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Optimization of Human Embryonic Stem Cells into Differentiation of Dopaminergic Neurons in Vitro: I. Additive Effect of Neurotrophic Factor on Human Embryonic Stem Cells

  • 이금실;김은영;이영재;신현아;조황윤;이훈택;정길생;박세필;임진호
    • 한국동물번식학회:학술대회논문집
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    • 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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    • pp.79-79
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    • 2003
  • Embryonic stem cells are capable of differentiating into a variety of cell lineages. However, the ultimate results of differentiation in vitro greatly depend on the duration of treatment and kinds of differentiating inducers added. In order to investigate the efficiencies of various differentiation inducers and the methods of treatment, we examined differentiation patterns of human embryonic stem cell (hESC, MB03) according to several different protocols. Exp. I) Upon differentiation using retinoic acid and ascorbic acid (RA/AA), embryoid bodies (EB, for 4days) derived from hESC was exposed to Rh (10$^{-6}$ M) and AA (50 mM) for 4 days, and were allowed to differentiate in N2 medium for 7, 14, 21, or 28 days. Exp. II) When bFGF was used, neuronal precursor cells were selected for 8 days in N2 medium after EB formation. After selection, cells were expanded at the presence of bFGF (20 ng/ml) for another 6 days followed by a final differentiation in N2 medium for 7, 14, 21 or 28 days. Exp. III) In addition, to examine the effects of neurotrophic factors in the production of mature neurons, groups of cells were exposed to either BDNF (5 ng/ml) or TGF-$\alpha$(10 ng/ml) during the 28 days of final differentiation. Differentiation patterns of RA/AA or bFGF treated groups were very similar; approximately 82% and 83% of the cells, respectively, were positive for anti-NF200 antibody, while it was about 10% and 11%, respectively, for anti-NF160 antibody in 28 days in N2 medium. Alsor, cells expressing TH were as low as 5%, while the cells doubled when matured at the presence of either BDNF or TGF-$\alpha$. Cells immunoreactive to anti-GAD antibody were approximately 20%. These results suggest that a maturation step rather than differentiation induction step, which is formation of EB, effects more decisively to the ultimate differentiation pattern.

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자외선의 상이한 선양을 조사한 브로일러 병아리의 다리 피부중 비타민 $C_3$ 함양의 경시적 변화 (Time Course Variation of Vitamin $C_3$ Content in Leg Skin of Broiler Chicks Exposed to Different Dose of UVB Light)

  • 장윤환;김강수;여영수;강훈석;조인호;배은경
    • 한국가금학회지
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    • 제20권2호
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    • pp.93-105
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    • 1993
  • 본 연구는 31 일령 broiler 병아리에 297 nm의 자외선을 조사하여 경과시간에 따라 다리의 피부를 채취하고 previtamin D$_3$(PreD$_3$), lumistero1$_3$(고), tachysterol$_3$(T$_3$), Vitamin D$_3$(VD$_3$) 및 provitaminD$_3$ (PreD$_3$) 함양을 측정코저 실시되었다. Broiler Hubbard계통 1일령 병아리 82수(2조사시간$\times$9 경과시간$\times$4회복+10대조구)를 무창약등 육계사에 넣고 VD-결핍사료로 315간 사육한 후 조사선양 0.204mJ/$ extrm{cm}^2$ (30분 조사) 또는 0.409mJ/$\textrm{cm}^2$(60분 조사)로 UVB 등을 조사하였으며 0, 6, 12, 18, 30, 42, 66, 90또는 138시간 후에 다리의 피부를 채취하였다. 9% ethyl acetate/n-hexane으로 처리하여 지질을 추출하였으며 Sep-Pak silica cartridge로 정제한 후 순상 HPLC로 PreD$_3$, L$_3$, T$_3$, VD$_3$ 및 ProD$_3$의 함양을 분석하였다. UVB를 조사하지 않은 대조구 병아리의 다리 피부중에는 PreD$_3$, L$_3$, T$_3$, VD$_3$가 검출되지 않았으며 PreD$_3$ 함양은 966$\pm$89ng/$\textrm{cm}^2$이었다. 0.204 또는 0.409mJ/$\textrm{cm}^2$(30분 또는 60분) .조사시 다리 피부중 PreD$_3$의 mole %(절대함량 ng/$\textrm{cm}^2$)는 열사직후 각각 4.67%(44ng/$\textrm{cm}^2$) 또는 3.97%(37ng/$\textrm{cm}^2$)이었으며 시간이 경과됨에 따라 점차 감소되었다. 병아리의 다리 피부 면적은 25.81$\pm$0.50$\textrm{cm}^2$이었으며 다리 피부전체에서 합성된 PreD$_3$는 각각 0시간대에 961ng/$\textrm{cm}^2$, 738ng/$\textrm{cm}^2$이었다. 다리 피부중의 은 0시간 경과 시 각각 1.24%(12ng/$\textrm{cm}^2$) 또는 0.92%(9ng/$\textrm{cm}^2$)이었으며 시간이 지남에 따라 점차 감소되었다 함량은 각각 0.58%(5ng/$\textrm{cm}^2$) 또는 0.57%(6ng/$\textrm{cm}^2$)이었으며 역시 시간의 경과와 더불어 감소되었다. VD$_3$함량은 0시간대에 각각 2.13(21ng/$\textrm{cm}^2$) 또는 0.97%(16 ng/$\textrm{cm}^2$), 12시간대에 4.30%(33ng/$\textrm{cm}^2$) 또는 6.40%(76ng/$\textrm{cm}^2$)을 나타내었고 0.204mJ/$\textrm{cm}^2$ 조사시에는 18시간대에, 0.409mJ/$\textrm{cm}^2$ 처리시에는 30시간 후에 최고치를 보였다 PreD$_3$가 VD$_3$로 열전환되는데 각각 18 또는 30시간이 소요된다는 것을 시사한다. 다리 피부중의 PreD$_3$함량은 0시간대에 각각 948또는 815ng/$\textrm{cm}^2$이었으며 대조구의 ProD$_3$ 함량 966ng/$\textrm{cm}^2$에 비하면 18 또는 151ng/$\textrm{cm}^2$의 PreD$_3$가 PreD$_3$, L$_3$, T$_3$ 및 VD$_3$이성화된 것이다. mole %로 보았을 때는 각각 8.63%, 6.43%가 광합성물로 전변 된 것이다 결논적으로 자외선을 2배 .무사하였을 때 VD$_3$등 광생성물이 2배는 아니지만 더 많이 생성된다는 것이 증명되었다.

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Relationships between Methionine Supply, Nitrogen Retention and Plasma Insulin-like Growth Factor-I in Growing Sheep Nourished by Total Intragastric Infusions

  • Li, Chong;Zhao, Guangyong
    • Asian-Australasian Journal of Animal Sciences
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    • 제24권10호
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    • pp.1393-1398
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    • 2011
  • Four 4-month old Charolais${\times}$Dorset male sheep (initial liveweight $25.0{\pm}1.1\;kg$), fitted with rumen and abomasal fistulas and nourished by total intragastric infusions, were used to study the relationships between methionine (Met) supply, nitrogen (N) retention and plasma insulin-like growth factor-I (IGF-I). Four graded levels of Met, i.e. 0 g/16 g N, 1.76 g/16 g N, 3.52 g/16 g N and 7.04 g/16 g N, were infused into abomasums as experimental treatments. The sheep and treatments were allocated in a $4{\times}3$ incomplete Latin square design (Yudon square design). The experiment lasted 3 periods and each period was 10 days. Quadratic correlations were found between Met level (x, g/16 g N) and N retention (y, g/d): y = $-0.03x^2$+0.41x+2.62, $r^2$ = 0.66, n = 12, p = 0.008, and between methionine level (x, g/16 g N) and plasma IGF-I concentration (y, ng/ml): y = $0.80x^2$-4.53x+190.24, $r^2$ = 0.51, n = 12, p = 0.009. No significant correlation was found between plasma IGF-I (x, ng/ml) and N retention (y, g/d) (p>0.05). It was concluded that Met level had a significant influence on N retention and plasma IGF-I concentration whereas IGF-I might not be an important mediator in the regulation of N metabolism by Met in growing sheep nourished by total intragastric infusions.

온라인 자동화 시료 전처리 및 HR Orbitrap LC/ESI-MS를 이용한 환경시료 중 잔류 의약물질 분석방법 확립 (Establishment of the Analytical Method for Residual Pharmaceuticals in Raw Water Using Online Sample Preparation and High Resolution Orbitrap LC/ESI-MS)

  • 황윤정;신상희;박종숙
    • 한국물환경학회지
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    • 제29권3호
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    • pp.409-419
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    • 2013
  • In this study, the analytical method for 27 residual pharmaceuticals in raw water was developed. Online sample preconcentration/extraction and analysis with high resolution Orbitrap mass spectrometry (LC-ESI/Orbitrap MS) were performed. The calibration curves showed good linearities (above $r^2$ = 0.998) in the range of 5 ~ 1,000 ng/L. The method detection limit and the limit of quantification were 1.1 ~ 10.0 ng/L and 3.4 ~ 31.7 ng/L, respectively. Recoveries of the target compounds were between 70.1% and 115.8% (except cefadroxil, cefradine, vancomycin, and iopromide (50.2 ~ 67.0%)). The optimized analytical method can be useful to determine the residual pharmaceuticals in raw water.

Spectrofluorimetric determination of Trimethoprim in pharmaceutical preparations

  • Amneen Mohammed Alsayegh;Abbas N. Alshirifi
    • 분석과학
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    • 제36권5호
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    • pp.250-257
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    • 2023
  • The development of a spectrofluorimetric method for the determination of trimethoprim according to the reaction between O-phthalaldehyde (OPA) in highly alkaline media, and 2-mercaptoethanol (2ME) and this reaction gives fluorescent product measured at (458) nm when excited at (342) nm. The optimization of the analytical parameters that influence intensity was investigated. The intensity of fluorescence of the formed product was linearly related to the concentration of trimethoprim in the (100-1200) ng mL-1 range. The limit of detection and limit of quantification were estimated to be (22.54) ng mL-1 and (75.15) ng mL-1 respectively. The utility of the proposed methods was successfully verified by analysis of trimethoprim in pure and real pharmaceutical preparations with high accuracy, the recovery percentages Re%, were found to be (100.5) % and (99.76) % for pure drug and pharmaceutical preparations respectively.

Plasma Concentrations of Morphine during Postoperative Pain Control

  • Park, Hahck-Soo;Kim, Jong-Hak;Kim, Yi-Jeong;Kim, Dong-Yeon
    • The Korean Journal of Pain
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    • 제24권3호
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    • pp.146-153
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    • 2011
  • Background: Morphine has been commonly used for postoperative pain control. We measured plasma concentrations of morphine and compared the efficacy and safety of continuous epidural analgesia (CEA) using morphinebupivacaine with intravenous patient controlled analgesia (IV-PCA) with morphine for 48 hrs after the end of the operation. Methods: Nineteen patients undergoing Mile's operation were assigned to receive a morphine loading dose of 5 mg followed by IV-PCA with 0.1% morphine (IV-PCA group, n = 9) or a morphine loading dose of 2 mg and 0.125% bupivacaine 10 ml, followed by CEA with 0.004% morphine and 0.075% bupivacaine at a rate of 5 ml/hr (CEA group, n = 10). The plasma concentrations of morphine were measured and visual analog scales (VAS) for pain were recorded at 1, 6, 12, 24, and 48 hr postoperatively and the effects on respiration and any other side effects were noted. Results: The mean maximal and minimal levels of plasma morphine were $40.2{\pm}21.2\;ng/ml$ and $23.4{\pm}9.7\;ng/ml$ for the IV-PCA group and $11.8{\pm}3.5\;ng/ml$ and $8.2{\pm}1.9\;ng/ml$ for the CEA group, respectively. Resting and dynamic pain scores were significantly lower in the CEA group than in the IV-PCA group. There were no significant differences for the effects on respiration and for any side effects between the two groups. Conclusions: We evaluated plasma concentrations of morphine with CEA using morphine-bupivacaine and IV-PCA using morphine for the postoperative pain control. The CEA group had better postoperative analgesia than that of the IV-PCA group and the incidence of side effects were not significantly different between the two groups.

RP-HPLC-PAD를 이용한 갈근(葛根)과 갈근(葛根) 함유 처방의 Puerarin과 Daidzin 분석 (Determination Method of Puerarin and Daidzin from Puerariae Radix by Reversed-Phase HPLC with Pulsed Amperometric Detection)

  • 초진영;권하정;정지선;이제현;홍선표
    • 대한본초학회지
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    • 제23권4호
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    • pp.171-177
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    • 2008
  • Objectives: This study presents a reversed-phase high-performance liquid chromatography- pulsed amperometric detection(RP-HPLC-PAD) method for the determination of puerarin and daidzin in Puerariae Radix extract and Chinese medicinal preparations. Methods: Chromatographic separation was performed using a 10% acetonitrile with a reversed-phase column(Unison UK-C18, $100mm{\times}2.0mm$ I.D.; $3{\mu}m$). The analyses were detected by pulsed amperometric detector(PAD) in alkaline conditions by combining with post-column NaOH solution. Geniposide was used as an internal standard. Results: The limit of detection(S/N=3) and the limit of quantification(S/N=10) were 0.025 ng, 0.075 ng for puerarin, and 0.05 ng, 0.15 ng for daidzin, respectively. The intra- and inter-day precisions(RSDs) were less than 6.5% and average recoveries of puerarin were 99.7-101.3% and those of daidzin were 101.0-102.8%. Conclusions: According to above results, we developed a determination method for puerarin and daidzin in Puerariae Radix with high sensitivity and selectivitely.

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경기 시흥안산지역 초등학교 고학년의 스마트폰 과의존 위험 수준에 따른 식태도, 식행동 및 스마트폰 사용관련 식습관 (Dietary Attitudes, Dietary Behaviors, and Smartphone Usage-related Dietary Habits According to Risk Level of Smartphone Overdependence of Elementary School Students in the Siheung and Ansan Areas)

  • 한소희;박소현;이은영;장경자
    • 한국식생활문화학회지
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    • 제32권5호
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    • pp.437-445
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    • 2017
  • This study was conducted to investigate the nutrition knowledge, dietary attitudes, dietary behaviors, smartphone usage-related dietary habits, and health-related lifestyles of higher grade students in elementary school according to risk level of smartphone overdependence. Subjects were 286 fifth and sixth grade students of elementary school in the Siheung and Ansan areas. Data were collected using self-administered questionnaires and analyzed using SPSS v. 20.0. Based on scores of S-scale (smartphone overdependence scale for adolescents), subjects who used smartphone were classified into an overdependence group (ODG, n=52) and a normal group (NG, n=234). The ODG showed a significantly longer usage time than NG on both weekdays and weekends (p<0.001). There were no significant differences in total scores of nutrition knowledge between the ODG and NG. Average total scores of dietary attitude and dietary behavior in the ODG were significantly lower than in the NG (p<0.001). There were higher risks of ODG for skipping meals (OR=8.3, 95% CI=4.027-17.099), changing eating speed (OR=4.4, 95% CI=2.209-8.822), and changing meal amount due to smartphone usage (OR=2.9 95% CI=1.233-6.623). Therefore, education programs are needed to ensure proper dietary behaviors and habits among smartphone overdependent elementary school students.

Determination of Ketorolac in Human Serum by High-performance Liquid Chromatography

  • Chun, In-Koo;Kang, Hyun-Hee;Gwak, Hye-Sun
    • Archives of Pharmacal Research
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    • 제19권6호
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    • pp.529-534
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    • 1996
  • A high-performance liquid chromatographic (HPLC) assay has been developed for the determination of ketorolac in human serum using a new extraction method with a good recovery. Human serum samples (1.0 ml) spiked with known concentrations of ketorolac tromethamine and 10${\mu}g$ of ketoprofen as the internal standard (IS) were acidified with 200${\mu}l$ of 1 N HCl and extracted with 7 ml of n-hexane-ether (7:3 v/v). Extracts were centrifuged and organic layer was back-extracted with 400${\mu}l$ of 0.1% tromethamine solution. Twenty .mu.l of centrifuged aqueous layer was injected onto a reversed-phase octyl column and eluted with a mixture of acetonitrile, water, methanol, and triethylamine [35:55:10:0.1 (v/v), pH 3.0] at a flow rate of 1.0 ml/min. Ultraviolet detection of ketorolac and IS was carried out at 300 nm. The calibration curve obtained using peak area ratios showed a good linearity (in concentration range 10-150 ng/ml $r^2$=O.9944; in range 50-2000 ng/ml, r$^{2}$=0.9998). The mean intra-day accuracy and precision for this HPLC method were found to be 3.6 and 3.7%, respectively. The mean inter-day accuracy and precision were found to be 4.0 and 3.7%, respectively, in the concentration range 50-2000 ng/ml. The recovery of ketorolac from serum was 92.0 $({\pm}5.7)$ % at the concentration of 100 ng/ml. This method proved to be readily applicable to the assay of ketorolac in human serum.

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