Different Concentrations Of Trehalose Research Articles

Structure and slow dynamics of protein hydration water with cryopreserving DMSO and trehalose upon cooling.

We study, through molecular dynamics simulations, three aqueous solutions with one lysozyme protein and three different concentrations of trehalose and dimethyl sulfoxide (DMSO). We analyze the structural and dynamical properties of the protein hydration water upon cooling. We find that trehalose plays a major role in modifying the structure of the network of HBs between water molecules in the hydration layer of the protein. The dynamics of hydration water presents, in addition to the α-relaxation, typical of glass formers, a slower long-time relaxation process, which greatly slows down the dynamics of water, particularly in the systems with trehalose, where it becomes dominant at low temperatures. In all the solutions, we observe, from the behavior of the α-relaxation times, a shift of the Mode Coupling Theory crossover temperature and the fragile-to-strong crossover temperature toward higher values with respect to bulk water. We also observe a strong-to-strong crossover from the temperature behavior of the long-relaxation times. In the aqueous solution with only DMSO, the transition shifts to a lower temperature than in the case with only lysozyme reported in the literature. We observe that the addition of trehalose to the mixture has the opposite effect of restoring the original location of the strong-to-strong crossover. In all the solutions analyzed in this work, the observed temperature of the protein dynamical transition is slightly shifted at lower temperatures than that of the strong-to-strong crossover, but their relative order is the same, showing a correlation between the motion of the protein and that of the hydration water.

Antagonistic yeast and trehalose-enriched gelatin film: A bioactive antifungal packaging film for cherry tomato preservation

To extend the shelf-life of cherry tomatoes infected with Botrytis cinerea, this study analyzed the effect of Pichia membranifaciens (PM) on a composite film of gelatin (Gel) and trehalose (Tre), and explored the effect of different concentrations of trehalose on the yeast in the bioactive film and its control effect on cherry tomatoes. Scanning electron microscopy showed that Tre/Gel-based film can be applied as effective carriers to protect the cellular integrity of PM. Fourier-transform infrared spectroscopy, thermogravimetric analysis, and tests of mechanical properties showed that there was interaction between PM and the Tre/Gel base film, which maintained the thermal stability of the base film. The addition of PM reduced the water vapor permeability, hygroscopicity, and moisture content of the Tre/Gel base film, effectively extending the active film's storage time. This study also explored the growth curve of PM and found that 15% trehalose was effective in maintaining PM activity and could result in a 30-day survival rate of 62% for PM in Tre/Gel films, with 72.41% inhibition of B. cinerea. Finally, the PM/Tre/Gel film significantly reduced the incidence of decay in cherry tomatoes and improved the storage quality and fungal resistance of the fruit. Thus, PM/Tre/Gel films may be applied as a new antifungal packaging material for the preservation of fresh fruit.

Effect of antioxidants in cryopreservation media on spotted halibut (Verasfer variegatus) sperm quality during cryopreservation

In fish hatcheries, sperm cryopreservation is a helpful tool for managing broodstock and artificial fertilization. However, cold shock can cause sperm injury resulting in decreasing sperm motility, low cell viability, structural damage to the plasma membrane and DNA fragmentation. This parameters are critical factor for the assessment of cryopreserved sperm for commercial scale application. In recent years, antioxidants have been increasingly useful for treating physiological issues in fish, including in reproduction and cryopreservation procedures. The purpose of this study was to examine the effects of supplementation of various antioxidants, including trehalose, reduced glutathione (GSH), Mito-TEMPO (MT), and rosmarinic acid (RA), during fish sperm cryopreservation on sperm motility, cell survival rate, and DNA damage. One-way ANOVA and Duncan's multiple range tests (p > 0.05) were used to compare the parameters. Sperm from adult fish were cryopreserved in 10% DMSO with 300 mM sucrose (1:1) supplemented with different concentrations of trehalose (0, 50, 100, 150, 200, and 250 mM), GSH (0, 2, 4, 6, 8, and 10 mM), MT (0, 25, 50, 75, 100, 125, 150, 175, and 200 μM), or RA (0, 25, 50, 75, and 100 μM). Compared with the control groups, antioxidant supplementation in sperm was more efficient in increasing post-thaw motility, maintaining cell survival rates, and inhibiting the increase in DNA damage that occurs during non-activated sperm during storage. The results demonstrated that the post-thaw motility of sperm was higher in 8 mM GSH (85.71 ± 0.04%), followed by 6 mM GSH (82.97 ± 1.30%), 75 and 25 μM RA (82.93 ± 0.93% and 82.76 ± 1.42%, respectively) and 100 mM trehalose (80.39 ± 0.61%) with not statistically different. The cell survival rate, VCL, and VSL of frozen–thawed sperm were higher in 100 mM trehalose resulted 84.22 ± 2.24%, 114.72 ± 2.00 μm/s, and 95.49 ± 0.70 μm/s, respectively. Mito-TEMPO at concentrations 100 μM resulted lower percentages of tail DNA (1.60 ± 0.08%) with not statistically different with 100 mM trehalose. In conclusion, the present study indicated that antioxidant 100 mM trehalose provided the most pronounced protective effect in improving frozen-thawed quality of frozen–thawed spotted halibut sperm.

Effects of different phosphorus-free water-retaining agents on the quality of frozen tilapia fillets.

Tilapia is an economically important fish worldwide, but its quality is affected by storage practices. To improve the quality of tilapia fillets during frozen storage, we examined the effect of pretreatment with various combinations of different concentrations of trehalose, potassium bicarbonate, and chitosan. Following pretreatment, we analyzed the tilapia fillets using quality indicators, including soaking weight gain, coating weight gain, water‐holding capacity, thawing loss, pH, Ca2+‐ATPase activity, and texture characteristics. Water distribution was analyzed using low‐field nuclear magnetic resonance and the optimal combination of water‐retaining agents was obtained using an L8(27) orthogonal experiment. The results showed that trehalose, potassium bicarbonate, and chitosan improved fillet quality at pretreatment concentrations of 5%–8%, 1.0%, and 0.5%, respectively. The optimal combination was 4% trehalose plus 1.2% potassium bicarbonate plus 0.2% chitosan. The Ca2+‐ATPase activity and mastication property of the frozen fillets that were pretreated with the optimized formulation were 1.39 μmol Pi/mg protein·h and 8.55 mJ, respectively, which were 43.3% and 80.0% greater, respectively, than that of the control group. Using a suitable concentration and combination of water‐retaining agents cannot only lock‐in the internal water content of frozen tilapia fillets but also improve their quality during frozen storage. These results can inform practical storage practices of similar aquatic products.

The Assessment of Various Factors Affecting the Postwarming Viability and Developmental Capability of Goat Metaphase II Oocytes Vitrified by Cryotop.

The effects of the equilibration time, the vitrification procedure, and the warming procedure on the quality of goat oocytes vitrified by Cryotop were assessed. In the first part of the study, oocytes were exposed to 10% dimethyl sulfoxide (DMSO) and 10% ethylene glycol (EG) for 1, 3, 5, or 10 minutes, respectively, followed by vitrification. In the second part, after equilibration in 7.5% DMSO +7.5% EG for 3 minutes, 10% DMSO +10% EG for 3 minutes, or 4% EG for 10 minutes, oocytes were equilibrated in 15% DMSO +15% EG, 20% DMSO +20% EG, or 35% EG for 30 seconds before vitrification. The vitrification procedures were designated as first vitrification procedure (VPI), second vitrification procedure (VPII), and third vitrification procedure (VPIII), respectively. In the third part, oocytes vitrified using VPIII were warmed by the three procedures (first warming procedure [TPI], second warming procedure [TPII], or third warming procedure [TPIII]) containing different concentrations of trehalose. The results showed that after equilibration for 1 or 3 minutes in 10% DMSO and 10% EG, the viability and developmental capability of vitrified oocytes were significantly superior to the groups after equilibration for over 5 minutes (p < 0.05). With the VPIII procedure, the frequencies with normal morphology, cleavage, and blastocyst formation of vitrified oocytes were 91.87% ± 4.14%, 76.51% ± 4.37%, and 39.84% ± 2.91%, respectively, demonstrating a significant increase compared to the VPI or VPII group (p < 0.05). The rates of vitrified oocytes with normal morphology and cleavage in the TPI group were higher than the TPII or TPIII group (p < 0.05). In conclusion, equilibration in 10% DMSO and 10% EG for <3 minutes benefits the viability of vitrified oocytes. EG may be more efficient for vitrification of goat oocytes compared to DMSO. Higher concentrations (more than 1 M) of trehalose enhance cryosurvival of goat oocytes when warming.

Trehalose: effect on cryopreservation of umbilical cord blood-derived hematopoietic stem cells

Introduction: Concerns over dimethyl sulfoxide (DMSO) toxicity, related to adverse reactions after hematopoietic stem cells (HSCs) therapy, warrant the development of an optimized DMSO-free or DMSO-reduced cryopreservation protocol for the quality and safety of HSCs. In this regard, the ideal concentration of trehalose, as a non-toxic natural cryoprotectant, is still an area of research. Based on the outcome of our previous study on lower concentrations of trehalose, the present study was focused on evaluating its cryoprotective efficacy at an increased concentration (0.5 M) on HSCs compared to 10% DMSO. This is a laboratory-based experimental study. Material and methods: The separated mononuclear cells collected from umbilical cord blood were set for culture up to two passages to get HSCs. The two different concentrations of trehalose, with and without 5% DMSO, were considered as freezing media for the preservation of the harvested HSCs in a slow freezing set up. Two sequential functional assays, viability followed by hematopoietic colony-forming unit assay, were performed with post-thawed cells of freezing media used in this study. Seventeen cord blood samples were selected. Results: Study results revealed 0.5 M trehalose and DMSO 5% showed the highest viability of 91.8 ±2.8% of HSCs. 5% DMSO inclusion to trehalose (0.5 M) ameliorated hematopoietic colonies such as erythroid and myeloid colonies with no significant difference from that of 10% DMSO. Conclusion: 0.5 M trehalose has proved to be a better concentration than 10% DMSO alone. This experimental study needs further transplantation-based clinical trials using post-thawed cells to ensure the safety of preserved HSCs from cord blood and other sources.

Trehalose: effect on cryopreservation of umbilical cord blood-derived hematopoietic stem cells

Introduction: Concerns over dimethyl sulfoxide (DMSO) toxicity, related to adverse reactions after hematopoietic stem cells (HSCs) therapy, warrant the development of an optimized DMSO-free or DMSO-reduced cryopreservation protocol for the quality and safety of HSCs. In this regard, the ideal concentration of trehalose, as a non-toxic natural cryoprotectant, is still an area of research. Based on the outcome of our previous study on lower concentrations of trehalose, the present study was focused on evaluating its cryoprotective efficacy at an increased concentration (0.5 M) on HSCs compared to 10% DMSO. This is a laboratory-based experimental study. Material and methods: The separated mononuclear cells collected from umbilical cord blood were set for culture up to two passages to get HSCs. The two different concentrations of trehalose, with and without 5% DMSO, were considered as freezing media for the preservation of the harvested HSCs in a slow freezing set up. Two sequential functional assays, viability followed by hematopoietic colony-forming unit assay, were performed with post-thawed cells of freezing media used in this study. Seventeen cord blood samples were selected. Results: Study results revealed 0.5 M trehalose and DMSO 5% showed the highest viability of 91.8 ±2.8% of HSCs. 5% DMSO inclusion to trehalose (0.5 M) ameliorated hematopoietic colonies such as erythroid and myeloid colonies with no significant difference from that of 10% DMSO. Conclusion: 0.5 M trehalose has proved to be a better concentration than 10% DMSO alone. This experimental study needs further transplantation-based clinical trials using post-thawed cells to ensure the safety of preserved HSCs from cord blood and other sources.

The combination of trehalose and glycerol: an effective and non-toxic recipe for cryopreservation of human adipose-derived stem cells

BackgroundAdipose-derived stem cells (ADSCs) promote tissue regeneration and repair. Cryoprotective agents (CPAs) protect cells from cryodamage during cryopreservation. Safe and efficient cryopreservation of ADSCs is critical for cell-based therapy in clinical applications. However, most CPAs are used at toxic concentrations, limiting their clinical application.ObjectiveThe aim of this study is to develop a non-toxic xeno-free novel CPA aiming at achieving high-efficiency and low-risk ADSC cryopreservation.MethodsWe explored different concentrations of trehalose (0.3 M, 0.6 M, 1.0 M, and 1.25 M) and glycerol (10%, 20%, and 30% v/v) for optimization and evaluated and compared the outcomes of ADSCs cryopreservation between a combination of trehalose and glycerol and the commonly used CPA DMSO (10%) + FBS (90%). All samples were slowly frozen and stored in liquid nitrogen for 30 days. The effectiveness was evaluated by the viability, proliferation, migration, and multi-potential differentiation of the ADSCs after thawing.ResultsCompared with the groups treated with individual reagents, the 1.0 M trehalose (Tre) + 20% glycerol (Gly) group showed significantly higher efficiency in preserving ADSC activities after thawing, with better outcomes in both cell viability and proliferation capacity. Compared with the 10% DMSO + 90% FBS treatment, the ADSCs preserved in 1.0 M Tre + 20% Gly showed similar cell viability, surface markers, and multi-potential differentiation but a significantly higher migration capability. The results indicated that cell function preservation can be improved by 1.0 M Tre + 20% Gly.ConclusionsThe 1.0 M Tre + 20% Gly treatment preserved ADSCs with a higher migration capability than 10% DMSO + 90% FBS and with viability higher than that with trehalose or glycerol alone but similar to that with 10% DMSO + 90% FBS and fresh cells. Moreover, the new CPA achieves stemness and multi-potential differentiation similar to those in fresh cells. Our results demonstrate that 1.0 M Tre + 20% Gly can more efficiently cryopreserve ADSCs and is a non-toxic CPA that may be suitable for clinical applications.

Protective effects of trehalose against Mn-induced α-synuclein oligomerization in mice: Involvement of oxidative stress and autophagy.

Overexposure to manganese (Mn) is widely known to induce alpha-synuclein (α-Syn) oligomerization, which has been attributed to the oxidative damage of α-Syn protein. Trehalose has been shown to induce autophagy and serve as a chemical chaperone, but little information has been reported about its effect on Mn-induced α-Syn oligomerization. In this study, we investigate whether trehalose can effectively interfere with Mn-induced α-Syn oligomerization, using different concentrations of trehalose (2% and 4% (g/vol [mL])) in a mouse model of manganism. After 6 weeks of exposure to Mn, both oxidative stress and autophagy were activated and resulted in α-Syn oligomerization and neuronal cell damage in the mouse brain tissue. Our results also revealed that pretreatment with trehalose significantly reduced the oxidative damage to α-Syn protein and increased autophagy activation. These findings clearly demonstrated that trehalose can relieve Mn-induced α-Syn oligomerization and neuronal cell damage through its anti-oxidative and autophagy-inducing effects.

Role of trehalose on antioxidant defense system and some osmolytes of quinoa plants under water deficit

BackgroundFor scavenging reactive oxygen species, plant possess effective system that protect them from destructive oxidative reaction. Parts of this system as osmoprotectants and antioxidative enzymes are key elements in the defense mechanisms. A field experiment was conducted to evaluate the potential of foliar treatment of trehalose (Tre) with different concentrations (0, 0.1 mM, or 0.5 mM) in improving antioxidant defense system of quinoa plant under normal irrigation and drought stress conditions.ResultsDrought stress caused significant increases in some osmoprotectants as glucose, trehalose, TSS, free amino acids, and proline. Meanwhile, trehalose foliar treatment with different concentrations significantly decreases in free amino acids and proline contents. More accumulation of the tested organic solutes of leaves (glucose, sucrose, trehalose, TSS) of the trehalose-treated plant in both normal irrigated and drought-stressed quinoa plants as compared with the corresponding controls. Treating quinoa plants with trehalose resulted in significant decrease in lipid peroxidation, hydrogen peroxide contents, and LOX activity in normal irrigated and drought-stressed plants. These decreases correlated with significant increases in total phenolic contents as compared with untreated control. Different concentrations of trehalose resulted in significant increases in antioxidant enzymes. Maximum increase antioxidant enzymes were observed by treating plants Tre at 0.5 mM either under normal irrigation or drought conditions.ConclusionIt could be concluded that foliar spray of trehalose was effective in improving quinoa performance by reducing hydrogen peroxide free radical and by enhancing antioxidant compounds (phenolics), compatible osmolytes, membrane stability, and antioxidant enzymes.

Developmental competence of in vitro matured ovine oocytes vitrified in solutions with different concentrations of trehalose.

This study aimed to determine the optimum concentration of trehalose in solutions used for vitrification of in vitro matured (IVM) ovine oocytes. IVM oocytes were randomly divided into four experimental (vitrified) and one control (fresh) groups. Experimental groups were treated with different concentrations (0.0, 0.25, 0.5 and 1.0M) of trehalose. After warming, some viable oocytes were exposed to 0.25% pronase to test zona pellucida hardening, whereas the others were fertilized and cultured in vitro for 8days to evaluate their developmental competence. Blastocysts quality was assessed by differential staining and TUNEL test. Survival and developmental rates of oocytes vitrified in the presence of 0.5M trehalose were significantly higher than those of the other vitrified groups. Furthermore, there was a significant difference between fresh and vitrified groups in total blastocyst rate. Analysis of blastocysts quality also revealed a significant difference between the group treated with 0.5M trehalose and other groups in terms of apoptotic index. Furthermore,zona pellucida digestion time period was longer in trehalose-free (0.0M) group compared to other groups. In conclusion, we found that IVM ovine oocytes vitrified in solutions containing 0.5M trehalose are fertilization-competent and are able to produce good-quality blastocysts with an apoptotic index comparable to that of the fresh oocytes. Therefore, 0.5M may be considered the optimum concentration of trehalose to be used in solutions prepared for vitrification of oocytes.

Cryoprotectant effect of trehalose in extender on post-thaw quality and in vivo fertility of water buffalo (Bubalus bubalis) bull spermatozoa.

This study was designed to ascertain the cryoprotectant effects of different concentrations of trehalose [0 (T0), 25 (T25), 35 (T35), 45 (T45) mm], egg yolk [20% (E20), 15% (E15) v/v] and glycerol [7% (G7), 5% (G5) v/v] in Tris-citric acid-based extender on post-thaw quality and in vivo fertility of buffalo bull spermatozoa. Twenty-five ejaculates were collected from five bulls and split into four parts. After that, the split ejaculates from each of the bull were diluted either in T0E20G7 (control) or T25E20G5 or T35E15G5 or T45E15G5 extender. Finally, the sperm suspension was frozen in 0.54-ml French straws. Post-thaw sperm total motility (%), progressive motility (%), rapid velocity (%), average path velocity (μm/s), straightline velocity (μm/s), curvilinear velocity (μm/s), linearity (%), plasma membrane and acrosome integrities (%) were higher (p<.05) in T45E15G5 extender as compared to other treatment groups and control. The fertility rate (56.8% versus 41.3%) was higher (p<.05) in buffaloes inseminated with semen doses cryopreserved in extender containing T45E15G5 combination of cryoprotectants than the control. In conclusion, addition of 45mm trehalose along with 15% egg yolk and 5% glycerol in extender improves the post-thaw quality and in vivo fertility of buffalo bull spermatozoa.

Trehalose improves rabbit sperm quality during cryopreservation

High levels of reactive oxygen species are associated with spermatozoa cryopreservation, which bring damage to functional spermatozoa. The aim of the present study was to investigate whether and how the freezing extenders supplemented with trehalose was beneficial for the survival of rabbit spermatozoa. semen was diluted with Tris-citrate-glucose extender addition of different concentrations of trehalose. Addition of 100 mM trehaose significantly improved post-thaw rabbit sperm parameters, such as motility, acrosome integriy, membrane integrity and mitochondrial membrane potential. Moreover, when freezing extenders supplemented with trehalose, activities of catalase (CAT), superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) of post-thaw spermatozoa were enhanced, meanwhile, reactive oxygen species (ROS) level and Malondialdehyde (MDA) content were decreased. The results suggest that freezing extenders supplemented with 100 mM trehalose resulted in less ROS level and MDA content, higher motility and mitochondrial membrane potential as well as the integrity of acrosome and plasma membrane. Supplementation of trehalose with freezing extenders is beneficial to the rabbit breeding industry.

Internal evaluation of impregnation treatment of waterlogged wood; relation between concentration of internal materials and relaxation time using magnetic resonance imaging

The purpose of this study is to clarify the degree of impregnation resulting from treatment of internal waterlogged wood samples using MRI. On a 1.5T MR scanner, T1 and T2 measurements were performed using inversion recovery and spin-echo sequences, respectively. The samples were cut waterlogged pieces of wood treated with various impregnation techniques which were divided into different concentrations of trehalose (C12H22O11) and polyethylene glycol (PEG; HO-(C2H4O)n-H) solutions. Then these samples underwent impregnation treatment every two weeks. From the results, we found that the slope of the T1-concentration curve using linear fitting showed the value of the internal area for PEG to be higher than the external area; internal, −2.73ms/wt% (R2=0.880); external, −1.50ms/wt% (R2=0.887). Furthermore, the slope of the T1-concentration curve using linear fitting showed the values for trehalose to have almost no difference when comparing the internal and the external areas; internal, −2.79ms/wt% (R2=0.759); external, −3.02ms/wt% (R2=0.795). However, the slope of the T2-concentration curve using linear fitting for PEG showed that there was only a slight change between the internal and the external areas; internal, 0.26ms/wt% (R2=0.642); external, 0.18ms/wt% (R2=0.920). The slope of the T2-concentration curve did not show a change in linear relationship between the internal and the external areas; internal, 0.06ms/wt% (R2=0.175); external, −0.14ms/wt% (R2=0.043). In conclusion, using visualization of relaxation time T1, it is possible to obtain more detail information noninvasively concerning the state of impregnation treatment of internal waterlogged wood.

Trehalose improves semen antioxidant enzymes activity, post-thaw quality, and fertility in Nili Ravi buffaloes (Bubalus bubalis)

Our objectives were to study the effect of trehalose in extender on (1) antioxidant enzymes profile during cryopreservation (after dilution, before freezing, and after thawing), (2) in vitro quality (after thawing), and (3) in vivo fertility of Nili Ravi buffalo (Bubalus bubalis) bull spermatozoa. Semen samples (n = 20) from four buffalo bulls were diluted in Tris-citric acid–based extender having different concentrations of trehalose (0.0, 15, 30, 45, and 60 mM) and frozen in French straws. At post dilution, profile of sperm catalase (U/mL) was higher (P < 0.05) in extenders containing 15, 30, and 45 mM of trehalose as compared to control. Although profiles of superoxide dismutase (U/mL) and total glutathione (μM) were higher (P < 0.05) in extenders containing 15 and 30 mM of trehalose as compared to control. At prefreezing, sperm catalase, superoxide dismutase, and total glutathione profiles were higher (P < 0.05) in all the treatment groups as compared to control. At post thawing, the profiles of catalase and total glutathione were higher (P < 0.05) in extender containing 30-mM trehalose as compared to other treatment groups and control. Whereas, profile of superoxide dismutase was higher (P < 0.05) in extenders containing 30, 45, and 60 mM of trehalose as compared to control and 15mM group. Post thaw total sperm motility (%) was higher (P < 0.05) in extender containing 30-mM trehalose as compared to control and 15 and 60-mM groups. Although sperm progressive motility (%), rapid velocity (%), average path velocity (μm/s), straight line velocity (μm/s), curvilinear velocity (μm/s), plasma membrane (structural and functional, %), acrosome (%), and DNA (%) integrity were higher (P < 0.05) in extender containing 30 mM trehalose as compared to other treatment groups and control. The fertility rates (61% vs. 43%) were higher (P < 0.05) in buffaloes inseminated with semen doses cryopreserved in extender containing 30 mM of trehalose than the control. It is concluded that addition of 30-mM trehalose in extender improves the semen antioxidant enzymes activity, post thaw quality, and fertility in Nili Ravi buffaloes.

Study of Lactate Dehydrogenase Kinetics in Trehalose Solutions. Temperature Effect

Some organisms may adapt to extreme environmental conditions such as drought, high salinity, freezing or high temperature. The accumulation of compatible solutes in bacteria, yeast and plants is a metabolic response to heat-shock and osmotic stress. Namely, compatible solutes are those molecules which at high concentrations does not interfere with structure or function of enzymes. Trehalose has been considered so long as a compatible solute, which has showed protective/stabilizing properties in structure and function of membranes and enzymes. Studies have demonstrated that trehalose displays the highest efficiency in protecting enzyme activity under dehydration and heat shock. However, there are few reports on the effect of trehalose in enzyme kinetic parameters. In this work, the effect of physiological concentration of trehalose on enzyme kinetics of lactate dehydrogenase (LDH) type II from rabbit muscle was studied. LDH kinetics was assayed in absence and presence of different concentrations of trehalose (0.2 to 0.8M) and at different temperatures (25-35 °C). The results showed that trehalose affects the kinetic parameters Km and Vmax, and the activation energy (Ea) for LDH reaction. Interestingly, trehalose effect increased with concentration. In contrast, the raise in temperature lead to slightly relieve LDH from such effects. Kinetic parameters Vmax and Km decreased as trehalose concentration was increased. Therefore, trehalose presence lead to diminish the maximum rate of catalysis while increasing affinity for substrate in LDH. Notably, the amount of energy required to overpass the transition state was increased by trehalose.

Effects of different concentrations of sucrose or trehalose on the post-thawing quality of cattle bull semen

Abstract Objective To examine the effect of different concentrations of trehalose or sucrose (50 or 100 or 200 mM) on post-thawed quality of bull semen, cryo-preserved in Tris-citric acid-egg yolk-fructose (TCYF). Methods Semen samples were diluted in TCYF extender, TCYF + trehalose (50, 100 and 150 mM/L) or TCYF + sucrose (50, 100 and 150 mM/L) to ensure 60 million motile spermatozoa mL-1, cooled slowly up to 5 °C and equilibrated for 4 h. Semen was packed into 0.25 mL polyvinyl French straws. The straws were placed horizontally on a rack and frozen in a vapor 4 cm above liquid nitrogen (LN2) for 10 minutes then dipped in liquid LN2. Frozen straws were thawed at 37 °C for 1 min. The parameters studied were sperm motility, sperm viability, sperm abnormality, sperm membrane integrity (HOST), percent of normal intact acrosome and DNA fragmentation. Results The output data demonstrated that addition of 50–100 mM of trehalose or sucrose/L TCYF after chilling at 5 °C had significantly ( P P Conclusion It is finally concluded that the addition of 50–100 mM trehalose or sucrose/L to TCYF have a beneficial effect in chilling diluted bull semen, while the use of 50 mM trehalose or 50–100 mM sucrose had their benefits on freezing-thawing of extended semen.

Effect of different concentrations of trehalose and glycerol on the freezability of ram semen using soybean lecithin-based diluents

This study was conducted to determine the effect of different levels of trehalose and glycerol on ram sperm cryosurvival using lecithin-based diluents. Ejaculates were collected from four rams, pooled after initial evaluation, diluted with Tris-soybean lecithin extender and split into nine equal parts. A total of 0 (T0), 50 (T50) or 100 (T100) mM of trehalose and 3% (G3), 5% (G5) or 7% (G7) of glycerol were added to each part. Sperm motility, viability, plasma membrane integrity and acrosome integrity were evaluated immediately after thawing (0 h), and subsequently after 3 h and 6 h post-thawing incubation at 37°C. Results indicated that there was interaction between trehalose and glycerol on sperm motility. In addition, interaction of trehalose and glycerol with incubation time on sperm motility, viability, plasma membrane integrity and acrosome integrity was not significant (P &gt; 0.05). Sperm motility was greatest in the sperm treated with 100 mM trehalose and 7% glycerol (27%; P &lt; 0.05). The effect of trehalose was significant on viability and plasma membrane integrity of ram spermatozoa (P &lt; 0.05). The main effect of trehalose showed that sperm viability was higher in T100 (47.06%) than T50 (53.96%; P &lt; 0.05). The highest membrane integrity was observed in T100 (47.04%; P &lt; 0.05). Membrane integrity was higher (P &lt; 0.05) in G5 (49.97%) than G3 (41.56%) and there was no difference between G7 (46.86%) and G3 (41.56%; P &gt; 0.05). The best sperm viability and plasma membrane integrity was observed at 0 h (65.75% and 51.58%, respectively). It was concluded that simultaneous use of 7% glycerol and 100 mM trehalose had a desirable effect on motility of ram frozen–thawed sperm.

Effects of Different Trehalose Concentrations in a Warming Medium on Embryo Survival and Clinical Outcomes in Vitrified Human Embryos

Background/Aims: In the present study, we examined the effects of different concentrations of trehalose in a warming medium on both embryo survival and clinical outcomes in vitrified-warmed embryo transfer cycles. Methods: We retrospectively analyzed a total of 209 vitrified-warmed cycles from 177 patients who underwent in vitro fertilization or intracytoplasmic sperm injection and embryo transfer. Embryos were cryopreserved by the vitrification method and warmed in solutions containing either 0.5 or 1.0 <smlcap>M</smlcap> trehalose. We compared the 0.5 and 1.0 <smlcap>M</smlcap> trehalose warming solution groups with respect to the embryo survival rate after warming and clinical outcomes. Results: The embryo survival rate in the 1.0 <smlcap>M</smlcap> trehalose group (96.5%) was significantly higher than that in the 0.5 <smlcap>M</smlcap> trehalose group (57.0%). The percentage of embryo transfers after warming in the 1.0 <smlcap>M</smlcap> trehalose group (94.3%) was significantly higher than that in the 0.5 <smlcap>M</smlcap> trehalose group (83.7%). The clinical pregnancy rate in the 1.0 <smlcap>M</smlcap> trehalose group (25.0%) was significantly higher than that in the 0.5 <smlcap>M</smlcap> trehalose group (11.1%). Conclusion: Embryo survival and clinical pregnancy rates were higher when a 1.0 <smlcap>M</smlcap> trehalose solution was used than when a 0.5 <smlcap>M</smlcap> trehalose solution was used during the embryo warming process.

Effect of trehalose on DNA integrity of freeze-dried boar sperm, fertilization, and embryo development after intracytoplasmic sperm injection

Freeze-drying (FD) medium containing ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid (EGTA) is reported to be beneficial for maintenance of sperm DNA integrity after FD. Recently, trehalose has also been reported to have notable ability to stabilize the protein structure and biomembranes of sperm in a dry state. In this study, we examined the effect of a combination of EGTA and different concentrations of trehalose in FD medium on sperm DNA integrity and the in vitro development of IVM porcine oocytes after intracytoplasmic sperm injection (ICSI) using freeze-dried boar sperm. Ejaculated sperm from a boar were suspended in basic FD medium supplemented with 0, 3.75, 7.5, 15, 30, 60, or 90 mM trehalose and freeze-dried. After rehydration, the sperm in all groups were subjected to DNA damage detection using a Halomax kit. It was found that the level of DNA damage in 15-mM group was significantly lower than that in 0-mM group, and no difference was observed between the 15-, 7.5-, and 3.75-mM groups. Moreover, there were no significant differences in the DNA damage level among 0, 3.75 mM, and other groups treated with trehalose. When freeze-dried sperm were used for ICSI, the fertilization rates and blastocyst formation rates (observed at 10 hours and 6 days of IVC after ICSI, respectively) in the 7.5- and 15-mM groups were not different from those in 0-mM group. These results suggest that FD medium supplemented with trehalose at appropriate concentrations improves sperm DNA integrity, but does not improve fertilization and preimplantation embryo development of IVM oocytes following ICSI.