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Freeze-induced stress causing aggregation of proteins has typically been primarily attributed to the ice-water interface. However, we hypothesize that the underlying observed and perceived detrimental effect of ice is, to some extent, attributed to air bubbles expelled from ice crystal lattices or to nanobubbles existing prior to freezing. The reduction of dissolved air was achieved via a deaeration process by placing samples in a reduced pressure chamber, while the reduction of nanobubbles was achieved by filtering samples via a syringe filter. The results showed that the reduction of both dissolved air molecules and stable colloidal nanobubbles in a bovine IgG solution prior to freezing led to a significant decrease in aggregation after thawing compared to untreated samples (∼6,000 vs. ∼ 40,000 particles/mL at a freezing rate of 100 K/s, respectively). The deaeration-filtration treatment works additively with cryoprotectants such as trehalose, further reducing the freeze-induced aggregation of IgG. The results also demonstrated that air-water interfacial aggregation of IgG in bulk liquid samples is a time-dependent process. The number of IgG subvisible particles increased with time and temperature, suggesting that random collisions of denatured molecules promoted the formation of aggregates with spherical morphology. In contrast, the IgG subvisible count after freeze-thawing had already reached its nominal value, suggesting a time-independent process where denatured protein molecules were compressed between ice crystals into filament-like aggregates. In summary, the findings shift the current paradigm from ice crystals being the main destabilizing factor during freezing to air bubbles, although the two are intertwined. From a translational aspect, this study underscores the value of deaeration-filtration as an essential supplemental process that can be applied in addition to formulation approaches such as the use of cryoprotectants to further reduce freezing stress on proteins and increase their stability., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: [Robert O. Williams III reports financial support was provided by TFF Pharmaceuticals Inc. Robert O. Williams III reports a relationship with TFF Pharmaceuticals Inc. that includes: consulting or advisory, equity or stocks, and funding grants. Zhengrong Cui reports that financial support was provided by TFF Pharmaceuticals Inc. Zhengrong Cui reports a relationship with TFF Pharmaceuticals Inc. that includes: equity or stocks and funding grants. Z Cui serves as the Editor of the International Journal of Pharmaceutics for North America. This paper was subject to all the Journal’s usual procedures. The peer review was handled, and the editorial decision was made independently of ZC and his research group.]., (Copyright © 2024. Published by Elsevier B.V.)

Cryopreservation is important in manufacturing of cell therapy products, influencing their safety and effectiveness. During freezing and thawing, intracellular events such as dehydration and ice formation can impact cell viability. In this study, the impact of controlling the ice nucleation temperature on intracellular events and viability were investigated. A model T cell line, Jurkat cells, were evaluated in commercially relevant cryoformulations (2.5 and 5 % v/v DMSO in Plasma-Lyte A) using a cryomicroscopic setup to monitor the dynamic changes cells go through during freeze-thaw as well as a controlled rate freezer to study bulk freeze-thaw. The equilibrium freezing temperatures of the studied formulations and a DMSO/Plasma-Lyte A liquidus curve were determined using DSC. The cryomicroscopic studies revealed that an ice nucleation temperature of -6°C, close to the equilibrium freezing temperatures of cryoformulations, led to more intracellular dehydration and less intracellular ice formation during freezing compared to either a lower ice nucleation temperature (-10 °C) or uncontrolled ice nucleation. The cell membrane integrity and post thaw viability in bulk cryopreservation consistently demonstrated the advantage of the higher ice nucleation temperature, and the correlation between the cellular events and cell viability., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Xiuling Lu reports financial support was provided by University of Connecticut. Xiuling Lu reports a relationship with International Journal of Pharmaceutics that includes: board membership. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Ice nucleation processes in the earth's atmosphere are critical for cloud formation, radiation, precipitation, and climate change. We investigated the physicochemical properties and ice nucleation potential of selected viral aerosols, including their RNA and proteins, using advanced techniques such as scanning-transmission electron microscopy (S/TEM), small angle X-ray scattering (SAXS), particle analyzers, and a peltier chamber. The experiments revealed that RNA particles obtained from MS2 bacteriophage had a mean freezing point of -13.9 ± 0.3 °C, comparable to the average ice nucleation temperature of global dust particles, which is approximatively -15 °C. RNA from MS2, Influenza, SARS-CoV-1 and SARS-CoV-2 demonstrated average ice nucleation temperatures of -13.9 ± 0.3 °C, -13.7 ± 0.3 °C, -13.7 ± 0.3 °C, and -15.9 ± 0.4 °C, respectively. SAXS analysis indicated a high local crystallinity value of 0.5 of MS2 RNA particles, hinting that high crystalline nature may contribute to their effectiveness as ice nuclei. Dilution experiments show that viral RNA consistently catalyzes ice nucleation. The addition of dust-containing particles, such as Fe 2 O 3 , CuO, and TiO 2 , to MS2 bacteriophage droplets enhanced ice nucleation, as did UV radiation. We herein discuss the implications of this work on ice nucleation and freezing processes.

The migration characteristics of nanoplastics (NPs) in the natural freezing process are complex and have attracted increasing attention in simulating natural freezing in recent years. However, simulated freezing conditions often fall short of replicating natural freezing processes, and studies on the vertical distribution of NPs remain inadequate. This study established a more realistic simulation of the natural freezing process in surface water by controlling both the air temperature ( T 1 ) and the water temperature ( T 2 ). Additionally, we introduced a new parameter, the local distribution coefficient ( K iw1 ), to compare with the effective distribution coefficient ( K iw2 = 1 °C. The results revealed the NPs concentration differed in ice, near-ice liquid, and far-ice liquid. Both properties of NPs and environmental factors could regulate the vertical distribution of NPs. The findings underscored the importance of freezing temperature regulated by K iw1 , elucidating the roles of various influencing factors on the vertical distribution characteristics of NPs and unraveling the mechanisms of NPs distribution in the ice-water system. This study can provide valuable insights for understanding the migration of NPs in surface water in cold regions.K iw2 for PS-500 nm were 0.18 and 0.21, respectively, at T 1 = -20 °C and T 2 = 1 °C. The results revealed the NPs concentration differed in ice, near-ice liquid, and far-ice liquid. Both properties of NPs and environmental factors could regulate the vertical distribution of NPs. The findings underscored the importance of freezing temperature regulated by T 1 and T 2 , elucidating the roles of various influencing factors on the vertical distribution characteristics of NPs and unraveling the mechanisms of NPs distribution in the ice-water system. This study can provide valuable insights for understanding the migration of NPs in surface water in cold regions.

In this study, we calculated the travel times of a thermal probe that descends through Europa's ice shell. The ice column is simplified to a conductive layer. Using a cellular automaton model, the descent of the probe was simulated by tracking temperature changes, with cell interaction dictated by heat conduction and cell state transition rules determined by cell temperatures. Validation tests, including a soil column simulation, and comparison with experimental data, support the reliability of the model. Simulations were performed with 2 different cell sizes, 19 constant probe temperatures, and 5 ice thermal conductivities. A smaller cell size ( Δ z = 3   mm) produced shorter travel times (between 22 days for a probe temperature T p = 600 K and ∼4 years for T p = 280 K ) than a larger cell size ( Δ z = 1   m), which produced travel times between 27 years ( T p = 600K) and ∼10 3 years ( T p = 280K). The ice shell's thermal conductivity has a modest impact on descent times. The results are generally consistent with previous approaches that used more detailed probe engineering considerations. These results suggest that a probe relying solely on heat production may traverse Europa's conductive ice shell within a mission's timeframe.

Background: The formation of ice crystals will have adverse effects on aquatic products, and the key to ensure the long-term preservation and better quality preservations of the product is to evaluate the intercellular ice crystal formation to find suitable refrigeration conditions and cryoprotectants., Results: The ice crystal formation was successfully captured by using an inverted microscope cryomicroscopic system equipped with a low-temperature stage, the ice crystals formed under different freezing methods between tuna muscle cells were observed directly, the deformation degree of muscle tissue pores during crystallization was evaluated, and the effect of freeze-thaw times on tuna samples was analyzed. The effects of the use of cryoprotectant such as cellobiose and carboxylated cellulose nanofibers on ice-growth inhibition were investigated, and the reliability of the ice crystal observation results was further verified by the determination of physical properties. The results showed that carboxylated cellulose nanofibers had the best ice-growth inhibition effect, they prevented about 50% cell deformation compared with the control group, and also reduced the minimum size of ice crystal formation. In addition, the addition of cellobiose and sodium tripolyphosphate gave the ice crystals a more uniform size and roundness., Conclusion: The experiment proposed a stable and clear observation method for the process of intercellular ice crystal formation, and the accuracy of the observation method was further verified by some physical indicators. This may help in the selection of suitable measurement methods to directly observe ice crystal formation behavior and screen cryoprotectants. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Controlling ice formation is critical in fields such as atmospheric science and biological cryopreservation. However, thermal heterogeneity during freezing and thawing in cryopreservation causes uneven ice crystallization and melting, leading to mechanical and thermal stress-induced damage. This study introduces biocompatible and biodegradable black phosphorus (BP)-polyethylene glycol-amine nanosheets (NS) to address this issue. BP NS primarily localize at ice grain boundaries, while amine groups of NH 2 -PEG-NH 2 form hydrogen bonds with H 2 O molecules, penetrating ice crystals. In situ cross-sectional observations confirmed that BP-PEG-NH 2 NS promotes uniform melting and facilitates ice cracks and boundaries. Heat transfer analysis using a bidirectional heating system revealed that the internal heat energy varies based on BP dispersion within the ice crystals. When applied to the cryopreservation of human tongue squamous cell carcinoma cells, BP-PEG-NH 2 NSs significantly improved post-thaw viability. It presents a promising strategy for designing thawing materials after cryopreservation of cells, tissues, and organs.

In 2012, the molecular structure of a new, broad class of ice-binding proteins, classified as "domain of unknown function" (DUF) 3494, was described for the first time. These proteins have a common tertiary structure and are characterized by a very wide spectrum of antifreeze activity (from weakly active to hyperactive). The ice-binding surface (IBS) region of these molecules differs significantly in its structure from the IBS of previously known antifreeze proteins (AFPs), showing a complete lack of regularity and high hydrophilicity. The presence of a regular, repeating structural motif in the IBS region of hitherto known AFP molecules, combined with the hydrophobic nature of this surface, promotes the formation of an ice-like ordering of the solvation water layer and, as a result, facilitates the process of transformation of this water layer into ice. It is, therefore, surprising that the newly discovered DUF3494 class of proteins clearly breaks out of this characteristic. In this paper, using molecular dynamics simulations, we analyze the solvation water structure of the IBS region of both DUF3494 family molecules and AFPs. As we show, although the IBS of DUF3494 molecules does not form an ice-like water structure in the solvation layer, this is compensated by the formation of the equivalent of "anchored clathrate water," in the form of a relatively large number of water molecules bound to the surface of the protein molecule and providing potential binding sites for it to the ice surface., (© 2024 Author(s). Published under an exclusive license by AIP Publishing.)

Bacterial ice nucleating proteins (INPs) are exceptionally effective in promoting the kinetically hindered transition of water to ice. Their efficiency relies on the assembly of INPs into large functional aggregates, with the size of ice nucleation sites determining activity. Experimental freezing spectra have revealed two distinct, defined aggregate sizes, typically classified as class A and C ice nucleators (INs). Despite the importance of INPs and years of extensive research, the precise number of INPs forming the two aggregate classes, and their assembly mechanism have remained enigmatic. Here, we report that bacterial ice nucleation activity emerges from more than two prevailing aggregate species and identify the specific number of INPs responsible for distinct crystallization temperatures. We find that INP dimers constitute class C INs, tetramers class B INs, and hexamers and larger multimers are responsible for the most efficient class A activity. We propose a hierarchical assembly mechanism based on tyrosine interactions for dimers, and electrostatic interactions between INP dimers to produce larger aggregates. This assembly is membrane-assisted: Increasing the bacterial outer membrane fluidity decreases the population of the larger aggregates, while preserving the dimers. Inversely, Dulbecco's Phosphate-Buffered Saline buffer increases the population of multimeric class A and B aggregates 200-fold and endows the bacteria with enhanced stability toward repeated freeze-thaw cycles. Our analysis suggests that the enhancement results from the better alignment of dimers in the negatively charged outer membrane, due to screening of their electrostatic repulsion. This demonstrates significant enhancement of the most potent bacterial INs., Competing Interests: Competing interests statement:The authors declare no competing interest.

Aircraft contrails, formed largely on soot particles in current flights, are important for aviation's non-CO 2 climate impact. Here we show that the activation of nonvolatile soot particles during contrail formation is likely determined by the sizes of primary soot particles rather than the effective sizes of soot aggregates as assumed in previous studies, which can explain less-than-unity fractions of soot particles forming contrail ice particles as recently observed during ECLIF (Emission and CLimate Impact of alternative Fuels) campaigns. The smaller soot primary sizes compared to aggregate sizes delay the onset of contrail ice formation, increase the maximum plume supersaturation reached in the contrail plume, and thus increase the probability of small volatile particles contributing to the total contrail ice particle number. This study suggests that the range of conditions for volatile plume particles to contribute significantly to the contrail ice number budget is wider than previously thought. As the aviation industry is moving toward sustainable aviation fuel and/or lean-burning engine technology, which is expected to reduce not only the emission index of nonvolatile soot particles but also the sizes of primary soot particles, this study highlights the need to better understand how the combined changes may affect contrail formation, contribution of volatile particles, and climate impacts.

Ice machines can harbor water-related organisms, and the use of ice or tap water for clinical care activities has been associated with infections in health care settings. During 2021-2022, a total of 23 cases of infection by Burkholderia multivorans (sequence type ST659) were reported at two southern California hospitals and linked to contaminated ice and water from ice machines. In addition to these 23 cases, this report also includes 23 previously unreported cases of B. multivorans ST659 infections that occurred during 2020-2024: 13 at a northern California hospital, eight at a hospital in Colorado, and two additional cases at one of the southern California hospitals. The same brand of ice machine and brands of filters, descaling, and sanitizing products were used by all four hospitals; B. multivorans was isolated from samples collected from ice machines in two of the hospitals. Whole genome sequencing indicated that all clinical and ice machine isolates were highly genetically similar (0-14 single nucleotide variant differences across 81% of the selected reference genome). Recommendations from public health officials to halt the outbreak included avoiding ice and tap water during clinical care activities. An investigation is ongoing to determine possible sources of ice machine contamination. During outbreaks of water-related organisms in health care facilities, health care personnel should consider avoiding the use of tap water, including ice and water from ice machines, for patient care., Competing Interests: All authors have completed and submitted the International Committee of Medical Journal Editors form for disclosure of potential conflicts of interest. Matthew J. Arduino reports honoraria and travel grants from the Society for Healthcare Epidemiology of America and the Association for Professionals in Infection Control and Epidemiology. Lindsay J. Caverly and John J. LiPuma report receiving grants from the Cystic Fibrosis Foundation during the investigation. No other potential conflicts of interest were disclosed.

Effectively controlling ice recrystallization (IR) during the frozen storage of food remains highly challenging. Inspired by the structural characteristics of antifreeze proteins in nature, silk fibroin (SF) derived from silk fibers has been developed. Through dual validation using the "splat" assay and "sucrose sandwich" assay, the IR inhibition activity of SF at various concentrations was confirmed, revealing that its regular alternating hydrophilic/hydrophobic domains endow SF with the potential to inhibit the axial growth of single ice crystal and significantly reduce the average maximum crystal size by approximately 67%. Additionally, the quality stability of frozen muscle foods treated with SF was comprehensively evaluated. In stark contrast to traditional commercial antifreeze agents (4% sucrose and 4% sorbitol), prepared steaks with the addition of 2% SF maintained rich juiciness and excellent color acceptability over a three-month frozen storage period. Thus, SF holds promise as a potential protective agent for frozen muscle foods, enhancing their quality during storage.

The present study compares two protocols for the cryopreservation of chicken semen. Both protocols had an initial low cooling rate in the first step, followed by higher cooling rates around ice nucleation (Protocol 1) or following the dissipation of the latent heat of fusion (Protocol 2) in the second step. Semen ejaculates obtained from 12 roosters were diluted with Rootex with 6% dimethylformamide and frozen following either Protocol 1 (from +5°C to -10°C at 5°C/min and from -10°C to -130°C at 60°C/min) or Protocol 2 (from +5°C to -35°C at 7°C/min and from -35°C to -140°C at 60°C/min). Compared with fresh semen, following both protocols, cryopreservation resulted in reduced post-thaw sperm quality (p < .001). Post-thaw percentage of sperm with an intact plasma membrane was greater using Protocol 2 than Protocol 1 (p < .05). The results suggest that high cooling rates around the time of ice nucleation are not recommendable., (© 2024 The Author(s). Reproduction in Domestic Animals published by Wiley‐VCH GmbH.)

Purpose: Healthcare workers (HCWs) wearing personal protective equipment (PPE) experience physiological strain that can impair motor and psychological functions, potentially affecting patient care. We assessed the effects of heat exposure on maximal strength and risk-taking behavior among PPE-wearing HCWs and the efficacy of ice slurry to alleviate adverse effects., Methods: Seventeen HCWS completed two experimental trials in a crossover design, consuming 5 g·kg -1 of body mass of ambient drink (AMB) or ice slurry (ICE) before donning PPE and undergoing 2 h of simulated decontamination exercise (wet-bulb globe temperature (WBGT): 25.9°C ± 0.8°C, PPE microenvironment WBGT: 29.1°C ± 2.1°C). Body core temperature ( Tc ), heart rate (HR), chest skin temperature ( Tsk ), ratings of perceived exertion (RPE), thermal sensation (RTS), maximal voluntary contraction (MVC), risk-taking behavior (balloon analogue risk-taking task (BART)), and salivary cortisol were assessed., Results: Predrinking to postdrinking ∆ Tc was greater in ICE (-0.2°C ± 0.1°C) than AMB (-0.0°C ± 0.1°C, P = 0.003). Post-drinking RTS was lower in ICE (2.7 ± 1.2) than AMB (4.1 ± 0.4, P < 0.001). ICE and AMB had similar Tc and HR (both P > 0.05), but Tsk was lower in ICE than AMB ( P = 0.049). A lower MVC (30.3 ± 6.7 vs 27.4 ± 4.9 kg, P = 0.001) and higher BART-adjusted total pump count (472 ± 170 vs 615 ± 174 pumps, P = 0.017) was observed pretrial to posttrial in AMB but absent in ICE (both P > 0.05). Salivary cortisol was similar between trials ( P = 0.42)., Conclusions: Heat-exposed PPE-wearing HCWs had impaired maximal strength and elevated risk-taking behavior. This may increase the risk of avoidable workplace accidents that can jeopardize HCWs and patient care. Ice slurry ingestion alleviated these heat-related impairments, suggesting its potential as an ergogenic aid., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American College of Sports Medicine.)

Neonicotinoids are widely used pesticides around the world, but the photolysis of neonicotinoids in cold agricultural region are still in blank. This paper aimed to study the influence of cold temperature over photolysis of neonicotinoids. To this end, the photolysis rates and photoproducts of dinotefuran and nitenpyram in water, ice and freeze-thawing condition were determined. Coupled with quantum chemistry calculation, the influence mechanisms of temperature and medium were investigated. The results showed the photolysis rates of neonicotinoids in water condition slightly declined with the lowered temperature due to the photolysis reactions were endothermic reactions. However, the photolysis rates increased by 89.8 %, 59.2 %, 49.4 % and 9.5 % for dinotefuran and nitenpyram in ice and thawing condition, respectively. This phenomenon was posed by the concentration-enhancing effect and change of photo-chemical properties of neonicotinoids in ice condition, which included lowered bond cleavage energy, lowered first excited singlet state energy and expanded light absorption range. The photolysis pathways of the two neonicotinoids did not change in different medium, but the concentration of carboxyl products was relatively higher than that of water condition due to the more amounts of reactive oxygen species in ice medium, which might increase the secondary pollution risk after ice-off in spring due to the higher ecotoxicity to nontarget organism of these photoproducts. The influence of cold temperature and medium change should be considered for the environmental fate and risk assessment of neonicotinoids in cold agricultural region., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: This study aimed to determine the effect of menthol ice application on thirst, dry mouth, mouth taste, and bad mouth odor in patients who underwent laparoscopic cholecystectomy in the postoperative period., Design: The study was conducted as a randomized controlled trial with an experimental design., Methods: The study sample consisted of 90 patients who underwent laparoscopic cholecystectomy and met the inclusion criteria. Patients were divided into three groups by randomization program. Patients in the menthol ice and ice popsicle (ice prepared with drinking water only) group were administered menthol ice/ice popsicle (10 mL) twice at 20 minutes intervals. Patients in the control group did not receive any intervention. Routine practices of the clinic were performed by the nurses of the general surgery clinic. Postoperative thirst intensity, severity of dry mouth, bad taste, and bad odor in the mouth were evaluated at 0, 20, and 40 minutes., Findings: No statistically significant difference was found between the sociodemographic and clinical characteristics of the patients in the control group, menthol ice group, and ice popsicle group (P > .05). A statistically significant difference was found between the 3 groups in terms of thirst intensity and severity of dry mouth at times at the 20th and 40th minutes after the application (P < .01). We found a statistically significant difference between the bad taste and bad odor sensation scores of the patients in the control and intervention groups at the 20th and 40th minutes after the application (P < .05)., Conclusions: The study concluded that menthol ice and ice popsicle application are effective strategies to reduce the intensity of thirst, severity of dry mouth, bad taste, and bad odor in postoperative patients., Competing Interests: Declaration of Competing Interest The authors declare no actual or potential conflict of interest., (Copyright © 2024. Published by Elsevier Inc.)

This study investigated the expediated transformation of halophenols in the presence of nitrite (NO 2 ) under slightly acidic conditions in ice, whereas such transformation was negligible in liquid water at 4 °C. We proposed that this phenomenon was attributed to the freeze-concentration effect, incurring a pH drop and the aggregation of NO - and halophenols within the liquid-like grain boundary layer amid ice crystals. Within this micro-environment, NO 2 - and halophenols within the liquid-like grain boundary layer amid ice crystals. Within this micro-environment, NO 2 - underwent protonation to generate reactive nitrous acid (HNO 2 drove oxidative coupling reactions, generating hydroxyl polyhalogenated diphenyl ethers (OH-PBDEs) and hydroxyl polyhalogenated diphenyls via C-O or C-C coupling. These two pathways were intricately intertwined. The presence of natural organic matter (NOM) mitigated the formation of nitrated products and completely suppressed the coupling products. This study offers valuable insights into the fate of halophenols in ice and suggests potential pathways for the formation of nitrophenolic compounds and OH-PBDEs in natural cold environments. These findings also open up a new avenue in environmental chemistry research.+ ) that facilitate the nitration and oxidation of halophenols. When 10 μМ halophenol was treated by freezing in the presence of 5 μМ NO 2 - , the total yields of nitrated products reached 2.4 μМ and 1.4 μМ within 12 h for 2-chlorophenol (2CP) and 2-bromophenol (2BP), respectively. NO + drove oxidative coupling reactions, generating hydroxyl polyhalogenated diphenyl ethers (OH-PBDEs) and hydroxyl polyhalogenated diphenyls via C-O or C-C coupling. These two pathways were intricately intertwined. The presence of natural organic matter (NOM) mitigated the formation of nitrated products and completely suppressed the coupling products. This study offers valuable insights into the fate of halophenols in ice and suggests potential pathways for the formation of nitrophenolic compounds and OH-PBDEs in natural cold environments. These findings also open up a new avenue in environmental chemistry research., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Junhe Lu reports financial support was provided by National Natural Science Foundation of China. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Controlling the formation and growth of ice is essential to successfully cryopreserve cells, tissues and biologics. Current efforts to identify materials capable of modulating ice growth are guided by iterative changes and human intuition, with a major focus on proteins and polymers. With limited data, the discovery pipeline is constrained by a poor understanding of the mechanisms and the underlying structure-activity relationships. In this work, this barrier is overcome by constructing machine learning models capable of predicting the ice recrystallisation inhibition activity of small molecules. We generate a new dataset via experimental measurements of ice growth, then harness predictive models combining state-of-the-art descriptors with domain-specific features derived from molecular simulations. The models accurately identify potent small molecule ice recrystallisation inhibitors within a commercial compound library. Identified hits can also mitigate cellular damage during transient warming events in cryopreserved red blood cells, demonstrating how data-driven approaches can be used to discover innovative cryoprotectants and enable next-generation cryopreservation solutions for the cold chain., (© 2024. The Author(s).)

Recent research has shown that microplastics are widespread in the atmosphere. However, we know little about their ability to nucleate ice and their impact on ice formation in clouds. Ice nucleation by microplastics could also limit their long-range transport and global distribution. The present study explores the heterogeneous ice-nucleating ability of seven microplastic samples in immersion freezing mode. Two polypropylene samples and one polyethylene terephthalate sample froze heterogeneously with median freezing temperatures of -20.9, -23.2, and -21.9 °C, respectively. The number of ice nucleation sites per surface area, n s ( T ), ranged from 10 -1 to 10 4 in a temperature interval of -15 to -25 °C, which is comparable to that of volcanic ash and fungal spores. After exposure to ozone or a combination of UV light and ozone, simulating atmospheric aging, the ice nucleation activity decreased in some cases and remained unchanged in others. Our freezing data suggest that microplastics may promote ice formation in cloud droplets. In addition, based on a comparison of our freezing results and previous simulations using a global transport model, ice nucleation by microplastics will impact their long-range transport to faraway locations and global distribution.-2 in a temperature interval of -15 to -25 °C, which is comparable to that of volcanic ash and fungal spores. After exposure to ozone or a combination of UV light and ozone, simulating atmospheric aging, the ice nucleation activity decreased in some cases and remained unchanged in others. Our freezing data suggest that microplastics may promote ice formation in cloud droplets. In addition, based on a comparison of our freezing results and previous simulations using a global transport model, ice nucleation by microplastics will impact their long-range transport to faraway locations and global distribution.

When cells are cryopreserved, they go through a freezing process with several distinct phases (i.e., cooling until nucleation, ice nucleation, ice crystal growth and cooling to a final temperature). Conventional cell freezing approaches often employ a single cooling rate to describe and optimize the entire freezing process, which neglects its complexity and does not provide insight into the effects of the different freezing phases. The aim of this work was to elucidate the impact of each freezing phase by varying different process parameters per phase. Hereto, spin freezing was used to freeze Jurkat T cells in either a Me 2 SO-based or Me 2 SO-free formulation. The cooling rates before ice nucleation and after total ice crystallization impacted cell viability, resulting in viability ranging from 26.7% to 52.8% for the Me 2 SO-free formulation, and 22.5%-42.6% for the Me 2 SO-based formulation. Interestingly, the degree of supercooling upon nucleation did not exhibit a significant effect on cell viability in this work. However, the rate of ice crystal formation emerged as a crucial factor, with viability ranging from 2.4% to 53.2% for the Me 2 SO-free formulation, and 0.3%-53.2% for the Me 2 SO-based formulation, depending on the freezing rate. A morphological study of the cells post-cryopreservation was performed using confocal microscopy, and it was found that cytoskeleton integrity and cell volume were impacted, depending on the formulation-process parameter combination. These findings underscore the importance of scrutinizing all cooling and freezing phases, as each phase impacted post-thaw viability in a distinct way, depending of the specific formulation used., (Copyright © 2024 Society for Cryobiology. Published by Elsevier Inc. All rights reserved.)

Large yellow croaker (Larimichthys crocea) is susceptible to oxidative denaturation during storage. This work is to investigate the quality alterations by analyzing its physicochemical changes and proteomics throughout preservation under refrigeration, frozen, and slurry ice (SI) conditions. Results revealed that the freshness of large yellow croaker, as evaluated by indicators such as total volatile basic nitrogen, total viable count, and thiobarbituric acid reactive substances, was well maintained while stored in the SI group. Meanwhile, the water distribution in the muscle tissue of group SI exhibited slower fluctuations, thereby preserving the integrity of fish muscle cells. Based on label-free proteomic analysis, a considerable downregulation was observed in the mitogen-activated protein kinase (MAPK) signaling pathway, indicating that SI decelerated this metabolic pathway and effectively delayed the deterioration of muscle. Therefore, the application of SI provides potential for maintaining the quality stability of large yellow croaker., Competing Interests: Declaration of competing interest The authors declare that they do not possess any recognized competing financial interests or personal relationships that could have potentially influenced the findings presented in this research., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Explosive ordnance disposal (EOD) technicians may be required to work in hot, humid environments while wearing heavy protective clothing. We investigated the ability of an ice vest to attenuate physiological strain and subsequently extend work tolerance., Methods: Eight male participants (24.3 ± 4.1 yr, 51.9 ± 4.6 mL kg -1 min -1 ) walked (4.5 km h -1 ) in simulated hot and humid conditions (35 °C; 50% relative humidity). Participants wore either an EOD suit (CON) or EOD and ice vest (IV). Heart rate, core and skin temperature were recorded continuously., Results: Participants walked longer in IV compared to CON (8.1 ± 7.4 min, p < .05). Over 90% of trials were terminated based on participants reaching 90% of their maximum heart rate. IV resulted in cooled skin (p < .001) and a physiologically negligible change in core temperature (p < .001). A condition by time interaction was identified for heart rate (p < .001), with a lower rate of rise in the IV condition., Conclusions: The cardiovascular inefficiency that limited performance was attenuated in the IV condition. The ice vest facilitated heat loss from the periphery; thus, the observed reduction in heart rate may reflect the preservation of central blood volume. The results identify the efficiency of a simple, inexpensive ice vest to assist EOD technicians working in the heat., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Creating molecules capable of inhibiting ice recrystallization is an active research area aiming to improve the freeze-thaw characteristics of foods and biomedical materials. Peptide mixtures have shown promise in preventing freezing-induced damage, but less is known about the relationship between their amino acid compositions and ice recrystallization inhibition (IRI) activities. In this article, we used Ni 2+ immobilized metal affinity chromatography (IMAC) to fractionate pulse protein hydrolysates, created by Alcalase and trypsin, into mixtures lacking and enriched in His, and Cys residues. The aim of this study was to fractionate pulse protein hydrolysates based on their amino acid compositions and evaluate their resulting physicochemical and IRI characteristics. Ni 2+ IMAC fractionation induced IRI activity in all of the evaluated soy, chickpea, and pea protein hydrolysates regardless of their amino acid composition. Ni 2+ IMAC fractionation produced chemically distinct fractions of peptides, differing by their molecular weights, amino acid composition, and IRI activities. The resulting peptide mixtures' molecular weight, amino acid composition, secondary structure, and sodium ion levels were found to have no correlation with their IRI activities. Thus, we demonstrate for the first time the ability of Ni 2+ IMAC fractionation to induce IRI activity in hydrolyzed pulse proteins., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interests., (Copyright © 2024. Published by Elsevier Ltd.)

An antifreeze protein's inclusion into ice can be used to purify it from other proteins and solutes. Domains that are covalently attached to the antifreeze protein are also drawn into the ice such that the ice-binding portion of the fusion protein can be used as an affinity tag. Here we have explored the use of ice-affinity tags on multi-subunit proteins. When an ice-binding protein was attached as a tag to multisubunit complexes a substantial portion of each multimer dissociated during overgrowth by the ice. The protein subunit attached to the affinity tag was enriched in the ice and the other subunit was appreciably excluded. We suggest that step growth of the advancing ice front generates shearing forces on the bound complex that can disrupt non-covalent protein-protein interactions. This will effectively limit the use of ice-affinity tags to single subunit proteins., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Accurate identification of ice phases is essential for understanding various physicochemical phenomena. However, such classification for structures simulated with molecular dynamics is complicated by the complex symmetries of ice polymorphs and thermal fluctuations. For this purpose, both traditional order parameters and data-driven machine learning approaches have been employed, but they often rely on expert intuition, specific geometric information, or large training data sets. In this work, we present an unsupervised phase classification framework that combines a score-based denoiser model with a subsequent model-free classification method to accurately identify ice phases. The denoiser model is trained on perturbed synthetic data of ideal reference structures, eliminating the need for large data sets and labeling efforts. The classification step utilizes the smooth overlap of atomic position (SOAP) descriptors as the atomic fingerprint, ensuring Euclidean symmetries and transferability to various structural systems. Our approach achieves a remarkable 100% accuracy in distinguishing ice phases of test trajectories using only seven ideal reference structures of ice phases as model inputs. This demonstrates the generalizability of the score-based denoiser model in facilitating phase identification for complex molecular systems. The proposed classification strategy can be broadly applied to investigate structural evolution and phase identification for a wide range of materials, offering new insights into the fundamental understanding of water and other complex systems.

Damage from ice and potential toxicity of ice-inhibiting cryoprotective agents (CPAs) are key issues in assisted reproduction of humans, domestic and research animals, and endangered species using cryopreserved oocytes and embryos. The nature of ice formed in bovine oocytes (similar in size to oocytes of humans and most other mammals) after rapid cooling and during rapid warming was examined using synchrotron-based time-resolved x-ray diffraction. Using cooling rates, warming rates and CPA concentrations of current practice, oocytes show no ice after cooling but always develop large ice fractions-consistent with crystallization of most free water-during warming, so most ice-related damage must occur during warming. The detailed behavior of ice at warming depended on the nature of ice formed during cooling. Increasing cooling rates allows oocytes soaked as in current practice to remain essentially ice free during both cooling and warming. Much larger convective warming rates are demonstrated and will allow routine ice-free cryopreservation with smaller CPA concentrations. These results clarify the roles of cooling, warming, and CPA concentration in generating ice in oocytes and establish the structure and grain size of ice formed. Ice formation can be eliminated as a factor affecting post-warming oocyte viability and development in many species, improving outcomes and allowing other deleterious effects of the cryopreservation cycle to be independently studied., (© 2024. The Author(s).)

Compromised heat loss due to limited convection and evaporation can increase thermal strain. We aimed to determine the effectiveness of ice slurry ingestion to reduce thermal strain following hyperthermia in a state of compromised heat loss. Twelve healthy males (age: 25 ± 4y) underwent hot water immersion to elevate rectal temperature (Trec) by 1.82 ± 0.08°C on four occasions. In the subsequent 60-min of seated recovery, participants ingested either 6.8 g·kg-1 of ice slurry (-0.6°C) or control drink (37°C) in ambient conditions (21 ± 1°C, 39 ± 10% relative humidity), wearing either t-shirt and shorts (2 trials: ICE and CON) or a whole-body sweat suit (2 trials: ICE-SS and CON-SS). Trec and mean skin temperature (Tsk) were recorded and a two-compartment thermometry model of heat storage was calculated. Heat storage was lower in ICE compared with CON at 20-40min (p ≤ 0.044, d ≥ 0.88) and for ICE-SS compared with CON-SS at 40-60 min (p ≤ 0.012, d ≥ 0.93). Trec was lower in ICE compared with CON from 30-60min (p ≤ 0.034, d ≥ 0.65), with a trend for a reduced Trec in ICE-SS compared with CON-SS at 40min (p = 0.079, d = 0.60). A greater Tsk was found in ICE-SS and CON-SS compared with ICE and CON (p < 0.001, d ≥ 3.37). A trend for a lower Tsk for ICE compared with CON was found at 20-40min (p ≤ 0.099, d ≥ 0.53), no differences were found for ICE-SS vs CON-SS (p ≥ 0.554, d ≤ 0.43). Ice slurry ingestion can effectively reduce heat storage when heat loss through convection and evaporation is compromised, relevant to those wearing personal protective equipment or those with compromised sweat loss. Compromised heat loss delays the reduction in heat storage, possibly related to ice slurry ingestion not lowering Tsk., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 O’Brien et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Oocyte vitrification has become a widely adopted method in clinical practice. However, the solidification behavior and its impact on oocytes during the ultrarapid cooling process remain poorly understood. In this study, we established a system and methodology to observe crystallization behavior in oocytes during quench cooling and warming. Subsequently, the threshold concentration of cryoprotective agents (CPAs) required for oocyte vitrification was determined through a visualization method. The results demonstrated that the ice front could not be observed in the image sequence when using 16.5% DMSO +16.5% EG during high-speed quench cooling (2821.58°C/min). Finally, oocytes were encapsulated with an antifreezing hydrogel (7.5% EG +7.5% DMSO +0.5% alginate) and subjected to high-speed quench cooling. No ice crystals appeared in the antifreezing hydrogel-encapsulated oocytes at a low concentration of osmotic CPA (2.4 M). This research opens up new possibilities for oocyte vitrification with a reduced concentration of CPA.

This study aimed to determine the bacteriological quality and presence of diarrheagenic Escherichia coli pathotypes (DEP) and nontuberculous mycobacteria (NTM) species in 85 packaged ice samples from 12 different states of central Mexico. Three samples had a pH of 9.8 and therefore fell outside of the acceptable range for pH. All samples were positive for aerobic-mesophilic bacteria, with limits ranging from 1 to 3.47 log CFU/mL. In total, 35, 11, and 3 ice samples were positive for total coliforms (TC), fecal coliforms (FC), and E. coli, respectively. In the samples, the TC concentration ranged from <1.1 to >23 MPN/100 mL and from <1.1 to 23 MPN/100 mL for FC and E. coli. In total, 38 (44.7%) ice samples were outside of Mexico's official guidelines. None of the 12 E. coli strains isolated from the three ice samples belonged to DEP. NTM were recovered from 20 ice samples and included M. neoaurum (n = 7), M. porcinum (n = 2), M. flavescens (n = 2), M. fortuitum (n = 1), M. abscessus (n = 1), M. senegalense (n = 1), M. conceptionense (n = 1), and M. sp. (n = 1). In the remaining four samples, two NTM were isolated simultaneously. Thus, we recommend that producers should evaluate the microbiological quality of purified water used as a raw material as well as that of the final product, the ice should be packed in thick bags to avoid stretching and tearing during transportation or storage to prevent environmental contamination of ice, personnel involved in the production, and handling of ice should be trained in relative hygiene matters and how ice-machines should be cleaned and disinfected and the implementation of hazard analysis and critical control points must be applied throughout the chain of production. Finally, regular inspection by the authorities is also of great importance. These recommendations can be applied in different countries with low microbiological quality packaged ice., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

During cryopreservation, the growth of ice crystals can cause mechanical damage to samples, which is one of the important factors limiting the quality of preserved samples. To enhance the preservation quality of biological samples, scholars have tried various engineering methods. Among them, an electric field is an essential factor affecting solution freezing. Dimethyl sulfoxide, as a commonly used cryoprotectant, can cause mechanical damage to cells due to ice crystals even when freezing at the optimal freezing rate. Water is a strongly polar dielectric material, and the applied alternating current (AC) electric field will affect the water freezing performance. Therefore, a quantitative study of ice crystal nucleation and growth during freezing of dimethyl sulfoxide solutions under different AC electric field conditions is needed to try to reduce ice crystal damage. We created a liquid-film device to approximate the ice crystal growth process as a two-dimensional image. The frequency of the AC voltage was set from 0 to 50 kHz. We measured the supercooling of the dimethyl sulfoxide solution under AC electric field conditions. As an objective and accurate quantitative analysis of the ice crystal growth process, we propose a Dilated Convolutional Segmentation Transformer for semantic segmentation of ice crystal images. It is concluded that the average area and the growth rate of single ice crystals decrease with increasing electric field frequency at a certain concentration of dimethyl sulfoxide solution. Lower concentrations of dimethyl sulfoxide solution in combination with an AC electric field can achieve similar ice suppression effects as when higher concentrations of dimethyl sulfoxide solution act alone. We believe that AC electric fields are expected to be an aid to cryopreservation and provide some theoretical basis and experimental foundation for its development.

The Winter Wonderland ice cave, located at an elevation of 3140 m above sea level in the Uinta Mountains of northern Utah, USA, maintains a constant sub-zero temperature. Seasonal snowmelt and rain enter the cave, freeze on the surface of the existing ice, and contribute to a 3-m-thick layered ice mass. This ice mass contains organic matter and cryogenic cave carbonates (CCCs) that date back centuries. In this study, samples of ice, liquid water, and exposed CCCs were collected to examine the bacterial communities within the cave and to determine if these communities vary spatially and between sample types. Flow cytometry showed that cell counts are an order of magnitude higher in liquid water samples than in ice. Epifluorescence microscopy and scanning electron microscopy imaging revealed potential coccoid and bacillus microbial morphologies in water samples and putative cells or calcite spherules in the CCCs. The diversity of bacteria associated with soil, identified through sequence-based analysis, supports the hypothesis that water enters the cave by filtering through soil and bedrock. A differential abundance of bacterial taxa was observed between sample types, with the greatest diversity found in CCCs. This supports a geomicrobiological framework where microbes aggregate in the water, sink into a concentrated layer, and precipitate out of the ice with the CCCs, thereby reducing the cell counts in the ice. These CCCs may provide essential nutrients for the bacteria or could themselves be products of biomineralization., (© 2024 The Author(s). MicrobiologyOpen published by John Wiley & Sons Ltd.)

Microorganisms' ice-binding structures (IBS) are macromolecules with potential commercial value in agriculture, food technology, material technology, cryobiology, and medicine. Microbial ice-structuring or microbial ice-binding particles, with their multi-applications, are simple to use, effective in low amounts, non-toxic, and environmentally friendly. Due to their source and composition diversities, microbial ice-binding structures are gaining attention because they are useable in various conditions. Some microorganisms also produce structures with dual ice-nucleating and anti-freezing properties. Structures that promote ice formation (ice nucleating particles- INPs) act as ice nuclei, lowering the energy barrier between supercooled liquid and ice, causing ice crystals to form. In contrast, anti-freeze particles (AFPs) prevent ice formation and recrystallization through several mechanisms, including disturbing the formation of string hydrogen bonds amongst water molecules, melting already formed ice crystals, and preventing crystal formation by binding to specific sites. Knowledge of the type and function of microbial ice-binding structures lends fundamental insight for possible scaling the production of cheap, functional, and advanced microbial structure-inspired mimics and by-products. This review focuses on microbial ice-binding structures and their potential uses in the food, medicinal, environmental, and agricultural sectors., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The ice phenomena are an inherent component of rivers in temperate, continental, and polar climate zones. Evident progress in global warming leads to a decrease in snow cover on land and ice phenomena in water bodies, disrupting the stability of the hydrological cycle and aquatic ecosystems. Although common observations indicate the disappearance of ice phenomena in rivers over recent decades, detailed quantitative research is lacking in many regions, especially in the temperate zone. In this paper, ice phenomena were analyzed on the rivers of southern Poland, located in the upland and mountain areas of the country, as no such studies have been conducted so far. The temporal changes in the annual number of days with ice (NDI) phenomena were studied in locations where ice phenomena were observed every year for at least 30 years between 1951 and 2021. Using straightforward but commonly accepted procedures, such as the Mann-Kendall test, statistically significant decreasing trends in the annual NDI were revealed for the majority of gauging stations. The Theil-Sen (TS) slope mean values were -1.66 (ranging from -3.72 to -0.56), -1.41 (from -3.22 to -0.29), and -1.33 (from -2.85 to -0.29) for the datasets representing the periods 1992-2020, 1987-2020, and 1982-2020, respectively. The results for the annual NDI were additionally presented within the context of meteorological characteristics such as annual and winter (Nov-Apr) air temperature, precipitation, and water temperature. Correlation and regression analyses revealed that the main factor triggering the decrease in NDI is the increase in the average winter air temperature. An increase in temperature by 1°C results in a decrease in NDI by up to twenty days. If these negative trends continue, ice phenomena may disappear completely from southern Polish rivers within few decades., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Kochanek et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Recrystallization is considered the main damaging mechanism during the frozen storage of biologic materials. In this study, furcellaran, a polysaccharide related to κ-carrageenan, was studied for its concentration-dependent effect on ice crystal growth and recrystallization. The structure and sulfate content of the utilized furcellaran was analyzed by 1 H nuclear magnetic resonance spectroscopy, ion chromatography, and high-performance size-exclusion chromatography. Additionally, the rheological properties of furcellaran solutions were investigated. Our findings demonstrate that furcellaran inhibits ice growth as effectively as κ-carrageenan. Furthermore, the rheological properties change with increasing furcellaran concentration, resulting in a gel-like consistency at 5 g/L, which coincides with decreased recrystallization inhibition activity and larger crystals. This suggests that gel formation or a gel-like consistency has to be avoided for optimal recrystallization inhibition activity.

Introduction: Acupressure and cold application are nonpharmacologic methods that midwives can use for labor pain. The purpose of this study was to determine the effects of ice massage applied to the SP6 acupressure point during labor on labor pain, labor comfort, labor duration, and anxiety., Methods: A single-masked, randomized controlled trial was conducted with 100 nulliparous women, including 50 in the intervention group and 50 in the control group. Rotational ice massage was applied to the SP6 point on both legs of the pregnant women in the intervention group at 4 to 5 cm, 6 to 7 cm, and 8 to 9 cm dilation amounts during 3 contractions. Routine oxytocin was administered to all pregnant women to promote progress of labor. Standard midwifery care was provided to the control group. Data were collected using a Personal Information Form, the Visual Analog Scale (VAS), a partograph form, the Childbirth Comfort Questionnaire (CCQ), and the State-Trait Anxiety Inventory State subscale., Result: The VAS pain scores of the pregnant women in the intervention group were significantly lower compared with the control group following the intervention at dilatations of 4 to 5 cm, 6 to 7 cm, and 8 to 9 cm (P = .001, P = .003, P <.001, respectively). The total CCQ and comfort level scores of the pregnant women in the intervention group at 8 to 9 cm cervical dilatation were significantly higher than the control group (P = 0.044, P = .027, respectively). Additionally, as the anxiety levels of the pregnant women increased, their total comfort scores decreased (P <.05)., Discussion: Ice application to the SP6 point during stage 1 of childbirth reduced labor pain, increased comfort, and reduced anxiety levels. No adverse events were found in the intervention group with ice massage applied to the SP6 point. Therefore, this method can be used as a safe and effective midwifery intervention in childbirth., (© 2024 The Authors. Journal of Midwifery & Women's Health published by Wiley Periodicals LLC on behalf of American College of Nurse Midwives (ACNM).)

Synthetic tubular grafts currently used in clinical context fail frequently, and the expectations that biomimetic materials could tackle these limitations are high. However, developing tubular materials presenting structural, compositional and functional properties close to those of native tissues remains an unmet challenge. Here we describe a combination of ice templating and topotactic fibrillogenesis of type I collagen, the main component of tissues' extracellular matrix, yielding highly concentrated yet porous tubular collagen materials with controlled hierarchical architecture at multiple length scales, the hallmark of native tissues' organization. By modulating the thermal conductivity of the cylindrical molds, we tune the macroscopic porosity defined by ice. Coupling the aforementioned porosity patterns with two different fibrillogenesis routes results in a new family of tubular materials whose textural features and the supramolecular arrangement of type I collagen are achieved. The resulting materials present hierarchical elastic properties and are successfully colonized by human endothelial cells and alveolar epithelial cells on the luminal side, and by human mesenchymal stem cells on the external side. The proposed straightforward protocol is likely to be adapted for larger graft sizes that address ever-growing clinical needs, such as peripheral arterial disease or tracheal and bronchial reconstructions.

Antifreeze proteins (AFPs) are remarkable biomolecules that suppress ice formation at trace concentrations. To inhibit ice growth, AFPs must not only bind to ice crystals, but also resist engulfment by ice. The highest supercooling, [Formula: see text], for which AFPs are able to resist engulfment is widely believed to scale as the inverse of the separation, [Formula: see text], between bound AFPs, whereas its dependence on the molecular characteristics of the AFP remains poorly understood. By using specialized molecular simulations and interfacial thermodynamics, here, we show that in contrast with conventional wisdom, [Formula: see text] scales as [Formula: see text] and not as [Formula: see text]. We further show that [Formula: see text] is proportional to AFP size and that diverse naturally occurring AFPs are optimal at resisting engulfment by ice. By facilitating the development of AFP structure-function relationships, we hope that our findings will pave the way for the rational design of AFPs., Competing Interests: Competing interests statement:The authors declare no competing interest.

Vitrification under isochoric (constant-volume or volumetrically confined) conditions has emerged as an intriguing new cryopreservation modality, but the physical complexities of the process confound straight-forward interpretation of experimental results. In particular, the signature pressure-based ice detection used in many isochoric techniques becomes paradoxical during vitrification, wherein the emergence of a sharp increase in pressure reliably indicates the presence of ice, but avoidance of this increase does not necessarily indicate its absence. This phenomenon arises from the rich interplay between thermochemical and thermovolumetric effects in isochoric systems, and muddies efforts to confirm the degree to which a sample has vitrified. In this work, we seek to aid interpretation of isochoric vitrification experiments by calculating thermodynamic limits on the maximum amount of ice that may form without being detected by pressure, and by clarifying the myriad physical processes at play. Neglecting kinetic effects, we develop a simplified thermodynamic model accounting for thermal contraction, cavity formation, ice growth, solute ripening, and glass formation, we evaluate it for a range of chamber materials and solution compositions, and we validate against the acutely limited data available. Our results provide both counter-intuitive insights- lower-concentration solutions may contract less while producing more pressure-undetectable ice growth for example- and a general phenomenological framework by which to evaluate the process of vitrification in isochoric systems. We anticipate that the model herein will enable design of future isochoric protocols with minimized risk of pressure-undetectable ice formation, and provide a thermodynamic foundation from which to build an increasingly rigorous multi-physics understanding of isochoric vitrification., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: This study investigates the thermal interactions between adjacent vials during freezing and assesses their impact on nucleation times., Methods: Various loading configurations were analyzed to understand their impact on nucleation times. Configurations involving direct contact between vials and freeze-dryer shelves were studied, along with setups using empty vials between filled ones. Additionally, non-conventional loading configurations and glycol-filled vials were tested. The analysis includes 2R and 20R vials, which are commonly utilized in the freezing and lyophilization of drug products, along with two different fill depths, 1 and 1.4 cm., Results: The investigation revealed that configurations with direct contact between vials and freeze-dryer shelves led to substantial thermal interactions, resulting in delayed nucleation in adjacent vials and affecting the temperature at which nucleation takes place in a complex way. In another setup, empty vials were placed between filled vials, significantly reducing thermal interactions. Further tests with non-conventional configurations and glycol-filled vials confirmed the presence of thermal interactions with a minimal inhibitory effect., Conclusions: These findings carry significant implications for the pharmaceutical industry, highlighting the role of thermal interactions among vials during freezing and their impact on the temperature at which ice nucleation occurs., (© 2024. The Author(s).)

Black ice, a phenomenon that occurs abruptly owing to freezing rain, is difficult for drivers to identify because it mirrors the color of the road. Effectively managing the occurrence of unforeseen accidents caused by black ice requires predicting their probability using spatial, weather, and traffic factors and formulating appropriate countermeasures. Among these factors, weather and traffic exhibit the highest levels of uncertainty. To address these uncertainties, a study was conducted using a Monte Carlo simulation based on random values to predict the probability of black ice accidents at individual road points and analyze their trigger factors. We numerically modeled black ice accidents and visualized the simulation results in a geographical information system (GIS) by employing a sensitivity analysis, another feature of Monte Carlo simulations, to analyze the factors that trigger black ice accidents. The Monte Carlo simulation allowed us to map black ice accident occurrences at each road point on the GIS. The average black ice accident probability was found to be 0.0058, with a standard deviation of 0.001. Sensitivity analysis using Monte Carlo simulations identified wind speed, air temperature, and angle as significant triggers of black ice accidents, with sensitivities of 0.354, 0.270, and 0.203, respectively. We predicted the probability of black ice accidents per road section and analyzed the primary triggers of black ice accidents. The scientific contribution of this study lies in the development of a method beyond simple road temperature predictions for evaluating the risk of black ice occurrences and subsequent accidents. By employing Monte Carlo simulations, the probability of black ice accidents can be predicted more accurately through decoupling meteorological and traffic factors over time. The results can serve as a reference for government agencies, including road traffic authorities, to identify accident-prone spots and devise strategies focused on the primary triggers of black ice accidents., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Hong, Yun. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Freezing of biological drug substance (DS) is a critical unit operation that may impact product quality, potentially leading to protein aggregation and sub-visible particle formation. Cryo-concentration has been identified as a critical parameter to impact protein stability during freezing and should therefore be minimized. The macroscopic cryo-concentration, in the following only referred to as cryo-concentration, is majorly influenced by the freezing rate, which is in turn impacted by product independent process parameters such as the DS container, its size and fill level, and the freezing equipment. (At-scale) process characterization studies are crucial to understand and optimize freezing processes. However, evaluating cryo-concentration requires sampling of the frozen bulk, which is typically performed by cutting the ice block into pieces for subsequent analysis. Also, the large amount of product requirement for these studies is a major limitation. In this study, we report the development of a simple methodology for experimental characterization of frozen DS in bottles at relevant scale using a surrogate solution. The novel ice core sampling technique identifies the axial ice core in the center to be indicative for cryo-concentration, which was measured by osmolality, and concentrations of histidine and polysorbate 80 (PS80), whereas osmolality revealed to be a sensitive read-out. Finally, we exemplify the suitability of the method to study cryo-concentration in DS bottles by comparing cryo-concentrations from different freezing protocols (-80°C vs -40°C). Prolonged stress times during freezing correlated to a higher extent of cryo-concentration quantified by osmolality in the axial center of a 2 L DS bottle., (© 2024. The Author(s).)

Background: Characterization of intracellular ice formation (IIF) in oocytes during the freezing and thawing processes will contribute to optimizing their cryopreservation. However, the observation of the ice formation process in oocytes is limited by the spatiotemporal resolution of the cryomicroscope systems., Objective: To observe the intracellular icing of oocytes during cooling and rewarming, and to study the mechanism of formation and growth of intracellular ice in oocytes., Materials and Methods: Mouse oocytes were frozen at different cooling rates to induce intracellular ice formation using a cryomicroscopy system consisting of a microscope equipped with a cryogenic cold stage, an automatic cooling system, a temperature control system, and a high-speed camera. The growth patterns of intracellular ice in oocytes were analyzed from the images recorded. Finally, the growth rate of intracellular ice formation in oocytes was calculated using an automatic intracellular ice tracking method., Results: The IIF temperature decreased gradually with the increase in cooling rate. Initiation sites of IIF could be classified into three categories: marginal type, internal type and coexisting type. There was a strong predominance for ice crystal initiation site in the oocytes, with up to 80% of the initiation sites located in the marginal region. The intracellular ice growth modes of darkening and twitching cells were characterized by "spreading" and "clustering", respectively. In addition, twitching cells started to recrystallize during rewarming, while darkening cells did not. The instantaneous maximal growth rate of ice crystals in twitching cells was about 10 times higher than that in darkening cells., Conclusion: By visualising the growth of ice crystals in mouse oocytes during cooling and rewarming, we obtained valuable information on the kinetics of ice formation and melting in these cells. This information can help us understand how ice formation and melting affect the viability and quality of oocytes after cryopreservation. Doi.org/10.54680/fr24310110412.

The main sources of redox gradients supporting high-productivity life in the Europan and other icy ocean world oceans were proposed to be photolytically derived oxidants, such as reactive oxygen species (ROS) from the icy shell, and reductants (Fe(II), S(-II), CH 4 , H 2 ) from bottom waters reacting with a (ultra)mafic seafloor. Important roadblocks to maintaining life, however, are that the degree of ocean mixing to combine redox species is unknown, and ROS damage biomolecules. Here, we envisage a unique solution using an acid mine drainage (AMD)-filled pit lakes analog system for the Europan ocean, which previous models predicted to be acidic. We hypothesize that surface-generated ROS oxidize dissolved Fe(II) resulting in Fe(III) (hydr)oxide precipitates, that settle to the seafloor as "iron snow." The iron snow provides a respiratory substrate for anaerobic microorganisms ("breathing iron"), and limits harmful ROS exposure since they are now neutralized at the ice-water interface. Based on this scenario, we calculated Gibbs energies and maximal biomass productivities of various anaerobic metabolisms for a range of pH, temperatures, and H 2 fluxes. Productivity by iron reducers was greater for most environmental conditions considered, whereas sulfate reducers and methanogens were more favored at high pH. Participation of Fe in the metabolic redox processes is largely neglected in most models of Europan biogeochemistry. Our model overcomes important conceptual roadblocks to life in icy ocean worlds and broadens the potential metabolic diversity, thus increasing total primary productivity, the diversity and volume of habitable environmental niches and, ultimately, the probability of biosignature detection., Competing Interests: Competing interests statement:The authors declare no competing interest.

Microplastics have littered the globe, with synthetic fibers being the largest source of atmospheric microplastics. Many atmospheric particles can act as ice nucleators, thereby affecting the microphysical and radiative properties of clouds and, hence, the radiative balance of the Earth. The present study focused on the ice-nucleating ability of fibers from clothing textiles (CTs), which are commonly shed from the normal wear of apparel items. Results from immersion ice nucleation experiments showed that CTs were effective ice nucleators active from -6 to -12 °C, similar to common biological ice nucleators. However, subsequent lysozyme and hydrogen peroxide digestion stripped the ice nucleation properties of CTs, indicating that ice nucleation was biological in origin. Microscopy confirmed the presence of biofilms (i.e., microbial cells attached to a surface and enclosed in an extracellular polysaccharide matrix) on CTs. If present in sufficient quantities in the atmosphere, biological particles (biofilms) attached to fibrous materials could contribute significantly to atmospheric ice nucleation.

Ice-binding proteins (IBPs) are expressed in various organisms for several functions, such as protecting them from freezing and freeze injuries. Via adsorption on ice surfaces, IBPs depress ice growth and recrystallization and affect nucleation and ice shaping. IBPs have shown promise in mitigating ice growth under moderate supercooling conditions, but their functionality under cryogenic conditions has been less explored. In this study, we investigate the impact of two types of antifreeze proteins (AFPs): type III AFP from fish and a hyperactive AFP from an insect, the Tenebrio molitor AFP, in vitrified dimethylsulfoxide (DMSO) solutions. We report that these AFPs depress devitrification at -80 °C. Furthermore, in cases where devitrification does occur, AFPs depress ice recrystallization during the warming stage. The data directly demonstrate that AFPs are active at temperatures below the regime of homogeneous nucleation. This research paves the way for exploring AFPs as potential enhancers of cryopreservation techniques, minimizing ice-growth-related damage, and promoting advancements in this vital field.

To inhibit the quality deterioration caused by the frozen storage of surimi products, this work investigated the effect of freezing methods, including raw-freezing-setting-heating, raw-setting-freezing-heating, and raw-setting-heating-freezing, on quality changes in surimi gel. The moisture loss, physical-chemical properties, and protein structure conformation of surimi gel derived from Bombay duck (BD) were assessed following frozen storage periods of 20, 40, and 60 days. The findings suggest that the raw-setting-heating-freezing method yielded optimal surimi gel properties with extended frozen storage time. Employing this approach led to a reduction in thawing loss, while cooking loss remained constant. After 60 days of frozen storage, the hardness exhibited an initial increase followed by a subsequent decrease, and water-holding capacity increased to 68.2%. Notably, the impact on surimi gel during the late stage of frozen storage was more pronounced throughout the formation of ice crystals, resulting in decreased disulfide bond content. Scanning hematoxylin-eosin (HE) staining slices of samples following thawing and heating demonstrated that the raw-setting-heating-freezing method could better resist the effect of ice crystals in frozen storage period on surimi tissue, while the gel on setting process could delay the erosion imposed on by ice crystals during frozen storage. This study provides a scientific foundation for the industrialization on frozen BD surimi products., (© 2024 Wiley Periodicals LLC.)