Chenopodium formosanum Koidz.) is an annual fast-growing underutilized pseudo cereal with a high percentage of starch content. In this study, djulis starch was extracted from the flour of dried grains by three different isolation procedures: (1) hydrochloric acid (HCl) isolation procedure (HP); (2) deionized water isolation procedure (WP); and (3) sodium hydroxide (NaOH) isolation procedure (NP), followed by investigation of the physicochemical properties of the isolated djulis starch. The amylose content of HP, WP, and NP was 22.14%, 24.15%, and 22.43%, respectively. For scanning electron microscopy (SEM) morphological observation, djulis starch presented a polygonal shape with granule sizes of 0.56–1.96, 0.74–3.02, and 0.62–2.48 μm, respectively. Djulis starch showed the classification of typical A-type x-ray patterns, and the relative degree of crystallinity for HP, WP, and NP was 33.15%, 36.17%, and 37.42%, respectively. Differential scanning calorimetry (DSC) analysis was used to determine the transition temperatures, transition range, and enthalpies of the gelatinization of starches. HP and WP isolated starch exhibited the highest ΔH 9.24 and 8.51 J/g, respectively, whereas NP starch showed the lowest ΔH of 6.95 J/g. The pasting temperatures of HP, WP, and NP isolated starch, which were analyzed by using a Rapid Visco Analyzer (RVA), were 71.70 °C, 72.80 °C, and 69.53 °C, respectively. The dependence of swelling power for the three isolated starches on temperature was tested at 10 °C with intervals between 60 °C and 90 °C. In short, the NP isolation procedure with a stable reaction is compelling from a technological point of view.Djulis ( Chenopodium formosanum Koidz.) belongs to the class Dicotyledoneae, a genus of Chenopodium that consists of more than 100 species of perennial and annual herbaceous flowering plants, is a native pseudo-cereal grain native grown in Taiwan [Djulis  glycosidic linkages with relatively few α-1→ 6) linked branches, while amylopectin is highly branched molecules of shorter α-(1→ 4) glycosidic linkage glucose molecules and more frequently α-(1→ 4) branches ,6. The m linked bThe physicochemical properties of starch including its morphological features, the characterization of the starch, amylose capacity, crystallization and thermal properties, swelling indexes, and pasting features establish the utilization variety of starch in food and non-food industries . ApproxiStarch isolated from different genotypes and botanical sources with suitable characteristics is gaining more attention in the food industry because it does not require chemical or genetic modification ,12. TherStarch can be isolated from native sources in several ways including alkali treatment, deionized water treatment, and enzyme digestion ,14. For Isolation methods intensely manipulate different starch extension characteristics. Hence, this study examined a suitable process to isolate starch from djulis grains. A hydrochloric acid treatment procedure (HP), deionized water treatment procedure (WP), and alkali treatment procedure (NP) was verified for producing a higher yield and recovery of djulis starch. Moreover, the effects of starch isolation procedures toward the chemical compositions and morphology, together with the crystalline, thermal, and pasting properties of starch granules, have been reported.Chenopodium spp. (Chenopodium quinoa) starch was testified to be much lower, which could be attributed to the different botanical species used, cultural conditions, and methods used for starch isolation [Chenopodium spp. ranged from 9 to 11%. The damaged starch content with the three isolation procedures ranged from 2.12 to 3.93%  in previous research [Chenopodium spp., which showed spherical and polygonal isolated quinoa starch with a submicron size. Djulis starch did not exhibit the cumuli conformation after the different starch isolation procedures. This conformation, in addition to the size and shape of the granules might affect the physicochemical properties of the samples. In short, the SEM images displayed that when the starch was in the djulis seed, it shaped clusters that were engrossed in a matrix formed by starch granules rich in amylopectin. Similarly, when the starch was separated, the starch formed was spherical and polygonal starch; both with almost the same size range were observed.For SEM morphological observation, the starch granules isolated by the three different methods exhibited a polygonal and angular shape A–F. The research . Figure research  toward tChenopodium species starch [Starch, a semi-crystalline material, could be acknowledged by characteristic x-ray diffraction patterns. The x-ray diffraction patterns of djulis starch is shown in s starch , which cs starch .Chenopodium starch. Regarding the crystallinity, previous studies have demonstrated that Chenopodium spp. is considered to be a semi-crystalline material due to its x-ray diffraction pattern, which expressed a broad peak, and the reports defined that this was due to the transposition of amorphous and crystalline layers from amylose and amylopectin that form the granule [The relative degree of crystallinity of djulis starch was 36.17% (WP), 37.42% (NP), and 35.25% (HP), respectively. The results are comparable with the study by Jan et al.  on Cheno granule  clarifie granule .The amylose content has a significant impact on the crystallinity of starch. The relative degree of crystallinity affects numerous practical properties and biochemical specifications of starch . HoweverC. quinoa starch [The swelling power of djulis starch extracted by the three different isolation procedures was measured at 10 intervals ranging in temperature from 60 °C to 90 °C . The resa starch . The inca starch , amylosea starch  or suppra starch . The var0, Tp, and Tc) in conjunction with an enthalpy of gelatinization (ΔH) and the gelatinization temperature range (ΔT) are summarized in 0, Tp, and Tc) of starch extracted by the three different isolation methods indicated non-significant differences, which make clear the scarce or null influence of the isolating procedures. The values found for djulis starch were: T0 = 59.45 ± 0.47 °C, Tp = 64.35 ± 0.62 °C, Tc = 72.19 ± 1.73 °C (HP), T0 = 60.27 ± 0.54 °C, Tp = 65.21 ± 0.22 °C, Tc = 72.95 ± 0.38 °C (WP), and T0 = 60.73 ± 0.89 °C, Tp = 66.02 ± 1.10 °C, Tc = 70.74 ± 1.35 °C (NP), respectively. Gelatinization is defined as the disruption of granules, double helical, and loss of crystalline structure of starch [Chenopodium spp. with the values confirmed as T0 from 44.7 to 57.4 °C, Tp from 50.7 to 61.9 °C, and Tc from 66.2 to 68.5 °C [Chenopodiaceae, nevertheless, the results were incomparable with the existing literature due to the different species.The djulis starch gelatinization was measured and documented the amount of heat involved by using DSC. The gelatinization parameters of starch are connected to a variety of circumstances as well as the size, proportion, and kind of crystalline organization, the shape of the starch granules as well as the architecture, size, and phosphate groups . Thermalf starch . These v 68.5 °C . NonetheStarch granules are initiated to swell and the inter-molecular hydrogen bonds of the starch molecules break down in the existence of adequate water and upraised temperature. At greater temperatures, the granule is obstructed entirely, and gelatinization is completed . The HP The ΔT was high for starch obtained by HP (12.74 ± 0.44 °C) and WP (12.68 ± 0.31 °C) when compared with NP (10.01 ± 0.25 °C). Taking into account that thermal properties are closely interrelated with the destabilizing effect of the amorphous zones (branched regions) that sponge up water before the crystalline zones (amylopectin magnificent zone). Thus, the attendance of alkali (NaOH) used in NP treatments may disturb the crystalline domains, triggering the starch to become more amorphous, consequently promoting the melting of the crystallites in a narrow range of temperature . The gelHP, WP, and NP starch exhibited the normal pasting feature of cereal and roots starch . NeverthThe measurement of the degree of re-association during cooling between the starch molecules concerning the amylose that leached from the swollen starch granules is called the setback viscosity. It is ordinarily practiced as a benchmark of the gelling ability or retrogradation tendency of starch . SetbackThe pasting temperatures of HP, WP, and NP isolated starch were 71.70 °C, 72.80 °C, and 69.53 °C, respectively, which were accurate to the gelatinization temperature acquired by DSC analysis. Thus, djulis starch isolated by NP may exhibit a low pasting temperature, but high consistency, and stability, which is an interesting feature, which is suitable for foodstuff and commodities that involve high viscosity at a lower processing temperature.Djulis starch isolated in this study were expressively different in terms of their physicochemical and functional properties. The three different starch isolation procedures used, HP, WP, and NP, produced high recovery starch with a low amount of damage caused. Generally, djulis starch isolated by the three procedures pretend to have an analogous morphological structure. The djulis starch granules were polygonal and angular in shape; the granule size was between 1.57 and 1.84, presenting a Maltese cross. Djulis starch can be classified as A-type, with a high relative crystallinity value (>30%), and has similar interaction between starch molecules. In conclusion, the NP isolation procedure with a steady and substantial reaction is fascinating from a technological point of view.Djulis starch had a lower swelling power, pasting, and gelatinization temperature, but higher values for amylose content and pasting viscosity when compared with the A-type x-ray diffraction pattern of cereal starch. The analysis of various properties of djulis starch by the different isolation procedures provides beneficial information accompanied by the functional properties for use in various food industries. Djulis starch might be favorable for enhancing the texture of pasta noodles, for example, as a gelling agent applied to high viscous foods, the replacement of chemical modified starches used in baking and other formulation foods as well as a fat substitute and a binder with oral active ingredients. Moreover, due to the small granule size, djulis starch might have broad utilization as an edible biodegradable film. The fascinating and exclusive functional peculiarity of djulis starch could advantageously be applied in the manufacture of specialty food production. Further analysis toward the chemical composition and bioactive components such as the phenolic and flavonoid compositions of djulis hulls, roots, and starch by high performance liquid chromatography (HPLC) is required for the development of djulis functional enriched foods.Fresh grains of djulis were cultivated on November 2018 at a local farm . The grains of djulis were indiscriminately harvested at a mature stage. All samples were transferred into transparent low-density polyethylene (LDPE) zipper bags to prevent any external contamination and stored at a cooler box to transfer back to the laboratory.Starch was detached from the djulis grain flours by the subsequent three procedures of (1) the hydrochloric acid (HCl) isolation procedure (HP); (2) deionized water isolation procedure (WP); and (3) alkali  isolation procedure (NP). Starch was separated from djulis flour as described ,34,35 wiThe moisture (934.01), protein (984.13), lipid (954.02), and ash (942.05) content of the isolated djulis starch was determined according to Association of Official Analytical Chemists (AOAC) methods. The dry matter content was obtained by subtracting the moisture content from the total weight of the sample.The starch content of djulis starch was determined by the American Association of Cereal Chemists (AACC) methods (1995), which is by using the Megazyme total starch content assay kit . The amylose content of djulis starch was examined by the iodine adsorption technique .The yield and recovery of djulis starch obtained by HP, WP, and NP were calculated according to Equations (1) and (2) below:The granule size of starch was measured by using a laser particle sizer . The starch samples were dissolved in deionized water at a concentration of 4%. The device output had the volume spreading of starch granules as the essential measurement. The output data D ‘4, 3’ indicate that the granule diameter resulted from the volume spreading.Scanning electron microscopy (SEM) involved the use of the instrument with a model ABT-150S system . Double-sided adhesive tape was used to attach the isolated starch on an aluminum stub. The sample was coated with gold-palladium . In addition, 15 kV of accelerating potential was used during SEM.For morphological analysis, a starch suspension (1% w/w) was ready with the addition of 50% glycerol. A drop of a starch suspension was placed on a slide and enclosed with a coverslip. An Olympus BX53 polarized light microscope (PLM) equipped with a CCD camera was used to observe the starch’s granule shape and Maltese cross.θ) 5–40° scanning range, 0.02 °/s scan speed, and 0.2 nm receiving slit width. The percentage crystallinity was calculated using Equation (3) as follows:X-ray diffraction of the starch samples was evaluated using a Siemens x-ray diffractometer  with the conducting requirement of 45 kV of target voltage, 30 mA of current, , peak (Tp) and concluding (Tc) gelatinization temperatures, enthalpy (ΔH) of gelatinization, and gelatinization range of dry starch were calculated automatically.Thermal properties of the starch samples isolated by the three different treatments were analyzed by testing with a differential scanning calorimeter . A total of 3 mg of starch samples and 9 mg of deionized water were loaded into 40 μL aluminum differential scanning calorimeter (DSC) crucibles that were hermetically sealed. The sealed crucibles were left overnight at room temperature before DSC analysis to acquire an equal spreading of water before heating the calorimeter. An unoccupied aluminum DSC crucible was used as a reference. The scanning temperature series was increased from 40 to 100 °C at a heating rate of 5 °C/min. The onset  was used to investigate the pasting properties of isolated starch. Starch slurries containing 8% w/w starch  were readied, then subjugated to a heating–cooling cycle. The starch slurries were equilibrated at 50 °C for 1 min, heated up to 95 °C at 6 °C/min, retained at a temperature at 95 °C for 5 min, then cooled to 50 °C at 6 °C/min, and then the temperature controlled at 50 °C for 2 min. The paddle speed was fixed at 960 rpm for the first 10 s of the experiment, followed by 160 rpm for the remainder of the experiment. The viscoamylograph elucidated the various features of the starch as well as the peak temperature, pasting temperature, peak viscosity, viscosity at 95 °C after 5 min holding, and the final viscosity at the end of the 50 °C holding period, which was accessed from a typical diagram of the Rapid Visco Analyzer pasting curves . Each sap < 0.05) on ANOVA, then a Duncan’s new multiple range test was used to correlate the treatment means.The mean (SD) of each analysis are reported. In the statistical analysis of this experiments, all data were evaluated by using a single-factor of ANOVA. If the F-value was significant (


Concerns have been raised that the transplants performed in the local context at the time of procedures reported in this article  may haveDetails as to the donor sources were not reported in . In respIn addition, the authors did not respond to inquiries about the availability of underlying data supporting this study.PLOS ONE Editors retract this article.Owing to the lack of documentation to demonstrate this study had prospective ethical approval, insufficient reporting, unresolved concerns around the source of transplanted organs and whether they included organs from prisoners, and in compliance with international ethical standards for organ/tissue donation and transplantation, the The corresponding author notified the journal that all authors disagree with the retraction. Xiao Xu, XL, QL, QW, ZL, LZ, SZ confirmed their disagreement, the other authors either could not be reached or did not respond directly.


Clostridium thermocellum JN4 and Thermoanaerobacterium thermosaccharolyticum GD17 coculture significantly improves overall cellulosic biofuel production efficiency. Here, we investigated this interaction and revealed the mechanism underlying the improved efficiency observed. In contrast to the previously reported mutualistic relationship, a harmful effect toward C. thermocellum JN4 was observed in these microbial consortia. Although T. thermosaccharolyticum GD17 relieves the carbon catabolite repression of C. thermocellum JN4 regarding obtaining more cellobiose or glucose released from lignocellulose, T. thermosaccharolyticum GD17 significantly hampers the growth of C. thermocellum JN4 in coculture. The increased formation of end products is due to the strong competitive metabolic advantage of T. thermosaccharolyticum GD17 over C. thermocellum JN4 in the conversion of glucose or cellobiose into final products. The possibility of controlling and rebalancing these microbial consortia to modulate cellulose degradation was achieved by adding T. thermosaccharolyticum GD17 stimulants into the system. As cellulolytic bacteria are usually at a metabolic disadvantage, these discoveries may apply to a large proportion of cellulosic biofuel-producing microbial consortia. These findings provide a reference for engineering efficient and modular microbial consortia for modulating cellulosic conversion.Characterizing and engineering microbial communities for lignocellulosic biofuel production has received widespread attention. Previous research has established that  A potentially imminent threat to mankind is our heavy reliability on fossil fuels for our energy, and the foreseeable depletion of these non-renewable resources . A solutClostridium thermocellum, which forms a highly organized extracellular multi-enzyme complex, the cellulosome, can efficiently degrade cellulose  and T. thermosaccharolyticum GD17 (CGMCC 1.5209) were isolated from cellulosic materials by our lab . Ground corncob was prepared by grinding 14 g of shredded corncob in a ball mill  for 1 h . The production of cellulase solutions from d method . ResazurClostridium thermocellum JN4, T. thermosaccharolyticum GD17 and the reconstructed coculture of the two bacteria were grown using CTFUD media in anaerobic tubes/serum bottles in an incubator at 60°C without agitation. The composition of CTFUD media is: sodium citrate tribasic dihydrate 3.0 g/L, ammonium sulfate 1.3 g/L, potassium phosphate monbasic 1.5 g/L, calcium chloride dihydrate 0.13 g/L, L-cysteine-HCl 0.5 g/L, MOPS sodium salt 11.6 g/L, magnesium chloride hexahydrate 2.6 g/L, ferrous sulfate heptahydrate 0.001 g/L, cellobiose 5.0 g/L, yeast extract 4.5 g/L, resazurin 0.5 ml/L  High-Performance Liquid Chromatography (HPLC) system and an Aminex HPX-87H column  from Bio-Rad Laboratories Inc. .Penicillium spp.  buffered in citrate buffer  and 500 μl citrate buffer was prepared. The enzymatic hydrolysis system was incubated for 6 days at 45°C, and the supernatant was extracted for glucose content determination.Analysis of residual cellulose in the cultures was carried out using either acid hydrolysis or enzymatic hydrolysis approaches with cellulose or filter papers, respectively, as the substrate. The dry cellulose pellet was prepared as follows: the pellet was subsequently washed three times with water to remove residual sugars and then dried at 105°C for 4 h in an electric oven. Acid hydrolysis of the dried cellulose residues followed methods described elsewhere . For enzC. thermocellum JN4 and C. thermocellum JN4-T. thermosaccharolyticum GD17 cocultures were performed in 80-ml cultures; for inoculation, 10% of the total volume of cellulose-grown seed cultures was used. Three individual biological replicates were carried out for each experiment. Analyses of glucose, cellobiose, lactate, acetate and ethanol content in cellobiose or glucose-grown C. thermocellum JN4 and T. thermosaccharolyticum GD17 were performed in 50-ml cultures; for inoculation, 10% of the total volume of cellobiose or glucose-grown seed cultures was used. Three individual biological replicates were carried out for each experiment. Determination of biomass accumulation for C. thermocellum JN4 and T. thermosaccharolyticum GD17 on sucrose or dextrin was performed by periodically measuring OD600 in three individual replicates. To compare cellulose degradation rates of cocultures grown on Avicel, Avicel + dextrin and Avicel + sucrose, we assayed residual cellulose using the acid hydrolysis approach. Three individual biological replicates were carried out for each experiment.Analyses of glucose, cellobiose, lactate, acetate, ethanol and residual cellulose in cellulose-grown 600 and the intracellular protein content using three individual replicates, C. thermocellum JN4 and T. thermosaccharolyticum GD17 were grown in media containing 0.5% glucose until an OD600 of 0.8–0.9 was reached. The cultures were then diluted with media to a final OD600 of 0.8000, 0.6000, 0.4000, and 0.2000. The cells were centrifuged at 12,000 rpm for 10 min to pellet the bacteria, after which total proteins were extracted using B-Per® Bacterial Protein Extraction Reagent . Determination of protein concentrations was carried out using a BCA protein assay kit .To determine the relationship between ODC. thermocellum JN4 monocultures and C. thermocellum JN4-T. thermosaccharolyticum GD17 cocultures grown using cellulose, cellulose + sucrose, cellulose + dextrin, or ground corncob with three biological replicates, 10 ml culture was centrifuged for 10 min at 12,000 rpm to pellet both the substrate and bacteria. The pellet was subsequently washed twice with double distilled water, and total proteins were extracted from using B-Per® Bacterial Protein Extraction Reagent . Determination of protein concentrations was carried out using a BCA protein assay kit .To analyze biomass in C. thermocellum JN4 and T. thermosaccharolyticum GD17 in coculture grown on cellulose, cellulose + sucrose, cellulose + dextrin or ground corncob with three biological replicates, total DNA was extracted from cocultures until mid-log phase  using a SoilGen DNA kit . 16S rDNA was then sequenced using a MiSeq or HiSeq2500 PE250 system . The species and abundance of sequenced 16S rDNA were determined.To assess the ratio of C. thermocellum JN4 in mono- and cocultures with T. thermosaccharolyticum GD17 was calculated from the relationship between the OD600 value and intracellular protein content, the assayed total protein content in cultures, and the cell ratio between the two bacteria in coculture.Biomass formation by C. thermocellum JN4 monoculture and C. thermocellum JN4-T. thermosaccharolyticum GD17 coculture, total RNA was extracted from mid-log phase mono- and cocultures grown in media containing 0.5% cellulose, glucose or cellobiose using an E.Z.N.A. Bacterial RNA kit. cDNA synthesis was carried out using a PrimeScriptTM RT reagent kit with gDNA Eraser  from Takara Bio Inc. . qPCR was performed using a Roche LightCycler 96 system  with FastStart Essential DNA Green Master  as the dye. The primers used for real-time PCR are listed in recA gene was used as the housekeeping gene   . Furtherradation . All theC. thermocellum JN4 was upregulated when grown together with T. thermosaccharolyticum GD17, we originally expected that if true mutualism exists between C. thermocellum JN4 and T. thermosaccharolyticum GD17, substrate utilization by C. thermocellum JN4 would also be promoted. This reasoning is because C. thermocellum JN4 supplies T. thermosaccharolyticum GD17 with the growth substrates glucose and cellobiose and consumes these substances that inhibit cellulose degradation in C. thermocellum JN4 thus benefiting C. thermocellum JN4 by promoting substrate consumption. Our analysis of cellulose degradation in mono- and coculture, however, suggested a different scenario. Evaluation of residual cellulose in cellulose-grown C. thermocellum JN4 monoculture and C. thermocellum JN4-T. thermosaccharolyticum GD17 coculture was carried out either by (1) degrading residual cellulose in the culture with acid or (2) degrading residual cellulose in the culture with Penicillium cellulases, followed by analysis of evolved glucose. Surprisingly, no significant difference in residual cellulose content and degradation rate between monoculture and coculture was observed using either method , we assumed that the same number of cells for each strain would lead to the same optical density at 600 nm. We then quantified the relationships between OD600 value and intracellular protein concentration for each bacterium, which suggested a very good linear relationship  of the cells are C. thermocellum JN4, and 42.7 ± 3.7%  of the cells are T. thermosaccharolyticum GD17. In corncob-grown cocultures, 21.1 ± 7.5%  of the cells are C. thermocellum JN4 and 78.9 ± 7.5%  of the cells are T. thermosaccharolyticum GD17. Based on these results, we further determined the biomass present in coculture grown on either cellulose or ground corncob, and the results showed strong growth repression of C. thermocellum JN4 in both cases , acetate (8.85-fold on glucose and 6.97-fold on cellobiose), and ethanol (11.92-fold on glucose and 10.60-fold on cellobiose). For each carbon atom in glucose, C. thermocellum JN4 transfers 0.53 ± 0.01 atoms  to lactate, acetate and ethanol, lower than the 0.77 ± 0.07 atoms  of T. thermosaccharolyticum GD17. For each carbon atom in cellobiose, C. thermocellum JN4 transfers 0.89 ± 0.03 atoms  to lactate, acetate and ethanol, lower than the 1.00 ± 0.03 atoms  of T. thermosaccharolyticum GD17. These results suggest that T. thermosaccharolyticum GD17 has more robust glucose and cellobiose metabolism than C. thermocellum JN4 and that it is more efficient in the conversion of glucose or cellobiose to the end-products lactate, acetate and ethanol. We can therefore conclude that T. thermosaccharolyticum GD17 has a clear and strong competitive metabolic advantage over C. thermocellum JN4 and that the better productive efficiency of T. thermosaccharolyticum GD17 is responsible for the improved alcohol and acid formation in coculture. This result is in contrast to the proposed mutualism between cellulolytic and non-cellulolytic bacteria.The capability of compared . T. therC. thermocellum JN4 suffers from growth inhibition when co-existing with T. thermosaccharolyticum GD17 in the presence of cellulose or corncob, which is in stark contrast to a generally believed mutualism in which the two organisms benefit each other. Repression of C. thermocellum JN4 is the result of competition on nutritional substrates such as glucose and cellobiose and the failure of C. thermocellum JN4 to obtain sufficient nutrients in the presence of T. thermosaccharolyticum GD17, which consumes them at a rate 6–8-fold higher than C. thermocellum JN4. Although the presence of T. thermosaccharolyticum GD17 leads to repression relief of C. thermocellum JN4 regarding cellulase-coding genes, this derepression cannot compensate for the lack of nutrients and subsequently the decrease in biomass. A tentative model of the interactions between C. thermocellum JN4 and T. thermosaccharolyticum GD17 can be generated according to these observations . These results are in agreement with the stimulatory effects of these two substances on the growth of T. thermosaccharolyticum GD17 compared to C. thermocellum JN4. Consequently, the re-balanced cocultures largely stopped degrading cellulose (T. thermosaccharolyticum GD17 inhibits, rather than benefits, C. thermocellum JN4 growth in coculture. They also demonstrate the possibility of modulating cellulose degradation and subsequently cellulosic biofuel formation by controlling the interaction of these two bacteria and of re-balancing the microbial community structure in coculture.To control the interaction between bstances . Supplemellulose . These rC. thermocellum and a non-cellulolytic companion is an improved strategy on cellulosic bioethanol production over the application of C. thermocellum monocultures (T. thermosaccharolyticum GD17 hindered C. thermocellum JN4 biomass accumulation when co-cultured with T. thermosaccharolyticum GD17 (C. thermocellum JN4 or T. thermosaccharolyticum GD17 culture was linearly correlated with the OD600 (C. thermocellum JN4 and T. thermosaccharolyticum GD17 ratio has been used for analyzing microbial community composition and identifying new bacteria (Results obtained in this and previous work clearly suggests that the use of a coculture of cultures . In contcum GD17 , as assehe OD600 , which hhe OD600 . Quantitbacteria .Clostridium species. For example, glucose increased the cellulolytic enzyme production in C. cellulolyticum (C. cellulovorans, glucose suppressed the expression of cellulase genes at the transcriptional level (C. thermocellum JN4 (Clostridium.It is well-known that glucose is an inhibitor on cellulase synthesis in many cellulosic degrading microorganisms, but this effect is different in olyticum . On the al level  as same llum JN4 . TherefoC. thermocellum prefers the transport of cellodextrin instead of cellobiose or glucose for metabolism (C. thermocellum to avoid metabolizing glucose and cellobiose directly, which can be viewed as a defense mechanism against non-cellulolytic bacterium. However, this defense mechanism can only partially recover the impaired growth of C. thermocellum because the rapid consumption of glucose and cellobiose by non-cellulolytic bacteria will thermodynamically promote more degradation of cellodextrin to glucose and cellobiose, therefore still effectively reducing the level of nutrients that C. thermocellum absorbs. As mentioned above, the concentration of cellodextrin fluctuated over time; therefore the type of interaction in this microbial consortium is far from straightforward. For example, when the amount of cellodextrin was elevated, it was indicated that C. thermocellum JN4 could absorb more amount of preferred carbon source, and then we speculated that there is an amensal relationship between C. thermocellum JN4 and T. thermosaccharolyticum GD17. When cellodextrin was almost exhausted, C. thermocellum JN4 lost the advantage over absorbing carbon source, leading to seriously damaged growth. Therefore, it was possible that there is a parasitism relationship between C. thermocellum JN4 and T. thermosaccharolyticum GD17. Anyway, a harmful effect toward C. thermocellum JN4 was observed when co-cultured with non-cellulolytic bacteria.It has been thoroughly established that tabolism . In addiC. thermocellum JN4 and T. thermosaccharolyticum GD17. In a hypothetical mutualistic relationship, T. thermosaccharolyticum GD17 can remove glucose and cellobiose for metabolism while benefiting C. thermocellum JN4 by lifting carbon catabolite repression induced by glucose and cellobiose yet leaving sufficient glucose and cellobiose for C. thermocellum JN4 to use; as the amount of substrate would be greater than when C. thermocellum JN4 is grown on cellulose in monoculture, it would grow better. However, leaving more glucose and cellobiose would defeat the purpose of “lifting cellulase synthesis inhibition” because more inhibition would occur in coculture than in monoculture. Thus, a paradox exists in a mutualistic relationship.Additional analyses could find the contradictory on the previously hypothetical mutualism between C. thermocellum JN4 and T. thermosaccharolyticum GD17 led us to address previously observed but unexplained phenomena regarding the following: C. thermocellum is difficult to separate from accompanying non-cellulolytic bacteria (C. thermocellum and non-cellulolytic bacteria (C. thermocellum commonly occurs in the presence of cellulose in anaerobic microbial consortia (The observed relationship between bacteria ; the probacteria ; C. theronsortia . Two keyBased on these discoveries, to achieve maximal results, microbial consortia for modulating cellulosic production need to be carefully balanced between cellulose degradation and biofuel formation. More specifically, the four microbial processes, cellulose degradation by cellulolytic bacteria, repression of cellulolytic bacteria, depression of cellulase formation, and enhancement of product formation by non-cellulolytic bacteria, need to be carefully balanced for maximizing rates and efficiency in cellulosic biofuel formation.C. thermocellum JN4 and T. thermosaccharolyticum GD17 based the discovery: adding external stimulants for the microbes. We describe a successful attempt to stimulate the growth and alter the proportion of T. thermosaccharolyticum GD17 and subsequently changed cellulose degradation. Pending the discovery of more stimulants and inhibitors for both C. thermocellum and non-cellulolytic bacteria, we believe that structural optimization of the microbial system and subsequent optimization of cellulosic biofuel production may be achieved.Previously published works have reported approaches for balancing such microbial communities, primarily via bioaugmentation of additional microbes in natural or designer microbial communities . In thisC. thermocellum and a non-cellulolytic companion, in contrast to the previously hypothesized mutualistic relationship, as well as the mechanism underlying improved cellulosic bioethanol formation, namely, that the non-cellulolytic bacterium outperforms C. thermocellum in sugar-to-ethanol conversion. We also proposed and verified an approach of stimulant addition for each bacterium in the microbial consortium to re-balance the microbial community structure. These discoveries serve as a basis for designing and optimizing microbial consortia between cellulolytic and non-cellulolytic bacteria to modulate cellulose conversion.These investigations reveal a new relationship between All datasets generated for this study are included in the manuscript and/or the XF, MW, and FW conceived the project and designed the experiments. XF, MW, and FW analyzed the data, and wrote and revised the manuscript. QZ, KN, SL, DH, and YL performed the experiments. All authors discussed the manuscript and agreed to publish.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Escherichia coli, and the enzyme was purified by means of standard chromatographic techniques. High-resolution NMR experiments revealed that HoLaMa is properly folded at pH 8.0 and 20°C. In addition, urea induced a cooperative folding to unfolding transition of HoLaMa, possessing an overall thermodynamic stability and a transition midpoint featuring ΔG and CM equal to (15.7 ± 1.9) kJ/mol and (3.5 ± 0.6) M, respectively. When the catalytic performances of HoLaMa were compared to those featured by the Klenow enzyme, we did observe a 10-fold lower catalytic efficiency by the HoLaMa enzyme. Surprisingly, HoLaMa and Klenow DNA polymerases possess markedly different sensitivities in competitive inhibition assays performed to test the effect of single dNTPs.We report here on the stability and catalytic properties of the HoLaMa DNA polymerase, a Klenow sub-fragment lacking the 3’-5’ exonuclease domain. HoLaMa was overexpressed in  First, a Mg2+-dNTP complex is bound and paired to the template DNA strand by the fingers domain, and the binding of a second divalent magnesium triggers the so-called fingers closure, i.e. a consistent conformational change leading to DNA elongation  + n[L0], with n representing the number of DNA molecules bound per enzyme molecule.Similar fluorescence experiments were performed to determine the dissociation constant, tal data :Q=Qmax*1H) NMR spectra were recorded both in 50 mM sodium phosphate, 50 mM NaCl (pH 8.0) and in 50 mM Tris-HCl, 50 mM NaCl (pH 8.0). The samples were prepared in a final volume of 500 μL containing 5% (v/v) D2O as field lock.High-resolution NMR spectroscopy was performed using a Bruker Advance III 600 MHz spectrometer equipped with a TCI-H/C/N triple resonance cryoprobe. All experiments were performed at 25°C. Water suppression was achieved by the Watergate sequence. Spectra were processed using TopSpin 2.1 software. One-dimensional proton  kJ/mol and the cooperativity of unfolding to m = (4.5 ± 0.5) kJ/mol*M. Accordingly, the midpoint for the chemical unfolding of HoLaMa is equal to CM = (3.5 ± 0.6) M of urea. Remarkably, this value of CM is very close to the value determined for the Klenow enzyme, i.e. CM = (3.6 ± 0.1) M [ΔG, and the cooperativity, m, of unfolding for Klenow, i.e. (20.9 ± 1.3) kJ/mol and (6.3±0.4) kJ/mol*M, respectively [ΔG and m determined for Klenow enzyme are most likely related to the higher molecular mass of this enzyme in comparison with that of its smaller derivative, HoLaMa. The ratio between the molecular mass of the two enzymes  is indeed equal to 0.69, and the ratio between the two m values equals 0.71.To further analyze the overall thermodynamic stability of HoLaMa, we also evaluated its sensitivity to denaturation as induced by increasing concentrations of urea. To this aim, we took advantage of the presence of two tryptophanes in HoLaMa, namely W866 and W924 (coordinates of DNA polymerase I primary structure). The fluorescence of protein tryptophanes is known to be affected by a number of factors , among w± 0.1) M . Howeverectively , were sin, as n = 1.24 ± 0.03, and the dissociation constant, KD, of the enzyme-DNA complex as equal to KD = (35.1 ± 6.1) nM, in reasonable agreement with the 67 ± 19 nM value we previously reported [Next, we tested whether the association with DNA does affect or not the thermodynamic parameters diagnostic of HoLaMa stability. First, we excited at 280 nm and detected the emission at 328 nm of samples containing 500 nM HoLaMa and increasing concentrations of the 40mer dsDNA  describing the folding to unfolding transition of HoLaMa was significantly affected by the binding of dsDNA to the enzyme. Indeed, we determined an overall thermodynamic stability of ΔG = (10.9 ± 3.5) kJ/mol, a cooperativity of unfolding of m = (3.3 ± 0.9) kJ/mol*M, and a transition midpoint of CM = (3.4 ± 1.4) M for the HoLaMa-DNA complex  s-1 (kobs = (64 ± 10) s-1 [In particular, we determined the fluorescence of W924 when an excess of enzyme was rapidly mixed to the 40mer polyA DNA see . The flu± 7) s-1 . This va 10) s-1 . However 10) s-1 , in agre-). In particular, we used for these assays three different 30mer DNAs featuring a template overhang of 15 bases  inactivating the proofreading activity  we determined v as equal to 13.08 ± 0.05, 11.04 ± 0.03, 17.98 ± 0.07 nM/s the activity of HoLaMa, Klenow, and Klenow exo-, respectively  was significantly higher when compared to that detected with 100 μM dTTP (v = 5.12 ± 0.02 nM/s). When the assay was repeated by using 1 μM 30mer 7AC DNA, 100 μM dTTP, and 100 μM dGTP the extension of the DNA substrate was expected to progress beyond the seventh position (v = (2.963 ± 0.003) nM/s, which is slower when compared to that detected in the presence of dTTP only (v = (1.880 ± 0.001) nM/s (v = (0.270 ± 0.005) nM/s which can be ascribed to the background, by unspecific reactions, of this series of assays  nM/s . TherefoWe used the 30mer 3AC DNA  to furth-,  the seventh position of the template overhang  Solutions containing 500 nM HoLaMa  and supplemented (blue line) or not (red line) with 7.6 M urea were excited at 280 nm. The emission spectra were recorded over the 290–400 nm interval and are reported in arbitrary units (a.u.). (B) Difference emission spectrum between native and denatured HoLaMa.(TIF)Click here for additional data file.


Glans resurfacing has been suggested as a treatment option for the surgical management of superficial penile cancer . In this article we describe in detail the glans resurfacing technique with skin graft for penile cancer in a video presentation and we review the current knowledge of the literature. The procedure is described in a stepwise fashion. Initially the patient is circumcised. The glans is marked in quadrants and completely stripped by dissecting and removing the epithelium and subepithelium layer of the glans. Deep spongiosal biopsies are taken to exclude invasion. Each quadrant is sent separately for biopsy. The surface of the graft size needed is estimated. A partial thickness skin graft is harvested from the thigh with a dermatome. The skin graft is then fenestrated. The graft is rolled over the glans and quilted with multiple sutures. A silicone 16F Foley catheter and a suprapubic catheter are placed. The penis is dressed with multiple gauzes and compressed with an elastic band. The patient is discharged the next day. The dressing and Foley catheter are removed in 7 days. The patient continues to use the suprapubic catheter for 7 more days. The patient refrains from any sexual activity for 6 weeks and is closely followed. Glans resurfacing is an emerging new appealing surgical technique that is already a recommendation in the EAU guidelines for the treatment of premalignant and superficial penile lesions. The overall satisfaction rate and recovery of the sexual function are acceptable, and it can be considered an ideal procedure to treat superficial penile cancer. Penile cancer (PeCa), though uncommon, presents a geographical variation in incidence and a strong relation with the human papilloma virus (HPV) , 2. One-The patient is placed in a supine position, with the legs slightly abducted. The procedure is performed under general anesthesia. Broad-spectrum antibiotics are administered preoperatively. The external genital organs and the inner thigh are shaved in the operating room and surgical skin preparation is performed using antiseptic solution. The surgical field is draped leaving the genitals and the thigh exposed. Usually the right thigh is preferred for a right-handed surgeon standing on the right side of the patient.The prepuce skin is marked, just below the corona and on the shaft of the penis . The excThe glans is marked in quadrants from the meatus to the corona . The undThe surface of the graft size needed is estimated by accurately placing a white paper over the glans circumference. Blood is absorbed from the paper defining the size of the graft bed . The papA silicone 16F Foley catheter and a suprapubic catheter are placed to prevent urine extravasation to the graft. The penis is dressed initially with a paraffin gauze and further covered with multiple gauzes and compressed with an elastic band .The patient is discharged the next day with instructions for bed rest for the following two days for better immobilization of the graft. The dressing and Foley catheter are removed in 7 days, and the graft is observed for inflammation or any necrotized tissue. The patient continues to use the suprapubic catheter for urine evacuation in order to keep the graft dry for 7 more days. The patient is advised to avoid any friction to the graft and refrain from sexual intercourse for 6 weeks. The patient is examined on a 3-monthly basis for 2 years, on a 6-monthly basis for a further two years, and then annually.PeCa diagnosis has a detrimental impact on the psychology of the patient, not only because a cancer has been diagnosed, but also due to the fact that the surgical management of the disease can affect adversely the cosmetic penile appearance, the sexual function, and the micturition of the patient . The maiGlans resurfacing is considered a new technique in the field of reconstructive urology and it has been originally used for the treatment of severe lichen sclerosis of the glans . CurrentThe major studies referring to glans resurfacing technique with a median follow-up of at least 30 months reported no cancer-specific deaths, while the rates of the local recurrence fluctuated between 0 and 6% . Glans rAfter performing the circumcision, the first important step is the exposure of the spongiosum by removing the epithelium and subepithelium tissue of the glans, from the meatus to the corona for each quadrant Step 3) [ 3, 6, 77, 9. TheThe next important stage of the operation is the estimation of the graft size, the harvesting and the “transplantation” of the selected graft (Step 4). Although graft-rejection in reconstructive operations is always a possibility, in TGR graft failure is not considered common , 6, 7. TA few key steps during this part of the operation can increase the success rate of inosculation. When the tourniquet is released, small bleeding vessels of the spongiosum should be coagulated meticulously and with accuracy in order to avoid any underlying hematoma, but at the same time taking care of the normal vascularity of the graft bed. Moreover, it is important to harvest a partial thickness graft ranging from 0,02 to 0.04 cm, carefully trimmed to fit and quilted with interrupted sutures in order to promote graft survival. Some authors have proposed immobilization of the skin graft without quilting but instead covering the neoglans with proflavine soaked gauge dressing anchored with tie-over sutures (TODGA-technique) . FurtherThe procedure is completed with the placement of a Foley catheter and a compressive dressing to the penis –7. We alGenerally the procedure is associated with minimal risk of intraoperative and postoperative complications like bleeding, infection, and hematoma , 5, 7. TGlans resurfacing is an emerging new appealing surgical technique that is already a recommendation in the EAU guidelines for the treatment of premalignant and superficial penile lesions. It is important to carefully select, harvest, and suture the graft to the recipient bed of the glans. Negative surgical margins are of great importance for oncological reasons and to avoid possible auxiliary procedure. The overall satisfaction rate and recovery of the sexual function are acceptable, and it could be considered an ideal treatment for the superficial penile cancer.


To improve tendon-to-bone healing in anterior cruciate ligament (ACL) reconstruction, a novel technique via calcium phosphate (CaP)-hybridized tendon graft using an alternate soaking process was developed. The purpose of this study was to evaluate the clinical results of anatomic single-bundle ACL reconstruction using the CaP-hybridized tendon graft with up to 2 years follow-up, and compare the outcome with conventional ACL reconstruction and preoperative data.n = 45) or the conventional method . At 1 and 2 years postoperatively, all patients were evaluated using KT-1000 arthrometry, pivot-shift test, International Knee Documentation Committee (IKDC) grade, Lysholm scale, and Tegner scale; at the same timepoints, bone tunnel enlargement was evaluated using computed tomography, and the tendon graft intensity was evaluated on magnetic resonance imaging. Tendon graft appearance was evaluated arthroscopically once after a period of up to 2 years postoperatively. Cases of re-rupture and adverse events were recorded in both groups.Ninety patients who required anatomic single-bundle ACL reconstruction were randomized to undergo either the CaP-hybridized tendon graft method (CaP group, In both groups, the KT-1000 arthrometry, pivot-shift test, IKDC grade, and Lysholm scale results at 1 and 2 years postoperatively were superior to preoperative data; these results did not significantly differ between groups at either timepoint. The rate of increase of the cross-sectional area of the femoral bone tunnel in the CaP group was smaller than that in the conventional group at 1 year postoperatively. The other results did not significantly differ between the two groups at any timepoint. There were two cases of re-rupture in the CaP group, and six cases of re-rupture in the conventional group. There were no adverse events during follow-up in either group.Anatomic single-bundle ACL reconstruction using a CaP-hybridized tendon graft was safe, and resulted in improved clinical outcomes at 2 years postoperatively compared with preoperative data; the outcomes were comparable with the conventional method. A longer follow-up is needed to clarify the clinical effects of the CaP-hybridized tendon graft in anatomic single-bundle ACL reconstruction.UMIN000019788 Registered 14 November 2015—Retrospectively registered.UMIN,  After anterior cruciate ligament (ACL) reconstruction using a soft tissue graft, only fibrous tissue that was mechanically inferior was noted between the grafted tendon–bone interface –3. To imIn anatomic single-bundle ACL reconstruction using the CaP-hybridized tendon graft, greater in-situ forces in the graft under applied anterior tibial loads were found at 6 months postoperatively in goats compared with an untreated tendon graft . A cliniTherefore, the purpose of the present study was to prospectively evaluate the clinical results of unilateral anatomic single-bundle ACL reconstruction using a CaP-hybridized tendon graft during 2 years of follow-up in comparison with the conventional method and with preoperative data. We hypothesized that anatomic single-bundle ACL reconstruction using the CaP-hybridized tendon graft would result in equivalent postoperative clinical results to the conventional method, and would reduce femoral bone tunnel enlargement compared with the conventional method.n = 45) or the conventional method . The randomization was done according to the days of the week when the patients first visited the outpatient. All patients were followed up for a minimum of 2 years. The ethics committee of Ichihara Hospital reviewed and approved the study . Informed consent was obtained from the enrolled patients. The study protocol was registered with the University hospital Medical Information Network Clinical Trials Registry (UMIN000019788). We excluded revision cases, multi-ligamentous surgery cases, and bilateral ACL reconstruction cases. All ACL reconstructions were performed by two experienced surgeons (T.K. and H.M.). The patient characteristics did not differ between the two groups regarding age, sex, height, weight, operative findings of meniscal injury, and duration from injury to operation  for 30 s. The grafts were subsequently soaked in a NaHPO4 solution  for 30 s. Before each soaking, the grafts were washed in a saline solution. This cycle was repeated ten times [The intraoperative CaP hybridization method was similar to that used in our previous study , 9. AftePostoperative rehabilitation was similar to that performed in our previous study . After sThe patients were assessed preoperatively, and at 1 and 2 years postoperatively. Anterior knee laxity was assessed using the KT-1000 arthrometer  at manual maximum anterior tibial load. The pivot-shift test, the Tegner scale , and theAll patients underwent computed tomography (CT) evaluation at postoperative 1 week, 1 year, and 2 years. The full evaluation was performed independently by a radiologist who was blinded to the study grouping. CT scans  were performed with the knee in full extension to assess the femoral and tibial bone tunnels. Initial volume acquisition was made with 2-mm cuts from 10 cm above the femoral tunnel to 10 cm below the tibial tunnel. Three-dimensional images were reconstructed using a Virtual Place Lexus workstation  , 12, 13.The bone tunnel enlargement evaluation was performed using a similar method to that used in our previous study . The tunThe magnetic resonance imaging (MRI) evaluation was similar to that described previously . MRI evaThe arthroscopic evaluation method was similar to that used in our previous report . Second-t test was used to analyze the results of KT-1000 arthrometry, Lysholm scores, and CT analyses between each group at the same timepoint. The chi-square for independence test was used to analyze the results of the pivot-shift test, Tegner scores, IKDC results, MRI evaluation, and arthroscopic appearance between groups at the same timepoint. The paired t test was used to analyze the KT-1000 arthrometry and Lysholm scores between preoperatively and 1 year postoperatively, and between preoperatively and 2 years postoperatively within the same group. The Wilcoxon signed-ranks test was used to analyze the pivot-shift test results, Tegner scores, and IKDC results between preoperatively and 1 year postoperatively, and between preoperatively and 2 years postoperatively within the same group. Differences were considered significant at P < 0.05.The Student’s A flow-chart of the present study is shown in Fig. Clinical results are summarized in Table Bone tunnel enlargement results are summarized in Table The results regarding the intensity of the tendon graft and the arthroscopic appearance are summarized in Table The main findings of the present study were that anatomic single-bundle ACL reconstruction using the CaP-hybridized tendon graft was safe, resulted in improved clinical results at 1 and 2 years postoperatively compared with preoperatively, and was as stable as the conventional method. The femoral bone tunnel increase rate at 1 year postoperatively in the CaP group was smaller than that in the conventional group. The postoperative intensity of the tendon graft and arthroscopic appearance in the CaP group were equivalent to those in the conventional group. The CaP hybridization technique is simple, and can be applied any patients ACL reconstruction is required. Moreover, the cost is inexpensive because it only involves the solutions preparation.The postoperative clinical results in the CaP group were improved compared with preoperatively, and were similar to the results after conventional anatomic single-bundle ACL reconstruction. Moreover, no adverse events occurred in the follow-up period. Therefore, the CaP-hybridized tendon graft safely and effectively improved the postoperative clinical results in anatomic single-bundle ACL reconstruction. In non-anatomic single-bundle ACL reconstruction, the clinical results at 1 and 2 years postoperatively in the CaP group were reportedly superior to the results after reconstruction via the conventional method . HoweverIn the present study, the CaP-hybridized tendon graft resulted in a reduced femoral bone tunnel enlargement compared with the conventional method at 1 year postoperatively; the lack of data at 2 years postoperatively in both groups may be the reason that no significant difference was detected between groups at 2 years. The articular ends of the bone tunnels are enlarged by bone resorption, which is associated with graft-tunnel motion after ACL reconstruction with suspensory fixation , 19, andThe tendon grafts in the two groups in the present study had a similar intensity on MRI and a similar arthroscopic appearance. The CaP hybridization method did not cause complications in the grafted tendons on MRI and arthroscopic analysis. This indicates that the CaP-hybridized tendon graft is safe for use in clinical practice. The intensity of tendon grafts on MRI and the arthroscopic appearance of the tendon grafts indicate the graft maturation process (ligamentization and revascularization) after ACL reconstruction , 24. We The present study had some limitations. The evaluations were only performed at three timepoints, and the follow-up period was short. Furthermore, there were missing data. Longer follow-up with a complete dataset is needed to investigate any increase in instability, bone tunnel enlargement, and graft maturation. Moreover, it is necessary to evaluate the relationship between bone tunnel enlargement, graft maturation, and knee instability.Anatomic single-bundle ACL reconstruction using the CaP-hybridized tendon graft is safe, and results in improved postoperative clinical results compared with preoperative data during 2 years of follow-up. The postoperative clinical results in the CaP group were as stable as those in the conventional group. The CaP-hybridized tendon graft can reduce postoperative femoral bone tunnel enlargement compared with the untreated tendon graft. A longer follow-up is needed to clarify the clinical effects of the CaP-hybridized tendon graft in anatomic single-bundle ACL reconstruction.


The B-granules of ZM18 and YZ4110 had less amylose content, although the difference among the total amylose contents of the three unfractionated starches was not significant. X-ray diffraction (XRD) patterns showed predominantly A-type crystallinity for all the starches. The A-granules showed sharper XRD patterns than the other starches. DSC analysis showed that the A-granules had broader ranges of gelatinization temperatures than the B-granules from the same wheat variety. The gelatinization enthalpy (ΔH) of A-granules was higher than that of B-granules. AK58 exhibited the smallest enthalpy, while ZM18 showed the largest enthalpy. In pasting tests, the A-granule starch of AK58 had higher peak, final and setback viscosity, lower breakdown and pasting temperature, and the B-granule starch and unfractionated starch of AK58 had lower peak, breakdown, final and setback viscosity and higher pasting temperature than ZM18 and YZ4110.Three starches from the wheat varieties AK58, ZM18 and YZ4110 were separated into large (A) and small (B) granules, which were characterized structurally and evaluated for their functional properties. SEM results showed that the size of A-granules from ZM18 and YZ4110 were about the same, but the sizes of A-granules and B-granules from AK58 were larger than those of ZM18 and YZ4110. FTIR spectra showed that all the samples exhibited a similar pattern, with seven main modes with maximum absorbance peaks near 3,500, 3,000, 1,600, 1,400, 1,000, 800, 500 cm Starch granules . The largranules . Starch granules . These d [et al.  conclude [et al. .Wheat varieties are being developed for different qualities in accordance with the development of genetic recombinations . In our The three kinds of unfractionated wheat starch granules showed bimodal size distributions , breakdown and setback viscosities and higher peak and final viscosities than did the A-granule starch, while the A-granule starch had greater pasting temperature (85.75 °C) than the B-granule starch.For YZ4110, the difference of peak viscosities among A-granule, B-granule and unfractionated starch was no obvious. The B-granule starch had the higher final viscosity and setback and than the A-granule starch and unfractionated starch, while the A-granule starch had the greater breakdown and pasting temperature than the B-granule starch and unfractionated starch.The differences in the pasting properties among three different wheat starches was significant. As shown in Three wheat varieties  were used in this study. They were provided as grains by the Wheat Research Center of Henan Institute of Science and Technology. The wheat grain was milled to straight-grade flour in a Senior mill  according to the approved method 26–31 . Other reagents and chemicals used were at minimum of analytical grade.et al. [The wheat starches were isolated following the method reported by Kasemsuwan et al. . The isoet al. [The A-and B-granules were separated by sedimentation using graduated cylinders (2 L) as described by Takeda et al. . The fraAmylose content of the sample was determined by using the method of Yoo and Jane . The staScanning electron microscopy (SEM) micrographs was recorded with a Quanta 200 environmental scanning electron microscope . The samples were evenly distributed on SEM specimen stubs with double adhesive tape. The micrographs were obtained with an accelerating potential of 15 kV under low vacuum. The micrographs obtained were used to detect any damage to the starch granules .−1 (mid-infrared region) at a resolution of 0.44 cm−1. The sample was diluted with KBr  before acquisition and the background value from pure KBr was acquired before the sample was scanned.In order to further determine the structure of the wheat starches, the FT-IR spectra were obtained using FT-IR . The spectra were recorded in transmission mode from 4,000 to 500 cmMonochromatic Cu Ka radiation (wavelength = 1.54056 Å) was produced by a D/MAX 2500V/PC X-ray diffractometer . The samples were incubated in a chamber at 100% RH for 24 h and then packed tightly in a rectangular aluminum cell. The samples were exposed to the X-ray beam from an X-ray generator running at 36 kV and 20 mA. The scanning regions of the diffraction angle, 2θ, were 0–30°, which covered most of the significant diffraction peaks of the starch crystallites. Other operation conditions included: step interval 0.02, scan rate 4°/min, Sollet and divergence slit 1°, receiving slit 1°, and scattering slit 0.16°. Duplicate measurements were made at ambient temperature. Radiation was detected with a proportional detector.Thermal properties of the starches were analyzed using a differential scanning calorimeter  equipped with a thermal analysis data station. Aluminum pans (Perkin-Elmer) were used for the analysis. Starch samples  were precisely weighed in the sample pans, mixed with distilled water (4 mg), and sealed. The heating rate was at 10 °C per min over the temperature range of 30–120 °C. Indium and zinc were used as the reference standards. Enthalpy change (△H), gelatinization onset temperature (To), peak temperature (Tp), and conclusion temperature (Tc) were measured. The data were averages of a minimum of three replicates of each starch sample.Pasting properties of the starch were determined by using a Rapid Visco-Analyzer (RVA) . Each starch suspension  was equilibrated at 50 °C for 1 min and then heated at a rate of 6 °C/min to 95 °C and then maintained at that temperature for 5 min. The sample was then cooled to 50 °C at a rate of 6 °C/min. A rotating speed of the paddle (160 rpm) was used except the paddle speed was 960 rpm at the first 10 s.The data obtained in this study were expressed as the mean of three replicate determinations and standard deviation (SD). Statistical comparisons were carried out using student t test. P values of <0.05 were considered to be significant.SEM results showed that the size of A-granules from ZM18 and YZ4110 were about the same, but the sizes of A-granules (16–28 µm) and B-granules (2.5–8.5 µm) from AK58 were larger than those of ZM18 and YZ4110. FTIR spectra showed that unfractionated starch and isolated A- and B- granules exhibited a similar pattern. The total amylose contents of the three unfractionated starches were about the same, but the A-granule starch of AK58 contained less amylose than that of ZM18 and YZ4110. XRD patterns showed predominantly A-type crystallinity for all of the starches. The A-granules possessed sharper XRD patterns than the B-granules. The A-granules had the broader ranges of gelatinization temperatures than the B-granules and the gelatinization enthalpy (△H) of A-granule was higher than that of B-granule starches. AK58 exhibited the smallest enthalpy while ZM18 showed the largest enthalpy. The A-granules starch of AK58 had higher peak, final and setback viscosity and lower breakdown and pasting temperature, and the B-granule and unfractionated starch of AK58 had lower peak, breakdown, final and setback viscosity and higher pasting temperature than that of ZM18 and YZ4110. These results showed that the structures and properties of the A- and B-granule starches were distinct, which suggested that biosynthesis of the A- and the B-granules differed.


The effects of black tea polyphenol extract (BTPE) on the retrogradation of starches from different plant sources were studied using differential scanning calorimetry (DSC) and X-ray diffraction (XRD). DSC analysis shows that the gelatinization temperature of maize starch and starches from different rice varieties increased with increasing BTPE level. After storage at 4 °C, BTPE at a concentration of 15% markedly retarded the retrogradation of maize starch and starches from different rice varieties. Native maize starch and starches from different rice varieties showed typical A-type X-ray diffraction patterns, while native potato starch showed a typical B-type X-ray diffraction pattern. Adding BTPE significantly affected the crystalline region and intensities of X-ray diffraction peaks of maize and rice starch granules. It is concluded that adding BTPE markedly inhibits the retrogradation of maize starch and starches from different rice varieties, but has no significant influence on the gelatinization and retrogradation characteristics of potato starch.  The starch granule is semicrystalline. When heated in the presence of water, the semicrystalline structure in granules transforms to an amorphous form, a process termed gelatinization. Gelatinized starch, however, tends to re-associate into an ordered crystalline structure during storage, which is called retrogradation . As the Camellia sinensis. It has attracted significant attention recently, both in the scientific and in consumer communities, for its health benefits for a variety of disorders, ranging from cancer to weight loss. Tea leaves are processed differently to give green, black, or oolong tea. Green tea constitutes about 20% of the tea manufactured in the World, oolong tea is about 2% and black tea is nearly 78% [Tea is a water extract of the leaves of the plant arly 78% . It is warly 78% . Howeveret al. [Many studies have demonstrated that due to their strong free radical scavenging ability, tea polyphenols exhibit antibacterial and antioxidative activities, and show good prospects for their use as preservatives and antioxidants ,12. In ret al.  studied et al. , and theet al. . HoweverThe amylose contents in maize starch (MS), potato starch (PS) and rice starch  were 27.6%, 20.3% and 28.3%, respectively, while the amylose contents in intermediate amylose rice starch (IAR) and low amylose rice starch (LAR) were 13.7% and 1.15%, respectively . There wThe polyphenol constituents in the BTPE sample are shown in To, Tp and Tc) of RS and MS significantly increased with increasing BTPE level, while the enthalpy value of gelatinization (ΔHgel) decreased significantly as the BTPE level increased. The addition of BTPE showed different effects on the R values of starches from different botanical sources. The R values of RS increased with the content of BTPE increasing, in contrast, that of MS decreased. The R values of the starches adding 0%, 5%, 10%, and 15% BTPEs ranged from 9.73 to 11.08 °C for RS, and 8.05 to 6.32 °C for MS, respectively. The gelatinization temperatures, R values, and ΔHgel of PS had no significant change before and after the addtion. This difference in the R value suggests that the degrees of heterogeneity of crystallites within granules of the studied starches are different [The gelatinization thermal properties of starches from different botanical sources with and without the addition of BTPE as investigated by DSC are shown in ifferent . Hret values of RS with 10% and 15% BTPEs still could not be detected after 5 days of storage. Whereas the ΔHret values of MS with only 15% BTPE could not be detected after 5 days of storage, and the ΔHret value of RS and MS with 5% BTPE showed a little decrease in comparison with their native counterpart starches without the addition of BTPE , IAR and LAR was further studied. The gelatinization transition temperatures and gelatinization enthalpy of native and added BTPE starches from different rice varieties are shown in eratures . LAR had [et al.  reported [et al. . To, Tp and Tc) increased significantly with the increasing content of BTPE. However, the enthalpy value of gelatinization (ΔHgel) decreased markedly as the BTPE level increased. The ΔHgel value ranged from 9.68 to 5.75 J/g for HAR, 10.45 to 6.11 J/g for IAR, and 11.86 to 7.04 J/g for LAR, respectively. After adding BTPE to MS and starches from different rice varieties, the increase in the gelatinization temperatures could be mainly contributed by the theaflavin content in BTPE. A study on the key polyphenols in teas indicated that theaflavins were present only in black tea samples . As enzyHret values of samples with 10% and 15% BTPEs still could not be detected after 5 days of storage, whereas the ΔHret values of samples with 5% BTPE were detected after 5 days of storage. After 20 days of storage, all samples showed retrogradation endotherms on the DSC. These results show that ΔHret and DR% began to decrease with the increasing concentration of BTPE, and BTPE at a concentration of 15% obviously retarded the retrogradation of HAR, IAR and LAR.When a starch gel stored at 4 °C is reheated in a DSC, an endothermic transition occurs that is not present in the DSC scan of the freshly gelatinized sample. The results obtained in the second run heating DSC showed that the retrogradation enthalpy and retrogradation ratio of gelatinized starches were significantly decreased by the addition of BTPE . CurrentX-ray diffraction patterns of native and retrograded starches from various botanical sources containing different concentrations of BTPE are shown in i.e., the BTPE levels had no significant effect on the intensity of the diffraction peak in retrograded potato starch. This observation agrees with the previous results obtained using the DSC after 10 days of storage. The investigation using X-ray diffraction further proves the inhibiting effect of BTPE on the retrogradation of maize and rice starches.After retrogradation treatment, rice and maize starches containing different concentrations of BTPE had a mixture of B-type and V-type (main peaks at 2θ = 19.68°) X-ray patterns, and potato starch containing different concentrations of BTPE showed similar diffraction patterns to native potato starch, The diffraction pattern of retrograded starch is clearly distinct from pattern of raw starch. Retrograded starch gives a B-type X-ray diffraction pattern due to the partial loss of granule crystallinity during heat treatment , which r® . Maize starch (MS), potato starch (PS), high amylose rice starch (RS or HAR), intermediate amylose rice starch (IAR), and low amylose rice starch (LAR) were purchased from Pure Biological Technology Co. Ltd. . BTPE was obtained as a fine brown yellow powder from Novanat Bioresources Co. Ltd. , Tea polyphenols, (–)-epigallocatechin gallate (EGCG), (–)-epigallo- catechin (EGC), (–)-epicatechin gallate (ECG), (–)-epicatechin (EC), (−)-gallocatechin gallate (GCG), (−)-catechin gallate (CG), and theaflavins were purchased from Sigma-Aldrichet al. [The amylose contents of rice, maize and potato starches were determined using the colorimetric method described by McGrance et al. . The moiet al. . 18 reversed-phase column . Mobile phases consisted of 0.1% methanoic acid in methanol (eluent A) and methanoic acid in water (eluent B). A gradient system was adopted from Wang and Zhou [The BTPE polyphenols were analyzed by high-pressure liquid chromatography (HPLC) with a DAD detector using a Cand Zhou  with a sTo, peak temperature Tp and conclusion temperature Tc were determined from the first run heating DSC curves, the range of gelatinization temperature (R = Tc − To) was calculated. Enthalpy of gelatinization (ΔHgel) was evaluated based on the area of the main endothermic peak. Then the gelatinized samples were stored at 4 °C for 5, 10, and 20 days and rescanned under the same conditions to determine the temperature and enthalpy changes due to retrogradation. The retrogradation temperature  and enthalpy (ΔHret) were determined from the second run heating. In addition, the degree of retrogradation (DR%) was calculated as the ratio of the retrogradation enthalpy to the gelatinization enthalpy in run heating [Gelatinization and retrogradation thermal properties of the samples were measured using a 61 DSC-Pyris Diamond instrument . Powdered starch and BTPE were mixed at the ratios of starch/BTPE for 2/0, 2/0.1, 2/0.2, and 2/0.3 (w/w) that equated with starch containing 0%, 5%, 10%, and 15% BTPEs (based on starch weight), respectively. The calorimeter was calibrated with an indium standard. Samples of the mixtures of starch and BTPE (about 2 mg) were accurately weighed into aluminum DSC pans, and deionized water was added by micropipette at a water-sample ratio of 2:1. The sample pans were sealed and equilibrated at room temperature for 24 h before analysis. For all DSC runs, a sealed empty aluminum pan was used as reference. The samples were heated. The scanning temperature range and the heating rate were 25–100 °C and 10 °C/min, respectively. The onset temperature  heating . AnalyseThe samples as same as DSC analysis were rice, maize, and potato starches containing 0%, 5%, 10% and 15% BTPEs (based on starch weight). Double deionized water was added to the samples and gelatinized by steam heating for 20 min in a closed thermostatic water bath. These samples were cooled to room temperature then stored at 4 °C for 10 days. The freeze-dried samples were ground and powder passed through a 100 mesh sieve before testing. The recrystallization analysis of samples was carried out using a Rigaku D-Max-2500 X-ray diffractometer  equipped with a copper tube operating at 40 kV and 250 mA. Diffractograms were obtained, the scanning region of the diffraction angle (2θ) was from 4° to 50° at a rate of 4°/min, a step size of 0.02°, 1°/1° divergence slit/scattering slit, 0.30 mm receiving slit. MDI Jade 5.0 was used to analyze the diffractograms. p < 0.05). The data reported in the tables were average of triplicate observations. Data obtained were analyzed by analysis of variance (ANOVA) using SPSS for windows version 13.0. Confidence interval of sample means was reported at the 95% confidence probability. Comparisons of means were made using least significant difference (LSD) and shortest significant ranges (SSR) at 5% significance level , but could not inhibit the retrogradation of PS. Further studies are required to elucidate the molecular mechanism that prevents retrogradation of these starches.


Starch is the main storage substance in rice caryopsis and its properties will determine the quality of rice. Super rice has been extensively studied due to its high-yield characteristics, but the knowledge of amyloplast development and starch quality in caryopsis of super rice especially with large panicle is limited.To address this, large panicle typed and normal panicle typed super rice cultivar Yongyou2640 (YY2640) and Nangeng9108 (NG9108) were investigated in this study. The development of amyloplast in YY2640 caryopsis was better than NG9108, showing faster degradation rate of pericarp amyloplast and better filling degree of endosperm amyloplast. Meanwhile, the starch granule of YY2640 presented as polyhedral shape with smooth surface and the granule size was slightly larger than NG9108. The starch of YY2640 exhibited the lower amylose content, ratio of amylose to amylopectin and the higher level of amylopectin short and long branch-chains compared with NG9108, but there was no significant difference in amylopectin branching degree between them. Two rice starches both showed the characteristics of A-type crystal, and the relative crystallinity and external ordered degree of YY2640 starch were higher than those of NG9108. Furthermore, YY2640 starch showed better pasting properties with lower pasting temperature, shorter pasting time, higher peak viscosity, trough viscosity, breakdown value and lower setback value because of lower apparent amylose content.Overall, the development and filling of amyloplast in YY2640 caryopsis were better than those of NG9108, thus leading to better starch quality of YY2640. Oryza sativa L.) is one of the most important cereal crops in the world, whose production ranks third, second only to maize and wheat. About 50% of global population consume rice as staple food and more than 20 million people take in energy and nutrition from rice and its by-products [Rice , which was close to the length of mature caryopsis. At the same time, the caryopsis grew in the lateral direction and gradually became wider. At 14 DAA, the width of caryopsis tended to be stable and then the volume of caryopsis increased slowly. At 17–20 DAA, the caryopsis was basically shaped and its morphology and size were no longer changed afterwards. The pericarp of rice caryopsis was greenish at early development stage and then faded with the duration of grain filling. At 23 DAA, the pericarp of the caryopsis was mostly waxy white and only the dorsal of caryopsis remained green. At full ripening period, the caryopsis was dehydrated and hardened, showing a transparent state  . As the Based on the above results, it was concluded that the development of amyloplast in YY2640 caryopsis was better than NG9108, which showed faster degradation in pericarp and more accumulation in endosperm.The morphology of starch granules from two rice caryopsis was observed and the granules size distribution was analyzed. Under light microscope, there was no significant difference in the morphology of two rice starch. Most of the starch granules were large and polygonal with irregular shapes, accompanied with some small spherical starch granules Fig. a, b. UndThe amylose content (AC) based on iodine colorimetric analysis is usually defined as apparent amylose content (AAC). The AAC of YY2640 starch was lower than that of NG9108. Due to the absorption of similar wavelengths by amylopectin-iodine and amylose-iodine complexes, AAC is usually an overestimate to the content of amylose. In order to exclude the influence of amylopectin long branch-chains on AAC, the true AC was also determined by assay kit. Although the AC of two rice cultivars exhibited a relatively low level compared with corresponding AAC, its trend of difference (NG9108 higher than YY2640) including the ratio of amylose to amylopectin (Am/Ap) was the same as for AAC Table .Table 3The molecular weight distributions of starch from caryopsis of two rice cultivars were determined by gel permeation column (GPC). A trimodal distribution with low, middle and high molecular weight peaks was designated as AP1, AP2 and AM fractions, respectively. The area ratios of three fractions were analyzed according to chromatograms and listed in Table X-ray diffraction (XRD) can be used to detect the crystal structure and qualitatively analyze the crystal type of starch . From th− 1 varies when starch conformation changes [− 1 is the characteristic structure of crystalline regions in starch, the band at 1022 cm− 1 is sensitive to the amorphous region in starch granules, and the band at 955 cm− 1 is related to the bonding in hydrated carbohydrate helices [− 1 is regarded as a measure to quantify the ordered degree of starch external structure and the ratio of 1022/995 cm− 1 can be used as an index of the proportion of amorphous to ordered carbohydrate structure in starch. As shown in Fig. − 1 and 1022/995 cm− 1 between two rice starch  spectroscopy can be used to study the external ordered structure of starch granules and the intensity of absorbance at 1045, 1022 and 995 cm changes . The ban helices . Thereforch Fig. d. ComparThe pasting properties of rice starch were determined by rapid viscosity analyzer (RVA) and the data is shown in Table Lodging is one of the important factors limiting the high yield of rice and the lodging resistance is mainly determined by plant height. The plant height is closely related to biological yield and the appropriate increase in plant height may lead to the increase of biomass, which is the material basis for the increase in kernel number and 1000-grain weight. Additionally, the thick degree of stalk and the amount of storage substance in stem-sheath can also affect the ability of resisting to lodging and play an important role on improving the seed setting rate and grain weight . PreviouThe base of caryopsis is thick and the upper part is thin in appearance. Then the upper part gradually becomes obtuse and smooth as the growth of caryopsis, which is basically as wide as the base part. Previous studies have found that the shape of caryopsis has a certain correlation with the quality of rice . The sleThe amyloplast in caryopsis is mainly divided into two types, namely pericarp amyloplast and endosperm amyloplast, which play different roles in the development of caryopsis. The pericarp amyloplast is a temporary energy storage that gradually degrades to provide energy for the growth of pericarp, while the endosperm amyloplast is the ultimate energy storage which reserves nutrient for the filial development , 35. TheThe physicochemical properties of starch such as particle size, morphology, crystal structure and external ordered degree are important factors affecting the functional properties of starch, that are closely related to the nutrition and processing quality of starch , thus deThe pasting of starch is the basis of starch utilization. During the pasting process, starch granules absorb water to irreversibly swell and eventually rupture. The crystal structure of starch is destroyed, showing unique physical properties. Therefore, pasting properties, as an important indicator of starch quality, are of great significance for the processing and utilization of starch . The pasIn this study, structural characteristics of amyloplast development in rice caryopsis were observed. The development of caryopsis amyloplast in YY2640 was better than NG9108, showing faster degradation rate of pericarp amyloplast and better filling degree of endosperm amyloplast. Meanwhile, physicochemical properties of rice starch were investigated and their relationship with structural development of amyloplast was discussed. YY2640 starch showed better pasting properties with lower pasting temperature, higher peak viscosity and lower setback value because of lower apparent amylose content. The results indicated that better development and filling of amyloplast in YY2640 might lead to better starch properties. Thus YY2640 may have the potential of good rice quality.The rice cultivars selected in this study were YY2640 and NG9108 which were provided by Ningbo Academy of Agricultural Science and Jiangsu Academy of Agricultural Sciences respectively. YY2640 and NG9108 are both super hybrid rice varieties with the characteristics of high yield and widely cultivated in the middle and lower reaches of the Yangtze River. YY2640 is a typical large panicle typed super rice while NG9108 has a relatively normal type of panicle. Compared with NG9108, YY2640 has a good growth in the field and its plant type is high and strong with large panicle and great number of grains Fig. . The ric2/KI solution at 25 °C for 30 s. All caryopses and their corresponding transections were photographed under a stereomicroscope  equipped with a digital camera. Meanwhile, the caryopses at different DAA were collected to determine fresh weight and dried in a fan-forced oven at 105 °C for 1 h to deactivate enzymes and then at 42 °C to attain constant weights for dry weight determination. Water content was calculated according to the formula below: water content = (fresh weight - dry weight)/fresh weight.Caryopses at different DAA were cut transversely from center and subsequently stained with IMature rice was harvested to determine panicle length, panicle weight, grain number per panicle, seed setting rate and 1000-grain weight.Caryopses at different development stage were collected and cut transversely into 2 mm thin slices from the middle with a razor blade and immediately immersed in 2.5% glutaraldehyde fixative at 4 °C for 4 h. Then, the samples were rinsed thrice with 0.1 M phosphate buffer (pH 7.2) and dehydrated in a graded ethanol series followed by propylene oxide replacement. Afterward, the samples were infiltrated and embedded in Spurr’s resin and polymerized at 70 °C for 12 h. Eventually, the samples were cut into 1 μm thickness slices using an ultramicrotome , pasted on glass slides, stained with 1% toluidine blue for 5 min, rinsed, dried, observed under a light microscope  and photographed by a camera . Each rice cultivar selected three caryopses as repeats, each of which was from different rice panicles.According to the method previously described by Chen et al. , Photoshg for 5 min, and then the supernatant was poured. The remaining precipitate was resuspended with 0.2% NaOH solution and the suspension was centrifuged at 3500 g for 10 min. The supernatant was discarded and the upper colored layer was scraped off. The starch was collected until the supernatant was clarified and dried in a 40 °C oven to constant weight. The dried starch was thoroughly ground into powder using a mortar and collected with a 100-mesh sieve.The mature caryopses were placed in a mortar and fully ground to homogenate with water. The homogenate was filtered through eight layers of gauze and a 100-mesh sieve. The filtrate was centrifuged at 3500 The determination of AAC referred to the method of Man et al. . Starch About 1 mg of dried starch and 500 μL of anhydrous ethanol were mixed and starch ethanol solution (20 μL) was pipetted onto a sample table filmed with aluminum foil and dried at 40 °C for 1 h. After the surface was plated with gold by a vacuum ion sputtering apparatus , the samples were photographed under a field-emission scanning electron microscope . A small amount of dried starch was scattered by 50% glycerol, and then pipetted onto a glass slide covered with a coverslip. The freely dispersible sample was photographed under a light microscope. Image-Pro Plus was used to measure the maximum axial length of starch granules in the image. The granule size distribution was plotted according to the percentage of starch granules in each granule size range. Each starch sample was randomly counted for 2000 starch granules and the measurement repeated thrice.The starch samples were deproteinized with protease and sodium bisulfite and debranched with isoamylase  according to the method previously described . The molAbout 200 mg of dried starch sample was placed in the groove of quartz sample table and flattened with a glass slide. The sample was scanned on an X-ray diffractometer  from the diffraction angle (2θ) ranged at 4° to 40° with a count time of 0.6 s to obtain an XRD spectrum. Photoshop and Image-Pro Plus software were used to analyze the relative crystallinity according to the method of Nara and Komiya  with min− 1 to 800 cm− 1 was recorded followed by deconvolution. The deconvolution enhancement factor was 1.9 and the half-band width was 19 cm− 1. The intensity of bands at 1045, 1022 and 995 cm− 1 was measured and the ratio of 1045/1022 cm− 1 and 1022/995 cm− 1 was calculated using Image-Pro Plus.30 mg of dry starch and 25 μL of ultrapure water were mixed in a centrifuge tube and the suspension was dropped on the probe of attenuated total reflection mode sample table equipped on Fourier transform infrared spectrometer . The spectrum of sample from 1200 cmThe pasting properties of starch were determined using RVA  according to the method previously reported . 3 g staP < 0.05 in SPSS. Origin and Photoshop were used to plot the figures.The experimental data was recorded using Microsoft Excel and significance test was performed using t-test at a probability significance level of


South African Journal of Physiotherapy in March this year. I follow a long line of editors, each of whom has worked hard to improve the standing of the journal. I hope that with the profession’s help I can continue the advancement of our journal.I had the privilege of being appointed as the editor-in-chief of the One of the most noteworthy changes to the journal occurred under the editorship of Prof. Josè Franz and that was the appointment of AOSIS (Pty) Ltd as the publisher. This has meant that the journal is now an online journal and has the support of a professional, experienced and enthusiastic organisation. This makes the future of the journal more secure and enables more progress to be made in improving the standard of the journal.Our profession has grown in leaps and bounds over the years in South Africa (and with much hard work!).We have gone from a diploma qualification at many of the training institutions to a 4-year bachelor’s degree and increasing numbers of physiotherapists now have higher degrees. These are both master’s and doctoral degrees. This means that there is an increasingly strong cohort of researchers and clinician researchers who are developing a local evidence base for our practice. These are the people we need to tap into to encourage them to consider this journal for their work. Quite rightly they want to publish in the best possible journals so it is incumbent on the editorial team to make this journal one of the best available.So where do we go from here? My aim over my term as the editor is to increase the quality and the visibility of the journal and ensure that it becomes a journal of choice for potential authors – obviously not the only journal that they consider for publication. That means that the journal must continue to be locally relevant whilst increasing its continental and international standing. Academics, in particular, need to publish in a variety of journals to ensure that they and their work reach as wide an audience as possible. So to make our journal one that they would consider as a first option means that much work needs to be done to improve both the quality and visibility of the journal.Steps were recently taken by Prof. Franz and the AOSIS (Pty) Ltd team to register the journal on PubMed, and the process is nearing its completion, with what we hope will be a positive outcome. This means that if we are registered, our articles will be listed on PubMed making our authors’ work internationally available. The next step is to get the journal listed on Medline and other international listings. This will require much work. The standard and number of submissions need to improve, and we need to get a wider, more international authorship and readership.To enable the above listings, we have already started on a number of endeavours to comply with the requirements to be listed. Firstly, our editorial board has been increased and now includes local, continental and international experts. The members of the board will help to advertise the journal amongst their colleagues, to increase both the readership and submission number. Secondly, we intend to solicit articles from a variety of experts that will be of value to our readership and at the same time increase the quality of the journal. Whilst doing this I am very conscious of the fact that we need to remain a friendly, supportive journal for young researchers in order that they may publish their articles with us. I hope that I, together with the editorial board, can provide this support. The satisfaction of publishing for the first time is one that we must support, as it is hoped that we will be facilitating these researchers to continue to publish.We also need to provide education and support for our reviewers so that we build up a strong body of competent reviewers. We have recently re-invited the whole list of reviewers to be on the reviewers’ list to ensure that those on the list are there because they want to support the journal. The minimum qualification to be a reviewer is to have a master’s degree and two publications. If readers feel that they would like to become reviewers and they have the above qualifications, they must please contact us so that they can be added to the list. At some point, we will have to consider running a course in reviewing and editing, I believe.We have also started on a programme to increase the readership of the journal. We would really like our profession to read and be aware of what is being done research-wise in our own environment so as to include local studies in their evidence-based practice. This programme includes a Twitter feed as well as a variety of advertisements at appropriate congresses. For example, we had a wonderful poster on the South African Society of Physiotherapy stand at the World Confederation of Physical Therapy in July this year. Other ideas are being considered and will be put in place in due course.Our journal has been in existence for over 40 years. This is an amazing statement of the dedicated professionals who have guided the profession over so many years. So our publishers have embarked on a programme to add all previous issues to the electronic database thus ensuring that this wonderful heritage will be there for future generations of physiotherapists. It also provides readers with the opportunity to see how the profession has grown and changed over the years.So there is much work to be done! I am sure that with the support of the profession, we can build on our solid foundation that has been developed over the years by many dedicated editors and their teams, so that we become a journal that is locally relevant but also has international standing, readership and submissions of articles.