Insights into the Supramolecular Structure and Degradation Mechanisms of Starch from Different Botanical Sources as Affected by Extrusion-based 3D Printing.

Extrusion-based 3D printing has emerged as the most versatile additive manufacturing technique for the printing of practically any material. However, 3D printing of functional materials often activates thermo-mechanical degradation, which affects the 3D shape quality. Herein, we describe the structural changes of eight different starch sources (normal or waxy) as a consequence of the temperature of an extrusion-based 3D printing system through in-depth characterization of their molecular and structural changes. The combination of size-exclusion chromatography, small-angle X-ray scattering, X-ray diffraction, dynamic viscoelasticity measurements, and in vitro digestion has offered an extensive picture of the structural and biological transformations of starch varieties. Depending on the 3D printing conditions, either gelatinization was attained ("moderate" condition) or single-amylose helix formation was induced ("extreme" condition). The stiff amylopectin crystallites in starch granules were more susceptible to thermo-mechanical degradation compared to flexible amorphous amylose. The crystalline morphology of the starch varieties varied from B-type crystallinity for the starch 3D printing at the "moderate" condition to a mixture of C- and V-type crystallinity regarding the "extreme" condition. The "extreme" condition reduced the viscoelasticity of 3D-printed starches but increased the starch digestibility rate/extent. In contrast, the "moderate" condition increased the viscoelastic moduli, decreasing the starch digestion rate/extent. This was more considerable mainly regarding the waxy starch varieties. Finally, normal starch varieties presented a well-defined shape fidelity, being able to form a stable structure, whereas waxy starches exhibited a non-well-defined structure and were not able to maintain their integrity after printing. The results of this research allow us to monitor the degradability of a variety of starch cultivars to create starch-based 3D structures, in which the local structure can be controlled based on the 3D printing parameters.

Natural Polymorphisms in Arabidopsis Result in Wide Variation or Loss of the Amylose Component of Starch.

Starch granules contain two Glc polymers, amylopectin and amylose. Amylose makes up approximately 10% to 30% (w/w) of all natural starches thus far examined, but mutants of crop and model plants that produce amylose-free starch are generally indistinguishable from their wild-type counterparts with respect to growth, starch content, and granule morphology. Since the function and adaptive significance of amylose are unknown, we asked whether there is natural genetic variation in amylose synthesis within a wild, uncultivated species. We examined polymorphisms among the 1,135 sequenced accessions of Arabidopsis (Arabidopsis thaliana) in GRANULE-BOUND STARCH SYNTHASE (GBSS), encoding the enzyme responsible for amylose synthesis. We identified 18 accessions that are predicted to have polymorphisms in GBSS that affect protein function, and five of these accessions produced starch with no or extremely low amylose (< 0.5% [w/w]). Eight further accessions had amylose contents that were significantly lower or higher than that of Col-0 (9% [w/w]), ranging from 5% to 12% (w/w). We examined the effect of the polymorphisms on GBSS function and uncovered three mechanisms by which GBSS sequence variation led to different amylose contents: (1) altered GBSS abundance, (2) altered GBSS activity, and (3) altered affinity of GBSS for binding PROTEIN TARGETING TO STARCH1-a protein that targets GBSS to starch granules. These findings demonstrate that amylose in leaves is not essential for the viability of some naturally occurring Arabidopsis genotypes, at least over short timescales and under some environmental conditions and open an opportunity to explore the adaptive significance of amylose.

Influence of the addition of different ingredients on the bioaccessibility of glucose released from rice during dynamic in vitro gastrointestinal digestion.

Rice represents a primary source of carbohydrates in human nutrition. Upon its consumption, the released sugars are mostly absorbed, categorising rice as a high glycemic index food. Addition of ingredients is common practice when cooking rice, which may affect rice digestibility and influence nutrients absorption in the gastrointestinal (GI) tract, enabling a controlled glucose release. In this sense, rice formulations were submitted to a dynamic in vitro GI model, constituted by reactors that simulates peristalsis coupled to filtration membranes, to evaluate carbohydrates hydrolysis and bioaccessibility. Addition of quinoa and wholegrains reduced carbohydrates hydrolysis (i.e. 38.5 ± 5.08% and 57.98 ± 1.91%, respectively) and glucose bioaccessibility (i.e. 25.92 ± 5.70% and 42.56 ± 1.39%, respectively) when compared with brown rice (i.e. 63.86 ± 2.96% hydrolysed and 44.33 ± 1.88% absorbed). Addition of vegetables significantly decreased sample chewiness and resulted in superior hydrolysis (71.75 ± 7.44%) and glucose absorption (51.61 ± 6.25%).

Starch Components, Starch Properties and Appearance Quality of Opaque Kernels from Rice Mutants.

Rice mutants with altered starch components and properties are important genetic resources in rice breeding programmes. In this study, 44 mutants with altered starch components were screened from 135 rice mutants with opaque kernels using a starch-iodine absorption spectrum method, and nine mutants from them were further selected for investigating their starch properties and kernel appearance quality. The results showed that the iodine absorption spectrum parameters, OD620, OD620/550, and λmax, could reflect the changes of starch components in rice mutants, and had significantly positive relationships with amylose content and negative relationships with the proportion of short branch-chains of amylopectin. The endosperm starches from nine mutants all showed A-type crystalline structure and similar short-range ordered structure, but had different relative crystallinities. The changes of starch components in mutants not only resulted in the different gelatinization properties of starch but also changed the appearance quality of brown rice kernels. This study provided abundant genetic plants for studying the molecular mechanism of starch synthesis and the quality regulation of rice kernels.

Unambiguous Ex Situ and in Cell 2D 13C Solid-State NMR Characterization of Starch and Its Constituents.

Starch is the most abundant energy storage molecule in plants and is an essential part of the human diet. This glucose polymer is composed of amorphous and crystalline domains in different forms (A and B types) with specific physicochemical properties that determine its bioavailability for an organism, as well as its value in the food industry. Using two-dimensional (2D) high resolution solid-state nuclear magnetic resonance (SS-NMR) on 13C-labelled starches that were obtained from Chlamydomonas reinhardtii microalgae, we established a complete and unambiguous assignment for starch and its constituents (amylopectin and amylose) in the two crystalline forms and in the amorphous state. We also assigned so far unreported non-reducing end groups and assessed starch chain length, crystallinity and amylose content. Starch was then characterized in situ, i.e., by 13C solid-state NMR of intact microalgal cells. Our in-cell methodology also enabled the identification of the effect of nitrogen starvation on starch metabolism. This work shows how solid-state NMR can enable the identification of starch structure, chemical modifications and biosynthesis in situ in intact microorganisms, eliminating time consuming and potentially altering purification steps.

WB1, a Regulator of Endosperm Development in Rice, Is Identified by a Modified MutMap Method.

Abnormally developed endosperm strongly affects rice (Oryza sativa) appearance quality and grain weight. Endosperm formation is a complex process, and although many enzymes and related regulators have been identified, many other related factors remain largely unknown. Here, we report the isolation and characterization of a recessive mutation of White Belly 1 (WB1), which regulates rice endosperm development, using a modified MutMap method in the rice mutant wb1. The wb1 mutant develops a white-belly endosperm and abnormal starch granules in the inner portion of white grains. Representative of the white-belly phenotype, grains of wb1 showed a higher grain chalkiness rate and degree and a lower 1000-grain weight (decreased by ~34%), in comparison with that of Wild Type (WT). The contents of amylose and amylopectin in wb1 significantly decreased, and its physical properties were also altered. We adopted the modified MutMap method to identify 2.52 Mb candidate regions with a high specificity, where we detected 275 SNPs in chromosome 4. Finally, we identified 19 SNPs at 12 candidate genes. Transcript levels analysis of all candidate genes showed that WB1 (Os04t0413500), encoding a cell-wall invertase, was the most probable cause of white-belly endosperm phenotype. Switching off WB1 with the CRISPR/cas9 system in Japonica cv. Nipponbare demonstrates that WB1 regulates endosperm development and that different mutations of WB1 disrupt its biological function. All of these results taken together suggest that the wb1 mutant is controlled by the mutation of WB1, and that the modified MutMap method is feasible to identify mutant genes, and could promote genetic improvement in rice.

Structures, physicochemical properties, and applications of amaranth starch.

Amaranth as a rediscovered "new" crop is becoming a research focus in the recent two decades. The major carbohydrate of some amaranth species is starch, which accounts up to around 60% of the dry grains. This review summarizes the present knowledge of the isolation, composition, structures, physiochemical properties, modifications, and applications of amaranth starches, and provides suggestions for research to further improve the utilization.

Dietary starch types affect liver nutrient metabolism of finishing pigs.

This study aimed to evaluate the effect of different starch types on liver nutrient metabolism of finishing pigs. In all ninety barrows were randomly allocated to three diets with five replicates of six pigs, containing purified waxy maize starch (WMS), non-waxy maize starch (NMS) and pea starch (PS) (the amylose to amylopectin ratios were 0·07, 0·19 and 0·28, respectively). After 28 d of treatments, two per pen (close to the average body weight of the pen) were weighed individually, slaughtered and liver samples were collected. Compared with the WMS diet, the PS diet decreased the activities of glycogen phosphorylase, phosphoenolpyruvate carboxykinase and the expression of phosphoenolpyruvate carboxykinase 1 in liver (P0·05). Compared with the WMS diet, the PS diet reduced the expressions of glutamate dehydrogenase and carbamoyl phosphate synthetase 1 in liver (P<0·05). PS diet decreased the expression of the insulin receptor, and increased the expressions of mammalian target of rapamycin complex 1 and ribosomal protein S6 kinase ß-1 in liver compared with the WMS diet (P<0·05). These findings indicated that the diet with higher amylose content could down-regulate gluconeogenesis, and cause less fat deposition and more protein deposition by affecting the insulin/PI3K/protein kinase B signalling pathway in liver of finishing pigs.

Development of formulae for estimating amylose content and resistant starch content based on the pasting properties measured by RVA of Japonica polished rice and starch.

We searched for the easy and simple method to measure the novel indicators which reflect not only AAC, but also (RS) based on pasting properties using RVA. Novel indexes such as SB/Con and Max/Fin (Maximum viscosity/Minimum viscosity) ratios had a very high correlation with proportion of intermediate and long chains of amylopectin; Fb1+2+3 (DP â‰§ 13). In Japonica polished rice, estimation formulae for AAC and RS content were developed using novel indexes based on pasting properties by RVA, and these equations showed determination coefficients of 0.89 and 0.80 for calibration and 0.71 and 0.75 for validation test. We developed the estimation formulae for AAC and RS content for Japonica starch samples. These equations showed determination coefficients of 0.86 and 1.00 for calibration and 0.76 and 0.83 for validation test, which showed that these equations can be applied to the unknown rice samples.

Amylose content decreases during tuber development in potato.

BACKGROUND: Potato starch is composed primarily of amylopectin and amylose in an approximately 3:1 ratio. Amylose is considered to be nutritionally desirable in North American and European markets, so there is interest in finding strategies to increase the amylose content of potato starch. There is also interest in marketing 'baby' potatoes, which are harvested when they are physiologically immature. This study was carried out to determine weekly changes in amylose content in potato tubers of 11 North American cultivars during the growing season. The trial was repeated across 3 years. RESULTS: We determined that amylose content is highest early and it decreases in a linear fashion as the growing season progresses. Mean amylose content across cultivars and years declined from 30.0% in late June to 26.8% in late August. The rate of decrease varied across years, with slopes of linear regression plots ranging from -0.17 in 2012 to -0.74 in 2011. Amylose content in tuber starch varied among cultivars, with the highest levels observed in Ranger Russet (30.7%) and White Pearl (31.6%); it was lowest in Kennebec (25.7%) and Langlade (25.6%). CONCLUSIONS: This study adds to a growing body of literature on the nutritional value of immature potato tubers. In addition to having higher levels of some phytonutrients, as reported in other studies, immature tubers have a higher proportion of amylose in the starch. This is nutritionally desirable in affluent regions where high fiber content is more important than calories from carbohydrates. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

A systematic review of the influence of rice characteristics and processing methods on postprandial glycaemic and insulinaemic responses.

Rice is an important staple food for more than half of the world's population. Especially in Asian countries, rice is a major contributor to dietary glycaemic load (GL). Sustained consumption of higher-GL diets has been implicated in the development of chronic diseases such as type 2 diabetes mellitus. Given that a reduction in postprandial glycaemic and insulinaemic responses is generally seen as a beneficial dietary change, it is useful to determine the variation in the range of postprandial glucose (PPG) and insulin (PPI) responses to rice and the primary intrinsic and processing factors known to affect such responses. Therefore, we identified relevant original research articles on glycaemic response to rice through a systematic search of the literature in Scopus, Medline and SciFinder databases up to July 2014. Based on a glucose reference value of 100, the observed glycaemic index values for rice varieties ranged from 48 to 93, while the insulinaemic index ranged from 39 to 95. There are three main factors that appear to explain most of the variation in glycaemic and insulinaemic responses to rice: (1) inherent starch characteristics (amylose:amylopectin ratio and rice cultivar); (2) post-harvest processing (particularly parboiling); (3) consumer processing (cooking, storage and reheating). The milling process shows a clear effect when compared at identical cooking times, with brown rice always producing a lower PPG and PPI response than white rice. However, at longer cooking times normally used for the preparation of brown rice, smaller and inconsistent differences are observed between brown and white rice.

Structures, gelatinization properties and enzyme hydrolyses of starches from transparent and floury grains of rices subjected to field natural extreme high temperature.

Three rice varieties underwent the field natural extreme high temperature (EHT) with daily average temperature over 30 °C from 21 to 89 days after sowing, and had transparent, chalky and floury grains. The structures, gelatinization properties and enzyme hydrolyses of starches from transparent and floury grains were investigated. Compared with control transparent grains, floury grains subjected to EHT markedly decreased the contents of amylose molecules, amylopectin A chains and amylopectin B1 chains and increased the contents of amylopectin B2 and B3+ chains and the average branch-chain length of amylopectin. Both transparent and floury grains had A-type starches, but floury grain starches exhibited higher relative crystallinity, gelatinization temperature, retrogradation and pasting viscosities than transparent grain starches. Floury grain starches had lower hydrolysis rates than transparent grain starches. Native starches were more resistant to digestion but gelatinized and retrograded starches were more prone to digestion in floury grains than in transparent grains.

Amylopectin chain length distributions and amylose content are determinants of viscoelasticity and digestibility differences in mung bean starch and proso millet starch.

This study aimed to reveal the underlying mechanisms of the differences in viscoelasticity and digestibility between mung bean starch (MBS) and proso millet starch (PMS) from the viewpoint of starch fine molecular structure. The contents of amylopectin B2 chains (14.94-15.09 %), amylopectin B3 chains (14.48-15.07 %) and amylose long chains (183.55-198.84) in MBS were significantly higher than PMS (10.45-10.76 %, 12.48-14.07 % and 70.59-88.03, respectively). MBS with higher amylose content (AC, 28.45-31.80 %) not only exhibited a lower weight-average molar mass (91,750.65-128,120.44 kDa) and R1047/1022 (1.1520-1.1904), but also was significantly lower than PMS in relative crystallinity (15.22-23.18 %, p < 0.05). MBS displayed a higher storage modulus (G') and loss modulus (G'') than PMS. Although only MBS-1 showed two distinct and discontinuous phases, MBS exhibited a higher resistant starch (RS) content than PMS (31.63-39.23 %), with MBS-3 having the highest RS content (56.15 %). Correlation analysis suggested that the amylopectin chain length distributions and AC played an important role in affecting the crystal structure, viscoelastic properties and in vitro starch digestibility of MBS and PMS. These results will provide a theoretical and scientific basis for the development of starch science and industrial production of low glycemic index starchy food.

Time-resolved in-situ observation of starch polysaccharide degradation pathways.

Analytical challenges in the direct time-resolved observation of starch metabolism have been addressed by using optimized multidimensional NMR experiments. Starch provides the main source of human dietary energy intake and is a raw material for beverage and renewable fuel production. Use of direct in situ observations of starch remodeling pathways could facilitate our understanding and control of processes of biotechnological, medical, and environmental relevance. Processes involving starch synthesis or degradation are difficult to monitor directly in aqueous solution, however, because starch consists of glucopyranosyl homopolymers that are built up from and degraded into structurally similar fragments that yield only small signal dispersion in optical and NMR spectroscopy. By focusing on acetal groups only, (1) H,(13) C HSQC experiments sampling narrow spectral windows in the highly resolved (13) C dimension have been employed in order to observe the amylopectin cleavage pathway in real time with a temporal resolution of 150 s. Quantifiable signals for more than 15 molecular species emerging during starch fragmentation by human saliva have been resolved and tracked over time in this manner. Altered accumulation of intermediates in the digestion of amylopectin in the presence of black tea acting as an effector have been monitored.

A novel factor FLOURY ENDOSPERM2 is involved in regulation of rice grain size and starch quality.

Rice (Oryza sativa) endosperm accumulates a massive amount of storage starch and storage proteins during seed development. However, little is known about the regulatory system involved in the production of storage substances. The rice flo2 mutation resulted in reduced grain size and starch quality. Map-based cloning identified FLOURY ENDOSPERM2 (FLO2), a member of a novel gene family conserved in plants, as the gene responsible for the rice flo2 mutation. FLO2 harbors a tetratricopeptide repeat motif, considered to mediate a protein-protein interactions. FLO2 was abundantly expressed in developing seeds coincident with production of storage starch and protein, as well as in leaves, while abundant expression of its homologs was observed only in leaves. The flo2 mutation decreased expression of genes involved in production of storage starch and storage proteins in the endosperm. Differences between cultivars in their responsiveness of FLO2 expression during high-temperature stress indicated that FLO2 may be involved in heat tolerance during seed development. Overexpression of FLO2 enlarged the size of grains significantly. These results suggest that FLO2 plays a pivotal regulatory role in rice grain size and starch quality by affecting storage substance accumulation in the endosperm.

Potential interactions between starch and fruit pomace may impact the expansion ratio of direct expanded extrudates.

Due to their dense characteristics, direct-expanded products fortified with insoluble fiber are generally not well accepted. Understanding the interactions between starch and fiber could help to effectively choose and modify ingredients to produce products containing high amounts of fiber. Therefore, this study aims to explain the interplay between two starches (native and waxy corn) and two pomace types (blueberry and cranberry). Blends up to 100% of pomace were extruded using a co-rotating twin-screw extruder. Raw material and milled extrudates were analyzed for their pasting and hydration properties. Fourier-transform infrared (FTIR) and solid-state nuclear magnetic resonance (NMR) spectroscopy were conducted to observe molecular changes. The expansion ratio (ER) significantly decreased as pomace was added and ranged from 3.85 for pure waxy corn starch to approximately 1 for blends that contained 80% pomace. Distinctions between the blends were observed. Particularly, at 20% of pomace inclusion, native corn starch with cranberry pomace showed a significantly higher ER. Different behaviors were also detected during the physicochemical analyses. A nonlinear trend between pomace level and water solubility as well as absorption was observed for native corn starch blends, suggesting that molecular interactions between the biopolymers occur. FTIR and NMR results give no evidence for new covalent bonds; hence, the most likely interactions occurring are hydrogen bonds. In addition to the dilution effect of pomace addition, the enhancement or weakening of such interactions between starch molecules by pomace compounds may reduce the ER.

The dent stage of maize kernels is the most conducive for fumonisin biosynthesis under field conditions.

The fungal pathogen Fusarium verticillioides infects maize ears and produces fumonisins, known for their adverse effects on human and animal health. Basic questions remain unanswered regarding the kernel stage(s) associated with fumonisin biosynthesis and the kernel components involved in fumonisin regulation during F. verticillioides-maize interaction under field conditions. In this 2-year field study, the time course of F. verticillioides growth and fumonisin accumulation in developing maize kernels, along with the variations in kernel pH and amylopectin content, were monitored using relevant and accurate analytical tools. In all experiments, the most significant increase in fumonisin accumulation or in fumonisin productivity (i.e., fumonisin production per unit of fungus) was shown to occur within a very short period of time, between 22/32 and 42 days after inoculation and corresponding to the dent stage. This stage was also characterized by acidification in the kernel pH and a maximum level of amylopectin content. Our data clearly support published results based on in vitro experiments suggesting that the physiological stages of the maize kernel play a major role in regulating fumonisin production. Here we have validated this result for in planta and field conditions, and we demonstrate that under such conditions the dent stage is the most conducive for fumonisin accumulation.

Deciphering molecular interaction and digestibility in retrogradation of amylopectin gel networks.

The impact of the internal part of aewx amylopectin on the gel network and digestibility during retrogradation was investigated using wx amylopectin as a reference. After ß-amylolysis for 60 min (aewx-60), greater shifts in both λmax value and absorbance of iodine binding profiles were observed, accompanied by an increment of short chains (DP 3-5) with reducing the external long chains (DP 17.2). For the amylopectin gels aged 7 days at 4 °C, aewx had greater intermolecular aggregation of double helices to form junction zones, resulting in remarkably higher G', which was significantly greater than that of wx amylopectin or aewx-60. Moreover, aewx amylopectin had a greater RS accompanied by a reduction in RDS after retrogradation. The gel network models of retrograded amylopectins were built to interpret more molecular interactions for aewx than those of wx. The results revealed that aewx amylopectin with a higher proportion of longer external chains prompted the flexibility to align and interact for the formation of double helices and enzyme-resistant structures.

Comparative study on physicochemical properties of starch films prepared from five sweet potato (Ipomoea batatas) cultivars.

Five different sweet potato (Ipomoea batatas) cultivars (Daeyumi, Gogeonmi, Sincheonmi [SCM], Singeonmi, and Sinyulmi [SYM]) were used to extract sweet potato starch (SPS) for developing starch-based films. After the chemical composition and amylose contents of all SPSs were evaluated, the morphological, moisture, mechanical, and barrier properties of the SPS-based films were investigated. As one of the film characteristics, the X-ray diffractograms revealed that the SCM-based film with the highest amylose content (26.34%) had the highest relative crystallinity (24.31%). The SCM-based film also showed higher tensile strength (3.05-fold) and elastic modulus (2.38-fold) than the SYM-based film with the lowest amylose content (21.84%). The water vapor and oxygen permeabilities of the SPS-based films were negatively correlated with the amylose content. Thus, the SCM-based film was less permeable for water vapor (3.16-fold) and oxygen (1.81-fold) than the SYM-based film. These results demonstrated that the sweet potato cultivar, especially the amylose content, plays a significant role in determining the physicochemical properties of the SPS-based films.

Morphological and physicochemical characterization of starches from underground stems of Trimezia juncifolia collected in different phenological stages.

In this study, starches from underground stems of Trimezia juncifolia were evaluated during dry season (DSS), wet season (WSS) and sprouting (SS). Results evidenced that drought stress did not interfere with the yield, amylose content and degree of polymerization (DP) of amylopectin. However, the extraction yield in SS was 58% lower, being observed and increase of 7.5% in the content of amylose, and 13.5% in DP values for SS amylopectin, with a predominance of A-chains. The amount of total sugar, the starch granules size as well as solubility and swelling properties varied as function of the phenological status. Also, starch granules changed from A-type polymorph in DSS and SS to a CA-type in WSS. Nevertheless, it was observed a crystallinity reduction from 56% in DSS to 37.1% in SS. In addition, thermograms evidenced the presence of amylose-lipid complexes, with endothermic transition temperatures being affected by drought stress and sprouting. Finally, results demonstrate that underground stems from T. juncifolia have adaptative strategies involving changes in the morphological and physicochemical properties of the starch granules.