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Dietary Fiber & Starch Analysis

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Dietary Fiber & Starch Analysis Dietary Fiber & Starch Analysis www.megazyme.com Contents This brochure features highlights from our range of analytical products. For the full catalogue, visit our website: www.megazyme.com Megazyme: Providing Analytical Solutions since 1988 Research is at the core of Megazyme’s product development. Continual innovation has allowed us to develop novel methodology and to introduce advances to accepted, industry-standard methods of analysis. Backed by best-in- class customer support, Megazyme products offer: • novel assays with excellent selectivity for the analyte in question • improved enzyme stability (resulting in a longer ‘shelf-life’) • unrivalled enzyme purity • reduced reaction times • extended cofactor stability Megazyme test kits - developed and manufactured in-house - have attracted worldwide acclaim for their novel methodologies and for the exceptional purity of the enzymes employed. Dietary Fiber Methods for the Measurement of Total Dietary Fiber 2 Choosing the Right Total Dietary Fiber Method 3 Assay procedure for the Rapid Integrated Total Dietary Fiber (RINTDF) Method 4 RINTDF: The Rapid Integrated Total Dietary Fiber Method 5 Total Dietary Fiber 6 Available Carbohydrates 7 Resistant Starch 9 b-Glucan 11 Polydextrose 12 Fructan and FOS 13 Starch Total Starch 15 Digestible Starch and Resistant Starch 16 Amylose/Amylopectin 18 Starch Damage 18 a-Amylase Activity 19 a-Amylase/b-Amylase Activity 19 b-Amylase Activity 20 Pullulanase/Limit Dextrinase Activity 20 Dietary Fiber Methods for the Measurement of Total Dietary Fiber Dietary fiber can generally be described as that portion of dietary carbohydrates that is not digested in the human small intestine. The most suitable analytical method to use will vary depending on which components are already known to be present within the sample. • HMWDF includes natural fibers such as cellulose, b-glucan, galactomannan and arabinoxylan • Resistant Starch comes from a variety of sources (see table below for types of Resistant Starch) • NDO includes inulin/FOS, GOS, polydextrose, resistant maltodextrin (such as Fibersol 2) Dietary Fiber Dietary Why does dietary fiber matter? Food manufacturers invest heavily into means of maximising the dietary fiber content of their products. However, resistant starch - a key dietary fiber component - is not measured accurately by the traditional methods used in dietary fiber analysis. Since the release of the internationally-recognised Codex definition of dietary fiber, Megazyme has developed two methods which address this issue. Our most advanced method - the Rapid Integrated Total Dietary Fiber (RINTDF) method - accurately measures all of the components of Total Dietary Fiber (as defined by Codex). Following interlaboratory evaluation, the RINTDF method has been adopted as AOAC Method 2017.16 and ICC Standard No. 185. The RINTDF method now stands as the new gold standard for comprehensive, accurate measurement of total dietary fiber in foods. Components of Total Dietary Fiber Comprehensive Total Dietary Fiber High Molecular Weight DF Low Molecular Weight DF Non-digestible Oligosaccharides “Classical” Fibers Resistant Starch (NDOs) • Cellulose • Fructooligosaccharides (FOS) • b-Glucan • RS (physically inaccessible starch) 1 • Galactooligosaccharides (GOS) • Galactomannan • RS (resistant starch granules) 2 • Polydextrose • Arabinoxylan • RS (retrograded starch) 3 • Resistant Maltodextrins (RMD) • Pectin • RS (phosphate-crosslinked starch) 4 • Xylooligosaccharides • Arabinogalactan Which methods accurately measure this component? RINTDF RINTDF RINTDF Prosky/Lee Prosky/Lee Prosky/Lee Matsutani Matsutani Matsutani 4 Choosing the Right Total Dietary Fiber Method Sample Details Information Required AOAC Method Megazyme Product Code Dietary Fiber Dietary A. Unknown TDF (HMWDF and SDFS) 2017.16 K-RINTDF IDF, SDFP and SDFS Modified 2017.16 K-RINTDF B. Devoid of RS HMWDF 985.29 (Prosky) K-TDFR / K-RINTDF IDF and SDFP 991.43 (Lee) K-TDFR / K-RINTDF HMWDF and SDFS 2001.03 (Matsutani) K-TDFR / K-RINTDF IDF, SDFP and SDFS 991.43 plus 2001.03 K-TDFR / K-RINTDF modification C. Devoid of HMWDF 985.29 (Prosky) K-TDFR / K-RINTDF NDO and RS IDF and SDFP 991.43 (Lee) K-TDFR / K-RINTDF Key to Acronyms: RS resistant starch NDO non-digestible oligosaccharides (or SDFS) TDF total dietary fiber IDF insoluble dietary fiber SDFP soluble dietary fiber that precipitates in the presence of 78% aqueous ethanol SDFS soluble dietary fiber which remains soluble in the presence of 78% aqueous ethanol HMWDF higher molecular weight dietary fiber which includes IDF and SDFP LMWDF lower molecular weight dietary fiber Notes: AOAC Method 2017.16 supersedes AOAC Method 2009.01. AOAC Method 2017.16 (modified to separately measure IDF and SDFP) supersedes AOAC Method 2011.25. 5 Purchase online at www.megazyme.com Assay procedure for the Rapid Integrated Total Dietary Fiber method: comparison of RINTDF with traditional methods Prosky/Lee + Matsutani RINTDF AOAC Method 985.29/991.43/2001.03 AOAC Method 2017.16 Heat-stable a-amylase 1. Enzyme Conditions: 98-100°C, pH 8.2, 30 min Pancreatic a-amylase Incubations Dietary Fiber Dietary Amyloglucosidase Protease Conditions: 37°C, pH 6.0, 4 h Conditions: 60°C, pH 7.5, 30 min Amyloglucosidase Protease Conditions: 60°C, pH 4.5, 30 min Conditions: 60°C, pH 8.2, 30 min 2. Alcohol precipitation, solvent washing, drying, filtration(2 residues) Protein Determination Ash Determination } 3. Ethanolic Filtrate 3. Ethanolic Filtrate Concentrate Concentrate Desalt in column In-line desalting (HPLC - TOSOH TSK) (HPLC - TOSOH TSK) “TDF” (HMWDF) HMWDF as defined by SDFS (NDO) as defined by RINTDF method (correct SDFS (NDO) } Prosky/Lee method } RS measurement) TDF as defined byMatsutani method TDF as defined byRINTDF method 6 RINTDF: The Rapid Integrated Total Dietary Fiber Method AOAC Method 2017.16 Dietary Fiber Dietary Product Code Product Name Pack Size Assay Kits K-RINTDF Rapid Integrated Total Dietary Fiber Assay Kit 100 assays Enzymes E-PANAA a-Amylase (Porcine pancreatic) 3 g 9 g E-AMGDFNG Amyloglucosidase (A. niger) Glycerol Free 20 mL 50 mL E-AMGDFPD Amyloglucosidase (A. niger) Powder 4 g E-BSPRPD Protease (Subtilisin A from B. licheniformis) Powder 1 g What’s different about Megazyme’s new RINTDF method? The Rapid Integrated Total Dietary Fiber Method (RINTDF) (AOAC Method 2017.16) is compared side-by-side with the traditional Prosky/Lee methods (AOAC Methods 985.29/991.43) in the figure opposite. The Prosky method measures only high molecular weight dietary fiber HMWDF( ), consisting of: • insoluble dietary fiber IDF( ) • soluble dietary fiber that precipitates in the presence of 78% v/v ethanol SDFP( ) • resistant starch (RS) which is underestimated In the Matsutani modification (AOAC Method 2001.03) of the Prosky method, non-digestible oligosaccharides (NDO) are also measured. NDO is soluble dietary fiber which remains soluble in the presence of 78% v/v ethanol SDFS( ). Addition of the HMWDF and SDFS fractions gives the total dietary fiber. Using the Prosky procedure, some of the resistant starch (RS2 and RS3) is hydrolysed under the extreme conditions required in the a-amylase incubation step. This leads to underestimation of some RS components, while RS4 (chemically crosslinked starch) is overestimated. The RINTDF method accurately measures all dietary fiber components, including resistant starch and non- digestible oligosaccharides. 7 Purchase online at www.megazyme.com Total Dietary Fiber Discover Megazyme’s bestselling range for total dietary fiber measurement Megazyme are experts in dietary fiber analysis, having developed two revolutionary AOAC-recognised methods for its measurement. Our dietary fiber range includes products suitable for use with all mainstream methods of dietary fiber analysis, recognising the 2009 Codex definition of total dietary fiber and methods designed for use with earlier definitions. AOAC Methods 2011.25/2009.01 Dietary Fiber Dietary Product Code Product Name Pack Size Assay Kits K-INTDF Integrated Total Dietary Fiber Assay Kit 100 assays Enzymes E-PANAA a-Amylase (Porcine pancreatic) 3 g 9 g E-AMGDF Amyloglucosidase (A. niger) 10 mL 40 mL 100 mL E-BSPRT Protease (Subtilisin A from B. licheniformis) 10 mL 40 mL 100 mL AOAC Methods 991.43/985.29 Our ultra-pure enzymes for use with this method are also available in formulations specifically designed for use with ANKOM auto-analysers. See our website (www.megazyme.com) for full details. Product Code Product Name Pack Size Assay Kits K-TDFR-100A Total Dietary Fiber Assay Kit 100 assays K-TDFR-200A 200 assays Enzymes E-BLAAM α-Amylase (B. licheniformis) 10 mL 40 mL 100 mL E-AMGDF Amyloglucosidase (A. niger) 10 mL 40 mL 100 mL E-BSPRT Protease (Subtilisin A from B. licheniformis) 10 mL 40 mL 100 mL 8 Available Carbohydrates Available carbohydrates is one of the key parameters required in nutritional tables. The UN’s Food and Agriculture Organisation (FAO) defines available carbohydrates as ”that fraction of carbohydrate that can be digested by human Dietary Fiber Dietary enzymes, is absorbed and enters into intermediary metabolism.” Available Carbohydrates Total Dietary Fiber Digestible Starch Total Starch Resistant Starch Maltodextrins Non-Digestible Oligosaccharides Sucrose ‘Classical’ Fibers D-Fructose D-Glucose Lactose In order to gain a complete picture of a foodstuff’s carbohydrate content, laboratories should measure and state the available carbohydrates alongside the total dietary fiber value for the sample. The available carbohydrates are defined
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