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Controlled manufacturing and long-term stability are key challenges in the development and translation of nanomedicines. This is exemplified by the mRNA-nanoparticle vaccines against COVID-19, which require (ultra-)cold temperatures for storage and shipment. Various cryogenic protocols have been explored to prolong nanomedicine shelf-life. However, freezing typically induces high mechanical stress on nanoparticles, resulting in aggregation or destabilization, thereby limiting their performance and application. Hence, evaluating the impact of freezing and storing on nanoparticle properties already early-on during preclinical development is crucial. In the present study, we used prototypic π electron-stabilized polymeric micelles based on mPEG- b -p(HPMAm-Bz) block copolymers to macro- and microscopically study the effect of different cryoprotective excipients on nanoformulation properties like size and size distribution, as well as on freezing-induced aggregation phenomena via in-situ freezing microscopy. We show that sucrose, unlike trehalose, efficiently cryoprotected paclitaxel-loaded micelles, and we exemplify the impact of formulation composition for efficient cryoprotection. We finally establish microfluidic mixing to formulate paclitaxel-loaded micelles with sucrose as a cryoprotective excipient in a single production step and demonstrate their stability for 6 months at −20 °C. The pharmaceutical properties and preclinical performance (in terms of tolerability and tumor growth inhibition in a patient-derived triple-negative breast cancer xenograft mouse model) of paclitaxel-loaded micelles were successfully cryopreserved. Together, our efforts promote future pharmaceutical development and translation of π electron-stabilized polymeric micelles, and they illustrate the importance of considering manufacturing and storage stability issues early-on during nanomedicine development. [Display omitted] [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Oocyte cryopreservation is not yet considered a reliable technique since it can reduce the quality and survival of oocytes in several species. This study determined the effect of different concentrations of antifreeze protein I (AFP I) on the vitrification solution of immature cat oocytes. For this, oocytes were randomly distributed in three groups and vitrified with 0 μg/mL (G0, 0 μM); 0.5 μg/mL (G0.5, 0.15 μM), or 1 μg/mL (G1, 0.3 μM) of AFP I. After thawing, oocytes were evaluated for morphological quality, and compared to a fresh group (FG) regarding actin integrity, mitochondrial activity and mass, reactive oxygen species (ROS) and glutathione (GSH) levels, nuclear maturation, expression of GDF9 , BMP15 , ZAR-1 , PRDX1 , SIRT1, and SIRT3 genes (normalized by ACTB and YWHAZ genes), and ultrastructure. G0.5 and G1 presented a higher proportion of COCs graded as I and while G0 had a significantly lower quality. G1 had a higher percentage of intact actin in COCs than G0 and G0.5 (P < 0.05). There was no difference (P > 0.05) in the mitochondrial activity between FG and G1 and they were both higher (P < 0.05) than G0 and G0.5. G1 had a significantly lower (P < 0.05) mitochondrial mass than FG and G0, and there was no difference among FG, G0, and G0.5. G1 had higher ROS than all groups (P < 0.05), and there was no difference in GSH levels among the vitrified groups (P > 0.05). For nuclear maturation, there was no difference between G1 and G0.5 (P > 0.05), but these were both higher (P < 0.05) than G0 and lower (P < 0.05) compared to FG. Regarding gene expression, in G0 and G0.5, most genes were downregulated compared to FG, except for SIRT1 and SIRT3 in G0 and SIRT3 in G0.5. In addition, G1 kept the expression more similar to FG. Regardless of concentration, AFP I supplementation in vitrification solution of immature cat oocytes improved maturation rates, morphological quality, and actin integrity and did not impact GSH levels. In the highest concentration tested (1 μg/mL), AFP maintained the mitochondrial activity, reduced mitochondrial mass, increased ROS levels, and had the gene expression more similar to FG. Altogether these data show that AFP supplementation during vitrification seems to mitigate some of the negative impact of cryopreservation improving the integrity and cryosurvival of cat oocytes. [Display omitted] • The use of AFP I in the vitrification solution of immature cat oocytes improved their maturation rate. • AFP I also improved the morphological quality and actin integrity of immature vitrified cat oocytes. • In the highest concentration tested (1 μg/mL), AFP I kept the gene expression more similar to fresh oocytes. • 1 μg/mL of AFP I seems to be the suitable concentration for improving oocyte vitrification in this species. [ABSTRACT FROM AUTHOR]

The preservation of microorganisms is pivotal in microbiological practice. Currently, cryopreservation is assumed to be an effective and inexpensive approach for the storage of microorganisms, including bacteria. The key point of cryopreservation is optimal cryoprotectant selection. In the present study, different cryoprotectant compositions were tested for long-term storage of 15 Enterobacterales bacterial strains at − 20 °C. The survival rates of the bacterial strains were evaluated in four different cryoprotectant solutions containing 70% glycerin only (cryoprotectants 1 and 4), 10% dimethyl sulfoxide (DMSO) with 70% glycerin (cryoprotectant 2), and 10% DMSO (cryoprotectant 3). In addition, cryoprotectants 1 and 2 contained peptone and yeast extract as nutritional supplements. The general survival rates of the bacterial strains were evaluated after 12 months of storage. After 12 months, the survival rates of the different cryoprotectants were as follows: cryoprotectant 1—88.87%; cryoprotectant 2—84.85%; cryoprotectant 3—83.50%; and cryoprotectant 4—44.81%. Thus, the composition of cryoprotectant 1 (70% glycerin with nutrient supplements) was optimal for preserving 15 tested strains of the order Enterobacterales. Despite these findings, the biochemical properties of the tested strains changed after cryopreservation for 12 months in the presence of 1 or 3 cryoprotectants. Alterations in the biochemical profile could be related to changes in environmental conditions and cold adaptation. We assume that the composition of cryoprotectant 1 can be optimal for storing the order Enterobacterales at − 20 °C. However, further investigations are needed to elucidate the problem of cryopreservation and to support our assumption. [ABSTRACT FROM AUTHOR]

Abstract The preservation of microorganisms is pivotal in microbiological practice. Currently, cryopreservation is assumed to be an effective and inexpensive approach for the storage of microorganisms, including bacteria. The key point of cryopreservation is optimal cryoprotectant selection. In the present study, different cryoprotectant compositions were tested for long-term storage of 15 Enterobacterales bacterial strains at − 20 °C. The survival rates of the bacterial strains were evaluated in four different cryoprotectant solutions containing 70% glycerin only (cryoprotectants 1 and 4), 10% dimethyl sulfoxide (DMSO) with 70% glycerin (cryoprotectant 2), and 10% DMSO (cryoprotectant 3). In addition, cryoprotectants 1 and 2 contained peptone and yeast extract as nutritional supplements. The general survival rates of the bacterial strains were evaluated after 12 months of storage. After 12 months, the survival rates of the different cryoprotectants were as follows: cryoprotectant 1—88.87%; cryoprotectant 2—84.85%; cryoprotectant 3—83.50%; and cryoprotectant 4—44.81%. Thus, the composition of cryoprotectant 1 (70% glycerin with nutrient supplements) was optimal for preserving 15 tested strains of the order Enterobacterales. Despite these findings, the biochemical properties of the tested strains changed after cryopreservation for 12 months in the presence of 1 or 3 cryoprotectants. Alterations in the biochemical profile could be related to changes in environmental conditions and cold adaptation. We assume that the composition of cryoprotectant 1 can be optimal for storing the order Enterobacterales at − 20 °C. However, further investigations are needed to elucidate the problem of cryopreservation and to support our assumption.

This pioneering study aimed to evaluate the cryopreservation of semen from P. falkneri (n = 4) and P. motoro (n = 4), maintained ex situ at the Sao Paulo Aquarium, Brazil. For this purpose, the animals were physically restrained, biometric data of the disc and clasper were obtained, and semen was collected through manual massage. Total motility and progressive motility parameters were evaluated using Computer-Assisted Sperm Analysis (CASA) with IVOS II equipment and Animal Breeders II software. The semen extenders INRA 96 and OptiXcell were used to assess their efficacy in sperm cryopreservation. INRA required the addition of 5% dimethyl sulfoxide (DMSO) as a cryoprotectant. The results indicated that there was no difference in semen motility values before and after freezing with INRA + DMSO (p = 0.6226). On the other hand, samples cryopreserved with OptiXcell showed a difference in semen motility post-thaw (p = 0.0156). These findings contribute to a broader study on optimizing cryopreservation protocols to ensure long-term viability and fertility of semen, enhancing genetic diversity and supporting wild population restoration. A multidisciplinary approach integrating reproductive biology, ecology, physiology, and assisted reproduction technologies, aligned with the One Conservation concept, is essential for advancing conservation and management strategies for these threatened species.

This pioneering study aimed to evaluate the cryopreservation of semen from P. falkneri (n = 4) and P. motoro (n = 4), maintained ex situ at the Sao Paulo Aquarium, Brazil. For this purpose, the animals were physically restrained, biometric data of the disc and clasper were obtained, and semen was collected through manual massage. Total motility and progressive motility parameters were evaluated using Computer-Assisted Sperm Analysis (CASA) with IVOS II equipment and Animal Breeders II software. The semen extenders INRA 96 and OptiXcell were used to assess their efficacy in sperm cryopreservation. INRA required the addition of 5% dimethyl sulfoxide (DMSO) as a cryoprotectant. The results indicated that there was no difference in semen motility values before and after freezing with INRA + DMSO (p = 0.6226). On the other hand, samples cryopreserved with OptiXcell showed a difference in semen motility post-thaw (p = 0.0156). These findings contribute to a broader study on optimizing cryopreservation protocols to ensure long-term viability and fertility of semen, enhancing genetic diversity and supporting wild population restoration. A multidisciplinary approach integrating reproductive biology, ecology, physiology, and assisted reproduction technologies, aligned with the One Conservation concept, is essential for advancing conservation and management strategies for these threatened species. [ABSTRACT FROM AUTHOR]

Summary: Testicular biopsies (9 mm3) from domestic cats (n = 10) submitted to orchiectomy were submitted to equilibrium vitrification in the presence of ethylene glycol (EG) alone or combined with dimethylsulfoxide (DMSO) as intracellular cryoprotectants, and sucrose or trehalose as extracellular cryoprotectants. The samples were vitrified with 40% EG or 20% EG + 20% DMSO, plus 0.1 M or 0.5 M of sucrose or trehalose. The study was divided into Step 1 and Step 2. In Step 1, intratubular cells (spermatogonia, spermatids, spermatocytes, and Sertoli cells) were quantified and classified as intact or degenerated (pyknotic and/or vacuolated cells). Cryodamage of seminiferous cords was determined by spermatogonia and Sertoli cell scoring of nuclei alterations, tubular basement membrane detachment, epithelium shrinkage, and tubular measures (total area, epithelium area, larger and smaller diameter, and height of the epithelium). In Step 2, Hoechst 33342 stain and propidium iodide (PI) fluorescent stain were used to assess the cell viability of the four best experimental groups in Step 1. The effect of treatments on all analyses was accessed using analysis of variance (ANOVA), and Fisher's post hoc test at P < 0.05 significance was considered. In Step 1, the mean percentage of spermatogonia and Sertoli cells morphological integrity did not show a difference when using both sugars at different concentrations, but their morphology was more affected when DMSO was used. EG use associated with 0.1 M of sucrose or trehalose positively affected spermatocyte and spermatid morphology, respectively. The larger diameter and epithelium height of seminiferous tubules were increased using DMSO plus 0.5 M sucrose and DMSO plus 0.1 M trehalose. The changes in spermatogonial/Sertoli nucleoli visualization were best scored in the EG groups, while the nuclei condensation was lower with sucrose. The basement membrane was satisfactorily preserved with 0.1 M sucrose. In Step 2, the percentage of cell viability was higher when EG plus 0.1 M sucrose was used. Therefore, DMSO's negative effect on the vitrification of testicular biopsies of adult domestic cats was evident. The EG plus 0.1 M of sucrose or trehalose associations are the most suitable CPAs to preserve the testicular histology structure of adult domestic cats in vitrification. [ABSTRACT FROM AUTHOR]

Fruit crops are an irreplaceable source of essential nutrients, macro- and microelements, vitamins, organic acids, and antioxidants. Today, the overwhelming part of fruit supply in the market is provided by foreign producers. Import substitution and meeting the demand of the Russian Federation population for fruit consumption by the domestic agro-industrial complex is impossible without expanding the geography of cultivation areas, including those in zones of risky agriculture, which requires breeding of frost-resistant (cold-resistant) cultivars (fruit crops). Application of modern biotechnological and molecular genetic methods in breeding work will increase the profitability of fruit growing by reducing the time required for obtaining plants with the desired traits and by complex evaluation of the prospects of genotypes of parental forms. The present review considers modern data on cold tolerance genes of various fruit and berry crops, summarizes the known mechanisms of their action, activation, and regulation.The review considers modern data on genes of fruit and berry crops resistance to low temperatures, including characterization of genes encoding key receptors, signaling, effector proteins, and transcription factors in apple, pear, peach, pineapple, and strawberry. The known mechanisms of their operation, activation, regulation are given, and signaling cascades are described.

During the freezing, storage, and thawing process, physicochemical changes occur in raw meat, which will inevitably lead to quality deterioration mainly including decreased water-holding capacity and the deterioration of color and texture. The quality deterioration can be effectively reduced by novel freezing and its assistive technologies. This article summarizes the quality deterioration mechanism of raw meat during freezing and frozen storage, and reviews the approaches for improving frozen meat quality, such as novel freezing techniques, cryoprotectants, novel packaging techniques and novel thawing techniques, which will provide a theoretical basis and technical guidance for improving the quality of frozen meat.

Cryopreservation is a valuable technique used to assist in the genetic improvement of cultured stocks and provide a continuous supply of good‐quality semen for artificial insemination. Conserving semen by cryopreservation serves several purposes (e.g. artificial reproductive technologies and species conservation) and is also used in the clinical treatment of human infertility. However, the lifespan of cryopreserved semen is influenced by a range of factors, including storage temperature, cooling rate, chemical composition of the extender, the concentration of cryoprotectant, reactive oxygen species, seminal plasma composition and hygienic control. The choice of cryoprotectant is a vital factor underlying the success of animal semen cryopreservation. In this regard, extensive research has been carried out on various cryoprotectants, such as egg yolk, dimethyl sulfoxide, methanol, ethylene glycol and dimethylacetamide. Recent studies have also described the use of a range of new cryoprotectants for cryopreservation, including compounds of plant origin (soy), amino acids, antifreeze proteins, carbohydrates and cyclodextrins. Moreover, semen cryopreservation and storage require the use of liquid nitrogen or ultralow refrigeration methods for both long‐ and short‐term storage. This review summarizes the general methods used for freezing semen and discusses the use of traditional and newly emerging cryoprotectants (permeable and non‐permeable) for the cryopreservation of semen in selected fish and mammalian species. [ABSTRACT FROM AUTHOR]

Copyright of Shipin Kexue/ Food Science is the property of Food Science Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Although several aspects like diapause determining factors, population structure, reproductive physiology, and genetics of diapause have been investigated, there is no clarity on carbohydrate energetics during larval diapause in Chilo partellus (Swinhoe). Present studies revealed significant variation between the nondiapausing and diapausing C. partellus for total carbohydrates, glycogen, sorbitol, and trehalose contents in different body parts, life stages, and for body parts × life stages interaction. Total carbohydrate content started declining, while sorbitol and trehalose increased in all the body parts as the C. partellus larvae progressed from prediapausing to diapausing state. However, glycogen content spiked in all the body parts at prediapausing stage, which then declined during diapause. Among the body parts, total carbohydrate content was significantly greater in the hemolymph as compared to other body parts of both larvae and pupae of C. partellus. Glycogen content was significantly greater in the larval fat bodies and pupal hemolymph as compared to their other body parts. In diapausing larvae, sorbitol and trehalose were greater in the integument than in other body parts. Furthermore, there was spike in trehalose and decrease in sorbitol in all the body parts of pupae from diapausing than those from nondiapausing larvae. These findings suggest that the diapause alterate and/or fluctuate major carbohydrates in different body parts of both larvae and pupae of C. partellus. This information will be helpful in better understanding the diapause energetics and overwintering metabolic cryoprotection in insects. [ABSTRACT FROM AUTHOR]

Cryopreservation is one of the reliable techniques for long‐term storage of sperm. The success of this technique depends on the choice of cryoprotectant; therefore, a plethora of literature has reported the effects of different cryoprotective agents so far. Kappa‐carrageenan (κ‐carrageenan) is a hydrocolloid polysaccharide extracted from red marine seaweed. Its unique property makes it a promising option as a non‐colligative cryoprotectant. The current study aims to evaluate the cryoprotective effect of k‐carrageenan along with glycerol on ram sperm quality both after equilibration and freezing. Nine Kajli rams were utilized in this experiment for semen collection through an artificial vagina maintained at 42°C. Qualified samples were diluted in tris egg yolk glycerol (TEYG) extender containing different concentrations of k‐carrageenan as 0 mg/mL (control), 0.2, 0.5, 0.8 and 1 mg/mL. Post‐thaw assessment was done at 37°C after 24 h of storage, which showed a significant improvement (p <.05) in sperm viability, motility, membrane and acrosome integrity in an extender containing k‐carrageenan at a concentration of 0.5 mg/mL compared to control. It is concluded from the current study that the combination of glycerol and 0.5 mg/mL concentration of k‐carrageenan improved the sperm post‐thaw quality. [ABSTRACT FROM AUTHOR]

The concept of superchilling, an alternative to traditional chilling and freezing methods, is to reduce the temperature 1-2 °C below the initial freezing point of the food enabling safe, and high-quality products. This work was aimed to explore the effects of superchilling (-2±0.5°C), chilling (4±1°C), and freezing (-18±1°C) on the quality and storage stability of dressed chicken and also to evaluate the role of cryoprotectants during superchilling for enhanced meat quality. In superchilled samples, improved water holding capacity (WHC) and lower shear force values were observed relative to frozen chicken. SDSPAGE analysis revealed 29 kDa protein bands in superchilled samples on 7th and 14th day, indicating troponin degradation and tenderization. Scanning electron microscopy (SEM) studies indicated minimal damage to muscle fibre ultrastructure in superchilled chicken relative to frozen samples. Integrity of fibre structure was well maintained in superchilled samples treated with sorbitol as a cryoprotectant relative to sucrose and polyphosphate. Results demonstrated the potential use of superchilling process along with cryoprotectants for improved quality and enhanced shelf-life of chicken. [ABSTRACT FROM AUTHOR]

(1) Background: DA-Gelucire® 50/13-based solid lipid nanoparticles (SLNs) administering the neurotransmitter dopamine (DA) and the antioxidant grape-seed-derived proanthocyanidins (grape seed extract, GSE) have been prepared by us in view of a possible application for Parkinson's disease (PD) treatment. To develop powders constituted by such SLNs for nasal administration, herein, two different agents, namely sucrose and methyl-β-cyclodextrin (Me-β-CD), were evaluated as cryoprotectants. (2) Methods: SLNs were prepared following the melt homogenization method, and their physicochemical features were investigated by Raman spectroscopy, Scanning Electron Microscopy (SEM), atomic force microscopy (AFM) and X-ray Photoelectron Spectroscopy (XPS). (3) Results: SLN size and zeta potential values changed according to the type of cryoprotectant and the morphological features investigated by SEM showed that the SLN samples after lyophilization appear as folded sheets with rough surfaces. On the other hand, the AFM visualization of the SLNs showed that their morphology consists of round-shaped particles before and after freeze-drying. XPS showed that when sucrose or Me-β-CD were not detected on the surface (because they were not allocated on the surface or completely absent in the formulation), then a DA surfacing was observed. In vitro release studies in Simulated Nasal Fluid evidenced that DA release, but not the GSE one, occurred from all the cryoprotected formulations. Finally, sucrose increased the physical stability of SLNs better than Me-β-CD, whereas RPMI 2650 cell viability was unaffected by SLN-sucrose and slightly reduced by SLN-Me-β-CD. (4) Conclusions: Sucrose can be considered a promising excipient, eliciting cryoprotection of the investigated SLNs, leading to a powder nasal pharmaceutical dosage form suitable to be handled by PD patients. [ABSTRACT FROM AUTHOR]

In recent decades, consumer preference and attention to foodstuff presented as healthy and with desirable nutritional information, has increased significantly. In this field, the meat industry has a challenging task since meat and meat products have been related to various chronic diseases. Functional ingredients have emerged in response to the increasing demand for healthier and more nutritious foods. On this matter, oligosaccharides such as fructooligosaccharides (FOS), xylooligosaccharides (XOS), galactooligosaccharides (GOS), and chitooligosaccharides (COS) have been presented as suitable ingredients for the meat industry with the aim of obtaining healthier meat derivatives (e.g. with low fat or sugar content, reduced amount of additives, and desirable functional properties, etc.). However, studies considering application of such oligomers in the meat sector are scarce. In addition, a large number of issues remain to be solved related both to obtaining and characterizing the oligosaccharides available in the industry and to the effect that these ingredients have on the features of meat products (mainly physicochemical and sensory). The study of new oligosaccharides, the methodologies for obtaining them, and their application to new meat products should be promoted, as well as improving knowledge about their effects on the properties of functional meat foods. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Refrigeration is the property of Journal of Refrigeration Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Abstract The objective of this work was to evaluate cryoprotectants on the physiological quality of Adenium obesum seeds preserved in liquid nitrogen. The following treatments were used: T1, control, without cryoprotectant and without immersion in liquid nitrogen; T2, with immersion in liquid nitrogen and without cryoprotectant; T3, with 0. 4 mol L-1 sucrose; T4, with 0.8 mol L-1 sucrose; T5, with 1.0 mol L-1 glycerol; T6, with 2.0 mol L-1 glycerol with PVS1; T7, with PVS1; T8, with PVS2; T9, with PVS2 + phloroglucinol (modified PVS2); and T10, with PVS3. Adenium obesum seeds show tolerance to immersion and storage in liquid nitrogen and do not need cryoprotectants to maintain their physiological quality when stored in a water content of 7.5%.

Anti-nucleolin (NCL) aptamer AS1411 is the first anticancer aptamer tested in clinical trials. Gold nanoparticles (AuNP) have been widely exploited for various biomedical applications due to their unique functional properties. In this study, we evaluated the colloidal stability and targeting capacity of AS1411-funtionalized AuNP (AuNP/NCL-Apt) against MCF-7 breast cancer cell line before and after lyophilization. Trehalose, mannitol, and sucrose at various concentrations were evaluated to determine their cryoprotection effects. Our results indicate that sucrose at 10 % (w/v) exhibits the best cryoprotection effect and minimal AuNP/NCL-Apt aggregation as confirmed by UV–Vis spectroscopy and dynamic light scattering (DLS) measurements. Moreover, the lyophilized AuNP/NCL-Apt at optimized formulation maintained its targeting and cytotoxic functionality against MCF-7 cells as proven by the cellular uptake assays utilizing flow cytometry and confocal laser scanning microscopy (CLSM). Quantitative PCR (qPCR) analysis of nucleolin-target gene expression also confirmed the effectiveness of AuNP/NCL-Apt. This study highlights the importance of selecting the proper type and concentration of cryoprotectant in the typical nanoparticle lyophilization process and contributes to our understanding of the physical and biological properties of functionalized nanoparticles upon lyophilization.

Fresh fermented milk products have a limited shelf life that can be extended by vacuum freeze-drying. Cryoprotectants are used to increase the survival of lactic acid microorganisms during freeze-drying. The most effective cryoprotectants are those of natural origin. Literature offers little information on the cryoprotective effects of fruit and vegetable purees. Therefore, we aimed to evaluate the effectiveness of fruit and vegetable purees in increasing the survival of lactic acid microorganisms during the freeze-drying and storage of fermented milk products. We studied bioyogurt samples containing pumpkin, fig, and banana purees. Rational modes of freezing and freeze-drying were established on the basis of thermal analysis. The cryoscopic temperature was determined by differential scanning calorimetry. The proportion of frozen moisture was calculated using the Nagaoka formula. Standard methods were employed to evaluate the sensory characteristics of bioyogurts and determine their protein, fat, and non-fat milk solids contents, as well as titratable acidity and microbiological indicators. The addition of pumpkin puree increased the cryoscopic temperature and reduced the freeze-drying stage and the total drying time by 13 h, depending on the amount of puree. However, the addition of sweet fig and banana purees decreased the cryoscopic temperature and increased the freeze-drying stage and the total drying time by 0.5–1.5 and 1.5–3 h, respectively. Based on the sensory evaluation of the freeze-dried bioyogurts, we selected the formulations with 15% of pumpkin and fig purees and 10% of banana puree. We found that the freeze-dried bioyogurts with puree had higher counts of lactic acid bacteria compared to the control. In the freeze-dried samples, the counts were higher at a storage temperature of 4 ± 2°C than at 20 ± 2°C. Pumpkin puree provided the best survival of lactic acid microorganisms during freeze-drying and storage.

Probiotic formula for the vaginal tract must contain high numbers of viable beneficial bacteria that maintain their characteristics during the production and further storage, in order to exert the claimed probiotic effect. Four probiotic strains—Lactobacillus gasseri CRL1320, Limosilactobacillus reuteri CRL1324, Ligilactobacillus salivarius CRL1328 and Lacticaseibacillus rhamnosus CRL1332—originally isolated from vagina of healthy women exhibiting beneficial properties were evaluated. Therefore, the optimization of growth parameters and dry biomass production with high viability while preserving probiotic features of strains is a great challenge. Different growth conditions in MRS medium were set up in a laboratory bioreactor; two initial pH and agitation were recorded speeds during static or controlled fermentations. Production conditions of 37 °C with controlled pH 6.5 and 5.5-MRS with 150 and 75 rpm stirring speeds were used for CRL1329 and CRL1332 strains, respectively, while static and free pH MRS for CRL1324 and CRL1328 probiotics allowed us to obtain maximal cell viability counts. However, during 12 h of fermentation, biomass yields of 19.3, 16.2 and 15.2 g/L were achieved when CRL1329, CRL1328 and CRL1324 probiotic strains were grown in static and free pH MRS. The highest biomass yield for CRL1332 strain was produced under controlled MRS-initial pH 6.5 and 75 rpm fermentation conditions. To preserve probiotic high biomass viability, freeze-drying was carried out in the presence of different cryoprotective agents. Thus, the highest viable numbers (10.9–11.8 log CFU/g) with survival rates between 91.3 and 95.6% were attained in the presence of 10% trehalose (L. reuteri and L. salivarius), lactose (L. rhammosus) and lactose + trehalose + sucrose mix (L. gasseri). When stability during post-freeze-drying storage was evaluated, probiotic strains showed a remarkably higher viability recovery when stored at 4 °C than at 25 °C for 12 and 3 months, respectively. In addition, surface characteristics of vaginal probiotics were affected to different extents during storage depending on the strain, protective agent and storage time/temperature. Critical factors for growth conditions, drying process and storage stability of probiotic lactobacilli strains were optimized in view to preserve cell high viability and surface features for the design of vaginal probiotic formula.

Peripheral blood hematopoietic stem cell (HSC) transplantation is a well-established procedure for the treatment of hematological, cancer and autoimmune diseases. In cancer patients, HSC transplantation allows the use of high-dose cytotoxic drugs in combination with radiation therapy during treatment, which provides a pronounced antitumor effect. The hematological toxicity of such treatment is eliminated by the sequential introduction of stem cells, which contribute to hematopoiesis restoration. Before transplantation, peripheral blood HSCs are subjected to collection and cryopreservation for further storage. An important requirement for cryopreservation is viable HSCs responsible for hematopoietic restoration. The aim of the review was to analyze the literature devoted to the influence of various methods of cryopreservation of human peripheral blood HSCs on the preservation of cell viability after thawing, as well as the development of adverse events in patients. Issues related to the use of various cryoprotectants, as well as methods for storing HSC grafts, are considered. The presented data indicate the need for further study of the effect of cryoprotectants on the human body and the cellular composition of the graft and improvement of protocols for HSC cryopreservation.

Abstract The transfusion of red blood cells (RBCs) is crucial in current medicinal research. The shelf‐life of donated RBCs preserved by the normal method is very short, limiting their clinical application. Cryopreservation is a reliable and effective technology for the long‐term storage of RBCs. During the process, ice formation will cause irreversible injuries to RBCs. Glycerol is used as a cryoprotectant (CPA) for RBCs to mitigate cryoinjuries. But it severely induces RBC hemolysis and deformation. This review introduces some biocompatible and effective CPAs used for RBC cryopreservation, outlining recent advances. Currently, there is no uniform standard to determine whether a CPA is suitable for RBCs. According to previous studies, we summarize for the first time comprehensive methods to evaluate the performance of CPAs by ensuring their safety and effectiveness. The safety of CPAs is defined as the degree of damage to RBCs, while the effectiveness is demonstrated by the properties of thawed RBCs, including membrane properties, protein activities, rheological properties, and metabolites levels. A novel CPA that is confirmed suitable for RBCs by these methods may be applied to other cells. We believe comprehensive methods can thoroughly evaluate the performance of CPAs and promote the development of transfusion therapy in clinic.

Probiotics are of great economic importance mainly because of the various health benefits it offers. It is important for the probiotics to maintain its viability and health-promoting properties during the manufacturing process and storage. Freeze- drying is a common method to preserve the probiotics. The probiotic bacteria were freeze-dried using different cryoprotectants and its effect on the stability and viability of the probiotics were studied. The freeze dried culture was stored at a temperature of -20°C and 4°C to check for the survival rate. The plate count method was used to determine the survival rate of bacterial cells before and after freeze-drying. The inactivation rate constant k, was determined. Production of vitamins - Riboflavin and Cobalamin in probiotic bacteria was analysed by HPLC and the presence was confirmed by FTIR. Yoghurt was prepared using monoculture and co-culture of Lactobacillus sp. and Lactococcus sp. The proximate analysis indicated that the yoghurt was good source of protein, fat and energy.

Cryopreservation of male gametes is one of the most important methods of assisted reproductive technologies, which allows preserving gametes for research or further use. However, the fertilizing ability of spermatozoa after cryopreservation decreases by 30–70%, which makes it urgent to search for new substances with cryoprotective properties. The review considers the main causes of cell damage during cryopreservation. The relevance of methods for assessing the formation of crystals and the physicochemical properties of cryoprotective media depending on various compositions is discussed. The problem of stabilization of the spermatozoa membrane during cryopreservation is considered. A possible solution to the problem of membrane integrity may consist in modification of the basic cryoprotective media with yolk emulsion or development of methods for saturation of the membrane phospholipid layer with cholesterol. [ABSTRACT FROM AUTHOR]

This study is aimed at evaluating the effect of different extenders on the cryopreservation of semen from Africanized honeybees (A. mellifera). Semen from honeybee drones from 10 different colonies was obtained by endophallus exposure technique and immediately evaluated for motility, viability using fluorescent probes, functional membrane integrity using the water test, and morphology. Samples from each colony were divided in three aliquots and subjected to a dilution ratio of 12:1 (diluent: semen) using Tris, Tris + egg yolk (Tris+EY), and Collins extender. Samples were cryopreserved and stored in liquid nitrogen for one week and then rewarmed and reevaluated. Immediate dilution provoked no significant effect on sperm motility and functional membrane integrity, regardless of the extender used; however, the greatest values (P < 0.05) for normal sperm morphology were found at the use of isolate Tris (69.3 ± 1.9%). After thawing, there were no significant differences among extenders with relation to the preservation of sperm motility, viability, and functional membrane integrity, but the Tris extender provided the highest post-thawing values (P < 0.05) for sperm normal morphology (49.2 ± 4.9%) while the Collins extender provoked the highest amounts (P < 0.05) of curled tail defects (67.5 ± 3.2%). Moreover, the Tris was the only extender at preserving the proportion of normal sperm after thawing similar to what was verified for fresh samples. In summary, we suggest the use of a Tris-based extender for the cryopreservation of Africanized honeybee semen. [ABSTRACT FROM AUTHOR]

The relative cryoprotective effects of flaxseed protein hydrolysate and pectin in comparison with conventional cryoprotectant (sucrose + sorbitol + sodium tripolyphosphates) on stabilization of proteins in surimi of Capoor (Cyprinus carpio) were investigated during freezing (−20°C for 4 months) and chilling storage (4°C for 10 days). Although pectin caused to improve water‐holding capacity (27.8%; 4°C and 21.5%; −20°C) on account of highly more inhibitory impact on the ice crystals growth, the protein denaturation may have occurred. It can be related to higher reduction in the amount of salt extractable protein (%) and the immeasurable value of thiol group in surimi formulation containing pectin compared with other cryoprotectants. The results of modeling surimi samples showed that salt extractable protein and sulfhydryl content were in good agreement with the first‐order reaction model at −20°C and second‐order kinetic model at 4°C. In comparison with other samples, samples treated with flaxseed protein showed the lowest reaction rate constant during chilled and frozen storage. The results confirmed that flaxseed protein with no sweetness and considerable caloric value had a cryoprotective effect similar to sucrose + sorbitol + polyphosphate and even better. [ABSTRACT FROM AUTHOR]

Deep eutectic solvents (DESs) and ionic liquids (ILs) are highly tailorable solvents that have shown a lot of promise for a variety of applications including cryopreservation, drug delivery, and protein stabilisation. However, to date, there is very limited information on the detailed interactions of these solvents with mammalian cells. In this work, we studied six DESs and one IL that show promise as cryoprotective agents, applying synchrotron macro–ATR–FTIR to examine their effects on key biochemical components of HaCat mammalian cells. These data were paired with resazurin metabolic assays and neutron reflectivity experiments to correlate cellular interactions with cellular toxicity. Stark differences were observed even between solvents that shared similar components. In particular, it was found that solvents that are effective cryoprotective agents consistently showed interactions with cellular membranes, while high toxicity correlated with strong interactions of the DES/IL with nucleic acids and proteins. This work sheds new light on the interactions between novel solvents and cells that may underpin future biomedical applications.

The origins of low-temperature tissue storage research date back to the late 1800s. Over half a century later, osmotic stress was revealed to be a main contributor to cell death during cryopreservation. Consequently, the addition of cryoprotective agents (CPAs) such as dimethyl sulfoxide (DMSO), glycerol (GLY), ethylene glycol (EG), or propylene glycol (PG), although toxic to cells at high concentrations, was identified as a necessary step to protect against rampant cell death during cryopreservation. In addition to osmotic stress, cooling and thawing rates were also shown to have significant influence on cell survival during low temperature storage. In general, successful low-temperature cell preservation consists of the addition of a CPA (commonly 10% DMSO), alone or in combination with additional permeating or non-permeating agents, cooling rates of approximately 1ºC/min, and storage in either liquid or vapor phase nitrogen. In addition to general considerations, cell-specific recommendations for hepatocytes, pancreatic islets, sperm, oocytes, and stem cells should be observed to maximize yields. For example, rapid cooling is associated with better cryopreservation outcomes for oocytes, pancreatic islets, and embryonic stem cells while slow cooling is recommended for cryopreservation of hepatocytes, hematopoietic stem cells, and mesenchymal stem cells. Yields can be further maximized by implementing additional pre-cryo steps such as: pre-incubation with glucose and anti-oxidants, alginate encapsulation, and selecting cells within an optimal age range and functional ability. Finally, viability and functional assays are critical steps in determining the quality of the cells post-thaw and improving the efficiency of the current cryopreservation methods.

Low temperature stress significantly threatens crop productivity and economic sustainability. Plants counter this by deploying advanced molecular mechanisms to perceive and respond to cold stress. Transmembrane proteins initiate these responses, triggering a series of events involving secondary messengers such as calcium ions (Ca2+), reactive oxygen species (ROS), and inositol phosphates. Of these, calcium signaling is paramount, activating downstream phosphorylation cascades and the transcription of cold-responsive genes, including coldregulated (COR) genes. This review focuses on how plants manage freezeinduced damage through dual strategies: cold tolerance and cold avoidance. Tolerance mechanisms involve acclimatization to decreasing temperatures, fostering gradual accumulation of cold resistance. In contrast, avoidance mechanisms rely on cryoprotectant molecules like potassium ions (K+), proline, glycerol, and antifreeze proteins (AFPs). Cryoprotectants modulate intracellular solute concentration, lower the freezing point, inhibit ice formation, and preserve plasma membrane fluidity. Additionally, these molecules demonstrate antioxidant activity, scavenging ROS, preventing protein denaturation, and subsequently mitigating cellular damage. By forming extensive hydrogen bonds with water molecules, cryoprotectants also limit intercellular water movement, minimizing extracellular ice crystal formation, and cell dehydration. The deployment of cryoprotectants is a key adaptive strategy that bolsters plant resilience to cold stress and promotes survival in freezing environments. However, the specific physiological and molecular mechanisms underlying these protective effects remain insufficiently understood. Therefore, this review underscores the need for further research to elucidate these mechanisms and assess their potential impact on crop productivity and sustainability, contributing to the progressive discourse in plant biology and environmental science. [ABSTRACT FROM AUTHOR]

Frozen storage is most widely adopted preservation method to maintain food freshness and nutritional attributes. However, at low temperature, food is prone to chemical changes such as protein denaturation and lipid oxidation. In this review, we discussed the reasons and influencing factors that cause protein denaturation during freezing, such as freezing rate, freezing temperature, freezing method, etc. From the previous literatures, it was found that frozen storage is commonly used to prevent freeze induced protein denaturation by adding cryoprotectants to food. Some widely used cryoprotectants (for example, sucrose and sorbitol) have been reported with higher sweetness and weaker cryoprotective abilities. Therefore, this article comprehensively discusses the new cryopreservation methods and providing comparative study to the conventional frozen storage. Meanwhile, this article sheds light on the freeze induced alterations, such as change in functional and gelling properties. In addition, this article could be helpful for the prolonged frozen storage of food with minimum quality related changes. Meanwhile, it could also improve the commercial values and consumer satisfaction of frozen food as well. [ABSTRACT FROM AUTHOR]

Several earthworm species are known to be able to withstand freezing. At the biochemical level, this ability is based on cryoprotectant accumulation as well as several other mechanisms. In this study, we used 1H NMR to investigate metabolomic changes in two freeze-tolerant earthworm taxa, Dendrobaena octaedra and one of the genetic lineages of Eisenia sp. aff. nordenskioldi f. pallida. A total of 45 metabolites were quantified. High concentrations of glucose were present in frozen tissues of both taxa. No other putative cryoprotectants were found. We detected high levels of glycolysis end products and succinate in frozen animals, indicating the activation of glycolysis. Concentrations of many other substances also significantly increased. On the whole, metabolic change in response to freezing was much more pronounced in the specimens of Eisenia sp. aff. nordenskioldi f. pallida, including signs of nucleotide degradation. [ABSTRACT FROM AUTHOR]

Despite its routine and frequent application, cryopreservation of human sperm is far from the desired efficacy, as freezing and thawing impair motility, viability, acrosomal unity, and DNA integrity. In this study, the authors aimed to investigate whether adding antioxidants, coenzyme Q10, and curcumin into the freezing medium provide better efficacy in the cryopreservation of human sperm. The semen samples from 40 healthy men aged 18–45 were collected in sterile containers by masturbation. Samples within normal reference values for sperm concentration (≥15 million/mL) and motility (progressive motile ≥ 32% and total motility ≥ 40%) were included in the study. Semen samples were equally divided into five groups and evaluated; i) pre-freezing sperm suspension, ii) frozen-thawed control (Ctrl) without any supplementation in freezing medium, iii) frozen-thawed with curcumin supplementation of 0.25 mM (Cur), iv) frozen-thawed coenzyme Q10 supplementation of 25 µM (CoQ10) and v) frozen-thawed curcumin (0.25 mM) plus coenzyme Q10 (25 µM) supplementation (CurCoQ10) into the freezing medium. Liquid nitrogen vapour freezing and rapid thawing were performed in each group (ii-v). Sperm motility, viability, acrosome integrity, and DNA fragmentation rates were compared and ultrastructural evaluations by transmission electron microscopy were undertaken between the groups. Additionally, the total antioxidant capacity/total oxidant capacity values were measured. According to CASA results, progressive motility was significantly higher in the CoQ10 group (9.4 ± 7.6) when compared with the Ctrl (7.1 ± 6.3), Cur (6.4 ± 4.8) and CurCoQ10 (8.1 ± 7.7) groups (p < 0.05). Flow cytometry results showed no difference in the viability and acrosome integrity values after thawing, but DNA fragmentation was significantly increased in the curcumin-added groups (p < 0.05). Acrosomal changes and sub-acrosomal defects were seen in all groups after thawing at the ultrastructural level. Mitochondrial membrane structure was preserved in CoQ10 and CurCoQ10 groups. Our results suggested that sperm ultrastructural morphology and motility were better preserved in the CoQ10 group during cryopreservation. In curcumin groups, DNA fragmentation and head defects were increased. [ABSTRACT FROM AUTHOR]

The surimi-like material made from duck (DSLM) was investigated on the freeze-thaw stability during 4 months of frozen storage using different low-sweetness cryoprotectants such as palatinose (PAL), trehalose (TRE), polydextrose (PLD), maltodextrin (MAL), sucrose and sorbitol (SS), and lactitol (LAC) at a 6% concentration. As compared to the CON, the inclusion of cryoprotectants significantly improved the water-holding capacity of all treated samples. Particularly, as compared to all of the cryoprotectants evaluated, the PLD and PAL samples displayed greater stability concerning Ca2+-ATPase activity, sulfhydryl concentration, and protein solubility. All samples' endothermic peaks declined over time, showing changes in thermal characteristics during storage. A scanning electron microscope (SEM) observation of the CON and LAC-treated samples after 4 months of frozen storage revealed an increase in the number of pores. These findings imply that low-sweetness PLD and PAL outperformed the usual cryoprotectant combination of sucrose and sorbitol (SS) and that they could be used in frozen DSLM. They provide greater water-holding capacity, anti-freezing characteristics, lowered sulfhydryl oxidation, increased protein solubility, and structural integrity, which renders them useful components for innovation in the frozen surimi-like material industry. [ABSTRACT FROM AUTHOR]

The present study aimed to identify the efficient cryoprotectants formulations on the survival of Lactobacillus plantarum CJLP133 microencapsulated in calcium alginate during freeze-drying. Trehalose, maltodextrin, sorbitol, sucrose, and soy peptone were selected to develop optimal formulations of cryoprotectants that were added to the calcium alginate microencapsulation using extrusion technology. The soy peptone after being combined with trehalose and maltodextrin was determined to have the highest cell viability than other combinations of sucrose and sorbitol during freeze-drying after microencapsulation (p ≤ 0.05). After storage for 12 weeks at 40 °C, the viability of microencapsulated L. plantarum was reduced by 1.03 log CFU mL− 1, while free cell viability was reduced by 1.68 log CFU mL− 1. Additionally, microencapsulated cells with cryoprotectants were resistant to simulated stomach–duodenum passage (SSDP) in vitro model, compared to free cells (p ≤ 0.05). This study highlighted that soy peptone was the most suitable cryoprotectant with trehalose and that the addition of maltodextrin during freeze-drying after microencapsulation of L. plantarum conferred enhanced survival during long-term storage at high temperatures and under adverse SSDP conditions. [ABSTRACT FROM AUTHOR]

The production of proteins capable of binding ice is one of the strategies evolved in biological organisms for survival in cold ecosystems where there is a risk of freezing. These proteins have an important ability to bind to the surface of ice, influence its growth and prevent cell damage and death. To understand the nature of interaction of such proteins with ice, it is necessary to know their structure. This study contributes to the understanding of the structural and dynamic mechanisms of action of ice-binding proteins that ensure the adaptation of organisms in critical conditions. The study of the contribution of proteins capable of binding ice to adaptation to cold conditions opens up wide opportunities in solving a number of important medical problems, including the development of effective cellular and organ cryoprotectants, as well as long-term storage of food products without loss of their consumer properties in the food industry. [ABSTRACT FROM AUTHOR]

Vacuum freeze-drying is one of the common methods to prepare probiotic powder. However, the low temperature and low moisture content could lead to the death of probiotics. The method of adding cryoprotectants is efficient to improve the survival rate of probiotics during vacuum freeze-drying. The synergistic combination of different cryoprotectants could play a better protective effect. However, the principle of the synergistic combination of different cryoprotectants during vacuum freeze-drying is not clear. In this study, literature review and analysis were firstly used to reveal commonly used cryoprotectants for coccus. Taking the price of cryoprotectants into account, the combination is preliminarily determined as sucrose/skim milk/sodium glutamate. Then, taking Streptococcus thermophilus 937 as the research target strain, the effect of this cryoprotectant combination was verified by a single-factor screening experiment and concentration optimization of cryoprotectants. It was found that the preferred cryoprotectant combination for Streptococcus thermophilus 937 is 6% sucrose/8% skim milk/4% sodium glutamate, with the freeze-dried survival rate and viable counts being 90.59% and 1.78 × 1011CFU/g, respectively. Meanwhile, this cryoprotectant combination also had a good protective effect on the other two cocci, Streptococcus thermophilus Grx02 and Enterococcus faecium 218. This result might have guiding and reference significance on the vacuum freeze-drying of coccus. [ABSTRACT FROM AUTHOR]

[Display omitted] The structural maintenance of therapeutic proteins during formulation and/or storage is a critical aspect, particularly for multi-domain and/or multimeric proteins which usually exhibit intrinsic structural dynamics leading to aggregation with concomitant loss-of-function. Protein freeze-drying is a widely used technique to preserve protein structure and function during storage. To minimize chemical/physical stresses occurring during this process, protein stabilizers are usually included, their effect being strongly dependent on the target protein. Therefore, they should be screened for on a time-consuming case-by-case basis. Herein, differential scanning fluorimetry (DSF) and isothermal denaturation fluorimetry (ITDF) were employed to screen, among different classes of freeze-drying additives, for the most effective stabilizer of the model protein human phenylalanine hydroxylase (hPAH). Correlation studies among retrieved DSF and ITDF parameters with recovered enzyme amount and activity indicated ITDF as the most appropriate screening method. Biochemical and biophysical characterization of hPAH freeze-dried with ITDF-selected stabilizers and a long-term storage study (12 months, 5 ± 3 °C) showed that the selected compounds prevented protein aggregation and preserved hPAH structural and functional properties throughout time storage. Our results provide a solid basis towards the choice of ITDF as a high-throughput screening step for the identification of protein freeze-drying protectors. [ABSTRACT FROM AUTHOR]

Probiotic formula for the vaginal tract must contain high numbers of viable beneficial bacteria that maintain their characteristics during the production and further storage, in order to exert the claimed probiotic effect. Four probiotic strains—Lactobacillus gasseri CRL1320, Limosilactobacillus reuteri CRL1324, Ligilactobacillus salivarius CRL1328 and Lacticaseibacillus rhamnosus CRL1332—originally isolated from vagina of healthy women exhibiting beneficial properties were evaluated. Therefore, the optimization of growth parameters and dry biomass production with high viability while preserving probiotic features of strains is a great challenge. Different growth conditions in MRS medium were set up in a laboratory bioreactor; two initial pH and agitation were recorded speeds during static or controlled fermentations. Production conditions of 37 °C with controlled pH 6.5 and 5.5-MRS with 150 and 75 rpm stirring speeds were used for CRL1329 and CRL1332 strains, respectively, while static and free pH MRS for CRL1324 and CRL1328 probiotics allowed us to obtain maximal cell viability counts. However, during 12 h of fermentation, biomass yields of 19.3, 16.2 and 15.2 g/L were achieved when CRL1329, CRL1328 and CRL1324 probiotic strains were grown in static and free pH MRS. The highest biomass yield for CRL1332 strain was produced under controlled MRS-initial pH 6.5 and 75 rpm fermentation conditions. To preserve probiotic high biomass viability, freeze-drying was carried out in the presence of different cryoprotective agents. Thus, the highest viable numbers (10.9–11.8 log CFU/g) with survival rates between 91.3 and 95.6% were attained in the presence of 10% trehalose (L. reuteri and L. salivarius), lactose (L. rhammosus) and lactose + trehalose + sucrose mix (L. gasseri). When stability during post-freeze-drying storage was evaluated, probiotic strains showed a remarkably higher viability recovery when stored at 4 °C than at 25 °C for 12 and 3 months, respectively. In addition, surface characteristics of vaginal probiotics were affected to different extents during storage depending on the strain, protective agent and storage time/temperature. Critical factors for growth conditions, drying process and storage stability of probiotic lactobacilli strains were optimized in view to preserve cell high viability and surface features for the design of vaginal probiotic formula. [ABSTRACT FROM AUTHOR]

Deep eutectic solvents (DESs) and ionic liquids (ILs) are highly tailorable solvents that have shown a lot of promise for a variety of applications including cryopreservation, drug delivery, and protein stabilisation. However, to date, there is very limited information on the detailed interactions of these solvents with mammalian cells. In this work, we studied six DESs and one IL that show promise as cryoprotective agents, applying synchrotron macro–ATR–FTIR to examine their effects on key biochemical components of HaCat mammalian cells. These data were paired with resazurin metabolic assays and neutron reflectivity experiments to correlate cellular interactions with cellular toxicity. Stark differences were observed even between solvents that shared similar components. In particular, it was found that solvents that are effective cryoprotective agents consistently showed interactions with cellular membranes, while high toxicity correlated with strong interactions of the DES/IL with nucleic acids and proteins. This work sheds new light on the interactions between novel solvents and cells that may underpin future biomedical applications. [ABSTRACT FROM AUTHOR]

Cryopreservation is the most prevalent method of long‐term cell preservation. Effective cell cryopreservation depends on freezing, adequate storage, and correct thawing techniques. Recent advances in cryopreservation techniques minimize the cellular damage which occurs while processing samples. This article focuses on the fundamentals of cryopreservation techniques and how they can be implemented in a variety of clinical settings. The article presents a brief description of each of the standard cryopreservation procedures, such as slow freezing and vitrification. Alongside that, the membrane permeating and nonpermeating cryoprotectants are briefly discussed, along with current advancements in the field of cryopreservation and variables influencing the cryopreservation process. The diminution of cryoinjury incurred by the cell via the resuscitation process will also be highlighted. In the end application of cryopreservation techniques in many fields, with a special emphasis on stem cell preservation techniques and current advancements presented. Furthermore, the challenges while implementing cryopreservation and the futuristic scope of the fields are illustrated herein. The content of this review sheds light on various ways to enhance the output of the cell preservation process and minimize cryoinjury while improving cell revival. [ABSTRACT FROM AUTHOR]