Genetical studies on the heterocyst and nitrogen fixation of the blue-green alga Nostoc muscorum

Summary The short-trichome-forming, non-heterocystous and non-nitrogen-fixing (het ^– nif^–) mutant of the nitrogen-fixing blue-green alga Nostoc muscorum was isolated by N-methyl-N-nitro-N-nitrosoguanidine (NTG)-mutagenesis after penicillin enrichment technique and characterized. The mutant did not grow and fix nitrogen in combined-nitrogen-free medium while in nitrate-containing medium it grew well (K=0.112/day, G=64.27 h), although its growth was comparatively poor than the parent alga (K=0.128/day; G=56.14 h). The mutant was stable and both the het ^– and nif ^– characters reverted to wild type (het ⁺ nif⁺) with the reversion frequency of 2.62×10^-7.The het ^– nif^– mutant tolerated 0.5 g/ml of streptomycin sulphate on the agar medium and its streptomycin resistant mutant capable of growing in presence of 10g/ml of streptomycin was isolated spontaneously with a frequency of 1.45×10^-8. These streptomycin resistant isolates (het ^– nif^– str^R) resisted 100 g/ml of streptomycin sulphate on the agar medium and 200 g/ml in liquid medium. Spontaneous virus-resistant mutant of het ^– nif^– str^R was isolated with a mutation frequency of 4.02×10^-4.The data of genetic recombination experiments suggested that there is transfer of both het and nif genes to het ^– nif^– strain with the frequency of 2×10^-6 to 2×10^-5 simultaneously. There was increase in recombination frequency with increasing the incubation period. The virus-resistance marker is also transferred to the sensitive recipient.Abbreviations CFU colony forming units - C–N Chu-10 medium without combined nitrogen - C+N Chu-10 medium with 0.232 g/l calcium nitrate - G generation time - het heterocyst differentiating genes - K specific growth rate constant - MOI multiplicity of infection - nif nitrogen-fixing genes - NTG N-methyl-N-nitro-N-nitrosoguanidine - PFU plaque forming units - str ^R streptomycin resistance - str ^R streptomycin sensitive     

Biotechnological production of unnatural L-amino acids from D,L-5-monosubstituted hydantions. II. L-α- and L-β-naphthylalanine

Summary Resting cells ofArthrobacter sp. (DSM 3745) with the ability to form L-tryptophan from D,L-5-(3-indolylmethy)hydantoin were used for the bioconversion of D,L-5-- and D,L-5--naphthylmethylhydantoin (D,L-5-- and D,L-5--NMH) to the corresponding L-amino acids. Under the optimal reaction conditions of pH 9.7 and 40°C specific productivities of 0.2 (-naphtylalanine) and 0.6 (-naphtylalanine) mM amino acid x g cell dry mass^–1 x h^–1 were obtained in a 0.1 M Na₂CO₃/NaHCO₃-buffer in a strirred bioreactor.     

Model aided design of repeated fed-batch penicillin fermentation

Structured models of antibiotic fermentation that quantify maturation and aging of product forming biomass are fitted to experimental data. Conditions of superiority of repeated fed batch cultivation are characterized on the basis of a performance criterion that includes penicillin productivity and costs of operation. Emphasis is placed on the relevance of such research to the model aided design of optimal cyclic operation.List of Symbols c IU/mg cost factor - D s^–1 dilution rate - J IU · cm^–3 · h^–1 net productivity - k _p IU · mg^–11 · h^–1 specific product formation rate - k _pm IU · mg^–1 · h^–1 maximum specific product formation rate - p IU/cm³ concentration of penicillin - T s final time of fermentation - t s fermentation time - X kg/m³ concentration of biomass dry weight - X ₁kg/m³ concentration of young, immature biomass - X ₂ kg/m³ concentration of mature product forming biomass - X _c kg/m³ biomass concentration of the end of growth phase - X _mkg/m³ maximum biomass concentration Greek Letters s^–1 specific maturation rate - s^–1 specific aging rate - s^–1 specific growth rate - _m s^–1 maximum specific growth rate - _p s^–1 specific growth rate during the product formation phase - s cycle time - % volume fraction of draw-off Abbreviations CC chemostat culture - RFBC repeated fed batch culture - RBC repeated batch culture     

Diverse effects of essential (n−6 andn−3) fatty acids on cultured cells

Fatty acids (FAs) have long been recognized for their nutritional value in the absence of glucose, and as necessary components of cell membranes. However, FAs have other effects on cells that may be less familiar. Polyunsaturated FAs of dietary origin (n–6 andn–3) cannot be synthesized by mammals, and are termed essential because they are required for the optimal biologic function of specialized cells and tissues. However, they do not appear to be necessary for normal growth and metabolism of a variety of cells in culture. The essential fatty acids (EFAs) have received increased attention in recent years due to their presumed involvement in cardiovascular disorders and in cancers of the breast, pancreas, colon and prostate. Manyin vitro systems have emerged which either examine the role of EFAs in human disease directly, or utilize EFAs to mimic thein vivo cellular environment. The effects of EFAs on cells are both direct and indirect. As components of membrane phospholipids, and due to their varying structural and physical properties, EFAs can alter membrane fluidity, at least in the local environment, and affect any process that is mediated via the membrane. EFAs containing 20 carbons and at least three double bonds can be enzymatically converted to eicosanoid hormones, which play important roles in a variety of physiological and pathological processes. Alternatively, EFAs released into cells from phospholipids can act as second messengers that activate protein kinase C. Furthermore, susceptibility to oxidative damage increases with the degree of unsaturation, a complication that merits consideration because lipid peroxidation can lead to a variety of substances with toxic and mutagenic properties. The effects of EFAs on cultured cells are illustrated using the responses of normal and tumor human mammary epithelial cells. A thorough evaluation of EFA effects on commercially important cells could be used to advantage in the biotechnology industry by identifying EFA supplements that lead to improved cell growth and/or productivity.Abbreviations AA arachidonic acid (20 carbons: 4 double bonds,n–6) - BHA butylated hydroxyanisole - BHT butylated hydroxytoluene - cAMP cyclic adenosine monophosphate - CHO Chinese hamster ovary - DAG diacylglycerol - DGLNA dihomo--linolenic acid (203,n–6) - DHA docosahexaenoic acid (226,n–3) - EFA essential fatty acid - EGF epidermal growth factor - EGFR epidermal growth factor receptor - EPA eicosapentaenoic acid (205,n–3) - FA fatty acid - FBS fetal bovine serum - GLNA -linolenic acid (183,n–6) - LA linoleic acid (182,n–6) - LNA -linolenic acid (183,n–3) - LT leukotriene - MDA malondialdehyde - NAD nicotinamide adenine dinucleotide - NDGA nordihydroguaiaretic acid - OA oleic acid (181,n–9) - PG prostaglandin - PKC protein kinase C - PUFA polyunsaturated fatty acid - SFM serum-free medium - TX thromboxane     

Determination of specific monoclonal antibody secretion rate during very slow hybridoma growth

A total cell recycling suspension perfusion reactor has been constructed for investigation of specific monoclonal antibody secretion rate of the 9.2.27 murine hybridoma under conditions of a very low growth rate. By rapidly recycling hybridoma cells using a thermostated tangential flow filter, 3.6 mg cell dry weight/cm³ could be maintained at growth rate of <0.05 _max for over 250 h. Under these conditions, secretion of lactate, ammonia and l-alanine were directly related to the rate of l-glutamine supply. Monoclonal antibody accumulated in the reactor to levels in excess of 1400 g/ cm³. Surprisingly, as specific growth rate decreased, the specific immunoglobulin secretion rate remained constant, implying that monoclonal antibody synthesis could be uncoupled from growth.List of Symbols CMF cm³/(min · cm²) cross membrane flow rate - D h^–1 dilution rate - DOT % air saturation dissolved oxygen tension - F _R cm³/min perfusion rate - GlcPR mg/min glucose provision rate - GlnPR mg/min l-glutamine provision rate - N _A mmoles O₂/(dm³ · h) oxygen transfer rate - q _ala mmoles/h l-alanine secretion rate - q _MAB mg MAB · 10^–8 viable cells ^–1 · day^–1 specific MAB secretion rate by viable cells - ¯q _MAB (dimensionless) ¯q _MAB/¯q _{MAB MAX} - ¯q _{NH 3} mmoles/h ammonia secretion rate - S _R mg/cm³ limiting substrate concentration - h^–1 specific growth rate - _app h^–1 apparent growth rate - ¯ (dimensionless) / _MAX - VC cells/cm³ viable cell number     

The economic production of anti-microbial factor from human promyelocytes in low serum containing medium under chemostat cultivation

It proves that a purifed Anti-Microbial Factor (AMF) from human promyelocytes has strong activity on Gram(–) and Gram(+) bacteria, showing 0.5 (g/ml) of Minimal Bacterical Concentration (MBC) on bothE. coli andS. aureus. For mass production of AMF, chemostat cultivation is recommended to accumulate cells out of the reactor since it is an intracellular protein and its system requires only 1% serum in the medium. Its production process proves to be closely growth-related. 1.7×10^–8 (g/viable cell/day) of maximum specific AMF production rate is estimated at 0.026 h^–1 of dilution rate, maintaining 6×10⁶ (viable cell/ml). Ca. 300 (mg/ml) of crude AMF can be obtained for 50 days of continuous cultivation under optimal conditions. The cell growth reaches relatively fast steady state.     

Cell recycling studies for α-amylase production by Bacillus amyloliquefaciens

Production of -amylase by a strain of Bacillus amyloliquefaciens was investigated in a cell recycle bioreactor incorporating a membrane filtration module for cell separation. Experimental fermentation studies with the B. amyloliquefaciens strain WA-4 clearly showed that incorporating cell recycling increased -amylase yield and volumetric productivity as compared to conventional continuous fermentation. The effect of operating conditions on -amylase production was difficult to demonstrate experimentally due to the problems of keeping the permeate and bleed rates constant over an extended period of time. Computer simulations were therefore undertaken to support the experimental data, as well as to elucidate the dynamics of -amylase production in the cell recycle bioreactor as compared to conventional chemostat and batch fermentations. Taken together, the simulations and experiments clearly showed that low bleed rate (high recycling ratio) various a high level of -amylase activity. The simulated fermentations revealed that this was especially pronounced at high recycling ratios. Volumetric productivity was maximum at a dilution rate of around 0.4 h^–1 and a high recycling ratio. The latter had to exceed 0.75 before volumetric productivity was significantly greater than with conventional chemostat fermentation.List of Symbols a proportionality constant relating the specific growth rate to the logarithm of G (h) - a ₁ reaction order with respect to starch concentration - a ₂ reaction order with respect to glucose concentration - B bleed rate (h^–1) - C starch concentration (g/l) - C ₀ starch concentration in the feed (g/l) - D dilution rate (h^–1) - D _E volumetric productivity (KNU/(mlh)) - e intracellular -amylase concentration (g/g cell mass) - E extracellular -amylase concentration (KNU/ml) - F volumetric flow rate (l/h) - G average number of genome equivalents of DNA per cell - k _l intracellular equilibrium constant - k ₂ intracellular equilibrium constant - k _s Monod saturation constant (g/l) - k ₃ excretion rate constant (h^–1) - k _d first order decay constant (h^–1) - k _gl rate constant for glucose production - k _st rate constant for starch hydrolysis - k _t1 proportionality constant for -amylase production (gmRNA/g substrate) - k ₁ translation constant (g/(g mRNAh)) - KNU kilo Novo unit - m maintenance coefficient (g substrate/(g cell massh)) - n number of binding sites for the co-repressor on the cytoplasmic repressor - Q repression function K₁/K₂Q1.0 - R ratio of recycling - R _s rate of glucose production (g/lh) - r _c rate of starch hydrolysis (g/(lh)) - R _eX retention by the filter of the compounds X: starch or -amylase - r intracellular -amylase mRNA concentration (g/g cell mass) - r _C volumetric productivity of starch (g/lh) - r _E volumetric productivity of intracellular -amylase (KNU/(g cell massh)) - r _r volumetric productivity of intracellular mRNA (g/(g cell massh)) - r _e volumetric productivity of extracellular -amylase (KNU/(mlh)) - r _s volumetric productivity of glucose (g/(lh)) - r _X volumetric productivity of cell mass (g/(lh)) - S ₀ free reducing sugar concentration in the feed (g/l) - S extracellular concentration of reducing sugar (g/1) - t time (h) - V volume (l) - X cell mass concentration (g/l) - Y yield coefficient (g cell mass/g substrate) - Y _E/S yield coefficient (KNU -amylase/g substrate) - Y _E total amount of -amylase produced (KNU) - substrate uptake (g substrate/(g cell massh)) - specific growth rate of cell mass (h^–1) - _d specific death rate of cells (h^–1) - _m maximum specific growth rate of cell mass (h^–1) This study was supported by Bioprocess Engineering Programme of the Nordic Industrial Foundation and the Center for Process Biotechnology, the Technical University of Denmark.     

Summer egg production rates of paracalanid copepods in subtropical waters adjacent to Australia's North West Cape

The biological oceanography of waters adjacent to Australia's North West Cape (21° 49 S, 114° 14 E) was studied during the austral summers of 1997/98 and 1998/99. We measured egg production rate (EPR) by the small paracalanid copepods that dominated the calanoid community. Bottle incubation experiments were conducted at a shallow (20 m) station in the mouth of Exmouth Gulf, and at a shelf-break station (80 m). In 1997/98, we measured EPR by Paracalanus aculeatus, P. indicus, Acrocalanus gracilis and Bestiolina similis, but in 1998/99, we concentrated on P. indicus. Maximal observed EPRs by Paracalanus and Acrocalanus species were 30 eggs female^–1 d^–1, but B. similis attained only 17 eggs female^–1 d^–1. Sporadic measurements of EPR by P. aculeatus minor (maximum 4 eggs female^–1 d^–1) and Parvocalanus crassirostris ( 9 eggs female^–1 d^–1) were also made. However, maximal EPRs were seldom achieved and were often less than 10 eggs female^–1 d^–1. There was no difference between EPR of either P. indicus or B. similis in 1997/98 and 1998/99, despite differences in temperature. Trophic resources severely limit copepod egg production in this area. We suggest that variability and skewness of egg production data derived from individual incubations may be used to judge the degree of food limitation of the population and the variability in feeding success between individuals. The dominance of small copepods and the invariance in their EPR suggest that pulses in physical forcing and subsequent primary production will be severely damped by trophodynamic processes before reaching larval fish.     

Adenosine triphosphate pool levels and endogenous metabolism in Arthrobacter crystallopoietes during growth and starvation

The adenosine triphosphate (ATP) content of Arthrobactery crystallopoietes was measured during growth, starvation and recovery from starvation. During exponential growth of the cells as spheres in a glucose salts medium, the level of ATP per cell remained constant at 8.0×10^-10 g/cell. Morphogenesis to rodshaped cells and an increased growth rate following addition of casein hydrolysate was accompanied by an almost two-fold increase in the ATP level. As division of the rod-shaped cells proceeded, the level of ATP declined. After growing as rods for 12–14 h the cells underwent fragmentation to spheres during which time the ATP level again increased to the original value of 8.0×10^-10 g/cell. As the spherical cells resumed growth on the residual glucose, their ATP content declined for a short period and then remained relatively constant. During starvation of sphere or rod-shaped cells for one week, the ATP level declined by approximately 70% during the first 40–50 h and then remained constant. The endogenous metabolism rate of spherical cells declined during the first 10–20 h of starvation and then remained constant at approximately 0.02% of the cell carbon being utilized per h. Addition of glucose to spherical cells which had been starved for one week increased both the ATP content per cell and their rate of endogenous metabolism. The ATP content fluctuated and then remained at a level higher than maintained during starvation while endogenous metabolism quickly declined.Non-Standard Abbreviations ATP adenosine triphosphate - GS glucose mineral salts - HC casein hydrolysate - PVP polyvinylpyrrolidone - DMSO dimethylsulfoxide - MOPS morpholinopropane sulfonic acid - EDTA ethylene diaminetetraacetic acid     

Cell recovery by continuous flotation

Summary Cell recovery by means of continuous flotation of the Hansenula polymorpha cultivation medium without additives was investigated as a function of the cultivation conditions as well as of the flotation equipment construction and flotation operational parameters. The cell enrichment and separation is improved at high liquid residence times, high aeration rates, small bubble sizes, increasing height of the aerated column, and diameter of the foam column. Increasing cell age and cultivation with nitrogen limitation reduce the cell separation.Symbols C_P cell mass concentration in medium g·l^–1 - C_R cell mass concentration in residue g·l^–1 - C_S cell mass concentration in foam liquid g·l^–1 - V equilibrium foam volume cm³ - V gas flow rate through the aerated liquid column cm³·s^–1 - V_F feed rate to the flotation column ml/min - ¹ V _S/V foaminess s - mean liquid residence time in the column s     

Kinetics of yeast growth and enzyme syntheses in a phosphate-limited continuous culture

Summary The effect of a deficiency of inorganic phosphate on the growth rate and on the invertase and phosphatase activities inSaccharomyces carlsbergensis was studied in a chemostat culture using a synthetic medium in which ethanol was the sole carbon source.The kinetic relationship between the growth rate and both the rates of phosphate uptake and the ethanol consumption agreed well with the threshold model but not the multicative model. The invertase activity of the yeast increased as the dilution rate decreased. As the phosphate concentration in the feed was reduced, the enzyme synthesis increased remarkably. Acid phosphatase activity was repressed completely above a critical molecular ratio, 0.015, of monopotassium phosphate to ethanol in the feed medium. As the phosphate concentration in the feed decreased, the maximum specific enzyme activity increased and the corresponding optimum dilution rate decreased. These experimental changes in enzyme synthesis were expressed mathematically using the modified operon models for enzyme regulation in terms of two fractions of limited inorganic phosphate; one which affects growth and the other which is incorporated in excess by the cells.Nomenclature A ethanol concentration in the culture (mM) - a, b, c, d exponents in the operon model - D dilution rate (h^–1) - E enzyme concentration in the culture (enzyme unit l^–1) - K_a, K_b, K_c, K_d, k equilibrium constants used in the operon model, see Toda (1976b) - o operator gene - P inorganic phosphate concentration in the culture (mM) - Pi limited inorganic phosphate concentration in the cells (mmole inorganic phosphate/g dry weight of cell) - Q specific enzyme activity, no units: (E/X)/(E/X)_max - Q_c, Q_d as defined in Eq. 12 - R repressor - r regulator gene - X cell concentration in the culture (dry cell weight l^–1) Greek Letters molecular ratio of inorganic phosphate to ethanol in the feed medium (mole/mole) - specific growth rate (h^–1) - A specific uptake rate of ethanol (mmole/g cell·h) - P specific uptake rate of inorganic phosphate (mmole/g cell·h) Suffix crit critical value - f feed - max maximum - min minimum - t total - 1, 2 number of species Superfix eff effective for cell growth - exc excess - str structural     

Feeding rate inhibition in crowded Daphnia pulex

Feeding rates of Daphnia pulex fed a range of levels of the alga Chlamydomonas reinhardi of 15 °C are strongly density-dependent. At lower densities, Daphnia (30 1^–1) fed at higher rates than crowded (270 1^–1) Daphnia which manifest a relatively depressed saturation feeding response. At 30 individuals/liter, Daphnia consumed 8.5 – 15.7 × 10⁴ cells d^–1h^–1 (on a volume basis, 12.1 – 22.2 × 10⁶ m³), at 270 L^–1 3.7 – 3.9 × 10⁴ (5.2 – 5.5 = 10⁶ m³ cells d^–1h^–1 when feeding on algae at 80 000 cells ml^–1 (11.3 × 10⁶ m³ ml^–1). The feeding rate data best fit an Ivlev feeding function. An autoallelopath might be causing the repression. Water preconditioned with crowded Daphnia completely repressed feeding in uncrowded Daphnia after six hours.     

Infrared analysis of eleven carrageenophytes from Baja California,Mexico

Infrared analyses of the carrageenan in ten species (representing four genera) of Gigartinaceae and one species of Hypneaceae in different reproductive phases from the northwestern coast of Baja California were studied. Cystocarpic samples of the Gigartinaceae presented varying degrees of a / hybrid. The degree of hybridization was determined based on the ratio between the peak absorbances at 805/845 cm^–1. A high correlation was observed between the 805/845 cm^–1 and 805/970 cm^–1 ratios. Tetrasporic samples of Gigartina leptorhynchos, Iridaea splendens, Rhodoglossum affine and R. roseum, presented a -carrageenan profile, whereas Gigartina tepida, G. exasperata, G. harveyana, G. canaliculata and G. spinosa presented a -carrageenan. The tetrasporic sample of Hypnea valentiae showed a -carrageenan with a very low degree of hybridization.     

Product inhibition during the feeding phasein fed-batch ethanol fermentation of sugar-cane blackstrap molasses

Summary The following equations represent the influence of the ethanol concentration (E) on the specific growth rate of the yeast cells () and on the specific production rate of ethanol () during the reactor filling phase in fed-batch fermentation of sugar-cane blackstrap molasses: = ₀ - k · E and v = v ₀ · K/(K +E) Nomenclature E ethanol concentration in the aqueous phase of the fermenting medium (g.L^–1) - E_m value of E when = 0 or = 0 (g.L^–1) - F medium feeding rate (L.h^–1) - k empirical constant (L.g^–1.h^–1) - K empirical constant (g.L^–1) - M_as mass of TRS added to the, reactor (g) - M_cs mass of consumed TRS (g) - M_e mass of ethanol in the aqueous phase of the fermenting medium (g) - M_s mass of TRS in the aqueous phase of the fermenting medium (g) - M_x mass of yeast cells (dry matter) in the fermenting medium (g) - r correlation coefficient - S TRS concentration in the aqueous phase of the fermenting medium (g.L^–1) - S_m TRS concentration of the feeding medium (g.L^–1) - t time (h) - T temperature (° C) - TRS total reducing sugars calculated as glucose - V volume of the fermenting medium (L) - V₀ volume of the inoculum (L) - X yeast cells concentration (dry matter) in the fermenting medium (g.L^–1) - filling-up time (h) - specific growth rate of the yeast cells (h^–1) - ₀ value of when E=0 - specific production rate of ethanol (h^–1) - ₀ value of when E=0 - density of the yeast cells (g.L^–1) - dry matter content of the yeast cells     

Initiation and growth characteristics of suspension cultures of Calendula officinalis cells

Callus cultures of marigold (Calendula officinalis L.) were induced on Murashige and Skoog medium with different concentrations of auxin (dichlorophenoxyacetic acid (2,4-D) or indole-3-acetic acid (IAA) and cytokinin (kinetin or 6-(,-dimethylallylamino)purine (2iP). Of all hormone combinations used in the medium, two were the most efficient in promoting callus development: 1.81 M (0.4 mg l^–1) 2,4-D and 1.85 M (0.4 mg l^–1) kinetin (0.4d–0.4k culture) or 0.45 M (0.1 mg l^–1) 2,4-D and 2.02 M (0.5 mg l^–1) 2iP (0.1d–0.5p culture). These combinations were selected to induce cell suspension cultures. The suspension cultures were maintained under light or dark conditions. The light stimulated cell aggregation in the cultures. In both cultures cells were undifferentiated under darkness, whereas in the light, rhyzogenesis was observed in 0.1d–0.5p culture. The cell growth and protein and oleanolic acid contents were determined. Initially, biomass production was similar under light and dark conditions, but after 7–8 months from the induction the cell growth was reduced by approximately 30% in the light, whereas the cell growth of the cultures maintained under darkness did not reveal any changes. The presence of oleanolic acid was detected in the suspension cultures kept in darkness. This compound reached two quantitative peaks: in the lag and stationary phases –- beyond the active growth phase of the culture cycle and its concentration was several times higher in 0.1d–0.5p culture than that in 0.4d–0.4k culture. It was for the first time that callus and suspension cultures were induced from the marigold plant.     

Abiotic metal stress enhances diosgenin yield in Dioscorea bulbifera L. cultures

Lower concentrations of CuSO₄ (25–75 M) in the MS medium supplemented with 0.1 mg l^–1 IAA+5.0 mg l^–1 Kn+500 mg l^–1 CH+10 mg l^–1 Cyst hyd enhanced the growth of regenerants of Dioscorea bulbifera L. CuSO₄ (75 M) induced an appreciable diosgenin yield in the regenerants compared to those obtained on media without Cu. The presence of Cu thus seems to stimulate diosgenin production. The regenerants also differentiated bulbils on lower concentrations of Cu. At CuSO₄ (100 M), however, cultures showed poor growth as well as a low diosgenin yield. Increased proline and protein contents were recorded in cultures grown on Cu-enriched media.     

Optimization of recombinantEscherichia coli fed-batch fermentation for bovine somatotropin

Summary Optimum specific growth rate for the production of recombinant bovine somatotropin (bST) was investigated in fed-batchE. coli fermentation with a controlled specific growth rate. Maximum specific bST concentration based on cell mass decreases linearly with the controlled specific growth rate. Productivity of bST is expressed by a quadratic equation as a function of the specific growth rate and has a maximum value at =0.23 hr^–1.     

Inhibition of proliferation of retrovirus-immortalized macrophages by LPS and IFN-γ: Possible autocrine down-regulation of cell growth by induction of IL1 and TNF

The GG₂EE macrophage tumor cell line was previously established by immortalization of C3H/HeJ mouse bone marrow cells with the J2 retrovirus which contains the v-myc and v-raf oncogenes. Studies on the control of GGZEE cell proliferationin vitro have recently been performed. We observed that the combination of 5–25 U/ml recombinant mouse interferon- (rmIFN-) plus 0.03 – 0.3 µg/ml lipopolysaccharide (LPS) markedly inhibited the proliferation of GG₂EE cells (by >95%)in vitro, while either agent alone inhibited only by <40% and 0–10%, respectively. Subsequent studies established that biologically active ILI-like (2–4 U/ml) and TNF-like (50–100 U/ml) activities were released into the supernatants of LPS-treated GG₂EE cells. The combination of IFN- + LPS induced more (6–8 U/ml) ILI release. These results suggested that the inhibition of proliferation of GG₂EE cells by IFN- + LPS could have been mediated in part by cytokines produced by the cells themselves. rhIL1 at a concentration of 10 U/ml inhibited GG₂EE proliferation by 25–30%, while rmIFN- (25 U/ml) + rhIL1 (10 U/ml) inhibited proliferation by 98%. Thus, 10 U/ml rhIL1 could completely replace LPS in the LPS + rmIFN- combination. Further, the combination of low doses of rhIL1 (0.1 to 1 U/ml) plus rmTNF (250 U/ml), which together inhibited proliferation by <20% synergized with doses of 5 to 25 U/ml rmIFN- to inhibit proliferation of GG₂EE cells by 98–99%. These results suggest that cytokines produced by the cells themselves can synergize with rmIFN- to inhibit the oncogene-driven proliferation of GG₂EE cells.     

New expanded bed adsorbents for the recovery of DNA

A 20–40 m pellicular high density (3.7 g cm^–3) expanded bed material has been designed for the capture of DNA and other large macromolecules. Anion exchangers fashioned out of these supports exhibited dramatically enhanced DNA binding capacities over commercial anion exchange adsorbents (6 mg ml^–1, c.f. 50 g ml^–1 at 10% breakthrough), due to a combination of small particle and fuzzy surface architecture created through the coupling of polyethylene imine chains.     

Interrelationships Between Nitrogen Supply and Photosynthetic Parameters in Vicia faba L.

We determined for Vicia faba L the influence of nitrogen uptake and accumulation on the values of photon saturated net photosynthetic rate (P _Nmax), quantum yield efficiency (), intercellular CO₂ concentration (C _i), and carboxylation efficiency (C _e). As leaf nitrogen content (N_L) increased, the converged onto a maximum asymptotic value of 0.0664±0.0049 mol(CO₂) mol(quantum)^–1. Also, as N_L increased the C _i value fell to an asymptotic minimum of 115.80±1.59 mol mol^–1, and C _e converged onto a maximum asymptotic value of 1.645±0.054 mol(CO₂) m^–2 s^–1 Pa^–1 and declined to zero at a N_L-intercept equal to 0.596±0.096 g(N) m^–2. fell to zero for an N_L-intercept of 0.660±0.052 g(N) m^–2. As N_L increased, the value of P _Nmax converged onto a maximum asymptotic value of 33.400±2.563 mol(CO₂) m^–2 s^–1. P _N fell to zero for an N_L-intercept of 0.710±0.035 g(N) m^–2. Under variable daily meteorological conditions the values for N_L, specific leaf area (_L), root mass fraction (R_f), P _Nmax, and remained constant for a given N supply. A monotonic decline in the steady-state value of R_f occurred with increasing N supply. _L increased with increasing N supply or with increasing N_L.