In this report, synthesis of the starch nanoparticles from underutilized and cheap sources viz: Horse chestnut (HS), Water chestnut (WS) and Lotus stem (LS) by using mild alkali hydrolysis and ultra-sonication process has been presented. The particles were characterized by Differential scanning colorimeter (DSC), X-Ray Diffraction (XRD), Rheology, Scanning electron microscopy (SEM) and Fourier transform infra-spectroscopy (ATR-FTIR). The particle size measurements, functional properties and antioxidant potential of starch nanoparticles were also analyzed. The experimental results revealed that the average particle size diameter of Horse chestnut starch nanoparticles (HSP), Water chestnut starch nanoparticles (WSP) and Lotus stem starch nanoparticles (LSP) was found to be 420, 606 and 535 nm, respectively. We observed a notable increase in the water absorption capacity but decreased capacity for oil absorption in the starch nano-particles. SEM images revealed damaged starch granules after size reduction. Additionally, loss of crystallinity and molecular order was observed from XRD and ATR-FTIR spectra. It was concluded that the starch nanoparticles have better thermal stability, increased viscosity and antioxidant properties. Starch is composed of linear chain molecule; amylose and branched chain molecule; amylopectin. These two starch components are assembled in the form of granules with the size ranging from 1 to 100 µm2. Starch has various applications, it has been used as thickening, gelling, stabilizing in a wide variety of foods and non-food products4. Starch is also used for drug and bioactive delivery systems6. The native form has however many limitations such as poor solubility, retro-gradation, limited digestibility and poor functional properties. For this reason, various physical, chemical and enzymatic methods have been employed to reduce these limitations or to add new attributes. Currently, starch nano particles are gaining more interest for improved quality and wide applications. They have been considered as the promising biomaterials for novel utilization in foods, cosmetics, and medicines as well as various composites7. There are various techniques for starch nano particle preparations including hydrolysis by acid, enzymes or combination of two, regeneration and mechanical treatments using extrusion, irradiation, ultrasound or precipitation by co solvent8. The easy and cost-effective methods for starch and starch derivative nanoparticles are always of paramount importance. Among various such methods, nano-precipation and ultra-sonication are very simple and reliable methods for nanoparticle production with desired size. The precipitation process involves a drop wise addition of a dilute starch solution into a non-solvent and ultra-sonication reduces the size by breaking the covalent bonds in polymeric material due to intense shear forces or mechanical effects associated with collapsing of micro bubbles by sound waves, it is simple, effective and environment friendly procedure10. Starch nanoparticles (SNP) were prepared by using ultrasound method without chemical additives from cassava, corn, yam and other sources of starch11. However the combination of physical and chemical processes produced nanoparticles with more desired properties12. So far starch from various sources like cereals, millets, tubers and others have been extracted and synthesized to nanoparticles15, but the novel and cheap sources are of great importance from commercial point of view. Therefore, we selected the extraction of starch from crops like horse chestnut, water chestnut and lotus stem, as they remain mostly underutilized at commercial level and huge quantum of produce generally goes waste. The starch nanoparticles were synthesized using novel methodology involving mild chemical and mechanical combinations of alkali hydrolysis and ultra-sonication. This technique is simple and convenient in terms of safety, cost and can give better yield with desired particle size. The morphological, rheological, thermal and functional properties of starch nanoparticles were investigated. The study also reported the anti-oxidant properties of starch nano-particles.Starch is a natural biopolymer, which is abundantly found in nature and also major component of our daily diet. It is mainly found in plant roots, staple crops and cereals such as rice, maize, wheat, barley, corn, tapioca, potato and others15 which could be easily broken down by ultra-sonication through a process called cavitation and therefore resulted in smaller size of the particles16. Furthermore, the zeta potential of all starch nano particles was found negative. Zeta potential predicts the long-term stability of the nanoparticle. The WSP displayed significantly higher negative zeta potential (−41.29 ± 3.23 mV) making them more stable than HSP and LSP. The high zeta potential increases the electrostatic repulsions between the particles, which results in the decrease of Van der Waals forces of attraction. The Van der Waals forces are responsible for particle agglomeration that eventually leads to bigger particles17.The average particle size, zeta potential and particle size distribution of starch nano particles of HSP, LSP and WSP are shown in Table 18. The OAC is associated with hydrophobic inner surface of loosely helical chains of amylose that is not able to hold water well, thus lipids or oils replaces them easily and the extent of hydrophobic channel depends on the size of intra helical included molecules19. The decrease in OAC can therefore be correlated with reduction in size and change in the polymeric intermolecular network influencing the hydrophobicity of starch nanoparticles.The water and oil absorption capacity of starch and starch nanoparticles are depicted in Fig. Data with different superscript above the bars are significantly different (p < 0.05).HSP, WSP and LSP represent starch nanoparticles while as HS, WS and LS represent native starch from Horse chestnut, Water chestnut and Lotus stem, respectively. WAC (water absorption capacity); OAC (Oil absorption capacity).−1 which is attributed to the -OH stretching20 and its breadth indicated the extent of formation of inter- and intra-molecular hydrogen. In starch nanoparticles, the peaks of O–H stretching shifted to higher wavelength for WSP, LSP and HSP. This could be attributed to the loss of the crystalline structure and exposure of -OH groups of the starch molecule due to alkalization and sonication process22. The other characteristic bands were observed at wavenumbers of around 2923 cm−1 assigned to -CH2 stretching vibrational modes bands and 1147, 1078 and 990 cm−1 associated with the stretching vibration of the C-O bond, C-O-H and C-O-C groups in the anhydrous glucose ring, respectively23. The characteristic peak at 1643 cm−1 is due to the presence of bounded water in starch and it does not show any evident change in the peak intensity after size reduction of starch. The characteristic peaks at 855, 856 and 867 cm−1 for LSP, WSP and HSP respectively, indicate the existence of β-glycosidic bonds in the samples24.Figure −1 with high peak intensity at 995 cm−1 possess shoulder at the wavenumber of 1018 cm−1 and 1047 cm−1 and final temperatures (TC). The peaks for WS, LS, HS, WSP, LSP and HSP have been recorded between 49.26–124.23 °C, 69.88–125.74 °C, 50.76–119.35 °C, 52.76–134.15 °C, 108.10–148.79 °C, and 51.99–144.61 °C respectively. The peak transition temperatures have been detected at 87.68 °C, 102.36 °C, 83.72 °C, 110.23 °C, 135.80 °C, and 121.81 °C for WS, LS, HS, WSP, LSP and HSP respectively. WSP showed a sharp peak at 135.80 °C corresponding to its melting point and its crystal structure. HSP starch curves displayed broad endotherm glass transition at vicinities 95 °C and a sharp endotherm melting transition has been shown by LSP at 160 °C. The broad glass transition could be ascribed to amorphous structure of starch in addition to endothermic events overlapping at gelation point of amylopectin27 WS, LS and HS showed lower transition temperature which could be attributed to the lower gelatinization enthalpy in native starches having values of −522, −685, −104 J/Kg respectively for WS, LS and HS. Ultra sonication of starch granules results in a serious disruption of the crystalline structure of clustered amylopectin and increase of hydrocarbon chain that can be related to an increase in Van der Waals and hydrogen bonds with greater stability and higher melting temperature apparently leading to nano-particles with low crystallinity or an amorphous character. Peak of the crystallization temperature was not recognized indicating that these nanoparticles have higher melting point and more amorphous structure. Similar results were reported by Hsanvand & Fathi28 in starch nanoparticlesThe thermal characteristics of native starch and starch nano-particles have been performed by DSC. The obtained values for each of the samples have been specified in Fig. 3, the obtained results and graphs were compared in this study with the nanoparticles of starch prepared from same source. The area under amorphous region and diffraction peaks has been calculated according to our previous method3 using Gaussian profile fits. XRD patterns revealed that the relative crystallinity (RC) of starch nano-particles decreased to 1.33, 5.56 and 14.46 in HSP, WSP and LSP, respectively. However, in the native starch the RC was found to be 13, 26 and 22% in HS, WS and LS respectively. The decrease in relative crystallinity can be related to an increase in amorphous region of the starch after size reduction. Nano sized starch showed either narrow or diminished diffraction peaks. The sharp or narrow peaks indicate large crystallite size. The crystallite size of the starch nanoparticles was estimated and found up to 3–10 nm. This also indicates that major peaks of diffraction do not exist and whole structure appeared like amorphous hump. Similar results were obtained from our previous studies where there is decrease in crystallite size of starch after sonication and alkalization of starch3. Methods like gamma irradiation, heat moisture treatment, microwave degradation and ultrasonic degradation have also shown decrease in crystallinity and crystallite size of starch32.Crystal structures of synthesized starch nano-particles have been determined by X-Ray Diffraction (XRD) analysis as shown in Fig. / and Loss modulus G// of starch was plotted against angular frequency (s−1). The G/ and G// of starch nanoparticles was higher than the values obtained for native starch. Further, the difference between G/ and G//of starch nanoparticles was observed higher than native starch samples (Data not shown) suggesting the more elastic behavior of starch nanoparticle suspensions than native starch. The results for frequency sweep tests were also in agreement with the flow rate test conducted against the shear rate ranged from 0–60 s−1. The graph shown in Fig. 34. The rheological properties of the starch are influenced by shape, size and distribution of the granules and also by the amylose content and granule–granule interaction35. According to obtained data, our study concluded with the results that decrease in size of starch at nano level increase its viscosity.Results for the rheological properties of native starch and starch nanoparticles are given in Fig. 36. The granular structure of the native starch particles was smooth and round or oval shaped as was also reported in our previous studies18. The micrographs revealed that starch nano-particle granules are irregular in shape, size with no uniform pattern, which has been ascribed to the cavitation phenomenon inducing very high pressures and shear forces responsible for degradation of the external crystalline and amorphous layers of starch granules and therefore small segments can be produced. The surface appearance change was also observed in oat starch nano-particles by Falsafi et al.37. HSP and LSP showed uneven exterior with voids and their granular structure was found to be disrupted to large extent. LSP had rough surface with many cavities and the surface of WSP was relatively smooth showing few large fragments besides the presence of small particles within granules.Scanning electron microscopy was used to observe the morphological analysis of the starch nano-particles including shape, size and porosity. The micrographs of native starch were also mentioned in our previous study. As shown in Fig. 38. Moreover, it is known from studies that low molecular weight and monosaccharide composition can contribute to the antioxidant activity of polysaccharides39. Formation of nano-particles is accompanied with a decrease of molecular weight, hence improving the antioxidant potential. It is reported that ultrasonic treatment reduces the molecular weight and viscosity of starch solution40. Starch from some sources contain flavonoids and several studies have shown that flavonoids possess significant superoxide radical scavenging activity42. Superoxide anions have been observed to directly initiate lipid peroxidation, thus leading to oxidative damage in proteins, lipids and DNA. Lipid peroxidation was reported in the following trend: WS (15.26 ± 0.85%), LS (18.51 ± 2.08%), HS (16.48 ± 0.16%), WSP (20.70 ± 1.38%), LSP (22.09 ± 0.85%) and HSP (27.80 ± 1.65%).The results obtained were statistically significant (P < 0.05). WS, LS and HS inhibited lipid peroxidation at low extent  and Himedia Laboratories Pvt Ltd. WS and LS was purchased from the local market of Srinagar, India, while HS were collected from the trees located in University of Kashmir, Srinagar, India at their maturity stage. The samples have been cleaned, de-hulled and then stored at 5 °C.3.The samples of HS, WS and LS were cut into small pieces after de-hulling/peeling and then grounded along with some quantity of water in a domestic blender. The starch has been extracted using the alkaline steeping methodThe synthesis of starch nano-particles has been achieved by ultra-sonication and mild alkali hydrolysis. Starch solutions (1.5%) was preheated at 80 °C in 0.1M NaOH solution and continuously stirred for 30 min using magnetic stirrer. The starch slurry obtained was then sonicated at 40 KHz using the fixed probe sonicator . The sonication time of 30 minutes was carried out at intervals of 5 minutes to avoid excessive heating. The resulting solution was then co-precipitated by ethanol in the ratio of 1:2 by drop wise addition of slurry to ethanol under continuous magnetic stirring. The precipitate was then collected by centrifugation of solution at 8000 rpm for 15 minutes and then lyophilized using freeze dryer . The powdered samples were stored at 4 °C for further analysis.In order to characterize particle suspension, particle size and zeta potential of starch nanoparticles is determined using particle size analyzer . The 0.01% of sample has been dispersed in millo-Q water and sonicated for 30 minutes at 40 KHz in sonicator bath before particle size measurement. For measurement of zeta potential the 0.01% of sample was suspended in 0.1 mM KCl with pH adjusted to 6 and the samples have been equilibrated overnight before measurement.One gram of each starch sample was mixed with 10 mL distilled water or vegetable oil and continuously stirred for 30 min at room temperature. The slurry was then centrifuged  for 15 minutes at 5000 rpm. The formed supernatant was decanted, and water/Oil absorption capacity was expressed as ratio of initial weight of sample to final weight of sample after oil/water absorption.−1. The FTIR spectra was examined by using Agilent Pro resolution software.The ATR-FTIR spectra of samples have been recorded on an Agilent FTIR-Cary 630 at room temperature at the wavelength region between 4000 and 650 cme systems, Mettler Toledo). The samples (5 mg) were placed in aluminum pans and 5 µl of milli Q water was added and mixed uniformly with the sample. The pan was hermetically sealed and heated from 20 °C to 200 °C at the rate of 10 °C/min. Characteristic temperatures of transitions were defined as To(onset), Tp (peak of gelatinization), Tc (conclusion) and enthalpy of gelatinization (ΔH) was recorded.The thermal behavior of samples was studied using DSC . The operating conditions were X-ray line (λ = 1.5418 Å), voltage 40 kV and current 30 mA. The samples were loaded on aluminum plate and X-ray diffractions recorded from 10° to 50° for 2θ with scanning rate of 0.02/min. The relative crystallinity (RC) of starch was calculated as described by Rabekα X-ray line, K = shape factor (=0.9), β = full width of the diffraction peak at half maximum, and θ = Bragg’s diffraction.The crystallite size of the starch nano-particles has been calculated using Scherrer equation as given below:The 6% starch nano-particles in water was heated to 90 °C for 20 minutes with constant gentle stirring till the gel was formed. The gel was allowed to cool at room temperature and rheological measurements were conducted using rheometer  at 25 ± 0.1 °C on parallel plate geometry . The flow properties were determined with increasing shear rate, from 0.1 to 100 s and the delay time was 10 s. The viscoelastic range was determined through stress sweep tests at constant frequency of 1 Hz with a logarithmic increase of shear stress from 0.01 to 100 Pa.Microstructure of starch nano-particles have been monitored by scanning electron microscope . The samples were gold plated and different micrographs of samples were recorded.The starch samples (20 mg/ml) was mixed with dimethyl sulphoxide (DMSO) and heated continuously till complete dissolution.et al.9 with modifications. Briefly, 200 µL of each sample was added to 200 µL of a 0.01% of DPPH solution dissolved in dimethyl sulphoxide (DMSO). The final volume was adjusted to 3 ml by adding 2.6 ml of DMSO and incubated at 37 °C for 30 min. After incubation, absorbance was measured at 517 nm against a control (without sample). Percentage inhibition was calculated by using the formula:The scavenging activity of 1,1-dihpenyl-2-picrylhydrazyl(DPPH) radical was measured according to the method described by Ahmad 2O2 (30 mM), 0.2 mL of ascorbic acid (100 mM) and 0.2 mL of ferric nitrate (20 mM). After this, the mixture was incubated at 37 °C in water bath for 1 h. The reaction was stopped by the addition of 1.0 mL TCA , following with 1.0 mL of TBA  and then tubes were placed in a boiling water bath for 20 min, which is followed by centrifugation at 5000 rpm for 15 minutes. The clear upper layer was collected and its absorbance was measure at 535 nm. Percentage inhibition was calculated by using equation.To measure the lipid peroxidation inhibition activity of samples, 100 µl of samples were added to1 mL of 0.2% linoleic acid, 0.2 mL of HAll the analytical experiments have been carried in triplicate and statistical analysis was performed using commercial statistical package SPSS.10.1 (USA) and the data were assessed by analysis of variance (ANOVA) using Duncan’s multiple range test at 5% significance level. The results are presented as means ± standard deviation (n = 3).


The COVID 19 pandemic has exposed the vulnerability of many families, including grandparent kinship families, to deal with a health/economic crisis. The fear of COVID-19 plus stay-at-home orders have increased individuals’ psychological distress. Moreover, school closures and homeschooling further increased parenting stress among caregivers. This study examined the relationship between material hardship and parenting stress among grandparent kinship providers and assessed grandparents’ mental health as a potential mediator to this relationship during the COVID-19 pandemic in the United States. Grandparent kinship providers (N=362) that took primary care of their grandchildren participated in a cross-sectional survey via Qualtrics Panels in June 2020 in the United States. Descriptive and bivariate analyses, binary logistic regression, and mediation analyses were conducted using STATA 15.0. Experiencing material hardship  was significantly associated with higher odds of parenting stress among grandparent kinship providers, and grandparents’ mental health  partially mediated this association. Addressing material and mental health needs among grandparent kinship providers is critical to decreasing their parenting stress.


Thoracic aortic graft infection (TAGI) is a rare and serious complication after surgery for which the treatment is controversial and challenging. Rather than following the traditional surgical strategy of graft replacement and extensive debridement, we have chosen to preserve the graft and cover it by a laparoscopic omental flap. In this article, we describe the clinical manifestation, diagnostic modalities, and treatment of this disease and analyze the role of laparoscopic omental flaps in its treatment.We present two cases of thoracic aortic graft infections that had undergone surgical graft replacement for acute Stanford type A dissection. Their clinical manifestation of infection was atypical, with computed tomography suggesting infection of the grafts. Both patients were successfully treated with debridement, laparoscopic omental transposition, and antibiotics. The first case, a 55-year-old male, was found to have an infection at the aortic arch. The second case is a 52-year-old male who was found to have infection at the ascending aorta and arch. Surprisingly, both intraoperative cultures were negative. The infections were brought under control and the patients recovered steadily after surgery. Early follow-up results showed no signs of graft infection.These findings suggest that graft replacement for the treatment of TAGI is not always necessary in selected patients. Conservative surgical treatment, including laparoscopic omental transposition, is effective and less invasive for treating TAGI. Graft infections are associated with aortic valve insufficiency, peripheral arterial embolus, false aneurysm, rupture, chronic draining of external sinuses, aortocutaneous fistula, aortobronchial fistula, hemolysis, graft deformity, chronic sepsis, septicemia, shock, and death [Thoracic aortic graft infection nd death . Most cond death . Here, w9 cells/L.A 55-year-old man was readmitted to our institute for poor chest wound healing on May 25, 2019. Four weeks previously, he had undergone a Bentall procedure using a No. 23 aortic valved graft  and a replacement of the ascending aorta and total arch with a 28-mm Dacron graft  and implantation of a frozen elephant trunk  under deep hypothermic circulatory arrest and selective antegrade cerebral perfusion for acute Stanford type A dissection. We used two chest plates to the fixed sternum in a closed procedure. He was finally discharged with a healed wound. Physical examination upon readmission showed a sternal upper middle section of skin ulcer about 2 × 4 cm, the chest exposed with a yellow purulent secretion, and normal temperature (36.8 °C). Laboratory investigation revealed mild leukocytosis of 10.8 × 10Healing was slow despite supportive therapy and debridement. The ulceration was found to be concentrated on the upper part of the sternum plate. Considering that the patient’s failure of wound healing was caused by an allergic reaction to the sternum plate, the sternum plate was taken out, the necrotic tissue was cut off, and the wound healed completely. However, a 1-cm fluctuant area developed in the upper part of the healed sternotomy incision. A few days later, a copious amount of yellow purulent fluid was drained and formed a sinus tract. Specimens from the pus collection cultures for bacterial screening were negative.We then performed a sinogram. The contrast agent was injected into the sinus tract until overflow. Then a 64-slice chest computed tomography (CT) scan was performed to determine the direction, length, and range of the sinus tract. CT examination showed that no graft was involved. Then we continued to plug the drainage strip after debridement every day. The sinus tract was still challenging to heal. CT showed soft tissue and ectopic gas surrounding the ascending aortic graft having a diameter of 14 mm at 7 weeks after surgery  in the Bentall procedure. Physical examination on readmission showed a skin ulcer on the upper sternum of about 1 × 1 cm with pale yellow purulent discharge. The patient’s body temperature was normal (37.1 °C), and the white blood cell count was 8.1 × 10The patient was treated with debridement daily. However, there was no progress in wound healing. The skin of the upper sternum formed a sinus tract, and the first sinogram with CT showed that infection did not involve the aortic graft. We continued to plug a drainage strip with daily dressing change, but the tract was slow to heal. Bacterial cultures were negative. The second sinogram showed some contrast agent surrounding the displaced aortal arch with a bit of gas Fig. . ThorougTAGI after surgery is a rare but serious complication, with an incidence of approximately 1–3% . PatientStaphylococcus aureus [Propionibacterium species. The most common etiology of graft infection is usually a surgical site infection (SSI), followed by seeding of distant infection sites [The primary causative pathogens are gram-positive organisms such as s aureus . Pineda s aureus  reportedon sites . The twoThe diagnosis of TAGI requires a multidisciplinary approach. Contrast-computed tomography (CT) is effective and can show perigraft fluid at low density, ectopic gas, perigraft structures with an increased amount of soft tissue (> 5 mm) between the graft and the surrounding sac, and pseudoaneurysm formation . CT-guidThere are various treatments for TAGI that are mainly divided into repeated in situ graft replacement and graft-sparing strategies. Historically, patients have been treated with radical debridement and in situ graft replacement. However, graft replacement is not available or practical in routine clinical practice, and it is also associated with severe complications and high mortality. In 1984, Hargrove et al.  reportedThe conservative approach of thorough debridement and greater omentum wrapping for revascularization is more practical. The key to the operation is the removal of infected and necrotic tissues and local revascularization. If the local blood supply is not enough, the infection will be difficult or worse to control, especially in patients with grafts. After debridement, the omentum, pectoralis major muscle flap, or rectus muscle flap can be used to fill the retrosternal defect. We used the omentum in both present cases. The omentum is a bulky, well-vascularized flap that can reach the base of the neck and fill the sternal defect. It has a strong anti-infection and repair function and can treat the defect well. Because of its special physiological functions, the greater omentum has been widely used in thoracic surgery, breast surgery, for burns, and in other fields , and itsWith the development and popularization of laparoscopic technology, we creatively adopted laparoscopy to obtain a greater omentum pedicle, which has the advantages of minimally invasive incision, favorable appearance, and fast postoperative recovery compared with traditional laparotomy . We founAlthough the bacterial culture results of case 1 and case 2 were negative, intraoperative findings combined with preoperative CT images still indicated a diagnosis of TAGI. Early literature indicated that a considerable number of patients (> 90%) show positive intraoperative microbiological specimens . HoweverLaparoscopic greater omental transplantation for the treatment of TAGI has been rarely reported around the world, though it has the following advantages: (1) There is relatively little surgical trauma, it is simple and easy to perform, and it is easily accepted by most elderly and frail patients. (2) The omentum is rich in vascular and lymphatic tissue, which gives it a strong ability to resist infection and repair. Auxiliary washing and systemic antibiotic treatment can maximize the control of infection. (3) The greater omentum has more bulk and stronger compliance than muscle flaps, and it can fill defects fully. (4) A laparoscopic omental flap reduces hemorrhage and the incidence of postoperative abdominal complications. However, disadvantages include (1) the abdominal cavity loses the function of the omentum to envelop inflammation, leading to peritonitis in the case of acute abdominal diseases, and (2) the infected area communicates with the abdominal cavity through the omentum, and there is a risk of abdominal infection. Neither patient in the present study had an abdominal infection.Some of our recommendations include the following: (1) Stop warfarin in time, and intravenous vitamin K antagonism is necessary to avoid excessive hemorrhage. (2) After anesthesia, diluted methylene blue can be injected through the sinus orifice for sinus tract staining to define the scope of the sinus tract and facilitate intraoperative debridement. (3) The sternum should be split as completely as possible during the operation. (4) Great care should used during debridement, because the graft becomes brittle after infection.TAGI presents a serious challenge to surgeons, and its management is still controversial. The greater omentum plays an important role in conservative treatment. Conservative surgical treatment, including laparoscopic omental transposition, is effective and less invasive for treating TAGI.


Given concern for increased aerosolization during intubation of patients with severe acute respiratory syndrome coronavirus, we sought to create a portable, inexpensive, and easily constructed device to help protect healthcare workers.A respiratory protection unit can be constructed in approximately 30 minutes and for less than 50 United States dollars in materials, using polyvinylchloride pipe and automobile collision wrap.This device provides possible increased protection during video laryngoscopy and can easily be replicated. Based on current evidence, the novel human coronavirus that is named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been found during experimentation to remain viable when aerosolized for three hours.The following materials were used to construct the portable respiratory protection unit (PRPU): polyvinyl chloride (PVC) ½ inch pipe; PVC joints; and 36-inch wide automobile plastic collision wrap .PVC pipe can be purchased in one 10-foot piece that can be cut up into 13 pieces with a miter saw or hacksaw. Completed box dimensions are 24 inches tall and 27 inches in length. Three-way fittings are used on superior surface for the four joints and two 90-degree fittings are used on base corners. On the posterior  surface, two “T” fittings are used. Following frame construction, automobile collision wrap is used to cover each side. Collision wrap is often used in the automotive industry to cover broken car windows and is adhesive on one side. The adhesive side of the wrap faces into the box. Each box can be constructed in approximately 30 minutes by two people . Total mWhat do we already know about this clinical entity?Given concern for healthcare worker exposure to aerosolization during intubation of the COVID+ patient, protective barriers have been suggested as a way to lessen risk.What makes this presentation of disease reportable?We present an inexpensive device that requires little construction.What is the major learning point?A portable respiratory protection unit can easily be replicated at low cost and used in the emergency department setting.How might this improve emergency medicine practice?Healthcare workers can be provided with some protection during airway management.During intubation, the box is laid over the patient to cover his head and upper chest. Using a knife or any other sharp device, two vertical incisions can be made to place hands through . EquipmeThe PRPU can be assembled rapidly and easily and materials should be readily available in most countries. If auto collision wrap is not readily available, a substitute plastic material could be used; however, depending on the opacity of the material it may further limit direct visualization of the patient. The PRPU use could be expanded beyond the emergency department setting. Other potential uses include protective covering for patients during emergency medical services transport or hospital transport.Given the ever-changing situation and time-sensitive nature of disseminating this model, we do not have time to trial this model. By no means does this model completely contain aerosol viral particles; therefore, appropriate personal protective equipment should still be worn when intubating any known positive or suspected SARS-CoV-2 patient.The PRPU is an inexpensive and quick to construct a protective device for healthcare workers to use during intubation of high-risk patients such as those with SARS-CoV-2 The device is easy to reuse and has many additional applications.


The best cross-validated PLSR model explained 56% of the measured variation and yielded a root mean square error (RMSE) of 17 mg g–1 DW. Model performance decreased when applying the trained model on the independent test set . Different variable selection methods did not increase model performance. Once validated in the field, the non-destructive spectral method presented here has the potential to detect large differences in leaf starch content of red clover genotypes. Breeding material could be sampled and selected according to their starch content without destroying the plant.Grassland-based ruminant livestock production provides a sustainable alternative to intensive production systems relying on concentrated feeds. However, grassland-based roughage often lacks the energy content required to meet the productivity potential of modern livestock breeds. Forage legumes, such as red clover, with increased starch content could partly replace maize and cereal supplements. However, breeding for increased starch content requires efficient phenotyping methods. This study is unique in evaluating a non-destructive hyperspectral imaging approach to estimate leaf starch content in red clover for enabling efficient development of high starch red clover genotypes. We assessed prediction performance of partial least square regression models (PLSR) using cross-validation, and validated model performance with an independent test set under controlled conditions. Starch content of the training set ranged from 0.1 to 120.3 mg g Lotus japonicus L.) or red clover  by bonds within specific functional groups of molecules. These absorptions are mirrored to the NIR region  are routinely used to estimate biochemical compounds such as lignin, cellulose, starch, sugars and proteins in numerous crops . These tR region . MultivaR region . NIRS orThe following study aimed at developing a non-destructive spectroscopic method to estimate leaf starch content in fresh leaf tissue of red clover. Although developed in the greenhouse under controlled conditions, such a method could, once validated in the field, enable to monitor starch turnover on the same genotype over a longer period, under different management regimes, and under various environmental conditions.® fluorescent lamps  at a ratio of 3:1; total light intensity 200 μmol m–2 s–1). Samples were taken at the end of the night , and at the end of the day . Single plants were selected from cultivars from Europe and from breeding germplasm. In addition, nine plants from an advanced breeding population previously shown to have a high variation for leaf starch content, were selected  of plants grown in two separate experiments. Plants from the two experiments were grown in a completely randomized design. The plants of the test and the training set were grown in spring and autumn 2018, respectively. All plants were clonally propagated using cuts that contained only one shoot and one root meristem to ensure comparable physiological states of all plants. These clonally propagated plants were grown in a climate chamber for 90 days before harvesting , and Groluxselected . Wet labThe training set included 18 genotypes, six thereof clonally duplicated . Starch Leaflets were cut using a round, sharpened tube with a diameter of 12 mm to standardize leaf area . These lAfter taking spectral measurements, whole plants from the training set were cut 2 cm above ground, flash-frozen and freeze-dried for starch quantification.Starch in leaf cuts and whole plants was quantified using a protocol of Statistical analyses were performed using the R statistical software version 3.6.0 . As assuSpectral analysis was realized using the R package simplerspec . The meaR2) and the slope (b) of a linear regression with intercept, by the root mean square error , by the bias or mean error (Eq. 2), and by the ratio of performance to deviation . The evaluation metrics were calculated by aggregating all holdout predictions from the repeated 10-fold cross-validation (yi) and corresponding observed values (yi) grouped by ncomp.Leaf reflectance data from the training set was modeled by PLSR with the orthogonal scores algorithm  is a measure of variable importance tailored to PLS regression . VIP scowaj are the PLS regression weights for the ath component for each of the wavelength variables and SSa is the sum of squares explained by the ath component (Eq. 4). The sum of squares SSa for the ath component was calculated from the score qa of the predicted variable y and the ta scores of the spectral matrix X (Eq. 5):where Variable influence on projection scores scores were also used to filter important predictors with a threshold of VIP > 1 within the training set, and the identified predictors were used to re-calibrate the test set and assess performance. This separation in the VIP filtering by independent tests was needed to avoid overfitting and over-optimistic assessment that typically occur when identifying subsets of features on the modeling data.r) were taken to re-perform PLS regression. Second, four wavelengths that were assigned to starch in previous literature were taken and normalized with the reflection at the wavelength that had the smallest standard deviation across the entire wavelength range, prior to performing a multiple linear regression  methods were tested on the independent test data set (60 samples). The predictive ability of these final models was again evaluated using –1 DW. For the samples harvested at the end of the night (EN), starch concentrations ranged from 0.1 to 47.8 mg g–1 DW, with a median of 9.6 mg g–1 DW. Starch concentrations for the test set ranged from 26.41 to 125.44 mg g–1 DW for ED harvested samples, with a median of 66.18 mg g–1 DW. Plants harvested at EN had lower starch concentrations, ranging from 3.66 to 79.51 mg g–1 DW, with a median of 23.28 mg g–1 DW. Differences between samples harvested at ED and EN were statistically significant (p < 0.5) for both sets (–1 DW (data not shown). Also, there was no substantial difference in dry matter content (dry weight/fresh weight) observed between samples from different plants or sampled at different time points, indicating that water content per se was not responsible for the differences in starch content observed by spectral analysis (data not shown).Starch concentration of samples of the training set harvested at ED ranged from 2.0 to 120.3 mg starch per g dry weight (DW), with a median of 46.3 mg goth sets . In ordep < 0.05) starch concentration when compared to mature leaves, old leaves or entire plants (p < 0.05), but a high variation within genotypes was observed , using seven PLS components (R2CV) of 0.56, a RMSE CV of 17 mg g–1, and a residual bias of −0.2  using the best training PLSR model calibrated with pre-processed spectra  yielded a substantially lower R2 of 0.36 and larger RMSE of 29 mg g–1 DW (Independent test set predictions (g g–1 DW . The thrg g–1 DW .Model development using different filtering methods such as variable importance in the projection (VIP), the top 50 starch correlated wavelengths (PCC), multiple linear regression (MLR) before performing partial least square regression (PLSR). Best model performance of each filtering method determined by five time’s repeated 10-fold-cross validation was used to estimate leaf starch content of an independent test set.R2) reached 0.99 for nitrogen and starch contend of cotton leaves , respectively, showing high variation within genotypes. The best PLSR training model explained 56% (R2 = 0.56) of the measured starch variation with an RMSE of 17 mg g–1 DW. The ratio of performance to deviation (RPD) followed the trend indicated by R2 values . Low and high weight sugars were estimated separately and best model prediction for the high weight sugars using PLS regression resulted in an R2 of 0.68 and a RMSE of 19.9 mg g–1. Assigning two-third of the data to calibration and using the remaining data for validation resulted in a slightly lower model performance [R2 = 0.63 and RMSE of 21.6 mg g–1 . These WSC cultivars can substantially increase animal performance and nitrogen use efficiency in pasture-based animal production systems . Red cloThis study is unique in developing and testing a non-destructive method to predict leaf starch content in red clover plants. The described method is suitable to differentiate between high and low starch content in red clover genotypes. Unfortunately, model performance is not sufficient to trace small changes in starch accumulation. Therefore, the method is only partially suited to monitor starch metabolism in detail or to investigate the effect of environmental influences or management regimes throughout an entire season on the same plant. We suggest follow up studies to enlarge the current red clover starch spectral library by means of additional measurements from different independent trials, covering both genotypic and phenotypic variation in starch and to validate the method under field conditions. Currently, the level of resolution is sufficient for the method to differentiate high variance in starch and thus, can be integrated into existing breeding programs to get a rough estimate on starch levels of different red clover cultivars under controlled conditions.https://zenodo.org/record/3598699#.XhOLvdko_d4.The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: RK, BS, and LF conceived the study. LF and PB performed the research, analyzed the data, and drafted the manuscript. RK, BS, and HA assisted with data analysis, interpreting the results, and drafting the manuscript. All authors read and approved the final version of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Over the past 20 years, vaccinologists, bioinformatics experts, and advanced programmers based in Providence, Rhode Island, USA have advanced the development of an integrated toolkit for vaccine design called iVAX, that is secure and user-accessible by internet. This integrated set of immunoinformatic tools comprises algorithms for scoring and triaging candidate antigens, selecting immunogenic and conserved T cell epitopes, re-engineering or eliminating regulatory T cell epitopes, and re-designing antigens to induce immunogenicity and protection against disease for humans and livestock. Commercial and academic applications of iVAX have included identifying immunogenic T cell epitopes in the development of a T-cell based human multi-epitope Q fever vaccine, designing novel influenza vaccines, identifying cross-conserved T cell epitopes for a malaria vaccine, and analyzing immune responses in clinical vaccine studies. Animal vaccine applications to date have included viral infections of pigs such as swine influenza A, PCV2, and African Swine Fever. “Rapid-Fire” applications for biodefense have included a demonstration project for Lassa Fever and Q fever. As recent infectious disease outbreaks underscore the significance of vaccine-driven preparedness, the integrated set of tools available on the iVAX toolkit stand ready to help vaccine developers deliver genome-derived, epitope-driven vaccines.Computational vaccinology includes epitope mapping, antigen selection, and immunogen design using computational tools. Tools that facilitate the  Over the past 20 years, academic researchers and commercial companies have developed immunoinformatics tools to discover the T cell epitope “triggers” that activate the immune system and have applied these tools to vaccine design. The pace of algorithm development for discovering epitopes and designing vaccines has recently accelerated, due to renewed interest in epitope-based vaccines generated by the introduction of immune checkpoint inhibitors to the field of oncology  and the This review will describe vaccine design tools developed by the authors and their application to epitope discovery, antigen engineering, and immunology research in wide-ranging collaborations, as illustrated in several case studies. Starting with T cell epitope discovery, we illustrate how T cell epitopes are relevant to vaccine safety, efficacy, antigen characterization, antigen engineering, and vaccine design. We believe that T cell epitopes deserve greater focus in vaccine development, because even in the absence of effective antibody response, T cell epitopes are important drivers of immune defense against pathogens and may also facilitate their escape from immune defense. Specifically, new information on cross-conservation between pathogen T cell epitopes and the human genome (and microbiome) is emerging, and has important implications for vaccine design. For example, “memory” of cross-conserved T cell epitopes has been defined as a key contributor to the strength of protection generated by vaccines –4. TheseOne of the newest tools described here addresses the concept of immune camouflage . BeginniJanusMatrix that triages human-like epitopes from others. We highlighted the important role of human-like T cell epitopes in the abrogation of effective vaccine response. Furthermore, we used a tool called OptiMatrix to design immune-engineered vaccines that enhance immunogenicity by removing human-like epitopes  and EpiCC (epitope content comparison), respectively. In the sections to follow, we will describe the newest iVAX tools, illustrate the application of the tools to current vaccine development programs, and highlight recent collaborations related to the development of vaccines against a range of human and animal pathogens.After validating the immune camouflage hypothesis in retrospective  and prosepitopes . In otheIn silico prediction of immune response to emerging infectious diseases and cancers can accelerate the design of novel and next generation vaccines. The iVAX toolkit is an integrated set of immunoinformatics algorithms that has been in development since 1998. It comprises a suite of immunoinformatics algorithms for triaging candidate antigens, selecting immunogenic and conserved T cell epitopes, eliminating potential regulatory T cell epitopes, and optimizing antigens for immunogenicity and protection against disease. While aspects of the toolkit have been published , class The “iVAX toolkit” is currently in use by EpiVax, Inc. for clients that include commercial firms and academic collaborators developing novel vaccines for malaria, Q fever and Burkholderia for commercial development and biodefense contracts, and by academic groups that are engaged in federally funded programs for analyzing immune responses to emerging/re-emerging pathogens such as influenza  and pertussis . Even more recently, EpiVax has created a seamless pipeline for the design of personalized cancer vaccines that capitalizes on iVAX and its vaccine development and T cell immunology expertise called Ancer. Collaborative studies carried out using iVAX have demonstrated the importance of computational tools for vaccine design and vaccine efficacy studies. Additional collaborations to evaluate the potential efficacy of existing vaccines against newly emerging variants, to predict immune response to candidate antigens from newly emerging strains of infectious diseases, and to design novel vaccines that have enhanced immunogenicity at an accelerated pace are possible and encouraged.Traditional cancer vaccines based on Tumor Associated Antigens (TAAs) have for the most part failed in clinical trials –82. EvidMeanwhile, the field of cancer therapy has undergone a major transformation in less than a decade due to the introduction of checkpoint inhibitors (CPI), next generation sequencing (NGS) and new emphasis on neo-epitopes. Neo-epitopes are T cell epitopes that contain tumor-specific non-synonymous amino acid mutations that effectively distinguish cancer (tumor) proteins from their normal counterparts. Neo-epitopes represent mutations to self-antigens that are recognized as “Non-Self” by the individual's own immune system, and are therefore capable of generating a potent immunogenic and clinical response. New strategies for developing personalized vaccines that direct immune responses to patient-specific T cell neo-epitopes are in development.Unlike TAAs, cancer neo-epitopes are not found in normal tissue. Also, unlike TAAs that may be present in more than one tumor type and across patients, the vast majority of neo-epitopes are unique to each patient's tumor. The cost of sequencing individual tumor genomes to discover neo-epitopes dropped precipitously to a few thousand dollars per genome in 2018. When combined with CPI, therapeutic vaccines based on neo-epitopes are highly likely to improve outcomes and may well be effective as single agents in certain clinical settings.Beginning in 2016, EpiVax developed an end-to-end immunoinformatics platform, separate from iVAX, that enables the rapid design of personalized, mutanome-directed therapeutic peptide cancer vaccines. These vaccines target multiple antigens specific to a patient's tumor, encoding several class I, and or class II HLA restricted neo-epitopes.This proprietary immunoinformatics pipeline, which includes EpiMatrix and JanusMatrix and a version of iTEM called J-iTEM (see above descriptions) is called Ancer, and has been exclusively licensed to EpiVax Oncology, Inc., a subsidiary of EpiVax, established in 2017. EpiVax Oncology will use Ancer to develop individualized cancer vaccine by screening each cancer mutanome to identify tumor-specific neo-epitopes using EpiMatrix, filter out neo-epitopes that may be tolerogenic using JanusMatrix, and rank the remaining neoantigens according to their optimal immunogenic profile. While the number of companies that are developing tools for identifying neo-epitopes is rapidly expanding, having access to high throughput, commercial-grade and validated tools that incorporate consideration of the rules of self-tolerance will be critically important for the design of more effective and safer vaccines for cancer immunotherapy.The developers of the iVAX toolkit and the Ancer pipeline can imagine a not-to-distant future when many “immunotherapies”—not only for cancer, but also for infectious diseases, autoimmune disease, and allergies, can be designed and administered within weeks of diagnosis. We also anticipate broader application of the tools to pathogens afflicting food animals, including fish, and companion animals, as quickly as additional epitope prediction capacity can be added. Twenty years of computational development and collaborative validation studies have built a strong foundation. Computational vaccinology has a bright future and enormous potential to improve human and animal health.martinb@epivax.com.The datasets analyzed in this article are not publicly available, as they are part of ongoing research and will be published at a later date. Requests to access the datasets should be directed to William Martin, All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.AN and VK are employees of Leidos, Inc., the prime contractor for the Malaria Vaccine Development Program (MVDP), Contract AID-OAA-C-15-00071, with the Office of Infectious Diseases, Bureau for Global Health, U.S. Agency for International Development (USAID). AD and WM are senior officers and shareholders, and LM, FT, AG, PH, GR, and MA are employees of EpiVax, Inc., a privately owned biotechnology company located in Providence, RI. These authors acknowledge that there is a potential conflict of interest related to their relationship with EpiVax and attest that the work contained in this research report is free of any bias that might be associated with the commercial goals of the company. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Durum wheat is an important food source in Mediterranean countries, and Italy is the major producer of durum wheat in Europe. The quality of durum wheat flours depends on the type and amount of gluten proteins and starch while flour nutritional value rests on metabolite contents such as polyphenols. In this work, two Italian cultivars, Iride and Svevo, were analyzed for two years (2016–2017) in four Italian regions to explore how the environment affects: (i) reserve proteome; (ii) starch content and composition; and (iii) free, conjugated, bound phenolics and antioxidant capacity. The impact of environmental and meteorological conditions was significant for many traits. Regardless of the cultivation site, in 2017, a year with less rainfall and a higher temperature during grain filling, there was an increase in low molecular weight glutenins, in the glutenin/gliadin ratio and in the A-type starch granules size, all parameters of higher technological quality. In the same year, the cultivars showed higher amounts of polyphenols and antioxidant capacity. In conclusion, the two wheat cultivars, selected for their medium to high yield and their good quality, had higher performances in 2017 regardless of their sowing locations. Triticum turgidum L. subsp. durum (Desf.) Huns) is an important food source with a relevant role in the human diet, providing carbohydrates, proteins and energy to the human diet [pro capite consumption rates of pasta in the world. Therefore, great effort has been utilized over the years to select durum wheat genotypes with the best agronomic and technological performances [Durum wheat : Iride and Svevo, grown in four Italian regions with different pedoclimatic conditions, in two growing years. In particular, the analysis involved: (i) the content and composition of storage gluten proteins; (ii) the amount of starch granules, both A and B-types; (iii) the amount of phenolic acids and of their antioxidant properties. The differences observed are discussed in relation to both genetic and environmental factors. This work reports a comparison of the flour quality of two Triticum durum Desf.) were utilized in this study: Iride and Svevo both of which were released in 1996 by PSB s.p.a. . Iride is one of the most cultivated durum wheat cv in Italy, in particular, for the exceptional fertility of the ear and the excellent environmental adaptability, whereas Svevo has an extraordinary aptitude for industrial transformation. The main characteristics of the two cvs are shown in more detail in Two cvs of durum wheat  in four locations: Argelato (BO) , Tolentino (MC) , Foggia (FG) , and S. Stefano Quisquina (AG) . The agronomic management followed a standard protocol for the Italian durum wheat variety trials. In each environment, rainfall distribution and temperatures were recorded during the 2016 and 2017 cropping seasons and are available within the national trial dataset. All grain samples were collected after harvesting and kindly provided by the Council for Agricultural Research and Economics, Research Centre for Cereal and Industrial Crops (CREA-CI) of Foggia .Gluten proteins , were extracted from wholemeal wheat semolina (30 mg) using a previously described sequential extraction procedure . The amoSDS-PAGE was performed in a Mini-PROTEAN Tetra Cell (Bio-Rad) using 12% for the LMW-GS and gliadins, and 8% for the HMW-GS of acrylamide gels, as previously reported . LMW-GS,Wheat starch was isolated from wholemeal semolina by using the protocol described by Budney and colleagues  with somg for 10 min at 4 °C. The pellet was then subjected to a fractionation procedure. From the pellet, the upper, loosely-packed slower sedimenting layer, which is characterized by a pale yellow-brownish color, was carefully removed with a spatula. The white main starch fraction was purified twice with 20 mL of water and centrifuged at 2300× g for 10 min at 4 °C. The starch sample was dried at 40 °C for 24 h.The liquid fraction was cooled to 3 °C for 24 h and centrifuged at 2300× https://imagej.nih.gov/ij/ open source program last accessed December 2019), and the diameter of starch granules was computed using the Excel program. The numbers of A-type and B-type starch granules were determined while assuming that starch granules had spherical shapes as a function of their diameter (<10 μm and >10 μm).Starch granules were visualized using an AXIO Image Z2 . Sample powder (10 µg) was sprinkled on a glass slide and covered with a single drop of an aqueous glycerol solution (1:1 water/glycerol) and covered with a coverslip. Micrographs were obtained at 400× magnification. Four slides were prepared for each sample, and three micrographs were selected from a total of ten for each sample, counting a total of 500 granules of starch. The granule area was measured using ImageJ software  scavenging capacity was performed according to Re et al. [•+ was produced by adding 2.45 mM of potassium persulfate to 7 mM of ABTS•+ solution and allowing the mixture to react in the dark at room temperature for 16 h. Then, the solution was diluted 1:80 with methanol to obtain an absorbance of 0.70 ± 0.02 OD at 735 nm. Fresh ABTS•+ solution was prepared for each assay. One hundred microliters of sample were added to 1 mL of ABTS•+ solution (A735nm 0.70 ± 0.02), and the A735nm was read after 5 minutes. Trolox was used as a standard (25–250 µM), and the determination of antioxidant activity was expressed as µM/g of Trolox equivalent (TE).The 2,2’-azino-bis-(3-ethylbenzothiazoline-6-sulfonate) radical cation . All data presented are the means of three replicates. For the evaluation of the storage protein amounts, densitometric analysis of bands, starch and antioxidant properties, first, we performed an analysis of variance (ANOVA). Then, all parameters were evaluated together by a multivariate analysis (MANOVA). Whenever a statistically significant effect was detected, the post-hoc Dunn’s test was applied to identify the differences among the pairs of samples. The amounts of the different gliadins, LMW-GS, and HMW-GS sub-units, which were calculated as reported in The four growing environments had very different meteorological conditions ,24. WithIt is well known that high temperatures during seed development represent a stressful condition which affects the duration and rate of grain filling ,29,30. IThe amount of rain was also recorded in all locations in the two growing seasons, and some differences were observed between the two years, mainly in the reproductive period . Plant dp < 0.05) were observed (>15%). Overall, Svevo had a significantly higher (p < 0.05) GPC than Iride (The gluten protein content (GPC %) was evaluated in both cvs in the two years in all locations. Each cv had comparable GPC values in the four locations , with than Iride . It is wThe accumulation of gluten protein is a complex process undergoing spatial and temporal regulation, which is affected by environmental conditions ,37. SyntIt was reported that the protein content may vary due to stressful environmental conditions during grain filling, affecting both kernel size and composition ,40. p < 0.05) in the four locations as compared to in 2016 (p < 0.05). Due to the differences in these two gluten fractions, the corresponding glu/gli and HMW-GS/LMW-GS ratios, which are important quality parameters [Therefore, the amount of each gluten protein fraction was compared in Iride and Svevo cultivated in 2016 and 2017 in the four locations. As expected, the gliadins fraction was the most abundant component, followed by the LMW-GS and the less abundant HMW-GS . The HMW in 2016 ; conversrameters , varied rameters ,43. In arameters ,45,46. NGlu-B1) is present with a band of 86 kDa (Bx7) and a band of 74 kDa with two isoforms (By8), while LMW-GS (locus Glu-B3) is represented by one main band of 42 kDa and minor bands  (LMW-2 profile) in accordance with previous studies [All gluten fractions were analyzed in both cvs by SDS-PAGE, and  studies ,48. This studies ,50. For p < 0.05) in relation to the year (Y) and to the interaction between genotype and year (G × Y) (p < 0.05) in 2017 than 2016 in both cvs, while By8 showed the opposite trend  : in all te trend A. Although HMW-GS only accounts for up to 10% of total flour protein, it affects the gluten viscoelastic properties. The variation in the amounts of Bx and By sub-units in the different years could also have had an impact on grain quality . In partp < 0.05) due to the combined effect of genotype and year, and for the 37 and 32 kDa subunits due to the year  in both cvs and at all cultivation sites. Proteins of 34 and 33 kDa, corresponding to α and α/β gliadins, were more abundant in 2017 than 2016 (p < 0.05). In particular, the 33 kDa sub-unit was undetectable in SDS-PAGE gel in 2016 in Svevo  and B (diameter <10 µm) granules A: A-granThese results are in agreement with several studies showing that the environmental conditions may have a strong effect on the median size of A and B granules and also on the total amount of starch ,56,57,58Temperature stress is critical for starch accumulation ; high teThe phenolic compositions of the investigated cvs are shown in p < 0.05), and samples from the year 2017 had a higher content than samples from the year 2016 (p < 0.05). The highest amounts were observed in Foggia and Quisquina: 339.30 and 483.11 µg GAE/g, respectively. It has been previously reported that in wheat bran, the amount of polyphenols is highly correlated with the antioxidant activity [+ scavenging capacity. Effectively, the highest ABTS+ scavenging capacity was observed in both cvs grown in Quisquina in all samples grown in 2017 (Bound phenolic acids, which are the most abundant class, showed variation in relation to year, location, and genotype ; in partactivity . Similar in 2017 . This re in 2017 . TherefoIn this study, we characterized two wheat cvs, which are among the most cultivated in Italy for their medium to high yield combined with the good quality of the flour . They pe


The results shown that there are three main mechanisms of dynamic softening of samples within the range of selected process parameters: dynamic recovery, dynamic recovery + dynamic recrystallization, and dynamic recrystallization, and the equiaxed dynamic recrystallization grain tends to be formed under higher temperature and higher strain rate. In order to accurately describe the dynamic recrystallization condition of Al-4.8Mg alloy under a wide range of hot deformation parameters, an improved dynamic recrystallization critical conditions model is proposed based on deformation activation energy and work-hardening rate. Additionally, a two–stage physically constitutive model considering the influence of work hardening–dynamic recovery and dynamic recrystallization is established. Comparisons between the predicted and experimental data indicate that the proposed model can adequately predict the flow stress in the range of selected process parameters with the average absolute relative error of 4.02%.The microstructure evolution and the mechanical behavior of Al–4.8Mg alloy were investigated by means of isothermal compression tests at various temperatures (280–520 °C) and strain rates (0.01–10 s Due to their characteristics of light weight, reasonable strength, good formability and high corrosion resistance, Al-Mg alloys are widely used in transportation, architectural decoration, food packaging and other industrial fields ,2,3. TheBecause of its high stacking fault energy, DRX of aluminum alloys is conventionally considered to be difficult to happen during hot deformation. However, with the in-depth research, DRX was proved to be an important softening mechanism during hot deformation of aluminum alloy. In recent years, the phenomena of DRX in different aluminum-based alloy systems were successively reported by many works, such as Al-Cu , Al-Li , Al-Zn-MGenerally, the thermal mechanical conditions for DRX of aluminum alloy are more severe than those for DRV. Therefore, there are should be a series of critical parameters from DRV to DRX. In the whole range of selected process parameters, if no DRX occurred in the material, the physical constitution based on DRV was used to describe the deformation behavior of the material. Such as, the deformation behavior of 5754 aluminum alloy reported by Huang . Moreove−1. An improved DRX critical conditions model of Al–4.8Mg alloy is proposed based on deformation activation energy and work-hardening rate. Further, a physically‒based model considering DRV and DRX mechanisms is established to describe the relationship between the flow stress and forming parameters.In order to accurately describe the DRX conditions of Al‒4.8Mg alloy under a wide range of hot deformation parameters, in this paper, the hot deformation behavior of Al‒4.8Mg alloy was investigated via a set of hot compression tests at temperatures of 280‒520 °C and strain rates of 0.01‒10 sThe experimental material employed in this paper is prepared by the industrial production line. It is cast into flat ingots after strict degassing and deslagging in the industrial process, and then the flat ingots were homogenized by 420 °C/2 h + 500 °C/12 h. The chemical composition (wt.%) of Al-Mg alloy  is 4.82Mg–0.49Mn–0.11Cr–0.29Fe–0.11Si– Al. The optical microstructure of the initial homogenized ingot exhibits uniform equiaxed grains, with an mean grain size of 180 um, as shown in –1 and strain of ~0.9. Prior to hot compression, all specimens were heated up to the target compression temperatures at a heating rate of 10 °C/s and held at the testing temperature for 3 min to obtain a uniform and stable temperature. After each compression test, all samples were quenched in cold water quickly to maintain the deformation microstructure.The homogenized ingot was machined to cylindrical specimens with 15 mm (height) × 10 mm (diameter) in size. The compression tests were carried out on a Gleeble–3500 thermal simulator  at temperatures from 280~520 °C, strain rates of 0.01~10 s4 + 90% C2H5OH solution and then observed using a Sirion 200 field-emission gun SEM  at 25 kV with a typical scan step size (1.5 µm). The TEM samples were first mechanically grinded and polished to less than 100 μm, then punched into discs with a diameter of 3 mm and finally twin–jet electropolished with an electrolyte solution of 30% nitric acid and 70% methanol at a voltage of ~15 V DC and a temperature below −25 °C.The compressed specimens were cut along the deformation axis for the microstructural observations. The microstructural characterizations were conducted on an AXIOScope A1 optical microscope , an electron back-scattered diffraction (EBSD) and a TecnaiG2 20 transmission electron microscopy . The OM samples were polished and anode–coated at a voltage of 20 V for 1 min using a solution of fluoroboric acid (10 mL) + water (400 mL). The EBSD samples were electrolytically polished at 20 V for 5 s in 10% HCHOWhen the softening rate and work hardening rate reach an equilibrium condition, the flow stress attains the maximum, i.e., the peak stress. In the steadily stage, the stress–strain curves show two change characteristics. If there only DRV occurs when the stress exceeds the peak strain, the flow stress tends to be stable after the dynamic equilibrium of WH and dynamic softening is achieved. If the dynamic softening rate caused by DRX is greater than the hardening rate, the flow stress will gradually decrease until the deformation reaches a certain strain point, the WH and dynamic softening reach a new equilibrium point, and the flow stress tends to be stable.−1, which represents the softening mechanism dominated by DRV. When the deformation temperature ranges from 400 °C to 460 °C and the strain rate ranges from 0 to 1 s–1, fine DRX grains and large numbers of sub-grains can be observed from the micrographs, as shown in –1 samples. This means the softening mechanism of this sample is dominated by DRX.−1, which dominated by DRV softening mechanism, represents large numbers of dislocations and almost no DRX grains, as shown in −1, there are many DRX grains in the sample and the recrystallization degree is obviously higher than that of the sample deformed at 400 °C /1 s−1. According to the observation results of EBSD and TEM, −1 is dominated by DRX. These results are consistent with the optical microstructures as shown in According to the microstructural characterizations, it can be inferred that the samples deformed at different conditions represent different softening mechanisms. Obviously, the variation of softening mechanisms with process parameters must be fully considered, when establishing the physical constitutive model. For instance, in this paper, in some conditions, the main softening mechanism of the experimental samples is DRV, while in some conditions the softening mechanism is dominated by DRV and DRX. Therefore, there must be a critical condition for DRX of the samples, which should be considered. However, the critical strain of DRX determined by the inflection point of the θ–σ curve proposed in  is only According to the microstructure characterizations, only when the temperature and strain rate meet a certain condition DRX in the experimental samples can occur. Whereas the temperature and strain rate are suitable for DRX, DRX occurs only after the strain achieves a critical value . Therefo−1), T is the temperature (K), Q denotes the activation energy (J mol−1), and R is a constant (8.314 J mol−1 K−1). In order to get lnZ under different deformation conditions, the Arrhenius constitutive equation was employed, as described by Equation (2) where :(3)f(σ)=1n, β, α, n are material constants and 1α = β/n. Substituting Equation (3) into Equation (2), then taking the natural logarithms of both sides, then Equations (4)–(6) are obtained:Therein, By transforming the Equations (4)–(6) and taking partial differentiation, the following expression are obtained:β can be calculated through the mean slopes by plotting β = 0.0817, and α = β/n and S can be obtained from n = 3.4519 and S = 6.0886. Substituting the above material coefficients into Equation (10), the average value of Q = 174.7 KJ/mol can be got, which is close to the result (168.61 KJ/mol) of Ref. −1, but nIt is generally believed that increasing temperature is beneficial to atom migration and dislocation movement, whereas increasing strain rate is beneficial to energy accumulation in the samples during hot deformation processing. The coupling effects of the deformation temperature and strain rate is complex. When describing the critical conditions of DRX, Z parameter reflects the sum of the contribution of strain rate and deformation temperature to DRX, however, it is a general relationship, and the weighting factors of deformation temperature and strain rate that contribute to DRX are not considered. Additionally, lnZ in Q is an important physical parameter for hot deformation processing. The higher the Q is, the more difficult of the sample is to be deformed, and vice versa. When DRX occurs, deformation is easy to perform. Therefore, Q may be more suitable than lnZ to determine the critical deformation temperature and critical strain rate: Generally, deformation activation energy Let:n is the slope of the curve of S is the slope of the curve of ln[sinh(ασ)] to 1/T at various strain rate as shown in n and T, S and Then, Equation (10) can be expressed as:n and S can be described as Equation (13) by linear fitting:Obviously, Q can be written as Equation (14):Substitute Equation (13) to Equation (12), Q under the whole deformation conditions, and the results are shown in Q of each deformation conditions, we can find that DRX occurred in the samples when the value of Q is below 183.9 KJ/mol, and when Q is higher than 183.9 KJ/mol, only DRV takes place. Therefore, Q can be used to describe critical temperature and strain rate of DRX.Equation (14) can be used to calculate the value of −1 in −1 and DRX takes place in As shown in According to the above comparative analysis, it can be judged that the temperature and strain rate coupled DRX critical conditions of Al-4.8Mg alloy can be described by Q as Equation (15):Equation (15) can be used to determine the critical temperature and strain rate of DRX occurrence. However, during the deformation process, even if the temperature and strain rate conform to Equation (15), DRX will occur in the samples only when the strain meets a critical value ,22. Genen of DRX ,22.As shown in −σ curve b, θ grad−σ curve d, then tBased on the above analysis, the minimum value of the By analyzing the data in The correlation coefficient between experimental value in Thus, the DRX critical conditions of sample under the whole deformation conditions in this paper can be established based on the deformation activation energy and work-hardening rate, as shown in Equation (17):In this paper, the dominating softening mechanism of the samples includes DRV, DRV+DRX and DRX. The critical conditions of DRX occurrence has been established in U is the WH, when the strain is defined in a certain value, it can be considered as a constant. The flow stress of metals is mainly affected by the internal dislocation density, while the variation of dislocation density in the matrix depends on the generation and elimination speed of dislocations. Therefore, the relationship among dislocation density, WH, DRV and true strain can be described as Equation (18) :(18)dρdεEquation  can be oEquation  can be or is a material constant, μ is the shear modulus, b is the Burgers vector. Plugging Equation (19) in to Equation (20), the flow stress of DRX samples can be obtained, as described by Equation (21):Therein, θ–σ curves of the DRV samples do not have inflection point, therefore, the peak stress of the DRV sample was approximated to their Saturation stress The DRV coefficient It was found by regression analysis that Thus, the constitutive model of the DRV stage of the experimental samples in the experimental range can be expressed as:t is the deformation time, D and nDk are the material constants, and generally, 0 < Dk < 1 < 0 < Dn ≤ 2. According to the published works [When the deformation strain of samples satisfies Equation (17), DRX occurs in the samples, and represents a higher softening rate. Generally, the volume fraction of DRX grains can be described as Equation (25) :(25)XD=1ns, Thus :(26)XD=1ed works , the volTherein, ion (28) :(28)σ=σrIn order to calculate the parameters of the model, the steady stress Combining Equations (26) and (27), Equation (29) can be obtained. Take logarithm on both sides of Equation (29) to obtain Equation (30). Thus, To verify the model, first, Equation (17) was applied to determine the softening mechanisms of the samples under the verifying deformation condition. Then, the parameters of the verifying deformation conditions in DRV state were substituted to Equation (24), and correspondingly, the parameters in DRX state were substituted to Equation (31). At last, the obtained predicted flow stress value was compared with the experimental flow stress. R) and the average absolute relative error (AARE) between the calculated flow stress and experimental flow stress were obtained though Equations (32) and (33) [N is the number of samples. By calculating, the correlation coefficient R is 98.9%, and AARE is 4.02%. This indicates the established constitutive model in this paper is of high accuracy.To further verify the model, the correlation between the experimental value and calculated value are shown in and (33) :(32)R=∑−1). The main conclusions can be summarized as follows:(1)There are three main dynamic softening mechanisms of samples within the range of experimental parameters: DRV, DRV + DRX and DRX, and the equiaxial DRX is more easily to form under higher deformation temperature and higher strain rate.(2)Based on the deformation activation energy and work-hardening rate, an improved DRX critical conditions model was proposed to describe the DRX conditions of Al-4.8Mg alloy under a wide range of hot deformation parameters.(3)According to the proposed DRX critical conditions model, a two–stage physically–based constitutive model including the WH-DRV stage and DRX stage was established. The 4.02% of AARE between the calculated values and experimental values indicates that the established physically–based constitutive model has a good prediction accuracy.In this study, the flow behavior of Al–4.8Mg alloy was investigated via thermo–mechanical tests in a range of deformation temperatures (280–520 °C) and strain rates (0.01–10 s


Age and high blood pressure are the two main determinants of arterial stiffness. In elderly hypertensives, large arteries stiffen and systolic and pulse pressures increase, due to wave reflections. A major reason for measuring arterial stiffness in clinical practice in elderly hypertensive patients comes from the repeated demonstration that arterial stiffness and wave reflections have a predictive value for CV events. A large body of evidence has been published during the last two decades, concerning the epidemiology, pathophysiology, and pharmacology of large arteries in hypertension in various settings of age. Particularly, two expert consensus documents have reviewed the methodological agreements for measuring arterial stiffness. The concepts of Early Vascular Aging (EVA) and Supernormal Vascular Aging (SUPERNOVA) help to better understand on which determinants of arterial stiffness it is possible to act, in order to limit target organ damage and cardiovascular complications. This review will address the issues of the cellular and molecular mechanisms of arterial stiffening in elderly hypertensives, the consequences of arterial stiffening on central systolic and pulse (systolic  Age and high blood pressure are the two main determinants of arterial stiffness. In elderly hypertensives, large arteries stiffen and systolic and pulse pressures increase, due to wave reflections. There are two major reasons for measuring arterial stiffness in clinical practice in elderly hypertensive patients. Firstly the repeated demonstration that arterial stiffness and reflected waves are predictive for CV events –4. An imVarious mechanisms are involved in aortic stiffening, in response to age and cardiovascular risk factors. They include breaks in elastin fibers, cross-links of the elastin network, and accumulation of collagen. In addition, fibrosis, inflammation, medial smooth muscle necrosis, calcifications, and diffusion of macromolecules within the arterial wall can play a role –11. Morein vitro  increases at the site of central and peripheral arteries. Because rhe arterial tree can be approximated to a viscoelastic tube with numerous branches and a high level of impedance (resistance in oscillatory conditions) of the tube end, retrograde waves, due to wave reflection, are generated. The higher the arterial stiffness, the higher the transmission velocity of both forward and reflected waves, thus the reflected wave arrives earlier in the central aorta and augments pressure in late systole, increasing central PP , reflection sites are closer to peripheral sites and reflection superimposes on forward wave, leading to amplification of pulse pressure. Because PWV is low, reflection comes late, in late-systole or early diastole in central arteries, limiting pulse pressure. By contrast, when the stiffness gradient disappears or is inverted , apparent reflection sites are much closer to central arteries, and pulsatile pressure is not sufficiently dampened at the central level, and the central-to-peripheral pressure amplification is attenuated .The physiological amplification phenomenon is attenuated by aging because of arterial stiffening. Indeed, by favoring early wave reflections, arterial stiffening increases peak- and end-systolic pressures in the ascending aorta. Thus, central systolic and pulse pressures are higher in elderly subjects than in young subjects and closer to the brachial SBP value, reducing the difference  Figures, 4A,B ExCross-sectional and longitudinal population studies have shown that both peripheral SBP and DBP trajectories increase progressively between adolescence and adulthood , 34. BefFor these reasons, isolated systolic hypertension (ISH), defined as SBP > 140 and DBP <90 mmHg, represents the most frequent subtype of hypertension after age 60 . Men andReference values for central SBP have been reported according to sex, age and BP categories . AccordiThe small/large artery cross-talk has a synergistic effect on target organs, in large part through a high pulsatile energy that is delivered as central systolic or pulse pressure. There is a large amount of evidence that central systolic and pulse pressures are the most deleterious elements of the blood pressure load on target organs. Their increase is better correlated with hypertensive target organ damage than either brachial systolic and pulse pressures or mean BP , 35, 36.A higher correlation has been observed with central SBP and PP than with brachial SBP and PP for left ventricular hypertrophy , systoliThe damage of large and small arteries can increase the risk of ischemic stroke, and, likely in parallel, white matter lesions, lacunar infarcts, and cognitive decline , 44. HigSeveral mechanisms can explain why an increased arterial stiffness can increase the risk of stroke: an elevated central PP, remodeling extracranial and intracranial arteries, in association with an increased carotid wall thickness and the development of stenosis and plaques  and the Myogenic tone in the renal circulation is impaired in the elderly hypertensive. The consequence is a loss of the autoregulation capacity and an increase in barotrauma due to high systolic BP, leading to glomerular injury. Because of its torrential nature (very low resistances), even small increases in peripheral blood pressure are transformed into high pulsatile energy that is transmitted to the kidney and dissipated in the microcirculation, leading to hyperfiltration and glomerulosclerosis. Clinical data are consistent with this pathophysiological approach. Significant relationships have been demonstrated between brachial pulse pressure and either glomerular filtration rate (GFR) or microalbuminuria ; betweenBecause the small/large artery cross-talk exerts a synergistic damaging effect on target organs, it is not surprising that arterial stiffness , 61–64, The Lancet Commission on Hypertension  , 67–69 trtension  used a sEVA can be diagnosed in patients who present an abnormally high arterial stiffness for their age and sex. Thus, EVA represents an altered capacity for repairing arterial damage in response to aggressors like mechanical stress and metabolic/chemical/oxidative stresses. In other words, arterial stiffness is an integrator of all damages done to the arterial wall. Moreover, aortic stiffness as a marker of arterial wall damage or arteriosclerosis, integrates both the effect of risk factors and susceptibility to those risk factors and duration of exposure. Thus, arterial stiffness measures not only the current arterial damage  but also its regression (when a therapeutic action is taken) or progression (when exposure continues or therapeutic actions fail). It differs from the usual “snapshot” that physicians get from their patients when they only measure BP, cholesterol and glycemia. This is why EVA-arterial stiffness has a higher predictive value for CV events than classical CV risk scores .Recent cross-sectional and longitudinal studies have extended the list of the epidemiological determinants of arterial stiffness. Most of these determinants belong to the classical risk factors, either non-modifiable such as ethnicity, sex, chronological age, family history and personal history, or modifiable such as blood pressure, diabetes mellitus, dyslipidemia, and smoking . AdditioMore recently, we extended the concept of extremes in vascular aging . Indeed,Arterial stiffness can be evaluated at the systemic, regional and local levels. Systemic arterial stiffness can only be estimated from models of the circulation. By contrast, regional and local arterial stiffness can be measured directly, and non-invasively, at various sites along the arterial tree. A major advantage of their evaluation is that they are based on direct measurements of parameters strongly linked to wall stiffness. A large number of reviews have been published on methodological aspects , 71, 72.Regional stiffness is mainly determined through pulse wave velocity between two arterial sites. The measurement of pulse wave velocity (PWV) is generally accepted as the most simple, non-invasive, robust, and reproducible method with which to determine regional arterial stiffness , 5. CaroAlternative methods have been proposed by measuring transit time between the arm and the leg , the heart and the leg (heart-ankle PWV), or between the finger and the toe (finger-toe PWV). Particularly, the measurement of brachial-ankle PWV - baPWV  has been developed as an automatic cuff-based method in order to increase easiness and acceptability . BrachiaSeveral reviews have made recommendations for adequate measurements of central BP , 78–82. Aortic pressure waveform can also be deduced from the common carotid artery waveform. Carotid tonometry requires a higher level of technical expertise, but a transfer function is not necessary since the arterial sites are very close and waveforms are similar . In addiThe measurement of arterial stiffness and wave reflections in clinical practice in hypertensive patients is comforted by the repeated demonstration that arterial stiffness has an independent predictive value for CV events , 5.Several longitudinal epidemiological studies have demonstrated the predictive value of arterial stiffness for CV events. The largest amount of evidence has been given for aortic stiffness, measured through carotid-femoral PWV. Aortic stiffness has independent predictive value for all-cause and CV mortality, fatal and non-fatal coronary events, and fatal strokes not only in patients with uncomplicated essential hypertension , 61, 62,Two meta-analyses –4 consisAortic stiffness has demonstrated an independent predictive value for CV events after adjustment to classical CV risk factors, including brachial pulse pressure. These data show that aortic stiffness predicts CV events to a larger extent than each of classical risk factors. In addition, aortic stiffness retains its predictive value for CHD events after adjustment to the Framingham risk score, demonstrating an added value to a combination of CV risk factors . Aortic An important issue is whether arterial stiffness retains its independent predictive value in elderly hypertensives. Ben Shlomo et al.  undertooSeveral reviews , 92 and Pharmacological treatments which are able to reduce arterial stiffness in humans include (a) antihypertensive treatment , such asAn open issue is whether, in the elderly, life-style changes and pharmacological treatments are effective in reducing arterial stiffness to the lower levels of younger subjects, and whether the reduction in arterial stiffness translates into a reduction in cardiovascular events. To our knowledge, no large randomized clinical trial has been performed in a specific group of elderly hypertensives, either above 60 or 70 years of age.Subjects older than 80 years represent a particular population since they are generally frail, with several comorbidities. Very few studies have been done in very elderly hypertensives. The HYVET study , one of Very few studies addressed the particular case of very elderly individuals. The PARTAGE study is an observational one , 105, thOne reason for which isolated systolic hypertension represents the most frequent subtype of hypertension in the elderly is because arterial stiffness increases with aging, in some more than others, a phenomenon leading to increased wave reflection and augmented central pulse pressure. High central systolic and pulse pressures damage target organs that ultimately leads to CV and renal complications. In the elderly, arterial stiffness has predictive value for CV and renal events. Even in the very old frail hypertensive, arterial stiffness retains its predictive value for cognitive decline and morbidity-mortality. The measurement of arterial stiffness may help the physician to better determine the risk of CV complications in elderly hypertensive and adapt the therapeutic strategy.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.