사과酒 醱酵中 褐變抑制와 Ployphenol Oxidase 電氣泳動像의 變化에 關하여

朴哲民,鄭基澤,方光雄 慶北大學校 1986 農業科學技術硏究所報 Vol.3 No.-

사과酒의 酵素에 依한 褐(변)의 基礎的 資料를 얻고자 polyphenol oxidase를 抽出하여 그 性質을 살펴보았으며, 사과酒 醱酵中 一般成分과 PPO 電氣泳動像의 (변)化 및 褐(변)抑制에 對해 살펴본 結果, 國光 PPO의 最適pH는 6.0, 最適溫度는 30℃였으며, 酵素의 活性은 70℃에서 1時間의 熱處理로 5%만이 남아있었다. 本 酵素의 主 基質은 o-diphenol이었으며, 이들中 chlorogenic acid가 最高活性을 나타내었다. 沮害劑로는 Na-metabisulfite, ascorbic acid, cystein이 良好하였으며, PPO活性을 完全히 沮害하는 濃度는 모두 5mM이었다. 사과酒 醱酵中 total phenol의 減少率은 SO₂處理에 依해 현저히 鈍化되었고, SO₂含量은 free와 tatal의 경우 모두 醱酵初期에 增加하다가 減少하기 경향을 나타내었다. 國光 PPO의 活性band는 總 8個였으며, 70℃에서 1時間동안 熱處理한 區와 하지 않은 區의 경우, 各各 2個와 6個의 band가 醱酵 終了時까지 남아있었으므로 이들이 醱酵中 酵素的褐변에 關與할 것으로 생각되어진다. In order to obtain preliminary data on enzymatic browning and of apple wine, prevention of browning some properties of polyphenol oxidase from apple(Ralls Janet) were investigated on electrophoretic patterns of polyphenol oxidase, and some factors related to browning of apple wine during fermentation. Optimum pH and temperature for the reaction of polyphenol oxidase were pH 6.0 and 30℃, respectively. The residual activity of the enzyme was only 5% after heating at 70℃ for 1 hr. The main substrates of the enzyme were o-diphenolic compounds, and chlorogenic acid of them showed the highest activity. Na-metabisulfite, ascorbic acid and cysteine were effective inhibitors and the concentration for complete inhibition of the enzyme was 5mM. Total phenol contents during fermentation decreased slowly by addition of SO₂in apple wine. The contents of free and total SO₂increased for first 2 days and decreased after 3 days during culture. The number of active bands of polyphenol oxidase from apple (Ralls Janet) was found to be eight, and two of them after heating at 70℃ for 1 hr and six of them in control remained active to the end of fermentation. These remaining bands seem to affect the enzymatic browning of apple wine during fermentation.

Killer 酵母 融合株 FWKS 260의 醱酵 特性 및 Killer Toxin의 精製에 關한 硏究

方光雄 慶北專門大學 1992 慶北專門大學 論文集 Vol.11 No.-

自然界에서 分離한 Killer 酵母의 同定 및 Killer Plasmid의 檢索

方光雄 慶北專門大學 1993 慶北專門大學 論文集 Vol.12 No.-

Ten strains out of about 1,000 yeast strains isolated from byproducts of alcoholic industries, milk products, fruits, greens, food-related industries and soils of nature, revealed the killer activities. Two strains which have excellent killer activities among them were isolated and identified so Saccharomyces cerevisiae B 15-1 and Hansenula anomala Y 33 by investigation of the morphological, cultural and physiological characteristics. The optimal conditions on these strains for the production of killer toxin were investigated. The strain B 15-1 showed the highest killer toxin activities when it was cultured up to the log phase of 48 hrs in YPD medium (pH 4.7) at 25℃. On the other hand, the strain Y 33 revealed the highest activities when it was cultured up to the stationary phase of 60 hrs in YPD medium (pH 4.0) at 20℃. The sensitive strain Kyokai 7 was found to be killed entirely by the killer toxin produced from the wild killer yeast B 15-1 when B 15-1 was co-cultured with the same cell concentration (10^(6)cells/ml) of Kyokai 7 after cultivation of 36 hrs, and with large concentration (9×10^(7) cells/ml) after 48 hrs. The dsRNA plasmids of the wild killer yeast B 15-1, Y 33, and the sensitive Kyokai 7 were extracted and then were observed by the agarose gel electrophoresis. The sensitive strain Kyokai 7 was found to have neither L nor M dsRNA plasmid. However, B 15-1 and Y 33 had both L and M dsRNA plasmid with 4.7 kb and 2.5 kb, respectively.

Biosensor의 原理와 食品工業的 應用

方光雄 慶北專門大學 1995 慶北專門大學 論文集 Vol.14 No.-

冷凍반죽을 利用한 醱酵빵 製造에 있어서 適正반죽의 組成

方光雄,鄭基澤,徐錫出,宋亨翼 慶北大學校 農業科學技術硏究所 1988 慶北大農學誌 Vol.6 No.-

Straight no-time method로 조제한 冷凍반죽을 利用한 醱酵빵제조에 있어서 가장 적절한 반죽의 組成을 製빵成積을 중심으로 검토하였다. 硬質밀가루 1,000g에 대하여 壓搾酵母 30g, 설탕 50g, 食鹽20g, 쇼트닝 40g, 브롬산칼륨 75mg, L-ascorbic acid 200mg, yeast food 3g, vital wheat gluten 30g, 제1인산칼슘 400mg, 스테아릴젖산나트륨 8g, 汲水量 680g 으로 冷凍반죽을 만드는 것이 製빵過程에서 膨脹力과 醱酵時間이 적절하고 높은 比容積을 얻을 수 있어서 가장 바람직하였다. Farinograph 성적상의 吸水率보다 높은 68%의 汲水量으로도 乳化劑나 品質改良劑의 多量添加로 반죽의 凍結障害를 억제할 수 있었다. We studied suitable dough formula for yeast-raised breadmaking using frozen dough prepared by straight no-time method, centering around breadmaking quality. The most suitable dough formula based on 1,000g of wheat flours was as follows: compressed yeast; 30g, sucrose; 50g, salt; 20g, shortening; 40g, potassium bromate; 75mg, L-ascorbic acid; 200mg, yeast food; 3g, vital wheat gluten; 30g, calcium phosphate, monobasic; 400mg, sodium stearoyl-2-lactylate; 8g, water; 680g. Breadmaking test employing this formula showed that gassing power and fermentation time were suitable and higher specific loaf volume was obtainable. By using much emulsifiers and dough conditioners, frozen injury of dough was controlable in spite of the addition of more content of water(68%) than that of water (62%) obtained from the farinograph data.

Killer 효모 융합주 FWKS 260 이 분비하는 Killer Toxin 의 정제

정기택,방광웅,우철주,정용진,김재근,송형익 한국미생물학회 1992 미생물학회지 Vol.30 No.3

원형질체 융합을 통하여 육성한 killer 효모 융합주 FWKS 260 의 killer toxin 을 ammonium sulfate fractionation, Amicon PM 10 concentration, Sephadex G-200 및 Sephadex G-75 column chromatography 를 행하여 정제한 결과 단일 단백질 band 를 보여 순수하게 정제되었음을 알 수 있었고, 단백질 분해효소를 처리한 결과 killer 활성이 소실되어 killer toxin 의 단백질 부분이 killer 활성을 나타냄을 알 수 있었다. 그리고 이 toxin 은 20.deg.C 에서는 거의 안정하였으나, 온도가 증가함에 따라 점차 활성이 소실되었고, pH 2.0-5.0 에서 비교적 안정하였다. 한편, SDS-polyacrylamide gel electrophoresis 결과 분자량은 약 13.000 임을 알 수 있었고, SDS polyacrylamide gel electrophoresis 를 행한 후 Schiffs reagent 로 염색한 결과 붉은 단일 band 를 보여 정제된 killer toxin 은 glycoprotein 임을 확인 할 수 있었다. Killer toxin from killer yeast fusant FWKS 260 developed by protoplast fusion between the wild killer yeast and alcohol-fermenting yeast was purified by ammonium sulfate fractionation. Amicon PM I0 concentration. Sephadex G-200 and Scphadcx G-75 column chromatography. The purified killer toxin showed a single band by SIX-polyacvlamide gel electrophoresis. The protein part of killer toxin was active site. which was found by treating the proteolytic enzyme such as pronase E and pepsin to killer toxin. The killer toxin was stable at pH 2.0-5.0 and 20$^{\circ}$C. but inactivated with increasing temperature. The molecular weight was determined to be approximately 13.000 according to the results obtained from the SDS-polyacrylamide gel electrophoresis. It was confirmed that the purified killer toxin is glycoprotein by showing a red single band after st'tining with Schiffs reagent.

Killer 효모와 알콜 발효효모간의 원형질체 융합주의 특성

정기택,방광웅,김재근,송형익,정용진 한국미생물학회 1990 미생물학회지 Vol.28 No.1

원형질체 융합을 통하여 killer 효모의 유전형질을 기존의 ethanol 발효효묘에 도입하므로서 야생의 killer 효모에 저항성을 가지고, 오염효모를 치사시킬 수 있으며, ethanol 발효능도 유지하는 새로운 효모균주를 개발하였다. 먼저 융합주의 생리적인 특성을 검토한 바, 융합주는 양천주에 비하여 세 적이 크고, DNA 함량이 높았으며, 증식도는 친주와 유사한 경향이었다. 또 융합주를 최소배지에서 보존하는 것이 안정성을 높이는 방법이었고, 7일 간격으로 7회 계대배양하므로서 유전적으로 안정화시킨 융합주를 6개월 간 계속 사용한 결과 segregant가 전혀 나타나지 않았으므로 매우 안정하였다. 또한 융합주는 핵 및 포자를 형성함을 관찰 할 수 있었고, TTC 정색반응에서 적색 및 pink 색을 띄었으며, 탄소원의 자화성 및 발효능은 천주와 유사하였는데, KCI, NaCl, sodium propionate alc cycloheximide 등에 대한 내성도 양친주의 한쪽을 따랐다. 그리고, 융합주를 20% glucose와 sucrose에서 72시간 및 60시간 배양했을 때, FWKS 260의 경우 각각 9.6v/v% 및 9.8v/v%의 ethanol을 생성하였고, 감수성주와 혼합배양한 결과 FWKS260의 경우 48시간 이후에는 감수성주를 거의 발견할 수 없었으며, 전주 및 융합주의 dsRNA plasmid를 추출하여 전기 2.5kb의 L 및 M dsRNA plasmidsms는 서로 연관성이 있으며, killer toxin 분비 및 저항성을 나타내는 유전자를 지배하는 것은 M dsRNA plasmid임을 확인할 수 있었다. 수 있었다. Cell volume and DNA contents of the fusants were similar to those of parents. Genetic stability of the fusants was increased when they were cultured on minimal medium (MM) rather than on complete medium (CM), and the fusants were stabilized by subculturing 7 generations each 7 day on MM agar. The finally selected fusants after being cultured for 6 months on CM were stable without segregation. The fusants could also form nuclein and ascospores, and show red and pink colors by the test of TTC colorization. Assimilability and fermentability of carbon sources of the fusants were similarto those of parents. The tolerance of KCl, NaCl, sodium propionate and cycloheximide showed the traits of one strain of parents. When the fusants were cultured for 72 hr and 60 hr in the medium containing 20% glucose and sucrose, respectively, the yield of ethanol for FWKS 260 was reached to 9.6 v/v% and 9.8v/v%, respectively. The sensitive strain Kyokai 7 was found to be killed entirely after cultivation of 48 hr by the killer toxin from the fusants. The recipient S 29 and Kyokai 7 were found to have neither L nor M dsRNA plasmid. However, K 52 and fusants had both L and M dsRNA plasmid of 4.7 kb and 2.5 kb, respectively. The curants treated by heat and cycloheximide did not contain M dsRNA plasmid, but had large amounts of L dsRNA plasmid of those of killer yeasts.

건조누에를 이용한 동충하초의 인공자실체 형성

이준우,방광웅,서부일,이은숙 대한본초학회 2004 大韓本草學會誌 Vol.19 No.4

Objectives : This study was carried out investigate cultural conditions for artificial fruiting body production of Paecilomyces japonica. Methods : The formation period of an artificial fruiting body of Paecilomyces japonica treated with dried-silkworm, silkworm pupa and naked barley were 30 days and 35 days, respectively. Results : The length and diameter of fruiting body were longer and thicker in media of the naked barley than that of dried-silkworm. The optimum illumination was 200 lux for the fruiting body formation. Conclusion : The growth of fruiting body was not affected by three subsequent transculture, and the optimum inoculum size was six percent(w/v).

.betha.-1, 3-glucanase 생성균의 분리 및 효소 생성 조건

정기택,방광웅,송형익,김재근,유대식 한국미생물학회 1986 미생물학회지 Vol.24 No.3

The bacteria, which were capable of producing ${\beta}-1$, 3-glucanase inducibly by utilizing cell wall of Aspergillus fumigatus as a sole carbon source, were isolated from soil in the campus of Kyungpook National University. Among them, the strain which produced the enzyme excellently was selected and identified to be Pseudomonas stutzeri KF 13 by morphological, cultural and physiological examination. The optimal conditions for the enzyme production from Pseudomonas stutzeri KF 13 were investigated. the enzyme production was reached maximum state shen the broth cultured for 72hr at $30^{\circ}C$. And the enzyme showed the highest activity in the medium containing 3.5% cell wall as an inducer, 15% yeast autolysate as a nitrogen source and 0.05% $MnSO_4$ at pH 7.5.

Chitinase를 생산하는 Pseudomonas sp. YS-542의 분리 및 특성

강신욱,이준우,방광웅,최희상 慶北專門大學(영주경상전문대학) 2002 慶北專門大學 論文集 Vol.21 No.-

A chitinase-producing bacterium was isolated from soil by selective enrichmentment culture medium. The bacterium was identified as Pseudomonas sp. based on the data obtained from its morphological, cultural and biochemical characteristics. The isolated strain was named Pseudomonas sp. YS-542. The medium for the production of chitinase was consisted of colloidal chitin 1.0%, tryptone 1.0%, MgS04·7H2O 0.2%, K2HP04 0.1%. The optimum cultural temperature, initial pH and time for the best production of chitinase were 25℃, 48hrs and pH 8.0, respectively.