• Asian-Australasian Journal of Animal Sciences
• /
• v.13 no.1
• /
• pp.10-18
• /
• 2000

This paper describes the magnetic orientation of the intact and demembranated bull sperm treated by DTT or heparin in a 5,400 G static field. Semen samples collected from four bulls (Japanese Black) were mixed to the same sperm density. One percentage triton X-100 was used to extract the plasma membrane. The intact and demembranated sperm suspensions were treated with 20, 200, 2,000 mM DTT, 100, 1,000 or 10,000 units heparin solutions at $4{^{\circ}C}$ for 6 days. The decondensation of the sperm nuclei treated by DTT or heparin was examined by measuring the sperm head area at 1, 3, and 6 days. After measuring the area, each sperm sample was exposed to a 5,400 G static magnetic field generated by Nd-Fe-B permanent magnets for 24 hours at room temperature. Results showed that the decondensation of bull sperm nuclei was not induced by the heparin treatment, however, incomplete decondensation was induced by the DTT treatment. During the magnetic orientation, bull sperms treated by DTT or heparin had low percentages of long axis perpendicular to the magnetic lines of force. However, different aspects were obtained for long axis perpendicular orientations following treatment of DTT or heparin. Through the DTT treatment, the decline of long axis perpendicularly oriented percentages was due to the increase of long axis parallel orientation with the head of the flat plane perpendicular to the magnetic lines of force, whereas, using the heparin treatment, the decline of long axis perpendicular orientation was due to the increment of long axis parallel orientation with the head of the flat plane parallel to the magnetic lines of force. Also, percentages of the head of the flat plane perpendicular were decreased by the heparin treatment. These findings suggest that maintaining the structure of protamine in the chromatin is necessary for the sperm head to orient with its flat plane perpendicular, and maintaining the disulfide bond in the chromatin is necessary for the long axis of sperm to orient perpendicularly.

• Microbiology and Biotechnology Letters
• /
• v.17 no.1
• /
• pp.35-45
• /
• 1989

Streptococcus thermophilus 510 was investigated as n potential source of $\beta$-galactosidase. Optimum cultural conditions for maximum enzyme production were 0.5% loctose as carbon source, initial pH 7.0, 37 $^{\circ}C$, and 18 hours of cultivation. The enzyme was purified to homogeneity by ammonium sulfate fractionation, protamine sulfate precipitation, Sephadex G-200 gel filtration, and DEAE-Sephadex A-50 ion exchange chromntography. The purified enzyme exhibited an optimum pH at 1.0, and an optimum temperature of 5$0^{\circ}C$. Metal ions such as Mn$^{2+}$ and $K^+$, dithiothreitol, and 2-mercaptoethanol stimulated $\beta$-galactosidase activity. Ethylenediamine tetraacetic add, 8-hydroxyquinoline, Hg$^2+$, Zn$^{2+}$, Co$^{2+}$, $Ca^{2+}$, and galactose were inhibitory. The $K_m$ and V$_{max}$ for o-nitrophenyl $\beta$-D-galactopyranoside were 1.25mM and 88.50$\mu$moles/min.mg protein, respectively. The molecular weight was estimated to be 520,000, and the amino acid composition indicated relatively high contents of glutamic acid, aspartic acid, leucine, and valine.

• Journal of Plant Biotechnology
• /
• v.47 no.3
• /
• pp.209-217
• /
• 2020

The presence of high amounts of seed storage proteins (SSPs) improves the overall quality of soybean seeds. However, these SSPs pose a major limitation due to their high abundance in soybean seeds. Although various technical advancements including mass-spectrometry and bioinformatics resources were reported, only limited information has been derived to date on soybean seeds at proteome level. Here, we applied a tandem mass tags (TMT)-based quantitative proteomic analysis to identify the significantly modulated proteins in the seeds of two soybean cultivars showing varying protein contents. This approach led to the identification of 5,678 proteins of which 13 and 1,133 proteins showed significant changes in Daewon (low-protein content cultivar) and Saedanbaek (high-protein content cultivar) respectively. Functional annotation revealed that proteins with increased abundance in Saedanbaek were mainly associated with the amino acid and protein metabolism involved in protein synthesis, folding, targeting, and degradation. Taken together, the results presented here provide a pipeline for soybean seed proteome analysis and contribute a better understanding of proteomic changes that may lead to alteration in the protein contents in soybean seeds.

• Clinical and Experimental Reproductive Medicine
• /
• v.45 no.1
• /
• pp.17-24
• /
• 2018

Objective: To investigate sperm chromatin/DNA integrity, global DNA methylation, and DNMT mRNA transcription in men with oligoasthenoteratozoospermia (OAT) compared with normozoospermic men. Methods: Semen samples from 32 OAT patients who comprised the case group and 32 normozoospermic men who comprised the control group were isolated and purified using a standard gradient isolation procedure according to World Health Organization criteria. DNMT1, DNMT3A, and DNMT3B transcripts were then compared between groups using real-time quantitative reverse-transcription polymerase chain reaction. Global DNA methylation in sperm was determined by an enzyme-linked immunosorbent assay. Protamine deficiency and the proportion of apoptotic spermatozoa were evaluated using chromomycin A3 (CMA3), aniline blue (AB), and toluidine blue (TB) staining, as well as the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. The p-values < 0.05 were considered to indicate statistical significance. Results: Significantly higher proportions of AB+, TB+, CMA3+, and TUNEL+ spermatozoa, as well as DNMT3A and DNMT3B transcription, were found in the OAT group. Positive correlations were detected between sperm parameters, DNA/chromatin damage, and DNMT3A and DNMT3B transcripts. Global DNA methylation was significantly higher in the OAT patients and had a significant correlation with abnormal results of all sperm chromatin integrity tests, but was not associated with DNMT1, DNMT3A, or DNMT3B expression. Conclusion: Oligoasthenoteratozoospermic men showed abnormal sperm parameters, abnormal chromatin/DNA integrity, and a higher global DNA methylation rate, as well as overexpression of DNMT mRNA.

• Journal of Veterinary Clinics
• /
• v.36 no.1
• /
• pp.74-77
• /
• 2019

A 10-year-old, 8.28 kg, intact female Cocker Spaniel was presented with continuous polyuria and polydipsia. The dog had proestrus bleeding 5 weeks earlier, and hyperglycemia, glucosuria, ketouria, and high level of serum fructosamine in laboratory findings. Based on hyperglycemia, glucosuria, ketouria, and ketosis, the patient was tentatively diagnosed as diabetes mellitus (DM) with ketouria. After diagnosis, 5 to 7 U/body porcine lente insulin was administered during 11 days from initial presentation as the remission of DM for the dog. But, blood glucose was still high level. Because there was no reaction to porcine lente insulin, it was replaced by 4-10 U/body neutral protamine Hagedorn (NPH) during 3 days. But, NPH also did not regulate blood glucose level. Because insulin therapy failed to regulate blood glucose level, the dog was considering insulin-resistant diabetes. The dog was tentatively diagnosed with progesterone-resistant DM on the basis of the history that had revealed proestrus bleeding 5 weeks earlier. Progesterone level was moderate high (43.7 ng/ml; reference range, 15.0-90.0 ng/ml). Ovariohysterectomy (OHE) was performed to remove the cause of the dog's diabetes. After OHE 11 days, blood glucose was gradually declined by insulin treatment. Consequently, blood glucose was well controlled in reference range without insulin treatment after 2 months. This case is a report on progesterone-induced DM treated with OHE and insulin treatment during the diestrus.

• Clinical and Experimental Reproductive Medicine
• /
• v.48 no.1
• /
• pp.34-42
• /
• 2021

Objective: Studies of the effects of estrogens on the male reproductive system have emphasized the role of these hormones in male fertility. Sesame oil has many phytoestrogenic compounds and may improve male fertility. This study investigated the effects of sesame oil and different concentrations of estrogen on sperm parameters and DNA integrity in male mice. Methods: Twenty old NMRI (The Naval Medical Research Institute) male mice (40 weeks; weight, 30-35 g) were treated with sesame oil or different concentrations of estrogen (estradiol, 1 and 10 μL/kg/day) or received no treatment (controls). After 35 days, sperm parameters and DNA integrity were assessed and analyzed. Results: Sperm count, progressive motility, and morphology were decreased in the group that received 10 μL/kg of estradiol. A remarkably lower percentage of DNA fragmentation and protamine deficiency were detected in the group that received 1 μL/kg of estradiol. In the groups that received sesame oil and 1 μL/kg of estradiol, the numbers of spermatogonia and Leydig cells were higher than in controls. The combination of sesame oil and 1 μL/kg of estradiol led to improved sperm parameters and chromatin and testicular structure. Conclusion: Based on this study, consumption of sesame oil and a low concentration of estradiol may improve testicular function in older mice.

• Clinical and Experimental Reproductive Medicine
• /
• v.51 no.3
• /
• pp.213-224
• /
• 2024

Objective: Some age-related testicular changes, such as Sertoli cell vacuolization and blood-testis barrier breakdown, reduce total sperm production and male fertility. Therefore, this study investigated the effect of vitamin E on restoring testicular function in aged mice. Sperm cryo-resistance was also assessed. Methods: Twenty-eight 48-week-old male Naval Medical Research Institute mice were divided into four groups for a daily gavage of vitamin E: the control group received distilled water, while the three treatment groups were administered 100, 200, and 400 mg/kg, respectively, for 4 weeks. Subsequently, semen analyses, DNA fragmentation index (DFI), and protamine deficiency tests were conducted. Testicular histology, tissue antioxidant enzyme activity, and gene expression levels were also assessed. Results: The two higher dosages of vitamin E were associated with a higher sperm count, greater progressive motility, and improved sperm morphology (p<0.05). These benefits were also evident after sperm freezing (p<0.05). Although chromatin abnormalities increased following vitrification, the treatment groups showed better outcomes (p<0.05). The tubular diameter, epithelium height, and luminal diameters remained unchanged with age. The tissue antioxidant capacity was greater in the groups receiving the high doses of vitamin E. Additionally, significant increases in inhibitor of DNA binding protein-4 (Id4) and GDNF family receptor alpha-1 (Gfra1) expression were observed in the higher vitamin E dosage groups, and promyelocytic leukemia zinc finger protein (Plzf) expression was notably present in the 400 mg/kg treatment group compared to the control group (p<0.05). Conclusion: Antioxidant supplementation might enhance reproductive outcomes in aging males. The observed effects included improved sperm cryo-resistance, which is advantageous for future applications such as sperm freezing or fertility preservation.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.14 no.4
• /
• pp.345-352
• /
• 1985

To characterize the protein from arrowroot leaf, proteins were extracted and separated from arrowroot leaf, then its amino acid composition and functional properties were studied. Protein in arrowroot leaf was consisted of 18.5% albumin, 33.5% globulin, 34.0% glutelin, 6.2% protamine and 7.8% insoluble residues. The rates of precipitation of proteins which extracted with water, 1M NaCl, and 0.015N NaOH as a solvent were 84.7% (at pH 3.0), 76.4% (at pH 2.5) and 86.4% (at pH 4.0), respectively. The extracted proteins were separated up to about 90% by organic solvents such as ethanol and acetone at 80% concentration, Composition of arrowroot leaf protein concentrates were: $1{\sim}2%$ moisture, $59{\sim}67%$ protein, $4{\sim}8%$ ash and $5{\sim}6%$ (dialyzed concentrates) or $1{\sim}2%$ (acetone-treated ones) lipid. Main amino acids of the concentrates were aspartic acid, glutamic acid and glycine. Solubility profile of the concentrates according to pH was typical. The minimum solubility (below pH 5.0) of acetone extracted protein concentrates was lower than that of unextracted ones, whereas the reverse was true for pH value above this region. Bulk density, water and fat absorption of the concentrates were attributable to correlation to the treatment of acetone. And the bulk density of the concentrates was negatively correlative to both water and fat absorption. Emulsifying and foaming properties were not varied with the treatment of acetone.

• Applied Biological Chemistry
• /
• v.22 no.4
• /
• pp.191-197
• /
• 1979

${\beta}-Tyrosinase$ was purified and crystallized from cells of Escherichia intermedia A-21 grown in a medium supplemented with 0.2% L-tyrosine. Molecular weight of its subunit, Km value and absorption spectra were determined. Crystallization methods were also studied to eliminate any unnecessary procedures. The results obtained were as follows: 1. The purification procedure included ammonium sulfate fractionation, dialysis against potassium phosphate buffer, pH 6.0 and pH 7.0, and DEAE-Sephadex column chromatography. In the column chromatography, 11 mg of protein was applied per ml of DEAE-Sephadex for efficiency. 2. Steps of protamine sulfate treatment and Sephadex G-150 gel filtration could be eliminated for this enzyme from the known procedures. 3. The purified enzyme was dissolved in 0.01M potassium phosphate buffer containing 2-mercaptoethanol, with a concentration of 20mg/ml. Crystalline enzyme, which appears as hexagonal rods, was obtained by adding solid fine powdered ammonium sulfate to the enzyme solution. 4. Absorption maxima of the enzyme appeared at 340 and 430nm when associated with pyridoxal phosphate. 5. Km value of the enzyme for L-tyrosine was $2.31{\times}10^{-4}M$ and the molecular weight of its subunit was determined by SDS-polyacrylamide electrophoresis to be approximately 50,000.

• Journal of Chest Surgery
• /
• v.3 no.2
• /
• pp.73-90
• /
• 1970

Clinical perfusion data on 16 cases of cardiopulmonary bypass using Sigmamotor pump and RyggKyvsgaard Oxygenator which performed at Seoul National University Hospital during the period of Aug. 1968 to Aug. 1970 was analized. AIl cases were hemodiluted and the perfusion was carried out under the normothermic condition. The age of the patients ranged between 6 and 43 years. The b:dy weight varied between 18.3 and 54.0 kg and the body surface area between 0.78 and 1. 59$M^2$. The priming solution was consiste:I with fresh ACD blood. Hartmann solution and Mannitol. The average amount of priming was approximately 2242 ml. The average hemodilution rate was 17%. The flow rate ranged from 1.7L to 3.5L/Min/$M^2$ and averaged 2.4L/Min/$M^2$ or 78mI/Min/kg. The duration of perfusion varied from 22 to 110 min with average of 56.9 minutes. Some hemodynamic responses were observed. The arterial pressure dropped immediately after the initiation of partial perfusion and was more marked after the total perfusion foIlowed by gradual increase to the safety level. The central venous pressure reflected the reduced blood volume especially in the cases of prolonged perfusion which lasted over 60 min. In most of the cases, red blood cell count decreased and white blood ceIl count increased after the perfusion. Hemoglobin level was decreased, averaging of 12.5mg%, Hct 3.3% and platelets count of 18% postoperatively. Plasma hemoglobin increased mildly, from pre-perfusion average value of 4. 06mg% to postperfusion value of 22.5mg%. Serum potassium was 4.4mEq/L pre-operatively and was decreased to 3.7mEq/L postoperatively. Five cases showed definite hypopotassemia immediately after the operation. Sodium and chloride decreased mildly. These electrolyte changes are thought to be related with hemodilution. diuretics and reduced blood volume during and after the perfusion. Arterial blood pH value revealed minimal to moderate elevation from preperfusion average value of 7.376 to 7.461 during perfusion and then 7.365 after perfusion. The pC02 and hicarbonate showed minimal to moderately lowered values. The total CO2 was decreased. Buffer base decreased during perfusion (Av. 42.6mEq/L) and further decreased after the perfusion (Av. 40.8mEq/L). These arterial blood acid base changes suggested that the metabolic acidosis was accompanied by respiratory alkalosis during and immediately after the perfusion. Authors belived that the acidosis could more effectively be corrected with the more additional dose of bicarbonate than we used by this study. The chest tune drainage during the first 24 hours following operation was 1158 ml in average. One case (Case No. 15) showd definite bleeding tendency and it was believed that the cause might be due to the defect of heparin and protamine titration. The average urinary out put during 24 hours post-perfusion was 1291ml. One case (Case No. ]) showed definite post perfusion oliguria. As conclusion hemodilution using fresh ACD blood. Hartmann and Mannitol solution added with Bivon and high flow rate unler normothermia. was thought to amelioratc the severity of mctabolic acidosis during and after perfusion with relatively satisfactory effect on the diuresis and bleeding tendency.