Starch and its products are used in a variety of ways for both the food and nonfood industries. A factorial experiment is carried out with two factors to explain the behavior of the percentage of starch, where the factors correspond to the extraction method and to the raw material. Three methods were used in triplicate: the first followed the official technique of the Association of Official Analytical Chemists (AOAC), to perform acid hydrolysis and quantification of starch by Titulation; the second method involved the colorful reaction with iodine using the UV equipment to measure the absorbance and calculate the percentage of starch; as a third method the FTIR was used, through which the concentration of the starch was calculated by the area under the curve obtained from the spectrum. Manihot esculenta. there is an effect of both the method and the raw material on the percentage of starch, while there was no effect of the interaction; the Tukey test indicates that the highest average percentage of extraction occurs with the extraction method by Titulation and with the starch of It is used as raw material. The method of quantification of starch by UV-VIS spectroscopy was the best for the study samples because it presented less deviation in relation to the FTIR and Titulation methods. The starch is a food reserve polysaccharide predominant in plants, it is the most important and abundant from the commercial point of view, and it is the most common way to include carbohydrates in our diet; the foods rich in it are a good source of energy ; starch The starch comes from different sources with different crystalline structures; cereal grains such as corn, wheat, or rice are sources of starch, such as roots and tubers; for example, tapioca, cassava root, and potatoes are frequently used in the preparation of gluten-free foods; the reversible transformations between the starch and glucose that intervene in the maturation and after the harvest have a remarkable influence on the quality, and the concentration of the starch varies according to the state of maturity , 6. StarStarch and its products are used in a variety of ways in both the food and nonfood industries. In food, it is used as an ingredient in different preparations and in the nonfood industry as a raw material for the elaboration of a wide range of products. The consumption of starch is destined approximately 75 percent to the industrial sector and 25 percent to the food sector –10. Musa paradisiaca, which belongs to the Musaceae family, grows abundantly in many developing countries, and as food is considered one of the most important sources of energy for people who live in the humble regions of many countries [ Manihot esculenta is detailed, it is a perennial woody plant; its stem is cylindrical and formed by knots (point where the leaf joins the stem) and internodes (portion of the stem between two knots). It can be multiplied better in a vegetative way; therefore the stems are important because when they are mature they are cut into stakes of 7 to 30 centimeters in length with which the plant propagates [ Dioscorea trífida; in this the leaves are heart shaped, alternate or opposite, long stalked; their stems are winged or oval cross section with small flowers in clusters or in panicles of three sepals and three stamens; although they are food species, they are characterized by poor flowering [In this work three raw materials are studied for the quantification of their starch. Among these isountries . Next, topagates . Finallylowering .Based on these three agricultural raw materials outlined above, it is necessary to quantify their starch by the three analytical methods most used in these cases: initially the Titulation (acid hydrolysis) typical of traditional analytical chemistry; UV spectroscopic methods; IR studied by instrumental analysis. To compare the methods of quantification, the support of statistical tools is necessary; however, a descriptive study is not sufficient for this purpose, due to the inherent variations in said process; therefore, the support of an inferential tool is necessary to identify the importance of the effect that these factors have on % starch. The experimental design is a very useful tool in the comparison of processes , while t Paradisiaca Musa, Manihot esculenta, and Dioscorea trífida, by means of Titulation, UV-VIS spectroscopy, and FTIR in order to demonstrate which of these raw materials provides the highest percentage of starch and which of the three techniques is most appropriate for each one.Taking into account the fact that it is very important to know the properties of starch and determine how these vary depending on the source from which it is obtained, by means of this work we quantifyThe method of wet starch extraction consisted basically of grinding the pulp and removing in liquid medium those components that are relatively larger, such as fibers and proteins, using sieves; subsequently, the elimination of the water by decantation is facilitated and the sedimented material was washed to eliminate the last different fractions of the starch and finally it was subjected to drying at room temperature. For the extraction of the starch, 2000 grams of each raw material was used, which was acquired from the market of the city of Monteria . The method developed consisted of the following stages .Washing: It was carried out using potable water. Musa paradisiaca, after peeling, it was disinfected with a solution prepared with 1% sodium hypochlorite for 10 minutes.Peeling: Separation of the pulp shell manually with knife. In the case of Musa paradisiaca and Dioscorea trífida; for Manihot esculenta this had a grating process.Maceration: The samples were divided into slices, and water was added to then decrease size to the maximum in a blender for two minutes for the case ofSieving: The product that was obtained is a mixture of starch, water, proteins, minerals, and impurities. To separate them, this slurry was passed through sieves.Decantation: The suspension obtained was deposited in a plastic container and left for 4 hours, then the supernatant was removed to ensure that all impurities had been removed, and the wet starch was washed and sieved three more times.Drying: The wet starch was dried in the environment for a day and then steamed to refine it.The quantification of the starch content of the three raw materials studied was carried out using the Titulation methods (acid hydrolysis), spectroscopic analysis (UV-VIS), and IR with Fourier transform (FTIR), which are described below.Titulation (acid hydrolysis): The technique 920.44 of the   was used Manihot esculenta starch, Musa paradisiaca, and Dioscorea trífida, the procedure was repeated in triplicate [Spectrophotometric analysis (UV-VIS): UV-Vis spectroscopy was one of the first physical methods applied to quantitative analysis and the determination of molecular structures. The technique of UV- Vis spectroscopy is widely used in quantitative analysis, although in qualitative analysis, in the determination of structures, it is surpassed by other techniques, such as infrared spectroscopy and nuclear magnetic resonance. The absorption of UV-Vis radiation by molecules and compounds is due to the electronic transitions of certain groups that make them called chromophobes. These are unsaturated groups, with a large number of electrons responsible for the absorption of UV and Vis radiation. This absorption of energy causes electronic transitions and the usual formation of excited states. Based on the type of electronic transitions that occur, the absorbing species can be classified and, in addition, it is possible to correlate the spectral behavior of a certain species with its chemical characteristics. The electronic transitions that originate from the absorption of UV-Vis radiation can be divided into electronic transitions between orbitals, charge transfer transitions, and ligand field transitions. This procedure of colorful reaction with iodine can only be used when the starch is completely dissolved (gelatinized). For this procedure, 3 mL of the starch solution was placed in a test tube and 1 mL of an I / KI solution was added; then the intensity of the blue color produced in a 600 nm spectrophotometer was measured against a reagent blank.; the amount of starch present in the sample was calculated from a standard curve prepared in the range of 10-50 mg of soluble starch / mL, treated in the same manner as the test sample. For each sample ofiplicate . Musa paradisiaca starch, Manihot esculenta, and Dioscorea trífida samples, using an infrared spectrophotometer with transformed Fourier equipped with a KBr tablet system and total reflectance system attenuated at a temperature of 25 ± 2°C; the solid sample of starch will be introduced in KBr tablets with different concentrations of starch, and finally it will be measured in the equipment in the region of 400 to 4,000 cm-1. We calculate the problem concentration from a calibration curve prepared in the range of 20-100 mg of starch/mg of KBr. For each sample of Manihot esculenta starch, Musa paradisiaca, and Dioscorea trífida, the procedure was repeated in triplicate [IR spectroscopic analysis with Fourier transformed (FTIR): The basis of near infrared spectroscopy consists essentially in the emission of a monochromatic beam of light on the sample, which, depending on its composition and the nature of the bonds present in its molecules, will perform a selective absorption of energy and it will reflect another determined quantity, which is quantified by some detectors present in the NIR instrument and will be used to indirectly quantify the amount of infrared energy absorbed. In this way, the spectra collected in the infrared region are represented graphically as the energy absorbed as a function of the wavelength. In this regard it should be noted that only molecules or part of molecules that vibrate with a frequency similar to that of the incident energy will absorb infrared radiation, so that their vibrational and rotational states are modified, with vibrations of light atoms taking place with strong molecular links. These characteristics correspond to the functional groups C-H, O-H, and N-H of the organic compounds that are part of both plant and animal tissues. For this reason, NIR spectroscopy is practically oriented to the determination and quantification of organic compounds that present the functional groups described above; consequently, from the spectra obtained in the infrared region, information about the chemical composition of the sample analyzed can be obtained. Therefore, the relationship of the energy absorbed with the analytical composition or known characteristics of the calibration samples allows obtaining prediction models for the automatic and instantaneous analysis of thousands of samples. In this sense it can be said that in the last ten years some analytical services based on NIRS technology have been consolidated within the agri-food industry, and many others have been created, both public and private, since NIRS technology or spectroscopy in near infrared is an analytical technique widely implemented in agri-food industries as a support tool for the quality control of raw materials and products, mainly due to its rapid analysis, low cost per sample, simplicity of handling, similar precision to the reference method, no or little preparation of the sample for analysis, etc. For the quantification of starch we proceeded to make a calibration curve and analyzed the spectra of∧2 with three repetitions, to determine the effect of the extraction method, the raw material, and the interaction between raw material and extraction. Finally, the Tukey test and the media graphs are performed to observe the behavior of the factors, a strategy of data analysis similar to that carried out by Baldiris et al. (2017) [Initially, the descriptive statistics of the percentage of starch are made, with respect to the raw material used and the extraction method; subsequently, an analysis of variance is carried out using a factorial experiment 3. (2017) .Musa paradisiaca, Manihot esculenta, and Dioscorea trífida) which were carried out using the Titulation methods (acid hydrolysis), analysis spectroscopic (UV- VIS), and IR with Fourier transform (FTIR).Next, we present the analysis of the results obtained by the procedures used to quantify the starch content of the three raw materials studied ; however, the greatest variations are observed in the method by Titulation and with the raw material of the Dioscorea trífida.The results of Before performing experimental design for starch percentages, the diagnostic graphs of the assumptions of the model are made as illustrated in Figures  Manihot esculenta there is a higher average extraction of starch percentage.The means test for the different raw materials shown in For the methods, there is a highly significant difference between the Titulation method and the UV and IR methods  as noted in  Manihot esculenta presented a higher yield of starch extraction, turning it into a potential source for this product as an alternative in the food industry with respect to the other raw materials studied which presented a lower extraction yield. The assumptions of normality, homogeneity of variance, and independence are fully met. As for the experimental design, it indicates that there is a highly significant effect of the raw material and the extraction method on the percentage of starch, but not on the interaction. Finally, the means tests show that the highest average starch percentages are obtained for the extraction method by Titulation and for the starch based on Manihot esculenta.According to the review of the literature, the results shown, and their discussion, the following conclusions can be obtained:


BioMed Research International has retracted the article titled “Treatment of Acute Pulmonary Embolism: Update on Newer Pharmacologic and Interventional Strategies” [https://doi.org/10.1016/j.jvir.2011.10.012. [William T. Kuo. “Endovascular Therapy for Acute Pulmonary Embolism”, Journal of Vascular and Interventional Radiology, 2012. http://journals.sagepub.com/doi/10.1177/1531003510379880). [Peter H. Lin, Huiting Chen, Carlos F. Bechara, and Panagiotis Kougias. “Endovascular Interventions for Acute Pulmonary Embolism”, Perspectives in Vascular Surgery and Endovascular Therapy, 2010.  https://doi.org/10.1161/CIR.0b013e318214914f. [Michael R. Jaff, M. Sean McMurtry, Stephen L. Archer, Mary Cushman, Neil Goldenberg, Samuel Z. Goldhaber, J. Stephen Jenkins, Jeffrey A. Kline, Andrew D. Michaels, Patricia Thistlethwaite, Suresh Vedantham, R. James White, Brenda K. Zierler, and on behalf of the American Heart Association Council on Cardiopulmonary, Critical Care, Perioperative and Resuscitation, Council on Peripheral Vascular Disease, and Council on Arteriosclerosis, Thrombosis and Vascular Biology, “Management of massive and submassive pulmonary embolism, iliofemoral deep vein thrombosis, and chronic thromboembolic pulmonary hypertension: a scientific statement from the American Heart Association”, Circulation, vol. 123, no. 16.  Registry”, American Journal of Cardiology, Volume 104, Issue 6, p. 198D, article TCT-535. ategies” . The art


The present study was intended to investigate changes in the microstructure and phase transformation of zirconia surfaces using etching and airborne-particle abrasion (AB) and the effects of these processes on the shear bond strength of dental resin cements to zirconia. Four groups were classified according to the surface treatment as follows: etching for 15 min (ET15), etching for 30 min (ET30), AB, and etching for 15 min following AB (ABET). These groups were then classified into two subgroups  according to the resin cement used for bonding, namely, Rely-X U200  or Panavia F 2.0 . Shear bond strength testing was performed using a universal testing device. Scanning electron microscopy (SEM) and X-ray diffraction (XRD) were also performed. When using Rely-X U200, ET15 exhibited the highest mean shear bond strength; the strengths of ET15, ABET, and ET30 were significantly higher than that of AB. When using Panavia F 2.0, ABET demonstrated the highest mean shear bond strength; the strengths of ABET and ET15 were significantly higher than those of ET30 and AB. The etched surface of zirconia could be observed using SEM, and the phase transformations resulting from each surface treatment were detected by XRD. Strong-acid etching of zirconia induced significant surface changes that increased the shear bond strength of resin cement, and the resin adhesive strength was higher when zirconia was etched with strong acid vs. AB alone. The production of dental prostheses using zirconia has been increasing in recent years. Zirconia exhibits mechanical properties comparable with those of metal dental materials, has a color similar to that of teeth, and has several physical and biocompatible advantages .Many studies investigating adhesion with zirconia have been performed, and some adhesion enhancement has been confirmed using several different surface treatments. Examples of such surface treatments for zirconia include airborne-particle abrasion (AB) –4, silicIt is expected that if AB and 10-methacryloyloxydecyl dihydrogen phosphate- (10-MDP-) containing luting agents are used adequately for cementing zirconia, then this will yield successful long-term clinical bonding –16. HoweRecently, studies examining the efficacy of employing chemical etching as a surface treatment for zirconia have been published , 22. EspFour groups were classified according to the surface treatment as follows: etching for 15 min (ET15), etching for 30 min (ET30), AB, and etching for 15 min following AB (ABET). These four groups were then classified into two subgroups each according to the resin cement used for bonding, i.e., Rely-X U200  or Panavia F 2.0 . A total of 8 groups were designated according to the zirconia surface treatment method and the resin cement used for bonding zirconia and the composite resin block. A total of 88 specimens were fabricated , with 11The Zircose E block  used in this study is a zirconium dioxide partially stabilized with 3 mol% yttria . The sin2 intensity for 20 s. After removing the cylindrical resin block from the tube, the resin block was photopolymerized for an additional 20 s and adjusted to a cylinder height of 3 mm  was poured into a cylindrical (6 mm inner diameter), polypropylene tube  and photopolymerized with a light-curing gun  of 1,000 mW/cm of 3 mm .μm Al2O3  particle, zirconia surfaces were abraded at a pressure of 2 bar at a distance of 10 mm for 10 s. After AB, the zirconia blocks were immersed in 96% isopropyl alcohol, sonically cleaned for 3 min, and thoroughly washed again with running distilled water.Using an AB device  and a 110 3) and 48% hydrofluoric acid and adding hydrogen peroxide (H2O2) to achieve a 10 wt.% mixed solution. The zirconia block was immersed in the etching solution and etched for 15 min or 30 min while being sonicated at a frequency of 30 kHz and a power of 100 W/cm2 at room temperature. After the zirconia block was thoroughly washed with running distilled water, annealing was performed in the furnace heated to 1150°C for 1 h to completely remove the etchant and to reduce the residual stress that was incurred during the sintering process.The zirconia block was etched using a strongly acidic solution, which was prepared by mixing 70% nitric acid  was first applied to the zirconia block surface and then air was gently applied using a three-way dental syringe. Following this, Rely-X U200 resin cement was applied according to the manufacturer's instructions. Alternatively, after applying Clearfil ceramic primer (Kuraray) in the same manner, adhesion was performed using Panavia F 2.0 according to the manufacturer's instructions. The cementation jig was made from putty (3M ESPE), and resin blocks were bonded to the center of the zirconia block using the cementation jig . While tA thermocycler  was used to artificially age the cemented zirconia-composite resin specimens for 5000 cycles between 5°C and 55°C. The mooring time at each temperature was 15 s, and the wait time was 2 s.Immediately after the artificial aging process, the shear bond strength was measured using a universal testing device  with a crosshead speed of 0.5 mm/min at the site of approximately 1 mm away from the zirconia surface until the adhered composite resin block fell off.The zirconia surfaces were assessed using a scanning electron microscope  at a magnification of 2,000x before and after the four surface treatments. The adhesion failure mode of the zirconia surface was assessed at a magnification of 40x.X-ray diffraction (XRD) experiments were performed to investigate the phase transformation of zirconia surface particles following the surface treatments. For this purpose, eight zirconia disks  were prepared. One untreated disk and seven disks from each of the surface treatments were observed and analyzed.P ≤ 0.05.For the comparison of the shear bond strengths, one-way analyses of variance with the Dunnett T3 validation method were performed using PASW version 18.0  for Windows . Differences were considered statistically significant at The mean and standard deviation of the shear bond strength between the dental resin cement and zirconia according to the surface treatment method are summarized in P=0.000, F = 17.15). When Panavia F 2.0 was used, the mean shear bond strength of ABET was the highest, and the mean shear bond strengths of ABET and ET15 were significantly higher than those of ET30 and AB .According to the results of our analyses of variance, the shear bond strengths were different according to the zirconia surface treatment for both cements . When ReOur SEM evaluations revealed that the surface roughness of the zirconia was greater when the surface was treated with etching and AB than it was when zirconia did not undergo surface treatment. The appearances of the etched and airborne-particle-abraded surfaces were different from each other. And the irregularity of the etched surface was more uniform than that of the AB surface at 2,000x magnification .After the shear bond strength testing of the dental resin cements to zirconia, the zirconia surface was observed at 40x magnification using SEM . In the θ) value was approximately 28° represents the main peak of the monoclinic phase  can be used not only for bonding with resin cement but also for increasing the bonding strength with porcelain veneers at the same time, and there will be no need to perform airbone-particle abrasion in the clinic before cementation.Within the limitations of the present study, the following conclusions were drawn: (1) strong-acid etching of zirconia caused significant surface changes that increased the shear bond strength of resin cement, and (2) the shear bond strength of resin cements was higher when zirconia was etched with strong acid than when AB was used alone.


This work investigates the stability of emulsions prepared by using octenyl succinic anhydride (OSA)-modified waxy maize starch in the form of granules, dissolved starch, and non-solvent precipitated starch as Pickering emulsion stabilisers. The aim of this study was to investigate the effects of different forms of starches on the stability of emulsion using light microscopy, light scattering, and static multiple light scattering. All starch samples were hydrophobically modified with 3% (w/w) n-octenyl succinyl anhydride (OSA). Starch polymer solutions were prepared by dissolving OSA- modified starch in water in an autoclave at 140°C. Non-solvent precipitates were obtained through ethanol precipitation of dissolved waxy maize. The stability of the oil/water emulsions were different for the three forms of starches used. The granule-based emulsions were unstable, with only a small proportion of the granules adsorbed onto oil droplets, as viewed under a light microscope. The emulsions were observed to cream after 2 hours. The dissolved starch and non-solvent precipitate-based emulsions were stable towards creaming for months, and they had almost 100% emulsifying index (EI = 1) by visual observation and EI ~ 0.9 by multiple light scattering measurements. The results from light microscopy and multiple light scattering measurements indicated the occurrence of coalescence for all three types of emulsions. The coalescence was fastest within days for the granule stabilised system while it was slower both for the dissolved starch and non-solvent precipitate-based emulsions. The latter demonstrated the least degree of coalescence over time. Thus, it was concluded that differences in starch particle size and molecular structure influenced the emulsion droplet size and stability. A decreased particle size correlates to a decrease in droplet size, thus increasing stabilisation against creaming. However, stability towards coalescence was low for the large granules but was best for the non-solvent precipitate starch indicating that there is a window of optimal particle size for stability. Thus, best emulsifying properties were obtained with the non-solvent precipitates (~ 120 nm particle size) where the emulsions remained stable after one year of storage. In conclusion, this study illustrated the potentiality of non-solvent precipitated starch as emulsion stabilizers. Emulsions are widely used in formulations consisting of two immiscible liquids dispersed in one another in the form of small droplets. Since emulsions are thermodynamically unstable, emulsifiers or stabilisers are required to stabilize the droplets to prevent rapid re-coalescing. The emulsion droplet stabilization is often achieved through the addition of surfactants or emulsifiers which decrease the interfacial tension between the phases and increase the steric hindrances and/or electrostatic repulsion between the droplets, thereby increasing the stability of the emulsion , 2. TradMany researchers concluded that larger solid particles (10 nm–5 μm) create a physical barrier between droplets, hence, produce a high desorption energy and adsorb irreversibly to the interface to a higher degree than other emulsifiers such as surfactants , 15. PicGeneral characteristics of starches which are derived from a renewable resource include low toxicity and tasteless. These characteristics enable its formulation application in food, beverages, cosmetics, pharmaceuticals, paints, and coatings , 17, 18.Furthermore, it has been reported that some emulsions which were stabilised using starch granules indicated long-term stability , 25, 26,In order to achieve adequate adsorption at the oil/water interface, both the starch granules and dissolved starch were made to be more hydrophobic. This technique is performed primarily using chemical modification with octenyl succinate anhydride . Using tOn the other hand, many researchers have been using waxy maize as a subject material and widely studied, making this starch type a popular source for comparison. Waxy maize has irregular and sharp edges, with graininess, pin-holes, and deep depressions on the surface of some granules , 30. TheSince small particle size is known to decrease the droplet size with constant parameters where in some cases it increases the stability, nano-sized particles could be of interest to produce Pickering emulsions. Nano-sized starch structures have been produced using non-solvent precipitation –35. A re3 (Sasol Germany). While the water phase was a 5 mM phosphate buffer with 0.2 M NaCl with a density of 1009.6 kg/m3. The n-octenyl succinyl anhydride (OSA) was obtained from Trigon Chemie, Germany.The native waxy maize starch used was supplied by Lyckeby Culinar, Sweden. The oil used was the Miglyol 812, a medium-chain triglyceride (MCT) oil with a density of 945 kg/mThe three different starch emulsifiers utilised in this study include; Starch granules based emulsions (SGE); Dissolved starch-based emulsions (DSE); and non-solvent precipitated starch-based emulsions (NPSE). The preparation of these emulsifiers is described below. All starches were OSA-modified.The amount of OSA used was based on the dry weight of starch measured by an IR-balance . The starch was dispersed in 1.5 to 2 times the amount of distilled water which was added during stirring. The pH was maintained at 7.4 and 7.8 where it was adjusted using citric acid, 1M HCl or NaOH. Then, 3–4% n-octenyl succinyl anhydride  was added in 4 portions with a 15 min delay between each portion. The automatic titration of 1M NaOH while stirring was set to maintain the pH close to 7.6. The titration step was terminated once the pH was stable for at least 15 min. The suspension was then centrifuged at 3000g for 10 min to recover the starch granules. The supernatant was discarded, and the starch was washed twice with water, once with citric acid (pH 4.5–5) and once more with water. The starch was frozen overnight and freeze dried.The determination of OSA level was carried out in a duplicate for modified and control samples. First, 2.5g of starch (dry weight) was wetted with a few drops of ethanol prior to the addition of 25ml 0.1M hydrochloric acid. The suspension was stirred for 30 minutes. Next, the suspension was centrifuged at 3000g for 10 min, where the supernatant discarded at the end. Pellet containing starch was washed with 25ml ethanol once, and with distilled water twice prior to dissolving it in 150ml distilled water and heated in a hot bath at 90°C for 10 min. The solution was rapidly cooled to 25°C using an ice bath, and the suspension was titrated while stirring with 0.1M NaOH until pH 8.3 was reached. The volume of NaOH used was recorded for the calculation of % OSA degree as below:Where V is the volume of NaOH required for titration (ml), M is the molarity of NaOH (0.1M), 210 is the molecular weight (g/mol) of octenyl succinate anhydride group (OSA) and W is the dry weight of sample used.Starch granules which were obtained from the supplier Lyckeby Starch were modified to increase the hydrophobicity using 3% of OSA based on dry weight of starch following the method described above. Following OSA modification, the wet samples were put into a freezer overnight and freeze-dried . The starting temperature was set at -20°C and was set to reach 20°C with an increase in temperature by 1°C/min over 4–5 days. The degree of OSA modification for starch granules was determined to be 3.1%The starting material for the dissolved starch was the OSA-modified starch granules prepared as described above. The OSA-modification was also the same as above. The OSA-modified starch granules were dissolved by adapting the dissolution methods reported by Perez-Rea et al. using an autoclave . This meNon-OSA modified starch was dissolved in the autoclave as described in the previous section. Non-solvent precipitated starch (NPS) was obtained by directly mixing ethanol into dissolved starch (8 mg/ml concentration) at a ratio of 1:1 (starch solution: ethanol). The precipitated starch was collected after centrifugation . Following centrifugation, the solvent was removed and samples were left in the fume hood overnight to completely evaporate the residual ethanol in the precipitated particles. The precipitates were then subjected to OSA modification using the similar method employed in OSA starch granules preparation as described above. The degree of OSA modification for starch granules was determined at 2.03%. The procedure for preparing NPS was adapted from a previous work .The emulsions produced consists of 5% (w/w) of oil/water emulsions containing an MCT oil for the dispersed phase and a phosphate buffer  for a continuous phase, with 200 mg of starch emulsifier per ml of oil. The emulsions were produced by mixing 70 mg of modified granule/non-solvent precipitated (in dry form) or dissolved starch (in solution) into the 0.35 g oil/6.65 g buffer . The emulsions prepared were mixed by vortexing for 10 s. Then, the suspensions were emulsified using a high-shear mixer  at 22000 rpm with a 6mm rotor diameter for 60 s. The stored emulsions that had not phase separated were observed after 24 hours, weekly for up to six weeks and also after a year.The particle size distribution of all emulsions was examined using static light scattering . A small quantity of sample was injected into the flow system connected to the pump circulating at 2000 rpm. The refractive index (RI) was set at 1.54 corresponding to starch , while fEmulsions were further observed with an optical microscope  equipped with a video camera. The samples were examined under Plan 2x, UMPlanFL 5x and 10x, and LMPlanFL 20x and 50x objectives (Olympus). One drop of sample was diluted with five drops of buffer solution. For particular samples, one drop of the diluted sample was placed uncovered on the glass plate for microscopic observation.A sample size of 4 ml starch-based emulsions was prepared to study the emulsion stability during storage. Dispersions of OSA-modified starches  and the MCT oil (oil in buffer) were also measured as a reference to compare with the emulsions. The stability study of the emulsions was evaluated using Turbiscan LabExpert  where it employs static multiple light scattering. The instrument operates by sending a light beam through a cylindrical glass cell containing 4 ml of the sample (12 mm height). The light source is an electroluminescent diode in the near infrared with a wavelength of 880 nm. Part of the incident light is then backscattered by the sample or transmitted through it and received by two sensors at different locations: the sensor that receives the transmitted light (TS) is located 180° from the incident radiation, while the sensor that receives the backscattered light (BS) is located 45° from the incident light. The TS and the BS were monitored as a function of time and cell height at 30°C. The TS and BS data were measured every 40 μm in % relative to standards (suspension of monodisperse spheres and silicone oil) as a function of the sample height (in mm).The BS measurement is directly dependent on the particles’ mean diameter and on their volume fraction. The main instability phenomena observed in colloidal systems are: particle migration  and increment in particle size  . BS profh|scani−scani−1) along the cell height. The sum of the variations (∑i) detected in the samples was calculated using the following equation:The Turbiscan stability index (TSI) was obtained using Turbisoft software (2.0.0.33) using mathematical computing based on the sum of scan-to-scan difference of the intensity of lights (∑The emulsifying index (EI) is the ratio between the volume of the emulsion layer and the volume of the whole sample. In the present study, the EI was calculated by measuring the height of the cell containing the emulsion layer (showing the backscattering peak) vs. the time from the Turbiscan profile. The emulsifying index was calculated as follows:Based on the described methodology, three different emulsion systems were investigated using waxy maize starch; OSA modified starch granules-based emulsions (SGE); OSA modified dissolved starch-based emulsions (DSE), and; OSA modified non-solvent precipitated starch-based emulsions (NPSE). SGE is a Pickering emulsion based on waxy maize granules that have a size of 5–18 μm with an average size around 15 μm  while DSThe macroscopic appearance of emulsions on day 1 is presented in The TSI-value  is a mea4,3 distribution. Hence, the larger particles possess a significant influence on the average droplet size. The discussion that follows focuses primarily on the average size and not the median. The SGE system exhibited a significant increase in In order to understand how the emulsions change with time, the changes in size distribution measured by static light scattering , includi dv, 0.5  were mon24 hours , while nThe change in particle size over time is illustrated in detail in the cumulative graph distributions . As a baOn the other hand, DSE possessed a nearly constant median droplet size (approximately 17 μm) until day 28, after which there was a significant increase to 85 μm. After 28 days, an increase in the number of droplets above 90 μm was observed . The droFor NPSE, The static multiple light scattering was measured using the Turbiscan LabExpert and the changes in backscattering during the first two hours is displayed in To better understand the results for the emulsions, a multiple light scattering baseline data were also obtained for samples containing only the starch emulsifiers  and oil When freshly dissolved OSA-modified starch was investigated , backscaThe backscattering profiles of samples containing only NPS exhibited that there were very small changes in the profile . Since tOn the other hand, in terms of stability, SGE was very unstable as displayed in the backscattering profiles Figs  and 4A. The BS profiles obtained for the DSE Figs  and 4B dAdditionally, the backscattering profiles of the NPSE Figs  and 4C eBased on the results above, a comparison between the three forms of starches can be discussed. The stability of the emulsion systems seemed to increase in the order of SGE << DSE < NPSE. This was true for stability towards both creaming and coalescence. The SGE showed clear phase separation and release of oil after 24 hours, while the emulsions stabilised by DSE were stable in this respect for 6 weeks and the NPSE were stable for at least one year.Since all the particles studied originated from the same starting material, the size and morphology of the emulsifiers play a role in affecting the emulsion stability as it was reported in previous studies . The lowAlthough the DS and NPS were in the same particle size range, the stability of the two emulsifiers varied from each other significantly. Based on previous studies, dissolved OSA waxy starch has been described as a good emulsifier, which was attributed to the adsorbed amylopectin inducing steric repulsion at the interface, as well as increased viscosity in the solution that to some extent hinders creaming , 23, 44.The lack of aggregates observed in the Turbiscan profile for the NPS compared with the profile of the DS indicates that the NPS affects the tendency for any free amylopectin to later retrograde and form new aggregates in solution. This allows NPS to remain stable for a longer period of time than that of DS. In addition, NPSE which has a smaller initial droplet size than DSE indicates NPS consisting of smaller particles can cover a larger surface area creating a higher affinity to the surface. One other explanation could be that the particles do not have to be closely packed at the surface in order to stabilise the emulsion. This is because, based on previous observations on the Pickering type of emulsions, the stability can still be obtained below full surface coverage . HoweverThe different sizes and shapes of the waxy maize-based emulsifiers  are capable of influencing the resulting emulsion droplet size and stability. The decrease in droplet size and the increase in stability towards coalescence and creaming were found to follow the order; SG , DS, and NPS . The observed stability of the emulsions towards coalescence increased for the three OSA modified starches in the order; SGE<< DSE<< NPSE. In general, based on the droplet size, size distribution, microscopic images and Turbiscan stability, NPSE is the most stable followed by dissolved starch and granules. Therefore, in conclusion, this study revealed the high potentiality of non-solvent precipitated starch, NPS as emulsifiers.S1 Fig(TIF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.


Triticum aestivum L.) was isolated and the protein subunit encoded by this gene was characterized as a truncated transit peptide (about 50% shorter than those of other plant AGPS1bs). In the present study, TaAGPS1b was fused with green fluorescent protein (GFP) in rice protoplast cells, and confocal fluorescence microscopy observations revealed that like other AGPS1b containing the normal transit peptide, TaAGPS1b-GFP was localized in chloroplasts. TaAGPS1b was further overexpressed in a Chinese bread wheat cultivar, and the transgenic wheat lines exhibited a significant increase in endosperm AGPase activities, starch contents, and grain weights. These suggested that TaAGPS1b subunit was targeted into plastids by its truncated transit peptide and it could play an important role in starch synthesis in bread wheat grains.ADP-glucose pyrophosphorylase (AGPase), the key enzyme in starch synthesis, consists of two small subunits and two large subunits with cytosolic and plastidial isoforms. In our previous study, a cDNA sequence encoding the plastidial small subunit (TaAGPS1b) of AGPase in grains of bread wheat ( Starch is one of the primary plant carbohydrate reserves in higher plants and it is composed of two different types of glucose (Glc) polymers: amylose  and amylopectin  ,3. DisprArabidopsis thaliana plants expressing a bacterial β-glucuronidase (GUS) gene under the control of the AGPS gene promoter, the expression patterns of GUS gene followed closely the AGPS protein location and starch production in plant tissues/cells, indicating the essential function of AGPase subunits in starch synthesis in higher plants . One unit was defined as amounts of AGPase that catalyzed production of 1 nmol NADPH per min.Activities of AGPase enzyme were assayed for the same transgenic wheat lines for analysis of the above transcript levels of eviously . Embryo 3 transgenic wheat lines were used for analysis on starch content and yield characteristics. Harvested grains were weighted after drying to equivalent moisture content. The starch contents were measured and calculated according to methods of Zhao and his colleagues [Mature grain harvested from wild and Tlleagues . The yielleagues .p < 0.05 level by SPSS (version 20.0) statistical software. Three biological replications were performed, and all data are the mean ± standard deviation (SD) of three independent experimental replicates. Data were statistically analyzed using one-way analysis of variance (ANOVA), and means were compared by using the Duncan's multiple range test at TaAGPS1b (EU586278), a wheat AGPase plastidial small subunit with a truncated transit peptide, was proved to be localized in chloroplasts and its overexpression transgenic wheat plants exhibited the improved AGPase activities, starch contents, and grain weights, indicating its important role in starch synthesis.


Clostridium thermocellum has been the subject of multiple metabolic engineering strategies to improve its ability to ferment cellulose to ethanol, with varying degrees of success. For ethanol production in C. thermocellum, the conversion of pyruvate to acetyl-CoA is catalyzed primarily by the pyruvate ferredoxin oxidoreductase (PFOR) pathway. Thermoanaerobacterium saccharolyticum, which was previously engineered to produce ethanol of high yield (> 80%) and titer (70 g/L), also uses a pyruvate ferredoxin oxidoreductase, pforA, for ethanol production.T. saccharolyticum pforA and ferredoxin into C. thermocellum. The introduction of pforA resulted in significant improvements to ethanol yield and titer in C. thermocellum grown on 50 g/L of cellobiose, but only when four other T. saccharolyticum genes  were also present. T. saccharolyticum ferredoxin did not have any observable impact on ethanol production. The improvement to ethanol production was sustained even when all annotated native C. thermocellum pfor genes were deleted. On high cellulose concentrations, the maximum ethanol titer achieved by this engineered C. thermocellum strain from 100 g/L Avicel was 25 g/L, compared to 22 g/L for the reference strain, LL1319 (adhA(Tsc)-nfnAB(Tsc)-adhEG544D (Tsc)) under similar conditions. In addition, we also observed that deletion of the C. thermocellum pfor4 results in a significant decrease in isobutanol production.Here, we introduced the pforA gene can improve ethanol production in C. thermocellum as part of the T. saccharolyticum pyruvate-to-ethanol pathway. In our previous strain, high-yield  ethanol production could be achieved with at most 20 g/L substrate. In this strain, high-yield ethanol production can be achieved up to 50 g/L substrate. Furthermore, the introduction of pforA increased the maximum titer by 14%.Here, we demonstrate that the The online version of this article (10.1186/s13068-018-1245-2) contains supplementary material, which is available to authorized users. Clostridium thermocellum is a promising candidate organism for the consolidated bioprocessing (CBP) of lignocellulosic biomass into biofuels, such as ethanol ; of; ofC. th deleted . These rC. thermocellum ferredoxins are compatible with T. saccharolyticum ferredoxin-utilizing enzymes [T. saccharolyticum ferredoxin into C. thermocellum in this study. We chose the Tsac_2084 ferredoxin because of its location adjacent to adhA and nfnAB  on the chromosome; given that these latter two enzymes were previously shown to be important for ethanol production, both in T. saccharolyticum [C. thermocellum [Another important component of the pyruvate to ethanol pathway that utilizes a Pfor enzyme is ferredoxin, which is responsible for electron transfer from pyruvate to nicotinamide cofactors, in the context of ethanol production . Previou enzymes . Howeverolyticum , 15 and T. saccharolyticum pforA would improve ethanol production in a strain of C. thermocellum that had been previously engineered with other T. saccharolyticum ethanol production enzymes . We also investigated whether the introduced PforA protein would be able to sustain ethanol production after the five known C. thermocellum Pfor enzyme and enzyme complexes were deleted  into C. thermocellum strains, and evaluating the effects that these genes had on enzyme activity, and on ethanol yield, titer, and maximum ethanol production rate, on soluble carbon sources as well as high loadings of crystalline cellulose.In this study, we hypothesized that introducing the ® HiFi DNA Assembly Master Mix, catalog number E2621). Purification of plasmid DNA or PCR products for cloning was done using commercially available kits from Qiagen, Zymo Research, or New England Biolabs. C. thermocellum strains were transformed using previously described methods [C. thermocellum was purified from Escherichia coli BL21 derivative strains  to ensure that the DNA was properly methylated [Table  methods . Plasmidthylated .Table 2LC. thermocellum strains were grown at 55 °C under anaerobic conditions, either in conical tubes in an anaerobic chamber , with previously described environmental conditions [All reagents used in this study were of molecular grade and obtained from either Sigma Aldrich or Fisher Scientific, unless otherwise noted. nditions , or in snditions .C. thermocellum cells in preparation for transformations, or for harvesting genomic DNA for strain resequencing.Complex medium (CTFUD) was prepared as previously described and used to culture The defined medium (MTC-5) was prepared as previously described  and usedSpecific growth rates, measured on MTC-5 medium with 5 g/L cellobiose as the main carbon source and using a microplate reader, were determined as previously described .600 between 0.6 and 1.0); cultures were then processed as previously described [C. thermocellum recA expression [Gene expression was determined using reverse transcription quantitative PCR (RT-qPCR); Additional file escribed . Gene expression  to allowpression , 22, 23.https://salis.psu.edu/software/reverse) [Translation initiation efficiencies were calculated using an online calculator from the Howard M. Salis group website (URL: reverse) , 25.Cell cultures were grown, harvested, stored, and lysed to obtain cell-free extract as previously described . ProteinAll enzyme assays were performed at 55 °C and at pH 7.0 under anaerobic conditions in an anaerobic chamber .−1 cm−1 was used to calculate activity [4, 0.5 mM DTT, 2 mM MgCl2, 0.4 mM coenzyme A, 0.4 mM thiamine pyrophosphate, and 1 mM benzyl viologen dichloride. Cell extract was added to this assay mixture first to establish a baseline; the reaction was then started by adding 2 mM sodium pyruvate. The final volume for all biochemical assays was 1200 µL.Pyruvate ferredoxin oxidoreductase (PFOR) activity was assayed by the reduction of benzyl viologen instead of methyl viologen, a modification of a previously described protocol . The redactivity . The assBioreactor experiments were carried out as previously described , with anThe fermentation products were measured by high-pressure liquid chromatography (HPLC) as previously described . For tubTo quantify extracellular amino acids, samples were first derivatized with a commercially available derivatization reagent . The derivatized samples were run on an HPLC equipped with the Waters AccQ∙Tag column  with fluorescence detection, using an excitation wavelength of 250 nm and emission wavelength of 395 nm, following the manufacturer’s recommended instrument method. Sample preparation was performed according to the manufacturer’s instructions .Residual cellulose (Avicel PH105) concentration was determined via quantitative saccharification . Pellet Volumetric ethanol production from bioreactor experiments was determined by fitting ethanol production data points for each fermentation with the five-parameter sigmoidal Richards equation , and calRoutine Sanger sequencing of plasmids was performed by Genewiz. Inc., with at least twofold coverage of the cloned regions. Whole genome resequencing of strains was performed by the Department of Energy Joint Genome Institute, using the Illumina Miseq sequencing platform, with an average of 100-fold coverage. Sequencing data were analyzed with the CLC Genomics workbench, using strain LL1319 as the reference genome (accession number SRP101300). Accession numbers for strains and plasmids are listed in Table Thermoanaerobacterium saccharolyticum pforA and ferredoxin were integrated into the C. thermocellum genome in strain LL1319, at a location immediately downstream of Clo1313_2637, and before the putative promoter for the Clo1313_2635 gene . Three promoters were tested to drive expression of the pforA–ferredoxin operon: the previously described C. thermocellum enolase promoter [T. saccharolyticum pforA promoter, and the Athe_2105 promoter from Caldicellulosiruptor bescii [pforA are reported in Additional file r bescii . PromoteT. saccharolyticum pforA and ferredoxin expression was observed in all three operon configurations , we proceeded forward with this strain and designated it as strain LL1391   and pfor4 (Clo1313_1353-1356) encoded for the main Pfor protein complexes in C. thermocellum [pfor1 and pfor4 encoding for important Pfor complexes in C. thermocellum was found when it was observed that BV:PFOR specific activity decreased by ~ 80% relative to wild-type C. thermocellum . Strains containing a deletion of either pfor2 or pfor5 did not show any significant differences in BV:PFOR specific activity, suggesting that they were not important for PFOR activity in C. thermocellum, or that pyruvate was not the primary substrate for these enzymes. The deletion of pfor3, which bears the most similarity to the T. saccharolyticum pforA, also resulted in ~ 40% decrease in specific BV:PFOR activity. Given these observations, pfor1 and pfor4 were therefore the first targets for gene deletion in strain LL1391.Previous work suggested that mocellum . FurtherT. saccharolyticum pathway, including pforA), deletion of pfor1 (strain LL1436) did not result in any significant effects on ethanol production or enzyme specific activity. Deletion of pfor4 (strain LL1437), however, showed a decrease in ethanol yield and a large decrease in titer   and LL1567 (LL1437 with pfor1 deleted). Since neither of these strains showed any significant difference in ethanol production relative to their respective parent strains (LL1438 vs. LL1436 and LL1567 vs. LL1437), the difference in ethanol production between LL1436 and LL1437 is likely due to the Clo1313_1483 mutation, and not the effect of the pfor4 deletion.To compare the effect of the pfor1/pfor4 deletion strain, LL1438, was still able to sustain the improved ethanol production observed in strain LL1391. To eliminate the possibility that this was due to the remaining three C. thermocellum Pfor enzymes compensating for the pfor1 and pfor4 deletions, the genes encoding for pfor3, pfor2, and pfor5 . Despitpfors with the T. saccharolyticum pforA and ferredoxin had improved the maximum ethanol titer and maximum volumetric production rate of ethanol, and to evaluate the performance of the strain on a cellulosic substrate, strains LL1319 and LL1570 were grown on 100 g/L Avicel microcrystalline cellulose.To determine if replacing of the native T. saccharolyticum pforA improved ethanol yield and titer . The differences in volumetric productivity of ethanol between strains LL1319 and LL1570  were not statistically significant  ), which was generated by eliminating the native competing carbon and electron pathways, followed by strain adaptation over ~ 2500 generations to increase growth rate, and which was reported to produce 27 g/L of ethanol from 95 g/L of Avicel [C. thermocellum pfors may be involved in the biosynthesis of isobutanol. Fermentation results from a set of C. thermocellum strains that contain a deletion of one of the five annotated pfors suggest that it is pfor4 that is associated with isobutanol production , whiT. saccharolyticum pforA and ferredoxin on ethanol production in C. thermocellum. There was no effect from expressing T. saccharolyticum ferredoxin in C. thermocellum. It is known that ferredoxins from one organism can often transfer electrons to proteins from another organism [C. thermocellum ferredoxins was sufficient for electron transfer from T. saccharolyticum Pfor protein.In this work, we investigated the effects of organism , so it wT. saccharolyticum pforA did not improve ethanol titer, but it slightly shifted the ethanol to acetate ratio in favor of ethanol production. When the T. saccharolyticum pforA was expressed alongside the previously introduced T. saccharolyticum adhA, nfnAB, and adhEG544D genes, ethanol production improved. These observations support the hypothesis that pforA is an important component of the T. saccharolyticum pyruvate-to-ethanol pathway. Furthermore, the pforA from T. saccharolyticum was able to functionally complement the deletion of the five annotated C. thermocellum pfor genes.Introducing just the C. thermocellum strains with single deletions of each of the five annotated pfors points to pfor4 being responsible for isobutanol production. Given the interest in producing isobutanol either for use as a biofuel or as a feedstock chemical [pfor4 is necessary for isobutanol production could be beneficial to further improve its production from cellulosic substrates.Isobutanol production in strain LL1570 was reduced below our limit of quantification (0.1 mM). Fermentation data from a set of chemical ; the knoT. saccharolyticum promoter to drive the expression of the pforA-ferredoxin operon is relevant to future work involving gene expression in C. thermocellum. Whereas native C. thermocellum promoters have been characterized [T. saccharolyticum pforA, these promoters may still be subject to native transcriptional regulation, and therefore may not be suitable if constitutive gene expression is desired. It should be noted that other examples of heterologous promoters being used in C. thermocellum have been reported [C. thermocellum [The ability to use a cterized , with threported . As hetemocellum , 22, 37,T. saccharolyticum pyruvate to ethanol pathway into C. thermocellum, we observe that there is still a ~ 40 g/L difference between the maximum ethanol titers achieved by engineered C. thermocellum (25–30 g/L) [T. saccharolyticum (60–70 g/L) [+ oxidoreductase (Fnor). It is possible that titer limitations in engineered C. thermocellum are due to an un-optimized Pfor–ferredoxin–Fnor module. Some promising directions for further research include characterizing the Pfor–ferredoxin–Fnor modules of C. thermocellum and T. saccharolyticum in more detail, or engineering the module with better enzymes or through protein engineering of the existing enzymes to overcome possible substrate or cofactor inhibitions [Having now introduced six genes from the –30 g/L)  and thosocellum 2–30 g/L [ibitions , 39.Additional file 1. Supplementary material for this study, including primer sequences (Table S1), promoter sequences (Table S2), fermentation data , chromosomal maps  and enzyme assay data .Additional file 2. List of mutations observed among the strains described in this studyAdditional file 3. Tabular presentation of the data from fermentation of high cellulose concentrations.


Nipponia nippon confined to two acclimation cages mimicking real habitats. We calculated the distance between GPS locations and the centroid of the cages as the positioning error, and used the 95% (95th percentile) positioning errors to define the accuracy. The positioning success averaged 92.0%, which is much higher than that of previous studies. Locations were not evenly distributed by Location Class (LC), with the LC A and B locations accounting for 88.7%. The observed 95% positioning error in the locations of LC A (9–39 m) and B (11–41 m) was quite accurate, while up to 6.9–8.8% of poor-quality locations were detected in LC C and D with >100 m or even >1, 000 m positioning error. Positioning success and accuracy were different between the test sites, probably due to the difference in vegetation structure. Thus, we argue that the tested transmitters could provide a large proportion of high-quality data for fine-scale studies, and a number of poor-quality locations that need attention. We suggest that the HPOD  or PDOP  be reported instead of the LC as a measurement of location accuracy for each location to ensure identification and filtering of implausible locations.GPS tracking has been increasingly used for wildlife studies in recent decades, but its performance has not been fully assessed, especially for newly developed lightweight transmitters. We assessed the performance of eight GPS transmitters developed in China by attaching them to Crested Ibises  GPS tracking has been increasingly used for wildlife studies in recent decades, which has revolutionized our understanding of individual wildlife movement, behaviour and spatial ecology . CompareDespite the high quality of tracking data, a number of factors may influence GPS location accuracy and positioning success (ratio between the observed and expected number of locations), including the effects of the terrestrial atmosphere , satellite constellation (influencing the number of satellites contacted), environment of the transmitters , and behaviour pattern and movement intensity of the tagged animal . TherefoTests of GPS transmitter performance have been carried out over the last two decades, mainly on large mammals due to the weight and size limitations of GPS devices . Due to Nipponia nippon. As part of a re-introduction project, captive Crested Ibises were GPS tagged and confined to two large acclimation cages where they could acquire survival skills in conditions mimicking real habitats before they were released. The tracking data would help understand the movement and habitat selection of the Crested Ibises in the acclimation cage, and thus contribute to evaluate whether these birds could survive in the wild  are evenly distributed in the cage. Therefore, transmitters tested at TC would be subjected to denser vegetation. In these acclimation cages, Crested Ibises can be free ranging, and take advantage of the mimicking real habitats and enough space to behave normally and acquire survival skills before release to the wild.Tests were conducted in two acclimation cages for Crested Ibis situated at Dongzhai National Nature Reserve  of Henan Province, China and Tongchuan  of Shaanxi Province, China . Both ofthe cage . The remDuring 2014–2015, five 25-g GPS-GSM transmitters  × 30 mm (width) × 25 mm (height)) and three 27-g GPS-GSM transmitters  × 27 mm (width) × 35 mm (height)) manufactured by Hunan Global Messenger Technology Corporation, Limited, China  were tested at DZ and TC. All of the transmitters were solar-powered and programmed to position every hour. The tracking data from the transmitters were transmitted through the GSM system once a day. If GSM service was unavailable, data were stored onboard until GSM transmission was possible, with the ability to store up to 12,000 locations.The transmitters were attached to Crested Ibises using a Teflon ribbon back harness. All the transmitters were glued with a cushion to ensure a total height of approximately 40 mm to reduce the masking of the solar panel by feathers. The total weight of the transmitter and harness was 1.6–1.9% of an ibis’s body mass. Birds were released into acclimation cages immediately after processing. All birds were confined to acclimation cages during the test period until they were released into the wild.The manufacturer provided a set of information for each successful location, including date, time, coordinates, speed, locomotion direction, altitude, temperature, voltage, and Location Class. Four Location Classes  from A to D with an advertised accuracy of 5 m, 10 m, 20 m and 100 m, respectively, computed from HDOP  or PDOP  at a probability of 95%, were reported by the manufacturer as a measurement of location accuracy  .We used the central column of the acclimation cage as the reference point, measured by the average of 30 position fixes collected with a handheld Garmin GPSmap 60CSx. We plotted all of the locations from transmitters in ArcView GIS  and used the XTools extension to compute the distance between each GPS location and the reference point as the location error. Since the advertised location accuracy was estimated at a probability of 95%, we calculated the 95th percentiles from the ranked location errors  for each LC to define the positioning accuracy. Because the tagged birds were free-ranging in the acclimation cage, the use of the central column of the cage as the reference point might incur a positive measurement error equals to the distance between the birds and the central column of the cage with a maximum of the radius of cage. This measurement error will result in an overestimate of the location error. To address this bias, we reported the positioning accuracy in a range, with the lower limiting value as the 95% location error minus the radius of the cage.We used a Chi-square test to compare differences in positioning success among transmitters and differences in the proportion of locations in each LC among transmitters. We checked the data for normality, and used a Mann–Whitney test to detect differences in positioning success between DZ and TC. We used a Kruskal Wallis test to compare differences in the positioning error among locations of the four LCs. All statistical analyses were performed using the SPSS statistical software package .All data collected as part of this study were approved by the National Bird Banding Center of China (No. NBBC20140210). This test was conducted as part of a re-introduction project for the Crested Ibis, under the supervision of a team of experienced ornithologists. The conditions in the cages and the acclimation procedure were in line with the IUCN Re-introduction Guidelines.χ2 = 19.073, df = 7, P = 0.008) and was higher at DZ than at TC . The proportion of locations in each LC differed among the eight transmitters . On average, the LC B locations accounted for up to 67.5%, while the LC D locations constituted the smallest proportion of 1.4%  and TC  .The distribution of locations relative to the test cages and bound 95% positioning error at DZ and TC is shown in < 0.001) . For the< 0.001) . TherefoA series of lightweight GPS transmitters have been newly developed in China in recent years to meet the increasing need for wildlife tracking. To the best of our knowledge, our study presented the first performance assessment of GPS transmitters from HQXS, one of the major brands in China, which will provide an essential prerequisite for the application of these transmitters and contribute to the proper use of GPS wildlife tracking data. Since animal behaviours such as locomotion, hiding and diving may influence GPS location accuracy and positioning success , it is mWe detected an average positioning success of 92.0%, which was much higher than that of GPS transmitters deployed on animals (69.3%) and similar to that in stationary tests (94.8%) based on a review of 35 journal articles . This inLocation accuracy is calculated by HDOP or PDOP and classified as four LCs for our tested transmitters . This clOur results revealed that there was lower positioning success and relatively more poor-quality locations at TC than at DZ, probably due to the relatively smaller cage and dense surrounding trees at TC. This was similar with many previous studies . Recio eIn conclusion, our study provided the first performance assessment of lightweight transmitters newly developed by HQXS in China, which could provide a large proportion of high-quality data. However, a number of factors, including vegetation structure, might have impacts on the positioning success and accuracy of these GPS transmitters, and result in a number of poor-quality locations. Thus, the tracking data need to be exploited properly and corrected carefully for fine-scale studies. We suggest the HPOD or the PDOP be reported instead of the LC as a measurement of location accuracy for each location to ensure the identification and filtering of implausible locations.10.7717/peerj.5320/supp-1Data S1Click here for additional data file.10.7717/peerj.5320/supp-2Data S2Click here for additional data file.


O-glycosides (TGFs) (quercetin (I), kaempferol (II), isorhamnetin (III)) from Ginkgo Biloba extracts (EGB) are evaluated. The adsorption and desorption properties of TGFs are studied on macroporous resins, including D101, D201, AB-8, HPD400, D301, and D311. Along with the results, AB-8 resin exhibits the best adsorption and desorption capacity for these three ginkgo flavonoid O-glycosides among the six resins. Adsorption isotherms are created on AB-8 resin and fit well to the Langmuir (R2 > 0.96) and Freundlich  models. After the treatment with gradient elution on AB-8 resin packed chromatography column, the contents of the three main ginkgo flavonoid O-glycosides  increase from 8.93%, 9.88%, and 6.11% in the extracts to 30.12%, 35.21%, and 14.14%, respectively, in the product. The recoveries of compounds I, II, and III are 88.76%, 93.78%, and 60.90%, respectively. Additionally, the anti-inflammatory effects of TGFs are evaluated in LPS-treated RAW 264.7 macrophages, and the result demonstrates that TGFs could significantly inhibit LPS-induced NO release in vitro in a dose-dependent manner compared with the control group. These findings suggest that TGFs could potentially be natural antioxidants and anti-inflammatory ingredients that could be used in pharmaceutical products and functional food additives.In the present study, the performance and separation characteristics of six macroporous resins for the enrichment and purification of total ginkgo flavonoid  Ginkgo biloba is called a living fossil and has been surviving on Earth for 180 million years, and EGB has been used for medicinal purposes in China for several thousand years. EGB is available in either a non-standardized form or as a standardized extract containing 24% flavone glycosides, as shown in After extraction, the purity of EGB still did not meet the purity demands, and further purification was carried out. Among all reported technologies , macropoEGB has been shown to have diverse bioactivities and different pharmacologic actions, such as antioxidation, anti-inflammatory, antitumor and immunomodulatory responses . Thus faThe object of the present work was to evaluate the adsorption and desorption characteristics of TGFs on different macroporous resins. The physical models of the Langmuir equation and the Freundlich equation, as well as the HPLC method, were used to analyze the kinetic characteristics and separation effects. Based on the information, an efficient method for the preparative separation of TGFs from EGB with an optimal resin. The anti-inflammatory potential effects of TGFs were evaluated in RAW264.7 macrophages.O-glycosides contain non-polar phenyl groups and polar multi-hydroxyl groups. Either non-polar resins or polar macroporous resins are applicable to the adsorption of TGFs. Thus, six macroporous resins ranging from non-polarity to polarity were used for adsorption of TGFs due to the physical and chemical properties, as shown in According to similarity–intermiscibility theory, total ginkgo flavonoid Six macroporous resins with distinct physical properties of TGFs from EGB were evaluated and are shown in The kinetics of adsorption that describe the solute uptake rate governing the contact time of the adsorption process is one of the key properties that describes the efficiency of adsorption. Hence, in the present study, the adsorption kinetics of TGFs were determined to understand the adsorption behavior of AB-8 resins. Generally, the adsorption capacity of TGFs increased transiently with adsorption time before reaching adsorption equilibrium. As shown in m value (obtained from the Langmuir isotherm) and 1/n value (obtained from the Freundlich isotherm). The correlations (0.9965–0.9969) for TGFs on AB-8 indicated that the two models were suitable for describing the tested adsorption system in the concentration ranges studied.To investigate the adsorption capacity and characterize the adsorption behavior of TGFs, sample solutions with various concentrations of EGB (15.87–37.18 mg/mL) were shaken with AB-8 resin at 25, 30 and 35 °C. Additionally, to interpret the adsorption experimental data, the Langmuir and Freundlich models were used. Generally, in the Freundlich equation, adsorption was easy to determine when the 1/n value was between 0.1 and 0.7, and it was hard to determine when the 1/n value was beyond 1 . As can O-glycosides hydrolyzed into H+ and their corresponding anions, thus resulting in a lower adsorption capacity. Consequently, the pH value of the sample solution was adjusted to 5.0 for subsequent tests [A key parameter affecting the adsorption and desorption capacity of the resins is the pH value of the initial sample solutions. The pH value influences the extent of ionization of solute molecules and therefore affects the affinity between the solutes and solutions . As shownt tests .Dynamic desorption was carried out with a gradient elution mode at a flow rate of 2.0 BV/h. Different elution solvents with the same volume (2 BV) were used to desorb TGFs when the sample loading amount was 2 BV. At 20% ethanol, the TGFs were barely desorbed. When the ethanol concentration was over 20%, the desorption ability increased sharply and achieved a maximum value at 40% ethanol. Thus, a gradient elution procedure with 40%, 60% and 80% ethanol, at a flow rate of 2 BV/h, was used for desorption of TGFs. In addition, the elution volume of each ethanol concentration was modified under the guidance of absorbance. The dried extract of EGB (4.0 g) was dissolved in 20% ethanol. The TGFs were enriched and purified from the sample solution with AB-8 resin. According to the content analysis of TGFs, the elution program was modified as 2 BV of 20% ethanol, 2 BV of 40% ethanol, 2 BV of 60% ethanol and 2 BV of 80% ethanol. As shown in Thus, the ultimate separation and purification methodology for TGFs was summarized as follows. Adsorption: Concentrations of TGFs in sample solution 9.64 mg/mL; pH 5.0; flow rate 2 BV/h; temperature 35 °C. Desorption: 20% ethanol and 40% ethanol, each 2 BV; then 2 BV of 60% ethanol; flow rate 2 BV/h.50  of TGFs, EGB, and ascorbic acid were 38.00 ± 1.06, 47.82 ± 1.05, and 5.73 ± 1.62 μg/mL, respectively. In the ABTS scavenging assay, the IC50 values of TGFs, EGB, and ascorbic acid were 24.34 ± 1.12, 42.08 ± 1.01, and 8.06 ± 1.05 μg/mL, respectively.DPPH and ABTS are relatively stable free radical compounds that have been broadly used to test the free radical-scavenging activities of various natural extracts. In the DPPH scavenging assay, the ICp > 0 05) affected by TGFs in the experimental concentration range.To determine the cytotoxicity of TGFs on Raws 264.7 cells, an MTT assay was performed after treatment with LPS and at various concentrations. The results showed that there was no difference among the 3.125, 6.25, 12.5, 25, 50 and 100 µg/mL TGFs and EGB-treated groups , which iThe anti-inflammatory effects of TGFs and EGB were further evaluated using LPS-treated Raws 264.7 cells. The NO reduction experiment indicated that TGFs and EGB at 50 µg/mL dramatically decreased the NO production of LPS-stimulated Raws 264.7 cells compared to the control group, as shown in EGB was provided by Jiangsu Kanion Pharmaceutical Co., Ltd. ; Macroporous resins code AB-8 D101 HPD400 D201 and D301 were purchased from Donghong Chemical Co., Ltd., Xuzhou, Jiangsu, China, Quercetin (I), kaempferol (II), and isorhamnetin (III) standards (98% purity) were purchased from Chengdu Must Biological Technology Co., Ltd. . Other chemical regents were analytical grade and obtained from Guoyao Chemical Reagent Co., Ltd. . B-100 rotary evaporator  was used for concentration of samples, ZHWY200D incubator shaker  was used for adsorption and desorption of resins, glass chromatographic columns   was used for dynamic desorption of AB-8 resin.Samples for TGFs analyses were prepared using the acid hydrolysis method described by Hasler and Sticher  with sliTGFsC, C1, C2 and C3 were the TGFs, quercetin, kaempferol and isorhamnetin concentration (mg/mL). TGFs were analyzed using HPLC 1200 system  and a reverse phase C18 column  with a solution system of A (water) and B (methanol). Gradient elution: At 50% B run 0–12 min, 50–80% B 13–15 min, 50% B 15–16 min. The flow ratio was 0.8 mL/min, column temperature was 38 °C, and detection wavelength was 360 nm [s 360 nm ,23. The In order to investigate the static adsorption and desorption properties of these resins, the tests were executed as follows : 5 g (weeQ)/desorption (dQ) and desorption ratios (D), which were calculated using the following equations [The preliminary choices of resins were based on their capacities of adsorption . dQ was the desorption capacity at adsorption equilibrium (mg/g dry resin). oC, e,C and dC were the initial, adsorption equilibrium and desorption concentrations of TGFs (mg/mL). iV and dV were the volume of the initial solution and effluent (mL), respectively. W was the weight of dry macroporous resin (g).The impact of pH on the adsorption capacity of TGFs was studied by mixing 5 g (wet weight) of resin with TGFs solutions (50 mL each) in the pH range of 3.0–9.0. The pH solution was adjusted using concentrated hydrochloric acid or ammonia. Then, the solution was shaken at 30 °C for 8 h in the constant temperature shaking incubator.So as to evaluate the effects of original concentration and temperature on TGFs adsorption, experiments of adsorption isotherm on AB-8 resin were executed. Five groups of 50 mL TGFs solutions with the pH value of 5 at different concentrations were brought into contact with 5 g of wet resins in a constant-temperature shaking incubator at 25, 30 and 35 °C for 8 h. After adsorption equilibrium, TGFs solutions were measured. The Langmuir and Freundlich models, used to evaluate the adsorption behavior, are described in the following equations .Langmuir isotherms:LK (mg/mL) was the Langmuir constants, mQ and 1/n were empirical contants, and FK was the Freundlich constants, which was an indicator of adsorption capacity [Freundlich isotherms:capacity .Dynamic adsorption and desorption tests of AB-8 resin were evaluated using glass columns (500 × 30 mm) with 80 g resin (wet weight). The bed volume (BV) of the resin was 100 mL. After adsorption equilibrium, the column was firstly washed with deionized water, and then eluted with different concentrations of ethanol using the optimized conditions, at a flow rate of 2.0 BV/h. The TGFs effluents were collected every 10 mL for a fraction. The elution volume of each ethanol concentration was adjusted using the HPLC method.2. Aspirin was used as a positive control. Compounds were dissolved in dimethyl sulfoxide to make stock solutions and were kept at −20 °C. The nitrate assay kit was purchased from Beyotime biotechnology company. The final concentration of the vehicle in the solution never exceeded 0.1% and had no effects on NO production and cell viability. 2,2′-Azino-bis (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) were purchased from Beyotime biotechnology company.The mouse Raw 264.7 macrophage cell line was obtained from Nanjing University and was cultured in RPMI 1640 medium  supplemented with 100 U/mL penicillin, 100 mg/mL streptomycin, and 10% heat-inactivated FBS (Beyotime biotechnology) at 37 °C under a humidified air atmosphere of 5% CO50 values were determined as reported above. The DPPH scavenging effect was calculated as follows:s is the absorbance of DPPH alone, and Ai is the absorbance of DPPH in the presence of a specific concentration of TGFs. EGB and ascorbic acid were prepared at the same concentrations and used as reference standards.The DPPH radical scavenging activity of TGFs was measured by the method described by Mishra K ,29 with s is the absorbance of ABTS solution, and Ai is the absorbance of ABTS solution with sample at different concentrations, respectively. EGB and ascorbic acid were also prepared at the same concentrations and were used as reference standards. The antioxidant activity was expressed as the IC50 (mg/mL), which is the TFE concentration that inhibits 50% of the free radicals.The ABTS radical scavenging activity of TGFs was assayed using the method of Dawidowicz and Olszowy  with min4 cells/well, serum-starved for 6 h, and then treated with different concentrations of compounds for 72 h. Next, MTT solution was added to each well, and the cells were incubated for another 4 h at 37 °C. Then, the media were removed, and DMSO was added to dissolve the purple precipitates. The absorbance was measured at a wavelength of 540 nm.For analysis of the cytotoxicity of TGFs, Raw 264.7 cells were plated in a 96-well plate at a density of 3 × 102-), an indicator of NO synthase activity, in culture supernatant fluids was measured based on Griess reaction [4) were seeded in 100 µL of RPMI 1640 into 96-well plates and co-incubated with different concentrations  of compounds in the absence or presence of LPS (500 ng/mL) for 48 h. Meanwhile, aspirin was also tested as a positive control. Culture supernatant fluids were mixed with 100 µL Griess reagent at room temperature for 5 min. Using NaNO2 to generate a standard curve, nitrite production was measured by an absorbance reading at 540 nm.Accumulation of nitrite (NOreaction . Brieflyn = 3). The results were expressed as the means ± standard deviation. The experiment data were analyzed statistically by GraphPad Prism 5. The data were subjected to t tests of column analyses. In all statistical analyses, p values ≤ 0.05 were regarded as statistically significant and p values ≤ 0.01 as very significant.All experiments were performed triplicate samples  in the EGB sharply increased from 8.93%, 9.88%, and 6.11% to 30.12%, 35.21%, and 14.14% in the TGFs. The recovery rate of compounds I, II, and III were 88.76%, 93.78%, and 60.90%. The results verified a good adsorption and desorption chromatography for the preparative enrichment and separation flavonoid O-glycosides from medical herbs. These results of the anti-inflammatory effect evaluation of TGFs showed it could significantly inhibit LPS-induced NO release in vitro compared with the control group in a dose-dependent fashion. This study offers a theoretical basis for further study on bioactive flavonoid O-glycosides from EGB, which shows potential for functional food and pharmaceutical applications in industry.In present study, a method for the preparative separation and purification of ginkgo flavonoid


Few, if any, patient reported symptoms have been shown to be related to objective measures of spine function. Recently, patient-reported measures of disability following spinal manipulative therapy have been associated with an immediate decrease in spinal stiffness obtained by instrumented L3 indentation. Given this novel relation, we anticipate that stiffness measures obtained from locations in addition to L3 may yield valuable information. As such, our research team has developed a new technique to acquire stiffness data continuously over an entire spinal region. The reliability of stiffness measurements obtained by this new technique has yet to be quantified.Continuous stiffness testing employs a weighted roller that moves uninterrupted over the spine while measuring the resulting spinal deflection along a subject-specific, laser-defined trajectory. A volunteer sample of asymptomatic participants were assessed in 2 sessions occurring 1 to 4 days apart, with each session scheduled at the same time of day. Each session consisted of 3 trials each beginning at a baseline of ~ 17 N then progressing to a maximally tolerable load as defined from pre-test familiarization trials . Reliability was evaluated with the intraclass correlation coefficient, the standard error of measurement and Bland & Altman analysis.A total of 17 asymptomatic participants  took part in the study. Overall, the within and between-session reliability of lumbar spine stiffness measures at the maximal tolerable load was excellent ranging from 0.95–1.00 and good to excellent ranging from 0.82–0.93, respectively. Trial averaging was found to reduce standard error of measurement by a mean of 35.2% over all measurement conditions compared to a single trial. Bland and Altman plots for agreement in lumbar spine stiffness measurements varied from − 0.3 +/− 1.2 at unloaded condition to − 0.2 +/− 1.2 at loaded condition. Data from two participants were removed due to the development of back pain between two sessions.This study introduced a new technique for measuring spinal stiffness over an entire spinal region in asymptomatic human participants. The new technique produced reliable measurements quantifying the load-displacement values for within-session and between-session assessments. A significant decrease in the mobility of lumbar spine has been reported as a common sign in individuals with low back pain (LBP) . PreviouDue to low levels of reliability and high variability related to clinical evaluation of spine stiffness, mechanical tools have been developed to quantify the applied loads and tissue displacement that occur during PA testing , 3, 5 thAs such, our research team has developed a novel device to improve on single-site spinal indentation by employing a loaded rolling wheel system. The reliability of stiffness measurements obtained by this new technique has yet to be quantified. Therefore, the objective of this study was to determine the within- and between-session reliability of lumbar stiffness measurements in asymptomatic participants using this new loaded rolling wheel system .A total of 17 consecutive volunteers were recruited using flyers distributed on campus at University of Alberta. The sample size calculation was based on an estimate used specifically for reliability studies . ThirteeStudy participants included asymptomatic males and females between the ages of 18 and 60 with no history of thoracic and lumbar pain within the last 6 months. Participants were excluded from the study if they could not tolerate the stiffness testing procedure, lay prone for 20 min, or had a history of the following: scoliosis, congenital spinal disorders, prior thoracic or lumbar surgery, spondylolisthesis, cauda equina syndrome, current pregnancy, severe respiratory disease, severe trauma, or a medical ‘red flag’ such as cancer, spinal infection, fracture, or systemic disease.A research assistant with 6 years of clinical experience in physical therapy and 1 year of experience using the testing device collected all measurements.The lumbar PA trunk stiffness was assessed with a mechanical device Fig.  whose coStepperonline.com, China) that is connected to a cable which raises and lowers the rollers in conjunction with a string potentiometer to quantify vertical position . Control of all motors and acquisition of signals is provided by in-house coding using LabVIEW   is used to position the roller along the X , Y  axes with built-in encoders to confirm motor position. The vertical Z axis employs a stepper motor system  before and after each session.Standardized instructions were given to the participants before testing which included information about how to hold their breath during testing (held expiration), to remain still during testing and to provide feedback if they experienced pain or felt they were resisting the roller wheels. An inter-wheel spacing of 29 mm was used for all participants in both sessions.To begin using the device, the examiner first manually identified and marked each spinous processes from S1 to T12. The examiner then used the laser system described previously to generate an XY trajectory for the wheels to follow  from S1 to T12. Additional mass was then added in ~11 N increments until a maximum of ~ 83 N or the maximal tolerable load for each participant was reached.Following the familiarization procedure, three trials were conducted per session using the unloaded condition and then three additional trials at the maximal tolerable load condition. Data from these trials were used in the reliability analysis. Figure In addition, before and during the session, participants were asked to rate any testing-related pain using the NPRS. A reported NPRS of ≥2/10 would stop the loading and the prior mass would be considered as the maximum tolerable load .These same procedures were repeated in the second session including the familiarization procedure and the reliability tests. All tests were conducted by the same examiner who was blinded to the stiffness assessment results of the first session. Between sessions, participants were asked to 1) maintain their usual physical activities and notice if any new activities had been undertaken between sessions or if new symptoms were present, and 2) to not wash the spinous process markings on their body so they could be used in the second session.The displacement value for each segment was automatically extracted from a custom program written in LabView and then exported to an Excel file. The roller landing and lifting trajectory points (S1 and T12) of all participants were discarded from the automated extracted data. From the remaining continuous displacement data, stiffness was determined at each of the lumbar spinous process locations with the unloaded roller mass defined as the weight of the apparatus (~ 17 N) and the maximum tolerable load considered as the maximum mass that participants could tolerate with no pain and discomfort  obtained from the familiarization process. Stiffness at each spinous process location was then calculated as a ratio between the applied force and the resultant displacement .3, k) was calculated to estimate the within-session reliability and the between-session reliability for stiffness values at each lumbar segment separately. ICC with k indicating 1 provided estimates of the relative reliability for a single trial, and at k = 3 provided estimates of the relative reliability for the average of 3 trials. This model of ICC was chosen because only one examiner was involved in this study, representing a fixed factor for rater [An Intraclass Correlation Coefficient  which is defined as an estimation of the variability expected for observed values when the actual value is held constant . The folBland and Altman graphs were plotted using the difference in spinal stiffness values between session 2 and session 1 (1 minus 2) against the mean of the 2 test sessions to provide a visual presentation of stiffness variability Fig.  16]. Th. Th16]. All statistical analyses were performed using IBM SPSS statistics, version 24 , . Intraclass Correlation Coefficient values were qualitatively interpreted using the following criteria: 0.00–0.50 = poor, 0.50–0.75 = moderate, 0.75–0.90 = good, and 0.90–1.00 = excellent .Seventeen asymptomatic participants, aged 19–43, and homogeneous in terms of age and body mass index were recruited in this study  for the single measures was estimated from 0.92 to 1.00 for the unloaded condition and from 0.95 to 1.00 for max tolerable load. In addition, the within-session reliability estimates  for the average of the 3 lumbar spine stiffness measurements ranged from 0.97 to 1.00 for the unloaded condition and from 0.98 to 1.00 for maximal tolerable load  ranged from 0.81 to 0.94 for the unloaded condition and from 0.83 to 0.92 for maximal tolerable load. The between-session reliability estimates of lumbar spine stiffness measurements for the mean of 3 trials  also ranged from 0.75 to 0.96 and 0.82 to 0.93 for unloaded and maximal tolerable load, respectively  shows that averaging three repeated measurements reduced the SEM by a mean of 35.2% over all measurement conditions  , and comBland and Altman plots show the majority of observations fall on or very near the mean resulting in a high level of agreement between the two measurement sessions. Any difference in stiffness between sessions may be attributed to individual differences between sessions or individual activities of the participants between sessions. Bland and Altman plots also show less reliability at higher stiffness measurements in both unloaded and loaded conditions. Possible explanations for this observation between sessions may include a variety of patient-based factors such as activity level and apprehension level.The unloaded conditions and the loaded  conditions did not differ significantly in terms of within-session and between-session reliability. This is shown by the ICC confidence intervals presented in Table Our study found that using an average of the three trials to create within-session stiffness values showed a reduction in SEMs as compared with a single trial. This is consistent with previous studies  that shoThe study protocol, which was designed for a research study on reliability, took 30–45 min including the familiarization procedure. Using single trials only, the total time to complete testing is ~ 12 min .While participants returned at similar times on separate sessions, it is currently unclear whether better control of inter-session time intervals and/or activities would improve between-session reliability results; it is impossible to know if a change in reliability in the second session is the result of differences in the participant over time, variability in the measurement process, or both. This is a drawback of reliability testing over multiple days. Furthermore, the measures obtained by a loading device such as this will always be influenced by the viscoelastic properties of the target tissues in their current state. As such, the reliability of this device is dependent on providing adequate recovery time between trials.While we expect that the reliability of the device may change when used to evaluate spinal pathology, this device may be contraindicated in specific pathologies as well . Further studies are needed to define relative and absolute contraindications for VerteTrack use. It is important to note that the reliability of the VerteTrack is likely decreased by patient-based factors such as voluntary/involuntary muscle contraction, changes in patient position during testing and inconsistent patient breathing procedures. Future identification of these factors and the magnitude of their impact is warranted.This study evaluated the reliability of a device capable of measuring spinal stiffness continuously over an entire spinal region in asymptomatic human participants. The new technique was shown to produce reliable measurements in quantifying load-displacement values for within-session and between-session assessments. The resulting data may have greater clinical utility than single site measures in that spinal stiffness can be obtained not only at one level, but over the entire spinal region of interest.


Science and Technology of Advanced Materials (STAM) celebrates its 20th anniversary this year. The journal was launched by veteran STAM editors Tsuyoshi Masumoto and Toyonobu Yoshida in 2000 [ in 2000 . STAM waSTAM the world’s leading open access journal focused on materials science. Further changes were initiated in 2016 by the former STAM Editor-in-Chief Shu Yamaguchi, including the launch of ‘Vision’ [Around that time NIMS also introduced new publication policies in order to widen the areas of research covered by the journal and improve the quality of articles. Those changes resulted in the impact factor rising from 1.267 in 2008 to 4.787 in 2017 – making ‘Vision’  and ‘His‘Vision’ ; and an STAM has been managed by NIMS with the Swiss Federal Laboratories for Materials Science and Technology (Empa) joining NIMS as a STAM sponsor in 2016; the journal is also supported by the World Materials Research Institute Forum (WMRIF). Since 2008, the content of STAM has been archived in the PubMed Central (PMC) database.Since 2008 the publication of STAM has become internationally recognized for publishing timely, seminal articles such as reviews by Hosono et al. [STAM authors also include Nobel Prize Laureates Dan Shechtman [Over the past 20 years, o et al. ,7 and Aro et al. ,9. STAM hechtman , Heinrichechtman , and Ei-hechtman .STAM articles.The publication team and editors are aware that much more work is required to enhance the journal’s visibility worldwide. Furthermore, the journal currently publishes only about 80 articles per year, so we need to increase the number of topics covered, whilst maintaining the quality of STAM as a valuable and readily accessible resource that offers critical insights into the latest discoveries in the constantly changing landscape of materials science.Finally, we welcome your continued support and suggestions for publishing