Simple photometric and fluorometric assays are insufficient for assessing UV-induced mutagenesis in genes controlling tryptophan synthesis in Escherichia coli

A new approach to detecting induced mutations was tested based on the assay of cell extracts and special growth media following cultivation of UV irradiated Escherichia coli cells. No correlation was found between the UV dose and the optical densities of cultural media or cell extracts prepared by Triton X-100 treatment. Blue fluorescence of concentrated cultural media varied with cell dose, according to a rather complex law, which differed substantially from the known dose-effect curves for induced mutations. Nevertheless, a certain extent of the brown staining of tryptophan containing medium could, presumably, serve as a quite sensitive indicator of the integral metabolic activity of bacteria grown in the medium. Besides, we observed that overnight lag phase cultures became gradually more transparent, when analysed in the spectrophotometer cuvette just after their dilution with fresh medium.     

Retinoic acid modification of cell culture used for reproduction of enteropathogenic viruses

The 0.001-0.005% retinoic acid injection into the growth cultural medium of prime and continue cell cultures 12-24h before inoculation considerably raised the cell sensitivity to animal entero- and coronaviruses. The entero- and coronaviruses concentrations in cultural medium increased by 10(1.58) and 10(1.68)TCID 50/1.0 respectively. The optimized parameters of the cell culture processing for the enteropathogenic viruses reproduction improvement are proposed.     

Antimicrobial activity of Laetiporus sulphureus strains grown in submerged culture]

Cultural conditions for growth and fruit body formation were elaborated to four strains of Laetiporus sulphureus isolated from nature. All strains demonstrated antimicrobial activity against a wide spectrum of gram-positive and gram-negative bacteria during agar and submerged cultivation including methicillin-resistant strain of Staphylococcus aureus (MRSA) and glycopeptide-resistant strain of Leuconostoc mesenteroides. Antifungal activity was not found. The level of antimicrobial activity during submerged cultivation reached maximum after seven days of growth on specific medium with soybean meal and corn liquid; the next four weeks its increasing was not so manifested. Antimicrobial activity correlated with orange pigment secretion and cultural liquid acidification to pH 2.0-2.8 that indicates on acid nature of synthesized products.     

微生物法生产丙烯酰胺的研究（Ⅰ）—腈水合酶产生菌株的培养和高活力的表达

研究了丙烯酰胺生产菌株的培养条件。通过对培养过程pH值调控、培养基补料以及诱导剂加入量的研究,使发酵液的腈水合酶的活力达到了6567u/mL菌液。这一酶活是国内外所见报道中最高的。进一步进行了丙烯腈的酶催化水合实验,产物中并没有发现副产物丙烯酸,说明在提高腈水合酶的同时,酰胺酶的活力并没有明显体现这一试验结果为工厂化生产改造以及新工艺的研究打下了基础。     

Phosphatase production byAspergillus ficuum

Summary Effects of nutritional and cultural conditions on cell growth and phosphatase production byAspergillus ficuum were studied.A. ficuum produced high levels of phosphatases when grown on a basal medium that contained a minimal amount (2 mg/100 ml) of phosphorus in an acidic growth medium. The organism produced a nonspecific acid phosphomonoesterase rather than phytin-specific phosphatase. The enzyme hydrolyzed a variety of phosphates and produced orthophosphate. The rate of phosphate hydrolysis was dependent on the pH of the reaction, where the pH optimum for acid phosphatase was 2.5 and that for phytase was 5.0. The organism slowly released the phosphatase, and the enzyme activity in the growth medium increased continually during a one-month growth period. For a high level of phosphatase production, low levels (1–5 mg%) of initial phosphorus were necessary and polyphosphates were the desired form rather than the monophosphate. The addition of surfactants, such as polyoxyethylene ethers and sodium oleate, to fungal culture medium markedly increased the level of phosphatase production.     

Changes in inducibility of ornithine decarboxylase activity in differentiating human neuroblastoma cells

Human SH-SY5Y neuroblastoma cells could be induced to differentiate morphologically and biochemically in the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA), retinoic acid (RA), or a combination of these two substances. The phenotypical changes induced by these substances differed, but one effect of both was an inhibition of the cell growth. Addition of TPA or RA to non-treated cells had no effect on the activation of ornithine decarboxylase (ODC, EC 4.1.1.17.), while a change to fresh medium stimulated the ODC to maximum activity after 4-6 h. The activity was not altered by the presence of RA in the fresh medium, but TPA partially inhibited the medium-stimulated ODC activity. Cells treated for 4 or 8 days with TPA or a combination of TPA and RA had a low ODC activity which could not be induced by fresh medium. However, RA-treated (and thus growth-inhibited) cells still responded to a change of medium by exhibiting an ODC activity of the same magnitude and duration as in medium-stimulated control cells. The results seem to suggest that the growth inhibition induced by TPA and RA, respectively, is mediated by different mechanisms.     

运用氨基酸分析法和DOE法优化抗PD-1单克隆抗体生产用培养基

本研究采用氨基酸分析法结合DOE设计法优化并获得高表达抗PD-1单克隆抗体生产用基础和补料培养基。通过对市售多种基础和补料培养基进行筛选,获得细胞生长状况较优的基础培养基和抗体表达较高的补料培养基,利用氨基酸分析法检测较优基础培养基和补料培养基中氨基酸消耗情况,确定影响细胞生长和抗体表达的关键氨基酸种类,利用DOE分析软件设计分别在较优基础和补料培养基中添加不同浓度的氨基酸种类及浓度,根据细胞生长及抗体表达,优化得到抗PD-1单克隆抗体的基础和补料培养基组合。最终优化后基础培养基配方为:Hycell CHO培养基中添加1.04 mmol/L L-天冬酰胺和0.76 mmol/L L-谷氨酰胺。最终优化后补料培养基配方为:OPM CHOCD Feed1补料培养基中添加38.7 mmol/L L-组氨酸,75.0 mmol/L L-酪氨酸,64.0 mmol/L L-丝氨酸,49.2 mmol/L L-谷氨酰胺和18.7 mmol/L L-半胱氨酸。经过3 L反应器培养验证,优化后的培养基比未优化时,最大活细胞密度(PVCD)提高了62.7%,抗PD-1单克隆抗体表达量提高了71.5%,且活性无明显差异。     

Effect of sodium dodecylbenzenesulfonate and antifoaming agents on the endonuclease activity of Serratia marcescens

The effect of sodium dodecylbenzene sulfonate (sulfonol) and certain froth breakers on the activity of endonuclease was studied in the cultural broth of Serratia marcescens in order to find out whether sulfonol could be used for limiting the infection. Sulfonol was found to have no effect on the cultural growth; it increased the activity of endonuclease in the cultural broth, and the peak of the activity appeared earlier than in the control medium. Propanol B-400 was shown to be the best froth breaker.     

Fatty Acid Composition of Escherichia coli as a Possible Controlling Factor of the Minimal Growth Temperature.

Shaw, Maxwell K. (University of California, Davis), and John L. Ingraham. Fatty acid composition of Escherichia coli as a possible controlling factor of the minimal growth temperature. J. Bacteriol. 90:141-146. 1965.-If Escherichia coli ML30 is shifted from 37 to 10 C during exponential growth in glucose minimal medium, a 4.5-hr lag results. During this lag, the proportion of unsaturated fatty acids increases in the cellular lipids. However, the adjustment of the fatty acid composition does not appear to be prerequisite to growth at 10 C. If shifts are made to 10 C into minimal medium containing glucose after starvation for glucose at 37 C for 0.5 and 16 hr, the lag periods at 10 C are 4.5 and 6 hr, respectively. Withholding glucose during the lag periods does not affect the duration of the lag periods, but no change in fatty acid composition occurs if glucose is not present. Supplementing the medium with glucose after the lag period permits immediate growth at 10 C; however, the fatty acid composition is still typical of cells grown at 37 C. It is concluded that the fatty acid composition of cells does not determine the minimal temperature of growth.     

Process development with nitrogenase-producing Escherichia coli C-M74 (pUS1) CellS

A process for the continuous fermentation of the genetically modified, nitrogenase-producing Escherichia coli C-M74 (pUS1)-strain has been developed. This strain, which is able to fix molecular nitrogen, has the nifgenes of the bacterium Klebsiella pneumoniae. Cell growth and nitrogenase activity of the enzyme have been optimized both in batch and continuous fermentations. For the fermentations, trial runs were performed by cultivating the E. coli cells in 50-ml culture bottles. The medium composition was varied in order to provide high biomass production and nitrogenase activity. For an effective fermentation control, an on-line analysis was built up for the substrates ammonium and glucose. Other medium components such as ampicillin, citric acid, acetic acid, nitrogenase activity, and protein were measured by using different off-line methods. Modern optical methods like in-line microfluorometry for monitoring the culture fluorescence and laser flow cytometry for the estimation of DNA and protein content were also employed. Plasmid stability was also determined.     

Derepression of certain aromatic amino acid biosynthetic enzymes of Escherichia coli K-12 by growth in Fe3+-deficient medium.

3-Deoxy-arabino-heptulosonic acid 7-phosphate synthase, prephenate dehydratase, tryptophan synthase, and 2,3-dihydroxybenzoylserine synthase enzyme activities are derepressed in wild-type Escherichia coli K-12 cells grown on Fe3+-deficient medium. This derepression is reversed when FeSO4 is added to the growth medium. Addition of shikimic acid to the Fe3+-deficient growth medium caused repression of the first three enzyme activities but not of 2,3-dihydroxybenzoylserine synthase activity. Addition of 2,3-dihydroxybenzoic acid to the Fe3+-deficient growth medium has no effect on any of the above-mentioned enzyme activities. The Fe3+ deficiency-mediated derepression of 3-deoxyarabino-heptulosonic acid 7-phosphate synthase activity is due to an elevation of the tyrosine-sensitive isoenzyme; the phenylalanine-sensitive isoenzyme is not derepressed under these conditions.     

Antimicrobial actions of degraded and native chitosan against spoilage organisms in laboratory media and foods

The objective of this study was to determine whether chitosan (poly-beta-1,4-glucosamine) and hydrolysates of chitosan can be used as novel preservatives in foods. Chitosan was hydrolyzed by using oxidative-reductive degradation, crude papaya latex, and lysozyme. Mild hydrolysis of chitosan resulted in improved microbial inactivation in saline and greater inhibition of growth of several spoilage yeasts in laboratory media, but highly degraded products of chitosan exhibited no antimicrobial activity. In pasteurized apple-elderflower juice stored at 7 degrees C, addition of 0.3 g of chitosan per liter eliminated yeasts entirely for the duration of the experiment (13 days), while the total counts and the lactic acid bacterial counts increased at a slower rate than they increased in the control. Addition of 0.3 or 1.0 g of chitosan per kg had no effect on the microbial flora of hummus, a chickpea dip; in the presence of 5.0 g of chitosan per kg, bacterial growth but not yeast growth was substantially reduced compared with growth in control dip stored at 7 degrees C for 6 days. Improved antimicrobial potency of chitosan hydrolysates like that observed in the saline and laboratory medium experiments was not observed in juice and dip experiments. We concluded that native chitosan has potential for use as a preservative in certain types of food but that the increase in antimicrobial activity that occurs following partial hydrolysis is too small to justify the extra processing involved.     

Selection of medium for esterase synthesis by Mycobacterium album by the method of mathematical planning of experiments]

The composition of a defined medium for the growth of Mycobacterium album Sohngen 726 was selected by the method of mathematic planning of the experiment. The specific activity of esterase during the growth of the culture on this medium is by 30 per cent higher than on the original medium.     

A basal-defined medium for the study of proteolytic activity of Serratia marcescens.

A simple defined basal medium is presented for the study of proteolytic activity, induction and repression, and protease purification with Serratia marcescens. Since the medium contains no protein, it does not interfere with or present artifact to protein assays, column chromatography, or electrophoresis. The medium consists of the basal salts and buffer medium of Bromke and Hammel (1979) plus a carbon-energy source such as glycerol, calcium chloride for the cation requirement for protease activity, and an amino acid, preferably leucine. Growth parameters and proteolytic activities are presented for unsupplemented medium and for the medium supplemented with each of 18 amino acids. Unsupplemented medium completes the logarithmic phase in 12.5 h of incubation and has a constitutive level of proteolytic activity. Supplementation with any amino acid, except cysteine and tryptophan, increases significantly the proteolytic activity, but has a varied effect on growth parameters.     

Investigation of the biological activity of solutions activated electrochemically in a membrane electrolyzer

The biological activity of the catholyte and anolyte of double distilled water was studied in experiments on the germination of wheat grains in the period from March to May. The activity of the solutions, which was characterized by a growth index, was high early in this period, then decreased almost to zero in the middle of the period, and then increased to about the initial value by the end of the period. Throughout, the efficiency of the anolyte of double distilled water generally exceeded the efficiency of the catholyte. Early and late in the period, the stimulatory effect of the anolyte exceeded that of the catholyte by a factor of 5–5.5. The changes in the biological activity of the catholyte and anolyte of double distilled water were also compared with the changes in the biological activity of the catholyte of nutrient medium M9. The stimulatory effect of the catholyte of the nutrient medium was evaluated from the change in the growth of E. coli cells. Early in the period at a cultivation temperature of 20°C, the stimulatory effect determined from the increase in the optical density of the cell suspension in the experiment with respect to a reference value was 55–60%. Next, the stimulatory effect decreased almost to zero in the middle of the period and increased to approximately initial value by the end of the period. It was assumed that the physicochemical mechanisms of action of the catholyte and anolyte of double distilled water on the wheat seed germination and of the catholyte of the nutrient medium on E. coli cell growth are of different nature.     

Evidence for the inhibition of platelet-derived growth factor induced rat smooth muscle cells DNA synthesis by fenofibric acid at the Go/G1 cell cycle level

It has been proposed that a platelet-derived growth factor (PDGF) may play an important role in the genesis of atherosclerosis by promoting proliferation of smooth muscle cells. The present study shows evidence that fenofibric acid inhibits the growth promoting activity of PDGF on cultured smooth muscle cells.When smooth muscle cells are in a quiescent state by feeding them a PDGF -and lipoproteins- deficient medium, addition of a platelet extract induces DNA synthesis in a synchronous fashion. A 6 hours' exposure of cells to this extract is sufficient for this effect. Fenofibric acid could inhibit this synthesis when the compound was present in the culture medium concomitantly with platelet extract. This result suggests that fenofibric acid target is a cellular event attendant on PDGF-induced growth promotion. Fenofibric acid is a hypolipidemic drug which inhibits 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (HMGCoA reductase) activity, the limiting step of endogenous cholesterol synthesis. If any endogenous cholesterol is available, smooth muscle cell cholesterol needs for proliferation can be supplied by LDL-cholesterol. As the addition of LDL to the culture medium did not overcome fenofibric acid inhibition, this demonstrates a cholesterol independent mechanism for the anti-PDGF growth promoting activity.The fact that fenofibric acid, a hypolipidemic drug, can also inhibit growth promoting activity of PDGF suggests that this drug can be effective on the prevention of atherosclerosis and cardiovascular diseases by at least two independent mechanisms.     

Composition of Methanospirillum hungatii GP1 during growth on different media

Growth of Methanospirillum hungatii GP1 as determined by optical density measurement wsa comparable to growth assessed by cell dry weight, ribonucleic acid content, and deoxyribonucleic acid content. Cultivation of M. hungatii on synthetic medium containing mineral salts, vitamins, and acetic acid indicated that, on a dry weight basis, cell constituents such as protein (71%), ribonucleic acid (15.8%), deoxyribonucleic acid (1.6%), and total carbohydrate (3.2%) did not vary significantly with the growth phase. Cells grown in the synthetic medium supplemented with yeast extract and tryptone had slightly higher protein content (76%), but the concentrations of the other cell constituents were similar and did not fluctuate much during growth. Nitrogen limiting growth resulted in somewhat lower ribonucleic acid content as well as slightly higher protein content than that in cells grown in nonlimiting medium. Methanospirillum hungatii did not accumulate any of the commonly known reserve materials under nitrogen or carbon and hydrogen limiting growth.     

Response surface optimization of the critical medium components for carbonyl reductase production by Candida viswanathii MTCC 5158

Culture conditions were optimized for the growth and carbonyl reductase production by a novel yeast strain Candida viswanathii. Response surface methodology was applied for the critical medium components (initial pH, mannitol, yeast extract and calcium chloride) identified earlier by one-factor-at-a-time approach. Central composite design was used for the optimization studies. Using this methodology, the optimal values for the concentration of mannitol, initial pH, yeast extract and calcium chloride were 1.9, 7.5, 1.6 and 4, respectively. This medium was projected to produce, theoretically, growth having an optical density of 1.1 (600 nm) and an enzyme activity of 81.5 U/ml. Using this optimized medium, an experimental growth of 1.1 OD (600 nm) and enzyme activity 80.9 U/ml verified the applied methodology. This approach for medium optimization led to an enhancement of the growth and enzyme activity by 1.3 and 2.3 times higher, respectively, as compared to the unoptimized media.     

Antimicrobial Actions of Degraded and Native Chitosan against Spoilage Organisms in Laboratory Media and Foods

The objective of this study was to determine whether chitosan (poly-β-1,4-glucosamine) and hydrolysates of chitosan can be used as novel preservatives in foods. Chitosan was hydrolyzed by using oxidative-reductive degradation, crude papaya latex, and lysozyme. Mild hydrolysis of chitosan resulted in improved microbial inactivation in saline and greater inhibition of growth of several spoilage yeasts in laboratory media, but highly degraded products of chitosan exhibited no antimicrobial activity. In pasteurized apple-elderflower juice stored at 7°C, addition of 0.3 g of chitosan per liter eliminated yeasts entirely for the duration of the experiment (13 days), while the total counts and the lactic acid bacterial counts increased at a slower rate than they increased in the control. Addition of 0.3 or 1.0 g of chitosan per kg had no effect on the microbial flora of houmous, a chickpea dip; in the presence of 5.0 g of chitosan per kg, bacterial growth but not yeast growth was substantially reduced compared with growth in control dip stored at 7°C for 6 days. Improved antimicrobial potency of chitosan hydrolysates like that observed in the saline and laboratory medium experiments was not observed in juice and dip experiments. We concluded that native chitosan has potential for use as a preservative in certain types of food but that the increase in antimicrobial activity that occurs following partial hydrolysis is too small to justify the extra processing involved.     

Studies on the activities of an acid phosphomonoesterase and an alkaline pyrophosphatase during the growth of Physarum polycephalum.

After an initial decrease, the specific activity of Physarum polycephalum acid phosphomonoesterase increases during the growth of the organism in an axenic medium. This increase is independent of the inorganic phosphate concentration in the culture medium. The specific activity of inorganic alkaline pyrophosphatase remains constant during the growth and is not modified by a high extracellular concentration of orthophosphate. During starvation in a non nutritive saline medium, the increase of acid phosphatase activity is immediate whereas pyrophosphatase activity remnins constant.