Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Freezing”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 577 records · Page 32Linked to original sources
PubMed1998

Characterization of the sucrose/glycine/water system by differential scanning calorimetry and freeze-drying microscopy.

The objective of this study was to characterize the thermal properties of systems containing various ratios of amorphous and crystalline components using both differential scanning calorimetry (DSC) and freeze-drying microscopy. The glycine/sucrose system was used as a model system, since it is routinely used in protein formulations. DSC analysis revealed that the addition of glycine to sucrose solutions resulted in a decrease in the glass transition (T'g) of the system. The T'g of a pure sucrose solution (7% w/v) decreased from -32.3 to -51.5 degrees C for a mixture containing a sucrose/glycine ratio of 2:5. The glass transition of the sucrose/glycine mixture decreased linearly as more glycine was added to the system. This decrease in glass transition resulted in severe collapse during freeze-drying of these mixtures above T'g. However, collapse was not observed during freeze-drying if the DSC thermogram of the sucrose/glycine mixture exhibited a transition resulting from recrystallization of the amorphous glycine. Mixtures having a sucrose/glycine ratio of 3:4 and 2:5 had a glass transition of -48 degrees C and -51.5 degrees C, respectively. Despite their low glass transition temperatures, these samples freeze-dried readily at a product temperature > T'g using a fast freeze-drying cycle (primary drying at a shelf temperature of +20 degrees C and chamber pressure of 100 mTorr) without any sign of collapse. The crystallization of the amorphous glycine from the frozen mixture of sucrose and glycine provided support during freeze-drying which prevented the macroscopic collapse of the final product. Freeze-drying microscopy visually revealed the crystallization and allowed for prediction of cake quality upon lyophilization. Although the freeze-drying microscope is not as sensitive as the DSC in detecting all transitions (it cannot detect a glass transition), it clarifies the interpretation of DSC, and together they provide valuable information regarding the relevance of each of the transitions to the final freeze-dried product elegance.

Calorimetry, Differential Scanning↗
PubMed1979

Influence of percent egg yolk during cooling and freezing on survival of bovine spermatozoa.

The influence of percent egg yolk during cooling from 37 to 5 C on ability of sperm to withstand subsequent freezing was studied with semen from each of eight bulls. Treatments were arranged factorially and included .5, 1, 2, 4, 8, 16, and 32% egg yolk, cooling times of .5, 1, 2, and 4 h, and freezing in ampules and straws. Percent egg yolk did not influence motility or percent intact acrosomes when semen was frozen in ampules. For sperm in straws, both post-thaw percentages of motile sperm and intact acrosomes were similar for sperm cooled with .5, 1, 2, 4, or 8% egg yolk and greater than with 16 or 32% egg yolk. Optimal egg yolk during cooling was not dependent upon rate of cooling, regardless of how semen was packaged for freezing. The influence of percent egg yolk during freezing was investigated in a second study. Treatments included eight bulls; 1, 2, 4, 8, 16, and 32% egg yolk; freezing times from +5 to -130 C of 3.5, 20, and 40 min; and freezing in ampules or straws. Optimal egg yolk during freezing in either ampules or straws was independent of the rate of freezing. For sperm frozen in ampules, differences in motility or the percent intact acrosomes due to egg yolk were small. However, the use of only 1 or 2% egg yolk depressed motility whereas 16 or 32% egg yolk decreased the percentage of intact acrosomes. Thus, extremely high or low egg yolk should be avoided. For sperm in straws, survival was optimal when the extended semen contained 4 to 32% egg yolk during freezing.

Animals↗
PubMed1981

Effect of five cycle rapid freeze-thaw treatment in conjunction with various chemicals for the reduction of Salmonella typhimurium.

A five cycle rapid freeze-rapid thaw process was used in conjunction with chemicals to reduce numbers of Salmonella typhimurium cells on poultry meat. The second portion of chicken wings consisting of ulna and radius with attached skin and muscle was inoculated with 400 to 900 colony forming units (CFU/g) of a nalidixic acid resistant strain of S. typhimurium. Chemicals used were 20 ppm chlorine, 5% potassium sorbate, 5% lactic acid, and 5% calcium propionate. The wings were either sprayed with or dipped into all chemicals before the freeze-thaw process. Wings were also chemically treated and not subjected to the freeze-thaw process. Numbers of S. typhimurium were determined by the most probable number procedure. The relative effectiveness of combinations of chemicals and the freeze-thaw process was compared to a control with the following percentage reductions of numbers of S. typhimurium cells: lactic acid, 98%; calcium propionate, 96%; potassium sorbate, 96%; chlorine, 95%; and freeze-thaw process without chemicals, 95%. There were no statistically significant differences among the treatments. In pilot plant study simulating commercial conditions, a carbon dioxide freezer was used for the rapid freeze and a microwave oven was used for the rapid thaw. Treatment of wings with 5% lactic acid plus freeze-thaw process resulted in statistically significant fewer numbers of S. typhimurium cells when compared to the freeze-thaw process without chemical treatment or to wings chemically treated without the freeze-thaw process.

Animals↗
PubMed2004

[Cryobiology and pathologic lesions induced by freezing-thawing processes in prostatic tissue. Second part].

Cryosurgery is an emerging technology consisting on controlled freezing of tissues. Good results, maintained in the long-term, have been referred in the treatment of prostate adenocarcinoma. A role as possible substitute of partial nephrectomy in the treatment of renal adenocarcinomas smaller than 4-5 cm is under research. There is no discussion that freezing destroys cellular machinery and triggers several events the final result of which is cell death by necrosis and apoptosis. The decrease of temperature makes extracellular liquid crystallize and creates a hyperosmotic environment, which induces water to go out of the cell producing intracellular dehydration. Intracellular ice is created with fast freezing speeds being attributed the most destructive effect on biological tissues with irreparable damage. In blood vessels, it directly induces endothelial cell death and mechanical lesions of the endothelium; the consequence is the formation of thrombi that obstruct the lumen of the vessel. In the post-thawing phase there is an increase in free radicals formation and neutrophil activity, which induces cellular membrane lipids peroxidation and new endothelium lesions. Tissue destruction is determined by: minimal temperature achieved, freezing speeds, freezing phase duration, number of freezing-thawing cycles provided, and distance to the freezing focus. As we move away from the freezing focus cells are affected in different ways, and there are several mechanisms proposed to explain the lethal action induced by temperatures higher than--40 degrees C. In our series pathologic findings were: necrosis, hemorrhagic areas either developed or not, fibrosis, hyalinization and increases in the relative number of hematic capillaries, microscopic calcifications, basal cells hyperplasia, and transitional or squamous metaplasia. Residual cancer is localized in the areas less affected by freezing. It should be emphasize the scarce morbimortality associated with the procedure. It does not require ICU admission or blood transfusions, no cerebral vascular accidents (CVA) or heart events have been described, and mortality is null, so that it is feasible in high surgical risk patients and without age limit.

Cryopreservation↗
PubMed1986

Freeze-dried embedded specimens for biological microanalysis.

The main problems associated with freeze-drying of biological material for electron microscopy concern the freeze-drying temperatures and times necessary to minimize artifacts. Due to the many parameters involved these problems have to be resolved experimentally. It can be shown that good morphological preservation of chemically unfixed material is possible when freeze-drying is done exclusively in a temperature range between -80 degrees C and -50 degrees C. OsO4 vapour fixation of the freeze-dried tissue is not necessary and should be avoided because it may cause ion redistribution artifacts. Embedding at low temperature of properly freeze-dried material does not seem to disturb structure and ion distribution of the freeze-dried material. Hence, sections of such freeze-dried material and embedded biological material seem to be suitable for microanalysis. Preliminary micro-analytical results obtained from sections of freeze-dried and Lowicryl K11M embedded muscle reveal an uneven distribution of potassium in the sarcomeres similar to the visualized uneven distribution of the electron dense thallium (potassium surrogate) in frozen hydrated cryosections. A comparison of different cryomethods shows that freeze-drying and embedding is the simplest way to obtain stable thin sections of chemically unfixed biological material which, for instance, may be used for future microanalytical investigation of the interaction of proteins with physiological and non-physiological ions.

Animals↗
PubMed2005

Three-dimensional stereotactic surface projection study of freezing of gait and brain perfusion image in Parkinson's disease.

Gait disturbance is a cardinal symptom in patients with Parkinson's disease. Among the gait disturbances, freezing of gait is a unique and troublesome symptom, but its mechanism is unclear. We compared brain perfusion images using three-dimensional stereotactic surface projection analysis of N-isopropyl-p-123I iodoamphetamine single photon emission computed tomography between Parkinson's disease patients with freezing of gait and those without. Twenty-four cases (freezing of gait group) with Parkinson's disease with freezing of gait, and 31 Hoehn and Yahr stage-matched cases (no freezing of gait group) with Parkinson's disease without freezing of gait were studied. Bilateral Brodmann area 11 perfusion of the freezing of gait group decreased significantly compared to that of the no freezing of gait group. The Brodmann area 11 may play important roles in gait, and impairment in this region may have a close relationship with freezing of gait in Parkinson's disease.

Aged↗
PubMed1997

Freezing phenomenon in patients with parkinsonian syndromes.

Freezing is a common symptom in parkinsonian syndromes, but its association with different causes of parkinsonism as well as with other symptoms has never been investigated. We conducted a database survey of the occurrence of freezing in parkinsonism. Of 347 patients with a clinical diagnosis of parkinsonism other than idiopathic Parkinson's disease and with specific data regarding freezing, 158 patients had freezing (46%). Freezing was significantly associated with progression of the disease as rated on the Hoehn & Yahr scale [odds ratio (OR), 1.69; p < 0.004]. Gender was not a risk factor for the development of freezing. Patients with drug-induced parkinsonism were at a very low risk for developing freezing (p < 0.00001; OR, 0.1). Freezing was found in a high frequency in patients with vascular parkinsonism (57%), normal-pressure hydrocephalus (56%), and generally in the group of patients who had parkinsonism resulting from neurodegenerative diseases (progressive supranuclear palsy, multiple system atrophy, and corticobasal ganglionic degeneration; 45%). Freezing was significantly associated with the presence of dementia, incontinence, and tachyphemia (OR, 2.01, 1.7, and 5.09, respectively).

Dementia↗
PubMed1997

Electron Microscopy of Modified Aluminum Alkoxide Microstructures on Freeze-Drying

Direct imaging of microstructures in chelated aluminum alkoxide systems subjected to freeze-drying and freeze-thaw-drying is reported using cryo-scanning electron microscopy (SEM) and SEM techniques. Aluminum sec-butoxide was chelated with a beta-diketoester in different molar ratios. This system was gelled by hydrolysis under intense sonication. The sonogels formed were subjected to cryo-SEM. Samples for cryo-SEM were made by freezing the chelated alkoxide samples in liquid nitrogen and freeze-fracturing them inside the cryo-SEM and monitoring the apparent morphological changes on exposure to the cryo-SEM environment including intentional sample heating. Microstructures of unchelated, 1:1 chelated and 1:2 chelated (alkoxide:chelate) sonogels after freezing, thawing, and drying were also imaged by SEM techniques. Heterogeneity introduced by chelation of alkoxide is proposed as the cause for the formation of flakes in freeze-thaw-dried chelated alkoxide gels. Microstructures of 1:1 chelated freeze dried alkoxide were compared with the freeze-thaw-dried gels. The absence of flakes in the chelated samples when subjected to in situ freeze-drying indicated reversible nature of the gels.

Journal Article↗
PubMed2004

Mapping of quantitative trait loci (QTLs) affecting autumn freezing resistance and phenology in Salix.

Quantitative trait locus (QTL) analysis was performed at different time points during cold-acclimation of a tetraploid F(2 ) Salix pedigree. The pedigree ( n=92) was derived from a cross between a frost-susceptible diploid female clone 'Jorunn' ( Salix viminalis) and a frost resistant hexaploid male clone 'SW901290' ( Salix dasyclados). Freezing resistance, height growth increment and number of new leaves were assessed at days 0, 12, 20, 24, 31 and 42 of a short day-low temperature (SD-LT) hardening regime, while the initiation of shoot tip abscission and shoot tip abscission were measured daily. Height increment, dry-to-fresh weight ratio and number of new leaves were also measured in a replicated field trial. Freezing resistance was determined from electrolyte leakage of leaf tissues and from visual injuries on stem segments, after exposure to a predetermined freeze-thaw stress. Using a genetic map of the F(2) composed of 432 single-dose AFLP markers, a total of 19 genomic regions controlling freezing resistance (10) and phenological traits (9) before and during cold-acclimation (SD-LT) were identified. The magnitude of the phenotypic variation explained by each freezing resistance locus varied over acclimation time (0-45%), and there was no time point at which all the QTLs could be detected. The single QTL detected for non-acclimated freezing resistance did not reach significance at any time point during cold-acclimation, suggesting an independent genetic relationship between non-acclimated and acclimated resistance to freezing in Salix. Five of the loci associated with freezing resistance shared common intervals with loci controlling phenological traits. Of the 14 QTLs controlling autumn freezing resistance and/or phenological traits in the indoors experiment, six (43%) were associated with autumn phenology-related traits, i.e. height increment, dry-to-fresh weight ratio and number of new leaves, measured in the field. A major locus with multi-trait association in both indoor and outdoor experiments was detected.

Acclimatization↗
PubMed1992

Limits of cryofixation as seen by Fourier transform infrared spectra of metmyoglobin azide and carbonyl hemoglobin in vitrified and freeze-concentrated aqueous solution.

The limits of cryofixation were probed by investigating metmyoglobin azide and carbonyl hemoglobin in approximately 5 wt% aqueous solution by Fourier transform infrared spectroscopy. Spectra of solutions cooled slowly and recorded in steps between 295 K and 190 K are compared with those obtained by "hyperquenching" either into their glassy states at 80 K, or into freeze-concentrated solution at 170 K. For metmyoglobin azide we conclude from an analysis of its covalently and ionically bound azide that it is impossible to freeze-in its high-spin/low-spin equilibrium even by hyperquenching, and that its vitrified state must correspond to a temperature T < 226 K for the Fe(II) site of the protein. In the amide I spectral region of carbonyl hemoglobin (HbCO), a band at approximately 1654 cm-1 due to alpha-helical structures is the dominant band in spectra recorded at ambient temperature and in the vitrified state, but in the spectrum of HbCO quenched at similar rates into a freeze-concentrated state, a band at approximately 1650 cm-1, tentatively assigned to unordered structures, becomes the dominant feature. This band is absent in the spectra of freeze-concentrated samples obtained by heating a vitrified sample to 170 K. We surmise that HbCO is dehydrated by freeze-concentration to a larger extent in solution quenched rapidly at 170 K than in a vitrified solution heated to 170 K, and that this dehydration is the primary cause for HbCO's perturbation. We conclude that freeze-concentration induced by heating a vitrified solution can cause less perturbations of a protein than does quenching into a freeze-concentrated state. Therefore it can be advantageous for the practice of freeze-etching to vitrify first a solution by "hyperquenching" and thereafter freeze-etch at e.g. approximately 170 K.

Animals↗
PubMed2003

Fos-like immunoreactive neurons following electrical stimulation of the dorsal periaqueductal gray at freezing and escape thresholds.

Electrical stimulation of the dorsal regions of the periaqueductal gray (PAG) leads to defensive reactions characterized as freezing and escape responses. Until recently it was thought that this freezing behavior could be due to the recruitment of neural circuits in the ventrolateral periaqueductal gray (vlPAG), while escape would be mediated by other pathways. Nowadays, this view has been changing mainly because of evidence that freezing and escape behaviors thus elicited are not altered after lesions of the vlPAG. It has been suggested that there are at least two pathways for periaqueductal gray-mediated defensive responses, one involving the hypothalamus and the cuneiform nucleus (CnF) which mediates responses to immediate danger and another one involving the amygdala and vlPAG which mediates cue-elicited responses, either learned or innate. To examine this issue further we measured Fos protein expression in brain areas activated by electrical stimulation of the dorsolateral PAG (dlPAG) at the freezing and escape thresholds. The data obtained showed that freezing-provoking stimulation caused increases in Fos expression in the dorsomedial PAG (dmPAG), while escape-provoking stimulation led to increases at both dmPAG and dlPAG. Surprisingly, neither escape- nor freezing-provoking stimulations altered Fos expression in the central nucleus of amygdala (CeA). Escape-provoking stimulation caused increased Fos expression in the ventromedial hypothalamus (VMH), dorsal premammilary nucleus (PMd) and in the cuneiform nucleus. Significant increases in Fos labeling were found in the dmPAG and PMd following freezing-provoking stimulation. Therefore, the present data support the notion of a neural segregation for defensive behaviors in the dorsal columns of PAG, with increased Fos expression in the dmPAG following freezing, while dlPAG is affected by both freezing and escape responses. dlPAG, CnF, VMH and PMd are part of a brain aversion network activated by fear unconditioned stimuli. The present data also suggests that the defensive responses generated at the dlPAG level do not recruit the neural circuits of the vlPAG and CeA usually activated by conditioned fear stimuli.

Animals↗
PubMed1996

Effects of typical and atypical antipsychotic drugs on freezing behavior induced by conditioned fear.

A typical antipsychotic drugs (atypical APDs), such as clozapine, ORG5222, and olanzapine, have been suggested to possess anxiolytic activity in the conflict test and elevated plus-maze test, while several studies have suggested that typical APDs are not anxiolytic in several models of anxiety. We investigated the effects of typical and atypical APDs on the acquisition and expression of conditioned fear-induced freezing. Drugs were administered subcutaneously to male Sprague-Dawley rats 30 min before foot shock stress (the VI60s schedule, 2.5 mA for 30 min). Twenty-four hours after foot shock, freezing behavior of rats was observed in the shock chamber without shocks. The atypical APD clozapine (0.3-10 mg/kg) dose-dependently inhibited the acquisition of conditioned freezing. Candidates for atypical APDs, ORG5222 (0.1-1 mg/kg), olanzapine (1-10 mg/kg), and raclopride (3-30 mg/kg), also reduced the acquisition of conditioned freezing in a dose-dependent manner. Typical APDs, haloperidol (3 mg/kg), spiperone (0.1-1 mg/kg) and nemonapride (1 mg/kg) had significant inhibitory effects on the acquisition of conditioned freezing, but their effects were reduced at higher doses. Chlorpromazine, a typical APD, showed about 50% inhibition of the acquisition of conditioned freezing at the dose of 10 mg/kg, but did not reveal significant inhibition at any of the doses (3-30 mg/kg). The ED50S (mg/kg) for inhibiting the acquisition of conditioned freezing significantly correlated with the Ki values for D4 dopaminergic receptors, but not with the Ki values for other monoamine and acetylcholine receptors. On the other hand, clozapine or haloperidol did not change the expression of conditioned freezing. These results suggest that protective effects of clozapine and other antipsychotic drugs on the acquisition of conditioned freezing may be mediated by blockade of D4 receptors.

Animals↗
PubMedDate not supplied

Cryopreservation of collagen-based tissue equivalents. II. Improved freezing in the presence of cryoprotective agents.

In Part I of this study we determined an optimal cooling rate for cryopreservation of collagen-based tissue equivalents (TEs) that preserves both the postthaw cell viability and mechanical properties, but results in tissue contraction and an overall loss of opacity. The empirically determined optimal cooling rate (5 degrees C/min) was obtained in a freezing medium consisting solely of phosphate-buffered saline (PBS) at physiological concentration (1x). In the present study we report the effect of freezing on TEs in the presence of PBS and two cryoprotective agents (CPAs) (glycerol and dimethyl sulfoxide [Me(2)SO]), at two different concentrations (0.5 and 1.0 M), to two different end temperatures (-80 and -160 degrees C), at a cooling rate of 5 degrees C/min. The controlled rate freezing experiments, postthaw cell viability, and mechanical property measurements were performed as described in Part I of this study. In addition to studying the effect of CPAs on the postthaw properties of TEs, we also investigated (1). the effect of freezing TEs attached to the substrate (as opposed to detached and floating in medium) to determine differences when freezing TEs subject to static mechanical stress via a mechanical constraint to contraction; (2). the effect of freezing glutaraldehyde-fixed TEs to determine differences in freezing-mediated damage to the microstructure; and (3). the effect of freezing more mature TEs that were incubated for 4 weeks in growth factor-supplemented medium as opposed to 2 weeks in basal medium. All TEs frozen at 5 degrees C/min to -80 degrees C in the presence of 0.5 M glycerol or Me(2)SO in PBS were found to be optimally cryopreserved in terms of maintaining opacity and structure as well as cell viability and mechanical properties as compared with unfrozen TEs. The postthaw mechanical properties were adversely affected by freezing to the lower end temperature of -160 degrees C in the presence of CPAs, with the samples frozen in the 1.0 M concentration of CPAs exhibiting a total loss of structural integrity on thawing. Furthermore, TEs frozen attached to the substrate showed decreased opacity and significant contraction as compared with TEs frozen detached from the substrate, as did cross-linked samples frozen without CPA.

Collagen↗
PubMed1992

Seasonal differences in freezing stress resistance of needles of Pinus nigra and Pinus resinosa: evaluation of the electrolyte leakage method.

Seasonal changes in freezing stress resistance of needles of red pine (Pinus resinosa Ait.) and Austrian pine (Pinus nigra Arnold) trees were measured by an electrolyte leakage method and by visual observation. During most of the year, freezing stress resistance determined by the two methods gave similar results. The electrolyte leakage method provided a good estimate of seasonal changes in freezing stress resistance except for red pine needles in their most winter-hardy state. To obtain a reliable estimate of freezing stress resistance in winter-hardy red pine needles it was necessary to combine the electrolyte leakage method with visual observations. When red pine needles survived exposure to -80 degrees C or lower, electrolyte leakage was never more than 30% even when the needles were exposed to a slow freeze-thaw stress of -196 degrees C. However, rapid freezing of red pine needles to -196 degrees C resulted in electrolyte leakage of over 80%. Red pine needles attained a much higher freezing stress resistance during the winter than Austrian pine. Red pine needles also acclimated and deacclimated faster than Austrian pine needles. An index of injury was developed based on the electrolyte leakage method ((R(2) + R(1))/2, where R(1) is the minimum % electrolyte leakage from noninjured tissue and R(2) is the maximum % electrolyte leakage at the highest injury) that reliably predicted freezing stress resistance of pine needles for most of the year. Important aspects for developing a successful index of injury for pine needles are: use of cut needles, vacuum infiltration and shaking during incubation in water.We conclude that: (1) during cold acclimation the cell wall properties of the pine needles changed and these changes, which appeared to differ in the two species, might explain the very low leakage of electrolytes from winter-hardy needles of red pine; (2) pine needles survive winter by developing the ability to tolerate extracellular ice formation, because after rapid freezing the needles were severely injured; and (3) red pine is adapted to a shorter growing season and colder winters than Austrian pine.

Journal Article↗
PubMed1995

Reversible Photoinhibition in Antarctic Moss during Freezing and Thawing.

Tolerance of antarctic moss to freezing and thawing stress was investigated using chlorophyll a fluorescence. Freezing in darkness caused reductions in Fv/Fm (ratio of variable to maximum fluorescence) and Fo (initial fluorescence) that were reversible upon thawing. Reductions in Fv/Fm and Fo during freezing in darkness indicate a reduction in the potential efficiency of photosystem II that may be due to conformational changes in pigment-protein complexes due to desiccation associated with freezing. The absorption of light during freezing further reduced Fv/Fm and Fo but was also reversible. Using dithiothreitol (DTT), which inhibits the formation of the carotenoid zeaxanthin, we found reduced flurorescence quenching during freezing and reduced concentrations of zeaxanthin and antheraxanthin after freezing in DTT-treated moss. Reduced concentrations of zeaxanthin and antheraxanthin in DTT-treated moss were partially associated with reductions in nonphotochemical fluorescence quenching. The reversible photoinhibition observed in antarctic moss during freezing indicates the existence of processes that protect from photoinhibitory damage in environments where freezing temperatures occur in conjunction with high solar radiation levels. These processes may limit the need for repair cycles that require temperatures favorable for enzyme activity.

Journal Article↗
PubMed1967

Freezing injury and uncoupling of phosphorylation from electron transport in chloroplasts.

Freezing of chloroplast membranes uncouples photophosphorylation from electron transport and inactivates the light-dependent and thiol-requiring ATPase, conformational changes and the light-dependent proton uptake. All of these energy requiring activities can be protected against inactivation by addition of sucrose prior to freezing. The direct relation to photophosphorylation is demonstrated by the quantitatively similar response of photophosphorylation and the other activities to sucrose protection. Salts interfere with the protection afforded by sucrose.In contrast to the light-dependent ATPase, the ATPase activities which are unmasked by digestion with trypsin show no significant response to freezing. Similarly, the chloroplast coupling factor, which is released from the membranes by ethylenediamine tetraacetic acid treatment, survives freezing. The membranes, which are depleted of the factor, are damaged by freezing.The results suggest that uncoupling of phosphorylation from electron transport is caused by interference of freezing with a structure involved in the formation of a non-phosphorylated high energy state of chloroplasts. They are best explained on the basis of Mitchell's theory of phosphorylation. Since freezing alters the permeability properties of chloroplast membranes-frozen membrane vesicles no longer function as osmometers-it may be assumed that freezing uncouples phosphorylation from electron transport by preventing the formation of a pH gradient across the vesicle membranes owing to proton leakage through the membranes. From the results, the basic injury caused by freezing appears to consist in the alteration of permeability properties of biological membranes due to the dehydration which accompanies freezing.

Journal Article↗
PubMed1969

Multiple freezing points as a test for viability of plant stems in the determination of frost hardiness.

A technique is presented for a simple, rapid, and reliable means of determining the viability of plant tissue subjected to freezing temperatures. Freezing curves of excised stems of Cornus stolonifera Michx., and several other genera were studied. Tissue temperature was recorded during freezing of plant stem sections. The heat of crystallization deflected the resultant freezing curves at points where tissue froze. Living stem sections of all genera studied revealed 2 freezing points, while dead tissue exhibited only 1. The influence of variables such as moisture content, sample size, thermocouple placement, and cooling rate on freezing curves was analyzed. Stem samples wrapped in moisture-proof film with a thermocouple inserted into the pith were frozen to a predetermined test temperature, thawed, and subjected to a second freezing cycle. The presence or absence of 2 freezing points in the second freezing cycle was used as a criterion for establishing viability. The results were immediately available and identical to results from regrowth tests which took about 20 days.

Journal Article↗
PubMed2000

Quantitative measurement and prediction of biophysical response during freezing in tissues.

Cryopreservation and cryosurgery are important biomedical applications used to selectively preserve or destroy cellular systems through freezing. Studies using cryomicroscopy techniques, which allow the visualization of the freezing process in single cells, have shown that a drop in viability correlates with the extent of two biophysical events during the freezing process: (a) intracellular ice formation and (b) cellular dehydration. These same biophysical events operate in tissue systems; however, the inability to visualize and quantify the dynamics of the freezing process in tissues has hampered direct correlation of these events with freezing-induced changes in viability. This review highlights two new techniques that use freeze substitution and differential scanning calorimetry to provide dynamic freezing data in tissue. Characteristic dimensions and parameters extracted from these new data are then used in a predictive model of biophysical freezing response in several tissues, including liver and tumor. This approach promises to help guide improved design of both cryopreservation and cryosurgical applications of tissue freezing.

Animals↗