To provide a stable surgical view in Minimally Invasive Surgery (MIS), it is necessary for a flexible endoscope applied in MIS to have adjustable stiffness to resist different external loads from surrounding organs and tissues. Pneumatic soft actuators are expected to fulfill this role, since they could feed the endoscope with an internal access channel and adjust their stiffness via an antagonistic mechanism. For that purpose, it is essential to estimate the external load. In this study, we proposed a neural network (NN)-based active load-sensing scheme and stiffness adjustment for a soft actuator for MIS support with antagonistic chambers for three degrees of freedom (DoFs) of control. To deal with the influence of the nonlinearity of the soft actuating system and uncertainty of the interaction between the soft actuator and its environment, an environment exploration strategy was studied for improving the robustness of sensing. Moreover, a NN-based inverse dynamics model for controlling the stiffness of the soft actuator with different flexible endoscopes was proposed too. The results showed that the exploration strategy with different sequence lengths improved the estimation accuracy of external loads in different conditions. The proposed method for external load exploration and inverse dynamics model could be used for in-depth studies of stiffness control of soft actuators for MIS support. In Minimally Invasive Surgery (MIS), rigid or soft surgical instruments need to be inserted into human body through incisions of minimal size or natural orifices for accessing the target sites. Compared to traditional open surgery which requires large incisions, MIS has been found to provide patients with a variety of benefits, including improved safety, cosmesis, shorter recovery time, shorter hospital stay, fewer postoperative complications, and less pain from the patient’s point of view . ConsequIn addition, some companies such as Ambu Inc. and Olympus Corp have already manufactured flexible endoscopes, which consists of a long flexible tube with a high-resolution camera and a light source at its tip. Their tips have a single degree of freedom (DoF) and can be controlled by means of a one thumb-controlled toggle or two thumb-controlled dials at their end. They can vary in diameter and length depending on the application and the patient. Flexible endoscopes are usually applied in thoracoabdominal and bladder-related surgical procedures ,5,6. Notably, the flexible endoscope is susceptible to external load from surrounding organs or tissues, especially when it must traverse or pass through several organs during a surgical procedure, such as a Transvesical Prostatectomy . In MIS Soft actuators can self-adjust their stiffness and are ideal for dealing with unstructured environments or interacting with humans because their softness allows them to deform around their surroundings. Soft actuators are also commonly employed in different fields, such as rehabilitation ,11,12,13The first type of pneumatic soft actuators could self-adjust stiffness rely on internal Shape Memory Alloys (SMA)  or a graIn some cases, when a pneumatic soft actuator with an endoscope is compressed and deformed by an external load or resistance from the surrounding organs and tissues, the pneumatic soft actuator should increase stiffness to resist the load. However, an over-stiffness generated by high air pressure to both antagonistic chambers may raise the risk of not only damage to the surrounding organs, but also air leakage from the soft actuator . TherefoIn recent years, there has been increasing interest in soft actuators embedded with sensing mechanisms ,27,28,29Self-sensing approaches based on NNs have been proposed, as NNs are capable of approximating functions with great precision. The defining characteristic of this method is that sensing is realized with neither independent nor embedded physical sensors; instead, it makes use of the relationship between target physical quantities and a set of state variables that are directly or indirectly affected by or interacting with the physical quantities. The relationship can be learned as a function from collected raw data of a soft actuator . A NN-baIn this study, we proposed a NN-based active load-sensing scheme and stiffness adjustment for a pneumatic soft actuator for MIS support with antagonistic chambers for three degrees of freedom (DoFs) of control (described as soft actuator for simplicity). An environment exploration strategy was studied for improving the robustness of the sensing. The minimal exploration sequence was determined by the estimation error and change in the smallest bending angle. A NN-based inverse dynamics model was established for estimating the air pressure from the predicted load and desired stiffness.This paper is organized as follows. The measurement approach for the soft actuator prototype experiments is depicted in As shown in ® aScopeTM 4 Broncho Slim, diameter: 3.8 mm; Ambu® aScopeTM 4 Broncho Regular, diameter: 5.0 mm; Ambu® aScopeTM 4 Broncho Cysto, diameter: 5.4 mm; Ambu Inc., Copenhagen, Denmark, displayed in Because endoscopes of varying diameters impede the expansion of the soft actuator’s chamber differently, the bending angle_o, bending angle_e, and -x deformation were all measured with the soft actuator inserted in three different types of endoscopes  from organs or tissues to provide a surgical view for surgeons. Therefore, the external load in the axial direction was very small, which allowed us to focus on the external load in the horizontal direction. Adjusting the stiffness on −x direction could help the flexible endoscope to resist different external loads in the −x direction. The overall stiffness in the −x direction , a sequence of actuation is implemented to explore the environment for dealing with the influence of the nonlinearity of the soft actuating system and the uncertainty of interaction between the soft actuator and its environment. Taking FD as an example, n means forward exploration , while a negative value means backward exploration , and m_step is a minimal exploration step (10 kPa in this study). Six different exploration sequences  were investigated. The biggest step number of exploration sequence is 6 and the smallest step number of exploration sequence is 2. The average of all the predicted external load values of all the steps in a sequence was used as the external load of the sequence.A positive value of In addition to the estimated external load, desired ic model . To contic model . Desired−3, and the total epochs are 5 × 104. This section employed the same data set as the active load-sensing scheme section for both the training and verification data; however, an extra target stiffness, external load and endoscope diameter value are included in the input data of model, and the desired stiffness of both sets was calculated according to Equation (1). At the same time, we removed bending angle_e from the input data of the model.x-axis (range 0–110 kPa) and antagonist (P_FU) which was the legend by Antag-0, 10, 20 and 30. As shown, when P_FU was constant, the bending angle_o increased as P_FD increased, and the rate of increase of the bending angle_o increase more rapidly from 70 kPa. In contrast, when P_FD was held constant, a greater P_FU reduced the bending angle_o, indicating that the P_FU had a negative influence on the bending angle_o. Although the angle of Antag-30 was greater than that of Antag-0, 10 and 20 at 70–110 kPa in x-axis (range 0–110 kPa) and antagonist (P_FU) which was the legend by Antag-0, 10, 20 and 30 without an endoscope. In addition, various sets of P_FU and P_FD resulted in varied bending angle_o values, which had significant influences on the soft actuator’s internal structure and further affected the stiffness of the soft actuator. The instantaneous load was the output of a NN that was only based on the present state and legend by instantaneous load. The synthesized estimation of external load with the different exploration strategies are the legend on the synthesized load (±n or +n). Ideally, different input sets  of the same external load should estimate almost the same instantaneous load through the trained NN. To confirm the effect of the optimal exploration sequence (±2), the input information of the optimal exploration sequence was concatenated as a total input vector to train another multi-step estimation NN, which means the multi-step estimation NN’s input size was . The single-step estimation NN was same as the instantaneous load. The NN-based inverse dynamics model for stiffness adjustment calculates the required P_FD when the desired stiffness was specified. The results are shown in The experiments were performed on the DoF 1 (composed of FU and an FD chamber) to further investigate the impact of various inflating circumstances on the soft actuator‘s stiffness and bending angle_o. As can be seen from Nonetheless, this inhibitory effect was not permanent and even diminished when the P_FD increased a. Due toMeanwhile, based on the properties of the endoscope, we knew that the endoscope’s stiffness could change by changing the endoscope’s bending angle; in particular, the endoscope’s stiffness could be higher when the endoscope’s bending angle was very larger. The stiffness of the soft actuator also could be changed by changing the bending angle_o as shown in At high P_FD (over 90 kPa), the soft actuator almost exhibited significant stiffness (over 30 N/m) due to the silicon material of the air chamber being massively stretched and rendered impossible to deform under external loads . In FiguTo improve the accuracy of the external load estimation utilizing the active load-sensing system, an environment exploration strategy was presented and employed in a prototype experiment. Moreover, according to On the other hand, multi-step estimation NN was trained use all the information of the optimal exploration sequence as input. The results in The NN-based inverse dynamics model could estimate appropriate P_FD for the desired stiffness, although only for pressures above 35 kPa. However, for pressures below 35 kPa, the model performs poorly, which might because when the P_FD was very low, the stiffness of the actuator varied very little, preventing the model from properly estimating the required P_FD.In the related research , the NN Furthermore, we also proposed an inverse dynamics model for stiffness, which could help surgeons easily adjust a suitable stiffness to resist different external loads during the MIS procedure and keep the surgical view stable. The proposed model could be used for in-depth study of soft actuators’ stiffness or impedance control in the future.The contributions of our study include:In this paper, we proposed a NN-based active load-sensing scheme and stiffness adjustment for a soft actuator to provide a stable surgical view during MIS. To properly estimate the external load and thus determine an appropriate stiffness for the soft actuator, we further optimized the estimation results using an environment exploration strategy.The results demonstrated that the exploration strategy had substantially enhanced the external load estimation accuracy and the successful estimation rate compared to the instantaneous estimation of NNs. The optimal exploration sequence was determined to be synthesized load (±2), with the highest successful estimation rate, smaller exploration range and shorter exploration time. An inverse dynamics model for stiffness adjustment based on a NN was proposed and implemented, with results demonstrating the model’s capacity to precisely calculate the air pressure P_FD for achieving the desired stiffness for the soft actuator.Future studies include expansion of the active load-sensing scheme to different directions of multi-DoFs soft actuators and to design a closed-loop feedback controller for desired angle and stiffness.


Suture or tape augmentation for anterior cruciate ligament (ACL) reconstruction has been described as a technique to increase biomechanical strength and potentially improve clinical outcomes. However, the suture or tape used for augmentation usually requires independent tibial fixation from the ACL graft in the form of an anchor or post. This may introduce the potential for graft and augment tension mismatch, while increasing surgical cost. We present our technique for ACL reconstruction with postless tape augmentation. The ACL graft and tape are fixed at the same tension with interference fixation using a single tibial sheath and screw construct, which allows for ACL augmentation without the need for an additional post or implant. This technique shows the case of an allograft anterior cruciate ligament (ACL) in a right knee with postless tape augmentation. The two ends of a posterior tibialis tendon allograft are each whipstitched with #2 high tensile strength suture . The graft is then loaded onto the femoral cortical button  by folding the graft over once through the suture loop of the button. A 2.5-mm tape suture  is also loaded through the suture loop of the button. Standard anterolateral (AL) and anteromedial (AM) arthroscopic knee portals are created. Diagnostic arthroscopy begins with a 30° arthroscope from a standard knee anterolateral portal. After a femoral socket and a tibial tunnel are created, the femoral button  is then passed into the knee from outside the tibial tunnel, into the joint, and into the femoral tunnel. The femoral button is flipped onto the lateral femoral cortex, the graft along with tape is advanced and docked into the femoral socket using the tensioning mechanism of the button. A sheath and screw  are used for tibial fixation. The surgeon or assistant maintains the knee in 30° of flexion and tensions the two limbs of the allograft, as well as the tape simultaneously, so that the sheath and screw can be inserted in the center between the limbs. First, a dilator is used and advanced into the tibial tunnel, followed by the placement of the appropriately sized sheath. The screw is then advanced through the sheath. It is essential to maintain tension on allograft and tape during all steps of tibial sheath and screw placement. No additional fixation is required as both the graft and the tape are well fixed with the sheath and screw. A clamp placed on the tape at the tibial tunnel aperture shows no creep during range of motion. The graft and tape are fixed at the same location with the same device eliminating concern over potential tension mismatch between the graft and tape. Anterior cruciate ligament (ACL) ruptures are among the most frequent injuries seen by orthopedic surgeons with a reported incidence of 69/100,000 in the United States.6,Several different femoral and tibial fixation methods exist for ACL reconstruction with varying literature supporting different techniques. Biomechanical studies suggest grafts with tibial interference fixation provide better time 0 stability when compared to cortical button fixation.,,To decrease failure risk and enhance protection of the reconstructed ACL, strategies, including ACL graft augmentation with suture or tape, lateral extra-articular tenodesis, and anterolateral ligament reconstruction have been instituted.Standard anterolateral (AL) and anteromedial (AM) arthroscopic knee portals are created. Diagnostic arthroscopy begins with a 30° arthroscope from a standard knee anterolateral portal. Any meniscal pathology is first addressed followed by the anterior cruciate reconstruction. Once the ACL remnant stump is debrided, the femoral tunnel is created. Although this technique can be performed with any femoral drilling technique, we prefer to create the femoral tunnel via anteromedial portal drilling with flexible curved guides . AlthougThis postless tape augmentation technique is performed using suspensory fixation on the femoral side. This technique can be performed with autograft  or allograft  using suspensory fixation. In this case example, the decision was made for an all soft-tissue allograft ACL reconstruction using posterior tibialis tendon. The allograft may be prepared by an assistant concurrently, while the surgeon completes the diagnostic arthroscopy and prepares the femoral and tibial tunnels. A posterior tibialis tendon allograft is first sized to a folded diameter of 9 mm. The two ends of the allograft are then each whipstitched with #2 high tensile strength suture  . The graAfter the ACL tunnels have been created, using a passing suture, the femoral button  is then passed into the knee from outside the tibial tunnel, into the joint, and into the femoral tunnel. The button is then flipped onto the lateral femoral cortex . AfterwaOnce the allograft and tape are docked into the femoral socket, attention is turned toward tibial fixation. A tibial sheath and screw are used for fixation  of both the graft and augment. We prefer to use polyetheretherketone material for the sheath and screw, but bioabsorbable options are also available. Since the allograft  has been folded once over the suture loop of the button, two limbs of the allograft that have previously been whipstitched along with the two limbs of the tape exit the tibial tunnel. We will typically first cycle the graft to remove any creep, and then the surgeon or assistant maintains the knee in 30° of flexion and tensions the two limbs of the allograft as well as the tape. For this case, the two limbs of the graft are separated, while the two limbs of the tape are kept together to create a three-limb construct  where thThe excess lengths from limbs of the graft and tape are cut flush with the tibial surface. Wounds are irrigated and closed, sterile dressings are applied, a hinged knee brace is placed, and a standard postoperative rehabilitation program for ACL reconstruction is pursued. Advantages and disadvantages , as wellVarious surgical modifications have been trialed in the past two decades to minimize ACL graft rerupture, including lateral extra-articular tenodesis,Biomechanically, suture or tape augmentation has been shown to significantly increase the force needed to cause graft failure by increasing load sharing between the graft and brace.While numerous augmentation techniques have been described in ACL reconstruction literature, all rely on either independent suture anchor tibial placement or individually tensioned suspensory fixation.In this technical note, we present a technique that benefits from the protection of tape augmentation while eliminating the potential of tension mismatch between the graft and augment. By using a postless interference fixation with a sheath and screw at the tibial tunnel, the graft and tape can be equally tensioned, avoiding the risks of augment over- or under-constraint. Intraoperative testing of the strength of fixation confirms the integrity of this system as no creep of the tape was found during stress testing. Avoidance of individual or independent graft and augment fixation may provide optimal protection of the young graft prior to incorporation, but more longitudinal research is required to investigate ACL reconstruction augmentation techniques.


This series lists a pictorial quiz pertaining to identification of normal and abnormal anatomical structures and landmarks at a given level on computed tomography (CT). Readers are expected to identify and appreciate the changes from normal anatomy and variations of a given pathology. The main structures assessed in this quiz are the pons, ventricular system of the brain, and the basal cisterns. Particular emphasis is placed on the presentations of intra-cranial haemorrhages, particularly sub-arachnoid and epidural haemorrhages, and masses around the region of the pons, midbrain and cerebellum. There is also a question pertaining to increased intracranial pressure. Differential diagnoses are also given where necessary to guide clinical practice and further learning. A Points to remember section details key clinical pearls. Furthermore, key resources have been cited as recommendations for further reading. It is anticipated that this series will enhance the understanding of sectional anatomy of the brain to aid in brain CT interpretation. AnswersA: Right temporal lobeB: PonsC: Right cerebellar hemisphereth ventricleD: 4E: Left sylvian fissureF: Pre-pontine cistern The pons is a midline structure spanning from below the midbrain and above the medulla. Anterior to the pons is the prepontine cistern, which contains cranial nerves V and VI, and lateral to the pons are the cerebellopontine angle cisterns, which contains cranial nerves VII and VIII. Pathology in these cisterns is usually from haemorrhage or masses which can compress the respective cranial nerves. The fourth ventricle is a chamber of the brain’s ventricular system which lies dorsal to the pons. It transmits CSF from the cerebral aqueduct rostrally to the obex caudally. A pontine mass may flatten the boundary between the fourth ventricle and pons (the floor of the fourth ventricle), typically a brainstem glioma in a child. The basal cisterns are chambers in the subarachnoid space created by a relatively greater distance between the pia and arachnoid maters, allow CSF to pool within these cisterns. Pathology is typically either the cisterns filling with blood from a subarachnoid haemorrhage or effacement of the cisterns can occur secondary to increased intracranial pressure and mass effects.1. Sciacca S, Lynch J, Davagnanam I, Barker R. Midbrain, Pons, and Medulla: Anatomy and Syndromes. RadioGraphics. 2019; 39(4): 1110-25.2. Ryan S, McNicholas M, Eustace SJ. Anatomy for diagnostic imaging e-book. 3rd Ed. New York: Elsevier Health Sciences; 2011.3. Currie S, Kennish S, Flood K. Essential Radiological Anatomy for the MRCS. Cambridge University Press. 2009.4. Moeller T, Reif E. Pocket atlas of sectional anatomy. Computed tomography and magnetic resonance imaging. Vol. 1 Head and neck. Stuttgart: Thieme; 2013.


This study examined the effects of low-temperature storage on the microstructural, absorptive, and digestive properties of cooked rice. Cooked rice was refrigerated and stored at 4 °C for 0.5, 1, 3, 5, and 7 days, as well as frozen and preserved at −20, −40, and −80 °C for 0.5, 1, 3, 5, 7, 14, 21, and 28 days. The results indicated that the stored rice samples generally exhibited a higher absorption capacity for oil, cholesterol, and glucose than the freshly cooked rice. In addition, after storage, the digestibility of the cooked rice declined, namely, the rapidly digestible starch (RDS) content and estimated glycemic index (eGI) decreased, whereas the slowly digestible starch (SDS) and resistant starch (RS) content increased. Moreover, the increment of the storage temperatures or the extension of storage periods led to a lower amylolysis efficiency. Scanning electron microscopy (SEM) analysis indicated that storage temperature and duration could effectively modify the micromorphology of the stored rice samples and their digestion. Moreover, microstructural differences after storage and during simulated intestinal digestion could be correlated to the variations in the absorption capacity and digestibility. The findings from this study will be useful in providing alternative storage procedures to prepare rice products with improved nutritional qualities and functional properties. As an important cereal grain, rice is a main carbohydrate and energy source for humans worldwide, especially for Asians. There are diverse types of rice available on the market that vary in quality. Rice quality is a comprehensive trait that includes the milling quality , the appearance , the cooking and eating quality , and the nutritional quality  ,2. AmongAs starch is the main constituent of rice, the starch digestion property of cooked rice is an important determinant of its nutritional quality . NormallIn recent years, increasing studies have been conducted to investigate rice starch digestibility ,8,9,10. Cooked rice is a metastable and non-equilibrium system that undergoes structural alterations during storage . Upon coBesides the occurrence of structural and compositional changes, the processing of low-temperature storage could also influence the functionality of starchy foods. Noteworthily, due to the richness of polysaccharides and dietary fibre in cereal and cereal fractions, an increasing number of studies have focused on investigating their absorption/binding capacity for glucose, oil/fat, cholesterol, and bile acid ,28,29, iIncreasing demand for convenient foods, coupled with preferences for potential health benefits and the availability of rice in abundance, stimulates the necessity to evaluate the changes that low-temperature storage treatment induces on the absorption capacity and digestibility of cooked rice. In this current study, the starch digestion rates and extents of cooked rice were detected by applying an in vitro-simulated intestinal digestion model. The morphological changes of cooked rice after storage and throughout digestion were determined with scanning electronic microscopy (SEM). The potential capacities of the cooked rice samples to absorb oil, cholesterol, and glucose were investigated. Moreover, the structural features and the absorptive and digestive properties of cooked rice were correlated to elucidate the mechanisms by which the low-temperature storage treatment exerted these effects. The findings of this study will expand the understanding of the microstructural, absorptive, and digestive profiles of low-temperature storage-modified cooked rice and provide valuable insight into the potential applications of low-temperature storage strategies to rice products.Oryza sativa L. cv. Longjing 26) were kindly provided by the Rice Research Institute of Heilongjiang Academy of Agricultural Sciences . The proximate analysis was performed according to AOAC method   , wherein0.549 HI .A scanning electron microscope (SEM) was utilised to visualise the morphological changes of cooked rice after storage and during the in vitro digestion process. The freshly cooked and stored rice samples were collected and pretreated as described in p < 0.05 was considered to indicate a significant difference. The SPSS 26.0 software , Origin Pro 2019 , and R software (R version 3.6.3) were used for statistical computations, mapping, and correlation analysis.All assays were implemented in triplicate and resulting data were expressed as mean values ± standard error. Two-way analysis of variance (ANOVA) followed by Tukey’s HSD test were conducted on means comparison and The cooked rice matrices revealed varying microstructures corresponding to the storage conditions . The freIn the case of cold storage, the rice samples displayed a coarse and discontinuous honeycomb-like structure during the first 3 days, and the pores became larger and more irregular with the extension of the storage time. This phenomenon indicated that the cooked rice had a greater disconnection of hydrogen bonds between the water and starch molecules due to the retrogradation process during cooling storage . As a reDiffering from the refrigerated cooked rice, most frozen cooked rice samples developed porous structures due to the formation of ice crystals during storage and exhibited a marked dependence on the storage temperature: the higher the temperature, the larger the pore size. Likewise, Song et al. revealed that the pore sizes of freeze-dried cooked rice stored at −70 °C were smaller than those of the samples stored at −20 °C and −10 °C . The difIn addition, for the rice samples stored at −20 °C after 14 d or at −40 °C after 21 d, the intensity of the agglomeration and weak porous architecture of the granules were apparent as the storage time increased, probably as a result of the progression of starch retrogradation. This recrystallisation process led to the gradual disruption of the starch granules into small crystallising particles. Consequently, distinctive phase segregation into starch-rich and starch-deficient regions was created due to the lower water diffusion and more flexible starch component mobility . Furtherp < 0.05) higher OAC values than their fresh counterpart, except for several frozen rice samples that exhibited slight increments after storing for 12 h. In addition, the oil absorption capacities of the 4, −20, −40, and −80 °C stored rice samples were in the range of 1.33~1.63, 1.31~1.78, 1.34~1.71, and 1.28~1.62 g/g, respectively, suggesting a greater oil absorption potential for cooked rice samples stored at −20 and −40 °C with respect to those stored at 4 and −80 °C. From the results, the stronger OAC may be due to the porous granule structure and the increased lipophilic tendency after low-temperature storage treatment. According to According to p < 0.05) for the rice samples stored at 4, −20, −40, and −80 °C, respectively. Herein lies the vital influence of storage temperature since higher absorbing performances are shown at 4 and −80 °C, which may be attributed to their greater pore density and aperture ratio. That is, according to the micromorphology described in As observed in p < 0.05) among the stored rice samples when compared with the freshly cooked rice. That is, the low-temperature storage treatment seemed to help to enhance the glucose absorption capacity of cooked rice. Moreover, in agreement with the tendency recorded in the cholesterol absorption, the rice samples stored at 4 and −80 °C were found to have a relatively greater potential for absorbing glucose. This may be attributed to the higher pore density and aperture ratio that appeared on the surfaces of the 4 and −80 °C stored rice samples, which may promote the exposure of more functional groups inside the cooked rice matrix, thus binding and entrapping more free glucose and reducing glucose concentration in the media [The GAC responses of the cooked rice samples are shown in he media .In this present study, for different cooked rice matrixes, their absorption capacity mainly relied on their surface structural characteristics, as well as the nature of the absorbed substances . The greImportantly, from To date, the link between the absorption capacity and microstructural characteristics of cooked rice is rarely described in the literature and no studies have focused on the specific role of low-temperature storage conditions in modulating the absorption capacity. The results from our study demonstrate the significant role of low-temperature storage conditions in affecting the oil, cholesterol, and glucose absorption capacities by regulating the microstructural properties of the cooked rice matrix. Improvements in living standards have been accompanied by an increased intake of fat, cholesterol, and glucose. As such, ways to reduce the excess lipids and glucose in the body have received increasing attention. In this current study, flours from low-temperature stored cooked rice have been confirmed to have potential in their in vitro hypolipidemic and hypoglycaemic effects. This signifies a promising application for these stored rice samples as a functional food to reduce the amount of glucose and lipids available in the intestinal lumen.The hydrolysis time curves obtained for white bread and all cooked rice samples are plotted in R2 ≥ 0.995) indicated that the starch digestion of all the rice samples followed first-order behaviour. The values of C∞ and k provided a good fit for the experimental data. In this context, the stored rice samples displayed evidently lower C∞ and k values compared to their fresh counterpart (p < 0.05), which was consistent with the above results of the hydrolysis profiles. Additionally, the C∞ and k values were found to be negatively related to the storage temperature and duration, and higher storage temperatures/longer storage periods elicited decreased C∞ and k values. In this study, the reduced k values under refrigerated/frozen storage might be mainly attributed to starch retrogradation and impaired digestibility by amylase, which depended strongly on the storage temperature. That is, a lower temperature induces faster retrogradation because nucleation and crystal growth are favoured [As shown in favoured .As shown in On storage, such changes in starch fractions could imply retrogradation occurrence and microstructural alterations, which could affect the susceptibility to enzymatic hydrolysis. Starch retrogradation during storage results in the formation of a perfect crystalline structure that increases the RS content or an imperfect crystalline structure that increases the SDS content . Based op < 0.05), varying between 81.82~91.31 and 84.63~89.84, respectively. Notably, the results also confirmed that a higher storage temperature/longer storage period had a more prominent effect on the eGI of cooked rice, which coincided with the digestion characteristics indicated above.As seen in In food processing, most starch-based products are usually subjected to heating and cooking. Therefore, it is more practical to investigate the GI values of starchy foods after cooking . OverallThe Pearson correlation coefficients for the relationship among the starch digestibility parameters of the stored cooked rice samples are presented in C∞, and k values of cooked rice showed a positive correlation with the RDS content, but a negative correlation with the RS and SDS contents. The results obtained, therefore, allow the inference that the starch fractions of cooked rice were susceptible to storage conditions and might play a role in influencing the glycaemic response and starch susceptibility to amylolysis. Although the findings of this current study are in agreement with several studies in the literature [In our case, cooling and freezing treatments were found to have relatively subtle impact on starch digestibility. As can be seen, the eGI, terature ,54,56, tterature , resultiTo further analyse the impact of low-temperature storage, the microstructural changes of the stored rice samples during simulated in vitro digestion as well as the counterpart fresh sample were examined using SEM . As digeAfter simulated intestinal digestion for 20 min, as shown in After additional digestion of 60 min, hydrolysis occurred and the morphology of the digested rice changed significantly. As seen in As the digestion progressed further, it was found that the residue of the cooked rice samples consisted mainly of much smaller pieces after digestion for 120 min , signifyGenerally, the storage temperature and period, as two important quality-determining factors, are believed to be involved in modulating the microstructural and digestive properties of starchy foods. However, the specific role of low-temperature storage conditions in regulating these properties of cooked rice, and the relationships or interactions between the micromorphology of stored rice samples and their digesta and the in vitro starch digestion rate and extent have rarely been studied so far. Thus, a systematic study of the microstructural characteristics and starch digestibility of cooked rice warrants special concern.In this current study, for all the cooked rice samples, their microstructural properties after storage and during digestion seemed to be closely related to the degree of starch digestion. On the one hand, according to the SEM morphologies before digestion , more amk and eGI values and the higher SDS and RS contents of rice samples that underwent a higher storage temperature or a longer storage period. Collectively, the microstructural features after storage and during the simulated intestinal digestion could explain why the cooked rice samples stored at a higher temperature or for a longer period showed lower digestion rates, as well as equilibrium starch hydrolysis. The findings of this study revealed that low-temperature storage conditions played a significant role in regulating the morphologies of the stored rice samples and their digesta, which could in turn affect the in vitro digestion rate.On the other hand, allied to the enzymatic action and mechanical shaking, the microstructural properties of the digested rice were altered . When thThis study elucidated how low-temperature storage processing influenced the absorption capacity and digestion characteristics of cooked rice samples by inspecting the microstructural changes after storage and throughout the starch digestion process. Through the usage of in vitro models, the results demonstrated that refrigerated and frozen storage temperatures and periods collaboratively modulated the absorption and digestion behaviours of cooked rice samples. Compared to freshly cooked rice, the stored rice samples exhibited lower RDS contents and eGI values, higher SDS and RS contents, as well as stronger absorption capacities for oil, cholesterol, and glucose, which positioned them as noteworthy vehicles for improving human health. Furthermore, the structure–property relationships indicated the significant role of the structural basis in controlling low-temperature storage-induced modifications in the absorption capacities and digestibility of the cooked rice samples. Nevertheless, this is a preliminary study which provides fundamental data for the rational use of low-temperature storage methods to develop rice products with an associated absorption capacity and digestibility. It is desirable that further investigations be conducted to reveal the structure–property relationships in greater detail and explore the potential application of low-temperature storage processing. Firstly, only SEM was used in this current study. More advanced techniques such as confocal laser scanning microscope, micro-CT, etc., should be combined to improve the understanding of how the microstructure of the refrigerated/frozen cooked rice modulates its absorption and digestion performances. Secondly, only the absorption and digestion properties were investigated in the present research. More physicochemical properties can be studied to comprehensively understand the influence of low-temperature storage treatment on cooked rice. Thirdly, since only in vitro models were applied in this paper, in vivo studies are needed to confirm the health benefits and physiological effects of refrigerated/frozen cooked rice. Fourthly, only one rice genotype was evaluated in this current study. More rice varieties could be employed to investigate their responses to low-temperature storage treatment and provide more evidence on the potential application of low-temperature storage processing.


Tetraselmis chui is a well-known starch producer, and was selected in this study for the production of biomass to be explored as an active ingredient in beer brewing. Cultivation was performed under nitrogen deprivation in 250 L tubular photobioreactors, producing a biomass containing 50% starch. The properties of high-starch microalgal biomass in a traditional mashing process were then assessed to identify critical steps and challenges, test the efficiency of fermentable sugar release, and develop a protocol for small-scale brewing trials. Finally, T. chui was successfully integrated at a small scale into the brewing process as an active ingredient, producing microalgae-enriched beer containing up to 20% algal biomass. The addition of microalgae had a noticeable effect on the beer properties, resulting in a product with distinct sensory properties. Regulation of pH proved to be a key parameter in the process.Microalgal biomass is widely studied for its possible application in food and human nutrition due to its multiple potential health benefits, and to address raising sustainability concerns. An interesting field whereby to further explore the application of microalgae is that of beer brewing, due to the capacity of some species to accumulate large amounts of starch under specific growth conditions. The marine species  Tetraselmis chui is particularly interesting for industrial applications in food as its upscaled cultivation is already well established for the production of feed ingredients [Tetraselmis species was proved to be notably rich in antioxidant content and activity [T. chui has been proved to be safe for human consumption [Tetraselmis have been successfully introduced as a novel ingredient in a wide variety of applications, such as bread [T. chui show enhanced bioactive properties and good technical properties, proving the feasibility of microalgal biomass use as an ingredient in staple foods [Microalgae cultivation is one of the emerging technologies in food sciences because of the rich nutritional profiles, impressive health benefits, and positive environmental impact of these photosynthetic organisms ,2. Amongredients . Biomassactivity ,5,6, andactivity . Additiosumption , and hasas bread ,10, soupas bread , gluten-as bread , and savas bread . Productle foods ,14. 2 capture and wastewater bioremediation [With the rapid development of the brewing industry, and of its environmental impact, microalgae are being widely studied for COediation , mainly ediation . HoweverSpirulina has reportedly been used on multiple occasions as a food colouring for seasonal beers, as, for example, in the case of the “Gimmicky Green” beer produced by the Captain Lawrence Brewery in New York. Furthermore, the Dutch microbrewery De Koperen Kat has been crafting an “Algenbier”, adding 5% Chlorella vulgaris biomass specifically cultured for this purpose [C. vulgaris biomass with the Brazilian spirit cachaça, has been tested on mice. The microalgae-enriched beverage was reported to have neuroprotective and antioxidant activity when compared to control cachaça, possibly mitigating the effect of alcohol on neural networks [The cyanobacteria  purpose . Despitenetworks . Tetraselmis species have been widely studied for their potential to produce high starch content under nitrogen deprivation, reaching up to 60% of the total dry weight [T. chui was recently reported to produce up to 59% starch under nitrogen deprivation [Microalgal biomass has the potential to be introduced as an active ingredient in brewing since microalgae are known to be highly efficient starch producers . Under ey weight , and T. rivation .The aim of this study was to test the feasibility of starch-rich microalgal biomass used as an active ingredient in fermented beverages. The effect of different algae-to-malt ratios on brewing protocols and on the final product was assessed by monitoring the release of fermentable sugars during mashing and fermentation phases, and finally, qualifying the beer characteristics and fermentation efficiencies. Providing insights and methods to implement microalgal starch in brewing may pave the way for an increased microalgal presence in industrial setups and in everyday products. Tetraselmis chui SAG 8−6 from the SAG Culture Collection of Algae  was kept on L1 medium agar plates [−2 s−1 irradiation. The algae were scaled up, maintaining exponential growth, to 0.1–0.25–0.5–1 L Erlenmeyer flasks in 2× F medium [−2 s−1 irradiance, on a shaking plate, and supplied with 1% CO2: air mixture diffused through a needle. The contents of the Erlenmeyer flasks were then used to inoculate first a GemTube RD1–25, 25 L tubular photobioreactor  at 25 ± 2 °C, pH 7.8 ± 0.2 (controlled by CO2 addition), and 100 µmol m−2 s−1 irradiance with 2× F medium. The 25 L culture was then finally scaled up to a GemTube RD1−250, 250 L tubular photobioreactor  at 25 ± 2 °C, pH 7.8 ± 0.2 (controlled by CO2 addition), and 100 µmol m−2 s−1 irradiance. r plates  at 22 °CF medium  at 50–10−1 (NO3−), MQuant® .The cultures were monitored daily by measuring absorbance at 750 nm and nitrogen content with nitrate strips 10–500 mg L−1, samples were extracted daily for starch analysis. Once the nitrate dropped below 50 mg Lg with an Evodos 10 centrifuge . The concentrated algal paste (~15–20% dry weight) was frozen at −20 °C and further freeze-dried in a FreeZone freeze dryer . Subsequently, the algae powder was milled in a Planetary Ball Mill PM400  at 400× g rpm, for three cycles of 6 min duration (3 + 3 in each direction). Between cycles, the containers were cooled on ice to prevent biomass overheating by friction. Milled biomass was mixed and aliquoted in 2 mL Eppendorf tubes according to protocols for compositional analyses and experiments.The culture was harvested after a total 8 days of growth, when starch content higher than 40% had been measured. The algae biomass was centrifuged at 3000× Starch content was assessed using the Megazyme Total Starch (AA/AMG) assay kit  on 10 ± 1 mg biomass, following the protocol supplied by the manufacturer, adapted to remove chlorophyll interference, as previously reported . ProteinThe first trials aimed at understanding whether the tightly packed, small, microalgal starch granules would be efficiently degraded by barley starch-degrading enzymes at mashing conditions.−1, thus adding 24 g of milled Pilsner malt  in 100 mL water. The flask was incubated for 1 h in a water bath set at 67 °C, shaking the bottle every 5 min. After incubation, the wort was centrifuged at 4700× g rpm for 5 min, and 1 mL of the enzyme-rich supernatant was pipetted into each of the pre-weighed microalgae and barley malt standard samples. The tubes were then incubated for 0, 1, and 2 h at 67 °C. At each time point, triplicates of each treatment were inactivated at 95 °C for 5 min, then centrifuged at 15,000× g rpm for 5 min. The supernatant was removed, and the pellets were analysed following the protocol described in Nine samples of milled starch-rich algal biomass, and nine samples of barley malt standard , as a control, were prepared, weighing 10 ± 0.2 mg in 2 mL screw-cap tubes. Enzyme-rich wort was prepared separately with a solid:liquid ratio of 0.24 kg LT. chui), barley malt, , and mixtures of microalgae plus barley malt, as shown in −1. Microalgae addition in the mash increased the pH to 8 ± 0.2, thus microalgae plus barley malt mixtures were tested with and without pH adjustment to pH 5.5 ± 0.1 with lactic acid, sampling at 0, 1, 2, 3, and 5 h from incubation. After studying microalgal starch degradation by malt enzymes, mashing trials were carried out with microalgae  for further HPLC quantification of maltose and glucose.At each time point, all triplicates were sampled by aliquoting 1 mL into an Eppendorf tube and incubating at 95 °C in a heating block for 5 min to inactivate all enzymes. The samples were then centrifuged at 15,000× Brewing with three malt:algae ratios was tested at a small scale against a 100% malt control. Each treatment was performed in triplicate, and the 12 beers were prepared over 2 consecutive days. Saccharomyces cerevisiae commercial strain US-05  was added, according to manufacturer’s instructions, and the bottles were closed with yeast locks.For each batch, malt and algae biomasses were weighed  and adde−1) was added to the beers before capping, and the bottles were incubated for 12 days at room temperature, and for 3 weeks at 4 °C, for maturation.The brews were incubated at 20 ± 1 °C in the dark and sampled daily via brix refractometry to evaluate progression of the fermentation (results not shown). Fermentation in both batches stopped after 5 days; however, the batches were bottled on the same date, on day 5 for Beers 3 and 4, and on day 6 for Beers 1 and 2. Upon bottling, samples for HPLC analysis were extracted, and the brews were gently decanted into 500, 330, and 250 mL dark glass bottles, removing most of the sediment. Carbonating sugar  instrument , consisting of a DMA 4500 M density meter, an Alcolyzer Beer ME module, a CarboQC ME module, and a PFD filling device. The instrument was operated with the aid of the Generation M software v2.42 .2SO4 and stored at 4 °C before further analysis. In some instances where sediments occurred in storage over time, samples were centrifuged at 14,800× g rpm for 2 min prior to analysis. Sugar concentrations were determined by HPLC using an Agilent Ultimate 3000  coupled to an ERC RefractoMax 520  refractive index (RI) detector. An organic acid resin column (Rezex ROA-Organic Acid H+ (8%), 300 × 7.8 mm, Phenomenex Inc., Torrance, CA, USA) was used for separation at a temperature of 65 °C and a flow rate of 0.6 mL min−1. As the mobile phase, 5 mM H2SO4 was used. Separation was performed in isocratic mode.All samples were initially diluted to 1:1 with 10 mM Hp < 0.001.Data were analysed using R Studio version 3.6.1 . One-way ANOVA was performed on the data after assessing homogeneity of variances with a Bartlett test. Where outliers were detected, a square root transformation was applied. If the transformation did not remove the effect of the outlier, the significance level to reject the null hypothesis was set at T. chui in a 250 L photobioreactor. After five days of cultivation, most of the nitrogen content was consumed (NO3 < 50 mg L−1), and the monitoring of starch content in the biomass began. After three days of starvation, starch content had quadrupled, reaching 50% of the total dry weight. This is in agreement with previously reported data, which indicate that Tetraselmis species under nitrogen deprivation can accumulate starch up to 62% of the total dry weight [Tetraselmis species grown under optimal conditions can reach up to 30% protein [Tetraselmis marina, which have shown a reduction in fatty acid content under nutrient stress [y weight ,21. The  protein . The lowt stress . Polyunst stress . Howevert stress .T. chui and barley malt, respectively, had been degraded. After two hours incubation, the starch in both treatments had nearly been depleted. Firstly, we tested whether the starch granules produced by the microalgae were efficiently digested by barley starch-degrading enzymes activated during the mashing phase . At the Mashing experiments were then performed on barley malt, microalgae, and microalgae–barley mixtures, with and without pH adjustments. In the 100% malt controls, the sugar release was directly proportional to the amount of malt used in the mashing, for both glucose and maltose a,b.To test the presence and activity of endogenous algal starch-degrading enzymes, a second mashing trial was performed with only starch-rich algal biomass, incubated in suspension at 67 °C for five hours. Although under nitrogen deprivation endogenous starch degrading enzymes may be synthesized in several microalgal species , in thisMeasurement of pH upon mashing with barley malt and algae mixtures highlighted a strong effect of algal biomass, resulting in pH values up to 8.0 ± 0.2. In traditional mashing, the pH is usually between 5.2–5.6; in our experiments, samples containing barley malt only (B1–4) fall within this range. Compared to the barley malt mashing a,b, the The results from the former trials were used to scale the experiment to 1 L bottles, adjusting the pH at the beginning of mashing to 5.5 ± 0.1. Mashing was performed by placing the bottles in a water bath set at 67 °C. The mashes were incubated for one hour after the temperature inside the mashes reached 66 ± 1 °C (45 min from starting point).The high deviations visible in the results a,b are ap < 0.05) differences between treatments were detected for all the parameters presented in 2 volume. Thus, higher content of algae resulted in stronger colour values, haze, and in a higher caloric content. Additionally, pH values were higher in all algae-enriched beers compared to the control, whilst remaining below pH 5. Further testing is needed to better isolate and evaluate the effect of adjustment with lactic acid and improve the pH management protocols.Despite algal biomass not hindering fermentable sugar release during mashing, the results clearly show an effect on the final product. Statistically significant (2 is also consequently lower in high-algae beer; however, no significant difference was evident for this variable.The alcohol content was lower in beers supplied with higher algal content. This is in agreement with the real and apparent degrees of fermentation, where the lowest algal content achieved similar values as the control, whereas high algal addition resulted in lower fermentation efficiency. The volume of COArthrospira platensis biomass was added to the dough, resulting in an enhanced protein profile of the food; however, at this concentration, the sensory profile was not appealing [T. chui biomass with a stabilising effect on dough; however, sensory scores were lower when compared to the 1–2% T. chui substitutions [Microalgal biomass application in food has proven to be a complex process because of its impacts on technical and sensory properties of the final products. For example, in savoury biscuits, up to 8.3% ppealing . Similarppealing . In bakippealing ,32. In gitutions . In our trials, the addition of higher microalgal content was aided firstly by the nature of beer brewing, which happens in suspension, reducing the impact of microalgae on texture. Secondly, some of the unpleasant aromas, which were clearly perceived at the beginning of brewing, were lost throughout the mashing, boiling, and fermentation processes. Finally, a proportion of microalgal biomass sedimented, and was partially removed from the final product during lautering and later bottling, thus possibly reducing the impact on taste and texture. It is also noteworthy that the addition of higher biomass resulted in thicker mashes, with some difficulties in the mixing and major issues in the lautering process, which led to consistent sediment formation. Thus, despite production at a small scale being successful, upscaling of the process is yet to be optimised.The tasting by brewing experts highlighted how the algae-enriched beers had a characteristic flavour compared to the controls, with increasing sourness upon higher microalgal biomass addition. The taste of algae was almost negligible at 5% addition, giving the beer a slight floral flavour, with hints of umami, while the algae taste was more distinct with increasing amounts. The addition of 12.5% had a clear umami taste with seaweed and marine flavours. Additionally hints of a sweet aroma, resembling canned corn, could be perceived. The 20% beer had the most intense profile, with a stronger seaweed flavour and aroma, marked syrupy notes, and strong umami taste. In the 5 and 12.5% beers, appearance was similar to the control, whereas green hues were visible in the 20% beer. These preliminary remarks on the profile of microalgae-enriched beer must be integrated in future studies, with a thorough sensory evaluation performed by a trained tasting panel.Based on this trial, a microalgae content ranging between 5–10% could be the most suitable for commercial purposes. However, testing on a larger scale, with specific brewing equipment is necessary to define in more detail, the colour, texture, and flavour in a final product.Tetraselmis chui biomass was grown under nitrogen-deprived conditions in 250 L reactors achieving the production of biomass with the desired characteristics for brewing. Stepwise trials were performed to assess the effect of microalgae on the mashing process, culminating in the introduction of up to 20% algal biomass as an active ingredient in brewing. Adjustment of pH before mashing proved to be a key step for successful beer brewing with microalgae. In the present study, starch from microalgae was successfully transformed into fermentable sugars during mashing, and further small-scale brewing trials proved the feasibility of microalgae-enriched beer production, yielding a final product that was palatable and had comparatively distinct sensory properties.Further research is required to fully understand the role of microalgal biomass in brewing and its effects on the final product. The assessment of different recipes and brewing conditions may yield important information on the behaviour and impact of microalgae on the technical properties, allowing one to further optimise and scale-up production. Contextually, a thorough sensory evaluation must be performed, in parallel to an analysis of volatile compounds, to understand in more depth the effects of algal biomass on sensory properties. Finally, the presence and role of bioactive compounds, such as antioxidants, in the final product could be tested to evaluate the possible benefits for the consumers and impacts on shelf-life.


Nodular regenerative hyperplasia (NRH) is associated with high morbidity and mortality in patients with common variable immunodeficiency (CVID). While liver biopsy is the gold standard for NRH diagnosis, a non-invasive technique could facilitate early disease recognition, monitoring, and/or immune intervention. We performed a cross-sectional analysis of ultrasound-based transient elastography (TE) in patients with CVID to evaluate liver stiffness and compared this between patients with (N = 12) and without (N = 6) biopsy-proven NRH. Additionally, these data were compared to a cohort followed at our institution for non-alcoholic fatty liver disease (NAFLD) (N = 527), a disease for which TE has routine diagnostic use. Clinical and pathologic features of NRH were evaluated as correlates of liver stiffness, and receiver operating characteristic curves were used to define a liver stiffness cutoff with diagnostic utility for NRH among CVID patients. CVID patients with NRH had a more severe disease presentation compared to those without. This included increased autoinflammatory disease comorbidities, combined B-cell and T-cell dysfunction, and abnormal liver biochemistries . Results of TE demonstrated a significantly elevated liver stiffness in CVID patients with NRH (mean 13.2 ± 6.2 kPa) as compared to both CVID patients without NRH (mean 4.6 ± 0.9 kPa) and non-CVID patients with NAFLD (mean 6.9 ± 5.5 kPa) (p < 0.01). No single or composite histopathologic feature of NRH correlated with liver stiffness including nodule size, nodule density, sinusoidal dilation, fibrosis, and/or lymphocytosis. In contrast, liver stiffness by TE was significantly correlated with clinical parameters of portal hypertension, including an elevated hepatic venous pressure gradient, an increased splenic longitudinal diameter, presence of varices, and presence of peripheral edema. A liver stiffness of greater than or equal to 6.2 kPa was a clinically significant cutoff for NRH in CVID patients. We propose that TE has diagnostic utility in CVID, particularly in the presence of immunophenotypic features such as combined B-cell and T-cell dysfunction, autoinflammatory comorbidities, and/or abnormal liver tests. Elevated liver stiffness by TE should raise suspicion for NRH in patients with CVID and prompt expedited evaluation by hepatology. It compculature , 8. Giveculature , 9, 10. culature .The gold standard for diagnosis of NRH is a liver biopsy. In patients with CVID, NRH is characterized histologically by a nodular pattern of alternating areas of hepatic plate expansion and atrophy. NRH-like changes are often accompanied by other histologic features of CVID such as a sinusoidal lymphocytic infiltrate, mild portal and lobular inflammation, and variably prominent sinusoidal fibrosis . NRH-lik® uses transducer-induced vibrations to create shear waves that move throughout the liver parenchyma. Calculations of the speed of these shear waves can estimate the degree of liver stiffness  , 15. Spetiffness . This aptiffness , 19. Addtiffness , 21. TE tiffness . More retiffness .Given the prevalence of NRH in CVID patients with liver disease, we hypothesized that TE may be a non-invasive imaging tool that has diagnostic utility. In this study, we performed TE among CVID patients and analyzed liver stiffness by kPa, comparing groups with and without biopsy-proven NRH. In addition, we analyzed pathologic and clinical features, such as the severity of portal hypertension, which correlated with the degree of liver stiffness by TE among CVID patients. Finally, we created and analyzed receiver operating characteristic (ROC) curves to define a liver stiffness cutoff by TE with diagnostic utility for NRH among CVID patients.This study was performed at Mass General Brigham under an Institutional Review Board-approved protocol (#2011P000940).Participants were recruited from Massachusetts General Hospital Immunology and Gastroenterology clinics from January 1, 2018, to March 14, 2022. CVID was diagnosed using International Consensus Document (ICON) criteria . In a CV® was performed by a trained ultrasonographer who completed 10 serial measurements with a median score reported. The raw median score in kPa was used for all subsequent analyses. In CVID patients, a ROC curve was generated to define the most accurate diagnostic kPa cutoff for NRH. A cutoff of ≥7.5 kPa was also assessed given its clinical significance in determining elevated liver stiffness among patients diagnosed with NAFLD  and were reviewed by a gastrointestinal (GI) pathologist blinded to the TE data and the initial pathology report. H&E and trichrome-stained slides were reviewed for each case. Histopathology was deemed to be generally consistent, or not, with NRH and additionally was classified based on the following characteristics: the number of nodules identified (N), diameter of the largest nodule (mm), nodule length (mm), nodule density (nodules/10 mm of core length), thickest hepatic plate (N cell layers), sinusoidal dilatation (±), sinusoidal lymphocytosis (±), centrilobular fibrosis, and/or portal fibrosis. Fibrosis was graded as absent, focal, diffuse, bridging, or cirrhosis.Participants’ clinical data were extracted from the electronic medical record. For all CVID participants, this included a review of complete blood count (CBC) with differential and liver biochemistries , total bilirubin, albumin, coagulation factors , and ammonia). Peak ALP was also recorded, included in the absence of other acute causes of elevation . Patient immunophenotype was reviewed including immunoglobulin levels, peripheral flow cytometry including T-cell, B-cell, NK-cell, class-switched memory B-cell, and naïve/memory T-cell counts  closest to the time of TE measurement. T-cell proliferation to mitogens, antigens, and anti-CD3 were also reviewed. Clinical parameters of liver disease were reviewed by the electronic medical recorded diagnosis, as follows: the presence of clinical portal hypertension, varices (grade 1–3), ascites (trace only), and/or peripheral edema. In individuals with transjugular liver biopsies, hepatic venous pressure gradient (HVPG) measurements were evaluated (N = 8). HVPG >5 and >10 mmHg were used to define any portal hypertension and clinically significant portal hypertension, respectively . In CVIDCVID was subcategorized as complicated or uncomplicated. Complicated CVID was defined by the presence of any autoinflammatory clinical feature (described above) or the presence of a combined deficiency immunophenotype .For all data recorded above, the measurement closest to the time of the TE was recorded. The median time from TE to liver biopsy was 359 days , to immunoglobulin level was 162 days (72–408 days), to flow cytometry was 185 days (78–675 days), to liver biochemistry was 39 days (15–147 days), and to CBC was 49 days (28–112 days).Data are represented as means ± SD, median (Q1–Q3), or proportions unless otherwise noted. The relationship between liver stiffness with immunologic and clinical variables was measured using one-way ANOVA with Tukey’s post-hoc correction and simple linear regression. A Mood’s median test compared median liver stiffness measurements. The chi-square test and logistic regression were used to compare categorical data. To account for the small sample size, log transformation was applied to continuous variables and Fisher’s exact test to categorical variables when estimating p-values. If linear regression assumptions were not met after transformation, Spearman’s correlation was performed. Figures were created utilizing GraphPad Prism 9.3.1  or BioRender.com. Statistical analyses were completed with SAS 9.4 ; a two-tailed p-value of <0.05 was considered significant.TE was performed at our single-center institute on 12 CVID patients with NRH, 6 CVID patients without NRH, and 527 non-CVID patients with NAFLD. The median age of all participants was 55  years, and 55.7% were female. There were no significant differences in age, race, and sex in the subgroups of CVID with NRH, CVID without NRH, and non-CVID with NAFLD. Within CVID subgroups, there was no significant difference between age at diagnosis and time from diagnosis to completion of liver stiffness measurement , presence of grade 1–3 varices , and presence of peripheral edema . Additionally, liver stiffness was higher among CVID patients with diagnosed portal hypertension, either clinically  or by increased HVPG  Table 1.Given the higher prevalence of complicated CVID in patients with versus without NRH who underwent TE, we evaluated if other clinical parameters correlated with the measure of liver stiffness in this patient demographic. The presence of CVID-related autoinflammatory disease was not associated with higher liver stiffness by TE. Specifically, there was no difference in a mean or median liver stiffness between CVID patients with GLILD, lymphadenopathy, enteropathy, or cytopenias. Additionally, liver stiffness values did not differ based on the history of GI infection or type of replacement immunoglobulin received Table 1.Previously, an ALP level in peripheral blood >1.5 times the upper limit of normal was suggested as a useful marker in the diagnostic workup of NRH for CVID patients, specifically to prompt further liver biopsy . We sougFinally, we analyzed peripheral blood immunophenotypes and liver biochemistries, comparing the liver stiffness cutoffs of 6.2 kPa (defined by our ROC above) and 7.5 kPa [a threshold for increased liver stiffness previously used in NAFLD ] TableTable3.This study demonstrates the utility of TE in diagnosing NRH among patients with CVID. While prior investigations have analyzed the relationship between TE and liver disease in CVID, this is the first study to our knowledge to do so in those with biopsy-confirmed NRH.It has been established that liver stiffness measurements are elevated in CVID-related liver disease. Crescenzi et al. previously investigated TE in CVID patients with liver disease demonstrating a mean liver stiffness of 7.5 kPa, with 75% of participants meeting the definition of complicated CVID used in this study. In the absence of enteropathy or polyclonal lymphoproliferation, the mean liver stiffness appeared to be below 6.2 kPa . Prior sIn this study, we identified significantly elevated liver stiffness by TE in CVID patients with NRH compared to both CVID patients without NRH and non-CVID patients with NAFLD. Interestingly, we did not find a statistically significant association between histopathologic features of NRH on liver biopsy and liver stiffness by TE in our CVID patient demographic. This is consistent with other studies of NRH in non-CVID populations, where liver stiffness by TE has not been associated with specific histopathologic features . This laThere are several limitations and sources of bias to consider in this study. While all eligible CVID participants with NRH were recruited, not all of those without NRH participated in the study. This may introduce selection bias in that CVID patients without NRH who consented to be included may be different from those who did not. This study is also cross-sectional in nature, and liver stiffness levels were not followed up over time, limiting causal inference. Furthermore, the study population was relatively small with recruitment from an academic tertiary care center, limiting statistical power and adding to potential selection bias or increasing type I error. Given the rarity of CVID, though, this is not unexpected and is a difficult limitation to overcome. It is important to note that four participants had splenectomy before this study, and thus, splenic diameter measurements were unavailable. As those with prior splenectomy likely had severe disease, censoring these patients in our analysis would have biased results towards the null, minimizing an association. While we did incorporate a comparison between liver stiffness by TE in those with CVID and NAFLD, mean liver stiffness measurements in NAFLD participants were below the established threshold of clinically significant fibrosis. We were unable to correlate liver stiffness measurements in those with NAFLD to the extent of fibrosis on biopsy given our data set. The influence of steatosis and other pathologic processes unrelated to NRH on liver stiffness measurements is difficult to measure and must be acknowledged as well. We attempted to correlate CAP as a surrogate of hepatic steatosis to liver stiffness and markers of portal hypertension, and while there did seem to be a positive correlation, this was not statistically significant, which we believe was due to the small sample size. Finally, most participants with NRH in this study had severe disease, with evident portal hypertension. It is therefore difficult to make conclusions related to the utility of TE as a predictor of early NRH. Future studies investigating the impact of early NRH diagnosis and the impact of underlying complicated CVID features in those without liver disease will be of great importance.The current standard of care in patients with CVID and NRH suffers from diagnostic delays and treatments that carry high morbidity and mortality  , 11. TE The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.This study was performed at Mass General Brigham under an Institutional Review Board-approved protocol (#2011P000940). The patients/participants provided their written informed consent to participate in this study.DD wrote the manuscript and performed the chart review and statistical analyses on the cohort. JS, PB, and KC provided patients from the gastroenterology clinic, as well as clinical expertise. RC reviewed liver biopsy histopathology and provided expertise in the creation of biopsy scoring for NRH. NY performed the chart review. JF conceived the project and supervised all aspects of the work. SB and JF provided expertise in immunology. All authors reviewed the manuscript, approved the final manuscript as submitted, and agree to be accountable for all aspects of the work.SB is supported by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health under Award Number K23AI163350.The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.JF holds investigator-initiated grants from Bristol Myers Squibb and Pfizer with no direct relation to the work presented.The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.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.


Statistical analysis was performed using one-way ANOVA and Tukey’s post hoc test at α = 0.05. All groups showed a predominant tetragonal phase, with a 450 nm average grain size. There was no significant difference between groups with regards to density, porosity, and microhardness (p > 0.05). The tilted group had the highest surface roughness (0.688 ± 0.080 µm), significantly different from the milled (p = 0.012). The horizontal group presented the highest contact angle (89.11 ± 5.22°), significantly different from the milled and tilted (p > 0.05). The BFS of the milled group (1507.27 ± 340.10 MPa) was significantly higher than all other groups (p < 0.01), while vertical and tilted had a similar BFS that was significantly lower than horizontal (p < 0.005). The highest and lowest Weibull modulus were seen with tilted and milled, respectively. Physical properties of all groups were comparable. The surface roughness of the tilted group was higher than milled. The horizontal group had the highest hydrophobicity. Printing orientations influenced the flexural strength of 3D-printed zirconia. Clinical implications: This study demonstrates how the printing orientation affects the physiomechanical characteristics of printed zirconia.The objective of this study is to examine the physiomechanical and surface properties of 3D-printed zirconia in comparison to milled zirconia. A total of 80 disc-shaped (14 × 1.5 ± 0.2 mm) specimens ) were manufactured from 3-mol% yttria-stabilized tetragonal zirconia. Five specimens per group were evaluated for crystalline phase, grain size, density, porosity, surface roughness, wettability, microhardness, and SEM analysis of the surface. Biaxial flexural strength (BFS) was measured ( Dental (3Y-TZP) . Y-TZP i(3Y-TZP) ,5.The increased demands for these new ceramic restorations over the past years resulted in a great advancement of the existing systems or the introduction of new restorative techniques . CeramicTo overcome the drawback of SM, additive manufacturing (AM) has been rapidly developing. AM offers multiple advantages in comparison to SM techniques. It involves building an object with consecutive thin layers that are either cured as they are built or receive post-layering curing. This approach of manufacturing uses CAD software and a 3D printer which are essential elements in the viewing of planned objects in a virtual environment and then producing the actual object ,12.The 3D-printing technology has brought many advancements into the dental field by improving the processing, and increasing the dental applications of polymers, ceramics, and metals within the dental treatment options ,14,15. SFurthermore, the mechanical and surface properties of materials produced by 3D printing is said to be affected by the printing technique ,21 and pExamining the microstructure and basic physical properties of these 3D-printed zirconia materials will enable us to determine the dependability and trueness of fabricating zirconia crowns using the AM process. Despite the reported potential success of the AM technique for ceramics, it is still in its initial stage of development. The purpose of this study was to evaluate the mechanical, physical, and surface properties of 3D-printed yttria–partially stabilized zirconia material and compare the results with those of milled zirconia specimens. The null hypotheses of the study are as follows: First, the different printing orientations will not affect the tested properties; crystalline structure, grain size, density, porosity, surface roughness, wettability, microhardness, and biaxial flexural strength (BFS) of 3D-printed zirconia. Second, 3D-printed zirconia specimens would have comparable results to those of milled specimens.https://clincalc.com/stats/samplesize.aspx, accessed on 7 September 2021). However, the number of specimens dedicated for BFS was increased to 15 to allow for Weibull analysis later as recommended by ISO standard #6872 [n = 5). All specimens were manufactured from 3 mol% yttria–partially stabilized tetragonal zirconia polycrystals (3Y-TZP), Based on the effect size detected between AM and SM zirconia ,32 and trd #6872 . AdditioThe specimens were milled in the green stage and therefore were sintered according to manufacturer’s directions. For AM specimens, the same STL file was sent to a commercial supplier to 3D print and sinter the specimens. The 3D-printed discs were produced out of 3Y-TZP paste  using a 3D printer , The specimens were tested as received from the manufacturer with no further modification except polishing of the specimens undergoing BFS. These specimens were polished using 1200-grit silicon carbide paper  followed by 3 μm polishing suspension  combined with a polishing cloth and machine . Polishing of all specimens was completed by a single operator and the dimensions of the specimens were confirmed after polishing using an electronic digital caliper . The specimens used to test the physical and surface properties were cleaned in an ultrasonic bath and air dried for 24 h between tests, and each test was completed by a single examiner.n = 2) were scanned between 2θ = 20–90° at a scan rate of 0.5°/min. The interplanar spacing was calculated using Bragg’s law while the crystallite size was found using Scherrer’s formula.These properties were measured using X-ray diffraction (XRD) ). The samples (n = 5) were dried in an oven at 100 °C for 1 h and then weighed individually using an analytical digital lab balance  with 0.001 g accuracy. Following that, the weight was recorded for the specimen after suspending in distilled water at 23 °C. Density was calculated using the following Formula (1):Density was determined using the Archimedes method ,38, wherD is the density of the specimen (g/cm3); m1 is the mass of dry specimen (g); m2 is the mass of the suspended specimen; and wD is the density of the immersion liquid. To calculate the apparent porosities (n = 5), the following Formula (2) was used [P is the apparent porosity (%) and m3 is the mass of the soaked specimen (g).was used :(2)P=m3−a) (n = 5) were undertaken at 5 separate locations, 1 at the center and 4 circumferentially on each specimen using a non-contact optical profilometer . The Ra value was averaged per specimen then per group [Measurements of the surface roughness (Rer group .n = 5). Measurements of the contact angles were performed at 20 °C using a goniometer  with a video camera. A drop of distilled water (10 µL) was generated on the sample’s surface using a pipette. Following that, images of the water drop were captured 15 s after the application and interpreted using software . The process was repeated three times for each specimen, then the values were averaged to calculate the contact angle per specimen and group [The wettability was determined using the sessile drop method  (n = 5).nd group .n = 5). Five indentations were made circumferentially in each disc using 9.81 N for 15 s. The average hardness value per specimen was calculated followed by averaging to obtain the group hardness value [Vickers indenter  was used to measure the microhardness  were individually placed on a supporting jig made of three hardened steel balls equally spaced on the circumference of a circle (120°). A universal testing machine  equipped with 5 KN load cell applied the load at the center of the specimen using a 1.5 mm flat-end piston, at a crosshead speed of 0.5 mm/min until specimen fracture. A piece of rubber spacer (thickness 0.2 mm) was placed between the specimen and the load. The load at fracture was recorded per specimen and the BFS was calculated using the following Formula (3) [σ is maximum flexural strength (MPa), P is maximum load causing fracture in (N),b is the thickness of specimen at fracture, all in mm. The specimens (mula (3) : (3)σ=−0n = 2) and fractured specimens (n = 2) was studied using scanning electron microscopy (SEM) . The samples were mounted onto an SEM stub using double-sided carbon tape. The SEM was operated at 20 kV and images of the gold-coated specimens were taken at multiple magnifications to highlight the various features and to disclose the grains of the specimens. However, representative images were displayed at magnifications of 500, 5000, and 20,000× for the as-received specimens, while fractured specimens were shown at 200 and 1000×.The surface morphology of the intact specimens  was used to perform all statistical analyses. Means and standard deviations were calculated. Following that, appropriate tests were used. A one-way ANOVA per tested property was run and when significant, Tukey’s post hoc test was employed to study pairwise comparisons. The level of significance was set at 0.05 for all tests.The mean values ± SD, of the tested properties  are summarized in According to the XRD analysis, F = 0.333, p = 0.802). The highest and lowest densities were reported with milled (6.056 ± 0.116 g/cm3) and tilted (5.942 ± 0.266 g/cm3), respectively. The apparent porosity of the specimens ranged between 0.923 ± 0.591% and 1.945 ± 1.509% for milled and tilted with no significant difference between the groups .The densities of the specimens in the four groups were not significantly different from each other . The tilted group had the highest Ra (0.688 ± 0.080 µm), significantly different (p = 0.017) from milled (0.542 ± 0.087 µm). However, no difference was detected between the milled and the other two printed groups (p > 0.05), The highest and lowest surface roughness values were reported with tilted and milled, respectively. The surface roughness of the specimens varied significantly . Pair-wise comparisons using Tukey’s post hoc test revealed a significantly higher contact angle with horizontal (89.11 ± 5.22°) than milled (p = 0.011) and tilted (p = 0.047), but not vertical (75.34 ± 9.24°). The vertical group was not different from milled or tilted either (p > 0.05), (The ANOVA results showed that the contact angles (wettability) of the specimens differed significantly between the groups , .F = 2.556, p = 0.092). The highest hardness value was reported for horizontal (1676.61 ± 37.77 VHN1) and the lowest for milled (1548.2 ± 62.32 VHN1). With regards to surface hardness, all groups showed similar hardness values with no significant difference , and the post hoc test revealed significant differences between milled vs. all other groups (p < 0.01), horizontal vs. all other groups (p < 0.01), while vertical and tilted were not significantly different from each other (p = 0.11). For BFS, the milled and vertical groups showed the highest and lowest values at 1507.27 ± 340.10 MPa and 521.51 ± 88.76 MPa, respectively. The ANOVA results indicated a significant difference among the groups  and the lowest for milled (5.1438), with vertical and horizontal ranking second and third, respectively, The SEM images of the as-manufactured specimens from each group are displayed in The higher magnification (5000×) of printed specimens’ surfaces showed clustering of zirconia grains into clumps in addition to the presence of few voids on the surface that ranged between 2 and 10 microns across (see black arrows). After further magnification , the milled specimens showed a dense structure with no voids and slightly polygonal grains in the range of 270–760 nm and an average of 450 nm. Smaller voids are detected in printed specimens (Voids are marked with black solid arrows. Crack lines are marked with a hollowed arrow.Images of fractured specimens at 200× and 1000× are shown in Comparisons of the available techniques (SM and AM) to fabricate zirconia restorations, the different printing orientations, and the determination of possible clinical uses and limitations are difficult, due to the limited publications on this topic. This study inspected and analyzed surface characteristics along with physical and mechanical properties of zirconia ceramic (3Y-TZP) manufactured by two different techniques: subtractive and additive. The results of the study suggested partial rejection of the first hypothesis with regards to contact angle and BFS. The second hypothesis was also partially rejected for roughness, contact angle, and BFS. Since the future is advancing rapidly in the field of 3D printing of restorations, soon, this technology will be available widely for ceramic restoration on a commercial level. Therefore, this study aimed to characterize one of the currently available zirconia materials and test a number of its properties that are relevant to the oral environment. According to the XRD analysis, the present study demonstrated that all zirconia groups (SM and AM) had comparable phase composition and pattern with tetragonal phase forming the bulk of the material, similar to what was reported by Nakai et al. . HoweverAll groups presented a grain size in the range of 240–920 nm with average of 420–460 nm. Although all printed groups received similar manufacturing steps, debinding and sintering temperatures, a slight difference in grain size was noted. These findings were aligned with Hofer et al. , who indThe surface porosities/voids seen in all printed groups might be linked to the manufacturing technique involving layering the specimen in thin sections which could entrap air during the process. This finding is inconsistent with Hofer et al. , who rep3 [Density is an important factor that influences the mechanical and optical properties of the material ,42. The 3 ,44, the 3 . The app3  suggesteThe aggregation of the nano-sized grains to form clumps has been seen in SEM images of AM specimens. This has been reported earlier by Li et al. , who relThe wettability of the surface has been linked to the surface topography and charge. Additionally, greater cell adhesion is linked to higher surface wettability and surface energy . The resThe hardness of a material represents its ability to resist plastic deformation upon indentation force . In our Since the current investigations of the 3D-printed zirconia will lay the foundation for future advancements and improvements to the material or technique before full adoption into the production streamline, it is very important to test the flexural properties of the 3D-printed material in different printing orientations and compare the results to those of SM. Accordingly, milled and vertical groups had the highest and lowest BFS, respectively. Those findings are consistent with previous reports . The higWeibull analysis was used to analyze the consistency of the tested specimen. The higher the value, the less variation is present between specimens . The higAt a clinical level, the produced restoration/crown must fulfil the mechanical and biological requirements as well as have good esthetics. According to the results of this study, the clinician must consider the most important feature/property of the restoration to match it with the appropriate printing orientation. If strength was the prime concern, a horizontally printed restoration is recommended, while it should be avoided if high wettability was the top interest of the dentist. However, there are other aspects of a material that are also considered prime for its success but could not be tested in this study. For example, the restoration must be accurate and precisely duplicate the original design in order to minimize the need for chair-side corrections. Alharbi et al.  and OsmaThe study aimed to characterize AM zirconia specimens printed at three different orientations and compare the results to those of SM zirconia. The findings play an important role in determining the most appropriate printing orientation for favorable mechanical and surface properties of the final prosthesis. However, among the encountered limitations is the use of only one type of zirconia per manufacturing technique, the limited number of tests, the use of standardized specimens that do not represent actual restorations, the absence of aging, in addition to the limited published papers involving different orientations of AM zirconia, which made comparisons with previous reports more difficult. Based on that, future studies are encouraged to test more zirconia brands, include more printing orientations, test specimens with a geometry closer to a dental restoration, test aspects related to color and translucency, evaluate microbial adhesion, test the strength of bonding to ceramic, and involve the specimens in aging and wear processes, in addition to the evaluation of the trueness/accuracy of different printing orientations. The 3D-printing orientation had an influence on the wettability and biaxial flexural strength of 3D-printed zirconia. However, all 3D-printed specimens achieved the minimum required strength for single (monolithic or substructure) restoration involving molars as recommended by ISO. Physical properties , and microhardness of all groups were comparable to each other. The roughness value of tilted specimens was higher than milled, while the wettability of horizontally printed specimens was the lowest.


Scanning laser Doppler vibrometry is a widely adopted method to measure the full-field out-of-plane vibrational response of materials in view of detecting defects or estimating stiffness parameters. Recent technological developments have led to performant 3D scanning laser Doppler vibrometers, which give access to both out-of-plane and in-plane vibrational velocity components. In the present study, the effect of using (i) the in-plane component; (ii) the out-of-plane component; and (iii) both the in-plane and out-of-plane components of the recorded vibration velocity on the inverse determination of the stiffness parameters is studied. Input data were gathered from a series of numerical simulations using a finite element model (COMSOL), as well as from broadband experimental measurements by means of a 3D infrared scanning laser Doppler vibrometer. Various materials were studied, including carbon epoxy composite and wood materials. The full-field vibrational velocity response is converted to the frequency-wavenumber domain by means of Fourier transform, from which complex wavenumbers are extracted using the matrix pencil decomposition method. To infer the orthotropic elastic stiffness tensor, an inversion procedure is developed by coupling the semi-analytical finite element (SAFE) as a forward method to the particle swarm optimizer. It is shown that accounting for the in-plane velocity component leads to a more accurate and robust determination of the orthotropic elastic stiffness parameters. The identification of elastic stiffness parameters is fundamental for structural design and health monitoring. Design recommended stiffness parameters might change over time due to cyclic thermal or mechanical exposure, or as the result of an abrupt impact event. Therefore, the implementation of fast and accurate characterization methods is essential in non-destructive evaluation (NDE) and structural health monitoring (SHM) . In viewIn vibration-based methods, the specimen is excited via a shaker or a transducer, and the frequency response at different points on the surface is measured. The measured frequency response is then used to extract modal parameters, such as natural frequencies, modal damping and mode shapes, which are analyzed to identify the stiffness parameters ,4. HowevIn wave propagation methods, a pair of transducers consisting of an emitting source and a receiving sensor is used to send and record an ultrasonic wave. The propagation behavior of the generated wave obviously depends on the amplitude and frequency content of the excitation signal, as well as on the geometrical, structural and elastic properties of the medium. In recent decades, a range of forward models were developed in the literature to predict wave propagation in various media and geometries, and to inversely estimate the genuine stiffness parameters. Among wave propagation-based characterization methods, the use of bulk waves in through-transmission ,6,7 or Ld) because of the infinite plate assumption, placing a lower boundary on the frequency range. Additionally, damping effects and fundamental homogenization conditions put an upper boundary for the “frequency × thickness” (fd) regime. Secondly, the effects of plate boundaries are neglected due to the inherent infinite thickness assumption which might lead to discontinuous phase shifts in transmitted signals. Even though bulk waves are independent of the frequency and plate thickness, abrupt layer transitions at interfaces lead to phase changes that need to be accounted for in calculating the time-of-flight as well as in the inversion of the stiffness constants terature , and theterature . It shoufd) range. Particle swarm optimization (PSO) was chosen for the inversion because of its speed and accuracy. The expected stiffness parameters are calculated by minimizing the absolute percentage error between the forward calculated (SAFE) wavenumbers and the wavenumbers deduced from actual or numerical wave propagation datasets using the matrix pencil decomposition method (MPDM). To obtain statistics on the results, the inversion procedure is repeated 20 times taking into account the heuristic nature of the optimization algorithm. The procedure is validated by using a series of COMSOL simulations and actual experiments on different materials. The numerical and experimental results show that it is possible to identify all nine stiffness parameters with an average error of less than 5%. Additionally, the study revealed that the identification of the out-of-plane stiffness  method as a forward model to estimate the characteristic Lamb wavenumber–frequency pairs in a pre-set frequency–thickness (


DOI: 10.1039/C6RA06210E.Retraction of ‘Enhanced electrocatalytic activity and durability of highly monodisperse Pt@PPy–PANI nanocomposites as a novel catalyst for the electro-oxidation of methanol’ by Özlem Karatepe  RSC Advances article due to concerns with the reliability of the data in the published article.The Royal Society of Chemistry hereby wholly retracts this International Journal of Hydrogen Energy article,1 and in Fig. 2 in a Journal of Cluster Science article,2 by the same author group representing different nanoparticles or synthetic methods. Fig. 1 in the International Journal of Hydrogen Energy article1 represents Pt(0)/DPA@GO NPs and Fig. 2 in the Journal of Cluster Science article2 represents Pt(0) NPs. The authors claim that this was a mistake and provided replacement data for consideration. However, an expert reviewed the author’s response and concluded that it did not satisfactorily address the concerns, and that the replacement figure did not fully support the conclusions. Given the significance of the concerns about the validity of the data, the findings presented in this paper are no longer reliable.The two high resolution transmission electron micrograph insets in Fig. 2 that represent Pt@PPy–PANI NPs polymer composites are identical. In addition, these insets are duplicated and scaled versions of the high-resolution transmission electron micrograph insets in Fig. 1 in an Sinan Eriş and Fatih Sen oppose this retraction. Handan Pamuk, Yunus Yıldız, Özlem Karatepe and Zeynep Dasdelen were contacted but did not respond.RSC AdvancesSigned: Laura Fisher, Executive Editor, rd September 2021Date: 23