L-glutamic Acid Fermentation Research Articles

廃糖蜜によるL-グルタミン酸醗酵(第11報)

廃糖蜜によるL-グルタミン酸醗酵(第11報)

L-Glutamic Acid Fermentation with Molasses

L-Glutamic Acid Fermentation with Molasses

L-Glutamic Acid Fermentation with Molasses

1) An L-glutamic acid (L-GA)-forming bacterium, Microbacterium ammoniaphilum, was cultured in the molasses medium with or without POEFE under agitation and aeration, to obtain L-GA-accumulating cells or nonaccumulating cells respectively. Protoplast-like bodies from each group of cells were obtained by their reaction with egg white lysozyme under conditions which were modifications of those in the previous paper. 2) Then the reaction mixture was divided equally into two parts and each part was submitted to L-GA-accumulating reaction in the presence of glucose, urea, NaNO3 and other salts, under higher or lower osmotic pressure. After 3hr of the reaction, the L-GA yields from glucose in various cases were compared. It was shown that the L-GA yield by the PLB from L-GA-non-accumulating cells was about the same as that by the PLB from L-GA-accumulating cells under the lower osmotic pressure condition, when the PLB-bursting ratios were high, but the former was considerably lower under the higher osmotic pressure condition, when the PLB-bursting ratios were low. 4) Further, the abilities of burst and nonburst PLB from both types of cells to accumulate L-GA were calculated from the results of the experiment. No significant difference was found in the ability of L-GA accumulation between burst PLBs from both origins. On the other hand, there was found great difference in the case of the nonburst PLB. From these results and the previous report, it is presumed that the molar ratio of saturated fatty acids/unsaturated fatty acids in the cell membrane would be closely related to the nature of cell membrane which determines the permeability for L-GA, and therefore it could affect the extracellular amount of L-GA accumulated.

L-Glutamic Acid Fermentation with Molasses

L-Glutamic Acid Fermentation with Molasses

L-Glutamic Acid Fermentation

It is well known that biotin has a marked effect on L-glutamic acid fermentation. The authors have intended to find strains which are independent of the amounts of biotin in the culture medium. As a result, oleic acid-requiring mutants were obtained from a strain of Brevibacterium thiogenitalis which is an auxotroph for biotin. The growth of the mutant was remarkably stimulated by Tween 20, 40, 60, Ca ions and a small amount of corn steep liquor. And also, the mutant was found to have lost its requirement for biotin and showed growth response only to oleic acid or unsaturated fatty acids.

Biochemical Effects of Fatty Acid and its Derivatives on L-Glutamic Acid Fermentation

1. For the growth of Brev. lactofermentum, oleic acid was completely replaced for biotin; trace amount of biotin was not required. This result applied to the case of many lactic acid bacteria except L. arabinosus. 2. Oleic acid had a growth-inhibitory effect as well as the stimulatory one. This dualistic property was based on the concentration of the acid in the culture medium and was ob-served generally with unsaturated fatty acids irrespective of their chemical structures, com-prising the position and the number of double bond and cis- and trans-form. 3. The growth inhibition caused by higher concentration of oleic acid was changed into the growth stimulation in the three different ways: (a) the prolongation of culture, (b) the esterification of oleic acid, and (c) the addi-tion of the hydrophilic ester of saturated fatty acid as a detoxicant. 4. When added into a medium containing oleic acid, Tween 60 had not only the de-toxificating effect but also synergetic effect; the oleic acid-sparing effect in the presence of small amount of oleic acid and the lag period. reducing effect. 5. The results obtained with Brev. lacta-fermentum confirmed other investigators' ob-servations on the properties of oleic acid and on the relation between saturated and un-saturated fatty acid for lactic acid bacteria.

L-Glutamic Acid Fermentation with Molasses

the correlation of the fatty acid composition of each cell fraction and the extracellular accumulation of L-GA was carried out as follows: 1. Five factors-pH, temperature, the con-centration of lysozyme, the addition of in-organic ion and EDTA-were studied in order to determine the conditions suitable for the cell wall lysis of the L-GA-producing bacteria. The result was that the cell wall of M. ammoniaphilum-A was undoubtedly attacked to some extent, but was doubtful to be completely broken down by the lysozyme. On the contrary, the cell wall lysis of M. ammoniaphilum-C was observed with electron microscope and phase-contrast microscope to be completed under the following conditions: Bacterial cell 4.5mg/ml (as dry weight) m/20 Na-phos- Lysozyme phate buffer (pH 6.2) emp. 30°C 14 hr. Lysozyme 0.33mg/ml NaNO3 0.3Mm/ml, EDTA 0 2. Each of the C strain grown in the glucose-biotin (biotin level: 2.5γ/1 and 20 γ/1) media and beet molasses-POEFE (POEFE level: free and 0.15 g/dl) media was submitted to the cell wall lysis under the above men-tioned conditions. After removal of the cell wall fraction by 6, 000×g, the protoplast was ground by quartz. The cell membrane fraction and “the proto-plasm fraction” were fractionated at 12, 000×g, to prevent the contamination of the cell mem-brane fraction with the protoplasm. Each fraction was hydrolyzed and methylated before being submitted to the gas chromato-graphy. The correlation of the resulting fatty acid composition of each fraction and the extra-cellular accumulation of L-GA was discussed. From these results, following thesis was proposed: A large amount of extracellular accumulation of L-GA took place when the ratio of Sat. FA./Unsat. FA. in the cell mem-brane fraction was above 1 and the ratio in the cell wall and the protoplasm had no direct relation to the L-GA accumulation.

Biochemical Effects of Fatty Acid and its Derivatives on L-Glutamic Acid Fermentation

Biochemical Effects of Fatty Acid and its Derivatives on L-Glutamic Acid Fermentation

Studies on Amino Acid Fermentation

The effect of biotin on glucose catabolism in a l-glutamic acid-producing bacterium, Brevibacterium ammoniagenes, was studied by a modified radiorespirometric method. The amount of glucose metabolized by way of the hexose-monophosphate shunt (HMP) was estimated to be 26% in biotin-deficient cells and 38% in biotin-sufficient cells, respectively. In both cases, the Embden-Meyerhof-Parnas Pathway (EMP) was found to be the major route for glucose catabolism. These results will exclude the present theory at least in our strain that the main route of glucose catabolism is HMP in biotin-deficient cells and EMP in biotin-sufficient ones, and that this is one of the main effect of biotin on l-glutamic acid fermentation.

L-Glutamic Acid Fermentation with Molasses

L-Glutamic Acid Fermentation with Molasses

Studies on Amino Acid Fermentation

The effect of biotin on glucose catabolism in a l-glutamic acid-producing bacterium, Brevibacterium ammoniagenes, was studied by a modified radiorespirometric method. The amount of glucose metabolized by way of the hexose-monophosphate shunt (HMP) was estimated to be 26% in biotin-deficient cells and 38% in biotin-sufficient cells, respectively. In both cases, the Embden-Meyerhof-Parnas Pathway (EMP) was found to be the major route for glucose catabolism. These results will exclude the present theory at least in our strain that the main route of glucose catabolism is HMP in biotin-deficient cells and EMP in biotin-sufficient ones, and that this is one of the main effect of biotin on l-glutamic acid fermentation.

Effects of Corn Steep Liquor and Thiamine on l-Glutamic Acid Fermentation of Hydrocarbons1

Effects of Corn Steep Liquor and Thiamine on l-Glutamic Acid Fermentation of Hydrocarbons1

廃糖蜜によるL-グルタミン酸醗酵(第3報)

甜菜廃糖蜜を糖源とするL-GA生産菌Microbacterium ammoniaphilumによるL-GAの菌体外著量蓄積にはPOEFEの添加が必要であるが,その存在がL-GA著量蓄積のために必要とされる時間について検討した結果, (1) POEFE(ノニオンS-6)はフラスコ段階において,添加濃度の如何によらず培地調製時添加が有効であり,培養3.5時間,すなわち培地上清中ビオチン残存量が1γ/l程度までの添加は, L-GA蓄積性への寄与に変りは認められなかったが,培養5時間以降は急速にL-GA蓄積への寄与性が低下し,特に9時間,すなわち菌体へのビオチン吸収終了後は極めて小さい.この現象はPOEFEの濃度を水溶性を保ちながら十分大きくしても変らない.これらのことは,フラスコ段階におけるL-GAの菌体外蓄積が培養7時間頃から始ることを考慮すれば, POEFEが単純な物理化学的な細胞膜透過性改善剤ではないと推定できる. (2) 培地調製時に添加したPOEFEは,培養約4時間,すなわち培地上清中のビオチン残存量が約1γ/lまでは培地中に存在することが必要であるが,約7時間,すなわちビオチンの菌体への吸収が終了する時間以降は存在する必要は認められなかった. (3) POEFEの添加時間及び存在必要時間を,従来L-GAの菌体外蓄積剤と認められているペニシリン及びCTABの最適添加時間と比較検討した結果,両者間の相異が明かとなり,両者のL-GA菌体外蓄積の機構は別個のものであることが推定される. (4) ノニオンS-6添加量とO.D.の関係を検討し,さらにL-GA著量蓄積条件下で生菌数を測定し, L-GAを蓄積せしめる濃度においては,菌生育,増殖に対し阻害的であることは認められなかった.

Biochemical Effects of Fatty Acid and its Derivatives on L-Glutamic Acid Fermentation

It has been found that although Brevibacterium lactofermentum No. 2256 is incapable of accumulating l-glutamic acid in a biotin sufficient medium, it produces a large quantity of the acid in the presence of sucrose fatty acid ester. In a biotin deficient medium, however, the ester brought the unfavorable diminution of l-glutamic acid accumulation caused by the decrease of glucose consumption in an incubation period. The undesirable effects were practically lost when the ester was added to the culture medium after more than eight hours in the course of incubation. This fact suggests that the ester is concerned with the growth of microorganism. It is very interesting to elucidate the interrelation between sucrose fatty acid ester and biotin. For the maximum accumulation of l-glutamic acid corresponding increase in amount of the ester to the increasing concentration of biotin was necessary. The proportional relation did not extend to excedingly high levels of the two implicating factors. The further observat...

廃糖蜜によるL-グルタミン酸醗酵(第2報)

廃糖蜜によるL-グルタミン酸醗酵(第2報)

Biochemical Effects of Fatty Acid and its Derivatives on L-Glutamic Acid Fermentation

It has been found that although Brevibacterium lactofermentum No.2256 is incapable of accumulating L-glutamic acid in a biotin sufficient medium, it produces a large quantity of the acid in the presence of sucrose fatty acid ester. In a biotin deficient medium, however, the ester brought the unfavorable diminution of L-glutamic acid accumulation caused by the decrease of glucose consumption in an incubation period. The undesirable effects were practically lost when the ester was added to the culture medium after more than eight hours in the course of incubation. This fact suggests that the ester is concerned with the growth of microorganism. It is very interesting to elucidate the interrelation between sucrose fatty acid ester and biotin. For the maximum accumulation of L-glutamic acid corresponding increase in amount of the ester to the increasing concentration of biotin was necessary. The proportional relation did not extend to excedingly high levels of the two implicating factors. The further observations concerning the effects of the individual fatty acid esters such as sucrose stearate remain unsatisfactory.

グルタミン酸醗酵液のグルタミンに就て(第1報)

グルタミン酸醗酵液のグルタミンに就て(第1報)

Studies on L-Glutamic Acid Fermentation

The authors have carried out a series of studies on L-glutamic acid fermentation with astrain of Brevibacterium divaricatum nov. sp. in the previous papers.In this paper, some metabolism of L-glutamic acid and oxidative decomposition of severalorganic acids concerning the tricarboxylic acid cycle by the resting cells have been studied.The results suggest that L-glutamic acid is one of the final fermentative products of thisbacterium, and the tricarboxylic acid cycle is working as a glutamic acid forming cycle.The presence of glucokinase, phosphoglucoisomerase, phosphofructokinase, aldolase, DPNlinkedglyceraldehyde-3-phosphate dehydrogenase, and TPN-linked glucose-6-phosphate dehydrogenase in cell-free extracts of this bacterium was also demonstrated.

L-GLUTAMIC ACID AND SUCCINIC ACID FERMENTATION BY BREVIBACTERIUM FLAVUM NO. 1996

It has been recognized that the conversion from L-glutamic acid fermentation to succinic acid fermentation can be performed with Brevibacterium flavum No. 1996.Succinic acid can be directly produced from glucose at a high yield of 28.3% for initial glucose in submerged culture (35.8% for consumed glucose).The optimal aeration rate and the optimal concentration of C.S.L. for the succinic acid production were 0.4 to 0.7×10-6 of Kd (g-mol/cc. min. atm.) and 1.0 to 2.0% of C.S.L., respectively.The activity of succinic decomposing enzyme in cell grown under the succinic acid producing condition was more feeble than that under the L-glutamic acid producing condition.Among the L-glutamic acid producing bacteria, genus Brevibacterium was able to perform this conversion of fermentations, while genus Bacillus was not.