• Reproductive and Developmental Biology
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• v.34 no.1
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• pp.41-46
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• 2010

The objective of this study was to evaluated the efficiency of sperm cryosurvival using each extenders Triladyl and LEY containing egg yolk to the cryopreservation of separated sperm by percoll in miniature pig. The ejaculated semen from miniature pig was separated by 65% percoll and un-separated sperm as a control before freezing. The freezing of diluted semen added with Triladyl containing egg yolk and LEY solution (solution I: 11% Lactose or Triladyl + egg yolk; solution II: solution I + glycerol + OEP). Analysis of sperm ability was estimated by viability, capacitation acrosome reaction using chlortetracycline (CIC) the morphologic abnormality and hypoosmotic swelling test(HOST). The groups were designed that as separated sperm by Percoll with Triladyl(ST) or LEY(SL) for cryopreservation. And unseparated sperm with Triladyl(UT) or LEY(UL). As a results, the viability was higher significantly(p<0.05) in ST, SL, UT than UL extender. The morphologic abnormality was measured significantly (p<0.05) lower in ST than other extenders. The AR-patterned in CTC analysis was measured significantly(p<0.05) lower in SL and UL than other extenders. In conclusion, using Triladyl extender resulted in viability and morphology of separated sperm by percoll that were effective than using LEY extender, but it resulted in capacitation acrosome reaction was lower than using LEY extender.

• Asian-Australasian Journal of Animal Sciences
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• v.21 no.2
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• pp.181-189
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• 2008

The aim of this study was to evaluate the effects of $Ca^{2+}$, $HCO_3{^-}$ and BSA on the in vitro capacitation-associated protein tyrosine phosphorylation, hyperactivation and acrosome reaction in guinea pig sperm. Caudal epididymal sperm were incubated in four different groups: modified TALP (Tyrode's albumin lactate pyruvate) or TALP without one of the medium constituents ($Ca^{2+}$, $HCO_3{^-}$ and BSA). After incubation for the required time (0 h, 0.5 h, 1 h, 3 h, 5 h, and 7 h), sperm were removed for further experiment. The capacitation effect was assessed by CTC (Chlortetracycline) staining. Western blotting and indirect immunofluorescence were used to analyze the level and localization of tyrosine phosphorylation. The results showed that guinea pig sperm underwent a time-dependent increase in protein tyrosine phosphorylation during the in vitro capacitation and the percentage of protein tyrosine phosphorylated sperm increased from 36% to 92% from the beginning of incubation to 7 h incubation. Also, there was a shift in the site of phosphotyrosine-specific fluorescence from the head of sperm to both the head and the flagellum. Moreover, an absence of $Ca^{2+}$ or $HCO_3{^-}$ inhibited in vitro hyperactivation and acrosome reaction and decreased the phosphorylation of the proteins throughout the period of in vitro capacitation. However, an absence of BSA could not influence these processes if substituted by polyvinyl alcohol (PVA) in the medium.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.38 no.6
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• pp.372-378
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• 2005

An analysis method of tetracyclines far fish and shellfish products containing large amount of low molecular materials and pigments was established. The recovery of the established analysis method for four tetracycline samples was $72-100\%$ and higher than other methods reported. Especially, proposed sample treatment protocol was shown to be effective for the removal of low molecular materials and pigments that tend to interfere with accurate analysis. The detection limit of oxytetracycline (OTC) and tetracycline (TC) from the sample was 0.02 ppm, and the detection limit of chlortetracycline (CTC) and doxycycline (DC) from the sample was 0.1ppm, To examine the efficiency of the established method and identify tetracycline usage in fish farms, tetracycline group antibiotics in the flounder being cultured was monitored. The improved method can be used for fish and shellfish products effectively and all surveyed fish farms have used tetracycline all the year round. The proposed method was adopted as official method for fishery products by Korean Food and Drug Administration in 2003 and it is being used by regulatory authority as National Fishery Products Quality Inspection Service.

• Clinical and Experimental Reproductive Medicine
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• v.27 no.2
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• pp.201-209
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• 2000

Objective: This study has been carried out to evaluation the effect of fertilization promoting peptide (FPP) on the kinematic parameters, capacitation and acrosome reaction of the frozen-thawed human spermatozoa. Methods: After FPP treatment, we examined kinematic parameters, capacitation and acrosome reaction, using the methods of computer-aided sperm analysis (CASA) and chlortetracycline (CTC) fluorescence analysis. Results: We have obtained the evidence that FPP can promote the capacitation and inhibit the spontaneous acrosome reaction of frozen-thawed human spermatozoa in vitro. FPP ($25{\sim}100$ nM) induced a significant increase in the proportion of B-pattem capacitated spermatozoa, and a significant decrease in the proportion of F-pattem uncapacitated ones without significant stimulation of acrosomal exocytosis. In the kinematic parameters treatment, FPP treated groups maint3ined higher LIN, BCF and STR than those of control. The VAP, VSL, VCL and ALH were not different. Therefore it is suggested that FPP in human seminal plasma may play a positive role in promoting human sperm function.

• Reproductive and Developmental Biology
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• v.37 no.4
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• pp.213-217
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• 2013

The objective of this study was to investigate the effect of bacterial contamination on elapsed time after preservation on boar semen. Known numbers of Escherichia coli (E. coli) were inoculated to freshly ejaculated semen and sperm parameters such as viability, motility, agglutination, acrosome integrity and hypo-osmotic swelling test were performed during 7 days of liquid preservation. Semen samples were prepared using antibiotic free BTS extender and 4 different levels of E. coli were treated to semen with following concentrations; 3,000, 5,000, 7,000, 10,000 CFU/ml of sperms. Semen samples were preserved at $17^{\circ}C$ for 7 days in semen storage until analyzed. Aliquots were subjected to measure the sperm viability, motility and agglutination using computer assisted sperm analysis (CASA) system, acrosome integrity was performed using chlortetracycline (CTC) staining method and hypo-osmotic swelling test was performed using hypotonic solution from day 1 (day of semen collection) to 7. Detrimental effects on sperm motility and viability were observed 3 days after preservation at the level of 5,000 CFU/ml (p<0.05). Percentage of sperm abnormality was higher (p<0.05) in over 5,000 CFU/ml groups. Sperm agglutination rate was also significantly higher (p<0.05) in groups of 5,000 and 7,000 CFU/ml. The rate of acrosome reacted sperm was higher as preservation time goes in all the samples but the pattern was clearly higher among E. coli contaminated groups (p<0.05). The sperm membrane integrity in terms of hypo-osmotic test, E. coli affects little compared to other sperm parameters. The deleterious effects observed due to the bacterial contamination in semen suggest that importance of hygiene protocol to minimize the bacterial contamination during semen collection and processing.

• Journal of Applied Biological Chemistry
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• v.66
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• pp.258-265
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• 2023

After application of veterinary antibiotics, they may be partially metabolized before they are excreted by feces or urine either as unaltered form or as metabolites. Residual antibiotics enter the adjacent agricultural environments by spraying manure-based compost and liquid fertilizer on farmlands and lead to secondary pollution. The objective of this study was to compare the residual concentrations of 6 veterinary antibiotics by different production stages of liquid fertilizer from livestock manure recycling facilities. The relationship between concentration change of the residual antibiotics and the amount of liquid fertilizer component was also assessed. Pre-treatment showed the recovery of 63.4-106.7% at ppb level and the limit of quantification of 0.009-0.037 ㎍/L.As the result of analyzing the relationship between the residual concentrations of antibiotics and the amount of N component in liquid fertilizer by different production stages, the residual concentrations of antibiotics and N tended to decrease as the stabilization period elapsed during the liquid fertilization process. Average concentrations of sulfamethazine in raw materials, middle and final products of liquid fertilizer were 40.85, 26.17, 3.54 ㎍/L, respectively. Those of chlortetracycline decreased from 2.32 to 1.25 ㎍/L. The other 4 antibiotics also showed a decreasing trend by different production stages of liquid fertilizer. The amount of liquid fertilizer component N decreased from 0.21 to 0.096% by production stages of liquid fertilizer. It is considered that the correlation between residual antibiotic concentrations and N content can be applied as basic data for setting antibiotic reduction indicators.

• Journal of Korean Society of Environmental Engineers
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• v.31 no.10
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• pp.845-854
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• 2009

There has been increased concern regarding the release of antibiotics to different environmental compartments due to the possibility of the development of antibiotic resistant bacteria. However, limited information is available regarding the occurrence, fate, and transport of antibiotics in Korea in both the aqueous phase and in solid phases such as sediment and soil. Therefore, this study was conducted to monitor the concentration of released antibiotics in surface water, sediment, and soil adjacent to a cattle manure composting facility in Korea. Specifically, the following six antibiotics were monitored: tetracycline (TC), chlortetracycline (CTC), oxytetracycline (OTC), sulfamethazine (SMT), sulfamethoxazole (SMX), and sulfathiazole (STZ). To extract and quantify the antibiotics from different environmental compartments, solid phase extraction (SPE) and high performance liquid chromatography mass spectrometry (HPLC/MS) techniques were adopted. The concentration of the six antibiotics ranged from below the detection limit (BDL) to 0.71 ${\mu}g$/L in surface water, from BDL to 27.61 ${\mu}g$/L in sediment, and from 0.12 to 157.33 ${\mu}g$/L in soil. In addition, higher concentrations of antibiotics were observed in surface water and sediment at locations closer to the composting facility indicating that composting is the source of the antibiotics found in the environment. Furthermore, higher concentrations of antibiotics were observed in the solid phase (sediment and soil) than the aqueous phase. These findings indicate that the possibility of antibiotic resistant bacteria is increased because such bacteria are more stable in the solid phase. Overall, longterm monitoring of the aqueous phase and solid phase is necessary to gain a better understanding of the impact of antibiotics from source on the environment in Korea.

• Reproductive and Developmental Biology
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• v.36 no.3
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• pp.147-154
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• 2012

This study was designed to determine whether low-density lipoproteins (LDL) from egg yolk and taurine, hypotaurine and trehalose as antioxidant in extender improve the freezability and fertility of Korean Jeju Black Bull semen. The semen was cryopreserved with tris egg yolk extenders containing 7% glycerol and treated 4% LDL, 20 mM taurine, hypotaurine and trehalose. Frozen-thawed sperm were evaluated motility, viability, membrane, and acrosome integrity and sperm penetration ability. The results were compared to semen cryopreserved in tris egg yolk extender only as control. Frozen-thawed semen evaluation cleary indicated that the addition of LDL and LDL-antioxidants (taurine, hypotaurine and trehalose) combination were significantly improved (p<0.05) the viability (%; with staining test using eosin-Y) compared to control spermatozoa. Also, in membrane integrity (%; with supravital hypo-osmotic swelling test), not only LDL-antioxiants combination but also LDL were significantly increased (p<0.05) the swelled sperm using HOST compared to control. Sperm acrosome integrity state was classified by CTC (chlortetracycline) staining test. F pattern was significantly increased in LDL-antioxidant combination than control (p<0.05) and B pattern was not significantly differences among all treatments and control. However, AR pattern was significantly decreased in LDL-antioxidants combination than control (p<0.05). Pronucleus formation and sperm penetration index (SFI) were significantly increased in LDL and LDL-antioxidants combination than control (p<0.05). Especially, LDL-taurine significantly improved pronucleus fomation and SFI than LDL (p<0.05). It was concluded that LDL and LDL-antioxidants in extender improved the freezability and fertility of Korean Jeju Black bull spermatozoa.

• Korean Journal of Animal Reproduction
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• v.25 no.4
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• pp.317-325
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• 2001

This study investigated the effect of ferrous sulfate (Fe$^{2+}$) and/or ascorbic acid (Asc) on fertilizing ability in vitro of frozen-thawed boar spermatozoa. Using chlortetracycline (CTC) fluorescence, the spermatozoa was treated in preincubation medium with control, Fe$^{2+}$(1 mM), Asc (0.5 mM) and Fe$^{2+}$Asc to assessed for acrosome reaction, and the oocyte penetration test to determine whether the Fe$^{2+}$ and/or Asc can promote the penetration ability in vitro. When frozen-thawed spermatozoa was washed with preincubation medium, there were significantly (P ＜ 0.05) more acrosome-reacted in medium with Fe$^{2+}$Asc (38%) than control (27%). The penetration rates were also significantly (P ＜ 0.05) higher in medium with Fe$^{2+}$Asc (76%) than control (55%). Next, the lipid peroxidation of sperm was evaluated on the basis of malondialdehyde production following same treatments. The addition of Fe$^{2+}$Asc to sperm suspension increases the formation of malondialdehyde. However, there were not significantly different under the all conditions. The sperm suspension were also treated with control, Fe$^{2+}$, Asc and Fe$^{2+}$/Asc and assayed for sulfhydry1(-SH) group content. In the Fe$^{2+}$/Asc group, sperm-SH group were higher than another groups. In spermatozoa treated with Fe$^{2+}$ and/or Asc, however, no changes in sperm -SH-groups were detected when compared to controls. In another experiment, the activity of sperm binding to zona pellucida was evaluated through binding to salt-stored porcine oocytes. In control and Asc treatment groups, sperm binding to zona pellucida were significantly (P ＜ 0.05) higher than in medium with Fe$^{2+}$. On the other hand, there is not a significant increase in binding to zona pellucida with spermatozoa treated by Fe$^{2+}$/Asc. In summary, the present study suggests that Fe$^{2+}$/Asc causes an enhancement in fertilizing ability that is associated with penetration rate increased without change of spermatozoa binding capacity to homologous zona pellucida.o homologous zona pellucida.