatechuic acid, luteolin-7-O-glucoside and p-coumaric acid, all other chemicals and HPLC-grade organic reagents were purchased from Sigma-Aldrich (Wicklow, Ireland).
The enzymes α-amylase, protease and amyloglucosidase were purchased from Megazyme (Wicklow, Ireland).
The enzymes α-amylase, protease and amyloglucosidase were purchased from Megazyme (Wicklow, Ireland).
5
Extraction and Fractionation of C. sinensis Polysaccharides
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The flowchart for the extraction and fractionation procedure was shown in Fig. 1a. Briefly, the powder of natural C. sinensis after exhaustively extracting with hot water was collected and dried.
It was then extracted with 0.5 mol•L -1 NaOH/0.01 mol•L -1 NaBH4 at 4 ºC two times, each for 12 h.
After centrifugation, all the supernatant was collected and neutralized with 1 mol•L -1 HAc. The solution was then centrifuged again to separate the supernatant, achieving the alkali extraction water-soluble fraction. After dialysis and precipitation with ethanol, a crude polysaccharide was obtained. Subsequently, the protein was removed by Sevag method (chloroform/1-butanol, v/v = 4:1) and protease (Megazyme, Ireland) hydrolysis, and then dialysis and further froze dry to get the alkali-extractable polysaccharide from natural C. sinensis. The alkali-extractable polysaccharide was then fractionated and purified by precipitating with ethanol repeatedly, and the supernatant, the major fraction, was collected for the following analysis.
It was then extracted with 0.5 mol•L -1 NaOH/0.01 mol•L -1 NaBH4 at 4 ºC two times, each for 12 h.
After centrifugation, all the supernatant was collected and neutralized with 1 mol•L -1 HAc. The solution was then centrifuged again to separate the supernatant, achieving the alkali extraction water-soluble fraction. After dialysis and precipitation with ethanol, a crude polysaccharide was obtained. Subsequently, the protein was removed by Sevag method (chloroform/1-butanol, v/v = 4:1) and protease (Megazyme, Ireland) hydrolysis, and then dialysis and further froze dry to get the alkali-extractable polysaccharide from natural C. sinensis. The alkali-extractable polysaccharide was then fractionated and purified by precipitating with ethanol repeatedly, and the supernatant, the major fraction, was collected for the following analysis.
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