TWO DISTINCT COLONIAL MORPHOTYPES OF AMPHORA COFFEAEFORMIS (BACILLARIOPHYCEAE) CULTURED ON SOLID MEDIA1

When cultured on different types of solid media, the marine-fouling diatom Amphora coffeaeformis (Ag.) Kütz. consistently formed two distinct colonial morphotypes named tight and fuzzy. Tight colonies were comprised mainly of small, morphologically distorted, nonmotile cells, whereas morphologically normal and highly motile cells formed the fuzzy colonies. Cells from tight colonies were less adherent to glass, grew more slowly in liquid media, and had a slightly decreased viability on plates with copper than cells from fuzzy colonies. Whereas the protein profiles of the two types of cells were nearly identical in polyacrylamide gels stained with Coomassie blue, cells from tight colonies produced a significantly lower amount of a protease-resistant, low M_r polysaccharide or glycoconjugate as detected in silver-stained gels. The frequency of appearance of the fuzzy and tight morphotypes was not influenced by the mode of nutrition or the type of substratum to which the algal cells adhered. However, certain formulations of solid medium and the presence of growth-inhibitory concentrations of copper in agar plates favored the formation of tight colonies. Due to their frequencies and patterns of appearance, it was clear that the two naturally formed morphotypes were not the consequence of spontaneous mutations, genetic rearrangement, or selection of stable natural variants, and we have hypothesized that they were linked to a normal physiological behavior. The tight colonial morphotype was used as a valuable marker to screen for true motility/adhesion mutants within an ultraviolet-mutagenized population of A. coffeaeformis. Seven mutants were isolated that were non-motile on agar plates, poorly adherent to glass, and distinguished from naturally formed cells from tight colonies by their inability to form fuzzy colonies upon subculture on solid media.     

Age-dependent aluminum accumulation in the human aorta and cerebral artery

The purpose of this study was to determine the extent of aluminum (Al) accumulation in the human aorta and cerebral arteries. The Al contents in the aortae and in the cerebral arteries from 23 human subjects was determined by inductively coupled plasma atomic emission spectrophotometry (ICP-AES). The subjects' age range was 45–99-yr-old; 15 of the subjects were males and 8 were females. Al was detected in twelve aortae and in six cerebral arteries, when the entire specimen was analyzed. Two specimens where Al was found in the cerebral arteries contained no Al in the aorta. No relationship to the subject's sex was found. When related to age, two groups were established. Group L (45–75 yr old) and group H (>75 yr old), which exhibited aortal Al concentrations of 33.3 and 72.7%, respectively. When the aortic wall was dissected into the tunica intima, media, and adventitia, Al was found mainly in the tunica media. In the aorta, significant relationships were found between Al and phosphorus (P) levels (r=0.801,p<0.01) and between Al and calcium (Ca) (r=0.661,p<0.05). We have concluded that Al accumulation is age-dependent and that it occurs both in the aorta and in the cerebral artery. In the aorta, accumulation occurs mainly in the tunica media. Both P and Ca appear to enhance aortal Al accumulation.     

Analysis of stimulatory and inhibitory amino acids for development of hamster one-cell embryos in vitro

Hamster embryo development to the blastocyst stage in vitro can be modulated by amino acids. This series of experiments employed both empirically and statistically designed approaches to elucidate which of 20 amino acids inhibit or stimulate development and to devise a complement of amino acids that best supports in vitro development of hamster 1-cell embryos. Development and/or mean cell number were significantly inhibited by the presence of leucine, tyrosine, valine, isoleucine, phenylalanine, arginine, methionine, or cysteine (at 0.5 mM) and isoleucine, phenylalanine, or tryptophan (at 0.05 mM). Three amino acids—glutamine, taurine, and glycine—were stimulatory and in combination improved development; the culture medium containing these amino acids was designated Hamster Embryo Culture Medium-5. Moreover, addition of another eight amino acids—asparagine, aspartic acid, serine, glutamic acid, histidine, lysine, proline and cysteine (medium designated HECM-6)—had a significant stimulatory effect on development over previously formulated culture media for hamster embryos. These results demonstrated that amino acids, alone and in combination, can markedly stimulate or inhibit hamster embryo development in vitro up to the blastocyst stage. Embryo transfer experiments showed that HECM-5 and ?6 (chemically defined, protein-free culture media) supported normal preimplantation embryo development in vitro. This study also indicates that empirically designed embryo culture media formulations can be as effective as those obtained by application of statistical methodologies. © 1995 wiley-Liss, Inc.     

Transport of bacteria in porous media: II. A model for convective Transport and growth

A model is presented for the coupled processes of bacterial growth and convective transport of bacteria has been modeled using a fractional flow approach. The various mechanisms of bacteria retention can be incorporated into the model through selection of an appropriate shape of the fractional flow curve. Permeability reduction due to pore plugging by bacteria was simulated using the effective medium theory. In porous media, the rates of transport and growth of bacteria, the generation of metabolic products, and the consumption of nutrients are strongly coupled processes. Consequently, the set of governing conservation equations form a set of coupled, nonlinear partial differential equations that were solved numerically. Reasonably good agreement between the model and experimental data has been obtained indicating that the physical processes incorporated in the model are adequate. The model has been used to predict the in situ transport and growth of bacteria, nutrient consumption, and metabolite production. It can be particularly useful in simulating laboratory experiments and in scaling microbial-enhanced oil recovery or bioremediation processes to the field. (c) 1994 John Wiley & Sons, Inc.     

Enhancement of oxygen transfer in highly viscous non-newtonian fermentation broths

The influence of short draft tubes covered by perforated plates on gas-liquid mass transfer was examined in external-loop airlift bioreactors. The volumetric mass transfer coefficients in a model external-loop airlift bioreactor were measured with water and non-Newtonian media. It was found that introduction of draft tubes covered with perforated plates in the riser significantly improved the mass transfer rate, particularly in higher viscous non-Newtonian fermentation media. The enhancement of mass transfer rate might be due mainly to an increase in bubble coalescence and redispersion. (c) 1994 John Wiley & Sons, Inc.     

Dramatic enhancement of enzymatic activity in organic solvents by lyoprotectants

When seven different hydrolytic enzymes (four proteases and three lipases) were lyophilized from aqueous solution containing a ligand, N-Ac-L-Phe-NH(2), their catalytic activity in anhydrous solvents was far greater (one to two orders of magnitude) than that of the enzymes lyophilized without the ligand. This ligand-induced activation was expressed regardless of whether the substrate employed in organic solvents structurally resembled the ligand. Furthermore, nonligand lyoprotectants [sorbitol, other sugars, and poly(ethylene glycol)] also dramaticaliy enhanced enzymatic activity in anhydrous solvents when present in enzyme aqueous solution prior to lyophilization. The effects of the ligand and of the lyoprotectants were nonadditive, suggesting the same mechanism of action. Excipient activated and nonactivated enzymes exhibited identical activities in water. Also, addition of the excipients directly to suspensions of nonactivated enzymes in organic solvents had no appreciable effect on catalytic activity. These observations indicate that the mechanism of the excipient-induced activation is based on the ability of the excipients to alleviate reversible denaturation of enzymes upon lyophilization. Activity enhancement induced by the excipients is displayed even after their removal by washing enzymes with anhydrous solvents. Subtilisin Carlsberg, lyophilized with sorbitol, was found to be a much more efficient practical catalyst than its "regular" counterpart. (c) 1993 John Wiley & Sons, Inc.     

Activity of mushroom polyphenol oxidase in organic medium

A kinetic study of the activity of mushroom polyphenol oxidase in an organic system was carried out to obtain detailed enzyme kinetic data in relation to optimization of reaction conditions and substrate specificity. A simple method for consistent measurement of reaction rates in the heterogeneous enzyme/organic solvent system (consisting of immobilized polyphenol oxidase and a hydrated solution of the substrate in chloroform) was designed. The aqueous content of the system was optimized using p-cresol as the substrate. With this system, a crude extract of Agaricus bisporus was used to hydroxylate and oxidize a range of selected p-substituted phenolic substrates, yielding o-quinone products. Michaelis-Menten kinetics were used to obtain apparent K(M) and V(max) values with respect to each of these substrates. Results from this analysis indicated a correlation between the enzymic kinetic parameters obtained and the steric requirements of the substrates, which could be rationalized in terms of the restricted flexibility of the enzyme when it is in chloroform and also in terms of substrate and solvent hydrophobicity. In the course of the investigation UV molar absorption coefficients of several o-quinones were measured by a novel method: (1)H nuclear magnetic resonance (NMR) spectroscopy was employed to determine component concentrations in reaction mixtures resulting from the transformation of phenols by polyphenol oxidase in chloroform. Thus the UV molar absorption coefficients could be obtained directly, avoiding the necessity to isolate the water-sensitive, unstable o-quinones. (c) 1993 John Wiley & Sons, Inc.     

Reactions Catalyzed by Peg-Modified α-Chymotrypsin in Organic Solvents. Influence of Water Content and Degree of Modification

-Chymotrypsin was modified with cyanuric chloride activated monomethoxypolyethylene glycol (MPEG) with molecular weights 1900 and 5000. Using the higher molecular weight MPEG a product that was soluble in benzene at moderate levels of modification was obtained, whereas with MPEG 1900 almost all the enzyme's amino groups had to be modified for dissolving the conjugate. The catalytic activity decreased with increasing degree of substitution. Apparent V_max was considerably higher for the less modified enzyme preparation than for the more modified one, while K_m,app stayed almost constant. The modified enzyme was used for peptide synthesis. The reaction was dependent on the content of dissolved water. Both V_max,app and K_m,app increased with increasing water content. It was possible to achieve a process with complete conversion of substrate to dipeptide.     

Cultures of bovine tracheal epithelium with differentiated ultrastructure and ion transport

Summary Tracheal epithelial cells were grown on Nuclepore filters coated with human placental collagen. When grown immersed in medium containing fetal bovine serum, cells displayed an undifferentiated ultrastructure (no cilia and a cell height of ∼ 10 μm). Short-circuit current (I_sc) was approximately 1/10 that of the native epithelium. By contrast, when grown in hormonally defined, serum-free medium with an air interface, cells showed I_sc equal to or greater than the original tissue, possessed cilia, and had a cell height of ∼ 50 μm. Responses in I_sc to mediators were similar to those of the original tissue, but differed from those of dog or human tracheal epithelium. Given the ready availability and low cost of the native tissues, bovine tracheal cultures grown in serum-free medium with an air interface should prove useful in studies of airway epithelial physiology.