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Development of Natural Antioxidants and Whitening Agents for Cosmeceuticals

  • Kim, Jong-Pyung
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 2007.11a
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    • pp.79-92
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    • 2007
  • Oxidative stress have known to be a risk factor for the degenerative processes and closely related to a lot of diseases. It is well established that antioxidants are good in protection and therapeutic means against oxidative damage. There is increasing interest in natural antioxidants and many natural antioxidants have been found and utilized as the possible protection for various diseases and skin aging. We have screened natural antioxidant agents for cosmeceuticals, nutraceuticals, and drugs as therapeutic and preventive means against oxidative stress, and have developed a number of novel antioxidants from various natural sources. A novel melanin synthesis inhibitor, Melanocin A, isolated from the metabolite of a fungal strain Eupenicillium shearii F80695 inhibited mushroom tyrosinase and melanin biosynthesis of B16 melanoma cells with $IC_{50}$ value of 9.0 nM and MIC value of $0.9\;{\mu}M$, respectively. Melanocin A also exhibited potent antioxidant activity by scavenging of DPPH and superoxide anion radicals. UV was found to increase the level of hydrogen peroxides and other reactive oxygen species (ROS) in skin tissues. This increase in ROS may not only alter the structure and function of many genes and proteins directly but may also modulate their expressions through signal transduction pathways and, ultimately, lead to skin damage. We investigated the effect of Melanocin A on UV-induced premature skin aging. Firstly, the effect of Melanocin A on UV-induced matrix metalloproteinase (MMP)-9 expression in an immortalized human keratinocyte cell line, HaCaT in vitro was investigated. Acute UV irradiation induced MMP-9 expression at both the mRNA and protein levels and Melanocin A suppressed this expression in a dose-dependent manner. We then investigated UV-induced skin changes in hairless mice in vivo by Melanocin A. Chronic exposure of hairless mouse dorsal skin to UV increased skin thickness and induced wrinkle formation and the gelatinase activities of MMP-2 and MMP-9. Moreover, Melanocin A significantly suppressed UV-induced morphologic skin changes and MMP-2 and MMP-9 expression. These results show that Melanocin A can prevent the harmful effects of UV that lead to skin aging. Therefore, we suggest that Melanocin A should be viewed as a potential therapeutic agent for preventing and/or treating premature skin aging. Terrein is a bioactive fungal metabolite isolated from Penicillium species. Terrein has a relatively simple structure and can be easily synthesized. However, the biologic effects of terrein are comparatively unknown. We found for the first time that terrein potently inhibit melanin production in melanocytes and has a strong hypopigmentary effect in a spontaneously immortalized mouse melanocyte cell line, Mel-Ab. Treatment of Mel-Ab cells with terrein (10-100 mM) for 4 days significantly reduced melanin levels in a dose-dependent manner. In addition, terrein at the same concentration also reduced tyrosinase activity. We then investigated whether terrein influences the extracellular signal-regulated protein kinase (ERK) pathway and the expression of microphthalmia-associated transcription factor (MITF), which is required for tyrosinase expression. Terrein was found to induce sustained ERK activation and MITF down-regulation, and luciferase assays showed that terrein inhibits MITF promoter activity in a dose-dependent manner. To elucidate the correlation between ERK pathway activation and a decreased MITF transcriptional level, PD98059, a specific inhibitor of the ERK pathway, was applied before terrain treatment and found to abrogate the terrein-induced MITF attenuation. Terrein also reduced the tyrosinase protein level for at least 72 h. These results suggest that terrain reduces melanin synthesis by reducing tyrosinase production via ERK activation, and that this is followed by MITF down-regulation.

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Seasonal color change of the oxyhydrous precipitates in the Taebaek coal mine drainage, south Korea, and implications for mineralogical and geochemical controls

  • Kim, J. J.;C. O. Choo;Kim, S. J.;K. Tazaki
    • Proceedings of the Mineralogical Society of Korea Conference
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    • 2001.06a
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    • pp.38-39
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    • 2001
  • The seasonal changes in pH, Fe, Al and SO$_4$$\^$2-/ contents of acid drainage released from coal mine dumps play a major role in precipitation of metal hydroxides in the Taebaek coal field area, southeastern Korea. Precipitates in the creeks underwent a cycle of the color change showing white, reddish brown and brownish yellow, which depends on geochemical factors of the creek waters. White precipitates consist of Al-sulfate (basaluminite and hydrobasaluminite) and reddish brown ones are composed of ferrihydrite and brownish yellow ones are of schwertmannite. Goethite coprecipitates with ferrihydrite and schwertmannite. Ferrihydrite formed at higher values than pH 5.3 and schwertmannite precipitated below pH 4.3, and goethite formed at the intermediate pH range between the two minerals. With the pH being increased from acid to intermediate regions, Fe is present both as schwertmannite and goethite. From the present observation, the most favorable pH that basauluminte can precipitate is in the range of pH 4.45-5.95. SEM examination of precipitates at stream bottom shows that they basically consist of agglomerates of spheroid and rod-shape bacteria. Bacteria species are remarkably different among bottom precipitates and, to a less extent, there are slightly different chemical compositions even within the same bacteria. The speciation and calculation of the mineral saturation index were made using MINTEQA2. In waters associated with yellowish brown precipitates mainly composed of schwertmannite, So$_4$ species is mostly free So$_4$$\^$2-/ ion with less AlSo$_4$$\^$+/, CaSo$\sub$(aq)/, and MgSo$\sub$4(aq)/. Ferrous iron is present mostly as free Fe$\^$2+/, and FeSo$\sub$4(aq)/ and ferric iron exists predominantly as Fe(OH)$_2$$\^$+/, with less FeSo$\sub$4(aq)/, Fe(OH)$_2$$\^$-/, FeSo$_4$$\^$-/ and Fe$\^$3+/, respectively Al exists as free Al$\^$3+/, AlOH$_2$$\^$-/, (AlSo$_4$)$\^$+/, and Al(So$_4$)$\^$2-/. Fe is generally saturated with respect to hematite, magnetite, and goethite, with nearly saturation with lepidocrocite. Aluminum and sulfate are supersaturated with respect to predominant alunite and less jubanite, and they approach a saturation state with respect to diaspore, gibbsite, boehmite and gypsum. In the case of waters associated with whitish precipitates mainly composed of basaluminite, Al is present as predominant Al$\^$3+/ and Al(SO$_4$)$\^$+/, with less Al(OH)$\^$2+/, Al(OH)$_2$$\^$+/ and Al(SO$_4$)$\^$2-/. According to calculation for the mineral saturation, aluminum and sulfate are greatly supersaturated with respect to basaluminite and alunite. Diaspore is flirty well supersaturated while jubanite, gibbsite, and boehmite are already supersaturated, and gypsum approaches its saturation state. The observation that the only mineral phase we can easily detect in the whitish precipitate is basaluminite suggests that growth rate of alunite is much slower than that of basaluminite. Neutralization of acid mine drainage due to the dilution caused by the dilution effect due to mixing of unpolluted waters prevails over the buffering effect by the dissolution of carbonate or aluminosilicates. The main factors to affect color change are variations in aqueous geochemistry, which are controlled by dilution effect due to rainfall, water mixng from adjacent creeks, and the extent to which water-rock interaction takes place with seasons. pH, Fe, Al and SO$_4$ contents of the creek water are the most important factors leading to color changes in the precipitates. A geochemical cycle showing color variations in the precipitates provides the potential control on acid mine drainage and can be applied as a reclamation tool in a temperate region with four seasons.

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Shielding for Critical Organs and Radiation Exposure Dose Distribution in Patients with High Energy Radiotherapy (고 에너지 방사선치료에서 환자의 피폭선량 분포와 생식선의 차폐)

  • Chu, Sung-Sil;Suh, Chang-Ok;Kim, Gwi-Eon
    • Journal of Radiation Protection and Research
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    • v.27 no.1
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    • pp.1-10
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    • 2002
  • High energy photon beams from medical linear accelerators produce large scattered radiation by various components of the treatment head, collimator and walls or objects in the treatment room including the patient. These scattered radiation do not provide therapeutic dose and are considered a hazard from the radiation safety perspective. Scattered dose of therapeutic high energy radiation beams are contributed significant unwanted dose to the patient. ICRP take the position that a dose of 500mGy may cause abortion at any stage of pregnancy and that radiation detriment to the fetus includes risk of mental retardation with a possible threshold in the dose response relationship around 100 mGy for the gestational period. The ICRP principle of as low as reasonably achievable (ALARA) was recommended for protection of occupation upon the linear no-threshold dose response hypothesis for cancer induction. We suggest this ALARA principle be applied to the fetus and testicle in therapeutic treatment. Radiation dose outside a photon treatment filed is mostly due to scattered photons. This scattered dose is a function of the distance from the beam edge, treatment geometry, primary photon energy, and depth in the patient. The need for effective shielding of the fetus and testicle is reinforced when young patients ate treated with external beam radiation therapy and then shielding designed to reduce the scattered photon dose to normal organs have to considered. Irradiation was performed in phantom using high energy photon beams produced by a Varian 2100C/D medical linear accelerator (Varian Oncology Systems, Palo Alto, CA) located at the Yonsei Cancer Center. The composite phantom used was comprised of a commercially available anthropomorphic Rando phantom (Phantom Laboratory Inc., Salem, YN) and a rectangular solid polystyrene phantom of dimensions $30cm{\times}30cm{\times}20cm$. the anthropomorphic Rando phantom represents an average man made from tissue equivalent materials that is transected into transverse 36 slices of 2.5cm thickness. Photon dose was measured using a Capintec PR-06C ionization chamber with Capintec 192 electrometer (Capintec Inc., Ramsey, NJ), TLD( VICTOREEN 5000. LiF) and film dosimetry V-Omat, Kodak). In case of fetus, the dosimeter was placed at a depth of loom in this phantom at 100cm source to axis distance and located centrally 15cm from the inferior edge of the $30cm{\times}30cm^2$ x-ray beam irradiating the Rando phantom chest wall. A acryl bridge of size $40cm{\times}40cm^2$ and a clear space of about 20 cm was fabricated and placed on top of the rectangular polystyrene phantom representing the abdomen of the patient. The leaf pot for testicle shielding was made as various shape, sizes, thickness and supporting stand. The scattered photon with and without shielding were measured at the representative position of the fetus and testicle. Measurement of radiation scattered dose outside fields and critical organs, like fetus position and testicle region, from chest or pelvic irradiation by large fie]d of high energy radiation beam was performed using an ionization chamber and film dosimetry. The scattered doses outside field were measured 5 - 10% of maximum doses in fields and exponentially decrease from field margins. The scattered photon dose received the fetus and testicle from thorax field irradiation was measured about 1 mGy/Gy of photon treatment dose. Shielding construction to reduce this scattered dose was investigated using lead sheet and blocks. Lead pot shield for testicle reduced the scatter dose under 10 mGy when photon beam of 60 Gy was irradiated in abdomen region. The scattered photon dose is reduced when the lead shield was used while the no significant reduction of scattered photon dose was observed and 2-3 mm lead sheets refuted the skin dose under 80% and almost electron contamination. The results indicate that it was possible to improve shielding to reduce scattered photon for fetus and testicle when a young patients were treated with a high energy photon beam.

A Study on the Nutritive Value and Utilization of Powdered Seaweeds (해조의 식용분말화에 관한 연구)

  • Yu, Jong-Yull;Lee, Ki-Yull;Kim, Sook-Hee
    • Journal of Nutrition and Health
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    • v.8 no.1
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    • pp.15-37
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    • 1975
  • I. Subject of the study A study on the nutritive value and utilization of powdered seaweeds. II. Purpose and Importance of the study A. In Korea the shortage of food will be inevitable by the rapidly growing population. It will be very important study to develop a new food from the seaweeds which were not used hitherto for human consumption. B. The several kinds of seaweeds have been used by man in Korea mainly as side-dishes. However, a properly powdered seaweed will enable itself to be a good supplement or mixture to certain cereal flours. C. By adding the powdered seaweed to any cereals which have long been staple foods in this country the two fold benefits; saving of cereals and change of dietary pattern, will be secured. III. Objects and scope of the study A. Objects of the study The objects will come under four items. 1. To develop a powdered seaweed as a new food from the seaweeds which have been not used for human consumption. 2. To evaluate the nutritional quality of the products the analysis for chemical composition and animal feeding experiment will be conducted. 3. Experimental cocking and accepability test will be conducted for the powdered products to evaluate the value as food stuff. 4. Sanitary test and also economical analysis will be conducted for the powdered products. B. Scope of the study 1. Production of seaweed powders Sargassum fulvellum growing in eastern coast and Sargassum patens C.A. in southern coast were used as the material for the powders. These algae, which have been not used for human consumption, were pulverized through the processes of washing, drying, pulverization, etc. 2. Nutritional experiments a. Chemical composition Proximate components (water, protein, fat, cellulose, sugar, ash, salt), minerals (calcium, phosphorus, iron, iodine), vitamins (A, $B_1,\;B_2$ niacin, C) and amino acids were analyzed for the seaweed powders. b. Animal feeding experiment Weaning 160 rats (80 male and 80 female rats) were used as experimental animals, dividing them into 16 groups, 10 rats each group. Each group was fed for 12 weeks on cereal diet (Wheat flour, rice powder, barley powder, potato powder, corn flour) with the supplementary levels of 5%, 10%, 15%, 20% and 30% of the seaweed powder. After the feeding the growth, feed efficiency ratio, protain efficiency ratio and ,organs weights were checked and urine analysis, feces analysis and serum analysis were also conducted. 3. Experimental cooking and acceptability test a. Several basic studies were conducted to find the characteristics of the seaweed powder. b. 17 kinds of Korean dishes and 9 kinds of foreign dishes were prepared with cereal flours (wheat, rice, barley, potato, corn) with the supplementary levels of 5%, 10%, 15%, 20% and 30% of the seaweed powder. c. Acceptability test for the dishes was conducted according to plank's Form. 4. Sanitary test The heavy metals (Cd, Pb, As, Hg) in the seaweed powders were determined. 5. Economical analysis The retail price of the seaweed powder was compared with those of other cereals in the market. And also economical analysis was made from the nutritional point of view, calculating the body weight gained in grams per unit price of each feeding diet. IV. Results of the study and the suggestion for application A. Chemical composition 1. There is no any big difference in proximate components between powders of Sargassum fulvellum in eastern coast and Sargassum patens C.A. in southern coast. Seasonal difference is also not significant. Higher levels of protein, cellulose, ash and salt were found in the powders compared with common cereal foods. 2. The levels of calcium (Ca) and iron (Fe) in the powders were significantly higher than common cereal foods and also rich in iodine (I). Existence of vitamin A and vitamin C in the Powders is different point from cereal foods. Vitamin $B_1\;and\;B_2$ are also relatively rich in the powders.'Vitamin A in ·Sargassum fulvellum is high and the levels of some minerals and vitamins are seemed4 to be some influenced by seasons. 3. In the amino acid composition methionine, isoleucine, Iysine and valine are limiting amino acids. The protein qualities of Sargassum fulvellum and Sargassum patens C.A. are seemed to be .almost same and generally ·good. Seasonal difference in amino acid composition was found. B. Animal feeding experiment 1. The best growth was found at.10% supplemental level of the seaweed Powder and lower growth rate was shown at 30% level. 2. It was shown that 15% supplemental level of the Seaweed powder seems to fulfil, to some extent the mineral requirement of the animals. 3. No any changes were found in organs development except that, in kidney, there found decreasing in weight by increasing the supplemental level of the seaweed powder. 4. There is no any significant changes in nitrogen retention, serum cholesterol, serum calcium and urinary calcium in each supplemental level of the seaweed powder. 5. In animal feeding experiment it was concluded that $5%{\sim}15%$ levels supplementation of the seaweed powder are possible. C. Experimental cooking and acceptability test 1. The seaweed powder showed to be utilized more excellently in foreign cookings than in Korean cookings. Higher supplemental level of seaweed was passible in foreign cookings. 2. Hae-Jo-Kang and Jeon-Byung were more excellent than Song-Pyun, wheat cake, Soo-Je-Bee and wheat noodle. Hae-Je-Kang was excellent in its quality even as high as 5% supplemental level. 3. The higher levels of supplementation were used the more sticky cooking products were obtained. Song-Pyun and wheat cake were palatable and lustrous in 2% supplementation level. 4. In drop cookie the higher levels of supplementation, the more crisp product was obtained, compared with other cookies. 5. Corn cake, thin rice gruel, rice gruel and potato Jeon-Byung were more excellent in their quality than potato Man-Doo and potato noodle. Corn cake, thin rice gruel and rice gruel were excellent even as high as 5% supplementation level. 6. In several cooking Porducts some seaweed-oder was perceived in case of 3% or more levels of supplementation. This may be much diminished by the use of proper condiments. D. Sanitary test It seems that there is no any heavy metals (Cd, Pb, As, Hg) problem in these seaweed Powders in case these Powders are used as supplements to any cereal flours E. Economical analysis The price of the seaweed powder is lower than those of other cereals and that may be more lowered when mass production of the seaweed powder is made in future. The supplement of the seaweed powder to any cereals is also economical with the criterion of animal growth rate. F. It is recommended that these seaweed powders should be developed and used as supplement to any cereal flours or used as other food material. By doing so, both saving of cereals and improvement of individual's nutrition will greatly be achieved. It is also recommended that the feeding experiment for men would be conducted in future.

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Crystal Structures of Dehydrated Partially $Sr^{2+}$-Exchanged Zeolite X, $Sr_{31}K_{30}Si_{100}A1_{92}O_{384}\;and\;Sr_{8.5}TI_{75}Si_{100}AI_{92}O_{384}$ (부분적으로 스트론튬이온으로 교환되고 탈수된, 제올라이트 X의 결정구조)

  • Kim Mi Jung;Kim Yang;Seff Karl
    • Korean Journal of Crystallography
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    • v.8 no.1
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    • pp.6-14
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    • 1997
  • The crystal structures of $Sr_{31}K_{30}-X\;(Sr_{31}K_{30}Si_{100}A1_{92}O_{384};\;a=25.169(5) {\AA}$) and $Sr_{8.5}Tl_{75}-X (Sr_{8.5}Tl_{75}Si_{100}A1_{92}O_{384};\;a=25.041(5) {\AA}$) have been determined by single-crystal X-ray diffraction techniques in the cubic space group $\=F{d3}\;at\;21(1)^{\circ}C$. Each crystal was prepared by ion exchange in a flowing stream of aqueous $Sr(ClO_4)_2\;and\;(K\;or\;T1)NO_3$ whose mole ratio was 1 : 5 for five days. Vacuum dehydration was done at $360^{\circ}C$ for 2d. Their structures were refined to the final error indices $R_1=0.072\;and\;R_w=0.057$ with 293 reflections, and $R_1= 0.058\;and\;R_w=0.044$ with 351 reflections, for which $I>2{\sigma}(I)$, respectively. In dehydrated $Sr_{31}K_{30}-X,\;all\;Sr^{2+}$ ions and $K^+$ ions are located at five different crystallographic sites. Six-teen $Sr^{2+}$ ions per unit cell are at the centers of the double six-rings (site I), filling that position. The remaining 15 $Sr^{2+}$ ions and 17 $K^+$ ions fill site II in the supercage. These $Sr^{2+}$ and $K^+$ ions are recessed ca $0.45{\AA}\;and\;1.06{\AA}$ into the supercage, respectively, from the plane of three oxygens to which each is bound. ($Sr-O=2.45(1){\AA}\;and\;K-O=2.64(1){\AA}$) Eight $K^+$ ons occupy site III'($K-O=3.09(7){\AA}\;and\;3.11(10){\AA}$) and the remaining five $K^+$ ions occupy another site III'($K-O=2.88(7){\AA}\;and\;2.76(7){\AA}$). In $Sr_{8.5}Tl_{75}-X,\;Sr^{2+}\;and\;Tl^+$ ions also occupy five different crystallographic sites. About 8.5 $Sr^{2+}$ ions are at site I. Fifteen $Tl^+$ ions are at site I' in the sodalite cavities on threefold axes opposite double six-rings: each is $1.68{\AA}$ from the plane of its three oxygens ($T1-O=2.70(2){\AA}$). Together these fill the double six-rings. Another 32 $Tl^+$ ions fill site II opposite single six-rings in the supercage, each being $1.48{\AA}$ from the plane of three oxygens ($T1-O=2.70(1){\AA}$). About 18 $Tl^+$ ions occupy site III in the supercage ($T1-O=2.86(2){\AA}$), and the remaining 10 are found at site III' in the supercage ($T1-O=2.96(4){\AA}$).

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The Usefulness of Pressure-regulated Volume Control(PRVC) Mode in Mechanically Ventilated Patients with Unstable Respiratory Mechanics (기계 호흡 중 불안정한 호흡역학을 보인 환자에서 압력조절용적조정양식(Pressure-regulated Volume Control Mode)의 효용)

  • Sohn, Jang-Won;Koh, Youn-Suck;Lim, Chae-Man;Shim, Tae-Sun;Lee, Jong-Deog;Lee, Sang-Do;Kim, Woo-Sung;Kim, Dong-Soon;Kim, Won-Dong
    • Tuberculosis and Respiratory Diseases
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    • v.44 no.6
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    • pp.1318-1325
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    • 1997
  • Background : Since the late 1960s, mechanical ventilation has been accomplished primarily using volume controlled ventilation(VCV). While VCV allows a set tidal volume to be guaranteed, VCV could bring about excessive airway pressures that may be lead to barotrauma in the patients with acute lung injury. With the increment of knowledge related to ventilator-induced lung injury, pressure controlled ventilation(PCV) has been frequently applied to these patients. But, PCV has a disadvantage of variable tidal volume delivery as pulmonary impedance changes. Since the concept of combining the positive attributes of VCV and PCV(dual control ventilation, DCV) was described firstly in 1992, a few DCV modes were introduced. Pressure-regulated volume control(PRVC) mode, a kind of DCV, is pressure-limited, time-cycled ventilation that uses tidal volume as a feedback control for continuously adjusting the pressure limit However, no clinical studies were published on the efficacy of PRVC until now. 'This investigation studied the efficacy of PRVC in the patients with unstable respiratory mechanics. Methods : The subjects were 8 mechanically ventilated patients(M : F=6 : 2, $56{\pm}26$ years) who showed unstable respiratory mechanics, which was defined by the coefficients of variation of peak inspiratory pressure for 15 minutes greater than 10% under VCV, or the coefficients of variation of tidal volume greater than 10% under PCV. The study was consisited of 3 modes application with VCV, PCV and PRVC for 15 minutes by random order. To obtain same tidal volume, inspiratory pressure setting was adjusted in PCV. Respiratory parameters were measured by pulmonary monitor(CP-100 pulmonary monitor, Bicore, Irvine, CA, USA). Results : 1) Mean tidal volumes($V_T$) in each mode were not different(VCV, $431{\pm}102ml$ ; PCV, $417{\pm}99ml$ ; PRVC, $414{\pm}97ml$) 2) The coefficient of variation(CV) of $V_T$ were $5.2{\pm}3.9%$ in VCV, $15.2{\pm}7.5%$ in PCV and $19.3{\pm}10.0%$ in PRVC. The CV of $V_T$ in PCV and PRVC were significantly greater than that in VCV(p<0.01). 3) Mean peak inspiratory pressure(PIP) in VCV($31.0{\pm}6.9cm$ $H_2O$) was higher than PIP in PCV($26.0{\pm}6.5cm$ $H_2O$) or PRVC($27.0{\pm}6.4cm$ $H_2O$)(p<0.05). 4) The CV of PIP were $13.9{\pm}3.7%$ in VCV, $4.9{\pm}2.6%$ in PVC and $12.2{\pm}7.0%$ in PRVC. The CV of PIP in VCV and PRVC were greater than that in PCV(p<0.01). Conclusions : Because of wide fluctuations of VT and PIP, PRVC mode did not seem to have advantages compared to VCV or PCV in the patients with unstable respiratory mechanics.

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Microbiological and Enzymological Studies on the Flavor Components of Sea Food Pickles (젓갈등속(等屬)의 정미성분(呈味成分)에 관(關)한 미생물학적(微生物學的) 및 효소학적(酵素學的) 연구(硏究))

  • Lee, Ke-Ho
    • Applied Biological Chemistry
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    • v.11
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    • pp.1-27
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    • 1969
  • More than thirty kinds of sea food pickles have been eaten in Korea. Out of these salted yellow tail pickle, salted clam pickle, salted oyster pickle, and salted cuttlefish pickle were employed for the analysis of their components, identification of main fermenting microbes, and determination of enzyme characteristics concerned. Also studied was the effect of enzymic action of microbes, which are concerned with the fermenting of pickles, on the production of flavorous 5'-mononucleotides and amino acids. The results are summarized as follows: 1. Microflora observed in the pickles are: (a) Total count of viable cells after 1-2 months of pickling was found to be $10^7$ and that after 6 months decreased to $10^4$. (b) Microbial occurence in the early stage of pickling was observed to be 10-20% Micrococcus spp., 10-20% Brevibacterium spp., 0-30% Sarcina spp., 20-30% Leuconostoc spp., ca 30% Bacillus spp., 0-10% Pseudomonas spp., 0-10% Flavobacterium spp., and 0-20% yeast. (c) Following the early stage of pickling, mainly halophilic bacteria such as Bacillus subtilis, Leuconostoc mesenteroides, Pediococcus halophilus and Sarcina litoralis, were found to exhibit an effect on the fermentation of pickle and their enzyme activities were in direct concern in fermentation of pickles. (d) Among the bacteria participating in the fermentation, Sarcina litoralis 8-14 and 8-16 strains were in need of high nutritional requirement and the former was grown only in the presence of purine, pyrimidine and cystine and the latter purine, pyrimidine and glutamic acid. 2. Enzyme characteristics studied in relation to the raw materials and the concerned microbes isolated are as follows: (a) A small amount of protease was found in the raw materials and 30-60% decrease in protease activity was demonstrated at 7% salt concentration. (b) Protease activity of halophilic bacteria, Bacillus subtilis 7-6, 11-1, 3-6 and 9-4 strains, in the complete media decreased by 10-30% at the 7% salt concentration and that of Sarcina litoralis 8-14 and 8-16 strains decreased by 10-20%. (c) Proteins in the raw materials were found to be hydrolyzed to yield free amino acids by protease in the fermenting microbes. (d) No accumulation of flavorous 5'-mononucleotides was demonstrated because RNA-depolymerase in the raw materials and the pickles tended to decompose RNA into nucleoside and phosphoric acid. (e) The enzyme produced in Bacillus subtilis 3-6 strain isolated from the salted clam pickles, was ascertained to be 5'-phosphodiesterase because of its ability to decompose RNA and thus accumulating 5'-mononucleotide. (f) It was demonstrated that the activity of phosphodiesterase in Bacillus subtilis 3-6 strain was enhanced by some components in the corn steep liquor and salted clam pickle. The enzyme activity was found to decrease by 10-30% and 40-60% at the salt concentration of 10% and 20%, respectively. 3. Quantitative data for free amino acids in the pickles are as follows: (a) Amounts of acidic amino acids such as glutamic and aspartic acids in salted clam pickle, were observed to be 2-10 times other pickles and it is considered that the abundance in these amino acids may contribute significantly to the specific flavor of this food. (b) Large amounts of basic amino acids such as arginine and histidine were found to occur in salted yellow tail pickle. (c) It is much interesting that in the salted cuttlefish pickle the contents of sulfur-containing amino acids were exceedingly high compared with those of others: cystine was found to be 17-130 times and methionine, 7-19 times. (d) In the salted oyster pickle a high content of some essential amino acids such as lysine, threonine, isoleucine and leucine, was demonstrated and a specific flavor of the pickle was ascribed to the sweet amino acids. Contents of alanine and glycine in the salted oyster pickle were 4 and 3-14 times as much as those of the others respectively. 4. Analytical data for 5'-mononucleotides in the pickles are as follows: (a) 5'-Adenylic acid and 3'-adenylic acid were found in large amounts in the salted yellow tail pickle and 5'-inosinic acid in lesser amount. (b) 5'-Adenylic acid, especially 3'-adenylic acid predominated in amount in the salted oyster pickle over that in the other pickles. (c) The salted cuttlefish pickle was found to contain only 5'-adenylic acid and 3'-adenylic acid. It has become evident from the above fact that clam and the invertebrate lack of adenylic deaminase and contain high content of adenylic acid. Thus, they were demonstrated to be the AMP-type. (d) 5'-Inosinic acid was contained in the salted yellow tail pickle in a significant concentration, and it might be considered to be IMP-type. 5. Comparative data for flavor with regard to the flavorous amino acids and the contents of 5'-mononucleotides are: (a) A specific flavor of salted yellow tail pickle was ascribed to the abundance in glutamic acid and aspartic acid, and to the existence of a small amount of flavorous 5'-inosinic acid. The combined effect of these components was belived to exhibit a synergistic action in producing a specific fiavor to the pickle. (b) A specific flavor of salted clam pickle has been demonstrated to be attributable to the richness in glutamic acid and aspartic acid rather than to that of 5'-mononucleotides.

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