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Study on stability test of in process sample of recombinant Protein A (재조합 단백질 A 제조공정시료의 안정성실험에 관한 연구)

  • Kim, Yoo Gon;Lee, Woo Jong;Won, Chan Hee;Shin, Chul Soo
    • Analytical Science and Technology
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    • v.25 no.6
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    • pp.483-491
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    • 2012
  • This study is to investigate the issues on how to secure stability during the purification process for the production of recombinant protein A. The final recombinant protein A is produced by passing through the cation exchange column (SP) and the anion-exchange column (Q) during the production process, for which the samples produced by the step-by-step processes can be exposed to trouble in securing stable storage in case the next process cannot be taken within the proper time period. Accordingly, this study aims to evaluate the proper storage conditions and length of time when storing samples produced in the production process. That is, in this study, how to store fair samples, how long the storage period should be set up, and how to evaluate the security of its quality depending on time are dealt with. The items to be experimented with were enodotoxin, SDS-PAGE, HPLC purity and concentration. Experimental results showed that after passing the cation exchange column, when stored at $4^{\circ}C$ or room temperature, SDS-PAGE showed a major band, endotoxin is 5.0 Eu/mg or less, and concentration is on average of 8.21 to 8.24 mg/mL and RSD% 0.10~0.62%. In addition, HLPC purity showed somewhat stable results; at the HPLC purity 214 nm, the average is 99.24% to 99.37% and RSD% is 0.22~0.29%, while the average is 89.72% to 89.80% and RSD% 0.62~1.26% at 280 nm. On the contrary, after passing the anion exchange column, when stored at $4^{\circ}C$ or room temperature, SDS-PAGE revealed the major band, endotoxin is 0.5 Eu/mg or less, and concentration is on average of 5.59 mg/mL and RSD% 0.03~0.10%. when it comes to HLPC purity, the result showed that at the HPLC purity 214 nm, the average is 99.74% and RSD% is 0.10~0.11%, while the average is 96.16% to 96.85% and RSD% 0.72~1.13%. In conclusion, the stability of fair samples of recombinant protein A during the manufacturing process could be obtained without substance decomposition for 7~8 days at $4^{\circ}C$ or 20~21 days at room temperature.

Validation of Method Determining Coixol in Coix lachryma-jobi var. ma-yuen Roots Extract (율무근 추출물의 Coixol 성분 분석법 검증)

  • Kwon, Jin Gwan;Seo, Changon;Choi, Yun-Hyeok;Choi, Chun Whan;Kim, Jin Kyu;Jeong, Wonsik;Lee, Ji Eun;O, Kyeong Hee;Hong, Seong Su
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.46 no.8
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    • pp.952-956
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    • 2017
  • An high performance liquid chromatography (HPLC) analysis method was developed for standard determination of coixol as a functional cosmetic material in Coix lachryma-jobi var. ma-yuen roots extract. HPLC was performed on a $C_{18}$ Unison US column ($4.6{\times}250mm$, $5{\mu}m$ column) using a gradient elution of 0.1% (v/v) trifluoroacetic acid and acetonitrile at a flow rate of 1.0 mL/min at $30^{\circ}C$. The analyte was detected at 290 nm. The HPLC method was validated in accordance with the International Conference on Harmonization guideline of analytical procedures with respect to specificity, precision, accuracy, and linearity. The limit of detection and quantitation were 0.07 and 0.25 mg/mL, respectively. Calibration curves showed good linearity ($R^2$>0.9995), and the precision of analysis was satisfied (less than 0.29%). Recoveries of quantified compounds ranged from 98.36 to 100.30%. This result indicates that the established HPLC method is very useful for the determination of a marker compound in C. lachryma-jobi var. ma-yuen roots extracts.

Changes in Shelf-Life, Water Activity, and Texture of Rye-Wheat Mixed Bread with Naturally Fermented Raisin Extract and Rye Sourdough during Storage (건포도 천연 발효액과 호밀 사워도우를 이용한 호밀-밀 혼합빵의 저장 중 저장수명, 수분활성도 및 조직감의 변화)

  • Kim, Mun-Yong;Chun, Soon-Sil
    • Korean journal of food and cookery science
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    • v.25 no.2
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    • pp.170-179
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    • 2009
  • This study examined effects on shelf-life extension and prevention of starch retrogradation in rye-wheat mixed bread (RWMB) samples prepared with substitutions of 20, 40, 60, and 80% rye sourdough (RSD) as well as a control made with the addition of naturally fermented raisin extract. These effects were investigated using mold growth, water activity, and textural characteristics during a storage period of 8 days at $20^{\circ}C$. The activities of the rye sourdough were examined in terms of pH, total titratable acidity, general bacteria, lactic acid bacteria, and yeast counts. As the incubation time of the sour dough increased, pH decreased, while total titratable acidity increased. General bacteria, lactic acid bacteria, and yeast counts increased with increasing incubation time. Mold grew on the external surface of the control group after 6 days and on the surface of the RSD bread samples after 8 days, respectively. With regard to water activity, the control group had increasing activity as the storage period increased. In addition, the 20, 40, 60, and 80% RSD samples had increasing water activities until the 4th day however, these strongly decreased by the 6th day. In terms of textural characteristics, as the storage period increased, hardness increased, while springiness, cohesiveness, and resilience decreased. There were no significant differences in gumminess among the samples. The control, 20, 40, and 60% RSD samples had decreases in chewiness with increasing storage time, however, the 40% RSD sample did not differ significantly during the storage period. Finally, water activity was negatively correlated with hardness (p<0.05). In conclusion, the results indicate that substituting rye sourdough in rye-wheat mixed bread has a prolongation effect on shelf-life, but no effect on the prevention of starch retrogradation.

Optimization of Analytical Methods for Ochratoxin A and Zearalenone by UHPLC in Rice Straw Silage and Winter Forage Crops (UHPLC를 이용한 볏짚 사일리지와 동계사료작물의 오크라톡신과 제랄레논 분석법 최적화)

  • Ham, Hyeonheui;Mun, Hye Yeon;Lee, Kyung Ah;Lee, Soohyung;Hong, Sung Kee;Lee, Theresa;Ryu, Jae-Gee
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.36 no.4
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    • pp.333-339
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    • 2016
  • The objective of this study was to optimize analytical methods for ochratoxin A (OTA) and zearalenone (ZEA) in rice straw silage and winter forage crops using ultra-high performance liquid chromatography (UHPLC). Samples free of mycotoxins were spiked with $50{\mu}g/kg$, $250{\mu}g/kg$, or $500{\mu}g/kg$ of OTA and $300{\mu}g/kg$, $1500{\mu}g/kg$, or $3000{\mu}g/kg$ of ZEA. OTA and ZEA were extracted by acetonitrile and cleaned-up using an immunoaffinity column. They were then subjected to analysis with UHPLC equipped with a fluorescence detector. The correlation coefficients of calibration curves showed high linearity ($R^2{\geq_-}0.9999$ for OTA and $R^2{\geq_-}0.9995$ for ZEA). The limit of detection and quantification were $0.1{\mu}g/kg$ and $0.3{\mu}g/kg$, respectively, for OTA and $5{\mu}g/kg$ and $16.7{\mu}g/kg$, respectively, for ZEA. The recovery and relative standard deviation (RSD) of OTA were as follows: rice straw = 84.23~95.33%, 2.59~4.77%; Italian ryegrass = 79.02~95%, 0.86~5.83%; barley = 74.93~97%, 0.85~9.19%; rye = 77.99~96.67%, 0.33~6.26%. The recovery and RSD of ZEA were: rice straw = 109.6~114.22%, 0.67~7.15%; Italian ryegrass = 98.01~109.44%, 1.65~4.81%; barley = 98~113.53%, 0.25~5.85%; rye = 90.44~108.56%, 2.5~4.66%. They both satisfied the standards of European Commission criteria (EC 401-2006) for quantitative analysis. These results showed that the optimized methods could be used for mycotoxin analysis of forages.

Signal compensation by the light scattering of sample aerosols in ICP-AES (ICP-AES에서 에어로졸의 광산란에 의한 신호의 보정)

  • Yeon, Pyung-Hum;Pak, Yong-Nam
    • Analytical Science and Technology
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    • v.25 no.4
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    • pp.223-229
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    • 2012
  • Analytical signal from ICP was compensated by the light scattering of sample aerosols. Reference scattering signal was generated by a He-Ne or diode laser, monitored for the amount of aerosol producing and used for the compensation of analytical signals. The result showed that significant improvement in precision could be achieved for the short-term signal (within 1 minute) from 3.4% to 0.9% RSD in signal and 14.9% to 4.2% for the long-term (10 minutes) for Be, Pb and Co. This method is very useful not only for the pulse type but for continuous type signals especially when a nebulizer is unstable. To improve long-term precision, higher stability is required in the scattering cell and detector as well as the reduction of noise from the line between a nebulizer and plasma.

Analysis of Cadaverine and Its Worker Honeybee Venom Content (Apis mellifera L.) (꿀벌(Apis mellifera L.) 일벌독의 생체아민 cadaverine 함량 및 분석법)

  • Choi, Hong Min;Kim, Hyo Young;Kim, Se Gun;Han, Sang Mi
    • Journal of Life Science
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    • v.31 no.2
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    • pp.144-148
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    • 2021
  • This study aimed to analyze the content and composition of a biological amine, cadaverine, isolated from the venom of worker honeybees (Apis mellifera L.). This biological amine―which has diverse functionality, such as anti-inflammatory and antibacterial effects―has not been previously reported in bee venom. An assay completed in 13 minutes was developed for the cadaverine present in the bee venom using an ultra-performance liquid chromatograph and a Halo C18 column with acetonitrile and water as the mobile phase. The specificity, accuracy, and precision of the assay were verified, and the assay was validated. The linearity for cadaverine in the bee venom was R2=0.99 or above, indicating a moderate level. The limit of detection and limit of quantification were both 0.3 ㎍/ml, and the rate of recovery was 97.6%-99.1%. The relative standard deviation (RSD) of the intra-day precision and inter-day precision for cadaverine was 0.25%-0.44% and 0.25%-1.25%, respectively, with an RSD that fell within 5% indicating excellent precision. Through this novel assay, it was found that the mean content of cadaverine was 1.10±0.05 mg/g. Our results indicated that the linearity, limit of detection, limit of quantification, and rate of recovery of the cadaverine assay were of a satisfactory level, and the cadaverine content of the bee venom was ably determined. This study provides basic data on cadaverine in bee venom, which will prove useful in further studies on the bioactivity of this component.

Evaluation of LA-ICP-MS Whole Rock Trace Element Analysis Using Fused Glass Bead of Silicate Rocks (규산염 암석의 알칼리 용융 유리원판에 대한 LA-ICP-MS 전암 미량원소 분석법 평가)

  • Kim, Myong Jung;Kim, Taehoon;Park, Kye-Hun;Lee, Ye Ji;Yang, Yun Seok;Moon, Jeongjin
    • The Journal of the Petrological Society of Korea
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    • v.24 no.2
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    • pp.141-147
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    • 2015
  • Using laser-ablation inductively coupled plasma mass spectrometry (LA-ICP-MS), we evaluated the reliability of trace element abundance data measured from the silicate rock references of JR-3, JG-3, JGb-1 and JB-1b using glass discs made by alkali fusion. For 28 elements including rare earth elements, relative standard deviations (RSD) are better than 7% in case when the concentrations of the elements in the rock samples are greater than 10 ppm. However, RSD shows somewhat increased values for the concentrations less than 10 ppm, but never exceeds 25%. Compared with previously reported averages of the compiled abundance data, our data display satisfactory results for the most cases with differences less than 10%. We suggest that LA-ICP-MS analysis using fused glass beads is a reliable, precise and time-saving method of trace element analysis for the silicate rocks spanning from mafic to felsic compositions.

Development of Jaspine B analysis using LC-MS/MS and its application: Dose-independent pharmacokinetics of Jaspine B in rats

  • Song, Im-Sook;Jeon, Ji-Hyeon;Lee, Jihoon;Lim, Dong Yu;Lee, Chul Haeng;Lee, Dongjoo;Choi, Min-Koo
    • Analytical Science and Technology
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    • v.34 no.2
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    • pp.37-45
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    • 2021
  • A rapid and simple LC-MS/MS analytical method in determining Jaspine B has been developed and validated in rat plasma. The standard curve value was 25 - 5000 ng/mL and the linearity, inter-day and intra-day accuracy and precision were within 15.0 % of relative standard deviation (RSD). The mean recoveries of Jaspine B ranged from 87.5 % to 91.2 % with less than 3.70 % RSD and the matrix effects ranged from 91.1 % to 108.2 % with less than 2.6 % RSD. The validated LC-MS/MS analytical method of Jaspine B was successfully applied to investigate the dose-escalated pharmacokinetic study of Jaspine B in rats following an intravenous injection of Jaspine B at a dose range of 1 - 10 mg/kg. The initial plasma concentrations and area under plasma concentration curves showed a good correlation with intravenous Jaspine B dose, indicating the dose independent pharmacokinetics of Jaspine B in rats. In conclusion, this analytical method for Jaspine B can be easily applied in the bioanalysis and pharmacokinetic studies of Jaspine B, including its administration at multiple therapeutic doses, or for making pharmacokinetic comparisons for the oral formulations of Jaspine B in small experimental animals as well as in vivo pharmacokinetic-pharmacodynamic correlation studies.

Optimization of Ascorbic Acid Extraction from Rugosa Rose (Rosa rugosa Thunb.) Fruit Using Response Surface Methodology and Validation of the Analytical Method

  • UM, Min;KIM, Ji-Woo;LEE, Jae-Won
    • Journal of the Korean Wood Science and Technology
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    • v.48 no.3
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    • pp.364-375
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    • 2020
  • In this study, ultrasound-assisted extraction was performed to extract ascorbic acid from rugosa rose (Rosa rugosa Thunb.) fruit. The optimal conditions were investigated by response surface methodology, using two variable including reaction time (16-44 min) and temperature (16-44℃). The ascorbic acid extraction was sensitive to the reaction time rather than the reaction temperature, and the optimal conditions for ascorbic acid extraction were 25℃ and 30 min. Ascorbic acid and gallic acid in the rugosa rose fruit extract were completely separated by HPLC, with a resolution factor of over 1.5 between the two. The correlation coefficient of the ascorbic acid was 0.999 in a linearity test for 50-150 ㎍/mL concentration of extract. The limit of detection and limit of quantification values were 0.16 ㎍/mL and 29.89 ㎍/mL, respectively. The relative standard deviations (RSD) for repeatability and reproducibility were determined, and each RSD showed good precision at less than 5% (N=6).

Validation, Measurement Uncertainty, and Determination of Bixin and Norbixin in Processed Foods of Animal Resources Distributed in Korea

  • Ga-Yeong Lee;Choong-In Yun;Juhee Cho;Young-Jun Kim
    • Food Science of Animal Resources
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    • v.43 no.6
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    • pp.949-960
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    • 2023
  • This research aimed to validate a high-performance liquid chromatography method for the quantitative determination of bixin and norbixin in various foods. The Diode Array Detector (495 nm) technique was used. Method was validated for specificity, linearity, limit of detection (LOD), limit of quantification (LOQ), precision, and accuracy, and the measurement uncertainty was assessed. The calibration curve showed excellent linearity (r2≥0.9999) over the tested concentration range of 0.2-25 mg/L. The LOD and LOQ were 0.03-0.11 and 0.02-0.05 mg/L for bixin and norbixin, respectively. The intra-and inter-day accuracies and precisions were 88.0±1.3-97.0±0.5% and 0.2%-2.6% relative SD (RSD) for bixin and 88.2±0.8-105.8±0.8% and 0.3%-2.7% RSD for norbixin, respectively. Inter-laboratory validation for accuracy and precision was conducted in three laboratories, and these results all met the AOAC guidelines. In addition, the relative expanded uncertainty (<22%) satisfied the CODEX recommendation. Furthermore, products distributed in Korea were monitored for annatto extracts using the proposed method to demonstrate its application. The developed analytical method is reliable for quantifying bixin and norbixin in various foods.