Barry V McCleary

• PhD, DScAgr
• Consultant at FiberCarb Consultants

The Dietary Fiber (fiber) Terminology Roundtable was organized to address crucial issues concerning current definitions of dietary fiber and the pressing need to resolve inconsistencies and ambiguities in fiber terminology. This publication captures valuable insights and diverse perspectives from a multidisciplinary group of experts who span resear...

A broad range of AOAC official methods of analysis (OMA) have been developed and approved for the measurement of dietary fiber (DF) and DF components since the adoption of the Prosky method (OMA 985.29). OMA 985.29 and other OMA were developed to support the Trowell definition of DF. However, these methods do not measure DF as defined by the "new",...

Background: A simple, accurate, and reliable method to measure available carbohydrate components of food products, including cereal and dairy products, fruits, vegetables, processed food, food ingredients and animal foods, was developed by Megazyme (product K-AVCHO, Bray, Ireland). A single-laboratory validation of the enzymatic method resulted in...

Background: A simple, accurate and reliable method for the measurement of total dietary fiber (TDF) according to the Codex definition (2009) was developed and successfully validated as AOAC Official Method of Analysis (OMA) 2017.16. Subsequently, OMA 2017.16 was modified to allow separate measurement of soluble and insoluble dietary fiber fraction...

Background: The AOAC Stakeholder Panel on Strategic Food Analytical Methods issued a call for methods in 2018 for the measurement of lactose in low-lactose and lactose-free products under Standard Method Performance Requirements (SMPR®) 2018.009. Megazyme's Lactose Assay Kit (K-LOLAC) was reviewed and accepted as a First Action Official Method in...

The primary quality assessor of wheat grain is the Hagberg Falling Number (FN) method. This is a viscometric test surrogate for α-amylase activity. Despite being used for over sixty years, FN has been increasingly scrutinised due to its low throughput, poor reproducibility and inability to differentiate between the causes of low FN including Pre-ha...

The Codex Alimentarius Commission, a central part of the joint Food and Agricultural Organization/World Health Organizations Food Standards Program, adopts internationally recognized standards, guidelines, and code of practices that help ensure safety, quality, and fairness of food trade globally. Although Codex standards are not regulations per se...

The primary quality assessor of wheat grain produced globally is the Hagberg Falling Number (FN) method, which dates from 1960.1 This is essentially a viscometric test that measures the ability of α-amylase in wheat extract to depolymerise a starch based slurry. FN results for a given wheat sample set the price paid to the grower. Unfortunately, α-...

A Single Lab Validation was previously reported for Megazyme’s Lactose Assay Kit (K-LOLAC) with analysis performed on 36 different commercial food and beverage products and a set of 10 certified reference materials. Parameters examined during the validation included Working Range and Linear Range, Selectivity, Limit of Detection (LOD), Limit of Qua...

A simple method for measurement of the amylolytic activity of malt has been developed and fully evaluated. The method, termed the Maltose Value (MV) is an extension of previously reported work. Here, the MV method has been studied in detail and all aspects of the assay (sample grinding and extraction, starch hydrolysis, maltose hydrolysis and deter...

Background The AOAC Stakeholder Panel on Strategic Food Analytical Methods issued a call for methods for the measurement of lactose in low-lactose and lactose-free products under SMPR 2018.009 (1). Megazyme’s Lactose Assay Kit (K-LOLAC) was developed specifically to address the need for accurate enzymatic testing in lactose-free samples. Objective...

Background The level of available carbohydrates in our diet is directly linked to two major diseases; obesity and Type II diabetes. Despite this, to date there is no method available to allow direct and accurate measurement of available carbohydrates in human and animal foods. Objective The aim of this research was to develop a method that would a...

The Ethanol Assay Kit is an enzymatic test kit developed by Megazyme for the determination of ethanol in a variety of samples. The kit has been validated in a single laboratory for use with Kombucha fermented drinks, fruit juices and low-alcohol beer samples. The commercially available Ethanol Assay Kit (Megazyme catalogue no. K-ETOH) contains all...

Stages of the brewing process, such as mash separation to produce wort and beer filtration, can in certain cases prove problematic due to the increased viscosity caused by high levels of the non-starch polysaccharides, primarily β-glucan and arabinoxylan. Of these two polysaccharides, β-glucan has been extensively studied, but arabinoxylan has been...

Background and objectives The importance of selectively measuring available and unavailable carbohydrates in the human diet has been recognized for over 100 years. The levels of available carbohydrates in diets can be directly linked to major diseases of the Western world, namely Type II diabetes and obesity. Methodology for measurement of total ca...

Colourimetric assays were used to measure the activities of six key hydrolases endogenous to barley: β‐glucanase, xylanase, cellulase, α‐amylase, beta‐amylase and limit dextrinase. The analysed barley malt samples were previously characterised by 27 conventional malt quality descriptors. Correlations between enzymatic activities and brewing paramet...

Most commonly used methods for the measurement of starch in food, feeds and ingredients employ the combined action of α‐amylase and amyloglucosidase to hydrolyse the starch to glucose, followed by glucose determination with a glucose oxidase/peroxidase reagent. Recently, a number of questions have been raised concerning possible complications in st...

Traditional enzyme-based methods for measurement of fructan were designed to measure just inulin and branched-type (agave) fructans. The enzymes employed, namely exo-inulinase and endoinulinase, give incompletely hydrolysis of levan. Levan hydrolysis requires a third enzyme, endolevanase. This paper describes a method and commercial test kit (Megaz...

The cover image is based on the Research Article A novel enzymatic method for the measurement of lactose in lactose‐free products by David Mangan et al., DOI: 10.1002/jsfa.9317. This cover was supported by Megazyme. The cover image is based on the Research Article A novel enzymatic method for the measurement of lactose in lactose‐free products by D...

This chapter discusses the evolution of the definition of dietary fiber and the methodology to service this definition. Cereals are an important source of fiber in the human diet and therefore accurate analyses of the compounds that make up this dietary component are needed. The need to quantify the amount of carbohydrate that affects blood glucose...

BACKGROUND In recent years there has been a surge in the number of commercially available lactose‐free variants of a wide variety of products. This presents an analytical challenge for the measurement of the residual lactose content in the presence of high levels of mono‐, di‐, and oligosaccharides. RESULTS In the current work, we describe the dev...

A method for measurement of total dietary fiber (TDF) has been validated. This method is applicable to plant materials, foods, and food ingredients as consumed, consistent with the 2009 CODEX definition (ALINORM 09/32/REP), and measures insoluble dietary fiber (IDF) and soluble dietary fiber (SDF), comprising SDF that precipitates in the presence o...

you can find the full SMPR on the AOAC web site:- http://www.aoac.org/aoac_prod_imis/AOAC_Docs/SMPRs/SMPR_2018_001.pdf

A method for measurement of total dietary fiber (TDF) (1,2), as defined by the Codex Alimentarius Commission (ALINORM 09/32/26 [4]), was validated for plant materials, foods, and food ingredients. The method measures insoluble dietary fiber (IDF) and total soluble dietary fiber (SDF), including SDF that precipitates from 78% aqueous ethanol (SDFP)...

In the shadow of a burgeoning biomass-to-fuels industry, biological conversion of lignocellulose to fermentable sugars in a cost-effective manner is key to the success of second-generation and advanced biofuel production. For the effective comparison of one cellulase preparation to another, cellulase assays are typically carried out with one or mor...

endo-1,4-β-Xylanase (EC 3.2.1.8) is employed across a broad range of industries including animal feed, brewing, baking, biofuels, detergents and pulp (paper). Despite its importance, a rapid, reliable, reproducible, automatable assay for this enzyme that is based on the use of a chemically defined substrate has not been described to date. Reported...

We report herein the development of a novel assay procedure for the measurement of β-glucanase and lichenase (EC 3.2.1.73) in crude enzyme extracts. Two assay formats based on a) a direct cleavage or b) an enzyme coupled substrate were initially investigated. The ‘direct cleavage’ substrate, namely 4,6-O-benzylidene-2-chloro-4-nitrophenyl-β-3¹-cell...

An α-l-arabinofuranosidase of GH62 from Aspergillus nidulans FGSC A4 (AnAbf62A-m2,3) has an unusually high activity towards wheat arabinoxylan (WAX) (67 U/mg; k cat = 178/s, K m = 4.90 mg/ml) and arabinoxylooligosaccharides (AXOS) with degrees of polymerisation (DP) 3–5 (37–80 U/mg), but about 50 times lower activity for sugar beet arabinan and 4-n...

α-Amylase content in milled wheat is a key quality parameter in the baking industry. This metric is usually determined using the Hagberg Falling Number (FN) method, the results of which can be used to segment grains into a range of quality categories. The authors describe herein an alternative to the traditional FN assay, namely the Amylase SD assa...

Phytic acid, or myo-inositol hexakisphosphate, is the primary source of inositol and storage phosphorus in plant seeds and has considerable nutritional importance. In this form, phosphorus is unavailable for absorption by monogastric animals, and the strong chelating characteristic of phytic acid reduces the bioavailability of multivalent minerals...

endo-1,4-β-Glucanase (endo-cellulase, EC 3.2.1.4) is one of the most widely used enzymes in industry. Despite its importance, improved methods for the rapid, selective, quantitative assay of this enzyme have been slow to emerge. In 2014, a novel enzyme-coupled assay that addressed many of the limitations of the existing assay methodology was report...

A robust and reliable method has been developed for the measurement of β-glucan in mushroom and mycelial products. Total glucan (plus free glucose and glucose from sucrose) was measured using controlled acid hydrolysis with H2SO4 and the glucose released specifically was measured using glucose oxidase/peroxidase reagent. α-Glucan (starch/glycogen)...

The most commonly used method for the measurement of the level of endo-xylanase in commercial enzyme preparations is the 3,5-dinitrosalicylic acid (DNS) reducing sugar method with birchwood xylan as substrate. It is well known that with the DNS method, much higher enzyme activity values are obtained than with the Nelson-Somogyi (NS) reducing sugar...

Plant biomass is the major substrate for the production of biofuels and biochemicals, as well as food, textiles and other products. It is also the major carbon source for many fungi and enzymes of these fungi are essential for the depolymerization of plant polysaccharides in industrial processes. This is a highly complex process that involves a lar...

The new definition of dietary fibre introduced by Codex Alimentarius in 2008 includes resistant starch and the option to include non-digestible oligosaccharides. Implementation of this definition required new methodology. An integrated total dietary fibre method was evaluated and accepted by AOAC International and AACC International (AOAC Methods 2...

The quality of wheat for baking is critically dependent on the level of α-amylase (1,4-α-D-glucan glucanohydrolase, EC 3.2.1.1), which can be present as “late maturity α-amylase” (LMA), or due to pre-harvest sprouting due to high rainfall and humidity at the time of harvesting. The most commonly used method to measure α-amylase in wheat grain is th...

Certain interactions between carbohydrate active enzymes and polysaccharides involve surface binding sites (SBS) situated on catalytic domains outside of the active site. We recently undertook to develop a toolbox for SBS identification and characterization. In affinity gel electrophoresis (AGE) SBS containing proteins are migrating slower in nativ...

The measurement of limit-dextrinase (LD) (EC 3.2.1.142) in grain samples such as barley, wheat or rice can be problematic for a number of reasons. The intrinsic LD activity in these samples is extremely low and they often contain a limit-dextrinase inhibitor and/or high levels of reducing sugars. LD also exhibits transglycosylation activity that ca...

Glucans are polymers of D-glucose with differing linkages in linear or branched sequences. They are constituents of microbial and plant cell-walls and involved in important bio-recognition processes including immunomodulation, anti-cancer activities, pathogen virulence and plant cell-wall biodegradation. Translational possibilities for these activi...

Background The measurement of α-amylase (EC 3.2.1.1) in sprout-damaged grains is a crucial analysis yet a problematic one due to the typically low α-amylase levels in these samples. A number of standardized methods such as the Falling Number Method and Ceralpha Method exist which are routinely used for the assay of α-amylase. These methods, however...

AOAC Official Methods 2009.01 and 2011.25 have been modified to allow removal of resistant maltodextrins produced on hydrolysis of various starches by the combination of pancreatic α-amylase and amyloglucosidase (AMG) used in these assay procedures. The major resistant maltodextrin, 63,65-di-α-D-glucosyl maltopentaose, is highly resistant to hydrol...

The concept of dietary fibre has been evolving over the past 20 years from a chemical to a physiological focus. This has led to changes in the definition of dietary fibre and the inclusion of carbohydrate components that previously were not considered to be dietary fibre, and, consequently, were not measured, or were partially measured. The inclusi...

Several in vivo approaches have been adopted for the measurement of resistant starch, including: the hydrogen breath test; direct collection of ileal effluent from patients (ileostomy patients) who have had the colon removed; and direct collection of the ileal effluent from healthy subjects using a long triple lumen tube. AOAC method is subsequentl...

Several in vivo approaches have been adopted for the measurement of resistant starch, including: the hydrogen breath test; direct collection of ileal effluent from patients who have had the colon removed; and direct collection of the ileal effluent from healthy subjects using a long triple lumen tube. An integrated method for the measurement of tot...

Phosphate cross‐linked starch, referred to as resistant starch 4 (RS 4 ) is hydrolyzed much more rapidly under physiological conditions (i.e., pH 6, 37°C, pancreatic α‐amylase) than under the conditions used in AOAC dietary fiber method 985.29/AACC International Approved Method 32‐45.01 (the Prosky method), in which samples are incubated with therm...

The Codex Committee on Methods of Analysis and Sampling recently recommended 14 methods for measurement of dietary fiber, eight of these being type I methods. Of these type I methods, AACC International Approved Method 32-45.01 (AOAC method 2009.01) is the only procedure that measures all of the dietary fiber components as defined by Codex Alimenta...

Several procedures are available for the measurement of endo-1,4-β-glucanase (EG). Primary methods employ defined oligosaccharides or highly purified polysaccharides and measure the rate of hydrolysis of glycosidic bonds using a reducing-sugar method. However, these primary methods are not suitable for the measurement of EG in crude fermentation br...

Microarrays are powerful tools for high throughput analysis, and hundreds or thousands of molecular interactions can be assessed simultaneously using very small amounts of analytes. Nucleotide microarrays are well established in plant research, but carbohydrate microarrays are much less established, and one reason for this is a lack of suitable gly...

A method for the determination of insoluble (IDF), soluble (SDF), and total dietary fiber (TDF), as defined by the CODEX Alimentarius, was validated in foods. Based upon the principles of AOAC Official Methods 985.29, 991.43, 2001.03, and 2002.02, the method quantitates water-insoluble and water-soluble dietary fiber. This method extends the capabi...

Interest in dietary fiber is a consequence of the belief that di- etary fiber contributes positively to the health/quality of life of the consumer. The physiological effects of dietary fiber are what make it of interest to the consumer, food nutritionists, and regu- lators. Because dietary fiber is a multicomponent mixture, it is essential that the...

A method for the determination of total dietary fiber (TDF), as defined by the CODEX Alimentarius, was validated in foods. Based upon the principles of AOAC Official MethodsSM 985.29, 991.43, 2001.03, and 2002.02, the method quantitates high- and low-molecular-weight dietary fiber (HMWDF and LMWDF, respectively). In 2007, McCleary described a metho...

A method for the determination of total dietary fiber (TDF), as defined by the CODEX Alimentarius, was validated in foods. Based upon the principles of AOAC Official Methods 985.29, 991.43, 2001.03, and 2002.02, the method quantitates high- and low-molecular-weight dietary fiber (HMWDF and LMWDF, respectively). In 2007, McCleary described a method...

An integrated total dietary fibre (TDF) method, consistent with the recently accepted CODEX definition of dietary fibre, has been developed. The CODEX Committee on Nutrition and Foods for Special Dietary Uses (CCNFSDU) has been deliberating for the past 8 years on a definition for dietary fibre that correctly reflects the current consensus thinking...

Plant cell walls are constructed from a diversity of polysaccharide components. Molecular probes directed to structural elements of these polymers are required to assay polysaccharide structures in situ, and to determine polymer roles in the context of cell wall biology. Here, we report on the isolation and the characterization of three rat monoclo...

Fermentation of beta-glucan fractions from barley [average molecular mass (MM), of 243, 172, and 137 kDa] and oats (average MM of 230 and 150 kDa) by the human faecal microbiota was investigated. Fractions were supplemented to pH-controlled anaerobic batch culture fermenters inoculated with human faecal samples from three donors, in triplicate, for...

Total Dietary Fibre: IntroductionSpecific Dietary Fibre ComponentsConclusions References

IntroductionEnzymatic Hydrolysis of GalactomannansProperties of Industrially Produced Galactose-Depleted Guar Galactomannans (Enzyme-Modified Guars)Properties of Oligosaccharides Produced on Hydrolysis of Guar Gum and Enzyme-Modified Guar Samples by β-MannanaseConclusions References

A method is described for the measurement of dietary fibre, including resistant starch (RS), non-digestible oligosaccharides (NDO) and available carbohydrates. Basically, the sample is incubated with pancreatic alpha-amylase and amyloglucosidase under conditions very similar to those described in AOAC Official Method 2002.02 (RS). Reaction is termi...

Procedures for the measurement of starch, starch damage (gelatinised starch), resistant starch and the amylose/amylopectin content of starch, β-glucan, fructan, glucomannan and galactosyl-sucrose oligosaccharides (raffinose, stachyose and verbascose) in plant material, animal feeds and foods are described. Most of these methods have been successful...

Arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis releases only C3-linked arabinose residues from double-substituted xylose residues. A genomic library of B. adolescentis DSM20083 was screened for the presence of the axhD3 gene. Two plasmids were identified containing part of the axhD3 gene. The nucleotide sequences w...

With the recognition that resistant starch (RS) and nondigestible oligosaccharides (NDO) act physiologically as dietary fiber (DF), a need has developed for specific and reliable assay procedures for these components. The ability of AOAC DF methods to accurately measure RS is dependent on the nature of the RS being analyzed. In general, NDO are not...

The 'gold standard' method for the measurement of total dietary fibre is that of the Association of Official Analytical Chemists (2000; method 985.29). This procedure has been modified to allow measurement of soluble and insoluble dietary fibre, and buffers employed have been improved. However, the recognition of the fact that non-digestible oligos...

Interlaboratory performance statistics was determined for a method developed to measure the resistant starch (RS) content of selected plant food products and a range of commercial starch samples. Food materials examined contained RS (cooked kidney beans, green banana, and corn flakes) and commercial starches, most of which naturally contain, or wer...

This study was conducted to evaluate the method performance of a rapid procedure for the measurement of α-amylase activity in flours and microbial enzyme preparations. Samples were milled (if necessary) to pass a 0.5 mm sieve and then extracted with a buffer/salt solution, and the extracts were clarified and diluted. Aliquots of diluted extract (co...

This book contains 16 chapters by individual authors reviewing feed enzymes within the context of their mode of action, interaction with intestinal physiology, economic and environmental impacts, and application of the technology to the diets of various livestock species.