A weak but still significant correlation was found between the number of commentaries by the EiC and the Ef of their journal . The reason for these correlations is unclear, and whether the methodology used to compute the IF and Ef should be modified needs further research.Journal impact factor (IF) inflation is suggested as a problem resulting from commentaries published by the editors in chief (EiCs) of their respective journals. However, it is unclear whether this is a systemic problem across the top thirty cardiovascular medicine journals. Therefore, the purpose of this investigation was to examine the relationship between the number of commentaries written by an EiC and their journal's IF and Eigenfactor (Ef). Utilizing Spearman rank partial correlations controlling for length of service as the EiC, significant moderate correlations were found between the number of commentaries and the number of first-author commentaries by the EiC and the IF of their journal (r=0.568,  A common method for determining the quality of a journal is the Journal Citation Reports' impact factor (IF) . DevelopDue to the simplicity of calculating the IF, it is possible to manipulate the score. Because the IF is simply a mean score, it can be skewed by outliers. Even one highly cited paper can significantly boost a journal's IF , 5. AnotWhile there is evidence of journal self-citation in the cardiovascular sciences, it is not clear what role editors in chiefs (EiCs) have in this problem . EiCs ofThe primary aim of this study was to determine the relationship between the number of commentaries written by an EiC and the IF of their journal. A secondary aim was to determine the relationship between the total number of publications by an EiC and the IF of their journal to account for the full influence of an individual editor's contributions to their field on their journal's IF. Tertiary aims were to determine whether the EiC being a first author influenced these correlations and to determine if an EiC's publication record also correlated with their journal's Ef.Utilizing the Journal Citation Reports database from Clarivate Analytics, we selected the top thirty cardiac and cardiovascular systems journals based on their IF in the year 2019. Cardiology was selected because of previous research indicating problems with self-citation in this field , 11. TabThe EiCs of journals were found using the journals' websites. PubMed was used to determine the following values for each EiC: number of commentaries published, number of commentaries for which the EiC was the first author, total number of publications, and total number of publications for which the EiC was the first author. The number of commentaries published was restricted to the EiC's journal, whereas total number of publications included all items published regardless of the journal. Commentaries were defined as any article written by the EiC published in their own journal that specifically said “no abstract” in PubMed. The approximate date of the appointment of each EiC was also determined to control for variation in publication counts based on the length of time each EiC served. If a journal had co-EiCs, the first EiC that appeared alphabetically was used.p<0.05.Data were analyzed using SPSS version 26. Data were tested for normality using the Shapiro-Wilk test. Partial correlations were computed using Spearman rank correlations. EiCs' publications were correlated with their journals' IF and Ef while controlling for the number of years the EiC had served to statistically remove the influence of length of EiC experience. Comparisons between correlation coefficients were performed by converting each correlation coefficient into a z-score using Fisher's r-to-z transformation. Asymptotic covariance of the estimates were computed, and these quantities were used in an asymptotic z-test . SignifiFor the journals included in this analysis, IFs ranged from 4.642 to 23.603, with a median IF of 6.730 and an interquartile range of 5.338 to 12.171. Efs ranged from 0.00164 to 0.20502, with a median Ef of 0.01884 and an interquartile range of 0.01132 to 0.02905. Spearman rank correlations between journal IF and Ef and their EiC's publication records are shown in p=0.261 and 0.193, respectively).Considering the IF, no significant correlation was found between the IF of a journal and the total number of publications by its EiC. However, weak to moderate significant correlations were found between the IF and the numbers of first-author publications by the EiC, commentaries by the EiC, and first-author commentaries by the EiC. Considering the Ef, the only significant correlation was between the Ef of a journal and the number of commentaries by its EiC. The magnitudes of the IF and Ef correlations with the number of commentaries and first-author commentaries by the EiC were not significantly different  of their journal while controlling for the number of years served by the EiC. We found weak to moderate, statistically significant correlations between the number of commentaries published by EiCs and these scientometric indices.Journal of the American College of Cardiology releasing a “Highlights of the Year” paper would allow their IF to rise by 0.309. This rise in IF would allow them to overtake Circulation, which at the time had the highest IF among cardiovascular journals, establishing the Journal of the American College of Cardiology as the journal with the highest IF in 2010 [Previous literature shows an association between journal self-citation and the IF . Journal in 2010 . While jHeneberg performed a citation analysis of publications appearing in eleven arbitrarily selected prominent general science and biomedical journals , which sBecause of the problems of journal self-citation and the use of different types of published items to calculate the numerator and denominator of the IF, Liu et al. suggest a number of solutions . These sSome limitations of the current study should be noted. Correlations were used to examine the relationship between EiCs' publications and their journals' IF and Ef. As such, a causal relationship between the two cannot be established. A few of the journals had two EiCs, and only the first one alphabetically listed was used for data collection. Due to the large number of publications analyzed, individual confirmation of whether each article was in fact a commentary was not performed—rather, whether the article had “no abstract” listed in PubMed was the criteria for denoting a commentary piece. Thus, a more precise and efficient method of enumerating commentaries is needed, such as developing a coding script to automate data collection. We examined the top cardiovascular medicine journals, but our analysis was not sensitive enough to determine whether particular journals engaged in excessive self-citation. A more detailed analysis of each individual journal would be required to obtain this information.In conclusion, we found significant moderate correlations between a journal's IF and the number of all commentaries and first-author commentaries published by its EiC. Also, a weak but statistically significant correlation was found between a journal's Ef and the number of commentaries published by its EiC. Whether the methodology used to compute the IF and Ef for cardiovascular medicine journals should be modified requires further research.


The establishment of international sanitary institutions, which took place in the context of rivalry among the great European powers and their colonial expansion in Asia, allowed for the development of administrative systems of international epidemiological surveillance as a response to the cholera epidemics at the end of the nineteenth century. In this note, I reflect on how a historical analysis of the inception of international epidemiological surveillance and pandemic management helps us to understand what is happening in the COVID-19 pandemic today. International relations play a central role in the scientific coordination necessary to halt the contagion of COVID-19 across the world. The two-week delay by Chinese authorities in notifying the World Health Organization of the outbreak of an uncommon form of pneumonia was the first element of international contention that emerged at the beginning of 2020. The US government leveraged that delay to claim compensation from the Chinese government, with the argument that an earlier notification of the outbreak would have stopped the spread of the virus and spared the death of hundreds of thousands of Americans Yang . In thisHistorically, international trade and epidemics went hand in hand. Indeed, since the foundation of the maritime republics of Venice, Pisa, and Genoa, commercial fleets of European powers reaching far-away countries proved to be a harmful vector of contagion Harrison . Quarant2Conseil Sanitaire Maritime et Quarantenaire, an international institution meant to regulate the traffic of the Suez Canal, through which commercial and military vessels of France, Britain, and other European powers reached Asian countries. The use of bacteriological analysis, steam sterilization, and medical inspections were conceived as ways to alleviate the use of quarantine and to safeguard the flow of commerce. In 1891, physician Justin Karlinski reported on the disastrous conditions of the Egyptian medical station of El-Tor, on the Red Sea, where Bosnian and Herzegovinian pilgrims were quarantined on their way back from Mecca. In the drinking water of the camp’s containers Karlinski counted more than “2 million germs per cubic centimetre” (Conférences Sanitaire Internationale At the 1892 Venice International Sanitary Conference, the scientific evidence developed a few years before by Louis Pasteur and Robert Koch about biological agents causing contagious diseases gained ground. Delegates convened in Venice to discuss the quarantine as a part of the activities of the Egyptian In 1893, at the International Sanitary Conference of Dresden, diplomats and physicians adopted the first international epidemiological surveillance system to prevent the spread of cholera epidemics in Europe. Indeed, between 1817 and 1893, five cholera outbreaks hit Asia, Africa, Europe, and the Americas, bringing their medical, social, and economic systems to their knees. The delegates who convened at the conference, Robert Koch included, unanimously decreed that telegraphic notification among diplomatic and national offices was the most important international protective measure against cholera. The compulsory weekly notification included the existence of a cholera outbreak on land, the precise place where it occurred, its starting date, the number of clinically observed cases, and the number of deaths (Conférences Sanitaire Internationale The epidemiological surveillance system of 1893 protected Europe from the sanitary crises of its colonies, and contributed substantially to the misattribution of responsibility for cholera epidemics to social groups that suffered from the disease without having any power to halt or slow the contagion. Despite the fact that John Snow revealed cholera’s means of diffusion in 1849, and that Filippo Pacini discovered its biological agent in 1854, the colonial administrators of Great Britain refused to recognize the structural, environmental causes of cholera outbreaks (D’Abramo et al. The European powers’ rejection of critical self-reflection in the nineteenth century produced a legacy of misguided assumptions about Western medical and administrative superiority whose consequences extend to the current health crisis. Looking at what has unfolded in the course of the COVID-19 global health crises since late 2019, one could argue that the same kind of perceived hegemony still animates many of today’s escalating disputes between the US and China. Over the last hundred years, politicians and diplomats, supported by administrative and biomedical experts, have contributed to the development of epidemiological surveillance systems in order to trace, isolate, cure, and prevent cases of infections and epidemics (D'Abramo and Neumeyer


Monilinia fructicola and Monilinia laxa species are the most destructive and economically devastating fungal plant pathogens causing brown rot disease on stone and pome fruits worldwide. Mitochondrial genomes (mitogenomes) play critical roles influencing the mechanisms and directions of the evolution of fungal pathogens. The pan-mitogenomics approach predicts core and accessory regions of the mitochondrial genomes and explains the gain or loss of variation within and between species. The present study is a fungal pan-mitogenome of M. fructicola (N = 8) and M. laxa (N = 8) species. The completely sequenced and annotated mitogenomes showed high variability in size within and between the species. The mitogenomes of M. laxa were larger, ranging from 178,351 to 179,780bp, than the mitogenomes of M. fructicola, ranging from 158,607 to 167,838bp. However, size variation within the species showed that M. fructicola isolates were more variable in the size range than M. laxa isolates. All the mitogenomes included conserved mitochondrial genes, as well as variable regions including different mobile introns encoding homing endonucleases or maturase, non-coding introns, and repetitive elements. The linear model analysis supported the hypothesis that the mitogenome size expansion is due to presence of variable (accessory) regions. Gene synteny was mostly conserved among all samples, with the exception for order of the rps3 in the mitogenome of one isolate. The mitogenomes presented AT richness; however, A/T and G/C skew varied among the mitochondrial genes. The purifying selection was detected in almost all the protein-coding genes (PCGs) between the species. However, cytochrome b was the only gene showing a positive selection signal among the total samples. Combined datasets of amino acid sequences of 14 core mitochondrial PCGs and rps3 obtained from this study together with published mitochondrial genome sequences from some other species from Heliotales were used to infer a maximum likelihood (ML) phylogenetic tree. ML tree indicated that both Monilinia species highly diverged from each other as well as some other fungal species from the same order. Mitogenomes harbor much information about the evolution of fungal plant pathogens, which could be useful to predict pathogenic life strategies. However, relatively limited data is available for whole mitochondrial genomes. For instance, only 793 mitochondrial genomes have been announced by NCBI Organelle Genome Resources2. Mitochondrial genomes present valuable information to explain both adaptative traits and the evolution of pathogens. Fungal mitochondrial genes can be targeted for plant disease management  differs among fungal groups, and these genes may have been transferred to the nuclear genomes in different eukaryotic species  that are known to cause brown rot disease. Mitogenomes were annotated as well as phylogenetics, and evolutionary selections were evaluated based on the protein-coding regions of the mitogenomes. This provides an essential foundation for future studies on population genetics, taxonomy, and crop protection strategies from the perspective of mitogenomics. disease , 2008. Td genera . Thus, wMonilinia species were selected from the collection of Dr. Ozkilinc. Original isolates were long term stored at −20°C on Whatman filter papers no 1. Isolates were obtained from infected peach fruits from different orchards in six cities of Turkey  (accession number: MN881998) was used as a reference to check the annotations. Annotated gene arrangements were analyzed by using MAUVE 2.3.1 software (M. fructicola were investigated by using the Geneious 9.1.8 program (Core protein-coding genes (PCGs), ribosomal RNA, transfer RNA, and introns were annotated using the online server MFannot  and Mitosoftware . Moreove program .The repetitive elements in the mitogenomes were determined by using Tandem Repeats Finder . The repThe percentages of the conserved and variable regions of the mitogenomes within each species were determined using bioinformatics tools Spine and AGEnt . Additiornl and rns) was assessed for A/T and G/C asymmetry by using the following formulas:The non-synonymous (Ka) and synonymous substitution rates (Ks) were calculated for all PCGs by using DnaSP v6.10.01 . Since aM. fructicola and M. laxa isolates were obtained based on the mitochondrial translation code data four using the Geneious 9.1.8 program , KJ434027.1 , KT283062.1 (Sclerotinia sclerotiorum), KF169905.1 (Glarea lozoyensis), NC_015789.1 , KF650572.1 (Rhynchosporium agropyri), KF650575.1 , KF650573.1 (Rhynchosporium commune), and KF650574.1 (Rhynchosporium orthosporum). The multiple protein sequences were concatenated by using the Geneious 9.1.8 program  had the largest mitogenome size with 179,780 bp, while the isolate (coded as T-B1-A5) from M. fructicola species had the smallest mitogenome size with 158,607 bp. Total GC% content of all mitogenomes ranged between 30.0 and 31.1%.Most of the mitogenomes were extracted as one contig. However, the mitogenomes of five isolates  were represented by four contigs. These contigs were mapped by using the other completed mitogenomes as reference. Statistics of QUAST reports for the assembled mitogenomes of 16 isolates of M. fructicola (BG-B1-A4) was chosen as representative sequence and submitted for the first time to the NCBI GenBank (accession number MT005827)  and ATP synthesis, including cytochrome c oxidase subunits 1, 2, and 3 (cox1-3), NADH dehydrogenase subunits 1–6 and 4L (nad1-6 and nad4l), ATP synthase subunits 6, 8, and 9 (atp6-8-9), and cytochrome b (cob or cytb). Furthermore, two ribosomal RNA genes for large and small subunits (rnl and rns), 32 transfer RNA (tRNAs), and a gene-encoding ribosomal protein S3 (rps3) were annotated. A set of 32 tRNAs of Monilinia isolates encoded for 20 essential amino acids were involved in the mitochondrial protein synthesis  . Besidesus study . Mitogenynthesis . Howeverticodons . The AT-ticodons . The mitM. fructicola were detected in cytochrome c oxidase (cox) subunits 1,2,3 and cytochrome b (cytb), ranging from 23.8 to 11 kb  and ribosomal protein (rps3) showed positive AT and GC skewness in all the mitogenomes of Monilinia samples 29 and (TC)18 located in the intergenic regions 17 in the mitogenome of M. laxa was detected previously  in the nad2 gene in comparison to the other samples. The nad5 gene in the mitogenomes of the two isolates (Ti-B3-A2 and Yolkenari-1) included two extra introns  to compare other isolates of the same species  and within coding regions (14.2%). The introns in mitogenomes of M. laxa isolates covered 38.9% of the whole mitogenome and were within the non-coding region (16.3%), and within coding sequences (22.6%).For all the eight mitogenomes of  M. laxa . IntronsM. fructicola isolates, the most intronic sequence carrying gene was the cytb with 72.8%, and intron rich genes followed by cox1 (66.5%), cox2 (61.5%), and cox3 (53%). The most intronic content was found in the cox2 gene with 65.4% and followed by nad1 (65.2%), cytb (62.1%), and cox3 (53%) for M. laxa isolates. Besides, group I and group II mobile introns were detected in the mitogenomes of M. fructicola , and repetitive sequences considered accessory regions of mitogenomes. However, many of these accessory elements were conserved within the species. The conserved regions formed a large portion of the mitogenome (ranging between 94 and 98%), while the variable regions covered 1.19–5.6% of the whole mitogenomes of the M. fructicola isolates. The most accessory-rich mitogenomes were detected in the isolates coded Ti-B3-A2 and Yolkenari-1 within M. fructicola  carried 0.7% of the total genome as a variable, which was not shared with any other isolate within this species  on mitogenome length variation. R2 values explaining dependent (mitogenome length) and independent (intron length) variables were found as 0.9851 and 0.995 for M. fructicola and M. laxa, respectively.According to the linear model test, intragenic introns had a significant effect  . The Coxelection . The cytelection .rps3 of the M. fructicola, M. laxa, as well as some other species from Heliotales . Besides, the mitogenome of M. laxa used in this study, was submitted in our previous research and can be found in GenBank (Accession number MN881998.1). All the mitogenomes was submitted to GenBank with following accession numbers for each isolate used in this study: MW794295 for isolate B5-A4, MW794296 for isolate BG-B1-A17, MW794297 for isolate SC-B2-A4, MW794298 for T-B1-A5, MW794299 for isolate Ti-B3-A2, MW794300 for isolate Ti-B3-A3-2, MW794301 for isolate Yolkenari-1, MW794302 for isolate Yildirim-2-10th, MW794303 for isolate Yildirim-1, MW794304 for isolate T-B1-A4-2, MW794305 for isolate MT-B1-A3-1, MW794306 for isolate MM-B4-A4, MW794307 for isolate MM-B2-A2, and MW794308 for isolate 2B1-A5.The first mitogenome of GY performed laboratory work and all the data analyses. HO designed the research, recommended, and directed the data analysis methods, and controlled all steps of the study. Both authors confirmed and discussed the results and wrote the article together.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.


Both types of analyzed starches also differently influenced the energy of activation of rotational movements of water molecules. On the basis of the analyses carried out with the use of LF NMR, it can be concluded that physically modified starch acts not only as a stabilizer, but also as an emulsifier, while acetylated starch does not exhibit good emulsifying properties.Starch is a widely known and used emulsion stabilizer. In order to improve its properties, various types of modifications are made that change its ability to emulsify and stabilize. This paper describes the analysis of the molecular dynamics of water using low-field nuclear magnetic resonance (LF NMR) in oil-in-water emulsions obtained with the use of physically or chemically modified potato starch. The analysis of changes in spin-spin and spin-lattice relaxation times depending on the temperature allowed the activation energy value of water molecules in the analyzed emulsions to be determined. It has been shown that the presence of starch influences the values of spin-lattice T Starch is one of the most abundant polysaccharides in nature, and is primary source of energy in our diet as well as a versatile, safe and cheap texture forming agent of food products. In this latter role, it is most often as used as modified starch preparation rather than in its native form. Starch can be modified by means of physical, chemical and/or biochemical methods. As a result, a wide variety of thickeners or gelling, bulking or water retention agents, as well as colloidal stabilizers, are available ,2. HydroThe applicability of different types of modified starches is estimated most often using rheological methods as it is believed that the functionality of starch relates mainly to its ability to increase the viscosity of the system ,6,7,8. TT2. The transfer of energy to the environment requires a longer time called spin-lattice relaxation time and marked as T1. The addition of large molecules, the dynamics of which are much lower, slows down the rotation of water particles that bind at the sorption sites, thus, the relaxation time is shortened. It is known that in systems such as, for example, muscle tissue, water protons are found in different states depending on the mobility of the water fraction that can be defined as free, bound, bulk or entrapped [T1 and/or T2 relaxation times are then the weighted average of the relaxation ratep is the fraction of protons associated with one type of water fraction, while T11, T21 as well as T12 and T22 are the relaxation times of individual proton fractions.There is increasing interest in low field nuclear magnetic resonance (LF NMR), a method designed to study the dynamics of protons ,19. It mntrapped . The meacτ. It describes the time for a molecule to rotate by 1 radian. Bloembergen–Purcell–Pound (BPP) equations (Equations (3) and (4)) [R1 (=1/T1) and spin-spin R2 (= 1/T2) relaxation rates and the average correlation time cτ:μ0 is the permittivity of free space, γ is the magnetogyric ratio, h is the Planck constant, r0 is the distance of the interacting nuclei, and ω is the resonance frequency .Relaxation times are macroscopic parameters. At the molecular level, the mobility of the studied molecules is demonstrated by a parameter called the mean correlation time and (4))  contain cτ, when molecules or functional groups perform rotational movements with a constant frequency, conditioned by their interaction with the environment at a specific temperature. If the rotational movements are thermally activated, it means that the molecules or ions must cross the potential barrier of the height ΔaE, which separates the adjacent minima of potential energy. The temperature dependence of the correlation time is described by the Arrhenius equation and has the form:aE is the energy barrier to the rotation of molecules, R is the gas constant, and T is the temperature on an absolute scale.The above equations are valid for a system characterized by one mean correlation time T12 and T22) that reflect proton the relaxation processes in the water phase and the short components (T11 and T21) related to the oil phase of the emulsions [T12 and T22 have values similar to those characteristic of starch pastes of a similar concentration of starch in water and recorded at a frequency of 15 MHz [T11 and T21 have similar values and additionally similar to those characterizing fats measured at low Larmor frequency [LF NMR shows particular advantages in studies of emulsions due to the strong correlation between their stability and relaxation phenomena occurring in both the oil and the water phase. Due to formation of an emulsion, which is a two-phase system, two components of both relaxation times are observed; the long components . Freshly prepared samples were put in the NMR tubes, closed by Parafilm® and placed in the NMR spectrometer.Beef or porcine fats  were used to form the oil phase of the studied oil-in-water type emulsions. Chemically modified starch E1420  or physically modified starch LU 1432  produced according to Polish Patent No. 207800 were applied for the formation of water phases. As the studied fats were solid at the room temperature, both the oil and water phases (starch suspensions) were heated to the temperature of 40 °C before mixing. Then, the resulting dispersion was heated to the temperature of 90 °C under constant stirring. The emulsions were formed along with the starch pasting process. The heat treatment at the temperature of 90 °C under constant stirring, using a mechanical stirrer MS-H-Pro , continued for one hour. The ratio of oil to water was set to 1/4 in all emulsions, and the concentration of the starch in water phase was set to 0.12 g, 0.17 g and 0.25 g (The fatty acid composition was determined by the gas chromatography technique according to the ISO 17059 standard . The chr1 and spin-spin T2) were performed using pulse NMR spectrometer PS15T  operating at 15 MHz, equipped in system of temperature control.Measurements of relaxation times (spin-lattice T1 were measured using inversion-recovery sequence 180−τ−90 [2 were measured using a CPMG impulse sequence (90−τ−180n) [The spin-lattice relaxation times T180−τ−90 . Spin-la180−τ−90 , allowin−τ−180n) ,43. The The preparation of the emulsion requires the use of an emulsifier a substance capable of reducing the interfacial tension at the oil–water interface. It is believed that starch as a polysaccharide that does not reveal surface activity acts only as a stabilizer and extends the durability of this two-phase dispersed system . PreviouThe type and the concentration of starch also affected the relaxation times in the oil phase of the emulsions, although this effect was smaller than in the water phase. This observation suggests that starch, although not dissolved in the oil phase, revealed an entrapping effect on fat molecules. This may be related to the higher viscosity of the emulsion than that of the oil phase, caused by the presence of the viscous water phase. The course of the changes in the values of relaxation times affected by type of fat used was more evident in the oil phase than in the water phase. This is especially evident for emulsions stabilized with chemically modified starch.For a detailed analysis of the molecular dynamics of proton-containing molecules in starch-containing emulsions, the protons in the oil phase and water phase were analyzed separately. Using analytical solutions of BPP equations (Equations (3) and (4)) , taking aE of the oil phase were recorded; however, both fats used in the research are characterized by a high content of C16:0 and C18:0 fatty acids. It is known, however, that in the presence of water, chains of fatty acids in emulsions create specific lamellar structures, and the use of E1420 starch eliminates the differences between the fatty acid composition of pork and beef. It should also be noted that aE correlates well with the temperature-dependent rheological proprieties of the starch pastes including those derived from both rheometers and viscorpaghs [It was shown that, when using chemically modified starch (E 1420), the molecular dynamics of the oil phase were characterized by an energy barrier approximately twice as high as the oil phase. Similar results were obtained for emulsions containing animal fat and native potato starch . The usecorpaghs . Moreovecorpaghs .As shown in The presence of starch in water phase of the emulsion affects spin-lattice and spin-spin relaxation times in both the water and the oil phases. The kind and the extent of these changes depends on the concentration and type of starch. The most noticeable difference was observed for spin lattice relaxation times in water phase, which was higher for physically modified starch. Physically and chemically modified starches affected the activation energy of rotational movements of molecules in different ways. LF NMR studies indicated that acetylated starch acted as emulsion stabilizer. In contrast, physically modified starch can be recognized as not only a stabilizer, but also as an emulsifier or fat replacer in emulsion studied. Relaxation phenomena are also partially affected by the fatty acids composition in studied fats.


However, radiological and intraoperative factors did not affect postoperative shoulder stiffness . Multivariate analyses revealed early postoperative stiffness was significantly associated with diabetes mellitus and timing of rehabilitation . The overall incidence of early postoperative stiffness following isolated ARCR in patients without preoperative stiffness was 14.2%. Diabetes mellitus and timing of rehabilitation were independent risk factors for early postoperative stiffness following ARCR.The purpose of this study was to investigate the incidence and risk factors of early postoperative stiffness in patients without preoperative stiffness undergoing isolated arthroscopic rotator cuff repair (ARCR). Two hundred seventy-four patients who underwent primary ARCR were included. At 3 months after surgery, criteria for shoulder stiffness was set as follows: (1) passive forward flexion < 120˚, or (2) external rotation at side < 30˚. Patients with preoperative stiffness or who underwent additional procedures were excluded. Patients-related, radiological (muscle atrophy and fatty infiltration), and intraoperative  risk factors were analyzed. Univariate and multivariate analyses were used to identify risk factors for postoperative stiffness. Thirty-nine of 274 patients (14.2%) who underwent ARCR developed postoperative stiffness. Univariate analyses revealed that early postoperative stiffness was significantly associated with diabetes mellitus ( Although most patients experience satisfactory clinical outcomes after arthroscopic rotator cuff repair (ARCR), potential complications have also been reported from a number of studies7. Among reported complications following ARCR, shoulder stiffness is one of the most common, with an incidence ranging from 2.3 to 28.5%12. Another frequently reported complication—shoulder stiffness—was more common in the early postoperative period  with a range from 11 to 35.4%14. Despite an otherwise successful ARCR, postoperative stiffness may lead to a distressing situation and dissatisfaction for both the patient and surgeon13. Therefore, the ability to better assess the risk and prevent postoperative stiffness is crucial, particularly in the early postoperative period.Rotator cuff tears are a common clinical problem with multifactorial etiology including degenerative changes and trauma. A commonly accepted and widely used treatment approach for rotator cuff tears involves arthroscopic repair16. Despite awareness of these risk factors, few studies have comprehensively analyzed patient-related, radiological, and intraoperative risk factors for early postoperative stiffness after ARCR. In addition, little is known about the incidence of early postoperative stiffness in patients without preoperative stiffness undergoing isolated ARCR.Various risk factors, such as preoperative shoulder stiffness, female, diabetes mellitus (DM), hypothyroidism, operative technique, additional procedures, prolonged immobilization, and glenohumeral joint (GHJ) synovitis, have been suggested as causes of postoperative stiffness after ARCRThe purpose of this study was to investigate the incidence and risk factors of early postoperative stiffness in patients without preoperative stiffness undergoing isolated ARCR. We hypothesized that the incidence of early postoperative stiffness would be higher than the previously reported values, and specific risk factors might be related to early postoperative stiffness.This study was approved by the institutional review board of our hospital (KMUDSH IRB No: 2020-07-080). The study was carried out in accordance with the Declaration of Helsinki. All patients signed informed consent forms.Between October 2013 and February 2020, 274 patients who underwent ARCR by a single surgeon were included in this study. Inclusion criteria were as follows: (1) patients with primary ARCR, (2) available medical records, and arthroscopic photos and videos. Exclusion criteria included patients who had: (1) preoperative shoulder stiffness, (2) open or mini-open rotator cuff repair, (3) additional procedures , (4) inadequate medical records , and rotator cuff repair was carried out. The number of anchors and repair techniques  used depended on the tendon mobility, tear size, and pattern. Subacromial decompression for the removal of inflamed bursa tissue and acromioplasty confined to the anterolateral aspect of the acromion was performed in all patientsPatient-related factorsThe following baseline data were collected: age, sex, body mass index (BMI), DM, thyroid disease, hyperlipidemia, duration of symptoms, dominant arm, history of trauma, the intensity of labor, level of sports activity, preoperative University of California, Los Angeles (UCLA) score, preoperative visual analog scale (VAS) pain score, and preoperative American Shoulder and Elbow Surgeons' scale (ASES) score. The intensity of labor was divided into four groups for analysis: (1) heavy labor, (2) light labor (included office work), (3) unemployed, (4) others. The level of sports activity was also divided into four groups: (1) none, (2) overhead sports, (3) contact sports, (4) non-contact sports.Radiological factors19.Plain radiography and MRI  were performed before surgery. Preoperative muscle atrophy was evaluated according to Thomazeau classification for each tendon, and fatty infiltration of muscle was evaluated according to Goutallier classification of each tendonIntraoperative factors20, GHJ synovitis was graded, with total GHJ synovitis scores ranging from 0 to 6. According to the grading system proposed by Jo et al.21, SAS synovitis was graded, and total SAS synovitis scores ranged from 0 to 5.Tear size was measured intraoperatively using a calibrated probe after debridement of the degenerated tendon edges. The anteroposterior dimension was measured at the lateral edge of the footprint, and medial retraction was estimated as the distance from the apex of the tear to the lateral footprint. According to the grading system proposed by Davis et al.22. An independent research coordinator performed a clinical examination. Criteria for shoulder stiffness was set as follows: (1) passive forward flexion < 120˚, or (2) external rotation at side < 30˚ according to the definition of Oh et al.22. Patients were included in the stiffness group if they exhibited either or both of these criteria at 3 months after surgery.To evaluate shoulder range of motion (ROM), passive motion in two directions  was measured with a goniometer. Forward flexion was measured in degrees between the arm and the thorax with the elbow held straight, and external rotation with 0˚ of shoulder abduction was measured with the elbow in 90˚ of flexion between the thorax and the forearmAll patients underwent active ROM of the finger, elbow and hand exercise from the first day after operation. An abduction brace was applied, and the duration was based on the tear size measured at the time of surgery. Patients with small- to medium-sized rotator cuff tears wore the abduction brace for 4 to 6 weeks and were allowed passive ROM of the shoulder from 2 to 4 weeks postoperatively, while patients with large to massive tears wore the brace for 6 weeks and were allowed passive ROM from 4 to 6 weeks postoperatively. Patients began active ROM exercises at 6 weeks after surgery, rotator cuff strengthening exercises at 3 months after surgery, and manual labor and sports activities at 6 months after surgery.2 of 0.15) for multiple linear regression. The independent t-test and chi-square test were used to compare baseline demographics data between both groups. Univariate analysis was used to identify risk factors associated with postoperative stiffness. Significant associations observed in univariate analysis and significant variables reported in the literature were included in the multivariate analysis24. Statistical significance was accepted for p-values of < 0.05.The SPSS statistical package  was used for data analysis. A power analysis indicated that a sample consisting of 230 patients would provide 95% statistical power with an α of 0.05 for medium effect size .All patients gave their oral and written consent for their clinical and radiological data to be analyzed and used in this study.All patients gave their oral and written consent for their clinical and radiological data to be published.Of the 274 patients treated with primary ARCR, 39 patients (14.2%) developed shoulder stiffness at 3 months after surgery.p > 0.05), however, the prevalence of DM was significantly different between the two groups (p = 0.030). DM was observed in 20.5% (8/39) of the stiffness group and 8.9% (21/235) of the non-stiffness group. There were no significant differences in preoperative UCLA score, VAS pain score, ASES score, and the timing of rehabilitation between the two groups  . The grade of fatty infiltration was also not significantly different between the two groups (p > 0.05). An analysis of intraoperative factors revealed that there were no significant differences in tear size, repair techniques, number of anchors, total GHJ score, or total SAS score between the two groups  (Table Muscle atrophy greater than grade 2 was observed in 20.4% (48/235) of patients in the non-stiffness group and 33.3% (13/39) of those in the stiffness group, however, this difference was not significant  . Additionally, DM and the timing of rehabilitation were independent risk factors for early postoperative stiffness following ARCR.12. Reasons for this wide range were the potential subjectivity of the criteria, and inconsistent timing of measurements. Several authors defined shoulder stiffness as passive forward flexion of less than 100˚ and external rotation less than 30˚25. Of the authors using this definition, Brislin et al.25 reported that 23 of 263 patients (8.7%) had shoulder stiffness 3 months after ARCR, and Parsons et al.11 noted that 10 of 43 patients (23.3%) experienced shoulder stiffness 6 to 8 weeks after ARCR. While Tan et al.8 reported that 32 of 290 patients (11%) experienced postoperative shoulder stiffness at their 3 months follow-up visit, 25/32 (78.1%) had resolution of stiffness by 9–12 months. Kim et al.26 defined shoulder stiffness as forward flexion of less than 140˚ or external rotation with the arm in 90˚ abduction of less than 40˚. They noted that 74 of 209 patients (35.4%) experienced postoperative stiffness within 6 weeks after ARCR. Chung et al.14 set the criteria of shoulder stiffness for forward elevation at less than 120˚, external rotation with the arm at the side at less than 30˚, or internal rotation at the back as lower than the third lumbar vertebral level, as previously described by Oh et al.22. According to these criteria, they reported that postoperative stiffness was observed in 18.8% (54/288) of patients at 3 months after ARCR, 2.8% (8/288) at 6 months, and 6.6% (19/288) at the final follow-up (mean 13.5 months)14. However, these previous studies included patients with preoperative stiffness or additional procedures that can affect postoperative stiffness. In the present study, 39 of 274 patients (14.2%) who underwent ARCR developed postoperative stiffness at 3 months after surgery. Unlike previous studies, our study included patients with ARCR only and excluded patients with preoperative stiffness, open or mini-open rotator cuff repair, and any who underwent additional procedure. Stiffness was defined as forward flexion of less than 120˚ and external rotation with the arm at the side of less than 30˚ in our study. These criteria were selected because they are easy to examine in the outpatient clinic. Several studies have included a motion of internal rotation in the definition of stiffness22. However, we excluded an internal rotation from the diagnostic criteria of postoperative stiffness because we thought that a hand-behind-the-back ROM might not accurately assess active and passive internal rotation of the shoulder27.Postoperative shoulder stiffness following ARCR may affect functional outcomes and a patient's satisfaction for the procedure. Because of the clinical importance of postoperative shoulder stiffness, an improved understanding of its incidence and a consistent definition is crucial for orthopedic surgeons. In a review of the literature, the incidence of postoperative stiffness following ARCR varies from 2.3 to 28.5%28. In particular, DM—which is a risk factor for frozen shoulder—has been widely studied for stiffness after ARCR29. Although some articles had not found an association between DM and postoperative stiffness14, several studies have reported that DM could be a potent risk factor for postoperative stiffness23. Blonna et al.15 reported that the overall incidence of postoperative stiffness was 29% (19/65) in patients who underwent ARCR or arthroscopic subacromial decompression. In their study, of the 12 patients who had DM or pre-diabetes conditions, 5 (42%) developed postoperative stiffness . Burrus et al.23 analyzed 232 of 19,229 patients (1.2%) who underwent lysis of adhesions or manipulation under anesthesia after isolated ARCR using the PearlDiver Patients Records Database. They reported that type-1 DM was a significant risk factor . In the present study, it was also noted that DM was independently associated with postoperative stiffness at 3 months; 20.5% (8/39) of DM patients were in the stiffness group compared to 8.9% (21/235) in the non-stiffness group.Although the etiology of stiffness after ARCR might be multifactorial and not completely understood, various risk factors  are reported to be associated with postoperative stiffness24. Parsons et al.11 retrospectively evaluated 43 patients who underwent full-time sling immobilization without formal therapy for 6 weeks after ARCR. They concluded that slower rehabilitation does not result in increased long-term stiffness, but 23% (10/43) patients were determined to have postoperative shoulder stiffness at 6 to 8 weeks after surgery. Koo et al.10 performed primary ARCR in 152 patients and patients with risk factors identified in the previous study  were enrolled in a modified rehabilitation protocol that added early overhead closed-chain passive motion exercises. They reported that no patients experienced postoperative stiffness at a mean of 8-month follow-up compared to a control group (7.8%). However, Galatz et al.30 found that early motion, even passive motion, may result in devastating consequences. This group reported a high percentage (94.4%) of recurrent defects in patients with early passive rehabilitation after ARCR.The present study revealed that rehabilitation was significantly associated with postoperative stiffness, and previous investigations also have shown that rehabilitation was closely related to postoperative stiffness33. Tan et al.8 reported that the GHJ synovitis score was independently associated with postoperative shoulder stiffness at 3 months after ARCR. Tauro9 analyzed 72 patients with rotator cuff tears and concomitant preoperative shoulder stiffness who underwent ARCR and also found that bursitis and articular synovitis were more advanced in the group with severe stiffness. However, scores of GHJ synovitis and SAS bursitis were not significantly different between both groups in the present study.Another potential variable is GHJ synovitis or SAS bursitisThis study has several limitations. First, it is a retrospective analysis, however, our data were collected prospectively by a single research coordinator. The second limitation is the short follow-up. Patients were followed up for 3 months. But, this decision was made since our primary goal was to analyze the incidence and risk factors of early postoperative stiffness. Third, patients in this study were not routinely assessed with postoperative imaging for the presence of cuff re-tear. Nevertheless, the strength of this study is that patients treated with isolated ARCR were included, and patients who underwent additional procedures or preoperative stiffness were excluded. Furthermore, the investigation was performed in a homogenous group of patients who underwent ARCR by a single surgeon, with a relatively large number of cases (274 patients) with various potential risk factors.The overall incidence of early postoperative stiffness following isolated ARCR in patients without preoperative stiffness was 14.2%. DM and timing of rehabilitation were identified as independent risk factors for early postoperative stiffness after ARCR. These findings may allow surgeons to adjust postoperative management in an attempt to prevent early postoperative stiffness. Additionally, surgeons may consider discussing these risk factors with patients before surgery.


P = 0.016), donor body mass index (P = 0.013), recipients' pretransplant serum creatinine (P = 0.036), and intrahepatic expression of heme oxygenase 1 (HO1) (P = 0.015) and tumor necrosis factor α (TNF-α) (P = 0.039) were independent predictors of EAD. Inferior graft and recipient prognosis were observed in patients who experienced EAD (P = 0.028 and 0.031) or received grafts with higher expression of sirtuin 1 (P = 0.005 and 0.013). The graft and recipient survival were worst in patients with both EAD and high expression of sirtuin 1 (P = 0.001 and 0.004). In conclusion, pretransplant intrahepatic expression of HO1 and TNF-α are associated with the incidence of EAD. The combination of EAD and EAD-unrelated proteins showed superiority in distinguishing recipients with worse prognosis.Early allograft dysfunction (EAD) after liver transplantation (LT) accompanies poor prognosis. This study aims to explore the relationship between pretransplant intrahepatic proteins and the incidence of EAD, and the value of combined EAD and protein profiles for predicting recipient and graft survival prognosis. Liver biopsy specimens of 105 pretransplant grafts used for LT were collected and used for immunohistochemistry analysis of 5 proteins. And matched clinical data of donor, recipient, transplantation, and prognosis were analyzed. The incidence of EAD was 41.9% (44/105) in this cohort. Macrovesicular steatosis ( Liver transplantation (LT) has been accepted as the only available curative treatment for patients with end-stage liver disease. With rapidly increasing demand for LT, there is a serious shortage of donor organs. This has promoted the usage of marginal or extended criteria donors such as donation after cardiac death (DCD), steatosis, and senile donor livers. However, the use of these suboptimal grafts leads to higher morbidity and mortality of recipients.Early allograft dysfunction (EAD) is a common complication following LT, with an incidence range between 20 and 40% . In receUp to now, there is still no widely acceptable approach for preventing or treating EAD. Many factors could contribute to the onset of EAD, such as donor age, donor body mass index (BMI), graft steatosis, cold ischemic time (CIT), warm ischemic time (WIT), donor serum sodium, and model for end-stage liver disease (MELD) score . RecentlIn the present study, we aim to explore the relationship between the expression level of the above pretransplant intrahepatic proteins and the onset of EAD, and to explore the predictive value for graft and recipient survival prognosis of the combination of EAD and protein profiles.A total of 561 patients who underwent DCD LT in the First Affiliated Hospital, Zhejiang University School of Medicine from January 2015 to July 2017 were enrolled. We collected clinical information including donor and recipient demographics, blood type, BMI, steatotic status of donor liver, donor hepatitis B virus (HBV) infection, pretransplant recipient MELD score and serum creatinine, operation time, CIT, WIT, blood loss, postoperative intensive care unit (ICU) length, and ventilator support time. The patients' condition should be stably maintained without the need for major interventions such as transcatheter arterial chemoembolization or an artificial liver support system during the interval between pretransplant tests and the day of LT, otherwise the case would be excluded. After the exclusion of pediatric LT, multiple-organ transplantation, split LT, re-transplantation, and missing pretransplant biopsy specimens or essential data, 105 cases were finally enrolled into our study. The median of follow-up duration was 16.3 months.EAD was defined as one or more of the following criteria: (1) serum bilirubin ≥ 10 mg/dL; (2) international normalized ratio (INR) ≥ 1.6 on posttransplant day 7; and (3) serum levels of aspartate aminotransferase (AST) or alanine aminotransferase (ALT) > 2,000 U/L within 7 days after LT. Each recipient was classified as “EAD” or “non-EAD” according to these criteria.The procurement of donor grafts was performed in accordance with the national guidelines for DCD in China. The procedure has been described in our previous studies . BrieflyImmunohistochemistry (IHC) staining was performed according to a standard protocol . Five inU-test or student's t-test. The chi-square test was used to compare categorical variables. Multivariate logistic regression analysis was performed to identify independent predictors of the onset of EAD, and the odds ratio (OR) was calculated. Survival analysis was performed to study the effects on graft and patient survival of each factors. All tests were two-sided, with a P-value of <0.05 considered statistically significant.The statistical analysis was performed using the statistical software IBM SPSS . Quantitative variables are expressed as the mean ± standard deviation or median and range depending on the distribution. Categorical variables are presented as values and percentages. Continuous variables were compared using the Mann-Whitney A total of 105 recipients who received primary LT were enrolled in this study. The incidence rate of EAD was 41.9% (44/105). The baseline characteristics of EAD and non-EAD recipients are summarized in P = 0.009), indicating that pretransplant renal function might affect the onset of EAD. Higher donor BMI rather than recipient BMI was associated with the development of EAD . In addition, macrovesicular steatosis ≥ 30% (P = 0.005), higher MELD score (P = 0.006), and longer CIT (P = 0.001) were identified as risk factors associated with the onset of EAD. Donor/recipient age , blood type compatibility (P = 0.208), and HBV infection (P = 0.740) were not associated with the onset of EAD in this analysis. The blood loss volume during operation in the EAD group was larger than that in the non-EAD group, although the difference was not statistically significant .Through univariate analysis, the EAD group tended to have higher pretransplant levels of serum creatinine than the non-EAD group  than the non-EAD group   .P < 0.1 in P = 0.016), pretransplant serum creatinine , and donor BMI  were independent predictors of EAD development. Moreover, HO1  and TNF-α  were two independent protein markers of the onset of EAD .To analyze independent predictors of EAD, all five proteins in t of EAD . AlthougTo study the effects on graft and recipient survival of EAD and those five protein markers, Kaplan-Meier analysis was performed . ResultsP = 0.013), CIT , EAD , and less blood loss volume  was associated with graft survival prognosis. Factors with P < 0.1 were further considered for multivariate Cox regression analysis, and results showed that only the expression level of SIRT1  and HO1 , EAD , and blood loss volume  were independent predictors of graft survival (P = 0.078).Univariate Cox regression analysis confirmed that the expression level of SIRT1 , but was a sensitive predictor of graft and patient survival prognosis . To address whether the combination of EAD and SIRT1 was a better predictor for graft or patient survival than each one of them alone, the cases were divided into four groups: (1) patients who received grafts with low expression of SIRT1 and did not develop EAD (n = 27); (2) patients who received grafts with high expression of SIRT1 and did not develop EAD (n = 34); (3) patients who received grafts with low expression of SIRT1 and developed EAD (n = 24); (4) patients who received grafts with high expression of SIRT1 and developed EAD (n = 20). As demonstrated in P = 0.001 and 0.004, respectively).An interesting phenomenon is that SIRT1 was not associated with the onset of EAD  in post-perfusion grafts was associated with the occurrence of clinical complications following LT in the first month . Kurian Hypoxia inducible factors are transcription factors whose expression can be regulated by oxygen supply . Since IHeme oxygenase-1 was reported to have anti-oxidative and anti-inflammatory functions . A recenSirtuin 1 is one of the downstream targets of HO1 and plays a vital role in cellular senescence, anti-inflammation, and stress response . SIRT1 dTLR4 was widely reported to have negative effects on liver graft function , 26. TLRTNF-α has critical roles in inflammation and hepatic IRI. Previous animal studies showed that dual-specificity phosphatase 14 could reduce hepatic IRI through quenching TNF-α-induced downstream NF-κB activation . TNF-α wFurther optimization of the EAD definition is urgently needed. Previous definitions of EAD established before 2010 are binary, which identified recipients as with or without EAD , 30–33. Nevertheless, there were several limitations in this study. First, this is a single-center retrospective study with a relatively small sample size, and we did not have enough cases for developing predicting models and for validation, thus the applicability of these results to other central populations remains to be verified. Second, there were some drawbacks to this technique, for example, the subjective nature of IHC and the fact that a small amount of specimen might be insufficient to evaluate the entire graft. And we recognize that expression patterns of proteins in different zonations of hepatic lobules might be of importance in developing a more accurate model for predicting the onset of EAD and the prognosis after LT. However, that is beyond the scope of this research, and further studies on that question should be carried out in the future. These limitations call for a more accurate and representative technique for evaluating the protein expression patterns of the entire graft, such as quantitative detection of released proteins in preservation solution. Finally, only five potential proteins were analyzed in this study due to the limited amount of specimen. Therefore, further studies should be made to identify the clinical value of other relevant proteins expressed in liver grafts. An integrated protein profile may be useful in guiding the evaluation and management of liver transplantation in the future.In conclusion, this study found that pretransplant intrahepatic expression of HO1 and TNF-α are independent predictors of the onset of EAD. And the combination of EAD and intrahepatic SIRT1 might be a superior predictor for graft and patient survival prognosis than using EAD alone.The original contributions presented in the study are included in the article/The studies involving human participants were reviewed and approved by Research Ethics Committee of the First Affiliated Hospital, College of Medicine, Zhejiang University. The patients/participants provided their written informed consent to participate in this study.XX: conception and design. SZ: administrative support. QW, JZ, and KW: data analysis and interpretation. All authors: provision of study material or patients, collection and assembly of data, manuscript writing, and final approval of manuscript.This work was supported by grants from the National Natural Science Foundation of China ; the Key Research & Development Plan of Zhejiang Province (Grant Numbers 2019C03050 and 2021C03118); and the Projects of Medical and Health Technology Program in Zhejiang Province (Grant Number WKJ-ZJ-2120).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 ofinterest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The starch acetates were then annealed at slightly lower temperatures than their pasting temperatures. The annealing process caused an almost twofold increase in the resistance to amylolysis and a threefold increase in the swelling power of the modified starch preparations. The heat of phase transition decreased almost two times and the range of starch pasting temperatures over two times, but the pasting temperature itself increased by ca. 10 °C. The 40 g/100 g addition of the modified starch preparation decreased the rate of drug release from a hydrogel by ca. one-fourth compared to the control sample.This study aimed to determine the effect of “annealing” acetylated potato starch with a homogenous granule size and various degrees of substitution on the thermal pasting characteristics (DSC), resistance to amylases, rheology of the prepared pastes, swelling power and dynamics of drug release. A fraction of large granules was separated from native starch with the sedimentation method and acetylated with various doses of acetic anhydride (6.5, 13.0 or 26.0 26 cm The process of native starch heating in a water suspension having a temperature that fits within the range of glass transition temperatures and the initial pasting temperatures is called annealing. The goal of this thermal modification is to bring starch temperature to such a value that would excite the motility of starch molecules but prevent starch transformation into a paste. It makes the crystalline structure of starch homogeneous, thereby protecting the structure of granules ,2,3. AnnThe annealed native starch of various botanical origin has been broadly investigated and described ,10. In tThe aim of this study was to determine how the annealing of acetylated starch with homogeneous granule size can affect its thermal pasting characteristics (DSC), resistance to amylases, rheology of the prepared pastes, swelling power and drug release dynamics.Potato starch granules differ significantly in size; their diameters can range from a few to over 100 µm. Granules of various sizes undergo pasting at different temperatures and for this reason the raw material used in our study to produce annealed acetylated starch was firstly homogenized in terms of granule size. Particle size distribution of native starch and of native starch with homogenized fraction size determined using a laser particle size diffraction analyzer (LSD) is presented in The mean granule diameter of the non-homogenized starch, expressed as the volumetric diameter D, reached 42.3 μm, and that measured after separating small granules reached 61.7 μm. In the present study, the acetylation of large starch granules with various doses of acetic acid anhydride resulted in the production of starch acetates which differed significantly in the degree of substitution and, consequently, in the initial pasting temperatures .The dependency between starch acetylation and the resulting decrease in pasting temperatures, including the greater decrease along with an increasing degree of starch substitution with acetic acid residues, has been extensively documented in literature ,17,18,19The annealing of acetylated starch led to a significant increase in the initial  and finaThe resistance of starch acetylated with various doses of acetic acid anhydride to amylolysis increased from a few to dozen per cents along with an increasing DS of starch acetates . The annThe annealing of native starch induces changes in its swelling powers. Trends and magnitude of these changes depend on the botanical origin of starch and process parameters ,33,34,35The modified properties of annealed acetylated starch prompted us to perform a pilot experiment to identify possibilities of the practical application of starch preparations modified in this way. Many authors work on bioderivative polymers used in the controlled drug delivery . Figure Experimental materials included: Superior Standard potato starch manufactured by PEPEES Łomża S.A. Acetic anhydride was purchased at POCH SA Gliwice company.A fraction of large granules was separated from native potato starch with the sedimentation method. A suspension was made from 5.3 g of starch per 100 g of the solutions in a 15-L vessel, with the height of the liquid column reaching 19.5 cm. After thorough stirring, the suspension was left for 3 min. Then, the precipitate was decanted and the missing volume was completed with distilled water. Another sedimentation and decantation cycle was repeated three times more. The precipitate of large granules was air-dried at a temperature of 30 °C for 48 h. Native starch and starch left after the separation of the large granule fraction were analyzed using a laser particle size analyzer  with a Hydro 2000 attachment.The large granule fraction was acetylated with the basic dose of acetic acid anhydride used for potato starch modification under industrial conditions (13 mL/100 g starch)  and withThe pasting characteristics (DSC) of acetylated starch was used to establish the initial pasting temperature, which was further used to selected annealing temperatures. A suspension from 10 g of starch/100 g of the solutions was prepared in a 2-L vessel from starch acetylated with three doses of the acetic acid anhydride . Under continuous stirring, the suspension was heated (water bath with stirrer) to the temperatures of 52 °C, 48 °C or 43 °C (for respective anhydride doses) with stirring speed 240 rpm for 8 h and left for 48 h. Afterward, it was rinsed three times with 5-L portions of distilled water. Starch precipitate was separated each time from the suspension using a Stratos flow centrifuge , then air-dried at 30 °C for 24 h, ground in a mortar, and sieved through a screen with mesh size of 400 μm.3 of distilled water and stirred. Then, phenolphthalein was added and a 0.1 M NaOH solution was instilled to obtain a light-pink color of the mixture that maintained for 1 min. Next, 25 cm3 of a 0.5 N NaOH solution were added and the reaction mixture was stirred on a shaker for 35 min. Afterward, it was titrated with a 0.5 M HCl solution with a known titer . The released drug amount was assessed via spectrophotometry  according to the available bibliography ,43.Results were statistically analyzed using Statistica 13.3 PL software package. Based on statistical computations , values of the least significant differences and standard deviation were calculated and equations of flow curves were determined. For statistical evaluation, the results were subjected to one-way analysis of variance at a significance level of 0.05. Values of the least significant difference (LSD) between the means were computed using the Duncan’s test at a significance level of 0.05 .The annealing of large granules of acetylated starch caused an increase in pasting temperatures, starch resistance to amylolysis and swelling power, but also a decrease in the heat of modified starch pasting. The magnitude of the observed changes depended on starch acetylation degree. The viscosity of pastes made from double-modified starch was determined mainly by the degree of starch acetylation. The annealing of starch with a lower DS caused an increase, whereas that of starch with the highest DS caused a decrease in the viscosity of the pastes. The 40% addition of modified starch decreased the rate of drug release from the hydrogel by 25% compared to the control sample. This promising discovery deserves in-depth investigations to identify its potential applicability in the pharmaceutical industry.


The ankle sprain is one of the most prevalent traumatic injuries in sport, associated with a significant risk of developing Chronic Ankle Instability (CAI) ,2. This n = 14 , age 22.0 ± 2.225 years) or absence of unilateral CAI  identified through the Ankle Instability Instrument and the Foot and Ankle Outcome Score. The ground reaction forces, and centre of pressure (CoP) were assessed during dominant and non-dominant single leg stance during 30 sec on an unstable surface while the participants performed the Stroop test. The measures of CoP displacement, root mean square (RMS), standard deviation, velocity and area and functional stiffness were assessed through the data obtained by a force plate. Functional stiffness was assessed through the relation between the moment of force and the angular position of the ankle according to the method proposed by Winter [A cross sectional study was performed with a sample of 28 athletes of modalities of increased risk for ankle sprain. Participants were divided into two groups according to the presence  but also when this limb was the non dominant one  and in the contralesional dominant limb . An increase in the RMS in the anteroposterior direction was observed in the lesioned limb , and a decrease in the CoP displacement in the non lesioned limb , in comparison to the control group but only when these limbs were dominant.The CAI group showed an increase in the functional stiffness for mediolateral direction in the lesioned limb when this limb was the dominant one (Subjects with CAI seem to present increased bilateral functional stiffness as a possible strategy to increase postural stability in single leg stance, however stiffness assessments based on kinematics should be performed to confirm these findings. Rehabilitation should consider strategies to promote more efficient bilateral postural control mechanisms in subjects with chronic ankle instability.


Starch-based hydrogels are natural polymeric structures with high potential interest for food, cosmeceutical, and pharmaceutical applications. In this study, the physical stability of starch-based hydrogels produced via high-pressure processing (HPP) was evaluated using conventional and accelerated methods. For this purpose, conventional stability measurements, namely swelling power, water activity, texture, and organoleptic properties, as well as microbiological analysis of rice, corn, wheat, and tapioca starch hydrogels, were determined at different time intervals during storage at 20 °C. Additionally, to assess the stability of these structures, accelerated tests based on temperature sweep tests and oscillatory rheological measurements, as well as temperature cycling tests, were performed. The experimental results demonstrated that the physical stability of starch-based HPP hydrogels was interdependently affected by the microorganisms’ action and starch retrogradation, leading to both organoleptic and texture modifications with marked reductions in swelling stability and firmness. It was concluded that tapioca starch hydrogels showed the lowest stability upon storage due to higher incidence of microbial spoilage. Accelerated tests allowed the good stability of HPP hydrogels to be predicted, evidencing good network strength and the ability to withstand temperature changes. Modifications of the rheological properties of corn, rice, and wheat hydrogels were only observed above 39 °C and at stress values 3 to 10 times higher than those necessary to modify commercial hydrogels. Moreover, structural changes to hydrogels after cycling tests were similar to those observed after 90 days of conventional storage. Data obtained in this work can be utilized to design specific storage conditions and product improvements. Moreover, the accelerated methods used in this study provided useful information, allowing the physical stability of starch-based hydrogels to be predicted. Hydrogels are a commercially widespread group of polymeric materials, consisting of three-dimensional crosslinked networks of hydrophilic or hydrophobic biopolymers capable of absorbing and retaining a significant amount of water . In receRecently, high-pressure processing technology (HPP) was proposed to produce natural-starch-based hydrogels ,13,14. HStarch-based hydrogels form thanks to the physical entanglement of polymer chains or other non-covalent interactions, giving a reversible nature to these structures, which represents an advantage in biomedical and food applications for these materials. However, due to their non-permanent bonds, HPP hydrogels are commonly considered weak structures with reduced rheological properties compared to chemical hydrogels. This hinders their utilization in applications where good mechanical properties and stability are desired . It is iNevertheless, to the best of our knowledge, no studies on the physical stability of these novel structures have been carried out, the determination of which is of the utmost importance in view of improving the design of these biomaterials, optimizing the processing conditions, and proposing their application in different industrial sectors.This work aimed at evaluating the stability and physicomechanical characteristics of starch-based HPP hydrogels with time by applying different stress conditions, using either specific conventional or accelerated detection methods, and determining the shelf life of hydrogels over short storage periods. Bearing in mind that the guidelines for stability testing  commonlGrowth curves of aerobic mesophilic microorganisms, yeasts, and molds on starch-based HPP hydrogels stored at 20 °C are shown in The initial microbial counts in all starch-based hydrogels tested were within the undetectable range, namely < 1 log CFU/g . MoreoveFurthermore, the results obtained gave interesting insights into the microbiological fingerprint of starch-based hydrogels produced by HPP, indicating that microbiological stabilization of these materials should be a matter of concern during processing.w). w values of corn, rice, wheat, and tapioca HPP hydrogels during storage at 20 °C.Water is the main solvent that influences the processing, quality, texture properties, and stability of starch-based hydrogels ,32,33,34w values (in the range of 0.974–0.969). Tapioca starch hydrogels showed the highest initial SP values (7.4 g H2O/g dry starch), followed by rice (6.5 g H2O/g dry starch), corn (5.4 g H2O/g dry starch), and wheat starch hydrogels (5.16 g H2O/g dry starch). These results can be explained by the higher capacity of tapioca starch granules to swell and solubilize with respect to cereal starch granules and the absence of amylose–lipid complex formation, as reported elsewhere [At the beginning of the storage period (0 days), starch-based HPP hydrogels presented similar Alsewhere ,22,35.2O/g dry starch for rice and −2.2 g H2O/g dry starch for tapioca starch hydrogel were measured. These results are in slight agreement with the ones reported by Torres et al. [w/w. However, it should be emphasized that either the gel formation conditions (60 °C for 30 min) or the storage temperature (4 °C) was more severe, and could have delayed the physical and microbiological deterioration of the hydrogels. In our case, it could be assumed that the differences observed among samples can be attributed to the different starch retrogradation extent in hydrogel samples. Retrogradation has been defined as a structural reorganization of the starch chains due to the crystallization of amylose and amylopectin molecules during storage, causing gel shrinking and phase separation (syneresis) [Nonetheless, an evident syneresis phenomenon was observed, as shown in s et al.  for potaneresis) . The ratneresis) ,38. Therw, which accounts for the status and dynamics of water in association or interaction with other molecules in complex matrices [w was observed during storage. Through Aw measurements, it was possible to evaluate the free water present on the surface of the sample, which changed only marginally with storage time. On the contrary, water loss, also quantified as free water in SP measurements, during storage time was much higher than that detected through Aw due to the centrifugal forces applied in this method. These discrepancies suggest that the water entrapped in the polymeric network experienced different changes upon storage, moving outwards layer by layer, reaching the hydrogel surface from where it was easily removed during SP measurements.Measurements of Amatrices , were caThe physical stability of polymeric materials is defined as the extent to which a product retains, within the specified limits, the same properties and physical characteristics possessed at the time of its packaging . TextureThe firmness gives information on the strength of a gel structure, whereas the adhesiveness accounts for the adhesive forces present in starch-based hydrogels . As can Similarly, as the storage time increased, the adhesiveness of rice, corn, and wheat starch hydrogels decreased due to the viscoelasticity losses of the gel network. Interestingly, the increased adhesiveness of tapioca starch hydrogels after 30 days of storage can be attributed to the leaching of water, amylose, and amylopectin, which determine adhesive forces in starch gels .Data for the organoleptic properties of rice, corn, tapioca, and wheat starch hydrogels measured during storage are reported in In agreement with the results of the previous sections, tapioca starch hydrogels showed more significant organoleptic changes than rice, corn, and wheat hydrogels. Syneresis and phase separation after the first month of storage, and consequently, the rupture of the structure after 60 days, were detected via the organoleptic evaluation as the main physical changes of tapioca starch hydrogels during storage. Interestingly, only slight organoleptic modifications of rice starch hydrogels were observed during the storage period . These fFurthermore, from conventional stability results, it can be hypothesized that the stiffer the starch-based HPP hydrogel is, the more marked the structural deterioration, meaning weaker physical stability can be expected when microbial spoilage promotes starch retrogradation. Starch, on which the hydrogels under investigation in this work are based, is a polymer chain of glucose molecules linked to each other through glycosidic bonds that can undergo important modifications during storage. The structural reorganization of its macromolecules and the attack of spore fungiform microorganisms that normally possess amylases  can syneIn accelerated stability tests, a product is subjected to high-stress conditions. For the sake of completeness, the applied stress required to cause starch-based hydrogels structure failure was determined in this work. These data can be utilized to predict the capability of products to keep their initial characteristics, as well as for processing improvements . In our Temperature cycling tests are accelerated physical methods that are commonly utilized in pharmaceutical sciences to provide information on a product’s instabilities that is not provided by isothermal tests . Cyclic p ≤ 0.05), whereas a sharp decrease in firmness was observed in tapioca starch hydrogels (−88%) (p < 0.05), as well as an increased adhesiveness (p ≤ 0.05) after the cycling tests. These results could be attributed to the leakage of water and solubilized amylose, or amylopectin molecules promoted by temperature changes (cycles from 4 °C to 40 °C), causing changes in the interactions occurring in the gel network and the adhesive forces. This is in agreement with the findings reported by Schirmer et al. [The data reported in r et al. . The autFurthermore, the results of the temperature cycling tests showed that physical changes in cream-like hydrogels are more related to the network weakening than to rupture, differently from what was observed in tapioca starch hydrogels. Remarkably, these physical changes are similar to those observed after the conventional storage period, highlighting the effectiveness and accuracy of this accelerated method to predict the physical instabilities of starch-based HPP hydrogels in shorter time periods.Rheological tests have been proposed to predict the stability of products and provide useful information to improve formulations or overcome the occurrence of certain instabilities in gels structures . For thiTo predict the thermal stability of the starch-based HPP hydrogels studied in this investigation, temperature sweep tests from 25 °C to 60 °C at a heating rate of 1 °C/min were performed. For the sake of comparison, thermal gels were also tested. From It is well known that gels obtained with physical methods exhibit a strong dependence of the G′ modulus on temperature, with significant decreases in elastic domains observed with increasing temperature, mainly due to the loss of interconnectivity of the network constituents . InteresMoreover, some studies have demonstrated that differing from thermal gelatinization, in HPP starch gels a characteristic retrogradation phenomenon is likely to occur due to the different dynamics of water and the presence of almost intact starch granules (absence of stirring). This prevents lower amylose leaching, which in turn is less prone to retrograde ,49,50. TThe network strength of starch-based HPP hydrogels, and for the sake of comparison of a commercial hydrogel, namely Carbopol, was evaluated through stress sweep tests. In All samples showed the typical behavior of gels structures, with elastic properties predominating over the viscous ones (G′ >> G″) ,14,53. IStarch-based HPP hydrogels are structures with high application potential in pharma, cosmetic, and food sectors. These novel structures are required to be physically stable at room temperature, and microbiological spoilage should be avoided. Thus, the determination of their stability is crucial to assess their performance with storage time. In this study, conventional and accelerated methods for the evaluation of starch-based HPP hydrogels were used. Conventional techniques demonstrated that the physical stability of starch-based HPP hydrogels depended on the occurrence of microbial spoilage and starch retrogradation, which synergically triggered the physical deterioration of the gel structures, such as network weakening and phase separation upon storage at room temperature.Accelerated methods were proven adequate to evaluate the stability of hydrogels, requiring a shorter testing period. They allowed us to predict good physical and thermal stability and good network strength for starch-based HPP hydrogels, superior to the results for commercial hydrogels. Rheological alterations of rice, wheat, and corn HPP hydrogels were demonstrated to take place only at temperatures higher than 39 °C.In conclusion, data obtained in this study enabled an understanding of the limits of these novel biomaterials and predictions of their physical stability, as well as improvements in their design and forecasting of their novel applications.Rice (S7260) , wheat (S5127) , and corn (S4126)  starch powders were purchased from Sigma Aldrich . Tapioca starch  was obtained from Rudolf Sizing Amidos do Brazil . Starch moisture content was determined according to AOAC guidelines (925.10). Amylose content was determined by an enzymatic rapid assay kit .w/w). To ensure sample homogeneity and avoid particles sedimentation, the starch suspensions were prepared immediately before HPP treatments.Based on previous experimental findings ,13, diffHPP treatments were performed in a U22 laboratory-scale high pressure unit  as described elsewhere .In each test, 10 g of the starch suspension was de-aerated, thoroughly mixed, and packed in flexible pouches, which were then sealed, loaded in the pressure vessel, and HPP-treated at 600 MPa for 15 min at 25 °C. This processing condition was sufficient for complete gelatinization and proper gel formation .HPP hydrogels were stored at room temperature (20 °C) before analyses.Rice, wheat, corn, and tapioca HPP hydrogels were stored at 20 ± 0.2 °C in an incubator . Microbial counts, organoleptic and texture properties, as well as swelling power and water activity levels were evaluated every four weeks. For all experimental determinations, three different samples of each starch-based hydrogel were used. Unless otherwise stated, all measurements were performed in triplicate.Hydrogels samples were analyzed to enumerate the numbers of mesophilic aerobic microorganisms, yeasts, and molds. Each bag containing 10 g of starch-based HPP hydrogels was aseptically opened, and together with 90 mL of buffered peptone water , was introduced in an aseptic stomacher bag. A Stomacher 400 unit  at 260 rpm was used for the complete homogenization of the samples. Further decimal dilutions were prepared with the same diluent and plated on appropriate growth media.To enumerate aerobic mesophilic microorganisms, 1.0 mL of each dilution was pour-plated in Plate Count Agar  and incubated at 30 °C for 72 h. To count yeasts and molds, 1.0 mL of each dilution was spread-plated on Dichloran Rose Bengal Chloramphenicol medium agar  and then incubated for 3–5 days at 25 °C.The microbial colonies were identified and quantified by standard methods. Microbiological data were transformed into logarithms of the number of colony-forming units (log CFU/g). The detection limit was 10 CFU/g (1.0 log CFU/g).The organoleptic properties—namely the appearance, homogeneity, integrity, color, and odor—of the samples during storage were monitored using visual observations. Additionally, the physical appearances of samples were recorded using a digital camera  in angular mode. Original pictures without editing and filtering were stored.Swelling Power (SP)The swelling power levels of hydrogels were determined according to the method reported by Kusumayanti et al.  and modig for 10 min, the supernatant was removed, and the pellet was weighed before and after drying for 6 h at 105 °C. The swelling power was evaluated as follows:Each hydrogel sample was centrifuged  at 1351× w) levels of samples were measured using a Novasina water activity instrument . Before Aw measurements, the instrument was calibrated at 25 °C using standard patterns. For the measurements, 1 g of hydrogel was poured into the center of the sampling chamber and consecutive readings were carried out until constant Aw values were attained.The water activity  equipped with a 5 kg load cell, connected to a microcomputer. The textures of hydrogel samples were determined according to the procedure reported by Larrea-Wachtendorff et al. . BrieflyThe cycling test was performed according to the method reported by Almeida and Bahia  with sli®, Eurofarma, Chile) and thermal gels. The latter were produced according to the method and processing conditions described by Larrea-Wachtendorff et al. [The rheological analysis of samples was carried out in a controlled stress rheometer  in a plate–cone geometry configuration  with a gap of 52 µm. The viscoelastic responses of hydrogel samples were recorded (immediately obtained after HPP treatments), namely the elastic (G′) and viscous (G″) moduli under different stress (0.01 to 1000 Pa at 25 °C) and temperature (25 °C to 80 °C at 1°C/min) conditions. Additionally, for the sake of comparison, the same measurements were also carried out on commercial hydrogel samples . All analyses were performed using Statgraphic Centurion XVI Statistical Software .The results were analyzed using statistical descriptive analysis (mean ± SD), one-way ANOVA, and post-hoc comparison using the Fisher least significant difference (LSD) test to determine significant differences among experiments (