• Clinical and Experimental Reproductive Medicine
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• 제28권1호
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• pp.25-31
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• 2001

연구목적: $Isolate^{(R)}$ gradient와 swim-up 방법이 정자의 형상 및 정밀정자형태 (strict morphology)에 미치는 영창을 비교분석하고, 이러한 정자처리방법이 정자의 동결-융해과정에 미치는 영향을 비교하고자 하였다. 연구재료 및 방법: 20명의 정상 정자를 대상으로 하였으며 각각의 정자는 두 가지 정자처리방법으로 나누어 정자의 형상과 정밀정자형태를 컴퓨터를 이용한 정자자동분석기를 통하여 측정하였고, 동결보호제의는 TYB 용액을 사용하였으며, 동결 및 융해는 cryo Magic사의 기계를 사용하였다. 통계는 SPSS PC+(version7.0)를 이용하였으며 통계학적인 유의성은 p<0.05로 하였다. 결과: 정자의 농도는 $Isolate^{(R)}$ gradient 처리군이 swim-up 처리군보다 유의성 있게 높았으나 ($51.2{\pm}40.1,\;156.6{\pm}64.3$), 운동성 VCL, VSL, VAP, Linearity, 및 ALH는 swim-up 처리군에서 유의성 있게 높았다. 정밀 정자형태는 swim-up 처리군과 $Isolate^{(R)}$ gradient 처리군에서 차이가 없었다 ($53.7{\pm}6.8$ vs $50.3{\pm}9.1%$). 동결-융해과정 중 두 가지 정자 처리군에서 정자의 형상들은 swim-up 처리군에서 전반적으로 높은 양상을 보였으나, 정밀정자형태는 $Isolate^{(R)}$ gradient 처리군이 swim-up 처리군 보다 감소율이 컸지만 두 군간에 유의한 차이는 없었다 ($12.8{\pm}8.5$ vs $8.6{\pm}6.6$). 결론: 정상 정자에서 swim-up 방법이 $Isolate^{(R)}$ gradient 방법보다 정자 회수율은 우수하였으나, 동결-융해과정 중 정밀정자형태에는 차이가 없어 두 방법을 상호보완적으로 사용할 수 있을 것으로 사료된다.

• Clinical and Experimental Reproductive Medicine
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• 제28권4호
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• pp.295-300
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• 2001

Objective : To assess the effects of progesterone and acetyl-L-carnitine used after treated with Isolate�� gradient before semen cryopreservation-thawing on sperm parameters and membrane integrity. Material and Methods : From April 2001 to July 2001, ten normal male partner of couples who were visited in vitro fertilization (IVF) clinics. the semens were treated with $Isolate^{(R)}$ gradient before cryopreservation, spermatozoa was incubated with progesterone (1, 5 and $10{\mu}M$), acetyl-L-carnitine (2.5, 5 and $10{\mu}M$), or both (progesterone, $1{\mu}M$; and acetyl-L-carnitine, $5{\mu}M$) for 30 min. Results: There were no differences in sperm parameters and vital stain among isolate only treated group, progesterone (1, 5 and $10{\mu}M$), acetyl-L-carnitine (2.5, 5 and $10{\mu}M$) and both (progesterone, $1{\mu}M$; and acetyl-L-carnitine, $5{\mu}M$). But, in high concentration of acetyl-L-carnitine ($10{\mu}M$) treated group, sperm parameters and vital stain were decreased. The statistical method was used ANOVA (Kruskal-Wallis test) and p value was <0.01. Conclusions : Neither progesterone nor acetyl-L-carnitine show to be protective effect on the cryodamage assessed by sperm parameters and vital stain (eosin-Y stain) in normal sperm. High concentration of acetyl-L-carnitine ($10{\mu}M$), however, was harmful effect on cryoprevention.

• Natural Product Sciences
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• 제15권3호
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• pp.144-150
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• 2009

Gradient centrifugal partition chromatography (GCPC) method was developed and applied to isolate 3,5-dimethoxyphenanthrene-2,7-diol (DMP) and batatasin-I (BA-I) from the dichloromethane soluble extract of Dioscorea opposita. In this method, the lower phase of n-hexane-methanol-water system (HMW, 10 : 9 : 1, v/v) was used as a mobile phase A (MpA) and water was used as a mobile phase B (MpB). This gradient CPC method is comparable to that of reversed-phase HPLC method in that the stationary upper-phase of HMW (10 : 9 : 1 v/v) works as if it were reversed-phase silica gel due to its hydrophobic property, while the lower phase (MpA) and water (MpB) functioned as hydrophilic mobile phases. The initial condition of the mobile phase was 20% MpA/80% MpB and maintained for 150 min to obtain DMP (1.2 mg), and then MpA was increased up to 50% to elute BA-I (1.7 mg). The purities of DMP and BA-I were 94.1% and 98.3% with the recovery yields of 83% and 86%, respectively. Similar results were obtained by linear-gradient CPC. The CPC peak fractions were identified by comparing their retention time to those of authentic samples of DMP and BA-I and their spectroscopic data ($^1$H NMR and $^{13}$C NMR) to those of literature values.

• Clinical and Experimental Reproductive Medicine
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• 제43권4호
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• pp.199-206
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• 2016

Objective: This study was carried out to investigate the correlations of the sperm DNA fragmentation index (DFI) with semen parameters and apoptosis, and to investigate the effects of density-gradient centrifugation (DGC) and magnetic-activated cell sorting (MACS) on reducing the proportion of sperm with DNA fragmentation and protamine deficiency. Methods: Semen analysis and a sperm DNA fragmentation assay were performed to assess the correlations between semen parameters and the DFI in 458 semen samples. Sperm with progressive motility or non-apoptosis were isolated by DGC or MACS, respectively, in 29 normozoospermic semen samples. The effects of DGC or MACS alone and of DGC and MACS combined on reducing the amount of sperm in the sample with DNA fragmentation and protamine deficiency were investigated. Results: The sperm DFI showed a significant correlation (r=-0.347, p< 0.001) with sperm motility and morphology (r=-0.114, p< 0.05) but not with other semen parameters. The DFI ($11.5%{\pm}2.0%$) of semen samples was significantly reduced by DGC ($8.1%{\pm}4.1%$) or MACS alone ($7.4%{\pm}3.9%$) (p< 0.05). The DFI was significantly further reduced by a combination of DGC and MACS ($4.1%{\pm}1.3%$, p< 0.05). Moreover, the combination of DGC and MACS ($1.6%{\pm}1.1%$, p< 0.05) significantly reduced the protamine deficiency rate of semen samples compared to DGC ($4.4%{\pm}3.2%$) or MACS alone ($3.4%{\pm}2.2%$). Conclusion: The combination of DGC and MACS may be an effective method to isolate high-quality sperm with progressive motility, non-apoptosis, high DNA integrity, and low protamine deficiency in clinical use.

• Journal of Applied Biological Chemistry
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• 제64권4호
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• pp.391-397
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• 2021

Four compounds were isolated from Salvia plebeia aerial parts. Silica gel open column chromatography with a gradient elution system was used to isolate and purify these compounds. Nuclear magnetic resonance spectroscopy and mass spectroscopy were used for structural elucidation and identification, while electronic circular dichroism was used to confirm the absolute configuration. The structures were determined to be 𝛽-sitosterol (1), (-)-1S,5S,8S,10R-1-acetoxy-8-hydroxy-2-oxoeudesman-3,7(11)-dien-8,12-olide (2), ursolic acid (3), and N-methylhydroxylamine (4). Compounds 2 and 4 were isolated for the first time from this plant. Compound 2 was quantitatively analyzed via HPLC/UV. The results showed that the methanol extract of S. plebeia had a higher content of compound 2 (1.20 mg/g) than the ethanol extract (0.55 mg/g). This study could be used as a preliminary step in conducting HPLC/UV analysis of sesquiterpenoids in S. plebeia extract to assess their bioavailability and potency.

• Journal of Microbiology and Biotechnology
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• 제30권7호
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• pp.1005-1012
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• 2020

Acid mine drainage (AMD) has been a serious environmental issue that threatens soil and aquatic ecosystems. In this study, an acid-tolerant sulfate-reducing bacterium, strain S4, was isolated from the mud of an AMD storage pond in Vietnam via enrichment in anoxic mineral medium at pH 5. Comparative analyses of sequences of the 16S rRNA gene and dsrB gene involved in sulfate reduction revealed that the isolate belonged to the genus Desulfovibrio, and is most closely related to Desulfovibrio oxamicus (with 99% homology in 16S rDNA sequence and 98% homology in dsrB gene sequence). Denaturing gradient gel electrophoresis (DGGE) analyses of dsrB gene showed that strain S4 represented one of the two most abundant groups developed in the enrichment culture. Notably, strain S4 was capable of reducing sulfate in low pH environments (from 2 and above), and resistance to extremely high concentration of heavy metals (Fe 3,000 mg/l, Zn 100 mg/l, Cu 100 mg/l). In a batch incubation experiment in synthetic AMD with pH 3.5, strain S4 showed strong effects in facilitating growth of a neutrophilic, metal sensitive Desulfovibrio sp. strain SR4H, which was not capable of growing alone in such an environment. Thus, it is postulated that under extreme conditions such as an AMD environment, acid- and metal-tolerant sulfate-reducing bacteria (SRB)-like strain S4 would facilitate the growth of other widely distributed SRB by starting to reduce sulfate at low pH, thus increasing pH and lowering the metal concentration in the environment. Owing to such unique physiological characteristics, strain S4 shows great potential for application in sustainable remediation of AMD.

• Journal of Microbiology and Biotechnology
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• 제23권11호
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• pp.1617-1626
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• 2013

The Bacterial community structure and its complexity of the enrichment culture during the isolation and screening of imidacloprid-degrading strain were studied using denaturating gradient gel electrophoresis analysis. The dominant bacteria in the original tea rhizosphere soil were uncultured bacteria, Rhizobium sp., Sinorhizobium, Ochrobactrum sp., Alcaligenes, Bacillus sp., Bacterium, Klebsiella sp., and Ensifer adhaerens. The bacterial community structure was altered extensively and its complexity reduced during the enrichment process, and four culturable bacteria, Ochrobactrum sp., Rhizobium sp., Geobacillus stearothermophilus, and Alcaligenes faecalis, remained in the final enrichment. Only one indigenous strain, BCL-1, with imidacloprid-degrading potential, was isolated from the sixth enrichment culture. This isolate was a gram-negative rod-shaped bacterium and identified as the genus Ochrobactrum based on its morphological, physiological, and biochemical properties and its 16S rRNA gene sequence. The degradation test showed that approximately 67.67% of the imidacloprid (50 mg/l) was degraded within 48 h by strain BCL-1. The optimum conditions for degradation were a pH of 8 and $30^{\circ}C$. The simulation of imidacloprid bioremediation by strain BCL-1 in soil demonstrated that the best performance in situ (tea soil) resulted in the degradation of 92.44% of the imidacloprid (100 mg/g) within 20 days, which was better than those observed in the ex situ simulations that were 64.66% (cabbage soil), 41.15% (potato soil), and 54.15% (tomato soil).