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Studies on the Amylase Production by Bacteria (세균(細菌)에 의(依)한 Amylase생산(生産)에 관한 연구(硏究))

  • Park, Yoon-Joong
    • Applied Biological Chemistry
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    • v.13 no.2
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    • pp.153-170
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    • 1970
  • 1. Isolation and identification of amylase-producing bacteria. The powerful strain A-12 and S-8 were respectively isolated from air and soil after screening a large number of amylase-producing bacteria. Their bacterial characteristics have been investigated and it has been found that all characteristics of strain A-12 and S-8 are similar to Bac. subtilis of Bergey's manual except for the acid formation from a few carbohydrates and the citrate utilization, i.e., the strain A-12 shows negative in the citrate utilization, and the acid formation from arabinose and xylose, S-8 shows negative in the acid formation from xylose. 2. Amylase production by Liquid cultures with solid materials. Several conditions for amylase production by strain A-12 in stationary cultures have been studied. The results obtained are as follows. (1) The optimum conditions are:temperature $35^{\circ}C$, initial pH 6.5 to 7.0 and incubation time 3 to 4 days. (2) The amylase production is not affected by the preservation period of the stock cultures. (3) Among the various solid material, the defatted soy bean is found to be the best for t1e amylase production. However, the alkali treatment of the defatted soy bean gives no effect contrary to the cage of defatted rape seed. The addition of soluble starch to the alkali extract of defatted soy bean shows the increased amylase production. (4) Up to 1% addition of ethanol to carbon dificient media gives the improved amylase production, whereas the above effect is not found in the case of carbon rich media. (5) The amylase production can be increased 2.5 times when 10% of defatted soy bean is admixed to cheaply available wheat bran. (6) The excellent effect is found for amylase production when 20% of wheat bran is admixed to defatted dry milk which is a poor medium. The activity is found to be $D^{40^{\circ}}_{30'}$ 7,000(L.S.V. 1,800) in 10% medium. (7) No significant effect is observed due to the addition of various inorganic salts. 3. Amylase production by solid cultures. Several conditions for amylase production by strain A-12 in wheat bran cultures have been studied and the results obtained are as follows. (1) The optimum conditions: are temperature $33^{\circ}C$, incubation lime 2 days, water content added 150 to 175% and the thickness of the medium 1.5cm, The activity is found to be $D^{40^{\circ}}_{30'}$ 36,000(L.S.V. 15,000) (2) No significant effect is found in the case of the additions of various organic and inorganic substances.

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Studios on Lipids in Fresh-Water Fishes 5. Distribution of Lipid Components in Various Tissues of Carp, Cyprynus carpio (담수어의 지질에 관한 연구 5. 잉어(Cyprynus carpio)의 부위별 지질성분의 분포)

  • CHOI Jin-Ho;RO Jae-Il;BYUN Dae-Seok;PYEUN Jae-Hyeung
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.18 no.2
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    • pp.149-156
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    • 1985
  • Distribution of lipid components in the tissue of meat, skin and viscera from carp(Cyprynus carpio) was analyzed using the techniques of column chromatography, thin layer chromatography and gas liquid chromatography according to the previous report(Choi, et al., 1984). Lipid content was varied by the portion such as $3.88\%$ in meat (free lipid, $2.47\%$ ; bound lipid, $1.41\%$), $8.02\%$ in skin(free lipid, $5.65\%$ ; bound lipid, $2.37\%$) and $6.18\%$ in viscera (free lipid, $3.54\%$ ; bound lipid, $2.64\%$). In the all portions of the body, free lipid was composed of $68\%\;to\;92\%$ in neutral lipid, $3\%\;to\;6\%$ in glycolipid and $4\%\;to\;18\%$ in phospholipid whereas bound lipid was composed of $8\%\;to\;20\%$ in neutral lipid, $2\%\;to\;7\%$ in glycolipid and $47\%\;to\;62\%$ in phospholipid. The free lipids of the tissues on the each portion were mostly represented by triglycerides and some diglycerides, but free lipids in viscera contained considerable amounts of free fatty acids. The bound lipids, on the other hand, commonly comprised appreciable amounts of esterified sterol and hydrocarbon, and triglycerides. The phospholipid was mainly consisted of phosphatidyl choline, phosphatidyl ethanolamine and phosphatidyl serine in the both free and bound lipids, and much more phosphatidyl choline in the bound lipid. The predominant fatty acids of free and bound lipids were $C_{16:0},\;C_{18:0},\;C_{20:4},\;C_{22:6}\;and\;C_{18:2}$ acids in polar lipids, and $C_{16:0},\;C_{16:1},\;C_{18:0},\;C_{18:1}\;and\;C_{18:2}$ acids in non-polar lipids, whereas those of neutral lipids were $C_{14:0}(2.54{\sim}6.98\%),\;C_{16:0}(11.20{\sim}21.13\%)$ and $C_{18:0}(1.58{\sim}12.76\%)$ of saturated acids, $C_{16:1}(7.06{\sim}20.70\%),\;C_{18:1}(21.68{\sim}30.50\%)$ and $C_{20:1}(1.76{\sim}6.27\%)$ of monoenoic acids, and $C_{18:2}(4.50{\sim}6.89\%),\;C_{20:4}(1.52{\sim}4.29\%)$ and $C_{22:6}(0.73{\sim}6.62\%)$, respectively. In conclusion, the fatty acid compositions revealed apparent differences between the free lipid and bound lipids in the tissues of body.

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Studies on the Factors Affecting Barley Injury Caused by Herbicides in Drained Paddy Field (제초제에 의한 답리작맥 약해발생 요인구명에 관한 연구)

  • Whan-Seung Ryang
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.14
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    • pp.147-157
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    • 1973
  • I. The effect of excessive soil moisture(at the time of germination) on germination of barley and crop damage of herbicides was investigated. Machete(Butachlor) and TOK(Nitrofen) were treated, respectively, at the rate of 150g ai/10a on each pot whose different soil moisture content was controlled by suppling 30, 40, 50 and 60ml of water per 100gr of air-dried soil, respectively. The results are summarized as follows: 1. Excessive soil moisture beyond field moisture capacity caused great inhibition, from 20 to 100%, of the germination of barley even at untreated pots(check pots). Also, further development of root and growth of barley were greatly inhibited even though the seeds germinated. 2. The same tendency in inhibition of germination and growth as at untreated pots was observed at treated pots, too. As a whole, however, the damage were heavier at treated pots. II. Wanju naked spring barley was seeded on four different soils and covered with soil to a depth of 1 em, and then Machete, TOK, Saturn and HE-314 were treated at the rate of 180, 150 and 200, 150, and 250g ai/10a, respectively, and the effect of soil texture on crop damage of the herbicides was investigated. The results are summarized as follows: 1. Machete(emulsion and granule, at 180g ai/10a) The degree of crop damage was quite different from one soil texture to another: while almost no crop damage was observed on a clay loam soil regardless of the type of formulation, the damage became heavier as the soil texture became sandier as sandy clay loam, volcanic ash loam and sandy loam, and great inhibition of growth was observed on sandy loam soil. In general heavier damage was caused by the application of emulsion than by granular formulation. 2. TOK(Wettable powder, at 150, 250g ai/l0a) Almost the same tendency as in the application of Machete was observed, and the damage became heavier as the application rate increased. 3. Saturn(at l50g ai/l0a) No great difference in crop damage among soil textures was observed. 4. HE-3l4(at 250g ai/l0a) Almost no difference in crop damage among soil textures was observed at this rate of 250g ai/l0a. III. To study a difference of crop damage on soil covering depth(4 levels), 9 herbicides(TOK, MO, HE-3l4, Machete, Saturn, Simetryne, Simazine, Gesaran, Lorox) were treated on the pots with two different soils, and the effect of soil covering depth on crop damage of the herbicides was investigated. The results obtained in this experiment are summarized as follows: Light Clay Soil 1. The growth of barley in relation to depth of soil covering at check pots followed the order vigorous to weak; lcm>1.5cm>0.5cm>0cm. And in case of 0 and 0.5cm covering the growth of barley was very poor. 2. The damage at 0 and 0.5cm covering at treated pots was very severe, but Saturn, Machete, MO and TOK at 100 to l50g ai/l0a, respectively and He-3l4 at 250 to 375g ai/l0a were relatively safe to barley at the depths of lcm and above. 3. Simazine, Lorox and Simetryne caused slight damage even at 1.5cm covering. Sandy Loam Soil The growth of barley in relation to depth of soil covering at untreated pots followed the order, from vigorous to weak; 1.5cm 0.5cm 3cm 5cm. While MO was safe to barley at 1.5cm covering, for other chemicals more than 3cm covering was require for safe use. Machete and Saturn at 100g ai/l0a, and HE-3l4 at 250g ai/l0a was relatively safe at more than 3cm covering. Simazine, Lorox, Simetryne and Gesaran were unsafe on sandy soil regardless of covering depth.

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Composition of Vitamin A, E, $B_l$ and $B_2$ Contents in Korean Cow's Raw Milk in Korea (국내산 원유 중 비타민 A, E, $B_l$$B_2$ 함량에 관한 연구)

  • Kwak Byung-Man;Kim Sung-Han;Kim Kang-Seob;Lee Ki-Woong;Ahn Jang-Hyuk;Jang Chi-Hoon
    • Food Science of Animal Resources
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    • v.26 no.2
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    • pp.245-251
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    • 2006
  • This study was performed to investigate the changes of vitamin A, vitamin E, vitamin $B_l$ and vitamin $B_2$ contents in cow's raw milk collected from dairy farms in Chungcheong-do and Jeolla-do for a year. The contents of fat soluble vitamin A and E were changed as seasonal effect, but water soluble vitamin $B_l$ and $B_2$ contents were not changed as seasonal effect. Vitamin A content in cow's raw milk was as follows [minimum ${\sim}$maximum (mean), ug/100 mL]; $35.1{\sim}59.0$ (44.4) in spring, $36.7{\sim}65.6$ (50.0) in summer, $28.7{\sim}61.2$ (46.8) in autumn and $29.9{\sim}57.8$ (43.1) in winter. In case of vitamin E was as follows [minimum${\sim}$maximum (mean), ug/100 mL]; $28.3{\sim}59.2$ (45.8) in spring, $39.6{\sim}69.9$ (58.8) in summer, $35.0{\sim}62.8$ (46.2) in autumn and $26.0{\sim}55.4$ (41.5) in winter. In case of vitamin $B_l$ was as follow [minimum${\sim}$maximum (mean), ug/100 mL]; $27.7{\sim}57.9$ (42.84) in spring, $32.4{\sim}66.1$ (49.39) in summer, $34.1{\sim}63.7$ (46.69) in autumn and $20.6{\sim}61.4$ (43.20 in winter. The amounts of vitamin $B_2$ in cow's raw milk was as follows [minimum${\sim}$maximum (mean), ug/100 mL]; $150{\sim}182$ (160 in spring, $145{\sim}185$ (163) in summer, $149{\sim}180$ (166) in autumn and $148{\sim}190$ (167) in winter.

A Study on the Growth Diagnosis and Management Prescription for Population of Retusa Fringe Trees in Pyeongji-ri, Jinan(Natural Monument No. 214) (진안 평지리 이팝나무군(천연기념물 제214호)의 생육진단 및 관리방안)

  • Rho, Jae-Hyun;Oh, Hyun-Kyung;Han, Sang-Yub;Choi, Yung-Hyun;Son, Hee-Kyung
    • Journal of the Korean Institute of Traditional Landscape Architecture
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    • v.36 no.3
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    • pp.115-127
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    • 2018
  • This study was attempted to find out the value of cultural assets through the clear diagnosis and prescription of the dead and weakness factors of the Population of Retusa Fringe Trees in Pyeongji-ri, Jinan(Natural Monument No. 214), The results are as follows. First, Since the designation of 13 natural monuments in 1968, since 1973, many years have passed since then. In particular, despite the removal of some of the buried soil during the maintenance process, such as retreating from the fence of the primary school after 2010, Second, The first and third surviving tree of the designated trees also have many branches that are dead, the leaves are dull, and the amount of leaves is small. vitality of tree is 'extremely bad', and the first branch has already been faded by a large number of branches, and the amount of leaves is considerably low this year, so that only two flowers are bloomed. The second is also in a 'bad'state, with small leaves, low leaf density, and deformed water. The largest number 1 in the world is added to the concern that the s coverd oil is assumed to be paddy soils. Third, It is found that the composition ratio of silt is high because it is known as '[silty loam(SiL)]'. In addition, the pH of the northern soil at pH 1 was 6.6, which was significantly different from that of the other soil. In addition, the organic matter content was higher than the appropriate range, which is considered to reflect the result of continuous application for protection management. Fourth, It is considered that the root cause of failure and growth of Jinan pyeongji-ri Population of Retusa Fringe Trees group is chronic syndrome of serious menstrual deterioration due to covered soil. This can also be attributed to the newly planted succession and to some of the deaths. Fifthly, It is urgent to gradually remove the subsoil part, which is estimated to be the cause of the initial damage. Above all, it is almost impossible to remove the coverd soil after grasping the details of the soil, such as clayey soil, which is buried in the rootstock. After removal of the coverd soil, a pestle is installed to improve the respiration of the roots and the ground with Masato. And the dead 4th dead wood and the 5th and 6th dead wood are the best, and the lower layer vegetation is mown. The viable neck should be removed from the upper surface, and the bark defect should undergo surgery and induce the development of blindness by vestibule below the growth point. Sixth, The underground roots should be identified to prepare a method to improve the decompression of the root and the respiration of the soil. It is induced by the shortening of rotten roots by tracing the first half of the rootstock to induce the generation of new roots. Seventh, We try mulching to suppress weed occurrence, trampling pressure, and soil moisturizing effect. In addition, consideration should be given to the fertilization of the foliar fertilizer, the injection of the nutrients, and the soil management of the inorganic fertilizer for the continuous nutrition supply. Future monitoring and forecasting plans should be developed to check for changes continuously.

Coffee consumption behaviors, dietary habits, and dietary nutrient intakes according to coffee intake amount among university students (일부 대학생의 커피섭취량에 따른 커피섭취행동, 식습관 및 식사 영양소 섭취)

  • Kim, Sun-Hyo
    • Journal of Nutrition and Health
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    • v.50 no.3
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    • pp.270-283
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    • 2017
  • Purpose: This study was conducted to examine coffee consumption behaviors, dietary habits, and nutrient intakes by coffee intake amount among university students. Methods: Questionnaires were distributed to 300 university students randomly selected in Gongju. Dietary survey was administered during two weekdays by the food record method. Results: Subjects were divided into three groups: NCG (non-coffee group), LCG (low coffee group, 1~2 cups/d), and HCG (high coffee group, 3 cups/d) by coffee intake amount and subjects' distribution. Coffee intake frequency was significantly greater in the HCG compared to the LCG (p < 0.001). The HCG was more likely to intake dripped coffee with or without milk and/or sugar than the LCG (p < 0.05). More than 80% of coffee drinkers chose their favorite coffee or accompanying snacks regardless of energy content. More than 75% of coffee takers did not eat accompanying snacks instead of meals, and the HCG ate them more frequently than LCG (p < 0.05). Breakfast skipping rate was high while vegetable and fruit intakes were very low in most subjects. Subjects who drank carbonated drinks, sweet beverages, or alcohol were significantly greater in number in the LCG and HCG than in the NCG (p < 0.01). Energy intakes from coffee were $0.88{\pm}5.62kcal/d$ and $7.07{\pm}16.93kcal/d$ for the LCG and HCG. For total subjects, daily mean dietary energy intake was low at less than 72% of estimated energy requirement. Levels of vitamin C and calcium were lower than the estimated average requirements while that of vitamin D was low (24~34% of adequate intake). There was no difference in nutrient intakes by coffee intake amount, except protein, vitamin A, and niacin. Conclusion: Coffee intake amount did not affect dietary nutrient intakes. Dietary habits were poor,and most nutrient intakes were lower than recommend levels. High intakes of coffee seemed to be related with high consumption of sweet beverages and alcohol. Therefore, it is necessary to improve nutritional intakes and encourage proper water intake habits, including coffee intake, for improved nutritional status of subjects.

Studies on Sclerotium rolfsii Sacc. isolated from Magnolia kobus DC. in Korea (목련(Magnolia kobus DC.)에서 분리한 흰비단병균(Sclerotium rolfsii Sacc.)에 관한 연구)

  • Kim Kichung
    • Korean journal of applied entomology
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    • v.13 no.3 s.20
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    • pp.105-133
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    • 1974
  • The present study is an attempt to solve the basic problems involved in the control of the Sclerotium disease. The biologic stranis of Sclerotium rolfsii Sacc., pathogen of Sclerotium disease of Magnolia kobus, were differentiated, and the effects of vitamins, various nitrogen and carbon sources on its mycelial growth and sclerotial production have been investigated. In addition the relationship between the cultural filtrate of Penicillium sp. and the growth of Sclerotium rolfsii, the tolerance of its mycelia or sclerotia to moist heat or drought and to Benlate (methyl-(butylcarbamoy 1)-2-benzimidazole carbamate), Tachigaren (3-hydroxy-5-methylisoxazole) and other chemicals were also clarified. The results are summarizee as follows: 1. There were two biologic strains, Type-l and Type-2 among isolates. They differed from each other in the mode of growth and colonial appearance on the media, aversion phenomenon and in their pathogenicity. These two types had similar pathogenicity to the Magnolia kobus and Robinia pseudoacasia, but behaved somewhat differently to the soybaen and cucumber, the Type-l being more virulent. 2. Except potassium nitrite, sodium nitrite and glycine, all of the 12 nitrogen sources tested were utilized for the mycelial growth and sclerotial production of this fungus when 10r/l of thiamine hydrochloride was added in the culture solution. Considering the forms of nitrogen, ammonium nitrogen was more available than nitrate nitrogen for the growth of mycelia, but nitrate nitrogen was better for sclerotia formation. Organic nitrogen showed different availabilities according to compounds used. While nitrite nitrogen was unavailable for both mycelial growth and sclerotial formation whether thiamine hydrochlioride was added or not. 3. Seven kinds of carbon sources examined were not effective in general, as long as thiamine hydrochloride was not added. When thiamine hydrochloride was added, glucose and saccharose exhibited mycelial growth, while rnaltose and soluble starch gave lesser, and xylose, lactose, and glycine showed no effect at all,. In the sclerotial production, all the tested carbon sources, except lactose, were effective, and glucose, maltose, saccharose, and soluble starch gave better results. 4. At the same level of nitrogen, the amount of mycelial growth increased as more carbon Sources were applied but decreased with the increase of nitrogen above 0.5g/1. The amount of sclerotial production decreased wi th the increase of carbon sources. 5. Sclerotium rolfsii was thiamine-defficient and required thiamine 20r/l for maximun growth of mycelia. At a higher concentration of more than 20r/l, however, mycelial growth decreased as the concentration increased, and was inhibited at l50r/l to such a degree of thiamine-free. 6. The effect of the nitrogen sources on the mycelial growth under the presence of thiamine were recognized in the decreasing order of $NH_4NO_3,\;(NH_4)_2SO_4,\;asparagine,\;KNO_3$, and their effects on the sclerotial production in the order of $KNO_3,\;NH_4NO_3,\;asparagine,\;(NH_4)_2SO_4$. The optimum concentration of thiamine was about 12r/l in $KNO_3$ and about 16r/l in asparagine for the growth of mycelia; about 8r/l in $KNO_3$ and $NH_4NO_3$, and 16r/l in asparagine for the production of sclerotia. 7. After the fungus started to grow, the pH value of cultural filtrate rapidly dropped to about 3.5. Hereafter, its rate slowed down as the growth amount increased and did not depreciated below pH2.2. 8. The role of thiamine in the growth of the organism was vital. If thiamine was not added, the combination of biotin, pyridoxine, and inositol did not show any effects on the growth of the organism at all. Equivalent or better mycelial growth was recognized in the combination of thiamine+pyridoxine, thiamine+inositol, thiamine+biotin+pyridoxine, and thiamine+biotin+pyridoxine+inositol, as compared with thiamine alone. In the combinations of thiamine+biotin and thiamine+biotin+inositol, mycelial growth was inhibited. Sclerotial production in dry weight increased more in these combinations than in the medium of thiamine alone. 9. The stimulating effects of the Penicillium cultural filtrate on the mycelial growth was noticed. It increased linearly with the increase of filtrate concentration up to 6-15 ml/50ml basal medium solution. 10. $NH_4NO_3$. as a nitrogen source for mycelial growth was more effective than asparasine regardless of the concentration of cultural filtrate. 11. In the series of fractionations of the cultural filtrate, mycelial growth occured in unvolatile, ether insoluble cation-adsorbed or anion-unadsorbed substance fractions among the fractions of volatile, unvolatile acids, ether soluble organic acids, ether insoluble, cation-adsorbed, cation-unadsorbed, anion-adsorbed and anion-unadsorbed. and anion-un-adsorbed substance tested. Sclerotia were produced only in cation-adsorbed fraction. 12. According to the above results, it was assumed that substances for the mycelial growth and sclerotial formation and inhibitor of sclerotial formation were include::! in cultural filtrate and they were quite different from each other. I was further assumed that the former two substances are un volatile, ether insotuble, and adsorbed to cation-exchange resin, but not adsorbed to anion, whereas the latter is unvolatile, ether insoluble, and not adsorbed to cation or anion-exchange resin. 13. Seven amino acids-aspartic acid, cystine, glysine, histidine, Iycine, tyrosine and dinitroaniline-were detected in the fractions adsorbed to cation-exchange resin by applying the paper chromatography improved with DNP-amino acids. 14. Mycelial growth or sclerotial production was not stimulated significantly by separate or combined application of glutamic acid, aspartic acid, cystine, histidine, and glysine. Tyrosine gave the stimulating effect when applied .alone and when combined with other amino acids in some cases. 15. The tolerance of sclerotia to moist heat varied according to their water content, that was, the dried sclerotia are more tolerant than wet ones. The sclerotia harvested directly from the media, both Type-1 and Type-2, lost viability within 5 minutes at $52^{\circ}C$. Sclerotia dried for 155 days at$26^{\circ}C$ had more tolerance: sclerotia of Type-l were killed in 15 mins. at $52^{\circ}C$ and in 5 mins. at $57^{\circ}C$, and sclerotia of Type-2 were killed in 10 mins. both at $52^{\circ}C$ or $57^{\circ}C$. 16. Cultural sclerotia of both strains maintained good germinability for 132 days at$26^{\circ}C$. Natural sclerotia of them stored for 283 days under air dry condition still had good germinability, even for 443 days: type-l and type-2 maintained $20\%$ and $26.9\%$ germinability, respectively. 17. The tolerance to low temperature increased in the order of mycelia, felts and sclerotia. Mycelia completely lost the ability to grow within 1 week at $7-8^{\circ}C$> below zero, while mycelial felts still maintained the viability after .3 weeks at $7-20^{\circ}C$ below zero, and sclerotia were even more tolerant. 18. Sclerotia of type-l and type-2 were killed when dipped into the $0.05\%$ solution of mercury chloride for 180 mins. and 240 mins. respectively: and in the $0.1\%$ solution, Type-l for 60 mins. and Type-2 for 30 mins. In the $0.125\%$ uspulun solution, Type-l sclerotia were killed in 180 mins., and those of Type-2 were killed for 90 mins. in the$0.125\%$solution. Dipping into the $5\%$ copper sulphate solution or $0.2\%$ solution of Ceresan lime or Mercron for 240 mins. failed to kill sclerotia of either Type-l or Type-2. 19. Inhibitory effect on mycelial growth of Benlate or Tachi-garen in the liquid culture increased as the concentration increased. 6 days after application, obvious inhibitory effects were found in all treatments except Benlate 0.5ppm; but after 12 days, distingushed diflerences were shown among the different concentrations. As compared with the control, mycelial growth was inhibited by $66\%$ at 0.5ppm and by $92\%$ at 2.0ppm of Benlate, and by$54\%$ at 1ppm and about $77\%$ at 1.5ppm or 2.0ppm of Tachigaren. The mycelial growth was inhibited completely at 500ppm of both fungicides, and the formation of sclerotia was checked at 1,000ppm of Benlate ant at 500ppm or 1,000ppm of Tachigaren. 20. Consumptions of glucose or ammonium nitrogen in the culture solution usually increased with the increment of mycelial growth, but when Benlate or Tachigaren were applied, consumptions of glucose or ammonium nitrogen were inhibited with the increment of concentration of the fungicides. At the low concentrations of Benlate (0.5ppm or 1ppm), however, ammonium nitrogen consumption was higher than that of the ontrol. 21. The amount of mycelia produced by consuming 1mg of glucose or ammonium nitrogen in the culture solution was lowered markedly by Benlate or Tachigaren. Such effects were the severest on the third day after their treatment in all concentrations, and then gradually recovered with the progress of time. 22. In the sand culture, mycelial growth was not inhibited. It was indirectly estimated by the amount of $CO_2$ evolved at any concentrations, except in the Tachigaren 100mg/g sand in which mycelial growth was inhibited significantly. Sclerotial production was completely depressed in the 10mg/g sand of Benlate or Tachigaren. 23. There was no visible inhibitory effect on the germination of sclerotia when the sclerotia were dipped in the solution 0.1, 1.0, 100, 1.000ppm of Benlate or Tachigaren for 10 minutes or even 20 minutes.

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