• Development and Reproduction
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• v.4 no.2
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• pp.161-173
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• 2000

The aim of this study was to assess the effect of the frequency of the L$N_2$ infusion on the ultrastructure, metabolism and development of the embryo after freezing and thawing by computerized cell freezer. Two-cell embryos of ICR mouse were randomly allocated into fresh (control), high-frequency freezing (group 1) and low-frequency freezing (group 2). For fresh and frozen-thawed intact 2-cell embryos, total ceil number in the blastocyst was counted by fluorescent microscope after Hoechst 33258 staining. Relative amount of $H_2O$$_2$ was measured by DCHFDA. Intracellular location and membrane potential of the mitochondria were evaluated by staining with rhodamine 123 and JC-1. The structure of actin filament was also evaluated by confocal microscope. DNA fragmentation was assessed by TUNEL method after development into blastocyst. The survival rate of intact embryo was higher in group 1 than group 2 (50.7％ vs. 34.6％ respectively, p<0.05). The blastocyst developmental rate was significantly low in group 2 (86.7％, 76.7％ vs. 44.0％ for control, group 1 and group 2 respectively, p<0.05). Total cell number in the blastocyst was also significantly lower in group 2 than control (79.5$\pm$12.9, 71.6$\pm$8.0, and 62.5$\pm$4.7 for control, group 1 and group 2 respectively, p<0.05). The relative amount of $H_2O$$_2$ was higher in group 2 than other groups (15.3$\pm$3.0, 16.6$\pm$1.6 vs. 23.4$\pm$1.8, p<0.05). After JC-1 staining, relative intensity of mitochondria with high membrane potential was significantly lower in group 2 than control and group 1 (17.2$\pm$3.8, 17.4$\pm$1.3 vs. 13.2$\pm$2.0, p<0.05). In group 2, partial deletion and aggregation of the actin filament was found. DNA fragmentation rate was also hieher for group 2 versus other groups (30.8％, 36.0％ vs. 65.6％, p<0.05). The frequency of the L$N_2$ infusion is an important factor for the development of frozen-thawed mouse embryo. High-frequency infusion may prevent damages of cytoskeleton and mitochondria in the embryo probably by preventing the temperature fluctuation during dehydration phase. We speculate that the application of high-frequency infusion method in human embryo may be promising.

• Journal of Embryo Transfer
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• v.26 no.1
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• pp.65-70
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• 2011

This study was conducted to investigate the survival rate of AndroMed and Tris-egg yolk extender for cryopreservation of Korean Native Bull Semen (Chick Cow). Semen was collected from a Korean Native Bull Semen over 3 year's old. The semen was diluted 1:1 by AndroMed and Tris-egg yolk extender. The pellet was diluted to final sperm concentration of $5{\times}10^7$ cell/ml by doubling in every 10 minutes at $4^{\circ}C$ cold chamber. The semen was equilibrated for 1 hrs at cold chamber and packed to 0.5 ml straw. The semen straws were located above 5 cm of liquid nitrogen for 5 minutes. And then the frozen straw was plunged to LN2. The presented straws were examined the viability and motility after thawed at $37^{\circ}C$ water bath. The survival rates was significantly higher (p<0.05) in Tris-egg yolk extender than AndroMed extender ($89.7{\pm}19.8$ vs. $73.4{\pm}11.2$). However, motility was no significant differences ($78.4{\pm}18.7$ vs. $67.9{\pm}14.6$). Survival rate in time of equilibration between visual and CASA program had higher in 2 h ($86.33{\pm}9.4$ vs. $92.32{\pm}12.4$) than in 5 h ($78.20{\pm}7.8$ vs. $88.28{\pm}13.1$) 15 h ($65.24{\pm}6.6$ vs. $76.48{\pm}17.3$) 20 h ($56.26{\pm}4.6$ vs. $67.73{\pm}18.4$). The development rates to cleavage was higher in Tris-egg yolk extender than AndroMed extender (82.2% vs. 81.7%). Similarly, the development rates to blastocyst was significantly higher (p<0.05) in Tris-egg yolk extender than AndroMed extender (42.3% vs. 29.6%). In conclusion, the obvious impact of this study will be its practical application to improve viability and fertilizing ability of cryopreserved spermatozoa used for in vitro fertilization (IVF) and AI, Which in turn will be beneficial to animal genetic resources conservation.

• Journal of Food Hygiene and Safety
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• v.35 no.3
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• pp.243-251
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• 2020

In this study, we investigated pesticide residues in 207 agricultural products distributed by direct trade in the northern area of Gyeonggi Province. A total of 94 general agricultural products and 113 eco-friendly agricultural products collected from local grocers and cooperative stores were analyzed by multiresidue method for 263 pesticides using GC (gas chromatography)/ECD (electron capture detector), GC/NPD (nitrogen phosphorus detector), GC-MS/MS (tandem mass spectrometry), LC (liquid chromatography)/PDA (photodiode array detector), LC/FLD (fluorescence detector), LC-MS/MS. All samples showing pesticide residues were general agricultural products collected from local food stores. The pesticide residue levels of 14 samples (6.8%) were below the maximum residue limits (MRLs) and one of them (0.5%) exceeded the MRLs. Sixteen pesticides were detected from samples of the following produce items: spinach, young cabbage, perilla leaves, mallow, cucumber, chives and water dropwort. The safety of the detected pesticides was assessed by monitoring the daily intake estimate (EDI) and the daily intake allowance (ADI) based on the amount of pesticides detected. The ADI percentage range (the ratio of EDI to ADI) was 0.0134-61.6259% and there was no health risk connected with consuming agricultural products in which pesticide residues were detected.

• Korean Journal of Food Science and Technology
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• v.39 no.2
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• pp.114-121
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• 2007

The purpose of this study was to investigate 253 kinds of pesticide residues in 58 commercial dried agricultural products in Seoul. The determinations of the pesticide residues were performed using multiresidue methods and were carried out by a gas chromatography-nitrogen phosphorus detector (GC-NPD), an electron capture detector ($GC-{\mu}ECD$), a mass spectrometry detector (GC-MSD) and high performance liquid chromatography-ultraviolet detector (HPLC-UV), and a fluorescence detector (HPLC-FLD). The pesticide residue detection rate in the commercial dried agricultural products was 24.1% (14 of 58 samples). Twelve pesticide residues without maximum residue limits (MRLs) were detected. In the vegetable groups, the frequency of pesticide residues was found to be in the increasing order of dried fruiting vegetables > dried leafy vegetables > dried stalk and stem vegetables. The pesticides used on dried red pepper in the dried fruiting vegetables were varied (7 kinds) and numerous (4 of 8 samples). The pesticide types detected in the commercial dried agricultural products were in the order of pyrethroid > organochloride > organophosphorus and insecticide > fungicide > herbicide ${\cdot}$ nematicide. The primary pyrethroid pesticide detected was cypermethrin. According to the producing areas of products, large numbers of pesticide residues were found in the order of Korea, China, North Korea, USA, and Vietnam.

• Reproductive and Developmental Biology
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• v.28 no.3
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• pp.167-172
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• 2004

This study was performed to examine the correlations among dog sperm viabilities evaluated by flow cytometry, by microscopic evaluation (ME), by carbo-xifluorescein diacetate and propidium iodide (CFDA/PI) and by hypoosmotic swelling (HOS) test. Semen were collected from 5 dogs ranging in age from 2 to 4 years. Each ejaculate was divided into 3 aliquots and different proportions of freeze-killed cells were added to each aliquot (1:0, 1:1 and 1:3). In the other experiment, semen was extended with Sweden extender containing 5% glycerol and equex STM paste, and frozen using liquid nitrogen vapor. Fresh and frozen-thawed dog sperm viability were assessed by flow cytometry using PI staining method. The accuracy of flow cytometry was evaluated by comparing with other classic assessments, microscopic evaluation, epifluorescence microscopic analysis using CFDA/PI, and HOS test. High correlations of sperm viabilities were found among flow cytometry, epifluorescence evaluation, HOS test (p<0.01) in fresh semen. Especially, sperm viability assessed by HOS test was highly correlated with viability by flow cytometry in all the ratios of live and dead spermatozoa, 1:0, 1:1 and 1:3 (p<0.01). The viability evaluated by ME were significantly correlated with that by flow cytometry in ratios of 1:0 and 1:3 (p<0.05) however, there was no significance in ratio of 1:1. The viability evaluated by C/p were highly correlated with that by flow cytometry in ratio of 1:0 and 1:1 (p<0.01) and significantly correlated in ratio of 1:3 (p<0.05). In frozen-thawed spermatozoa, the viability determined by HOS test was considerably correlated with that by flow cytometry (p<0.01). There was significant correlation between the viabilities by ME and by flow cytometry (p<0.05). But the viability evaluated by CFDA/PI was not correlated with viability by flow cytometry. The result from this study validate the use of flow cytometry as a precise method for assessing the viability of fresh and frozen-thawed dog spermatozoa.

• Reproductive and Developmental Biology
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• v.35 no.3
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• pp.377-383
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• 2011

The objective of this study was to examine effect of ethylene glycol for semen cryopreservation in Korean Jeju Black Bull. The semen was cryopreserved with extenders containing cryoprotectants (7% glycerol and 3%, 5%, 7% ethylene glycol) and packed to 0.5 ml straws. The semen straws were located above 3 cm of liquid nitrogen for 5 min, 5 cm for 10 min and 8 cm for 10 min. And then frozen straw was plunged into $LN_2$. Post-thawed sperm motility, viability and membrane integrity were significantly higher in 5% ethylene glycol ($72.5{\pm}5.00%$, $54.88{\pm}0.66%$ and $46.00{\pm}2.40%$; p<0.05). Motility and viability were similar between 7% glycerol and 5% ethylene glycol. However, the membrane integrity was significantly higher in 5% ethylene glycol ($34.69{\pm}4.64%$ vs $46.00{\pm}2.40%$; p<0.05). The viability and membrane integrity were significantly higher in 5 cm for 10 min and 8 cm for 10 min than 3 cm for 5 min (viability: $55.81{\pm}2.94$, $55.19{\pm}3.34$ vs $47.94{\pm}3.48%$; p<0.05 and membrane integrity: $44.94{\pm}3.51$, $46.06{\pm}2.25$ vs $40.38{\pm}1.03%$; p<0.05). The percentage of capacitated sperm assessed by CTC staining, percentage of F pattern was higher in 7% glycerol, 5% and 7% ethylene glycerol, and AR pattern was significantly higher in 3% ethylene glycol. F pattern was significantly increased in 5 cm for 10 min and 8 cm for 10 min (p<0.05), but AR pattern was significantly increased in 3 cm for 5 min (p<0.05).

• Journal of Animal Science and Technology
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• v.48 no.6
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• pp.797-804
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• 2006

The present study was undertaken to investigate the post-thawed survivability of bovine embryo depending on different dose of ethylene glycol and sucrose. Ovaries were collected at local slaughterhouse and the cumulus-oocyte-complexes aspirated from ovaries were in vitro matured, fertilized and cultured at 39°C in an atmosphere of 5% CO2 incubator. For conventional slow-freezing, d 7 or 8 expanded blastocysts were collected. Embryos were equilibrated in 1.5 M and 1.8 M ethylene glycol(EG) with 0.1 M and 0.3 M sucrose in Dulbecco's phosphate-buffered saline(D-PBS) supplemented with 0.5% bovine serum albumin. Embryos were then loaded individually into 0.25ml-straw and placed directly into cooling chamber of programmable freezer precooled to 󰠏7°C, after 2 min, the straw was seeded, maintained at 󰠏7°C for 8 min, and then cooled to 󰠏35°C at 0.3°C/min, plunged and stored in liquid nitrogen for at least 3 days. For thawing, the straw containing embryos were warmed in air for 10 sec and exposed to 37°C water for 20 sec. Straws were then removed from 37°C water. Rates of blastocyst survive and hatching were evaluated at 24 to 72 h post-warming. No difference of the survivability was shown between 1.5 M and 1.8 M EG (71 and 70%, respectively). Addition of 0.1 M sucrose to 1.5 M and 1.8 M ethylene glycol in the freezing solution did not differ significantly embryo survival (74 and 77%, respectively), whereas survival rates was higher(89%) in freezing solution contained 0.3M sucrose to 1.8M EG compared with 0.3M sucrose to 1.5M EG group(71%). However, there was no difference in the overall total cell number between the two groups (122±1.8 vs 131±1.4, respectively). In conclusion, the results suggest that 0.3 M sucrose in 1.8 M EG may be optimal condition for freezing and thawing methods with in vitro produced embryos and may be applied to on-farm conditions for embryo transfer.

• Journal of Food Hygiene and Safety
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• v.30 no.1
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• pp.81-86
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• 2015

The purpose of our paper was to investigate the migration level of 4,4'-MDA(4,4'-methylenedianiline), 2,4-TDA(2,4-toluenediamine), aniline, acrylonitrile and methylmeth acrylate from plastic cookwares into food simulants and to evaluate the safety of each monomers. The test articles for monomers were PA (polyamide) items for 4,4'-MDA, 2,4-TDA and aniline, PU (polyurethane) items for 4,4'-MDA, ABS (acrylonitrile-butadiene- styrene) items for acrylonitrile, and acrylic resin items for methylmethacrylate. All the article samples of 321 intended for contact with foods were purchased in domestic market. 4,4'-MDA, 2,4-TDA and aniline were analyzed by LC-MS/MS (liquid chromatography -tandem mass spectrometer), acrylonitrile by GC-NPD (gas chromatography-nitrogen phos phorus detector) and methyl methacrylate by GC-FID (gas chromatography-flame ionization detector). The migration level of monomers were within the migration limits of Ministry of Food and Drug Safety (MFDS). As a result of safety evaluation, our results showed that the estimated daily intake (EDI, mg/kg bw/day)s were $2.39{\times}10^{-9}$ and $1.20{\times}10^{-9}$ for 4,4'-MDA and 2,4-TDA of PA, $4.32{\times}10^{-9}$ for acrylonitrile of ABS and $2.27{\times}10^{-7}$ for methylmethacrylate of acrylic resin. Reference Dose (RfD, mg/kg bw/day) of acrylonitrile and tolerable daily intake (TDI, mg/kg bw/day) of methacrylate were established respectively as 0.001 by EPA (US Environmental Protection Agency) and as 1.2 by WHO (World Health Organization). When comparing with RfD and TDI, the EDIs of acrylonitrile and methylmethacrylate accounted for $4.32{\times}10^{-4}%$ and $1.89{\times}10^{-5}%$ respectively.

• Asian-Australasian Journal of Animal Sciences
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• v.13 no.11
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• pp.1619-1637
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• 2000

A review was undertaken to obtain information on the range of beak-trimming methods available or under development. Beak-trimming of commercial layer replacement pullets is a common yet critical management tool that can affect the performance for the life of the flock. The most obvious advantage of beak-trimming is a reduction in cannibalism although the extent of the reduction in cannibalism depends on the strain, season, and type of housing, flock health and other factors. Beak-trimming also improves feed conversion by reducing food wastage. A further advantage of beak-trimming is a reduction in the chronic stress associated with dominance interactions in the flock. Beak-trimming of birds at 7-10 days is favoured by Industry but research over last 10 years has shown that beak-trimming at day-old causes the least stress on birds and efforts are needed to encourage Industry to adopt the practice of beak-trimming birds at day-old. Proper beak-trimming can result in greatly improved layer performance but improper beak-trimming can ruin an other wise good flock of hens. Re-trimming is practiced in most flocks, although there are some flocks that only need one trimming. Given the continuing welfare scrutiny of using a hot blade to cut the beak, attempts have been made to develop more welfare friendly methods of beak-trimming. Despite the developments in design of hot blade beak-trimmers the process has remained largely unchanged. That is, a red-hot blade cuts and cauterises the beak. The variables in the process are blade temperature, cauterisation time, operator ability, severity of trimming, age of trimming, strain of bird and beak length. This method of beak-trimming is still overwhelmingly favoured in Industry and there appears to be no other alternative procedures that are more effective. Sharp secateurs have been used trim the upper beak of both layers and turkeys. Bleeding from the upper mandible ceases shortly after the operation, and despite the regrowth of the beak a reduction of cannibalism has been reported. Very few differences have been noted between behaviour and production of the hot blade and cold blade cut chickens. This method has not been used on a large scale in Industry. There are anecdotal reports of cannibalism outbreaks in birds with regrown beaks. A robotic beak-trimming machine was developed in France, which permitted simultaneous, automated beak-trimming and vaccination of day-old chicks of up to 4,500 chickens per hour. Use of the machine was not successful because if the chicks were not loaded correctly they could drop off the line, receive excessive beak-trimming or very light trimming. Robotic beak-trimming was not effective if there was a variation in the weight or size of chickens. Capsaicin can cause degeneration of sensory nerves in mammals and decreases the rate of beak regrowth by its action on the sensory nerves. Capsaicin is a cheap, non-toxic substance that can be readily applied at the time of less severe beak-trimming. It suffers the disadvantage of causing an extreme burning sensation in operators who come in contact with the substance during its application to the bird. Methods of applying the substance to minimise the risk to operators of coming in contact with capsaicin need to be explored. A method was reported which cuts the beaks with a laser beam in day-old chickens. No details were provided on the type of laser used, or the severity of beak-trimming, but by 16 weeks the beaks of laser trimmed birds resembled the untrimmed beaks, but without the bill tip. Feather pecking and cannibalism during the laying period were highest among the laser trimmed hens. Currently laser machines are available that are transportable and research to investigate the effectiveness of beak-trimming using ablasive and coagulative lasers used in human medicine should be explored. Liquid nitrogen was used to declaw emu toes but was not effective. There was regrowth of the claws and the time and cost involved in the procedure limit the potential of using this process to beak-trim birds.

• Asian-Australasian Journal of Animal Sciences
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• v.25 no.5
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• pp.690-700
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• 2012

The experiment was conducted by in vitro fermentation and bacterial community analysis to investigate the reduction of odorous compounds in response to the use of feed additives (FA) during carbohydrate overload in growing pigs. Soluble starch at 1% (control) and various FA at 0.1% Ginseng meal (FA1); Persimmon leaf (FA2); Gingko nut (FA3) and Oregano lippia (FA4) were added to fecal slurry and incubated anaerobically for 12 and 24 h. In vitro parameters and microbial diversity of the dominant bacteria following fermentation were analyzed using Denaturing Gradient Gel Electrophoresis (DGGE), band cloning and sequencing of the V3 region. Results showed that total gas production increased with the advancement of incubation (p<0.05). pH values of FAs and control groups were decreased except the FA4 group which increased somewhat from 12 to 24 h (p<0.05). Ammonia nitrogen ($NH_3$-N) and $H_2S$ gas concentrations were comparatively lower in both stages in FA4 treatment than in the other groups (p<0.05). Hence, $NH_3$-N concentrations in liquid phases were increased (p<0.05) from 12 to 24 h, but the trend was lowest in FA4 than in the other groups at both stages. The total VFA production was comparatively lower and butyrate levels were moderate in FA4 group than in the the other groups during both stages (p<0.05). Indirect odor-reducing compounds such as $NO_2$, $NO_3$ and $SO_4$ concentrations were higher in the FA4 and FA3 than in the other groups at 24 h (p<0.05). After fermentation, ten dominant bands appeared, six of which appeared in all samples and four in only the FA4 treated group. The total number of DGGE bands and diversity was higher in the FA4-group compared to other groups. Additionally, similarity indices were lowest (71%) in the FA4, which represented a different bacterial community compared with the other groups. These findings indicate that $NH_3$-N, $H_2S$ and VFA production was minimal, and pH was also better in the FA4 group than in the other groups. Furthermore, the conversion of odor-reducing indirect compounds or their intermediates was higher in the FA4 group in compared to the other groups. FA4 group generated less odorous products and more indirect products by in vitro fermentation at 24 h, and their microbial pattern appeared to differ from that of the other groups. These findings suggest that this particular FA could change the microbial population, which may have a beneficial effect on odor reduction. It is recommended that the oregano lippia may be supplied to growing pigs as FA along with excess carbohydrate sources to reduce the production of odorous compounds.