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Total Starch HK Assay Kit

Product code: K-TSHK

100 assays per kit

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Content: 100 assays per kit
Shipping Temperature: Ambient
Storage Temperature: Short term stability: 2-8oC,
Long term stability: See individual component labels
Stability: > 2 years under recommended storage conditions
Analyte: Total Starch
Assay Format: Spectrophotometer
Detection Method: Absorbance
Wavelength (nm): 340
Signal Response: Increase
Linear Range: 4 to 80 µg of D-glucose per assay
Limit of Detection: 1 g/100 g
Total Assay Time: ~ 90 min
Application examples: Cereal flours, food products and other materials.

The Total Starch HK (Hexokinase) Assay Kit for total starch determination in cereal flours and food products.

This assay kit contains an improved α-amylase that allows the amylase incubations to be performed at pH 5.0 (as well as pH 7.0). The method has been further modified by adjusting the D-glucose determination to a hexokinase/glucose-6-phosphate dehydrogenase/NADP+ based format.

See our full range of dietary and starch assay kits.

Scheme-K-TSHK TSHK Megayzme

  • Very competitive price (cost per test) 
  • All reagents stable for > 2 years after preparation 
  • Simple format 
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing 
  • Standard included
Certificate of Analysis
Safety Data Sheet
Assay Protocol Data Calculator Product Performance Validation Report
Megazyme publication
Measurement of carbohydrates in grain, feed and food.

McCleary, B. V., Charnock, S. J., Rossiter, P. C., O’Shea, M. F., Power, A. M. & Lloyd, R. M. (2006). Journal of the Science of Food and Agriculture, 86(11), 1648-1661.

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 successfully subjected to interlaboratory evaluation. All methods are based on the use of enzymes either purified by conventional chromatography or produced using molecular biology techniques. Such methods allow specific, accurate and reliable quantification of a particular component. Problems in calculating the actual weight of galactosyl-sucrose oligosaccharides in test samples are discussed in detail.

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Megazyme publication
Measurement of total starch in cereal products by amyloglucosidase-alpha-amylase method: collaborative study.

McCleary, B. V., Gibson, T. S. & Mugford, D. C. (1997). Journal of AOAC International, 80, 571-579.

An American Association of Cereal Chemists/AOAC collaborative study was conducted to evaluate the accuracy and reliability of an enzyme assay kit procedure for measurement of total starch in a range of cereal grains and products. The flour sample is incubated at 95 degrees C with thermostable alpha-amylase to catalyze the hydrolysis of starch to maltodextrins, the pH of the slurry is adjusted, and the slurry is treated with a highly purified amyloglucosidase to quantitatively hydrolyze the dextrins to glucose. Glucose is measured with glucose oxidase-peroxidase reagent. Thirty-two collaborators were sent 16 homogeneous test samples as 8 blind duplicates. These samples included chicken feed pellets, white bread, green peas, high-amylose maize starch, white wheat flour, wheat starch, oat bran, and spaghetti. All samples were analyzed by the standard procedure as detailed above; 4 samples (high-amylose maize starch and wheat starch) were also analyzed by a method that requires the samples to be cooked first in dimethyl sulfoxide (DMSO). Relative standard deviations for repeatability (RSD(r)) ranged from 2.1 to 3.9%, and relative standard deviations for reproducibility (RSD(R)) ranged from 2.9 to 5.7%. The RSD(R) value for high amylose maize starch analyzed by the standard (non-DMSO) procedure was 5.7%; the value was reduced to 2.9% when the DMSO procedure was used, and the determined starch values increased from 86.9 to 97.2%.

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Physicochemical characterization and microbiota modulatory potential of brewer’s spent grain and arabinoxylan-derived fractions: A valorization study.

Calvete-Torre, I., Sabater, C., Montilla, A., Moreno, F. J., Riestra, S., Margolles, A. & Ruiz, L. (2023). LWT, 185, 115107.

Brewer’s spent grain (BSG), one of the largest by-products generated by brewing industries, holds a wealthy nutritional value due to its content in proteins and fibres. Although some initiatives have explored alternatives for leveraging the nutritional value of BSG, it mostly remains an underexploited source of valuable ingredients for human nutrition, being mainly dedicated to animal feed and landfilling. BSG is rich in hemicellulosic material, composed of arabinoxylans (AX), which have been postulated as emergent prebiotics, capable to confer health benefits through modulating the gut microbiome. In this work we comprehensively investigated in vitro the composition and gut microbiome modulatory potential of BSG and AX fractions on human microbiota of two population groups by means of fecal batch fermentations and 16S rRNA gene sequencing. The gut microbiome modulatory effects was influenced by the specific BSG/AX fraction; the population group (healthy or Crohn’s disease (CD) patients) and by interindividual variation. For instance, Dorea and Eubacterium halli which include well-known fibre degrading, and health promoting bacteria, were only stimulated in fermentations of fractions with high arabinose and xylose contents. This work demonstrates diverse modulatory capacity of distinct AX fractions, diversifying alternative BSG valorization strategies through exploitation of novel prebiotics tailored to specific groups.

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Low-dosage enzymatic hydrolysis of organic municipal waste for sugar and ethanol production.

Rudnyckyj, S., Chaturvedi, T. & Thomsen, M. H. (2023). Biomass Conversion and Biorefinery, 1-17.

Enzymatic hydrolysis (EH) of the organic fraction of municipal solid waste (OFMSW) is challenging due to its complexity and heterogeneous character. This study investigated the effects of OFMSW compositional characteristics and pretreatment techniques on the efficiency of EH. The experiments involved varying enzymatic dosages and pretreatment methods, including heat pretreatment and anaerobic incubation of raw OFMSW. Remarkably, after thermal pretreatment, a saccharification yield exceeding 50% was achieved with only 1 filter paper unit (FPU) of Cellic® Ctec3 and 0.5 glucoamylase unit (AGU) of AMG® 300 L BrewQ per g of dry OFMSW. This resulted in the recovery of 239 g of sugar per kilogram of dry OFMSW. The study revealed that the decomposition of OFMSW by enzymes is limited after reaching about 50% of the sugar recovery yield, and high concentrations of inhibitors such as organic acids, alcohols, and potentially other organics and inorganics may contribute to this limitation. These compounds, however, serve as excellent preservatives, achieving contamination control during biological conversion. Sugar-rich hydrolysates were subjected to ethanol fermentation using Saccharomyces cerevisiae, leading to the conversion of most hexose sugars into ethanol, reaching 331 g of ethanol per kilogram of dry OFMSW, and indicating that the limitation of OFMSW saccharification was not due to product inhibition. These findings highlight the need for further investigation of the saccharification process of biowaste to overcome inhibitory effects and subsequently achieve higher efficiency of the following bioprocesses.

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Kabuli chickpea seed quality diversity and preliminary genome‐wide association study identifies markers and potential candidate genes.

Mugabe, D., Frieszell, C. M., Warburton, M. L., Coyne, C. J., Sari, H., Uhdre, R., Wallace, L., Ma, Y., Zheng, P., McGee, R. J. & Ganjyal, G. M. (2023). Agrosystems, Geosciences & Environment, 6(4), e20437.

Malnutrition due to macro- and micro-nutrient deficiencies is one of the major global health concerns, especially in developing countries. Using genomics-assisted breeding to enhance the nutritional value of important crops such as chickpea (Cicer arietinum L.) can help to address the problem. In this study, we conducted genome-wide association studies to identify genes associated with protein, starch, oil, and fiber in chickpea to create resources to speed the breeding process. The USDA kabuli chickpea mini-core of 88 accessions was genotyped using genotyped-by-sequencing, and 36,645 single nucleotide polymorphisms (SNPs) were identified across the eight chromosomes of the chickpea genome. A genome-wide marker-trait analysis using the FarmCPU model was conducted to identify SNP markers that can enable marker-assisted breeding for seed protein, fiber, oil, and starch concentrations. The most significantly associated markers for seed protein concentration (p = 8.82E-12), starch (p = 2.79E-12), fiber (p = 7.65E-12), and oil (p = 1.37E-08) were found on chromosomes 1, 2, 6, and 7, controlling 11%, 12%, 20%, and 16% of the phenotypic variation, respectively. Validation of the SNP markers in a broader set of plant genetic resources and environments will be needed to determine their usefulness in breeding for end-use characteristics.

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Mining the AE Watkins Wheat Landrace Collection for Variation in Starch Digestibility Using a New High-Throughput Assay.

Zafeiriou, P., Savva, G. M., Ahn-Jarvis, J. H., Warren, F. J., Pasquariello, M., Griffiths, S., Seung, D. & Hazard, B. A. (2023). Foods, 12(2), 266.

Breeding for less digestible starch in wheat can improve the health impact of bread and other wheat foods. The application of forward genetic approaches has lately opened opportunities for the discovery of new genes that influence the digestibility of starch, without the burden of detrimental effects on yield or on pasta and bread-making quality. In this study we developed a high-throughput in vitro starch digestibility assay (HTA) for use in forward genetic approaches to screen wheat germplasm. The HTA was validated using standard maize and wheat starches. Using the HTA we measured starch digestibility in hydrothermally processed flour samples and found wide variation among 118 wheat landraces from the A. E. Watkins collection and among eight elite UK varieties (23.5 to 39.9% and 31.2 to 43.5% starch digested after 90 min, respectively). We further investigated starch digestibility in fractions of sieved wholemeal flour and purified starch in a subset of the Watkins lines and elite varieties and found that the matrix properties of flour rather than the intrinsic properties of starch granules conferred lower starch digestibility.

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Aqueous ethanol organosolv process for the valorization of Brewer’s spent grain (BSG).

Parchami, M., Agnihotri, S. & Taherzadeh, M. J. (2022). Bioresource Technology, 362, 127764.

Brewers spent grain (BSG), the main solid byproduct of brewing, is annually generated by ca 37 million tons worldwide, which due to limited application, mostly ends up in landfills. This study aims to separate BSG’s fractions (lignin, cellulose, and hemicellulose) by ethanol organosolv pretreatment. Lignin-rich fractions were recovered using a two-step separation technique. The effects of temperature, retention time, and ethanol concentration on the quantity and quality of fractions were studied. The temperature considerably impacted the quality and quantity of obtained fractions, while other parameter effects greatly depended on the temperature. Substantial hemicellulose removal (90 %) along with lignin removal (56 %) and recovery (57 %) were obtained at 180°C. The highest lignin purity (95 %) was obtained at the pretreatment conditions of 180°C, 120 min, and 50 % ethanol concentration. This work provides an alternative route for BSG utilization, mitigating its environmental impact while enhancing the economy of a brewery.

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Effects of grape phylloxera leaf infestation on grapevine growth and yield parameters in commercial vineyards: a pilot study.

Wilmink, J., Breuer, M. & Forneck, A. (2022). OENO One, 56(1), 197-208.

Grape phylloxera (Daktulosphaira vitifoliae) can infest both roots and leaves of Vitis species. In commercial vineyards planted with Vitis vinifera scions grafted on rootstocks, grape phylloxera infestation is generally limited to root feeding. Vineyards are, however, increasingly subjected to vineyard-wide foliar infestations that last throughout the growing season. While some vineyards are affected by the infestation pressure of external leaf-feeding populations, other annually affected V. vinifera vineyards do not have these in their vicinity. Much is known about the damage potential of grape phylloxera root feeding; however, data on how phylloxera leaf infestation affects V. vinifera grapevines in commercial vineyards are lacking. This study, therefore, aimed to assess whether grapevine growth and yield are affected due to leaf infestation as it occurred in three commercial vineyards in the study area. Treatments were based on phylloxera leaf infestation and additional defoliation. Single-leaf carbon acquisition was measured with gas exchange analyses on healthy and galled leaves. Pruning weight and internode length were measured to assess the effect of leaf infestation and the effect of plant growth and vigour on leaf gall outbreaks. Yield quantity and quality were measured, and grapes were vinified for sensory analyses. Furthermore, using enzymatic analyses, non-structural carbohydrates were analysed in perennial wood. A significant decrease in sugar content in grapes (10 %) and starch reserves in perennial wood (11 %) was found in the most heavily infested vineyard. Grape must of infested plants in another vineyard furthermore showed a significantly higher level of titratable acid (7.5 %). Significant infestation effects seen in one vineyard were not significant in the other two vineyards. No significant differences were seen for carbon acquisition, harvest quantity, wine sensory analysis, pruning weight or internode length. The overall effect of phylloxera leaf infestation in the studied vineyards was, therefore, marginal. Grapevine vigour did not differ between infested vines, insecticide-sprayed vines, and vines on which no leaf infestation outbreaks took place. By analysing phylloxera leaf infestation under field conditions, these preliminary results form a basis for future long-term field studies about phylloxera leaf feeding on Vitis vinifera within the context of other biotic and abiotic plant stresses.

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Prebiotic Effect of Maitake Extract on a Probiotic Consortium and Its Action after Microbial Fermentation on Colorectal Cell Lines.

De Giani, A., Bovio, F., Forcella, M. E., Lasagni, M., Fusi, P. & Di Gennaro, P. (2021). Foods, 10(11), 2536.

Maitake (Grifola frondosa) is a medicinal mushroom known for its peculiar biological activities due to the presence of functional components, including dietary fibers and glucans, that can improve human health through the modulation of the gut microbiota. In this paper, a Maitake ethanol/water extract was prepared and characterized through enzymatic and chemical assays. The prebiotic potential of the extract was evaluated by the growth of some probiotic strains and of a selected probiotic consortium. The results revealed the prebiotic properties due to the stimulation of the growth of the probiotic strains, also in consortium, leading to the production of SCFAs, including lactic, succinic, and valeric acid analyzed via GC-MSD. Then, their beneficials effect were employed in evaluating the vitality of three different healthy and tumoral colorectal cell lines (CCD841, CACO-2, and HT-29) and the viability rescue after co-exposure to different stressor agents and the probiotic consortium secondary metabolites. These metabolites exerted positive effects on colorectal cell lines, in particular in protection from reactive oxygen species.

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Fortified blended foods prepared from fermented milk and cereal: Effect of storage conditions on composition, color, and pasting behavior.

Shevade, A. V., O’Callaghan, Y. C., O’Brien, N. M., O’Connor, T. P. & Guinee, T. P. (2021). Journal of Food Processing and Preservation, 45(5), e15419.

Fortified blended food powders (FBF) were prepared by blending fermented milk with parboiled wheat (FBFw), barley (FBFb), or oats (FBFo), incubating the blend, drying, milling, and fortifying the powder with vitamins, minerals, and refined soya oil. FBFs were stored at 15°C, 30°C, or 37°C for 0-18 months, and evaluated for compositional and functional properties. FBFo had lower contents of lactose and galactose, and higher contents of starch and fat, water-holding capacity (WHC) and pasting viscosity than FBFw or FBFb. Storage time and temperature affected composition (lactose, galactose, and lysine), color, WHC, pasting viscosity, and flow to a degree dependent on cereal type. FBFs stored at 15°C were generally stable over the 18 months, while those stored at 37°C underwent a rapid deterioration in color, WHC, and viscosity after storage times of ≤4 months. Storage of FBF at 15°C is recommended, whereas storage at 37°C should be avoided.

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Mixture design applied for formulation and characterization of vegetal-based fermented products.

Chekdid, A. A., Kahn, C. J., Prévot, E., Ferrières, M., Lemois, B., Choquet, C. & Linder, M. (2021). LWT, 146, 111336.

The present study consists on the characterization of six different flours in order to evaluate their ability to ferment alone and in mixtures. A discrepancy in the composition and physico-chemical characteristics of the flours was recorded. Three of these flours (oats, chickpea and coconut) were selected for the construction of the mixture design, which was used to formulate a base for a fermented vegetable dessert. Oat and chickpea flours have an effect on color accentuation (ΔE = 17.37) as well as on increasing water retention (75.31 g/100 g). During storage, post-acidification of the media to pH 4.42 was recorded, with a water holding capacity ranging from 56.64 to 66.12 g per 100 g, indicating a change in the texture of the mixtures. The fermented plant product showed a high viability of lactic acid bacteria, more than 107 CFU/mL for L. delbrueckii subsp. Bulgaricus and more than 108 CFU/mL for S. thermophiles after 30 days of storage. The use of flours in the formulation of fermented products was demonstrated as an alternative strategy for the development of new vegetable-based desserts.

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Starch and protein recovery from brewer's spent grain using hydrothermal pretreatment and their conversion to edible filamentous fungi-A brewery biorefinery concept.

Parchami, M., Ferreira, J. A. & Taherzadeh, M. J. (2021). Bioresource Technology, 125409.

This study aimed at recovering a highly concentrated starch and protein stream from the brewer’s spent grain (BSG). The effect of pretreatment temperature and retention time on the solubilization of starch and protein; and the generation of fermentation inhibitors were studied. Then, the application of recovered streams for fungal cultivation was evaluated using different edible fungi Aspergillus oryzae, Neurospora intermedia, and Rhizopus delemar. The hydrothermal pretreatment resulted in the highest solubilized starch concentration, 43 g/L, corresponding to 83% solubilization of initial BSG starch content. The highest protein concentration was 27 g/L (48% solubilization of initial BSG protein content). Cultivation with Neurospora intermedia on the recovered streams from the two best pretreatment conditions, 140°C for 4 h and 180°C for 30 min, resulted in pure fungal biomass with the highest protein content 59.62% and 50.42% w/w, respectively. Finally, a brewery biorefinery was proposed for the valorization of BSG.

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Soft Durum Wheat as a Potential Ingredient for Direct Expanded Extruded Products.

Gu, B. J., Kerr, C. J., Morris, C. F. & Ganjyal, G. M. (2021). Journal of Cereal Science, 103184.

Soft durum wheat was extruded to assess the influence of independent parameters (moisture content, screw speed, and barrel temperature each) on system responses (back pressure, torque value, and specific mechanical energy input) and product responses (expansion ratio, unit density, water absorption index (WAI) and water solubility index (WSI)). The lowest barrel temperature (120 °C) and moisture content (16%) with the highest screw speed (250 rpm) resulted in the highest specific mechanical energy input, expansion ratio, and water solubility index in the extrudates. Among the three independent parameters evaluated, moisture content was the most influential factor on the system and product responses. Energy efficiency on solubility (EES) was defined, and the notable impact of moisture content on EES was observed. Extrudates made from soft durum wheat had a reasonable expansion ratio, proving its potential usage in snack and cereal products.

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Solid-state fermentation of stale bread by an edible fungus in a semi-continuous plug-flow bioreactor.

Wang, R., Gmoser, R., Taherzadeh, M. J. & Lennartsson, P. R. (2021). Biochemical Engineering Journal, 169, 107959.

Stale bread, one of the major food wastes, holds great potential to produce new food products by solid-state fermentation using the edible filamentous fungus Neurospora intermedia. Yet, this process is limited to small-scale batch production. In this study, a plug-flow bioreactor was developed and successfully operated semi-continuously without addition of external inoculum. Two critical process parameters namely residence time and inoculum-to-substrate ratio (ISR) were studied in two experimental set-ups, 48 h and ISR 10:65 or 24 h and 20:55. The fermentation performance was assessed by the CO2 evolution rate and starch degradation in the substrate. Both experiments resulted in a relatively stable CO2 evolution rate up to 10 days and the starch content was reduced from 65 % in bread to 40.6 % and 43.2 %, respectively, in the product. Performance of the plug-flow bioreactor was compared to batch fermentation in tray bioreactors using the same ratio of inoculum. No significant differences of the final starch content were observed between the bioreactors, which indicate improved productivity by the semi-continuous process compared to the batch. The fungal biomass yield was calculated to range from 0.12 to 0.5 mol Cbiomass/mole Csubstrate. A material balance of the process revealed that 220 g dry bread and 200 ml water were required to produce 400 g fermented product with a fungal biomass content of 15 % in 24 h.

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Purification of Sucrose in Sugar Beet Molasses by Utilizing Ceramic Nanofiltration and Ultrafiltration Membranes.

Sjölin, M., Thuvander, J., Wallberg, O. & Lipnizki, F. (2020). Membranes, 10(1), 5.

Molasses is a sugar mill by-product with low value that today is used primarily for animal feed. However, molasses contains large amounts of sucrose which, if purified, could be used for other purposes. In this study, purification by membrane filtration using ceramic tubular ultrafiltration (UF) and nanofiltration (NF) was examined. NF purifies sucrose by removing small compounds, whereas UF removes larger compounds. Based on our results, high filtration fluxes could be obtained, and it was possible to clean the membranes sufficiently from fouling compounds. Sucrose was separated from other compounds, but the separation efficiency was generally higher with diluted molasses compared with concentrated molasses. This could be explained by more severe fouling when filtering dilute molasses or potentially due to aggregate formations in the molasses as our analysis showed. Overall, this study shows the potential of ceramic UF and NF membranes for sucrose purification from molasses.

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Safety Information
Symbol : GHS05, GHS08
Signal Word : Danger
Hazard Statements : H314, H334, H360
Precautionary Statements : P201, P202, P261, P264, P280, P284, P304+P340, P342+P311, P501
Safety Data Sheet
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