Total Starch HK Assay Kit

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.

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%.

A novel rotating disk bioreactor was designed and manufactured to produce bacterial cellulose (BC) using Komagataeibacter rhaeticus. Optimal conditions-45% disk submersion, mechanically etched disks with rotation speed of 20 rpm, spacing of 35 mm and 0.5 vvm air supply- achieved a yield of 1020 mg BC/disk with commercial glucose. Bread waste enzymatic hydrolysates improved BC production by 133.3%, highlighting the potential of waste valorization in sustainable biopolymer production. BC was further modified into nanostructures (BNCs) using H₂SO₄ (BNC1), H₂SO₄- HCl (BNC2), and cellulases (BNC3). FTIR spectra of BC and BNCs revealed typical cellulose vibration-bands while dynamic light scattering showed bimodal or trimodal size distributions (hydrodynamic radiuses of 60-2969 nm). TEM imaging of BNC1 and BNC2 demonstrated rodlike/needlelike nanostructure with widths of 33.1 ± 18.2 nm and 24.8 ± 14.2 nm respectively. BNC3 presented buddle of flat ribbons (width of 56.4 ± 26.3 nm). The maximum degradation temperature of BC (295°C) decreased after its ex-situ modification (271–294°C). The enhanced production and tailored structural modifications of BC highlight its potential for diverse applications in materials science. This transformative approach that integrates bread waste valorization and innovative bioreactor design paves the way for high-value advancements in biotechnology and environmental sustainability.

Carotenoids are essential pigments in plants, playing critical roles in photosynthesis, photoprotection, and stress tolerance, particularly under environmental conditions such as high light intensity and drought. To enhance β-carotene content in durum wheat (Triticum durum Desf.), a TILLING approach was used to generate null mutants for the lycopene ε-cyclase (LCYE) and β-hydroxylases 1 (HYD1) genes, which are key players in carotenoid biosynthesis. Homozygous mutants for both genes were obtained by crossing single homeoallelic mutant lines, resulting in three distinct mutant lines (LxH_1, LxH_2, LxH_3). Carotenoid metabolism and antioxidant-related genes expression were analyzed during seed ripening, revealing significantly reduced expression of LCYE and HYD1, while violaxanthin de-epoxidase (VDE) gene was upregulated at later stages. The mutant lines also showed significantly higher β-carotene accumulation in seeds, with an increase of up to 245% compared to the control, while lutein content was reduced by over 99%. In leaves, β-carotene levels remained unchanged, but zeaxanthin and violaxanthin accumulated at significantly higher levels compared to the control plants. Chlorophyll content was reduced in the mutant leaves, leading to altered chlorophyll a/b ratios and an overall decrease in total carotenoid levels. Although photosynthetic efficiency was lower in the mutants, gas exchange parameters remained unaffected, suggesting that primary carbon assimilation was not severely compromised. Phenotypic analysis revealed a reduction in plant height, spike length, and spikelet number; however, key yield traits were largely preserved. Notably, the mutant lines exhibited albinism under cold acclimation conditions, a phenotype absent in the control plants, likely due to the crucial role of lutein in photoprotection at low temperatures. These findings demonstrate that the pyramiding of mutations in LCYE and HYD1 effectively alters carotenoid composition, impacts photosynthesis-related traits, and influences plant responses to environmental stresses. This study provides valuable insights for breeding programs aimed at enhancing carotenoid content in wheat, with potential applications in improving both nutritional quality and stress resilience in cereal crops.

A two-stage pretreatment toward lignocellulosic biomass fractionation was devised. The process consisted of dilute acid hydrolysis using oxalic acid, followed by oxalic acid–assisted ethanol organosolv pretreatment. A biomass mixture consisting of four regional lignocellulosic materials, namely, brewer’s spent grain, tomato waste biomass, cucumber waste biomass, and spent coffee grounds, was used. In the first stage, the optimum mixture composition was determined using a full factorial design coupled with a simplex-centroid design. The interactive effects of the solid-to-liquid ratio, holding time, acid type, and concentration were also considered. In the attempt to lower solid yields and increase hemicellulose dissolution, elevated levels of furfural (15.73 g/L) and 5-hydroxymethylfurfural (8.56 g/L) were formed, due to increased pretreatment severity (180 min, 135C, 83.15 mg oxalic acid/g biomass, and 100 g biomass/L). The solid yield achieved was 50.53%. In the second stage, the effect of ethanol-to-water solvent ratio, holding time, and temperature were investigated using a central composite experimental design. Solid yields ranged between 72.57 and 85.20% (w/w), mainly due to lignin removal. Pretreatment with 75% (v/v) ethanol at 120 min and 190°C resulted in the highest lignin recovery (44.69%). Post-experimental verification runs were performed to evaluate the validity of the response surface models with a maximum error of 15.17%. Characterization by X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), thermogravimetric analysis (TGA), and heteronuclear single quantum coherence spectroscopy (HSQC) were conducted to assess biomass fraction integrity and structural changes during pretreatment.

Global warming will disturb dormancy release and growth resumption of trees. To better understand this process, it is important to investigate physiological and biochemical traits related to these stages. We examined dormancy release and growth resumption in Japanese cedar (Cryptomeria japonica [L.] D. Don), an evergreen needle-leaved tree, in the warm-temperate zone by evaluating budbreak under growth-promoting conditions, and simultaneously examining respiration rates and contents of carbohydrates and phytohormones in shoots from November 2022 to March 2023. A long time to budbreak and the lowest budbreak rates of 75% in November indicated shallow dormancy. Budbreak rates of 98%, short time to budbreak, and first appearance of budbreak in the field in March indicated growth resumption. Continuous changes in budbreak rates and time to budbreak between dormancy and growth resumption indicated dormancy was gradually released. Surges in budbreak rates in December indicated dormancy was almost completely released by early winter. Contents of abscisic acid (ABA) and salicylic acid (SA) decreased from November, remained low in March, and were strongly associated with budbreak rates according to principal component analysis. It was suggested that the depletion of SA led to the depletion of ABA, contributing to dormancy release and growth resumption. Fructose and trans-zeatin accumulated until February, and low levels of starch, indole-3-acetic acid, jasmonic acid, and jasmonic acid-isoleucine during winter was followed by accumulation in March. Although these biochemical traits were less related to budbreak rates compared to ABA and SA, they seemed to assist either dormancy release or growth resumption.

Background and Objective: This study aimed to assess the potential of incorporating ultrasonication treatment into the soaking process before germination as a method to enhance the nutritional profile of amaranth. While ultrasound has been used to expedite germination in various grains, its impact on germinated grains from a food science perspective remains underexplored. The research focused on the effects of ultrasonication on the composition, functional, and rheological properties of amaranth grains post-germination. Findings: The study revealed that germination alone significantly increased γ-aminobutyric acid (GABA), protein content, phenolic acids, amino acids, and total dietary fiber while reducing phytic acid and rheological parameters. Incorporating ultrasonication during soaking further elevated amino acids, GABA, protein, and phenolic acid levels. Notably, ultrasonication increased protein content by 25%, antioxidants by 35%, and dietary fiber by 20%, and attributed to cell wall breakdown and enhanced enzymatic activity during germination. Phytic acid levels decreased by up to 95% with prolonged ultrasonication, enhancing nutritional quality. Additionally, GABA levels revealed a significant rise, with an 82% and 87% increase in the Annapurna and Durga varieties, respectively. The study also found that ultrasonication led to reduced starch content, decreased pasting properties, and increased sugar content in the grains. Muffins made with ultrasonicated amaranth showed lower water activity levels, indicating better shelf stability, though specific volume decreased due to starch breakdown and increased amylase activity. Conclusion: The study demonstrates that ultrasonication combined with germination is an effective method for enhancing the nutritional and functional properties of amaranth grains. This method significantly boosts health-promoting components like GABA, proteins, and antioxidants while also affecting key baking quality parameters.

Understanding rye and wheat digestion is vital for evaluating impacts on nutrient availability and glycaemic responses. This study investigated the disintegration of processed high-fibre rye foods and refined wheat products during simulated intestinal digestion, aiming to link product characteristics with nutrient liberation. The overarching aim was to elucidate how these rye products contribute to the observed benefits in human intervention studies, particularly regarding satiety, weight loss, and metabolism. Analysis included four wholegrain rye products and three refined wheat products, spanning yeast-fermented breads and un-leavened cereal products. Microstructure examination revealed larger, partially intact digesta particles in wholegrain rye products after 120 min of digestion, alongside more aggregated and less degraded starch granules compared to refined wheat bread. Fermented rye bread exhibited greater degradation of subaleurone cell wall fragments than un-fermented rye bread. Viscosity assessments indicated lower viscosity for wheat products than for rye products, with yeast-fermented soft rye bread and rye crispbread showing notably lower viscosity than unfermented rye products. Post-digestion carbohydrate analysis uncovered higher glucose and maltose release during digestion for wheat products. PCA analysis confirmed negative correlations between glucose and maltose release and rye products, characterized by larger post-digestion bolus particles and higher dietary fibre. The elevated cell wall content in rye products acted as a protective barrier for starch granules, mitigating swelling and amylose release, explaining the observed viscosity differences between wheat and rye products and potentially influencing starch digestibility. Consequently, rye products undergo slower and less complete digestion than wheat, aligning with findings from human intervention studies.

This study demonstrates the sustainable advancement of fermentation media by blending the organic fraction of municipal solid waste (OFMSW) with organosolv beechwood cellulose. Investigations examined the effects of enzyme dosages and OFMSW integration into organosolv beechwood cellulose on sugar yield. The findings indicate that OFMSW inclusion and Cellic® CTec3 dosage significantly influence hydrolysis across two different batches of beechwood cellulose. Experimental data showed that OFMSW inclusion levels of 35% and 45% (w/w) produced sugar levels comparable to pure beechwood cellulose, achieving 58% to 68% (w/w) saccharification with sugar concentrations of 44 to 46 g/L. This highlights OFMSW's potential as a buffer substitute during the enzymatic conversion of organosolv cellulose. The resulting sugar-rich hydrolysates, derived from OFMSW-cellulose blends and pure cellulose, were evaluated for ethanol and cell biomass production using Saccharomyces cerevisiae and Mucor indicus, yielding 30 g of ethanol/L hydrolysate. Furthermore, OFMSW inclusion in beechwood cellulose proved to be an excellent alternative to synthetic nitrogen agents for S. cerevisiae cell production, reaching 12.2 g of biomass/L and surpassing the biomass concentration from cultivation on cellulose hydrolysate with nitrogen supplementation by threefold. However, M. indicus did not grow in the OFMSW-cellulose blend, suggesting that the inhibitory compounds of OFMSW may be a bottleneck in the proposed process. The present study demonstrates the benefits of incorporating OFMSW into cellulose material, as it enhances both cost-effectiveness and sustainability. This is attributed to the natural buffering properties and nitrogen content of OFMSW, which reduces the need for synthetic agents in fermentation-based lignocellulose biorefineries.

This work explores the upcycling of stale bread into bio-based, low-density porous materials with partial mesoporosity, produced through gelatinization and drying, using either supercritical CO₂ (aerogels) or low-vacuum conditions (xerogels). Cryogels were also fabricated via freeze-drying for comparison purposes. Stale bread particles (Bread) were subjected to proteolytic gluten depletion (Gluten-Depleted Bread, GDB) or particle size reduction (Finely milled Bread, FB) to investigate the effect of protein removal or particle size on porous materials’ properties. Porous materials made from wheat starch (WS) and wheat flour (Flour) were also examined for comparison. The solvent exchange induced volume shrinkage (SE-VS), which accounted for over 87% of the total shrinkage, ranged from 62% in GDB to 78% in WS. Bread-based porous materials presented comparable specific surface area (~40 m²/g) and water absorption capacity (~400%) to WS materials, but outperformed in resistance to volume shrinkage, resulting in lower density. FB porous materials possessed a higher specific surface area than Bread materials, indicating the benefits of particle size reduction. Furthermore, gluten depletion resulted in GDB-aerogels with the highest specific surface area (~80 m²/g), highlighting the benefits of gluten depletion. However, WS materials exhibited significantly greater maximum compressive stress (>2.0 MPa) and compressive modulus (>6 MPa) than stale bread-based porous materials. Importantly, the porous properties of xerogels and aerogels were similar (differences < 10%), indicating the feasibility of using low vacuum drying to produce new porous materials with partial mesoporosity (surface area 60–80 m²/g) from stale bread at a lower cost.

The combined drought and heat (DH) stress have devastating effects on plant physiological and biochemical processes, leading to poor yield. The aims of this study were to identify the physio-biochemical mechanisms employed by edamame (Glycine max L. Merrill) for survival and to establish better performing cultivars under DH stress. The impact of DH stress on the photosynthesis efficiency and osmolytes production in three edamame cultivars (UVE14, UVE17 and AGS429) was investigated. Non-destructive measurements were performed to determine the photosynthesis attributes, while pigments, non-structural carbohydrates (starch, glucose, sucrose, trehalose) and proline were extracted and quantified spectrophotometrically. The results showed that chlorophyll a (Chl-a), Chl-b, total chlorophyll and carotenoids in AGS429 were not affected under DH stress, which corresponded to increased normalised difference vegetative index (NDVI). Positive correlations between the carotenoids and total chlorophyll contents suggest that in AGS429, prevention of chlorophyll degradation under DH stress could be attributed to the increased carotenoids because they have antioxidative function. Additionally, AGS429 and UVE14 had increased trehalose, suggesting high osmotic adjustment under DH stress. An increase in starch production maintained glucose balance in AGS429 and UVE14, demonstrating higher photosynthetic efficiency compared to UVE17. The DH stress reduced photochemical reactions and carbohydrate accumulation in UVE17. This study shows that AGS429 and UVE14 protected the photosystems and photosynthetic pigments during DH stress, which led to higher photosynthetic capacity and accumulation of carbohydrates. Thus, maintaining the photosynthesis efficiency and carbohydrate metabolism processes in the AGS429 and UVE14 were adaptation features under DH stress.

The study investigated the potential of the organic fraction of municipal solid waste (OFMSW) for microbial biomass production. The compositional analysis of OFMSW showed richness in sugars, proteins, lipids, organic acids, and ethanol, suggesting promising cheap cultivation feedstock if inhibitory compounds are sustainably detoxified. The enzymatic hydrolysis with Cellic® CTec3 and AMG® 300 L BrewQ (Novozymes A/S) demonstrated excellent saccharification of sugar polymer, reaching 92% glucan hydrolysis and 70% xylan hydrolysis. However, higher enzymatic dosages led to a rise in the total organic acids content, potentially causing increased microbial inhibition. Full hydrolysate and hydrolysate after solids removal were cultivated with seven robust microbial strains. Cultivation on hydrolysate with solids showed consumption of sugars and organic acids solely by commercial backer yeast Saccharomyces cerevisiae. Removal of solids from hydrolysate resulted in increased performance of tested strains, showing consumption of measured organic acids and ethanol by S. cerevisiae, Yarrowia lipolytica DSM 8218, and Cutaneotrichosporon oleaginosus ATCC 20509. Remarkably, the investigation of biomass production revealed superior cell mass formation and detoxification by S. cerevisiae, resulting in 18.9 g of biomass/L hydrolysate with 50% of crude protein (w/w) in shake flasks and 13.2 g/L of hydrolase with 46% of crude protein (w/w) in a 5-L bioreactor. Furthermore, bioreactor cultivation confirmed organic acids and ethanol conversion into biomass, highlighting S. cerevisiae’s suitability for utilizing OFMSW for microbial biomass production. These findings contribute to advancements in biowaste-to-fodder conversion, promoting the development of a more sustainable circular economy.

Building up from our previous findings on deep eutectic solvents (DES) as reaction promoters for the acetylation of pure wheat starch, the current work explored combinations of reaction time, temperature and acetic anhydride: bread molar ratios to acetylate macromolecules within bread particles relying solely on macromolecule solvation and the slightly basic environment provided by the eutectic mixture. High degree of substitution with acyl groups (DS_acyl, 0.73–1.09 as quantified by ¹H NMR and confirmed by ¹³C NMR, and FTIR) was achieved within a short timeframe of 36 min and < 16 acetic anhydride:bread molar ratio. Spectroscopy discarded a major presence of side-reaction products starch carbamates and acetyl urea in reacted samples. Nevertheless, DES acetylation resulted in reacted samples with starch weight average molecular weight (M_w) ranging from 2.62 × 10⁶ to 1.39 × 10⁶ g/mol for the most degraded sample and the partial wash-off of starch and cell wall polysaccharides, which resulted in increased gluten content. Notably, TGA (coupled to real-time FTIR analysis of the evolved gases) suggested a potential enhancement in hydrophobicity and glass transition temperature (T_g) in the reacted samples, with a positive correlation between DS_acyl and T_g. These findings emphasize the potential of DES as reaction promoters in complex starchy matrices for esterification reactions.

The global average daily minimum temperatures are increasing at a quicker pace than the average daily maximum temperatures, which are predicted to increase in severity impacting global food production. This study focuses on elucidating the physiological and transcriptional response to high night-time temperature (HNT) stress in 12 US commercial maize (Zea mays) hybrids using unique field-based infrastructure. Our experimental objectives were to (i) impose an accurate and uniformly distributed post-flowering HNT stress of +4.0°C until physiological maturity, (ii) quantify the impact of HNT stress on physiological and yield-related traits, (iii) establish the impact on end-use quality of maize kernels formed under HNT stress, and (iv) analyze the differential expression of genes involved in grain starch metabolism. Accurate and uniformly distributed HNT stress of 3.8°C higher than the ambient night-time temperature throughout the grain-filling period reduced yield (−14%), kernel weight (−8%), and significantly reduced kernel nutrient content, specifically magnesium in the susceptible hybrids. HNT significantly increased the expression of key genes involved in starch metabolism in the tolerant hybrid. Although HNT stress had a negative impact on yield and quality in field grown maize, two hybrids had physiological and transcriptional regulation that favored higher level of resilience which lays the platform for developing climate smart maize hybrids.

Hawthorn fruits (Crataegus pinnatifida) present high content of high-methoxyl pectin, able to gel under high-sugar acidic conditions. In this work, the proximate and phytochemical composition of two cultivars of hawthorn fruit and the gelling ability of their unrefined (not further processed) dried powders and their extracted pectins were evaluated and systematically compared with citrus pectins (CP1 and CP2). Mianqiu (MI), a less known cultivar, showed two-fold higher pectin content and titratable acidity than Dajinxing (DA), one of the most common cultivars. DA showed higher starch, insoluble dietary fiber, pasting viscosity and total and extractable (EPP) phenolic compounds. EPP content was almost two-fold higher in DA than MI, resulting in stronger antioxidant properties. All high-methoxyl sugar acid gels exhibited a predominantly elastic response. MI resulted in hawthorn-powder gels with higher elastic modulus (G′) after gel-making (initially stronger gels), and lower G′ increase during storage (hardening) than DA. Citrus pectins (CP2 > CP1) showed higher gel-strength and faster gelling ability than hawthorn pectin gels (DA > MI) based on the lower G’ and lack of gel formation after 90 min of cooling in hawthorn pectin-based gels. The gelation results were closely linked to the starch-to-pectin ratio, purity, and degree of methyl esterification.