• ALGAE
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• 제37권2호
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• pp.175-183
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• 2022

Polar microorganisms produce physiologically active substances to adapt to harsh environments, and these substances can be used as biomedical compounds. The green microalga Micractinium variabile KSF0031, which was isolated from Antarctica, produced phytol, a natural antimicrobial agent. Furthermore, several polyunsaturated fatty acids (PUFAs), including omega-3, exhibit antioxidant properties. Here statistical methods (Plackett-Burman design and Box-Behnken design) were used to optimize the culture medium of KSF0031 to improve biomass production, and K₂HPO₄, MgSO₄·7H ₂O, and ammonium ferric citrate green (AFCg) were selected as significant components of the culture medium. Changes in the concentration of K₂HPO₄ and MgSO₄·7H ₂O as positive factors and AFCg as a negative factor affected cell growth to a remarkable degree. The biomass production in a 100 L culture using the optimized medium for 24 d at 18℃ was improved by 37.5% compared to that obtained using the original BG-11 medium. The quantities of PUFAs and phytol obtained were 13 mg g^-1 dry cell weight (DCW) and 10.98 mg g^-1 DCW, which represent improved yields of 11.70% and 48.78%, respectively. The results of this study could contribute to an improved production of phytol and fatty acids from Antarctic microalgae in the biomedical industry.

• 응용통계연구
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• 제25권1호
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• pp.215-223
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• 2012

반응표면 방법론은 어떤 공정을 개선하거나 최적화하는데 이용되는 아주 유용한 통계적방법이다. 이러한 최적조건을 추정하기 위하여 최적조건이 있으리라 예상되는 실험구역을 탐색하여 실험을 실시한다. 그런데 이 실험구역은 실험의 환경의 제약 그리고 연구자의 선택 등으로 그 모습이 다양하게 달라질 수 있다. 반응표면 설계는 실험구역의 모양에 따라 보통 둥그런 모양의 "구형설계"와 육면체 모양의 "입방형설계"로 구분한다. 구형설계는 회전성을 만족하거나 회전성에 상당히 근접하는 "유사회전성"을 갖는 특징이 있다. 반응표면 설계에서 가장 많이 사용되는 중심합성설계는 실험구역이 구형인 5수준 실험설계이다. 이 때, 축점의 ${\alpha}$값을 ${\alpha}=\sqrt{k}$ 대신 ${\alpha}=1$로 조정하면 5-수준이 아닌 3-수준 입방형 중심합성설계를 얻을 수 있다. 그러나 입방형 중심합성설계는 실험구역이 구형이 아니므로 회전성을 만족하지 못하는 문제가 있다. 이러한 이유로, 변수들의 수준 수를 3으로 제한하면서 실험구역은 구형인 실험설계가 필요할 때가 많다. 이에 대한 대표적 실험설계가 바로 박스-벤켄 실험설계이다. 이 실험설계는 구형의 실험구역으로 회전성을 만족하나 실험구역의 크기가 변수의 개수가 증가해도 제자리 수준으로 좁은 특징이 있다. 현실적으로 실험구역의 가상 자리 부분에 대한 예측에 관심이 있을 경우 변수의 개수가 많아지면 이에 비례하여 실험구역이 커지는 실험설계가 바람직하다. 본 논문은 3-수준 입방형설계에 비하여 실험구역이 유달리 좁은 박스-벤켄 실험설계를 보완하여 구형설계를 만족하면서도 다른 한편으로는 변수 수에 따라 실험반경이 커지는 3-수준 구형 반응표면 설계를 소개하고자 한다. 이 방법을 기존의 실험설계들과 비교한 결과 변수수가 비교적 작을 경우 실험횟수 등을 고려하여 응용가치가 있음을 확인하였다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제10권3호
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• pp.192-197
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• 2005

The effects of macronutrients $(NO_3^-,\; NH_4^+\;and\;PO_4^{3-})$ on cell growth and triterpenoids production in Centella asiatica cell suspension cultures were analyzed using the Box­Behnken response surface model experimental design. In screening and optimization experiments, $PO_4^{3-}$ as a single factor significantly influenced cell growth where increasing the phosphate level from 0.1 to 2.4 or 2.6 mM, elevated cell growth from 3.9 to $14\~16g/L$. The optimum values predicted from the response surface model are 5.05mM $NH_4^+$, 15.0mM $NO_3^-$ and 2.6mM $PO_4^{3-}$, yielding 16.0g/L cell dry weight with $99\%$ fitness to the experimental data. While the $NH_4^+-NO_3^-$ interaction influenced cell growth positively in the optimization experiment, $NH_4^+$ and $NO_3^-$ as single factors; and interactions of $NO_3^--PO_4^{3-},\;NH_4^+-PO_4^{3-}$ and $NH_4^+-NO_3^-$ were all negative in the screening experiment. Cell growth and the final pH level were positively affected by $PO_4^{3-}$, but negatively affected by $NH_4^+\;and\;NH_4^+-PO_4^{3-}$ interactions. The different effects of factors and their interactions on cell growth and final pH are influenced by a broad or narrow range of macronutrient concentrations. The productions of triterpenoids however were lower than 4mg/g cell dry weight.

• Natural Product Sciences
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• 제22권4호
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• pp.270-274
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• 2016

Green tea, the leaves of Camellia sinsneis (Theaceae), is generally acknowledged as the most consumed beverage with multiple pharmacological functions including antioxidant activity. This study was performed to analyze the effect of extraction conditions of green tea on its antioxidant effects using DPPH assay. Three extraction factors such as extraction solvent (EtOH, 0 - 100%), extraction time (3 - 15 min) and extraction temperature ($10-70^{\circ}C$) were analyzed and optimized extraction condition for antioxidant activity of green tea extract (GTE) was determined using response surface methodology with three-level-three-factor Box-Behnken design (BBD). Regression analysis showed a good fit of data and the optimal conditions of extraction were found to be 57.7% EtOH, 15 min and $70^{\circ}C$. Under this condition, antioxidant activity of experimental data was 88.4% which was almost fit to the ideal value of 88.6%. As epigallocatechin gallate (EGCG) is known for the major ingredient for antioxidant activity of green tea, we investigated the effect of EGCG on antioxidant activity of GTE. EGCG showed antioxidant activity with the $IC_{50}$ value of $4.2{\mu}g/ml$ and a positive correlation was observed between EGCG content and the antioxidant activity of GTE with $R^2=0.7134$. Interestingly, however, GTE with 50 - 70% antioxidant activity contain less than $1.0{\mu}g/ml$ of EGCG, which is much lower than $IC_{50}$ value of EGCG. Therefore, we suppose that EGCG together with other constituents contribute to antioxidant activity of GTE. Taken together, these results suggest that green tea is more beneficial than EGCG alone for antioxidant ability and optimal extraction condition of green tea will be useful for the development of food and pharmaceutical applications

• Preventive Nutrition and Food Science
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• 제22권1호
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• pp.37-44
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• 2017

Sulfur-containing amino acids play important roles in good flavor generation in Maillard reaction of non-enzymatic browning, so aqueous model systems of glucosamine and cysteine were studied to investigate the effects of reaction temperature, initial pH, reaction time, and concentration ratio of glucosamine and cysteine. Response surface methodology was applied to optimize the independent reaction parameters of cysteine and glucosamine in Maillard reaction. Box-Behnken factorial design was used with 30 runs of 16 factorial levels, 8 axial levels and 6 central levels. The degree of Maillard reaction was determined by reading absorption at 425 nm in a spectrophotometer and Hunter's L, a, and b values. ${\Delta}E$ was consequently set as the fifth response factor. In the statistical analyses, determination coefficients ($R^2$) for their absorbance, Hunter's L, a, b values, and ${\Delta}E$ were 0.94, 0.79, 0.73, 0.96, and 0.79, respectively, showing that the absorbance and Hunter's b value were good dependent variables for this model system. The optimum processing parameters were determined to yield glucosamine-cysteine Maillard reaction product with higher absorbance and higher colour change. The optimum estimated absorbance was achieved at the condition of initial pH 8.0, $111^{\circ}C$ reaction temperature, 2.47 h reaction time, and 1.30 concentration ratio. The optimum condition for colour change measured by Hunter's b value was 2.41 h reaction time, $114^{\circ}C$ reaction temperature, initial pH 8.3, and 1.26 concentration ratio. These results can provide the basic information for Maillard reaction of aqueous model system between glucosamine and cysteine.

• Natural Product Sciences
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• 제24권2호
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• pp.103-108
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• 2018

The usage of wild ginseng (Panax ginseng C.A. Meyer) has been limited due to short supply and high price. Therefore, sufficient production as well as efficient extraction of mountain ginseng are required for the development as products. In this study, wild ginseng adventitious root cultures were prepared for efficient production with advantages of fast growth and stable production. Treatment of methyl jasmonate (MJ) to wild ginseng adventitious root cultures increased the extraction yield and antioxidative activity. Further investigation on effect of extraction conditions suggested the importance of ethanol concentration on antioxidative activity and extraction yield of MJ-treated wild ginseng adventitious root cultures. Optimized extraction condition of MJ-treated wild ginseng adventitious root cultures for maximum extraction yield and antioxidative activity was determined using response surface methodology with three-level-three-factor Box-Behnken design (BBD). Extraction of 1 g MJ-treated wild ginseng adventitious root culture with 30 ml of 9% ethanol at $30^{\circ}C$ produced 310.2 mg extract with 71.0% antioxidative activity at $100{\mu}g/ml$. Taken together, MJ-treated wild ginseng adventitious root culture is valuable source for wild ginseng usage and optimized extraction condition can be used for the development of functional products or folk remedies.

• Journal of the Korean Wood Science and Technology
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• 제48권4호
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• pp.548-561
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• 2020

Paecilomyces japonica is widely cultured to produce mycelium for medicinal and health food use. Illumination is an important factor in the growth and production of mycelium in submerged culture. The effects of different light-emitting diode (LED) combinations on the growth and cordycepin content as bioactive substances of mycelium were investigated. The results showed that the mycelium dry weights were lower under dark condition and red LED treatments. Dark condition, fluorescent light, and ultraviolet-A failed to increase the cordycepin content. Blue light was necessary to increase the cordycepin content, and a red-to-blue ratio of 3:7 induced the highest cordycepin content. The cordycepin contents of mycelium in submerged culture were significantly higher in a 12 h/day illumination time under red and blue (red-to-blue ratio of 3:7) LED treatments, showing an increase of up to 38% compared with those under the fluorescent-light control condition. The results demonstrated the roles of light with different wavelengths on the biosynthesis of cordycepin as bioactive substances. The low-heat release and replacement of traditional fluorescent lights with low-energy-consuming LEDs could increase the contents of bioactive substances. After optimization of the cordycepin production using response surface methodology (Box-Behnken design) to its canonical form, the optimum combination was found to be as follows: illumination time = 17.7 h/day, sugar content in the medium = 9.7 g/50 mL, and incubation time = 61.2 h. The model predicted a maximum response of 3779.2 ㎍/mL cordycepin yield.

• 분석과학
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• 제30권1호
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• pp.39-48
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• 2017

Roxithromycin (RXT), which is an antibiotic used to treat respiratory tract and urinary infections, is official in Korean Pharmacopoeia (KP) and is marketed in various dosage forms including tablet, granule, suspension, and tablet for suspension in Korea. This study presents how a universal and reliable method to quantify RXT in bulk drug and formulations was developed. Effects of factors including column type, buffer concentration, type and concentration of organic solvent, buffer pH, and type and concentration of mobile phase additive, were examined, and some categorical or crucial factors including the types of column, organic solvent, mobile phase additive and the buffer pH were optimized by one-factor-at-a-time approach. Subsequently, concentrations of the buffer and additive and column temperature were optimized by response surface methodology using Box-Behnken design aiming to acquire the RXT peak of good shape. The optimized method employed a Phenomenex Gemini $5{\mu}$ C18 110A ($150{\times}4.60mm$, $5{\mu}m$) maintained at $30^{\circ}C$ with the mobile phase consisting of 25 mM phosphate buffer (pH 6.0) with 0.3 % tetrabutylammonium hydroxide and methanol at a ratio of 37:63 (v/v). Method validation results showed that the developed method was linear, precise, and accurate. Compared to the compendial methods in KP 10 that exhibited a significant tailing of the RXT peak despite using unfavorably high buffer concentrations and were not harmonized among bulk drug and formulations, this method could be universally applied to RXT bulk drug and marketed products in various dosage forms and thus was adopted in KP 11.

• Journal of Microbiology and Biotechnology
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• 제26권8호
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• pp.1428-1438
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• 2016

In the present work, Arthrobacter sp. 25, a lead-tolerant bacterium, was assayed to remove lead(II) from aqueous solution. The biosorption process was optimized by response surface methodology (RSM) based on the Box-Behnken design. The relationships between dependent and independent variables were quantitatively determined by second-order polynomial equation and 3D response surface plots. The biosorption mechanism was explored by characterization of the biosorbent before and after biosorption using atomic force microscopy (AFM), scanning electron microscopy, energy dispersive X-ray spectroscopy, X-ray diffraction, and Fourier transform infrared spectroscopy. The results showed that the maximum adsorption capacity of 9.6 mg/g was obtained at the initial lead ion concentration of 108.79 mg/l, pH value of 5.75, and biosorbent dosage of 9.9 g/l (fresh weight), which was close to the theoretically expected value of 9.88 mg/g. Arthrobacter sp. 25 is an ellipsoidal-shaped bacterium covered with extracellular polymeric substances. The biosorption mechanism involved physical adsorption and microprecipitation as well as ion exchange, and functional groups such as phosphoryl, hydroxyl, amino, amide, carbonyl, and phosphate groups played vital roles in adsorption. The results indicate that Arthrobacter sp. 25 may be potentially used as a biosorbent for low-concentration lead(II) removal from wastewater.

• 한국미생물·생명공학회지
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• 제49권2호
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• pp.181-191
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• 2021

The present study addresses isolation, optimization, partial purification, and characterization of a haloalkaline serine protease from a newly isolated haloarchaeal strain isolated from Wadi El Natrun in Egypt. We expected that a two-step sequential statistical approach (one variable at a time, followed by response surface methodology) might maximize the production of the haloalkaline serine protease. The enzyme was partially purified using Hiprep 16/60 sephacryl S-100 HR gel filtration column. Molecular identification revealed the newly isolated haloarchaeon to be Natrialba hulunbeirensis strain WNHS14. Among several tested physicochemical determinants, casamino acids, KCl, and NaCl showed the most significant effects on enzyme production as determined from results of the One-Variable-At-A-time (OVAT) study. The BoxBehnken design localized the optimal levels of the three key determinants; casamino acids, KCl, and NaCl to be 0.5% (w/v), 0.02% (w/v), and 15% (w/v), respectively, obtaining 62.9 U/ml as the maximal amount of protease produced after treatment at 40℃, and pH 9 for 9 days with 6-fold enhancement in yield. The enzyme was partially purified after size exclusion chromatography with specific activity, purification fold, and yield of 1282.63 U/mg, 8.9, and 23%, respectively. The enzyme showed its maximal activity at pH, temperature, and NaCl concentration optima of 10, 75℃, and 2 M, respectively. Phenylmethylsulfonyl fluoride (PMSF, 5 mM) completely inhibited enzyme activity.