Combination Of Ethylene Glycol Research Articles

Developmental competence of frozen-thawed yak (Bos grunniens) oocytes followed by in vitro maturation and fertilization

In the present study, we examined the ability of immature germinal vesicle (GV) and subjected to in vitro matured (MII) yak oocytes to survive after cryopreservation as well as their subsequent development following in vitro maturation and fertilization. Both GV and MII oocytes were cryopreserved by using two different vitrification solutions (VS); VS-I contained 10% ethylene glycol (EG) and 10% dimethylsulfoxide (DMSO) in TCM-199+20% (v/v) fetal calf serum (FCS) whereas VS-II contained 40% EG+18% Ficoll+0.5M sucrose in TCM-199+20% FCS. The percentage of oocytes found to be morphologically normal was greater (P<0.01) in VS-I group than in VS-II group. Rates of cleavage (30.6–42.2%) and blastocyst formation (2.9–8.9%) did not differ among groups, but were lower than in unfrozen control (55.7% and 25.4%, P<0.01). These results show that a combination of EG and DMSO or EG, Ficoll and sucrose can be used to cryopreserve yak oocytes in French straws.

The effect of minimal concentration of ethylene glycol (EG) combined with polyvinylpyrrolidone (PVP) on mouse oocyte survival and subsequent embryonic development following vitrification

Vitrification techniques employ a relatively high concentration of cryoprotectant in vitrification solutions. Exposure of oocytes to high concentrations of cryoprotectant is known to damage the oocytes via both cytotoxic and osmotic effects. Therefore, the key to successful vitrification of oocytes is to strike a balance between the usage of minimal concentration of cryoprotectant without compromising their cryoprotective actions. The minimal concentration of ethylene glycol (EG) on mouse oocyte survival and subsequent embryonic development was evaluated following vitrification-warming and parthenogenetic activation. Polyvinylpyrrolidone (PVP) combined with EG on mouse oocyte survival and subsequent embryonic development as well as morphology of the spindle and chromosome alignment were also evaluated. Vitrification system was adapted with JY Straw and the cooling rate was approximately 442-500°C/min. In contrast, the warming rate was approximately 2,210-2,652°C/min. Survival rate of oocytes increased significantly when 15% EG was combined with 2% PVP in vitrification solution (VS). The effect of combination of EG and PVP was not significant when the concentration of EG was 20% and higher. Although there were no significant differences in embryonic development, the percentage of abnormal spindle and chromosome alignment was significantly higher in the oocytes without 2% PVP in VS. Our data provide a proof of principle for oocyte vitrification that may not require a high concentration of cryoprotectant. There are synergic effects of EG combined with PVP for oocyte vitrification, which may provide important information to the field in developing less cytotoxic VS.

Melt Polycondensation and Analysis of Hydroxyl-Terminated Poly(L-lactic Acid) Using Small Molecular Diols as Terminating Agents

The synthesis of hydroxyl-terminated poly (L-lactic acid) (PLLA) by direct melt polycondensation of L-lactic acid and use of small molecular diols as terminating agents was investigated. L-lactic acid was dewatered to prepare the dehydrated prepolymer at first. Then PLLA was prepared through melt polycondensation in a melt with catalyst system including TSA, SnCl2and Sn (Oct)2. Finally, ethylene glycol, 1,4-butanediol, and 1,8-octanediol, respectively, were used in hydroxyl termination of the PLLA. The molecular weights and end-group structures of all products were characterized by gel permeation chromatography (GPC) and1H-NMR. DSC and Disc Polarimeter were used to characterize crystallinity and racemization, respectively. The results show that two kinds of hydroxyl-terminated PLLA polymers were obtained: complete hydroxyl-terminated PLLA prepared by using ethylene glycol or 1,4-butanediol, incomplete hydroxyl-terminated PLLA prepared by using 1,8-octanediol or combination of two kinds of diols. Combination of ethylene glycol or 1,4-butanediol with 1,8-octanediol can reach high hydroxyl termination of PLLA, and maintain the molecular weight by decrease of alcoholysis at the same time.1H-NMR was found as an effective estimating way for extents of reactions. The hydroxyl-terminating reactions of PLLA by the diols reduced the crystallinity and resulted in racemization to some extent.

Comparison of Ethylene Glycol and Propylene Glycol for the Vitrification of Immature Porcine Oocytes

Our aim was to optimize a cryoprotectant treatment for vitrification of immature porcine cumulus-oocyte complexes (COCs). Immature COCs were vitrified either in 35% ethylene glycol (EG), 35% propylene glycol (PG) or a combination of 17.5% EG and 17.5% PG. After warming, the COCs were in vitro matured (IVM), and surviving oocytes were in vitro fertilized (IVF) and cultured. The mean survival rate of vitrified oocytes in 35% PG (73.9%) was higher (P<0.05) than that in 35% EG (27.8%). Oocyte maturation rates did not differ among vitrified and non-vitrified control groups. Blastocyst formation in the vitrified EG group (10.8%) was higher (P<0.05) than that in the vitrified PG group (2.0%) but was lower than that in the control group (25.0%). Treatment of oocytes with 35% of each cryoprotectant without vitrification revealed a higher toxicity of PG on subsequent blastocyst development compared with EG. The combination of EG and PG resulted in 42.6% survival after vitrification. The maturation and fertilization rates of the surviving oocytes were similar in the vitrified, control and toxicity control (TC; treated with EG+PG combination without cooling) groups. Blastocyst development in the vitrified group was lower (P<0.05) than that in the control and TC groups, which in turn had similar development rates (10.7%, 18.1% and 23.3%, respectively). In conclusion, 35% PG enabled a higher oocyte survival rate after vitrification compared with 35% EG. However, PG was greatly toxic to oocytes. The combination of 17.5% EG and 17.5% PG yielded reasonable survival rates without toxic effects on embryo development.

CaO/Y2O3 pellets for reversible CO2 capture in sorption enhanced reforming process.

Pellets of a novel, high temperature CO2 sorbent, made of CaO-impregnated porous Y2O3, were prepared. Yttria pellets were synthesized from a mixture of yttria powder and a softener. A combination of ethylene glycol, used as a softener, and a casting method yielded mechanically stable pellets after calcination at 800ºC. Treatment of the Y2O3 support at higher temperatures further increases the pellets strength. The same effect of pellets strengthening was observed after yttria impregnation with CaO. Sorption capacity of the pellets with CaO content of 9 wt. %, measured at isothermal conditions of 740ºC, reaches 7.5 wt. % for shorter recarbonation time of 20 min and 8.5 wt. % for a longer time of 1 hour. In this respect, sorption properties of pelletized CaO/Y2O3 are similar to those of powdered material. A distinctive feature of the pelletized CaO/Y2O3 sorbent pretreated at high temperatures is the increase in capacity during the initial cycles.

Hybrid Titanium-Sapphire: Dye laser

On this work we present a Titanium:Sapphire laser with an intracavity dye amplifier. The system was pumped with 5 W of 532 nm laser radiation from an intracavity doubled Nd: YVO4 laser. The dye used was styryl 14 dye dissolved in a 15%:85% combination of ethylene glycol and propylene carbonate solvents inside a flow cell. The laser was tuned using a three-stage birefringent filter. We observe the behavior of the system in which the intracavity amplifier, depending on the pump power, behaved as an amplifier, hybrid laser or saturable absorber.

Morphological changes, DNA damage and developmental competence of in vitro matured, vitrified-thawed buffalo (Bubalus bubalis) oocytes: A comparative study of two cryoprotectants and two cryodevices

Present study was designed to investigate the impact of two cryodevices; french mini straw (FMS) and open pulled straw (OPS) using two different cryoprotectants; ethylene glycol (EG) and propylene glycol (PG) on morphological damage, recovery rate, DNA damage and developmental competence of in vitro matured vitrified-thawed buffalo oocytes. In vitro matured oocytes were divided into three groups: (a) no cryoprotectant (unfrozen, control), (b) vitrified in FMS and (c) in OPS using EG/PG. After thawing, recovered oocytes were subjected to morphological evaluation, cryoinjury at DNA level and their developmental competence. Results showed that recovery rate from both the groups (b and c) were almost same. Amongst the morphological damaged oocytes, zona pellucida crack, oocyte shrinkage and splitting were significantly (P<0.05) higher in FMS with PG as compared to FMS with EG (group b) while, OPS with EG was significantly (P<0.05) better as it maintained the architecture of oocytes and hardly any damage was found except some cytoplasmic shrinkage and change in shape. The number of oocytes displaying DNA damage was significantly (P<0.05) higher in FMS with PG. Cleavage and blastocysts production rate was significantly higher (P<0.05) for the oocytes recovered from OPS as compared to FMS with PG or EG. OPS with EG gave best cleavage and blastocysts rate amongst all the groups. In conclusion, combination of EG with OPS gives the best result in terms of better recovery and survival rate, least morphological damages with good developmental competence of vitrified matured buffalo oocytes post-thawing.

Stallion semen cryopreservation using different cryoprotective agents on the skim milk trehalosa extender

Cryoprotective agents (CPAs), protect the sperm during cryopreservation. There are two general classes of CPAs, at first penetrating cryoprotectants, these pass through the sperm membrane and act both intracellular and extracellularly, and the second non-penetrating cryoprotectants, these act only extracellularly. The objective of this study was to evaluate the addition of different CPAs namely glycerol (G), combination of ethylene glycol with glycerol (EG) and dimethilformamide (DMF) using skim milk trehalosa extender. The semen collected from 3 stallions using artificial vagina twice aweeks. The semen was evaluated, centrifugated and diluted in skim milk extender supplemented with 50 mM trehalose and three different CPAs with the concentration of sperm were 200x10 6 ml -1 . Extended semen was then packed into minitub 0.3 ml and equilibrated at 4 o C for 2 hours, freeze in the liquid nitrogen vapor for 10 minutes and stored in liquid nitrogen container -196 o C. After 24 hours, the semen was thawed at 37 o C for 30 second. Results of this experiment indicated that the percentage of motile and viable sperm in skim trehalosa extender using DMF and glycerol better than combination of ethylene glycol and glycerol. Key Words : Cryopreservation, Stallion Sperm, Cryoprotectant

P-199: The comparison of combination of EG and PROH to EG as cryoprotectant

This study was conducted to investigate whether combination of ethylene glycol (EG) and propanediol (PROH) has influence on the survival and spindle presence compared to EG only when mice oocytes were vitrified-thawed. Animal study. Oocytes were collected from superovulated BD F1 mice. The metaphase II (MII) oocytes (n=148) divided into two groups. After the meiotic spindle was evaluated by Polscope, group I was vitrified in DPBS with 20% hFF, 0.3 M sucrose, and 1.5 M EG and group II was vitrified in DPBS with 20% hFF, 0.3 M sucrose, 1.5 M EG, and 1.5 M PROH. The vitrified oocytes were thawed in DPBS with 20% hFF and 0.5 M sucrose. The meiotic spindle of the survival oocytes in each group was evaluated by Polscope 2 h after thawing. The recovery rate of thawed oocytes was 90.9% (80/88) in group I and 86.7% (52/60) in group II. The survival rate of recovered oocytes was 77.5% (62/80) in group I and 73.1% (38/52) in group II. The rate of meiotic spindle captured in group II was 100% (22/22), which was higher than that in group I (68.6%, 28/44). This result suggested that combination of ethylene glycol (EG) and propanediol (PROH) could cryopreserve more stable spindle than using EG only.

Amorphous Copolyesters Containing Monomers Derived from Bisphenols

Amorphous copolyesters based on combinations of ethylene glycol (EG) and 1,4-cyclohexanedimethanol (CHDM) as diols with terephthalic acid have excellent properties, such as toughness, clarity, and chemical resistance and are well known commercial polymers. Several different hydoxyethoxylated bisphenols were copolymerized into the backbone of these amorphous copolyesters in order to raise the glass transition temperature (Tg). The incorporation of bis[4-(2-hydroxyethoxy)phenyl]sulfone (SEO-2) into the amorphous copolyester backbone led to significant Tg increase and improvement in resistance to lipids. Melt viscosity and other mechanical and physical properties were unchanged by the addition of SEO-2.

Developmental capacity of vitrified immature porcine oocytes following ICSI: effects of cytochalasin B and cryoprotectants

In the present study, effects of concentration and pretreatment time of cytochalasin B (CB), and of two types of cryoprotectant solutions on the nuclear maturation of vitrified–warmed porcine oocytes were examined. Also, the developmental capacity of vitrified immature porcine oocytes following intracytoplasmic sperm injection (ICSI) was investigated. The nuclear maturation rate (46.8%) of the vitrified–warmed oocytes treated with 7.5 μg/mL CB for 30 min was significantly higher ( P < 0.05) than those (13.9–39.2%) of the vitrified–warmed oocytes treated with 0, 2.5, or 5.0 μg/mL CB for 10 or 30 min. Additionally, the nuclear maturation rate of oocytes treated with CB and vitrified in ethylene glycol (EG) (37.1%) was significantly higher ( P < 0.05) than that of EG + dimethyl sulfoxide (Me 2SO) (23.9%). However, no significant differences were observed in the cleavage and blastocyst development rates among the control (45.2 and 20.0%, respectively), the EG group (37.8 and 13.5%, respectively) and the EG + Me 2SO group (39.3 and 14.3%, respectively). These results demonstrated that: (1) pretreatment with 7.5 μg/mL CB was beneficial for the vitrification of immature porcine oocytes; (2) the combination of EG and Me 2SO as a cryoprotectant was not advantageous for in vitro maturation (IVM) of vitrified immature porcine oocytes; and (3) vitrified–warmed porcine oocytes matured after IVM, developed to the blastocyst stage without distinct differences compared to fresh oocytes following ICSI.

Incorporation of penetrating cryoprotectants in diluents for pellet-freezing ram spermatozoa

Glycerol may be toxic to frozen-thawed ram spermatozoa and reduce their fertilizing capacity. This study examined the cryoprotective effects of dimethyl sulphoxide (DMSO), ethylene glycol, glycerol and propanediol alone and in combinations with each other in Triscitrate-glucose diluents on the post-thaw motility and acrosome integrity of pellet-frozen ram spermatozoa. The 4 cryoprotectants were examined in diluents at 5 concentrations (0, 1.5, 3.0, 6.0, 12.0% v/v). Post-thaw motility of spermatozoa was higher in diluents containing ethylene glycol (1.5 to 6.0% v/v), glycerol (at all levels tested) and propanediol (1.5 and 3.0% v/v) than in diluents without cryoprotectant (P<0.001), but there was no effect of DMSO on post-thaw motility. Motility of spermatozoa was higher in diluents containing ethylene glycol or glycerol than DMSO or propanediol (P<0.001). In diluents containing the 4 cryoprotectants at 3 concentrations (1.5, 3.0, 6.0% v/v), better recovery of spermatozoa was found with the addition of 18.0 than 4.5% v/v egg yolk. Combinations of ethylene glycol and/or propanediol (0 to 6.0% v/v) with glycerol (0 to 6.0% v/v) in diluents were also examined. In the presence of glycerol at all levels tested, increasing levels of ethylene glycol and/or propanediol decreased motility and acrosome integrity of spermatozoa (P<0.001). We conclude that the compounds examined exert a cryoprotective effect on pellet-frozen ram spermatozoa, except for DMSO which had no effect. In this study, glycerol remained the single most effective cryoprotectant, and there was no enhancement of this cryoprotection by addition of the other compounds.

High-performance liquid chromatography of proteins on deformed nonporous agarose beads: immuno-affinity chromatography, exemplified with human growth hormone as ligand and a combination of ethylene glycol and salt for desorption of the antibodies

An affinity chromatography column packed with nonporous agarose beads derivatized with human growth hormone via carbonyldiimidazol was used for the purification of antibodies against human growth hormone from antiserum. Desorption with 1 M sodium chloride in 60% ethylene glycol at pH 9.8 gave 100% total recovery of the antibodies, as measured by radioimmunoassay. The adsorption/desorption process is discussed in terms of hydrophobic and electrostatic interaction (these interactions may be involved in the bond between antibody and antigen in a cooperative fashion). The binding capacity of the column was estimated at about 50 micrograms of antibodies per gram sedimented agarose beads.

Purification of eukaryotic RNA polymerase II by immunoaffinity chromatography. Elution of active enzyme with protein stabilizing agents from a polyol-responsive monoclonal antibody.

Active eukaryotic RNA polymerase II (RNAP II) was purified by immunoaffinity chromatography, using a monoclonal antibody (mAb) that reacts with the highly conserved heptapeptide repeat of the largest subunit. This mAb (designated SWG16) was conjugated to CNBr-activated Sepharose and used to purify RNAP II from wheat germ and calf thymus. The subunit composition of the immunoaffinity-purified enzyme was essentially the same as RNAP II purified by conventional chromatography except that it contained only the form with the unproteolyzed largest subunit. Active enzyme could be eluted from the SWG16-Sepharose, at pH 7.9, with combinations of low molecular weight polyols and nonchaotropic salts. The superior eluting procedure used combinations of ethylene glycol (30-40%) and ammonium sulfate (0.5-0.75 M). Active enzyme also could be eluted with a synthetic peptide containing four repeats of the heptapeptide; however, the peptide was not as effective as the polyol and salt combinations for eluting the enzyme. This mAb should be useful for purifying RNAP II from many eukaryotic species. Because the elution of enzyme from the immunoadsorbent seems to be dependent upon the presence of a polyol, this antibody is referred to as a "polyol-responsive mAb." A procedure that helps to identify a polyol-responsive mAb and to optimize the eluting conditions is described. Polyol-responsive mAbs might have broad applicability to the purification of many labile enzymes by immunoaffinity chromatography.

Scrapie agent contains a hydrophobic protein.

The scrapie agent causes a degenerative nervous system disorder of sheep and goats. Considerable evidence indicates that the scrapie agent contains a protein that is necessary for infectivity [Prusiner, S. B., Groth, D. F., Cochran, S. P., Masiarz, F. R., McKinley, M. P. & Martinez, H. M. (1980) Biochemistry 19, 4883-4891], but direct demonstration of a protein moiety has been hampered by lack of sufficiently purified preparations. Employing preparations of the scrapie agent enriched 100- to 1000-fold with respect to protein, we found that digestion by proteinase K destroyed more than 99.9% of the infectivity. Diethylpyrocarbonate, which chemically modifies amino acid residues in proteins with high efficiency, also inactivated the scrapie agent in these purified preparations. Reductions of infectivity by proteinase K and diethylpyrocarbonate were not observed with less purified preparations. The agent bound to phenyl-Sepharose could not be eluted with 8.5 M ethylene glycol; however, a combination of ethylene glycol and detergents did release the agent. These observations provide good evidence for a protein and for hydrophobic domains within the scrapie agent. Whether the protein required for infectivity is the same protein responsible for the hydrophobic properties of the scrapie agent remains to be established.