An aberrant fused in sarcoma liquid droplet of amyotrophic lateral sclerosis pathological variant, R495X, accelerates liquid-solid phase transition.

Intracellular aggregation of fused in sarcoma (FUS) is associated with the pathogenesis of familial amyotrophic lateral sclerosis (ALS). Under stress, FUS forms liquid droplets via liquid-liquid phase separation (LLPS). Two types of wild-type FUS LLPS exist in equilibrium: low-pressure LLPS (LP-LLPS) and high-pressure LLPS (HP-LLPS); the former dominates below 2 kbar and the latter over 2 kbar. Although several disease-type FUS variants have been identified, the molecular mechanism underlying accelerated cytoplasmic granule formation in ALS patients remains poorly understood. Herein, we report the reversible formation of the two LLPS states and the irreversible liquid-solid transition, namely droplet aging, of the ALS patient-type FUS variant R495X using fluorescence microscopy and ultraviolet-visible absorption spectroscopy combined with perturbations in pressure and temperature. Liquid-to-solid phase transition was accelerated in the HP-LLPS of R495X than in the wild-type variant; arginine slowed the aging of droplets at atmospheric conditions by inhibiting the formation of HP-LLPS more selectively compared to that of LP-LLPS. Our findings provide new insight into the mechanism by which R495X readily forms cytoplasmic aggregates. Targeting the aberrantly formed liquid droplets (the HP-LLPS state) of proteins with minimal impact on physiological functions could be a novel therapeutic strategy for LLPS-mediated protein diseases.

The Enrichment of Docosahexaenoic Acid from Microalgal Oil by Urea Complexation via Rotary-evaporation Crystallization.

Urea complexation is a widely used method for enriching polyunsaturated fatty acids, and cooling is the traditional approach for urea crystallization. This study aimed to investigate the potential of rotary-evaporation under vacuum as an alternative method for urea crystallization in urea complexation to enrich docosahexaenoic acid (DHA). DHA-containing microalgal oil was converted to ethyl esters (EE) as the raw material. In comparison to cooling, rotary-evaporation crystallization, as a post-treatment method for urea complexation, led to higher DHA contents in the non-urea included fractions. The ratios of urea to EE converted from DHA-containing microalgal oil was found to be the primary factors influencing urea complexation when using rotary-evaporation crystallization. Through an orthogonal test, optimal process conditions were determined, including a urea/EE ratio of 2, an ethanol/urea ratio of 7, and a rotary-evaporation temperature of 75℃. Under these conditions, a concentrate containing more than 90% DHA could be obtained.

Exploring the Physical Properties of Lipid Membranes with Polyhydroxy Oxanorbornane Head Group Using NBD-Conjugated and DPH Fluorescent Probes.

The present study focuses on exploring the physical properties of lipid membranes based on the polyhydroxy oxanorbornane (PH-ONB) headgroup, designed as synthetic analogues of naturally occurring archaeal lipid membranes. Specifically, we study two variants of PH-ONB headgroup-based lipids differing in the number of hydroxy groups present in the headgroup, with one having two hydroxy groups (ONB-2OH) and the other having three (ONB-3OH). These lipids form stable bilayer membranes. The study begins with a comprehensive analysis of the fluorescence characteristics of nitrobenzoxadiazole (NBD)-tagged ONB-based lipids in different solvent environments and within a model lipid membrane 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC). Subsequently, the physical properties of the ONB-based membranes were examined by using an NBD-tagged ONB-based probe and a commonly used extrinsic 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescent probe. The steady-state and time-resolved fluorescence properties of the NBD-tagged ONB-based probe and DPH were used to compare the physical properties of the ONB-based membranes, including polarity, fluidity, phase transition, order, hydration, location, heterogeneity, and rotational diffusion. The solid gel to liquid crystalline phase transition temperatures of ONB-2OH and ONB-3OH lipid membranes are found to be (68 ± 1) °C and (74 ± 1) °C, respectively. The variation in organization (size), fluidity, and phase transition temperature of ONB-based lipid membranes is explained by the extent of hydrogen bonding interactions between lipid head groups. ONB-based membranes exhibit characteristics similar to those of phospholipid membranes and possess a notably high phase transition temperature. These properties make them a promising and cost-effective synthetic alternative to archaeal lipid membranes with a wide range of potential applications.

Fragility and Tendency to Crystallization for Structurally Related Compounds.

The present study was designed to investigate the physical stability of three organic materials with similar chemical structures. The examined compounds revealed completely different crystallization tendencies in their supercooled liquid states and were classified into three distinct classes based on their tendency to crystallize. (S)-4-Benzyl-2-oxazolidinone easily crystallizes during cooling from the melt; (S)-4-Benzylthiazolidine-2-thione does not crystallize during cooling from the melt, but crystallizes easily during subsequent reheating above Tg; and (S)-4-Benzyloxazolidine-2-thione does not crystallize either during cooling from the melt or during reheating. Such different tendencies to crystallize are observed despite the very similar chemical structures of the compounds, which only differ in oxide or sulfur atoms in one of their rings. We also studied the isothermal crystallization kinetics of the materials that were shown to transform into a crystalline state. Molecular dynamics and thermal properties were thoroughly investigated using broadband dielectric spectroscopy, as well as conventional and temperature-modulated differential scanning calorimetry in the wide temperature range. It was found that all three glass formers have the same dynamic fragility (m = 93), calculated directly from dielectric structural relaxation times. This result verifies that dynamic fragility is not related to the tendency to crystallize. In addition, thermodynamic fragility predictions were also made using calorimetric data. It was found that the thermodynamic fragility evaluated based on the width of the glass transition, observed in the temperature dependence of heat capacity, correlates best with the tendency to crystallize.

Interfacial crystallized oleogel emulsion with improved freeze-thaw stability and tribological properties: Influence of cooling rate.

In this study, the influence of cooling rate on the freeze-thaw stability, rheological and tribological properties of interfacial crystalized oleogel emulsion was investigated. Results showed that slower cooling rate could promote formation of larger crystals and stronger network in oleogels. Additionally, oleogel emulsions showed higher freeze-thaw stability than those stabilized solely by emulsifiers. The slower cooling rate resulted in larger crystals adsorbed at the droplet surface. This led to greater steric hindrance that prevented the migration of oil droplets with higher resistance to disruption by ice crystals. The rheological and tribological measurements suggested that with appropriate amount of crystals, the tribological properties were better maintained for emulsions prepared at slow cooling rate after freeze-thaw treatment. This strategy greatly enriched oleogel emulsion formulations and provided important clues for potential applications in food products involved with freeze-thaw treatment.

Energy based techno-economic and environmental feasibility study on PV/T and PV/T heat pump system with phase change material-a numerical comparative study.

A sustainable, affordable, and eco-friendly solution has been proposed to address water heating, electricity generation, space cooling, and photovoltaic (PV) cooling requirements in scorching climates. The photovoltaic thermal system (PV/T) and the direct expansion PV/T heat pump (PV/T DXHP) were numerically studied using MATLAB. A butterfly serpentine flow collector (BSFC) and phase change material (PCM) were assimilated in the PV system and MATLAB model was developed to evaluate the economic and enviroeconomic performance of the PV/T water system (PV/T-W), PV/T PCM water system (PV/T PCM-W), the PV/T DXHP system, and the PV/T PCM heat pump system (PV/T-PCM-DXHP). In this study, annual energy production, socioeconomic factors, enviro-economic indicators, and environmental characteristics are assessed and compared. Also, an economic, environmental, and enviro-economic analysis was conducted to assess the commercial viability of the suggested system. The PV/T PCM-DXHP demonstrated the highest electrical performance of 53.69%, which is comparatively higher than the other three configurations. The discounted levelized cost of energy (DLCOE) and payback period (DPP) of the PV/T PCM-DXHP were ₹2.87 per kW-h and 3-4 years, respectively, resulting in a total savings of ₹67,7403 over its lifetime. Furthermore, installing this system mitigated 280.72 tonnes of CO2 emissions and saved the mitigation cost by ₹329,700 throughout its operational lifecycle.

Irreversible lipid phase transition detected in a porcine oocyte at chilling.

Under physiological conditions, the membranes and lipid droplets of germ cells are in a conformationally disordered phase. Typically, during cooling, lipids undergo the transition to ordered phases and, upon heating, melt into a disordered phase. In this communication, we report the lipid phase transition in lipid droplets observed in porcine oocytes. Upon cooling, a sharp lipid phase transition from conformationally disordered to ordered state was detected within the temperature range between 20 and 15 °C. Subsequent heating to 45 °C does not return lipids to their original phase state. To the best of our knowledge, this is the first observation of an irreversible phase transition in lipid droplets of biological cells with native lipid composition.

Adaptive cooling strategy via human hair: High optothermal conversion efficiency of solar radiation into thermal dissipation.

Natural species have developed complex nanostructures in a hierarchical pattern to control the absorption, reflection, or transmission of desired solar and infrared wavelengths. This bio-inspired structure is a promising method to manipulating solar energy and thermal management. In particular, human hair is used in this article to highlight the optothermal properties of bio-inspired structures. This study investigated how melanin, an effective solar absorber, and the structural morphology of aligned domains of keratin polymer chains, leading to a significant increase in solar path length, which effectively scatter and absorb solar radiation across the hair structure, as well as enhance thermal ramifications from solar absorption by fitting its radiative wavelength to atmospheric transmittance for high-yield radiative cooling with realistic human body thermal emission.

Impact of Processing Methods on the Physico-chemical Properties of Posaconazole Amorphous Solid Dispersions.

BACKGROUND & PURPOSE: Different methods have been exploited to generate amorphous solid dispersions (ASDs) of poorly water-soluble drugs. However, the impact of processing methods on drug stability and dissolution hasn't been studied extensively. The purpose of the current study is to investigate the impact of the two common ASD processing methods, hot-melt extrusion (HME) and spray drying, on the chemical/physical stability and supersaturation of Posaconazole (Posa) based ASDs. METHODS & RESULTS: ASDs with 25% drug loading in hydroxypropylmethylcellulose acetate succinate were prepared using HME, and two types of spray dryers, a Procept Sprayer (ASD-Procept) and a Nano Sprayer (ASD-Nano). The relative physical stability of these ASDs upon exposure to heat and crystalline API seeding followed the order: ASD-Nano > ASD-Procept ≈HME. ASD-Procept and ASD-Nano showed similar chemical stability, slightly less stable than HME under 40°C/75%RH. All three ASDs demonstrated similar supersaturation induction times, and de-supersaturation kinetics with or without crystalline seeds. CONCLUSIONS: Posa ASDs prepared via spray drying were chemically less stable compared with HME, which can be attributed to their smaller particle size and hollow structure allowing oxygen penetration. For ASD-Procept and HME, the detailed phase changes involving recrystallization of amorphous Posa and a solid-solid phase transition from Posa Form I to Form Ia during the seed-induced studies were proposed. Similar dissolution and supersaturation-precipitation kinetics of three Posa ASDs indicated that any residual nanocrystals in the bulk ASDs were not enough to induce crystallization to differentiate ASDs made by three processing methods.

Microscopic Cracks Modulate Nucleation and Solid-State Crystallization Tendency of Amorphous Celecoxib.

Drugs have been classified as fast, moderate, and poor crystallizers based on their inherent solid-state crystallization tendency. Differential scanning calorimetry-based heat-cool-heat protocol serves as a valuable tool to define the solid-state crystallization tendency. This classification helps in the development of strategies for stabilizing amorphous drugs. However, microscopic characteristics of the samples were generally overlooked during these experiments. In the present study, we evaluated the influence of microscopic cracks on the crystallization tendency of a poorly water-soluble model drug, celecoxib. Cracks developed in the temperature range of 0-10 °C during the cooling cycle triggered the subsequent crystallization of the amorphous phase. Nanoindentation study suggested minimal differences in mechanical properties between samples, although the cracked sample showed relatively inhomogeneous mechanical properties. Nuclei nourishment experiments suggested crack-assisted nucleation, which was supported by Raman data that revealed subtle changes in intermolecular interactions between cracked and uncracked samples. Celecoxib has been generally classified as class II, i.e., a drug with moderate crystallization tendency. Interestingly, classification of amorphous celecoxib may change depending on the presence or absence of cracks in the amorphous sample. Hence, subtle events such as microscopic cracks should be given due consideration while defining the solid-state crystallization tendency of drugs.

Encounter between Gyroid and Lamellae in Janus Colloidal Particles Self-Assembled by a Rod-Coil Block Copolymer.

Controlling the internal structure of block copolymer (BCP) particles has a significant influence on its functionalities. Here, a structure-controlling method is proposed to regulate the internal structure of BCP Janus colloidal particles using different surfactants. Different microphase separation processes take place in two connected halves of the Janus particles. An order-order transition between gyroid and lamellar phases is observed in polymeric colloids. The epitaxial growth during the structural transformation from gyroid to lamellar phase undergoes a two-layered rearrangement to accommodate the interdomain spacing mismatch between these two phases. This self-assembly behavior can be ascribed to the preferential wetting of BCP chains at the interface, which can change the chain conformation of different blocks. The Janus colloidal particles can further experience a reversible phase transition by restructuring the polymer particles under solvent vapor. It is anticipated that the new phase behavior found in Janus particles can not only enrich the self-assembly study of BCPs but also provide opportunities for various applications based on Janus particles with ordered structures.

Interfacial Aß fibril formation is modulated by the disorder-order state of the lipids: The concept of the physical environment as amyloid inductor in biomembranes.

The behavior of amphiphilic molecules such as lipids, peptides and their mixtures at the air/water interface allow us to evaluate and visualize the arrangement formed in a confined and controlled surface area. We have studied the surface properties of the zwitterionic DPPC lipid and Aß(1-40) amyloid peptide in mixed films at different temperatures (from 15 to 40 °C). In this range of temperature the surface properties of pure Aß(1-40) peptide remained unchanged, whereas DPPC undergoes its characteristic liquid-expanded â†’ liquid-condensed bidimensional phase transition that depends on the temperature and lateral pressure. This particular property of DPPC makes it possible to dynamically study the influence of the lipid phase state on amyloid structure formation at the interface in a continuous, isothermal and abrupt change on the environmental condition. As the mixed film is compressed the fibril-like structure of Aß(1-40) is triggered specifically in the liquid-expanded region, independently of temperature, and it is selectively excluded from the well-visible liquid condensed domains of DPPC. The Aß amyloid fibers were visualized by using BAM and AFM and they were Thio T positive. In mixed DPPC/Aß(1-40) films the condensed domains (in between 11 mN/m to 20 mN/m) become irregular probably due to the fibril-like structures is imposing additional lateral stress sequestering lipid molecules in the surrounding liquid-expanded phase to self-organize into amyloids.

Multiplex Ultrasound Imaging of Perfluorocarbon Nanodroplets Enabled by Decomposition of Postvaporization Dynamics.

Despite the real-time, nonionizing, and cost-effective nature of ultrasound imaging, there is a dearth of methods to visualize two or more populations of contrast agents simultaneously─a technique known as multiplex imaging. Here, we present a new approach to multiplex ultrasound imaging using perfluorocarbon (PFC) nanodroplets. The nanodroplets, which undergo a liquid-to-gas phase transition in response to an acoustic trigger, act as activatable contrast agents. This work characterized the dynamic responses of two PFC nanodroplets with boiling points of 28 and 56 °C. These characteristic responses were then used to demonstrate that the relative concentrations of the two populations of PFC nanodroplets could be accurately measured in the same imaging volume within an average error of 1.1%. Overall, the findings indicate the potential of this approach for multiplex ultrasound imaging, allowing for the simultaneous visualization of multiple molecular targets simultaneously.

Alkylbenzoic and Alkyloxybenzoic Acid Blending for Expanding the Liquid Crystalline State and Improving Its Rheology.

Thermotropic mesogens typically exist as liquid crystals (LCs) in a narrow region of high temperatures, making lowering their melting point with the temperature expansion of the mesophase state an urgent task. Para-substituted benzoic acids can form LCs through noncovalent dimerization into homodimers via hydrogen bonds, whose strength and, consequently, the temperature region of the mesophase state can be potentially altered by creating asymmetric heterodimers from different acids. This work investigates equimolar blends of p-n-alkylbenzoic (kBA, where k is the number of carbon atoms in the alkyl radical) and p-n-alkyloxybenzoic (kOBA) acids by calorimetry and viscometry to establish their phase transitions and regions of mesophase existence. Non-symmetric dimerization of acids leads to the extension of the nematic state region towards low temperatures and the appearance of new monotropic and enantiotropic phase transitions in several cases. Moreover, the crystal-nematic and nematic-isotropic phase changes have a two-step character for some acid blends, suggesting the formation of symmetric and asymmetric associates from heterodimers. The mixing of 6BA and 8OBA most strongly extends the region of the nematic state towards low temperatures (from 95-114 °C and 108-147 °C for initial homodimers, respectively, to 57-133 °C for the resulting heterodimer), whereas the combination of 4OBA and 5OBA gives the most extended high-temperature nematic phase (up to 156 °C) and that of 6BA and 9OBA (or 12OBA) provides the existence of a smectic phase at the lowest temperatures (down to 51 °C).

Anomalous lateral diffusion of lipids during the fluid/gel phase transition of a lipid membrane.

A lipid membrane undergoes a phase transition from fluid to gel phase upon changing external thermodynamic conditions, such as decreasing temperature and increasing pressure. Extremophilic organisms face the challenge of preventing this deleterious phase transition. The main focus of their adaptive strategy is to facilitate effective temperature sensing through sensor proteins, relying on the drastic changes in packing density and membrane fluidity during the phase transition. Although the changes in packing density parameters due to the fluid/gel phase transition are studied in detail, the impact on membrane fluidity is less explored in the literature. Understanding the lateral diffusive dynamics of lipids in response to temperature, particularly during the fluid/gel phase transition, is albeit crucial. Here we have simulated the phase transition of a single component lipid membrane composed of dipalmitoylphosphatidylcholine (DPPC) lipids using a coarse-grained (CG) model and studied the changes of the structural and dynamical properties. It is observed that near the phase transition point, both fluid and gel phase domains coexist together. The dynamics remains highly non-Gaussian for a long time even when the mean square displacement reaches the Fickian regime at a much earlier time. This Fickian yet non-Gaussian diffusion (FnGD) is a characteristic of a highly heterogeneous system, previously observed for the lateral diffusion of lipids in raft mimetic membranes having liquid-ordered and liquid-disordered phases co-existing together. We have analyzed the molecular trajectories and calculated the jump-diffusion of the lipids, stemming from sudden jump translations, using a translational jump-diffusion (TJD) approach. An overwhelming contribution of the jump-diffusion of the lipids is observed suggesting anomalous diffusion of lipids during fluid/gel phase transition of the membrane. These results are important in unravelling the intricate nature of lipid diffusion during the phase transition of the membrane and open up a new possibility of investigating the most significant change of membrane properties during phase transition, which can be effectively sensed by proteins.

Enhanced heat transfer by medical gauze for cell vitrification with French straw.

BACKGROUND: Film boiling occurs in the cooling process of samples with liquid nitrogen, which limits heat transfer and decreases the cooling rate. OBJECTIVE AND METHODS: The study developed a method to enhance convective heat transfer by wrapping the French straw (FS) with a layer of medical gauze upon cooling to eliminate film boiling. A numerical model was used to study the thermodynamic mechanism in the method. RESULTS: Numerical simulation based on heat transfer and crystallization equations indicated that wrapping the FS with medical gauze could suppress film boiling. Experimental verification showed the increased cooling rate and better cell survival when wrapped FS was used. CONCLUSION: Numerical simulation and experimental verification demonstrated the efficacy of wrapping FS with medical gauze for better cell cryopreservation. Doi.org/10.54680/fr23510110312.

RNAs undergo phase transitions with lower critical solution temperatures.

Co-phase separation of RNAs and RNA-binding proteins drives the biogenesis of ribonucleoprotein granules. RNAs can also undergo phase transitions in the absence of proteins. However, the physicochemical driving forces of protein-free, RNA-driven phase transitions remain unclear. Here we report that various types of RNA undergo phase separation with system-specific lower critical solution temperatures. This entropically driven phase separation is an intrinsic feature of the phosphate backbone that requires Mg2+ ions and is modulated by RNA bases. RNA-only condensates can additionally undergo enthalpically favourable percolation transitions within dense phases. This is enabled by a combination of Mg2+-dependent bridging interactions between phosphate groups and RNA-specific base stacking and base pairing. Phase separation coupled to percolation can cause dynamic arrest of RNAs within condensates and suppress the catalytic activity of an RNase P ribozyme. Our work highlights the need to incorporate RNA-driven phase transitions into models for ribonucleoprotein granule biogenesis.

Regulation of Protein Flexibility and Promoting the Cod Protein Gel Formation Using Ultrasound Treatment.

In order to obtain a soft-textured protein gel suitable for the elderly, the cod protein gel was prepared by improving the protein flexibility under ultrasound treatment. It has been found that the increase in ultrasonic power, protein flexibility, particle size, ζ-potential, surface hydrophobicity, and α-helix content of preheated cod protein exhibited an increasing trend. The improvement of protein flexibility promoted uniformity and density of the gel network, water retention, and texture properties. The flexibility of preheated cod protein increased to 0.189, the water holding capacity of the gel reached up to 99.41%, and the hardness increased to 49.12 g, as the ultrasonic power level increased to 400 W. Protein flexibility was correlated well with the cohesiveness of the gel. The storage modulus (G') initially decreased and then increased during the heating-cooling process. The attractive forces forming between the flexible protein molecules during cooling in the ultrasound treatment groups promoted protein self-assembly aggregation and formed the cod protein gel. The gel obtained at 100-400 W could be categorized as Level 6─soft and bite-sized according to the International Dysphagia Diet Standardization Initiative (IDDSI) framework, indicating that the cod protein gel has potential as an easy-to-swallow diet for the elderly.

Multi-scale molecular simulation of random peptide phase separation and its extended-to-compact structure transition driven by hydrophobic interactions.

Intrinsically disordered proteins (IDPs) often undergo liquid-liquid phase separation (LLPS) and form membraneless organelles or protein condensates. One of the core problems is how do electrostatic repulsion and hydrophobic interactions in peptides regulate the phase separation process? To answer this question, this study uses random peptides composed of positively charged arginine (Arg, R) and hydrophobic isoleucine (Ile, I) as the model systems, and conduct large-scale simulations using all atom and coarse-grained model multi-scale simulation methods. In this article, we investigate the phase separation of different sequences using a coarse-grained model. It is found that the stronger the electrostatic repulsion in the system, the more extended the single-chain structure, and the more likely the system forms a low-density homogeneous phase. In contrast, the stronger the hydrophobic effect of the system, the more compact the single-chain structure, the easier phase separation, and the higher the critical temperature of phase separation. Overall, by taking the random polypeptides composed of two types of amino acid residues as model systems, this study discusses the relationship between the protein sequence and phase behaviour, and provides theoretical insights into the interactions within or between proteins. It is expected to provide essential physical information for the sequence design of functional IDPs, as well as data to support the diagnosis and treatment of the LLPS-associated diseases.

Freeze-Induced Phase Transition and Local Pressure in a Phospholipid/Water System: Novel Insights Were Obtained from a Time/Temperature Resolved Synchrotron X-ray Diffraction Study.

Water-to-ice transformation results in a 10% increase in volume, which can have a significant impact on biopharmaceuticals during freeze-thaw cycles due to the mechanical stresses imparted by the growing ice crystals. Whether these stresses would contribute to the destabilization of biopharmaceuticals depends on both the magnitude of the stress and sensitivity of a particular system to pressure and sheer stresses. To address the gap of the "magnitude" question, a phospholipid, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), is evaluated as a probe to detect and quantify the freeze-induced pressure. DPPC can form several phases under elevated pressure, and therefore, the detection of a high-pressure DPPC phase during freezing would be indicative of a freeze-induced pressure increase. In this study, the phase behavior of DPPC/water suspensions, which also contain the ice nucleation agent silver iodide, is monitored by synchrotron small/wide-angle X-ray scattering during the freeze-thaw transition. Cooling the suspensions leads to heterogeneous ice nucleation at approximately -7 °C, followed by a phase transition of DPPC between -11 and -40 °C. In this temperature range, the initial gel phase of DPPC, Lß', gradually converts to a second phase, tentatively identified as a high-pressure Gel III phase. The Lß'-to-Gel III phase transition continues during an isothermal hold at -40 °C; a second (homogeneous) ice nucleation event of water confined in the interlamellar space is detected by differential scanning calorimetry (DSC) at the same temperature. The extent of the phase transition depends on the DPPC concentration, with a lower DPPC concentration (and therefore a higher ice fraction), resulting in a higher degree of Lß'-to-Gel III conversion. By comparing the data from this study with the literature data on the pressure/temperature Lß'/Gel III phase boundary and the lamellar lattice constant of the Lß' phase, the freeze-induced pressure is estimated to be approximately 0.2-2.6 kbar. The study introduces DPPC as a probe to detect a pressure increase during freezing, therefore addressing the gap between a theoretical possibility of protein destabilization by freeze-induced pressure and the current lack of methods to detect freeze-induced pressure. In addition, the observation of a freeze-induced phase transition in a phospholipid can improve the mechanistic understanding of factors that could disrupt the structure of lipid-based biopharmaceuticals, such as liposomes and mRNA vaccines, during freezing and thawing.