• Microbiology and Biotechnology Letters
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• v.28 no.1
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• pp.33-38
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• 2000

In order to produce ascorbic acid-2-phosphate from ascorbic acid, bacteria capable of transforming ascorbic acid to ascorbic acid-2-phosphate were isolated from soils and the stock cultures in our laboratory. Among them, a newly isolated bacterium LSH-3 having the best ability of producing ascorbic acid-2-phosphate was selected and partially identified as Pseudomonas sp. The optimum conditions for the production of ascorbic acid-2-phosphate from ascorbic acid and using its resting cells as the source os enzyme were investigated. The results were summarized as follows: The optimum cultivation time and the cell weight for the production of ascorbic acid-2-phosphate was 14 hours and 100g/I(wet weight), respectively. And 0.1%(v/v) Trition X-100 was the most effective surfactant. The optimum concentrations of ascorbic acid and pyrophosphate were 400mM and 500mM, respectively, which led to produce 14.54g/I of ascorbic acid-2-phosphate. The most effective buffer was 50mM sodium acetate. The optimum pH and temperature were 4.5 and $40^{\circ}C$, respectively. Under the above conditions, 17.71 g/I of ascorbic acid-2-phosphate was produced from ascorbic acid after 32 hour-incubation, which corresponded to 17.5% of conversion rate based on ascorbic acid.

• Microbiology and Biotechnology Letters
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• v.25 no.3
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• pp.271-276
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• 1997

An ascorbic acid phosphorylating enzyme, which catalyzes the formation of ascorbic acid-2-phosphate from ascorbic acid and pyrophosphate, was purified 32.7-folds to homogeneity from a cell-free extract of Cellulomonas sp. AP-7. The combination of DEAE- Sephacel ion exchange chromatography and Sephacryl S-200 get filtration was used for their purification. The molecular weight of the native protein was estimated to be 96.lkDa on high performance gel filtration chromatography. The SDS-PAGE analysis indicated that the protein consisted of four identical subunits of 24.6 kDa. The purified enzyme showed the optimal tempeature of 40$\circ$C and optimal pH of 4.5. The Km for ascorbic acid and pyrophosphate were 119 mM and 11.9 mM, respectively. The addition of 5,5'-dithiobis-(2-nitrobenzoic acid) into the reaction mixture resulted in the reduction of the enzyme activity at 51%. The enzyme also had a phosphatase activity at weakly acidic pH and the Km for ascorbic acid-2-phosphate in phosphatase activity was 7.9 mM.

• Microbiology and Biotechnology Letters
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• v.24 no.5
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• pp.613-618
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• 1996

Microorganisms capable of producing ascorbic acid-2-phosphate (AsA2P) from ascorbic acid (AsA) and pyrophosphate (PPi) were screened from the culture collection of this laboratory. Among them, Cellulomonas sp. AP-7 showed the highest productivity of AsA2P. The optimal conditions for the production of AsA2P from AsA and PPi with cell-free extract as an enzyme source were investigated. The reaction mixture for the maximal production of AsA2P consisted of 21 g protein of cell-free extract per liter as the enzyme source, 250 mM AsA, 200 mM sodium pyrophosphate, 150 mM sodium acetate buffer (pH 4.5). By using this reaction mixture, 31.9 mM of AsA2P, which corresponded to a 12.76% yield based on AsA, was produced after incubation of 48 hr at 33$\circ$C.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.13 no.1
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• pp.29-35
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• 1987

Purely synthesized L-ascorbic acid 2 phosphate Mg salt (1 AsA PMg) improved the weak point of ascorbic acid which is easily decomposed in water solution. This compound is hydrolyzed with phosphatase of skin to corresponding ascorbic acid giving Vitamine C activities. The buffer solution of potassium acetate 0.5% and citric acid 0.005% and the sodium sulfite respectively showed good stabilizing effect of the AsA PMg solution. Compared to the other ascorbic acid derivatives the good solubility of AsA PMg gives broad application to cosmetic field.

• Journal of Microbiology and Biotechnology
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• v.17 no.5
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• pp.769-773
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• 2007

With the aim to produce ascorbic acid-2-phosphate(AsA-2-P) from L-ascorbic acid(AsA, Vitamin C), nine bacteria conferring the ability to transform AsA to AsA-2-P were isolated from soil samples alongside known strains from culture collections. Most isolates were classified to the genus Brevundimonas by 16S phylogenetic analysis. Among them, Brevundimonas diminuta KACC 10306 was selected as the experimental strain because of its the highest productivity of AsA-2-P. The optimum set of conditions for the AsA-2-P production from AsA using resting cells as the source of the enzyme was also investigated. The optimum cultivation time was 16 h and the cell concentration was 120g/l(wet weight). The optimum concentrations of AsA and pyrophosphate were 550mM and 450mM, respectively. The most effective buffer was 50mM sodium formate. The optimum pH was 4.5 and temperature was $40^{\circ}C$. Under the above conditions, 27.5g/l of AsA-2-P was produced from AsA after 36 h of incubation, which corresponded to a 19.7% conversion efficiency based on the initial concentration of AsA.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.23 no.2
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• pp.97-117
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• 1997

L-Ascorbic acid 및 그 유도체는 항산화 작용과 미백작용 및 섬유아세포의 생장 촉진과 콜라젠 생합성의 증가시키는 효과가 있으므로 화장품에서 오래 전부터 사용되어 왔다. 본 연구에서는 생체에 대한 안전성과 안정성이 우수한 L-Ascorbic acid 유도체를 개발하기 위하여 인지질과 유사하게 L-Ascorbic acid 에 3-Aminopropane phosphoric acid를 결합하여 L-Ascorbic acid-3-aminopropane phosphoric acid diester를 합성하였다. ASA-APPA 는 2-Chlorotetrahydro-2H-1, 3, 2-oxazaphosphorine P-oxide 와 5, 6-isopropylidene L-Ascorbic acid 를 반응 시킨 후 산 사수분해 반응을 통하여 얻을 수 있었다. ASA-APPA는 수용액에서 안정성이 우수하였으며, 독성실험에서 무독성 물질이며, 인체 첩포 실험에서도 무자극 물질임을 확인하였다. 그리고, ASA-APPA 는 L-Ascorbic acid 와 3-APPA 의 혼합물과 거의 유사한 섬유아세포의 증식효과를 나타내었으며, melanoma 에 대한 멜라닌 생성 억제 실험에서 L-Ascorbic acid phosphate magnesium salt 와 유사한 효과를 보였다. 따라서 ASA-APPA 는 멜라닌 생성 억제 효과와 섬유아세포의 증식 효과를 갖는 새로운 화장품 원료로서 사용이 가능할 것으로 생각된다.

• Applied Biological Chemistry
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• v.36 no.2
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• pp.86-92
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• 1993

Myrosinase from radish was purified by DEAE Bio-Gel, Con-A, and Superose-6 column. The purified myrosinase(II) possessed 2 subunits, and their molecular as determined by SDS-polyacrylamide gel electrophoresis were 53 and 39 KD, respectively. The specific activity of purified enzyme was 37,500 units/mg. The enzyme was purified approximately 44-fold compared to the crude enzyme. Optimum pH of the myrosinase was $6.5{\sim}7.0$ in phosphate and Tris-HCl buffer solutions. Optimum temperature of the enzyme was $37{\sim}38^{\circ}C$. The enzyme was stable at pH 7.0, and less than $30^{\circ}C$. Cu or Hg ion significantly inhibited the enzyme activity, but ascorbic acid enhanced, resulting in a maximum activity by 1 mM ascorbic acid. Among the ascorbic acid analogues, dehydroascorbic acid did not affect, whereas others showed a little effect, but less than ascorbic acid itself. Individual 2-mercaptoethanol and dithiothreitol (reducing agents) did not enhance the enzyme activity. but 2-mercaptoethanol effect was enhanced when mixed with ascorbic acid.

• Journal of the Korean Society of Food Science and Nutrition
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• v.24 no.4
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• pp.563-569
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• 1995

Myrosinase from Korean cabbage(Bogdoli) was purified and its enzymatic properties were investigated. Myrosinase from the Korean cabbage was purified by DEAE Bio-Gel Sepharose, Concanavalin-A, and Mono-Q column chromatography and exhibited a 55KD molecular weight with a single band on the gel of SDS-PAGE. The enzyme was purified about 21-fold compared to its crude enzyme and a specific activity of purified enzyme was 15, 120units/mg. Optimum pH of the myrosinase was 7.0 in both phosphate and Tris-HCl buffer solutions, the enzyme was stable at pH 6.5~7.0. Optimum temperature of enzyme was 37~38$^{\circ}C$. The enzyme activity was significantly inhibited by Cu2+ and Hg2+, but enhanced by ascorbic acid, resulting in a maximum activity at 1mM ascorbic acid. Among the ascorbic acid analogues, dehydro-ascorbic acid did not affect, whereas others showed a little effect on the enzyme activity, but less than ascorbic acid itself. Reducing agents such as 2-mercaptoethanol and dithiothreitol had no effect on the enzyme activity, but the enzyme activity was enhanced when 2-mercaptoethanol was mixed with ascorbic acid.

• Proceedings of the SCSK Conference
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• 2003.09b
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• pp.479-479
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• 2003

We investigated the tyrosinase inhibitory effect using whitening materials such as arbutin, ethyl ascorbyl ether, glabridin, kojic acid, magnesium ascorbyl phosphate and ascorbic acid. Tyrosinase inhibition rate were determined varying the enzyme concentration, reaction time, reaction temperature and pH. The optimal conditions to measure the inhibitory efficacy were as follows. : enzyme concentration 1,500 or 2,000IU/mL, reaction time 15min(for the enzyme concentration 1,500 IU/mL) and l0min(for the enzyme concentration 2,000IU/mL), reation temperature 42$^{\circ}C$, pH 6.5. Under these conditions $IC_{50}$/ of arbutin, ethyl ascorbyl ether, glabridin, kojic acid, magnesium ascorbyl phosphate and ascorbic acid were calculated. In the case of magnesium ascorbyl phosphate, the inhibitory effect of tyrosinase was very low and the $IC_{50}$/ of magnesium ascorbyl phosphate could not be calculated. Other five materials showed good inhibitory effect of tyrosinase and can be used for the whitening materials.

• Journal of the Korean Society of Food Science and Nutrition
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• v.25 no.4
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• pp.687-694
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• 1996

Myrosinase was purified from Korean mustard seed(Brassica juncea) by a sequential process of DEAE-cellulose, concanavalin A-sepharose, and Superose 6 chromatography. The molecular weight of puri-fied myrosinase(II-2) determined by SDS-polyacrylamide electrophoresis was 67KD. About a 248-fold purification for myrosinase II-2 was obtained after Superose 6 chromatography. Optimum pH of the myrosinase was 7.0 and optimum temperature of the enzyme was $3^{\circ}C.$ The enzyme was stable at pH 7.0, and below $30^{\circ}C.$ Cu, Hg and Fe ion significantly inhibited the enzyme activity, but ascorbic acid enhanced, resulting in a maximum activity by 1mM ascorbic acid. Among tile ascorbic acid ana-logues, dehydroascorbic acid inhibited the enzyme activity, whereas others showed a little effect. Reducing agents such as 2-mercaptoethanol and dithiothreitol inhibited the enzyme activity, but the reducing agents with ascorbic acid was enhanced enzyme activity.