The Arctic is experiencing rapid changes in sea-ice seasonality and extent, with significant consequences for primary production. With the importance of accurate monitoring of spring phytoplankton dynamics in a changing Arctic, this study further examines the previously established critical relationship between spring phytoplankton bloom types and timing of the sea-ice retreat for broader temporal and spatial coverages, with a particular focus on the Pacific Arctic for 2003–2019. To this end, time-series of satellite-retrieved phytoplankton biomass were modeled using a parametric Gaussian function, as an effective approach to capture the development and decay of phytoplankton blooms. Our sensitivity analysis demonstrated accurate estimates of timing and presence/absence of peaks in phytoplankton biomass even with some missing values, suggesting the parametric Gaussian function is a powerful tool for capturing the development and decay of phytoplankton blooms. Based on the timing and presence/absence of a peak in phytoplankton biomass and following the classification developed by the previous exploratory work, spring bloom types are classified into three groups . Our results showed that the proportion of under-ice blooms was higher in the Chukchi Sea than in the Bering Sea. The probable under-ice blooms registered as the dominant bloom types in a wide area of the Pacific Arctic, whereas the marginal ice zone bloom was a relatively minor bloom type across the Pacific Arctic. Associated with a shift of sea-ice retreat timing toward earlier dates, we confirmed previous findings from the Chukchi Sea of recent shifts in phytoplankton bloom types from under-ice blooms to marginal ice zone blooms and demonstrated that this pattern holds for the broader Pacific Arctic sector for the time period 2003–2019. Overall, the present study provided additional evidence of the changing sea-ice retreat timing that can drive variations in phytoplankton bloom dynamics, which contributes to addressing the detection and consistent monitoring of the biophysical responses to the changing environments in the Pacific Arctic. Phytoplankton blooms in the Arctic have typically been defined as enhanced growth of phytoplankton populations that occur along the seasonally retreating ice edge, with water column stratification from sea-ice melt providing sufficient light for photosynthesis . Such maArctic annual surface air temperatures in 2014–2018 were higher than in any prior years since observational records began in 1900 . The mosa (chla) concentration, which is a proxy of phytoplankton biomass at the sea surface, Lowry et al. [a concentrations at the same time of the sea-ice retreat, indicating that there had occurred phytoplankton growth under the ice, whereas PUI blooms were defined as low chla concentrations for the first several weeks of the ice-free period because UI blooms assumed to be matured and depleted nutrients before the ice retreat. MIZ blooms were characterized as low chla concentrations at the time of sea-ice retreat and those chla concentrations increased with time to have blooms days to weeks after the ice retreat. Using a simple bloom classification approach based on the timing when the satellite-derived chla reached a bloom threshold of 2.5 mg m−3 following the retreating sea-ice edge, Lowry et al. [Based on satellite-retrieved time-series of chlorophyll-y et al.  exploredy et al.  reportedThe Pacific Arctic, specifically the region extending from the northern Bering Sea to the southern Chukchi Sea , is one a required to distinguish accurately between spring phytoplankton bloom types [a threshold [With the importance of accurate monitoring spring phytoplankton dynamics in a changing Arctic, this study examines the previously established critical relationship between the spring phytoplankton bloom types and timing of the sea-ice retreat for broader temporal and spatial coverages, with a particular focus on the Pacific Arctic for 2003–2019. To this end, time-series of satellite-retrieved phytoplankton biomass during the period of seasonal ice edge retreat were fit to an expected growth curve to identify the phase of the bloom using a parametric Gaussian function, as an effective approach to capture the development and decay of phytoplankton blooms –23. The om types . Moreovehreshold , our appThe current study is a contribution to a special issue on the Distributed Biological Observatory (DBO), specifically to highlight the potential for satellite remote sensing in supporting point and transect-based measurements in a complex environment. The DBO is a change detection array for select ecosystem variables along eight sampling transects in the Pacific Arctic . The oveRrs(λ)) at 9-km resolution at four wavelengths , obtained by the Moderate Resolution Imaging Spectroradiometer (MODIS) sensor onboard the Aqua satellite, were downloaded from NASA’s Ocean Color website (https://oceandata.sci.gsfc.nasa.gov), for the period of 2003–2019. In addition, the Advanced Microwave Scanning Radiometer (AMSR) Unified daily sea-ice concentrations gridded at a 12.5-km resolution for the period of 2003–2019 were downloaded from the National Snow and Ice Data Center website (https://nsidc.org/). AMSR Unified sea-ice concentrations were created by reprocessing the Advanced Microwave Scanning Radiometer-Earth Observing System (AMSR-E) sensor onboard the Aqua satellite during 2003–2011 and Advanced Microwave Scanning Radiometer 2 (AMSR-2) sensor onboard the Global Change Observation Mission 1st-Water (GCOM-W1) during 2013–2019. To align the spatial resolution of satellite data used in this study, daily sea-ice concentrations were resampled onto the 9-km grid using nearest-neighbor interpolation [Level-3 daily standard mapped images of remote-sensing reflectance (a (chla) concentration was computed from Rrs(λ) using the Arctic-OC3L algorithm developed and validated in the Chukchi Sea [a and b are empirically derived values of 0.3364 and −3.4388, respectively, and R is the base-10 logarithm of the maximum band ratio of Rrs(λ) among the three potential blue wavelengths  and the green wavelength .Chlorophyll-kchi Sea . The Arca over a 20-day of MIZ retreat period following the date of sea-ice retreat (DSR) were fit to an expected growth curve to identify the phase of the bloom, i.e., UI, PUI, or MIZ bloom. It is noteworthy that the 20-day of MIZ retreat period was widely used in previous studies [a was captured at a given location, phytoplankton blooms in spring were classified only as such if the pixel contained a minimum of one chla value per week and at least one chla concentrations within two days after onset of ice retreat [a time-series during the MIZ period was modeled as a Gaussian function using MATLAB lsqnonlin  on a pixel-per-pixel basis as follows:t represents the date, chla0 is the background chla, chlam is the amplitude of the chla peak observed on the date tm, and σ corresponds to the number of days chla(t) exceeded half of chlam , the chla time-series was classified into either an UI bloom or MIZ bloom. The distinction between UI and MIZ blooms is based on the model parameter tm: a chla time series was classified as an UI blooms if tm was observed before DSR (tm < DSR); in contrast if tm was equal to or after DSR (tm ≥ DSR) the bloom was classified as a MIZ bloom. In the case that the models were statistically insignificant (p ≥ 0.05), chla time-series is expressed as a constant at chla0 as follows:Then, the chlom types  based ona time-series as a single term of chla0 were classified as PUI blooms. This is based on the assumption that the pixels had already experienced a bloom before DSR, and thus the chla retrievals during the MIZ retreat period were remnants of the UI bloom [Modeled chlUI bloom .The classification was based on the assumption that pre-bloom surface nutrient concentrations are high throughout the Pacific Arctic, such that a phytoplankton bloom would be expected in waters with sufficient light availability. This assumption is reasonable because the vertical turbulent mixing of nutrient-rich Anadyr Water and Bering Shelf Water provide sufficient nutrients throughout the water column for phytoplankton growth in spring  and sucha. To ensure the robustness of our approach, we conducted a sensitivity analysis using the minimum required number  of chla concentrations during the MIZ retreat period. First, a sample chla time-series during the MIZ retreat period was generated by averaging daily chla time-series during the MIZ retreat period for all pixels classified as UI blooms over the entire study period from 2003 through 2019. Then, five chla concentrations were randomly selected from the sample chla time-series as input data, and were modeled using the a time-series during the MIZ retreat period for PUI and MIZ blooms were also generated to examine the consistency of the resulting bloom type.In the Arctic Ocean, the extensive cloud cover limits ocean-color data availability, and in turn may hamper parametric modeling of time-series variations in satellite-derived chlaMIZ denotes averaged chla during the MIZ retreat period for 2003–2019 on a pixel-by-pixel basis .Initial value, and lower and upper bounds of model parameters apply to Eqs  and 3)..3). Thesp < 0.05) for 43% of the grid cells in the study area  for UI and MIZ blooms varied substantially, suggesting this parametric modeling approach induces substantial uncertainties in estimating these bloom characteristics. In contrast, temporal variations in chla for PUI blooms were accurately fitted by the parametric approach, because this bloom type can be expressed by a single model parameter .The results of our sensitivity analysis are shown in p = 0.14; PUI bloom, p = 0.05; MIZ bloom, p = 0.75). Proportions of each bloom type in individual years are summarized in p < 0.05) decreasing and increasing trends in the proportions of UI blooms and MIZ blooms for 2003–2019 (p < 0.05) positive and negative correlations with the proportions of UI and MIZ blooms, respectively. These findings demonstrate the influence of interannual variations in DSR on bloom typology in the Pacific Arctic. Note that the mean proportion of pixels satisfying the parametric model criteria for 2003–2019 was 7.7 ± 4.1% of ocean surface in the Pacific Arctic  in the proportions of UI and MIZ blooms accompanied by the changing timing of sea-ice retreat for the period 2003–2019  and at high concentrations (>0.5 mg m−3) within a distance of ~10–20 km from the ice edge, respectively, whereas the sub-pixel contamination results in overestimation at moderate to high concentrations (>0.05 mg m−3). Therefore, the adjacency effect could lead to an underestimation of UI blooms, whereas the sub-pixel contamination could lead to an overestimation of UI blooms. By comparing the resulting proportions of three bloom types using a more conservative ice retreat threshold, Lowry et al. [a retrievals are less significant in determining spring bloom types.Sea-ice related error impacts on satellite-retrieved chly et al. . The seay et al. : the adjy et al.  demonstry et al. . AlthougThe magnitude of the zooplankton grazing on phytoplankton growth is primarily controlled by grazer biomass which was very low in spring, when zooplankton was low , 53. Inda concentration may be subject to errors due to the optical complexity of the Arctic waters [a concentrations. Therefore, calibrating/validating the performance of our approach with in situ time-series data, such as at a mooring location, would further help assess the broader validity of these satellite-based findings. It is important to reinforce that this study provided an important insight into satellite-based monitoring of phytoplankton communities in the Pacific Arctic, which can contribute to developing and managing plans and strategies for in situ observation and/or related field activities in the future.Our sensitivity analysis demonstrated a robust performance of the parametric approach in identifying spring phytoplankton bloom types in the Pacific Arctic . Howeverc waters . Moreovec waters , hamperiAlthough monitoring phytoplankton communities using satellite always contain uncertainties, the present study demonstrated robust performance of a parametric, Gaussian fitting function for the satellite time series of phytoplankton biomass in identifying spring phytoplankton bloom types . Through


We describe a recipient with 7DS, successfully managed with emergentABO-incompatible (ABOi) retransplant. The patient provided informed consent topublish case.Seventh-day syndrome (7DS) is a rare but fatal acute complication of livertransplantation. It is characterized by an abrupt rise in liver enzymes and massivehepatocyte necrosis at around a week post-transplant, leading to fulminant hepaticfailure in a previously well-functioning graft.A 47-year-old man with well-compensated cirrhosis (Na-MELD 18) and hepatoma secondaryto autoimmune hepatitis underwent uncomplicated liver transplant with a brain-deaddonor liver. Immunosuppressive induction was methylprednisone. Tacrolimus wasintroduced on postoperative day (POD) 1.He had a routine recovery until POD 6 (with tacrolimus level 9.4 ug/L), when theliver enzymes abruptly increased (AST > ALT) . DopplerAn ABOi offer  was accepted due torapid deterioration and lack of blood group matched offers. His clinical conditioncontinued to worsen, requiring inotropic support, initiation of hemodialysis andoperative devascularization of the graft with temporary portocaval shunt. On POD 8,retransplant was performed with simultaneous splenectomy.Prior to ABOi transplant, 2 sessions of plasmapheresis were completed. Inductiontherapy was thymoglobulin and methylprednisone. He received 5 sessions ofplasmapheresis and IV Immunoglobulin G on alternate days. Surveillance biopsies onPODs 10 and 21 were unremarkable. At 10 months posttransplant, he was doing wellwith no evidence of rejection and normal graft function. There has been no demonstrated association with age, wait time, etiology,MELD score, type of allograft or surgical factors. Massive coagulative necrosis without major inflammation is the most prominenthistological feature. The postulated pathogenesis involves an undefined immuneresponse and activation of apoptosis pathways, as supported by mice models demonstrating increased hepatocyte expression ofFas receptor and induction of hepatocyte apoptosis by systemic injection ofmonoclonal antibody to Fas receptor. The mortality rate is above 80%, and retransplantation is the only curative option with 80% 1-year survival.Seventh-day Syndrome is an uncommon transplant complication, with incidence between0.5% to 1.68% and male preponderance.In North America, ABOi transplant has largely been reserved for patients withfulminant hepatic failure in the absence of a blood group-compatible organ.Intensive desensitization using IVIG, plasmapheresis, and rituximab vs splenectomyhas improved graft and patient survival after ABOi transplantation.Our case describes the rare entity of 7DS, characterized by sudden graft necrosis andfailure. This represents a posttransplant emergency, highlighting the need tourgently act upon development of markedly abnormal liver biochemistry and evidenceof liver failure, despite initial normal graft function and in the absence ofvascular thrombosis. Fortunately, certain mortality was avoided with the successfulapplication of emergent ABOi transplantation.


Capsaicin was microencapsulated in six different wall systems by spray drying whey protein and citrate mung bean starch at various ratios  to improve its stability and water solubility and reduce its pungency. The morphological, rheological, storage stability, and physicochemical properties of capsaicin emulsion and capsaicin microcapsules were characterized. As a result, the yield of six capsaicin microcapsules was 19.63–74.99%, the encapsulation efficiency was 26.59–94.18%, the solubility was 65.97–96.32%, the moisture content was lower than 3.63% in all systems, and particle size was broadly distributed in the range of 1–60 μm. Furthermore, microcapsules with high whey protein content in the encapsulation system had an excellent emulsifier effect and wetness, smooth particle surface, and higher lightness (L*). Moreover, the system formed by composite wall materials at a ratio of whey protein to citrate mung bean starch of 7:3 had the highest retention rate and the best stability. The overall results demonstrate that whey protein combined with citrate mung starch through spray drying could be a promising strategy to produce microcapsules of poorly water-soluble compounds such as capsaicin. Capsicum annuum), known for their hot and spicy flavor and belonging to the family Solanaceae, are among the most commonly consumed spices [2 in 2019. As a result, pepper accounts for China’s largest vegetable planting area [40H56O3, with a molecular weight of 584.87 g/mol [Chili peppers  and provide good protection against oxidation. However, native starch in microcapsules lacks emulsification properties and weak flavor retention ability ,13. TherWhey protein plays a vital role in anti-oxidation; it also has good emulsification and film-forming properties . In addiBecause of the limitation of modified starch as a single wall material, it can be combined with protein. For example, Tontul and Topuz reported that the combination of modified starch and whey protein endowed flaxseed oil microcapsules with a high entrapment rate . In addiIn the present work, whey protein compounded with citrate mung bean starch was used as complex wall materials to construct capsaicin microcapsules through spray drying. The impacts of the two wall materials on the microcapsule system’s morphology, physical properties, and stability were assessed, and the better wall material was chosen. Finally, principal component analysis (PCA) was used to compare component differences in the microcapsules. This work aimed to produce capsaicin microcapsules with reduced pungency and improved functional performance to expand the application of capsaicin.Mung bean , whey protein , medium-chain triglyceride , capsaicin , Nile red , DMSO, citric acid, absolute ethanol, phenolphthalein, sodium hydroxide, hydrochloric acid, and so on were chemically pure.Mung bean seeds were washed, put in warm water at 30–40 °C, and soaked at room temperature for 6–10 h. The soaked seeds were ground and sieved (100-mesh), and the material under the sieve was collected and stirred thoroughly. Then, the starch slurry was allowed to settle for 4–5 h, after which the upper layer of water was removed, and water was added to the settled starch. After thoroughly stirring, the solution was passed through a 200-mesh sieve to collect the material under the sieve. The under-sieve material was repeatedly washed and settled with water until the top slurry layer was transparent. The supernatant was removed, and the sunken starch was collected. The obtained starch was dried at 40 °C, ground with a high-speed universal pulverizer, and sieved (100-mesh) to obtain a sample of mung bean starch .w/w). Next, it was transferred into a stainless-steel tray and dried at 130 °C for 5 h, and the dry mixture was repeatedly washed with deionized water. Finally, the washed starch was dried, ground, and sieved (100-mesh) again, then stored in a vacuum bag. The degree of substitution (DS) of mung bean starch modified by citric acid was 0.167 ± 0.004.Citrate starch was obtained based on Lee et al.  with litProtein was dissolved in deionized water and agitated for 12 h to allow it to completely swell. Modified starch was dissolved in deionized water, heated for half an hour at 70 °C with continuous stirring, and then stirred for 12 h at 25 °C. The modified starch was mixed with protein and stirred for 40 min to obtain an aqueous phase with 396 g. The ratio of protein mass to esterified starch was 10:0, 9:1, 7:3, 5:5, 3:7, 1:9, and 0:10. The protein was added with modified starch as a composite emulsifier, and its mass concentration was 5%; the mass concentration of the oil phase was 1%.According to the preliminary experiment, capsaicin  was added to MCT (4.0 g) and heated at a high temperature (180 °C) until it dissolved to obtain the oil phase. After cooling, the oil phase was added slowly to the aqueous phase and stirred for 10 min. Finally, the above oil–water mixture was homogenized  and ultrasonically treated  for 5 min to obtain the capsaicin emulsion.Capsanthin microcapsule powder was obtained after spray drying  . The ratThe morphology of the capsaicin emulsion was observed with an optical microscope  at a magnification of 400×.First, the fluorescent dye solution was obtained by dispersing 1 mg of Nile red (excitation wavelength at 488 nm) in 1 mL of DMSO. Next, 20 μL of dye solution was mixed with 0.5 mL of freshly prepared emulsion, and then 9 μL was placed on a glass slide, covered with a cover glass, and allowed to stand for 2 h. After natural drying, the distribution of oil phase was observed under the 20× objective lens of the fluorescence microscope .−1.The rheological properties of capsaicin emulsion were measured by adding 1.25 mL of capsaicin emulsion to a DHR-1 rotary rheometer . The test conditions were as follows: the selection mode was flow scanning, the clamp was a 40 mm parallel aluminum plate, the temperature was 25 °C and 50 °C, and the shear rate was 0.01–100 so + kγn) was applied to fit shear stress/shear rate curves, and the rheological parameters of the samples were obtained, where τ represents the shear stress, Pa; τo represents the yield stress, Pa; K represents the consistency coefficient, Pa·sn; γ represents the shear rate, s−1; n represents the fluid index; R2 represents the degree of fit [The Herschel–Bulkley model (τ = τe of fit .The morphology of the microcapsule particles was observed using a Nano SEM-450 scanning electron microscope (SEM) . The dried sample was fixed on a carrier with conductive adhesive and sprayed with gold, observed, and photographed.−1 at a resolution of 4 cm−1. Samples were ground and laminated with KBr at a ratio of 1:100. The test was carried out with a KBr sheet as the background.FTIR spectra of the capsaicin microcapsules were scanned on a Vetex 70 Fourier transform infrared spectrometer  in the range of 4000 to 400 cmY) is [The yield of capsaicin microcapsules was calculated according to the following formula. The formula of yield (Y) is :(1)Y (%)aM: total mass of capsaicin in microcapsules, g; bM: total sample mass (capsaicin) added before spray drying, g.In the formula, 2 = 0.9999).An accurately weighed 50 mg sample of capsaicin was dissolved in absolute ethanol and diluted to a final volume of 500 mL in a brown volumetric flask to obtain a standard capsaicin solution with a 10 mg/100 mL concentration. Gradient concentrations of 0.0, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, and 9.0 mg/100 mL were respectively prepared by using anhydrous ethanol to dilute standard solutions of 0.0, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, and 9.0 mL to 10 mL. The absorbance value was measured at a wavelength of 460 nm. The standard curve was drawn with the absorbance value as the ordinate and the concentration of capsaicin as the abscissa. The standard curve of capsaicin was plotted as follows: A = 0.0811C + 0.004 (Rv/v) as the control to calculate the total content of capsaicin.The capsaicin content was determined by ultraviolet spectrophotometry. Determination of total capsaicin content in microcapsules: 100 mg of capsaicin microcapsule was dissolved in 5 mL of deionized water, placed in an ultrasound bath, and then put into a 100 mL brown volumetric flask with absolute ethanol at constant volume. The mixture was centrifuged  for 10 min ,26. The Surface capsaicin was quantified by dissolving 200 mg of capsaicin microcapsules with 20 mL of absolute ethanol in a 50 mL centrifuge tube. After centrifugation  for 10 min, 5 mL of supernatant was taken and diluted to 25 mL in a brown volumetric flask with absolute ethanol. The absorbance values were determined at a wavelength of 460 nm, with ethanol solution as the control to calculate the total content on the surface.The method of Kuang et al.  with a sThe moisture content in microcapsules was determined by the vacuum-oven method.S) was calculated using the following Equations:M is sample mass (g); 1M is aluminum box mass (g); 2M is the total mass of the insoluble substance and aluminum box (g).A certain mass of capsaicin microcapsules (dry basis) was dissolved in deionized water and centrifuged  for 10 min, and the supernatant was poured out. The residue was transferred into an aluminum box and dried to constant weight. Solubility  calibrated with a black and white standard plate. A 10 mm reflection ring was selected as the measuring hole. The value of L*  ranges from 0 (black) to 100 (white). The parameter a* represents the level of red (+a*) or green (−a*) color, and the value of b* measures the level of yellow (+b) or blue (−b*) color. All measurements were conducted three times for each sample. The 10W-0OP sample was taken as the standard sample to calculate the color difference (ΔE) value. The calculation formula is as follows:0*, a0*, and b0* represent the color of microcapsule samples prepared with protein as a single wall material.In the formula, L*, a*, and b* represent the color of the capsaicin microcapsule sample; LThe particle size of capsaicin microcapsules was measured through laser particle size analyzer , and the microcapsule samples were dissolved in deionized water. The universal liquid module (ULM) was selected as a sample processing module.The thermal properties of the capsaicin microcapsules were analyzed by a differential scanning calorimeter  under a nitrogen atmosphere at a flow rate of 20 mL/min. A 3 mg sample was hermetically sealed in an aluminum pan and heated from 30 °C to 250 °C at a heating rate of 10 °C/min to obtain the DSC curve, and an empty hermetic pan was used as a control. The DSC curve was processed and analyzed with TA software.R) of capsaicin in the microcapsules. The following formula was used to calculate the retention rate:The three kinds of capsaicin microcapsules with the highest EEs were stored in a dark and sealed environment at 25 °C or 50 °C for 15 day, and a 100 mg sample was taken every 3 day to determine the total capsaicin content and calculate the retention rate , SPSS 20.0 was used for principal component analysis (PCA), and Origin 8.0 was used for graphing.All experiments were carried out in triplicate, and the final results are presented as the mean ± standard deviation. The data were processed with Excel 2013, Minitab 18.1 was used for statistical analysis , the tiny oil droplets aggregated together. Small oil droplets adhered to each other or grew on the surface of large oil droplets to be fused, which indicates that emulsions emulsified with more citrate mung bean starch were poor. Kierulf et al. [f et al.  also men2–G2. This figure shows that oil droplets in the emulsion system with more protein content were smaller and evenly distributed, which shows that the emulsifier has a better emulsifying effect. This further demonstrates that protein is an excellent emulsifier [The fluorescence microscope (FM) observes the distribution of the oil phase in the emulsion by staining the oil phase specifically. Capsaicin was dissolved in MCT and stained with Nile red (labeled green). The FM photographs of whey protein–citrate mung bean starch–capsaicin emulsion are shown in ulsifier . When th−1 at 50 °C and 25 °C is displayed in The shear rheological behavior of the capsaicin emulsion at a shear rate of 0–100 so + Kγn) was used to fit the obtained shear rheologic curve. The fitting results are recorded in 2) ranged from 0.9960 to 0.9997, which highlights that the model is very suitable for fitting the rheological curves of this study [n < 1.The Herschel–Bulkley equation , indicating that the system with more modified starch content could form a strong and compact three-dimensional network structure, which improved the thickening effect. At the same time, it also showed the interaction between starch and protein, which improved their compatibility [When the ratio of whey protein to citrate mung bean starch was 1:9 and 0:10, the consistency coefficient (K) was significantly higher than those of other emulsion systems (tibility . Furthertibility  found th3–F3). This may be caused by the rapid dehydration of the emulsion during spray drying, which results in the shrinkage of the wall materials of the microcapsules, leading to dents around the powder.The appearance and morphology observed with SEM of the six microcapsule systems composed of whey protein and citrate mung bean starch are shown in When the ratio of whey protein to citrate mung bean starch was 10:0, 9:1, and 7:3, the surface of the microcapsules was smooth. They also exhibited a high degree of integrity with no noticeable wrinkles or cavities on the surface. In addition, the EE of these microcapsules was favorable, indicating that the capsaicin was well encapsulated. However, with the increased content of modified starch in the wall materials, some holes appeared on the surface of some microcapsules. Generally, holes can enhance the permeability of microcapsules and weaken the microencapsulation degree, thus leading to the oxidation and degradation of microcapsules, which become easily affected by the environment . Further−1 of natural mung bean starch represent the characteristic vibrational stretches of –OH groups [−1; a strong absorption peak observed at 1650 cm−1 is ascribed to water combined with starch.The Fourier transform infrared spectrometer (FTIR) detects whether capsaicin microcapsules introduce new functional groups or form new chemical bonds, and the molecular structure can be characterized. The obtained infrared spectrum is shown in H groups . The str−1 and 1572 cm−1 correspond to functional group vibrations of –C=O and RCOO–, respectively. This proves that the starch introduced new groups after esterification and verifies the esterification of native starch with citric acid in an alkaline environment. The sharp and tiny peaks at 2960 cm−1, 2930 cm−1, and 2850 cm−1 of whey protein represent –CH stretching and bending vibration bands; there were two prominent characteristic peaks at 1530 cm−1 and 1650 cm−1. In addition, the spectrum contained peaks at 1153 cm−1, 1750 cm−1, 2850 cm−1, and 2930 cm−1, which are characteristic absorption peaks of pure capsaicin.For citrate mung bean starch, the absorption peaks observed at 1730 cm−1 than pure capsaicin, with greater intensity and narrower peaks caused by C=O stretching vibration. This is connected to the encapsulation of capsaicin; it also indicates that the active components of capsaicin formed strong bonds with the compound materials and could form a stable microcapsule system. The mechanism of capsaicin encapsulation by starch and protein is shown in In addition to the characteristic peaks of the wall materials in the microcapsules, there were also more substantial absorption peaks near 1750 cm−1, 2930 cm−1, and 2960 cm−1 were sharper and stronger than the characteristic peaks of whey protein. In addition, a distinct peak was formed at 1153 cm−1, which was related to capsaicin microcapsules.Moreover, the absorption peaks of microcapsule powder products at approximately 2850 cmp < 0.05). When the ratio of whey protein to citrate mung bean starch was 1:9, the yield was the lowest, and the water content reached the peak . The shallow moisture content makes the microcapsules less susceptible to mildew, absorption of moisture, and aggregation; decreases the flow and dispersity of active constituents; reduces the possibility of microbial growth and lipid oxidation; and improves the storage stability of capsaicin. Higher moisture content may be ascribed to the rapid formation of the shell structure of the modified starch during the faster drying process. As a result, the internal water did not diffuse to the surface in time, making it hard for the moisture to evaporate.In addition, it can be seen in p < 0.05). The hydrophobic properties of citrate mung bean starch resulted in decreased solubility of the capsaicin microcapsule powders with the addition of citrate starch [Solubility is a crucial element in measuring the quality of powder ingredients. Products with poor solubility are not easily processed and will cause economic losses. As shown in e starch . NaturalThe wetness of microcapsules refers to the capacity of powder to be wetted by water to reflect its rehydration properties, usually recorded as the time that it takes for the powder to completely dissolve in water . It was p < 0.05), which were 93.38, 93.49, and 92.59, respectively. Meanwhile, we observed that the particle sizes of the two samples with ratios of whey protein to citrate mung bean starch of 10:0 and 9:1 were also the smallest. Other researchers have found a similar phenomenon in studies of pigment powders and red pepper powders; the L* values increased as the particle size decreased [p < 0.05) in the L* values was observed for the other samples as the whey protein content in the wall materials gradually decreased. The samples with higher whiteness indicate that the less the amount of capsaicin exposed on the surface of microcapsules, the better the EE, which is accordant with the above EE results  and ΔE of capsaicin microcapsule powder prepared with the combination of whey protein and citrate green bean starch are shown in ecreased . This maecreased . There w results .p < 0.05). When the ratio of whey protein to citrate mung bean starch was 1:9, the a* value reached a maximum of 14.87, indicating that the color tended to be red; under such conditions, the b* value also reached the maximum of 34.86, illustrating that the color of the product tended to be yellow. In terms of ΔE, there was no apparent difference between samples with ratios of whey protein to modified starch of 10:0 and 9:1; when the ratio of whey protein to citrate mung bean starch was 7:3, 5:5, 3:7, and 1:9, there was a significant difference from the first two types of microcapsules (p < 0.05). The difference increased with decreasing whey protein content in the wall materials, showing that the color of the microcapsules tended to be dark orange.The values of a* and b* increased with the proportion of modified starch in the encapsulation system (except for ratios of whey protein to citrate mung bean starch of 10:0 and 9:1). There was a significant difference between the samples  and  of microcapsules with whey protein as the only wall material were 1.12 μm and 0.83 μm, respectively. When the ratio of whey protein to citrate mung bean starch was 9:1,  and  were 1.20 μm and 0.87 μm, respectively. With the increasing content of modified starch in the wall materials, the mean grain size of microcapsules markedly increased (p < 0.05). A high-viscosity emulsion can form larger droplets during atomization. The particle size of the microcapsule powder was increased after spray drying, corresponding to the viscosity observed in the analysis of the rheological properties of the capsaicin emulsion described above. This may be due to increased particle size distribution at higher drying air temperatures [The mean particle size and particle size distribution of capsaicin microcapsule powder formed by spray drying are recorded in eratures . The micIn terms of the three microcapsule products with higher EE , when the ratio of whey protein to citrate mung bean starch was 7:3, the mean particle size of the capsaicin microcapsule powders was more extensive, which was helpful in improving the stability of microcapsules. The results are consistent with the storage stability of capsaicin microcapsules described below. Smaller particles have a larger specific surface area, increasing the possibility of contact between microcapsules and the external environment and decreasing the stability of microcapsule products. and d  of microcapsules were close to each other, indicating that the particle shape of microcapsules was more regular compared to the others. The particle size was more uniform, while the particle size uniformity of other products was poor, and the distribution range was wide.In addition, the viscosity of the emulsion also affects the particle size distribution of microcapsule particles. It was previously reported that as the viscosity of the solutions increased, the number of large particles increased, while the number of small particles decreased . In geneUsing TA software to analyze and compare the obtained DSC curves to study the thermal properties of capsaicin microcapsules is of great significance in evaluating product quality, safety, and stability. The DSC curves and glass transition temperatures of the six capsaicin microcapsules are presented in g). This demonstrates that the glass state of capsaicin microcapsules could remain stable at room temperature. Thus, the structure of capsaicin microcapsules could remain intact after heat treatment, which might be attributed to the compound having a stable structure composed of starch, protein, and capsaicin. Compared with the first four microcapsules with higher EE, when the ratio of whey protein to citrate mung bean starch was 7:3, the maximum glass transition temperature of the product was 120.79 °C. This indicates that the microcapsules required the highest amount of heat for phase change and also implies that the microcapsules prepared with composite wall materials have good thermal stability.The dissolution temperature of natural capsaicin was 210.02 °C. The six capsaicin microcapsules formed after spray drying showed the first endotherm peak near 120 °C, the glass transition temperature  compares the differences among the components of capsaicin microcapsules. There is no variable directly related to the principal component. The contribution value of the first principal component (PC1) was 85.40%, which principally affected the yield of microcapsules, moisture content, EE, solubility, color value, particle size distribution, glass transition temperature, rheological properties, and storage stability. The contribution value of the second principal component (PC2) was 14.60% , which wp < 0.05), which was attributed to the hydrophobicity of starch. Light and temperature significantly affected the retention rate of capsaicin microcapsules. Moreover, the system formed by the composite wall materials with a ratio of whey protein to mung bean starch citrate of 7:3 had the highest retention rate and the best stability.This study used capsaicin as the core substance and whey protein composited with citrate mung bean starch as wall materials to prepare oil-in-water-type microcapsules based on spray drying. The aim of this study was to improve the stability and water solubility of active components of the core material. The results show that capsaicin emulsion is a typical non-Newtonian fluid. The emulsification performance of the microcapsule system with high whey protein content was excellent, and the surface of the microcapsule particles was smooth. The yield, EE, and lightness (L*) were also increased compared to microcapsules with low whey protein content. The moisture content was lower than 3.63%, and the solubility of microcapsules increased significantly and decreased with the increase in citrate mung bean starch content in the wall materials (These findings show that capsaicin encapsulation by spray drying whey protein mixed with citrate mung bean starch as wall materials is efficient. Capsaicin encapsulated in microcapsules has satisfactory stability, good water solubility, and reduced irritability and can be added to other foods to improve its functional properties. Our findings expand the application of capsaicin and reveal a new way to apply microcapsules with starch-based wall materials for delivering functional ingredients. There is great potential for designing food-grade delivery systems to encapsulate natural lipophilic pigments or ingredients.


A new metric for the quantitative and qualitative evaluation of bone stiffness is introduced. It is based on the spectral decomposition of stiffness matrix computed with finite element method. The here proposed metric is defined as an amplitude rescaled eigenvalues of stiffness matrix. The metric contains unique information on the principal stiffness of bone and reflects both bone shape and material properties. The metric was compared with anthropometrical measures and was tested for sex sensitivity on pelvis bone. Further, the smallest stiffness of pelvis was computed under a certain loading condition and analyzed with respect to sex and direction. The metric complements anthropometrical measures and provides a unique information about the smallest bone stiffness independent from the loading configuration and can be easily computed by state-of-the-art subject specified finite element algorithms. The fracture risk was shown to be correlated with bone stiffness9. The stiffness is also a key component for analyzing the skeletal or intraskeletal adaptation and changes in response to a given loading10. The stiffness may also help to better understand the dynamics of the human pelvis during side falls11 or alike injury. Moreover, age and sex dependence of stiffness was shown14: osteoporosis was investigated by analyzing the stiffness15.The stiffness is usually defined as a resistance to deformation. This is a natural mechanical behavior and it describes the capacity of a structure to retain an amount of deformation energy. The stiffness should be considered as a combination of both material and structural properties, which form a mechanical response to a given load. The stiffness is a one of the key measures in bone mechanics. A correlation between stiffness and bone strength was shown by several authorsThe bone quality directly influences the patient’s health  and hence novel methods for predicting bone mechanics in vivo, ideally non-invasively, are of high interest in contemporary trauma, orthopedics and endocrinology research.digital twin. Although the paradigm of digital twin provides a step towards real clinical applications, the bone mechanics must be precisely known before make fully use of digital models.Digital models are often used in biomechanics community to analyze bone mechanics virtually. Nevertheless, such computer models should be driven by clinical patient’s data according to a paradigm of so-called 17, or computational approaches such as finite element (FE) models20. In this study, we introduce an alternative way to estimate bone stiffness. The key idea is to apply spectral decomposition of a stiffness matrix  based on the previous authors’ study21. The resultant eigenpairs composed from eigenvalues and associated eigenvectors contain a unique information about the modal as well as static stiffness. These pairs have following properties: (A)rotation/translation invariant,(B)naturally considering the structural/intrinsic bone properties ,(C)under certain conditions provide the minimal bone stiffness for given boundary conditions together with associated deformation shape,(D)does not necessarily require boundary conditions defined.(A) This property allows to compare arbitrarily rotated bone geometries, and it has furthermore been demonstrated in shape analysis23. (B) The stiffness is constructed often by FE method, where the geometry and material properties are discretized together and represented by a big sparse matrix. (C) The stiffness matrix should be symmetric (positive definite) and constant to obtain minimum static stiffness and corresponding eigenvector. (D) If no kinematic boundary conditions are provided, the spectrum of eigenvalues close to zero contains the eigenvalues corresponding to rigid movement of the body  followed by regular deformable eigenvalues. The rigid eigenvalues will not be in the focus of this study.Bone stiffness is generally location and direction-dependent, and requires a precise and often complex experimental protocols21 provide high sex classification accuracy. Moreover, we compared them with common anthropometric measures and natural frequencies of pelvic bone. We further analyzed the smallest static stiffness of the pelvis with respect to the applied boundary conditions. Our new metric can be considered as modal stiffness, which allows to reconstruct the static stiffness of a bone. Through the study, modal stiffness, smallest static stiffness and natural frequency are called eigenmetric.In this study, we stated a hypothesis that newly formulated mechanical metrics in24. The robustness is predominantly analysed for long bones, for which the bending stiffness is considered as most relevant26 and, according to our approach, the bending stiffness is the smallest one possible for long bones21. Moreover the stiffness of long bone can be uniquely expressed for almost any crossection. Nevertheless, it is not clear or it is difficult to define the stiffness for geometrically complex bones or bone assemblies (such pelvic bone or foot24 or hip-spine complex). At this point, our eigenmetric comes into play, because it provides the direction  and magnitude (quantitative measure) of stiffness based only on bone morphology and bone material properties. The eigenvector in our study can be seen as a resultant deformation after aplying certain bone load, which produces a corresponding modal/static stiffness-this raises the question whether loading modes of pelvic bones within the range of physiological loads corresponded to some eigenvectors. Analyzing such load correspondence could help to better understand anthropological or anatomical variations/evolution of bone, for example by proving that bone is physiologically loaded under its static smallest stiffness. The eigenmetric can be also be benefitial in analysis of hip-spine syndrome and load disbalance after hip total replacement or spinal fusion30. Another potential use of our eigenmetric can be seen in forensic biomechanics as unique bone descriptor .It has been shown that bone robustness (the ratio of stiffness and weight) reflects the evolution of skeletal system and how the skeletal system maintains its mechanical function under different enviromental and genetical factorsThe first eigenvector for model FEMP-I might represent a torsion deformation with maximal value located in proximity to the pubic tubercle and minimal value below the termination of the anterior gluteal line, see Fig. S and natural frequency f are given in Table S as well as natural frequency f is reached for first eigenpair. The best accuracy 0.62 for side classification was reached by static stiffness metric S for the first eigenpair. In Fig. 31 and modal stiffnesses are shown. Only moderate correlations were observed for both sexes with the maximal positive value 0.49 with . Practically, only a few eigenpairs are required to capture most of the information about stiffness. In this study, ten modal stiffnesses close to zero were presenting 94% of the bone static stiffness. The smallest eigenvalue corresponded to the smallest natural frequency 21 and in32. This relation can be also explained by a so-called modal reduction technique, where the quantities of interest are projected on to a reduced space spanned by a few eigenvectors to decrease computational complexity while maintaining model as accurate as possible21.The smallest static stiffness can be approximated by modal stiffness if there exist a load and kinematic boundary conditions that produce a deformation of bone, which is similar to an eigenvector. This relation was already demonstrated in the authors’ previous studyThis fact corresponded to an origin of eigenmetrics, which can be seen as spectral components of bone shape and material information. In fact, the eigenmetrics include apart from geometry information the topological and intrinsic characteristics of bone, which cannot be simply captured by the gold-standard method. Further, we include a side classification as a complementary test to validate tested metrics. As it was expected, no significant side difference was observed from classification tests with tested metrics. The sex classification accuracy of defined metrics comparing to gold-standard seems considerable for individual eigenvalues and depends on the boundary conditions defined. Nevertheless, including used eigenvalues simultaneously into classification algorithm, the final accuracy is excellent (ACC/ACU 98.1%/97.1% ± 0.01%). Moreover, the sex classification accuracy of gold-standard anthropometric landmarks was also comparably high (ACC/ACU 93.3%/92.1% ± 0.01%).33. Apart from these advantages, the drawback of FEMP-II model is the difficulty in interpreting the static stiffness as it is computed within an ill-posed static problem. One interpretation could be in an analogy with an alternative model with suitable boundary conditions defined that produces a deformation corresponding to FEMP-II eigenvector. This hypothesis was partially touched in previous author’s study on beam bending analysis21.We demonstrated that natural frequencies, modal and static stiffnesses (eigenmetrics) can be computed for a configuration without boundary conditions defined (FEMP-II). This presents a serious advantage, given the proper modelling of often complicated anatomical boundary conditions is difficult and introduces an additional uncertainty into the model. Moreover, results showed that our metrics computed on the model FEMP-II have better description and classification accuracy than a constrained model FEMP-I. In a sex classification, the FEMP-II metrics contain four sensitive eigenvalues Fig. , while F35 and as a reliable experimental protocol to calibrate computational models36. In fact, the squared angular frequencies computed from modal analyses corresponded with the eigenvalues in this study up to scale constant given by modal mass contribution . The eigenvectors are, up to a constant of the same shape as shape from modal analysis. The natural frequencies were considered as a metric as well, which complements modal and static stiffness metrics. Further, our computed natural frequencies of pelvic bones in the dataset are consistent with those in literature36.The static stiffness is related to modal stiffness in following way. The static stiffness is computed as an inverse sum of modal compliances (compliance is an inverse of stiffness), see relation . The mod28. This unwanted interaction could be explained through lumbosacral-pelvis junction, which can be considered as load-sharing mechanical node30. In other words, lumbosacral-pelvis complex could be responsible for load balance in hip and lumbar spine. Therefore, what happens to load balance after total hip replacement or lumbar spine fusion? The overlly stiff hip implants  “reinforce” femoral and pelvic bones. Fusing two vertbraes or inserting overly stiff cage between them lead again to increased stiffness (or lower flexibility). Such artificial stiffness may cause a disbalance of the load distribution and hence overloading of other hip-spine regions. This implies implants should be designed such they minimize the load disbalance. Nevertheless, the implant desing strategy is unclear and our stiffness metric could be a good starting point as it provides quantitative (direction) and qualitative (magnitude) of minimall stiffness of bone or bone-implant assembly. Therefore, the influence of implant stiffness on load balance of hip-spine complex is an important topic for next study.The current clinical experience indicates that prior surgical treatment of lumbar spine decreases the outcome of total hip replacement38 of used CT. The lack of individual calibration of density–elasticity relationship may potentially decrease the accuracy and reliability of the finite element models used. This potential source of bias will be analyzed in future experiments. Further, although the proposed metrics can be used without boundary conditions defined, we demonstrated that these metrics’ changes are sensitive to boundary conditions. The boundary conditions in this study roughly mimic physiological conditions (model FEMP-I) and their precise definition  must be included to extend the usability our proposed metrics. We have obtained that smallest modal stiffness corresponds to a smallest static stiffness. Nevertheless, this observation is rather heuristic as we were not able to provide a suitable prove that the first eigenvector always points to a smallest stiffness, see the study21. On the other hand, this limitation does not decrease a potential of proposed metric for studying bone mechanics, because the smallest stiffness can be still found by careful analyzing of stiffness spectra or optimization-based approach proposed in21. The smallest static stiffness and corresponding load configuration of a bone have been yet demonstrated on digital models only and will be experimentally verified in a future study. In this study, we simplified material model of bone to be linear and isotropic. This simplification together with density–elasticity relationship could introduce potential sources of errors, which should be carefully investigated in future study. The bone  contains a microstructure that is the source of anisotropy in mechanical properties and significantly alters elastic response and hence the stiffness as well41. The direction of anisotropy is known for long bones and for mandible43. Nevertheles, little is known for the pelvic bone. The artifitial direction43 or X-ray based direction field44 was considered for mandible bone with good accuracy in mechanical response. These approachech could be the good initial attemp for pelvic bone. Of course, there is also a question how the smallest stiffness is sensitivite to material anisotropy. Therefore, the sensitivity of developed eigenmetrics with respect to material anistropy should be carefully investigated.The presented material model of bone has been defined based on the density–elasticity relation from literature with internal calibrationThe heterogeneous sample population CT data obtained from anonymized routine CT scans  consisting of 46. The masks from CT segmentation were converted to an STL (stereolitography format) representation (VTK library47). Consequently, the volume finite element meshes were automatically built with library TetWild48, see Fig. 49. To estimate Young’s modulus, the CT scans were calibrated internally resulting in hydroxyapatite content in bone38. The density–Young’s modulus in 52. The Poisson’s ratio 49 over the whole bone domain. Two sets of boundary conditions were built. The first model FEMP-I described the fixed boundary conditions, see Fig. 53 and mimic the physiological conditions up to some extent. The second model considers no kinematic and no force boundary conditions, called FEMP-II. Based on the bone geometry, isotropic linear model of bone and boundary conditions defined, the stiffness and mass matrices 54), while 55. The homogeneous fixed boundary conditions are injected into stiffness matrix 54). Once the stiffness matrix is built up, the following generalized eigenvalue problem  can be formulated:i’th eigenvalue and eigenvector associated with of sample m, respectively. In order to compute those eigenpairs, LOBPCG solver from SLEPc library was used to find 10 smallest real eigenpairs56. The overall computational framework is written within problem solving environment library Fenics 2018.154.The CT data was resampled to have a uniform isotropic resolution m’th sample is defined as a pointwise norm:k(degrees of freedom in FE model) of point. This corresponds with looking for maximum of compliance:k we can compute i’th modal stiffness of m’th sample ask is computed in virtue of modal superposition described in21 and can be expressed as:S were based on the theoretical and computational study21.The norm of the eigenvector 31. These landmarks and distances have been adopted from DSP2 tool (Diagnose Sexuelle Probabiliste57). DSP2 is capable of sexing the bone specimens by computing the probability of being male or female using the combination of sex sensitive variables. Chosen landmarks and distances do not only describe the shape of bone, additionally they provide a set of sex-specific information. Pelvic morphology is complex, sexually dimorphic, and hence is considered as a good bone model for analysis of eigenmetric proposed in this study. Additionaly, the information about the side (left/right pelvis bone) was included in the study.The shape of the pelvic bones were parametrized by a series of landmarks called 31, see Fig. 58 and consequently between the template bone and all bones in datasets the symmetric, diffeomorphism map was computed by Symmetric registration method (SyN) with demon metric (ANTs library59). The correlation between automatically and manually computed anthropometric distances is shown in Table To decrease the operator error in construction of large number of FE models, the automatized generation of landmarks and boundary conditions was employed with the help of deformable registration algorithm60. The Pearson’s correlation was used to measure the degree of correspondence of automatized and manually measured anthropometric distances Ensemble random forest method was used to binary classify sex and side. The relation between anthropometric variables and eigenmetrics was analyzed with Spearman’s correlation on a significance level 95%. The classification is evaluated with sensitivity/specificity/area under curve (AUC) metrics summarized in receiver operation characteristic (ROC) based on threefold cross-validation procedure


Surface treatments are expected to be a reason for alteration in fracture resistance of zirconia. This study evaluated the effect of post-sintering processes on the fracture toughness of different types of monolithic zirconia.Classical- (Cz) and high-translucent (Hz) monolithic zirconia discs  were prepared, and randomly divided for surface treatments with 1) as-glazed (AG); 2) finished and polished (FP); 3) finished, polished, and overglazed (FPOG); and 4) finished, polished, and heat-treated (FPHT) technique (n=15/group). Fracture toughness (KIC) was determined with indentation fracture toughness method at load 1 kg for AG, FPOG and 10 kg for FP, FPHT with 15 sec dwelling time. Weibull analysis was applied for survival probability, Weibull modulus (m), and characteristic toughness (K0). Microstructures were examined with SEM and XRD. ANOVA and multiple comparisons were determined for significant differences (α=0.05).p<0.05). The Cz indicated a significantly higher KIC than Hz. The FP significantly enhanced KIC, while OG was unable to raise KIC. HT reduced KIC due to reverse phase transformation.The mean±sd value of KIC (MPa.m1/2), m, and K0 were 1.60±0.19, 7.27, 1.71 for CzAG; 9.57±0.89, 9.97, 10.96 for CzFP; 1.61±0.15, 10.56, 1.68 for CzFPOG; 6.45±0.31, 20.31, 6.60 for CzFPHT; 1.45±0.13, 10.91, 1.51 for HzAG; 6.58±0.24, 27.00, 6.70 for HzFP; 1.24±0.05, 23.90, 1.27 for HzFPOG; and 5.07±0.16, 30.51, 5.15 for HzFPHT. The KIC was significantly affected by the post-sintering process, type of zirconia (Post-sintering processes caused alteration in fracture resistance of zirconia. Fracture toughness was enhanced with FP, but not with either OG or HT process for both Cz and Hz. Surface treatment of zirconia through a finished-polished process is recommended, while glazing and heat-treated are not suggested. Key words:Fracture toughness, glazing, heat treatment, polishing, post-sintering process, zirconia. All-ceramic restoration has become popular and plays a significant role in contemporary restorative dentistry as a result of modern technology of computer-assisted design and computer-assisted manufacturing (CAD-CAM) and the advancement of ceramic nanomaterials, that are capable of providing aesthetic and load-bearing restorations ,2. ZircoPost-sintering processes are clinical procedures that every clinician needs to perform on the zirconia restoration before delivery to the patient. The restorations need to be ground, adjusted, finished, polished, glazed, or heat-treated -13. The Fracture toughness is a mechanical characteristic of brittle material to resist crack propagation under applied stress. It is measured by the amount of energy required for fracture which is quantified by the stress-intensity factor (KIC) ,14. Frac-Preparation of zirconia specimensThe zirconia blanks were milled into a cylindrical shape with an 18 mm diameter (Ø) from pre-colored classical translucent monolithic zirconia , and high translucent monolithic zirconia  and sectioned into a circular disc shape of 1.6 mm in thickness using a low-speed diamond cutting machine . The zirconia discs were compensated for sintering shrinkage with the enlargement factor of 1.2302 for Cz and 1.2334 for HZ. The discs were ground flat to remove surface irregularities with silicon carbide abrasive grit # 500, 800, and 1200, respectively, with water coolant, then sintered in a furnace  according to the manufacturer’s firing schedules. One hundred and twenty zirconia discs of thickness 1.2±0.2 mm and Ø 14±0.2 mm were derived. The discs were sandblasted with 50 µm alumina oxide abrasive with a pressure of 30 psi at a distance of 15 mm from the blasting tip and then cleaned in distilled water. The mixture of glazing paste and liquid  was applied over the blasted surface and fired in a ceramic furnace  to produce a glazed surface.-Post-sintering surface treatmentAll specimens were randomly divided into four groups according to post-sintering surface treatment: AG (as-glazed), FP (finished and polished), FPOG , and FPHT  groups. The specimens in the AG group did not receive any surface treatment. The specimens in the FP group were ground by fine diamond finishing bur  by an air-rotor with a speed of 400,000 rpm and water coolant 50 ml/min. The contact pressure was exactly 50 grams, and the finishing time for each step was 30 seconds in a continuous stroke. The horizontal movement was conducted in one direction with the custom-made load and direction-controlled machine with a fixture for holding the grinding handpiece Fig. A. The buDetermination of fracture toughnessIndentation fracture (IF) was performed in the Vicker’s hardness tester  by placing the treated surface of zirconia against the indenter Fig. B and loaK_IC=0.016 (E⁄Hυ)1/2 (P⁄c3/2) )............. Equation 1Hυ=P/2a2................... Equation 2P is indentation load in newton, c is the radius of crack length (m), Where: KIC is the fracture toughness (MPa•m1/2), E is Young’s modulus (GPa), Hυ is hardness (MPa), a is indent half-diagonal (m), -Statistical analysisThe mean and standard deviation (sd) of KIC for each group of monolithic zirconia were calculated, compared, and then further analyzed using ANOVA in conjunction with a post hoc Bonferroni multiple comparisons using statistical software  to determine significant differences in the KIC of monolithic zirconia materials with different post-sintering processes. The result was considered statistically significant at a 95% confidence interval (CI). Weibull analysis was used to determine the reliability of KIC and to estimate characteristic toughness (K0) as well as the Weibull modulus (m) using Weibull++® statistics  according to equation 3.Pf(KIC)=1-exp (-KIC/K0 )m…………….....Equation 3where: Pf is toughness probability, KIC is fracture toughness, K0 is characteristic toughness, and m is Weibull modulus.-Microscopic examinationThe specimens were coated with gold at a current of 10 mA and a vacuum of 130 m-torr for three minutes, then dried in a desiccator, and finally evaluated the microstructures with a scanning electron microscope . The crystalline phases of zirconia were determined by using an X-ray diffractometer . The crystal structures were examined at a diffraction angle of 20–90º with a 0.02º step size per second intervals using copper k-alpha radiation. The phase was analyzed by cross-reference with the standards database and determined the intensity of peaks using X’Pert Plus software .p<0.05) (p<0.05) . The mean±sd values of KIC of zirconia upon post-sintering surface treatment with AG, FP, FPOG, and FPHT were 1.53±0.18, 8.07±1.70, 1.39±0.21, and 5.76±0.77 MPa•m1/2, respectively  . The res05) Fig. B. The poely Fig. . No signely Fig. . Howeverely Fig. . For nonely Fig. , Fig. 2CThe XRD analysis of the crystalline contents and phases of the Cz and Hz was illustrated in This study indicated that post-sintering processes significantly affected fracture toughness of different types of monolithic zirconia. Therefore, the null hypothesis was rejected for the post-sintering processes, types of zirconia, and their interactions. Fracture toughness is the characterizing value of stress absorption in the material at the crack site before the catastrophic failure occurs ,16. The Post sintering adjustment of restoration through grinding, finishing, and polishing procedures is a stepwise method, which is necessary to proceed from the coarsest grit to the finest grit size. These procedures are needed to achieve a smooth, mirror-like surface that provides less susceptibility to bacterial plaque accumulation and minimizes deleterious effects of low-temperature degradation (LTD) and wears of antagonist natural dentition ,23-26. Set al. 2014 showed that fracture toughness of polished zirconia with fine polishing silicone bur was lower than the coarse polishing silicone bur and machining zirconia. However, the surface roughness from machining and coarse polishing zirconia was not acceptable in a clinical situation  and polished surface (FP and FPHT groups). The glazed surface consisted of low-fusing fluorapatite glass-ceramics that cannot withstand the load as much as the zirconia surface does. The 1 kg load on the glazed surface was capable of producing crack length, which similar to the glass-based dental ceramics . While 1ituation . The glaituation .Conventional translucence zirconia has high KIC especially in the CzFP group and lowers respectively for CzFPHT, CzAG, and CzFPOG groups. This related with the phase transformation as evidenced by the broad peaks of r-phase at 101-crystalline plane for CzFP, which responsible for the t- → r- phase transformation mechanism, resulting in KIC increasing, compared to the CzAG group. The r-phase might affect the crack healing ability or the ability to resist crack propagation . The CzFThe Weibull analysis provided the m, K0, and survival probability. The m in ceramic was used to determine the reliability of the material and the distribution of flaws. A higher m had higher reliability or homogenous distribution of flaws . Most ofWithin the limitations of this study, the fracture toughness was influenced by types of zirconia, post-sintering processes. Each post-sintering process resulted in a different superficial zirconia surface. The bare zirconia that occurred from the multi-stepwise finishing and polishing process provided the highest fracture toughness due to the phase transformation ability and reduction of surface flaws. The overglazing provides a luster surface, but it does not enhance fracture toughness because it is based on low-fusing nano-fluorapatite glass ceramics that is indicated for glazing on the zirconia. Moreover, the adherence between glazing and zirconia was occurred by mechanical retention, so, this layer can chip out and removed over time. The heat treatment after polishing provides the bare zirconia surface, so, the fracture toughness is high. The heat treatment influences the reverse phase transformation, then the fracture toughness was lower than the finished-polished process. Nevertheless, the long-term service life of heat-treated zirconia may better than only the grinding and polishing process. Monolithic zirconia which can be used in the posterior  and anterior region (high-translucent zirconia), the finishing polishing procedure is unavoidable, it provides adequate strength to deliver to the patient while the overglazing or heat treatment is an optional procedure.


The waxy locus controls both the non-waxy and waxy rice phenotypes. Rice starch can be altered into various forms by either reducing or increasing the amylose content, depending on consumer preference and region. Low-amylose rice is preferred by consumers because of its softness and sticky appearance. A better way of improving crops other than downregulation and overexpression of a gene or genes may be achieved through the posttranslational modification of sites or regulatory enzymes that regulate them because of their significance. The impact of posttranslational GBSSI modifications on extra-long unit chains (ELCs) remains largely unknown. Numerous studies have been reported on different crops, such as wheat, maize, and barley, but the rice starch granule proteome remains largely unknown. There is a need to improve the yield of low-amylose rice by employing posttranslational modification of Wx, since the market demand is increasing every day in order to meet the market demand for low-amylose rice in the regional area that prefers low-amylose rice, particularly in China. In this review, we have conducted an in-depth review of waxy rice, starch properties, starch biosynthesis, and posttranslational modification of waxy protein to genetically improve starch quality in rice grains.The  Oryza sativa L.), one of the most vital crops, is a primary meal for more than half of the world’s population and also serves as a source of energy and nutrition for millions of consumers. It is a significant staple food in Asia, West Africa, Latin America, and the Caribbean; the main end-use of rice is human consumption [Rice  protein in rice grains is crucial for amylose synthesis [waxy (GBSSI) serves as a vital/critical gene, but it has a minor influence on the temperature of gelatinization [Wx) locus-coded GBSSI in plants gives rise to low waxy or waxy rice [Waxy rice is the primary focus of plant breeders in China and is generally used in brewing and conventional Chinese cuisine . The devynthesis . For botnization . The absnization . Granulenization . The GBSnization . GBSSI cnization . In stornization ,27, whilnization ,27. Waxynization . Downregnization . The nonaxy rice . Overexpaxy rice . Althougaxy rice .In addition to the approaches listed above, a better way of improving crops may be achieved through the posttranslational modification (PTM) of sites or regulatory enzymes that regulate them because of their significance  and alsoPosttranslational modification (PTM) processes play vital roles in determining the functional performance of the genome and gene transcription . VirtualIn this review, we have conducted an in-depth review of waxy rice, starch properties, starch biosynthesis, and posttranslational modification of waxy protein to genetically improve starch quality in rice grains.Starch, the primary carbohydrate content in plants, is a crucial natural source of feed, raw materials for industry, and food . It playStarch has two major constituents: α-polyglucan amylose, which is essentially linear, and α-polyglucan amylopectin, which is branched ,51. The Indica varieties (waxy) are more resistant to rapid amylase hydrolysis and therefore have a high glycemic index (GI). In the management of diabetes and other diet- or lifestyle-related diseases, in which there is a focus on slowing down digestion of starch and delaying the pace at which glucose breaks down, significant attention has been paid to resistant starch (RS).Starch modification includes an effort to boost or minimize the starch content and adjust the composition and component structure to improve the starch properties by reducing or increasing its content to satisfy consumer preferences and suit particular end uses in industries . Three twaxy gene. Huang et al. [waxy gene have been prolonged used for the modification of amylose content, and most plant breeders commonly targeted this gene for starch improvement in plants. The starch of waxy mutant contains amylopectin and amylose [Wx gene encoding the enzyme GBSSI primarily regulates the synthesis of amylose in seed production, and the amount of amylose in the grain is closely correlated with the amount of GBSSI in the endosperm seed [Wxa and (b) Wxb, which is distinct from a significant discrepancy in the gene quality responsible for Wx content in mature seeds [Wx protein control is characterized by two functional alleles identified based on Wx protein quantity that accumulates in mature seeds [Wxa, mainly found in Indica rice, and Wxb, usually found in Japonica rice, and is found to prevail in high- and low-amylose rice at the waxy locus, respectively [Wx include Wxa, Wxb, Wxin, Wxop, Wxmp, and Wx; these variations influenced the geographical variance in amylose content and consumer preferences depending on the region. Waxy proteins, the major regulators of amylose biosynthesis, have been documented [Wxa allele in the grain is responsible for high amylose content, while the Wxb allele in the grain is responsible for low to moderate amylose content. The allele that regulates the generation of minor/curtailed levels of Wx protein is Wxb, which is primarily expressed in O. sativa subsp. Japonica [Wxa allele generates approximately ten times more Wx protein than Wxb and is commonly dispersed in domestic rice, such as O. indica sativa, O. glaberrim, and the wild progenitors thereof [Wxb synthesizes little amylose owing to the mutation on the 5À intron 1 splice site [Wx locus encoding the GBSSI in plants leads to crops with low or no amylose in storage tissues, without apparent total starch content [Indica, while the T present at the junction leads to cryptic splicing, lower GBSS performance, and low amylose levels in Japonica [Wx) [The amylose quality of starch is influenced by many mutated rice genes , but theg et al.  reported amylose . Low-amy amylose . Amylose amylose . The Wx erm seed . The waxerm seed . There are seeds . Rice Wxre seeds , includiectively . Zhang eectively  reportedcumented ,69. Zhancumented  submitteJaponica . The Wxa thereof . Wxb synice site . There aice site . Active ice site . Molecul content . Park et content  observed content . Heilers content  tested l content . Maize i content . It is t content . Waxy ma content . In rice content . The exiJaponica . To imprJaponica  or by doJaponica ,77. SuppJaponica . The amyica [Wx) , which eica [Wx) .waxy (GBSSI) and ALK (SSS2A) are the major genes responsible for determining the nature of rice cooking and eating quality by influencing the gel consistency, amylose content, and temperature of gelatinization temperature. ALK is a major gene that regulates the temperature of grain gelatinization and a minor gene, affects the quality of the gel and amylose content [ISA and SBE3 [SSIIa and Wx, and a variety of minor genes, SSIV, SBEI, isoamylase, and SBEII3, influence eating and cooking quality have been identified in rice through the study of various varieties [AGPlar, APGiso, AGPsma, SS1, S-II-2, SSII-1, SSIII-2, SSIII-1, SSIV-2, SSIV-1, SBE1, SBE3, SBE4, ISA, PUL, and GBSSII [Wx-1160+111, ISA-1499, ISA-1326, PUL+855, SSII-3+3796, SSI+3216, SSIV-2+437, and SSIII-2-1078 [SuS2, SuS4, SuS3, AGPL2, AGPL1, AGPS1, SSSII-3, SSSI, SSSIII-2, SBEI, SBEIV, and SBEIII [PUL, SSS3A, SSS1, and AGPlar affect the amylose level, AGPiso affects the quality of gelatinization, and SSS4B (SSSIV-2) affects the temperature of gelatinization [Wx haplotype for each related gene and vice versa [Wx haplotype was controlled [SSII-3 has a paramount significance for the gelatinization temperature properties of GT, which suggests that SSII-3 performs a vital role in controlling the gelatinization temperature [SSII-3 alleles: SSII-3-II and SSII-3-I [SSII-3-II had GT values less than those of SSII-3-I [SSII-3, a further study established ISA, SSIV-2, and Wx as minor genes influencing gelatinization temperature additively [SBE1 and SBE3 levels, while overexpression of SBE1 and SBE3 increases the palatability of cereal [SSS1, SSS2A, SBE1, and SBE3, especially SBE3 and SBE1, could improve eating quality, but overexpression of GBSSI diminished the eating quality of cereal [SSS2 genes, two SSS3 genes, one SSS1 gene, and two SSS4 genes in rice plants, the SSS2A action specifies the form of amylopectin formation of rice starch to be either the Japonica or Indica type by performing a particular function in long B1 chain synthesis by extending the short A and B1 chains [TaSSIVb-D in starch granule synthesis in leaves at various developmental stages and reported that TaSSIVb-D mutations decreased the expression of genes and the amount of leaf starch granules.Tian et al.  reported content . The genand SBE3 . Two maiarieties ,80. In ad GBSSII . The fund GBSSII  stated tI-2-1078 , with ead SBEIII ,81. In ace versa . The meantrolled . SSII-3 perature . There aSSII-3-I . CategorSSII-3-I . In addiditively . Grain p1 chains . Guo et 1 chains  used homshrunken 2 gene encodes a modified broad AGP subunit with reduced sensitivity to its adverse allosteric effector, orthophosphate, and more robust connection with small and large subunits, leading to an increased weight of grain per plant up to 38% [shrunken 2 (encoding large AGPase subunits) and brittle 2  at the same time in maize raised the starch content to a higher degree than when both genes were expressed individually [The transformation of wheat with a changed variant of maize p to 38% . Overexpvidually .SUS, AGPase, and SSS, and they are depleted by their enzyme activity [Numerous reports have shown that plant hormone control genes encode enzymes that are essential for starch synthesis enzymes . The proactivity . The effactivity . The traactivity , while aactivity , increasactivity , and impactivity . The intactivity , are lishttps://www.uniprot.org/keywords/KW-0750 (accessed on the 20 January 2021)), starch (glucans and amylopectins) is synthesized via the ADP-glucose pathway by three main enzymes: (1) starch synthase, (2) ADP-glucose pyrophosphorylase, and (3) starch branching enzyme. To generate amylopectins, the arbitrarily branched glucan molecules are precisely debranched through the debranching enzyme. Both glucose-derived sucrose and glucose-derived fructose are synthesized in the cytoplasm and subsequently transferred to the cytosol for sucrose degradation by the invertase enzyme [Starch biosynthesis from sucrose mainly influences the yield of cereal and rice quality in the developing endosperm . It is ae enzyme . Starch e enzyme . Of the e enzyme . Zhu et e enzyme  also repe enzyme . Most bie enzyme . As a pee enzyme .shrunken 2 (Sh2) and the huge brittle 2 (Bt2) gene subunits, mutations in AGPase contribute to a significant decrease in starch content. Starch synthase (SS) can be classified as GBSS, which synthesizes amylose and the extra-long chain fraction of amylopectin, and soluble starch synthase (SSS), which synthesizes amylopectin [The commencement of biosynthesis of starch in storage organs involves sucrose mobilization into glucose-6-phosphate and the importation of G6P into the amyloplast by inorganic phosphate (Pi) exchange; by plastidial phosphoglucomutase (Pi) action, glucose-1-phosphate (G1P) is formed by the transformation of G6P . Geigenblopectin . Geigenblopectin  reportedlopectin . Both amlopectin . Amyloselopectin . Two grolopectin . The bralopectin .SBE, which hydrolyzes an α- linkage within a chain and then catalyzes the creation of an α- connection between the decreasing end of the cut glucan chain and another residue of glucose, presumably one from the hydrolyzed chain, creates the α- branches in starch polymers . RemarkaGenetic findings demonstrate that various SS, SBE, and debranching enzyme isoforms are likely to play unique roles in establishing complex starch structures . To syntPTMs are covalent modifications that alter the principal protein structure in a manner that is sequence-specific, through acylation, phosphorylation, nitration, glycosylation, and ubiquitination, by removal and reversible addition of functional classes . This meDue to the physical–chemical characteristics of reactive amino acids and the cellular atmosphere, including oxygen, pH, metabolites, and reactive oxygen species, or as a result of the activity of particular modifying enzymes, PTMs may occur spontaneously  . SignifiThe reversible phosphate group covalently binds to the hydroxyl group of hydroxyl amino acids such as threonine, serine, and tyrosine but rarely covalently binds to hydroxyproline in protein phosphorylation . Tyr, ThC3 phosphorylation accounts for 20 to 30% of phosphate monoesters, while about 70 to 80% of phosphate monoesters are attached to the C6 position of the glucosyl unit . In ordeWx has rendered it easier to identify more suitable target locations for editing and enabled the amylose content of crops to be controlled more specifically in a more effective manner [Poaceae family, the first cys (Cys337) in GBSSI of rice, rye, corn, sorghum, wheat, and barley is conserved, whereas in non-Poaceae plants such as soybean, pea, Arabidopsis thaliana, cassava, kidney stone, sweet potato, and potato, it is substituted with valine [hpWx directly affects the Wx gene and subsequently influences Haopi amylose content. In addition, by editing the CpG site in the waxy promoter, DNA methylation modification can give rise to rice with preferable eating quality and healthy rice with minimized GI [2 fixation and starch metabolism by balancing the rate of biosynthesis of starch with the availability of carbon and energy in numerous tissues of plants under different environmental conditions [The degree of phosphorylation of starch differs greatly because of the sources of starch in the plant . Hebelste manner . The rede manner ,137. Phoe manner . Liu et e manner  reportede manner . LC-MS/Me manner . Huang ee manner  reportede manner  located e manner . Kringele manner  reportede manner . Phosphoe manner . Gluten’e manner . In prote manner . The line manner . The sece manner . In the h valine . In culth valine . The conh valine . Consumeh valine . It is wh valine . The dish valine . Liu et h valine  reportedmized GI . DNA metmized GI . Lysine mized GI . Amylosemized GI . Out of mized GI . Lysine,mized GI . In recemized GI . Becausemized GI . During mized GI . Many ofmized GI . Meng etmized GI  also obsmized GI . In ordemized GI . Multiplnditions .In wheat, Chen et al.  detected3 aqueous solution [In maize, a phosphorylation-specific dye study of the granule phosphoproteome showed that starch phosphorylase, GBSS, and BEIIb are phosphorylated as they exist in the granule . Crosslisolution . STMP-aqsolution . Phosphosolution .In Arabidopsis, the low MNF-Pin-39 variety activity indicates that the replacement of Ser-570Tyr is counterproductive to activity and, considering significant GBSS abundance, is attuned with the very low amylose content . The subStGWD [StGWD overexpression heterologously leads to a 10-fold increase in grain starch-bound phosphate [In potato tubers, starch phosphorylation can induce both starch degradation and starch synthesis . Due to StGWD . This liStGWD . In the hosphate . The devhosphate .SBEII-b, SSIII, and AGPL I, were substantially decreased at 15 DPA, which is in line with GBSSI, indicating that the impact of drought stress on the biosynthesis of starch occurs at the stage of transcription, translation, and phosphorylation of proteins. Starch crosslinking greatly increases the temperature of gelatinization, whereas the enthalpy is not significantly affected [Nabeshima et al.  observedaffected . The To,affected . Owing taffected .ALK are the major genes responsible for determining the nature of rice cooking and eating quality by influencing the gel consistency, amylose content, and temperature of gelatinization. Rice starch can be altered into various forms by either reducing or increasing the amylose content, depending on consumer preference and region. The majority of consumers prefer waxy and low-amylose rice varieties because of their cooking and eating qualities. It is regionally desired and selected for consumption as cooked rice because of its softness and stickiness. To meet the market demand, which is increasing every day, it is important to focus more on yield improvement of Wx and low-amylose rice because increases in rice production rely largely on the improvement of yield, which could be achieved by posttranslational modification rather than an extension of the planting field. Therefore, there is a need for government, private, and parastatal organizations to invest more in research to improve the palatability of rice for consumers.Significant advancements have been made through the downregulation and overexpression of genes, which are comparatively brute force methods that sometimes contribute to unwanted trade-offs between plant development and stress resilience, and a better way of improving crops may be achieved through the posttranslational modification of sites or regulatory enzymes that regulate them because of their significance. The waxy locus controls both the non-waxy and waxy rice phenotypes. Active granule-bound starch synthase is essential for amylose synthesis, while amylopectin is synthesized as a result of a complex pathway involving various isoforms of starch-debranching enzymes, starch branching enzymes, and starch synthase. An interaction study of 18 genes involved in gelatinization temperature, starch synthesis, gel consistency, and amylose content revealed that waxy and