Revealing isochoric water nucleation: a visual study.

The phenomena of water freezing at constant volume, or isochoric, is becoming more and more fascinating. However, because the system is subjected to extremely high pressures, it is exceedingly challenging to investigate it visually. Fewer properties have been found visually up till now, but many have been found through other means. Nevertheless, we were able to design a reactor so that it could be observed through a microscope as water was frozen and thawed at constant volume, reaching temperatures as low as - 12 °C and pressures up to 129 MPa. In this study, we observed critical characteristics visually, focusing on the location of the ice nucleus, its shape, and dynamics. Phase transitions from liquid to solid state are essential mechanisms in the physical sciences. The creation of ice stands as the quintessential and pervasive example of nucleation, playing a central role in diverse disciplines such as geology, biology, aviation, and climate research.

Prototype isochoric preservation device for large organs.

This paper presents the design and prototype of a constant volume (isochoric) vessel that can be used for the preservation of large organs in a supercooled state. This prototype is a preliminary version of a more advanced design. The device consists of a cooling bath operated by a mechanical vapor compression refrigeration unit and an isochoric chamber made of stainless steel. The preservation of organs using supercooling technology in an isochoric chamber requires a continuous temperature and pressure monitoring. While the device was initially designed for pig liver experiments, its innovative design and preservation capabilities suggest potential applications for preserving other organs as well. The isochoric reactor may be used to accommodate a variety of organ types, opening the door for further research into its multi-organ preservation capabilities. All the design details are presented in this study with the purpose of encouraging researchers in the field to build their own devices, and by this to improve the design. We chose to design the device for isochoric supercooling as the method of preservation to avoid the ice formation.

Isochoric Supercooling Organ Preservation System.

This technical paper introduces a novel organ preservation system based on isochoric (constant volume) supercooling. The system is designed to enhance the stability of the metastable supercooling state, offering potential long-term preservation of large biological organs at subfreezing temperatures without the need for cryoprotectant additives. Detailed technical designs and usage protocols are provided for researchers interested in exploring this field. The paper also presents a control system based on the thermodynamics of isochoric freezing, utilizing pressure monitoring for process control. Sham experiments were performed using whole pig liver sourced from a local food supplier to evaluate the system's ability to sustain supercooling without ice nucleation for extended periods. The results demonstrated sustained supercooling without ice nucleation in pig liver tissue for 24 and 48 h. These findings suggest the potential of this technology for large-volume, cryoprotectant-free organ preservation with real-time control over the preservation process. The simplicity of the isochoric supercooling device and the design details provided in the paper are expected to serve as encouragement for other researchers in the field to pursue further research on isochoric supercooling. However, final evidence that these preserved organs can be successfully transplanted is still lacking.

Preservation of black grapes by isochoric freezing.

Fruits are perishable. It's crucial to have an efficient preservation technique that may extend storage duration while maintaining the physical quality and nutritional values to avoid wastage. The majority of long-term storage solutions for fruits use refrigeration. In this study, we evaluate the potential of isochoric freezing as an alternative method of preservation for black grapes (Vitis vinifera L.). We compare the properties of black grapes preserved for 7 days in trehalose solution at -4 °C in isochoric conditions (average pressure 34.2 MPa) with those of fresh black grapes and with grapes preserved isobarically in four conditions (room temperature, in the fridge, in the freezer, and in a plastic bag filled with trehalose solution). The results indicate that grapes preserved by isochoric freezing at temperatures below the freezing point of water do not lose weight; on the contrary, they resulted in a very small (2%) weight gain. Freezing under isochoric conditions did not result in significant changes in terms of macroscopic appearance, colour, firmness, °Brix values, or pH. We consider that isochoric freezing has the potential to be used as a preservation method for grapes while maintaining physicochemical parameters similar to those of fresh fruits.

An exploratory study on isochoric supercooling preservation of the pig liver.

This study was motivated by the increasing interest in finding ways to preserve organs in a supercooled state for transplantation. Previous research with small volumes suggests that the isochoric (constant volume) thermodynamic state enhances the stability of supercooled solutions. The primary objective of this study was to investigate the feasibility of storing a large organ, such as the pig liver, in a metastable isochoric supercooled state for clinically relevant durations. To achieve this, we designed a new isochoric technology that employs a system consisting of two domains separated by an interior boundary that can transfer heat and pressure, but not mass. The liver is preserved in one of these domains in a solution with an intracellular composition, which is in osmotic equilibrium with the liver. Pressure is used to monitor the thermodynamic state of the isochoric chamber. In this feasibility study, two pig livers were preserved in the device in an isochoric supercooled state at -2°C. The experiments were terminated voluntarily, one after 24 h and the other after 48 h of supercooling preservation. Pressure measurements indicated that the livers did not freeze during the isochoric supercooling preservation. This is the first proof that organs as large as the pig liver can remain supercooled for extended periods of time in an isotonic solution in an isochoric system, despite an increased probability of ice nucleation with larger volumes. To serve as controls and to test the ability of pressure monitoring to detect freezing in the isochoric chamber, an experiment was designed in which two pig livers were frozen at -2°C for 24 h and the pressure monitored. Histological examination with H&E stains revealed that the supercooled liver maintained a normal appearance, even after 48 h of supercooling, while tissues in livers frozen to -2°C were severely disrupted by freezing after 24 h.

Temperature-pressure correlations of cryoprotective additives for the design of constant volume cryopreservation protocols.

In the recent years, the use of constant volume (isochoric) cryopreservation, in medicine and biotechnology has captured more attention from the research community and now there is an increasing interest in the use of this new technology. It has been established that the thermodynamics of isochoric freezing is different from that of isobaric (constant pressure) freezing. This study provides researchers in the field experimental results for various compositions of cryoprotectants commonly used in isobaric cryopreservation, in terms of temperature-pressure-molar concentration correlation. It also reveals experimental isochoric thermodynamic data for the following cryoprotectants, commonly used in isobaric cryopreservation: dimethyl sulfoxide, trehalose, ethylene glycol and diethylene glycol. Currently, the data on the pressure-temperature correlation in an isochoric system of cryoprotectants used in isobaric cryopreservation is not available. Our new experimental results indicate that the studied concentrations for each of the CPAs, lower and expands the range of temperatures in which cryopreservation by isochoric freezing can be safely practiced. We consider that these experiments will aid researchers developing new isochoric cryopreservation protocols.

Exergetic Analysis of a Cryogenic Air Separation Unit.

This case study analyzes a cryogenic air separation unit (ASU) with a production of V˙O2=58,300 [m3Nh] of gaseous oxygen with a concentration greater than 98.5%, operating in Romania on a steel plant platform. The goal of the paper is to provide an extensive model of exergetic analysis that could be used in an optimization procedure when decisional parameters are changed or structural design modifications are implemented. For each key part of the Air Separation Unit, an exergetic product and fuel were defined and, based on their definition, the coefficient of performance of each functional zone was calculated. The information about the magnitude of the exergetic losses offers solutions for their future recovery. The analysis of the exergy destructions suggests when it is worth making a larger investment. The exergetic analysis of the compression area of the ASU points out an exergy destruction and loss of 37% from the total plant's electrical energy input. The exergy loss with the heat transferred to the cooling system of compressors can be recovered; for the exergy destruction portion, the challenge between investment and operating costs should be considered. The exergy destruction of the air separation columns found the High Pressure Column (HPC) to be more destructive than the Low Pressure Column. The share of the exergy destruction in the total plant's electrical energy input is 8.3% for the HPC. The local COP of the HPC, calculated depending on the total exergy of the local product and fuel, is 62.66%. The calculus of the air separation column is performed with the ChemSep simulator.

Glucose and glycerol temperature-pressure correlations for the design of cryopreservation protocols in an isochoric system at subfreezing temperature.

There is growing interest in the use of isochoric (constant volume) freezing for cryopreservation of biological matter. The goal of this study is to generate fundamental experimental data on the pressure temperature relation during the freezing of an isochoric system of aqueous solutions of two compounds, glucose and glycerol. Glucose and glycerol are commonly used as cryoprotectants in conventional isobaric (constant pressure) cryopreservation protocols. Earlier studies have shown that the increase in pressure during isochoric freezing is detrimental to biological matter and limits the range of temperatures in which isochoric freezing can be used for preservation to temperatures corresponding to pressures below 40 MPa. In physiological saline solution this pressure corresponds to a temperature of - 4 °C. Our new experimental data shows that the addition of 2 M glycerol to the saline solution lowers the temperature at which the isochoric freezing pressure is 40 MPa to -11 °C, 3 M glycerol to - 16.5 °C, and 4 M glycerol to - 24.5 °C, thereby substantially expending the range of temperatures in which cryopreservation by isochoric freezing can be practiced.

Pressure in isochoric systems containing aqueous solutions at subzero Centigrade temperatures.

OBJECTIVE: Preservation of biological materials at subzero Centigrade temperatures, cryopreservation, is important for the field of tissue engineering and organ transplantation. Our group is studying the use of isochoric (constant volume) systems of aqueous solution for cryopreservation. Previous studies measured the pressure-temperature relations in aqueous isochoric systems in the temperature range from 0°C to - 20°C. The goal of this study is to expand the pressure-temperature measurement beyond the range reported in previous publications. MATERIALS AND METHODS: To expand the pressure-temperature measurements beyond the previous range, we have developed a new isochoric device capable of withstanding liquid nitrogen temperatures and pressures of up to 413 MPa. The device is instrumented with a pressure transducer than can monitor and record the pressures in the isochoric chamber in real time. Measurements were made in a temperature range from - 5°C to liquid nitrogen temperatures for various solutions of pure water and Me2SO (a chemical additive used for protection of biological materials in a frozen state and for vitrification (glass formation) of biological matter). Undissolved gaseous are is carefully removed from the system. RESULTS: Temperature-pressure data from - 5°C to liquid nitrogen temperature for pure water and other solutions are presented in this study. Following are examples of some, temperature-pressure values, that were measured in an isochoric system containing pure water: (- 20°C, 187 MPa); (-25°C, 216 MPa); (- 30°C, 242.3 MPa); (-180°C, 124 MPa). The data is consistent with the literature, which reports that the pressure and temperature at the triple point, between ice I, ice III and water is, - 21.993°C and 209.9 MPa, respectively. It was surprising to find that the pressure in the isochoric system increases at temperatures below the triple point and remains high to liquid nitrogen temperatures. Measurements of pressure-temperature relations in solutions of pure water and Me2SO in different concentrations show that, for concentrations in which vitrification is predicted, no increase in pressure was measured during rapid cooling to liquid nitrogen temperatures. However, ice formation either during cooling or warming to and from liquid nitrogen temperatures produced an increase in pressure. CONCLUSIONS: The data obtained in this study can be used to aid in the design of isochoric cryopreservation protocols. The results suggest that the pressure measurement is important in the design of "constant volume" systems and can provide a simple means to gain information on the occurrence of vitrification and devitrification during cryopreservation processes of aqueous solutions in an isochoric system.

A comparison of freezing-damage during isochoric and isobaric freezing of the potato.

BACKGROUND: Freezing is commonly used for food preservation. It is usually done under constant atmospheric pressure (isobaric). While extending the life of the produce, isobaric freezing has detrimental effects. It causes loss of food weight and changes in food quality. Using thermodynamic analysis, we have developed a theoretical model of the process of freezing in a constant volume system (isochoric). The mathematical model suggests that the detrimental effects associated with isobaric freezing may be reduced in an isochoric freezing system. To explore this hypothesis, we performed a preliminary study on the isochoric freezing of a produce with which our group has experience, the potato. METHOD: Experiments were performed in an isochoric freezing device we designed. The device is robust and has no moving parts. For comparison, we used a geometrically identical isobaric freezing device. Following freezing and thawing, the samples were weighed, examined with colorimetry, and examined with microscopy. RESULTS: It was found that potatoes frozen to -5 °C in an isochoric system experienced no weight loss and limited enzymatic browning. In contrast the -5 °C isobaric frozen potato experienced substantial weight loss and substantial enzymatic browning. Microscopic analysis shows that the structural integrity of the potato is maintained after freezing in the isochoric system and impaired after freezing in the isobaric system. DISCUSSION: Tissue damage during isobaric freezing is caused by the increase in extracellular osmolality and the mechanical damage by ice crystals. Our thermodynamic analysis predicts that during isochoric freezing the intracellular osmolality remains comparable to the extracellular osmolality and that isochoric systems can be designed to eliminate the mechanical damage by ice. The results of this preliminary study seem to confirm the theoretical predictions. CONCLUSION: This is a preliminary exploratory study on isochoric freezing of food. We have shown that the quality of a food product preserved by isochoric freezing is better than the quality of food preserved to the same temperature in isobaric conditions. Obviously, more extensive research remains to be done to extend this study to lower freezing temperatures and other food items.

Isochoric and isobaric freezing of fish muscle.

We have recently shown that, a living organism, which succumbs to freezing to -4 °C in an isobaric thermodynamic system (constant atmospheric pressure), can survive freezing to -4 °C in an isochoric thermodynamic system (constant volume). It is known that the mechanism of cell damage in an isobaric system is the freezing caused increase in extracellular osmolality, and, the consequent cell dehydration. An explanation for the observed survival during isochoric freezing is the thermodynamic modeling supported hypothesis that, in the isochoric frozen solution the extracellular osmolality is comparable to the cell intracellular osmolality. Therefore, cells in the isochoric frozen organism do not dehydrate, and the tissue maintains its morphological integrity. Comparing the histology of: a) fresh fish white muscle, b) fresh muscle frozen to -5 °C in an isobaric system and c) fresh muscle frozen to -5 °C I in an isochoric system, we find convincing evidence of the mechanism of cell dehydration during isobaric freezing. In contrast, the muscle tissue frozen to -5 °C in an isochoric system appears morphologically identical to fresh tissue, with no evidence of dehydration. This is the first experimental evidence in support of the hypothesis that in isochoric freezing there is no cellular dehydration and therefore the morphology of the frozen tissue remains intact.

The nematode Caenorhabditis elegans survives subfreezing temperatures in an isochoric system.

This study is the first experimental evidence showing that a living multicellular organism, the nematode Caenorhabditis elegans, can survive subfreezing temperatures in an isochoric (constant volume) thermodynamic system, while immersed in a simple isotonic solution, without the addition of cryoprotectants. Some of the test conditions were more extreme than those found at the ice/water interface of the Antarctic subglacial Vostok lake. On earth, life takes place in an isobaric (constant pressure) environment. In isobaric systems, subfreezing temperature survival of organisms in nature and subfreezing temperature preservation of living material for biotechnology and medicine, is made possible by use of cryoprotective chemicals additives. Our theoretical thermodynamic studies suggested that in an isochoric system, living biological material could survive subfreezing temperatures, without any cryoprotective chemicals. By confirming the theoretical predictions, this paper suggests a new technology for subfreezing preservation of cells, organs and organisms of possible value for biotechnology and medicine as well as new possible mechanisms of living organism survival in nature.