Surfactants in microbiology and biotechnology: Part 2. Application aspects

Surfactants are amphiphilic compounds which can reduce surface and interfacial tensions by accumulating at the interface of immiscible fluids and increase the solubility, mobility, bioavailability and subsequent biodegradation of hydrophobic or insoluble organic compounds. Chemically synthesized surfactants are commonly used in the petroleum, food and pharmaceutical industries as emulsifiers and wetting agents. Biosurfactants produced by some microorganisms are becoming important biotechnology products for industrial and medical applications due to their specific modes of action, low toxicity, relative ease of preparation and widespread applicability. They can be used as emulsifiers, de-emulsifiers, wetting and foaming agents, functional food ingredients and as detergents in petroleum, petrochemicals, environmental management, agrochemicals, foods and beverages, cosmetics and pharmaceuticals, and in the mining and metallurgical industries. Addition of a surfactant of chemical or biological origin accelerates or sometimes inhibits the bioremediation of pollutants. Surfactants also play an important role in enhanced oil recovery by increasing the apparent solubility of petroleum components and effectively reducing the interfacial tensions of oil and water in situ. However, the effects of surfactants on bioremediation cannot be predicted in the absence of empirical evidence because surfactants sometimes stimulate bioremediation and sometimes inhibit it. For medical applications, biosurfactants are useful as antimicrobial agents and immunomodulatory molecules. Beneficial applications of chemical surfactants and biosurfactants in various industries are discussed in this review.     

Inhibitors of the tyrosine kinase EphB4. Part 1: Structure-based design and optimization of a series of 2,4-bis-anilinopyrimidines

A series of bis-anilinopyrimidines have been identified as potent inhibitors of the tyrosine kinase EphB4. Structural information from two alternative series identified from screening efforts was combined to identify the initial leads.     

Evaluation of secondary amide replacements in a series of CCR5 antagonists as a means to increase intrinsic membrane permeability. Part 1: Optimization of gem-disubstituted azacycles

Replacement of a secondary amide with an N-acyl or N-sulfonyl gem-disubstituted azacyle in a series of CCR5 antagonists led to the identification of compounds with excellent in vitro HIV antiviral activity and increased intrinsic membrane permeability.     

Genetic aspects regarding piglet losses and the maternal behaviour of sows. Part 1. Genetic analysis of piglet mortality and fertility traits in pigs

In spite of the improvement in management and the breeding goal of increasing the number of piglets born alive, piglet mortality is still a substantial problem in pig breeding. The objective of the first part of the study was to estimate genetic parameters for different causes of piglet losses and to investigate the relationship to litter-size traits. Data were collected on a nucleus herd from January till December 2004. Records from 943 German Landrace sows with 1538 pure-bred litters and 13 971 individually weighted piglets were included. Four different causes of piglet losses (LOSS) were evaluated. Additional analysed traits were underweight and runting. Furthermore, the fertility traits number of piglets born alive, born in total and stillborn piglets as well as the individual birth and weaning weights were analysed. The different LOSS were treated as a binary trait and subsequently the heritabilities were estimated using a threshold model. The most important LOSS was crushing under the sow (12.4%). The survival rate and crushing had a heritability of h2 = 0.03. The fertility traits piglets born alive, born in total and stillborn piglets were analysed with a linear model and heritabilities rank from h2 = 0.05 (stillborn) to h2 = 0.10 (born alive). The estimated heritabilities for birth- and weaning weight were both h2 = 0.10. The genetic correlations between number of piglets born alive and each LOSS trait were analysed bivariately. Of all piglets born alive 84.3% survive the lactation period. Survival decreased with increasing litter size (rg = -0.54 up to -0.78) and the probability of being crushed under the sow increased.     

Novel GlyT1 inhibitor chemotypes by scaffold hopping. Part 2: Development of a [3.3.0]-based series and other piperidine bioisosteres

This Letter describes the development and SAR of a novel series of GlyT1 inhibitors derived from a scaffold hopping approach, in lieu of an HTS campaign, which provided intellectual property position. Members within this new [3.3.0]-based series displayed excellent GlyT1 potency, selectivity, free fraction, and modest CNS penetration. Moreover, enantioselective GlyT1 inhibition was observed, within this novel series and a number of other piperidine bioisosteric cores.     

Evaluation of amide replacements in CCR5 antagonists as a means to increase intrinsic permeability. Part 2: SAR optimization and pharmacokinetic profile of a homologous azacyle series

Replacement of a secondary amide with a piperidine or azetidine moiety in a series of CCR5 antagonists led to the discovery of compounds with increased intrinsic permeability. This effort led to the identification of a potent CCR5 antagonist which exhibited an improved in vivo pharmacokinetic profile.     

Part of the series: from dietary antioxidants to regulators in cellular signalling and gene expression. Role of reactive oxygen species and (phyto)oestrogens in the modulation of adaptive response to stress

There is increasing evidence that reactive oxygen species (ROS) are not only toxic but play an important role in cellular signalling and in the regulation of gene expression. We, here, discuss two examples of improved adaptive response to an altered cellular redox state. First, differences in longevity between males and females may be explained by a higher expression of antioxidant enzymes in females resulting in a lower yield of mitochondrial ROS. Oestrogens are made responsible for these phenomena. Oestradiol induces glutathione peroxidase-1 and MnSOD by processes requiring the cell surface oestrogen receptor (ER) and the activation of pathways usually involved in oxidative stress response. Second, oxygen radicals produced during moderate exercise as performed during training up-regulate the expression of antioxidant enzymes in muscle cells. An increased level of these enzymes might prevent oxidative damage during exhaustive exercise and should, therefore, not be prevented by antioxidants. The relevance of these findings is discussed in the context with observations made in transgenic animals overexpressing MnSOD or catalase.     

Physiological responses of Aureoumbra lagunensis and Synechococcus sp. to nitrogen addition in a mesocosm experiment

Aureoumbra lagunensis is the causative organism of the Texas brown tide and is notable because it dominated the Laguna Madre ecosystem from 1990 to 1997. This species is unusual because it has the highest known critical nitrogen to phosphorus ratio (N:P) for any microalgae ranging from 115 to 260, far higher than the 16N:1P Redfield ratio. Because of its high N:P ratio, Aureoumbra should be expected to respond to N additions that would not stimulate the growth of competitors having the Redfield ratio. To evaluate this prediction, a mesocosm experiment was performed in the Laguna Madre, a South Texas coastal lagoon, in which a mixed Aureoumbra–Synechococcus (a cyanobacterium) community was enclosed in 12 mesocosms and subjected to nitrogen addition (6 controls, 6 added ammonium) for 16 days. After day 4, added nitrogen did not significantly increase Aureoumbra specific growth rate but the alga retained dominance throughout the experiment (64–75% of total cell biovolume). In control mesocosms, Aureoumbra became less abundant during the first 4 days of the experiment but rebounded by the end of the experiment and was dominant over Synechococcus. Despite the lack of a strong positive growth response, Aureoumbra did respond physiologically to N addition. By the end of the experiment, the average N:P ratio of the Aureoumbra-dominated community was 86 in the N+ treatment and 41 in the control, indicating that the alga became less N-limited in the N+ treatment. The average C:N ratio was 6.6 in the N+ treatment (8.6 in the control) and suggests that the alga was not N-limited, however, C:N ratio may not be a good indicator of nitrogen limitation since this alga can produce significant quantities of carbon-containing extracellular polysaccharides, depending on growth conditions. Both Aureoumbra cellular chlorophyll fluorescence and cell size increased in response to added N, indicating a reduction in N limitation. It appeared that the N additions were not large and/or frequent enough to stimulate Aureoumbra growth. The main competitor, the unicellular cyanobacterium Synechococcus, responded positively to the nitrogen addition by increased specific growth rate. Unlike Aureoumbra, no significant effect on Synechococcus cellular pigment fluorescence or cell size was noted. Literature data suggest that Synechococcus, like Aureoumbra, may have a critical N:P ratio much higher than 16:1, which could explain its response.     

Electron transfer in cyanobacterial photosystem I: I. Physiological and spectroscopic characterization of site-directed mutants in a putative electron transfer pathway from A0 through A1 to FX

The Photosystem I (PS I) reaction center contains two branches of nearly symmetric cofactors bound to the PsaA and PsaB heterodimer. From the x-ray crystal structure it is known that Trp697PsaA and Trp677PsaB are pi-stacked with the head group of the phylloquinones and are H-bonded to Ser692PsaA and Ser672PsaB, whereas Arg694PsaA and Arg674PsaB are involved in a H-bonded network of side groups that connects the binding environments of the phylloquinones and FX. The mutants W697FPsaA, W677FPsaB, S692CPsaA, S672CPsaB, R694APsaA, and R674APsaB were constructed and characterized. All mutants grew photoautotrophically, yet all showed diminished growth rates compared with the wild-type, especially at higher light intensities. EPR and electron nuclear double resonance (ENDOR) studies at both room temperature and in frozen solution showed that the PsaB mutants were virtually identical to the wild-type, whereas significant effects were observed in the PsaA mutants. Spin polarized transient EPR spectra of the P700+A1- radical pair show that none of the mutations causes a significant change in the orientation of the measured phylloquinone. Pulsed ENDOR spectra reveal that the W697FPsaA mutation leads to about a 5% increase in the hyperfine coupling of the methyl group on the phylloquinone ring, whereas the S692CPsaA mutation causes a similar decrease in this coupling. The changes in the methyl hyperfine coupling are also reflected in the transient EPR spectra of P700+A1- and the CW EPR spectra of photoaccumulated A1-. We conclude that: (i) the transient EPR spectra at room temperature are predominantly from radical pairs in the PsaA branch of cofactors; (ii) at low temperature the electron cycle involving P700 and A1 similarly occurs along the PsaA branch of cofactors; and (iii) mutation of amino acids in close contact with the PsaA side quinone leads to changes in the spin density distribution of the reduced quinone observed by EPR.     

Nutrient cycling in a tropical seasonal rain forest of Xishuangbanna, Southwest China. Part 1: tree species: nutrient distribution and uptake

Tropical rain forests are characterized by large numbers of the species with diverse growth habits. The objective of the present study was to determine the distribution of nutrient content in the major trees of the tropical rain forests in Xishuangbanna. This will improve the understanding of the nutrient losses from such sites that result from harvesting and flow of nutrients within the ecosystem and lead to the development of effective and rational forest management strategies. Based on the results in this study, the distribution of nutrients among biomass components of trees varied: The ordering of major elements concentrations was K>N>Mg>Ca>P in branch, stem and root tissues but was N>K>Mg>Ca>P in leaves. The maximum amount of all nutrients per ha occurred in the stems followed by branches, roots and leaves. Of the total uptake of 6167.7 kg ha⁻¹ of all nutrients, the contribution of various nutrients was found to be N (2010.6 t ha⁻¹), P (196.3 t ha⁻¹), K (2123.8 kg ha⁻¹), Ca (832 kg ha⁻¹) and Mg (1005 kg ha⁻¹). However, comparing the nutrient uptake of other tropical and sub tropical forests, the results indicated that rates for the Xishuangbanna forests were 20–35% lower than previously reported values.     

Effects of Multiple Disturbances (Fire and Hurricane) on Epiphyte Community Dynamics in a Subtropical Forest,Florida, U.S.A.1

We addressed the interacting effects of a natural large–scale fire and a subsequent major hurricane on relative positions of epiphytes in a subtropical forest. In Everglades National Park, subtropical hammocks (hardwood tree “islands”; burned and unburned) during the Ingraham Fire (1989) were surveyed for trees and epiphytic bromeliads (Tillandsia spp.) one year before, as well as one and five years after, Hurricane Andrew (1992). We measured trees (species, diameter, and status [alive/dead]) and epiphytes (species, height, host tree characteristics, substrate life status, and density). The fire decreased the height of epiphytes during the hurricane because branches and bark of trees killed by the fire were unstable epiphyte substrates in the high winds. Proportions of epiphytes on Quercus virginiana were equally increased after the hurricane in both unburned and burned hammocks; the large size and bark characteristics resulted in greater proportional survival of epiphytes on this species. During the five years following the hurricane, changes in the distributions of epiphytes generally were toward pre–hurricane distributions, but recovery was faster in unburned than burned hammocks. We conclude that disturbances that kill trees are likely to amplify the vertical reduction of epiphytes during a subsequent hurricane and that effects of a single disturbance on plant populations can be influenced by the disturbance history of the system, including different types of disturbances.     

Genetic diversity and differentiation processes in the ploidy series of Olea europaea L.: a multiscale approach from subspecies to insular populations

Geographical isolation and polyploidization are central concepts in plant evolution. The hierarchical organization of archipelagos in this study provides a framework for testing the evolutionary consequences for polyploid taxa and populations occurring in isolation. Using amplified fragment length polymorphism and simple sequence repeat markers, we determined the genetic diversity and differentiation patterns at three levels of geographical isolation in Olea europaea : mainland-archipelagos, islands within an archipelago, and populations within an island. At the subspecies scale, the hexaploid ssp. maroccana (southwest Morocco) exhibited higher genetic diversity than the insular counterparts. In contrast, the tetraploid ssp. cerasiformis (Madeira) displayed values similar to those obtained for the diploid ssp. guanchica (Canary Islands). Geographical isolation was associated with a high genetic differentiation at this scale. In the Canarian archipelago, the stepping-stone model of differentiation suggested in a previous study was partially supported. Within the western lineage, an east-to-west differentiation pattern was confirmed. Conversely, the easternmost populations were more related to the mainland ssp. europaea than to the western guanchica lineage. Genetic diversity across the Canarian archipelago was significantly correlated with the date of the last volcanic activity in the area/island where each population occurs. At the island scale, this pattern was not confirmed in older islands (Tenerife and Madeira), where populations were genetically homogeneous. In contrast, founder effects resulted in low genetic diversity and marked genetic differentiation among populations of the youngest island, La Palma.     

Possible role of a short extra loop of the long-chain flavodoxin from Azotobacter chroococcum in electron transfer to nitrogenase: Complete 1H, 15N and 13C backbone assignments and secondary solution structure of the flavodoxin

Summary The ¹H, ¹⁵N and ¹³C backbone and ¹H and ¹³C beta resonance assignments of the long-chain flavodoxin from Azotobacter chroococcum (the 20-kDa nifF product, flavodoxin-2) in its oxidized form were made at pH 6.5 and 30°C using heteronuclear multidimensional NMR spectroscopy. Analysis of the NOE connectivities, together with amide exchange rates, ³J_Hn_H coupling constants and secondary chemical shifts, provided extensive solution secondary structure information. The secondary structure consists of a five-stranded parallel -sheet and five -helices. One of the outer regions of the -sheet shows no regular extended conformation, whereas the outer strand 4/6 is interrupted by a loop, which is typically observed in long-chain flavodoxins. Two of the five -helices are nonregular at the N-terminus of the helix. Loop regions close to the FMN are identified. Negatively charged amino acid residues are found to be mainly clustered around the FMN, whereas a cluster of positively charged residues is located in one of the -helices. Titration of the flavodoxin with the Fe protein of the A. chroococcum nitrogenase enzyme complex revealed that residues Asn¹¹, Ser⁶⁸ and Asn⁷² are involved in complex formation between the flavodoxin and Fe protein. The interaction between the flavodoxin and the Fe protein is influenced by MgADP and is of electrostatic nature.Abbreviations SQ semiquinone - FMN riboflavin 5-monophosphate; nif, nitrogen fixation - TSP 3-(trimethylsilyl)propionate sodium salt - DSS 2,2-dimethyl-2-silapentane-5-sulfonate sodium salt Supplementary Material is available on request, comprising a Materials and Methods section for the expression and purification of the A. chroococcum flavodoxin, a Table S1 containing the parameters of the titration of A. chroococcum flavodoxin with the Fe protein, and a Table S2 containing the ¹⁵N, H^N, ¹³C, ¹H, ¹³C, ¹H and ¹³CO chemical shifts.To whom correspondence should be addressed.     

Observation of the terminal methyl group in fatty acids of the linolenic series by a new 1H NMR pulse sequence providing spectral editing and solvent suppression. Application to excised frog muscle and rat brain

A new 1H NMR pulse sequence is described that combines water suppression with the selective observation of signals from coupled spin systems. The pulse sequence is easy to set up and compensates for pulse width inhomogeneity in the biological sample. Suppression of the water signal is achieved by pulses that return the water spins to their equilibrium position; spectral editing is based on the J modulation present in spin-echo spectra and its inhibition by coherent decoupling at one of the resonances of the spin system of interest. The pulse sequence, which was designed for 1H NMR spectroscopy of tissue, was tested at 470 MHz on excised frog muscle and rat brain. The lactate methyl resonance of caffeine-treated frog sartorius muscle was observed selectively by irradiation at the position of its alcoholic proton. The terminal methyl signal of linolenic acid, along with other fatty acids of the linolenic series (first double bond in the omega-3 position), was observed selectively by irradiation at the position of its omega-1 methylene group. 1H NMR spectra of rat brain were edited to reveal the terminal methyl of either linolenic series or all other fatty acids. The results suggest that the terminal methyl groups of fatty acids of the linolenic series (mostly docosahexaenoic acid, 22:6) have higher mobility than those of all other fatty acids.     

Application of Markov Chain Model to Calculate the Average Number of Times of Use of a Material in Society. An Allocation Methodology for Open-Loop Recycling. Part 1: Methodology Development (7 pp)

- Preamble. In this series of two papers, a methodology to calculate the average number of times a material is used in a society from cradle to grave is presented and applied to allocation of environmental impact of virgin material. Part 1 focuses on methodology development and shows how the methodology works with hypothetical examples of material flows. Part 2 presents case studies for steel recycling in Japan, in which the methodology is applied and allocation of environmental impact of virgin steel is conducted. - Abstract Goal, Scope and Background. It has been recognized that LCA has a limitation in assessing open cycle recycling of materials because of inevitable subjective judgments in setting system boundary. According with the enforcement of recycling laws, there has been a rapid increase in recycling ratio of materials at the end-of-life of products in many industrialized countries. So, materials' life cycle is getting more complicated, which makes it difficult to quantify the environmental impacts of materials used in a product in an appropriate way. The purpose of this paper is to develop a methodology to calculate the average number of times a material is used in a society from cradle to grave. The method developed in this paper derives the average number of times material is used; this value could be used for allocation of environmental burdens of virgin material as well as an indicator for assessing the state of material use in a certain year, based on material flow of material in that year. Main Features Our methodology is based on Markov chain model using matrix-based numerical analysis. A major feature of this method is that it creates transition probability matrices for a material from the way in which the material is produced, consumed, and recycled, making it possible to simply elicit indicators that assess the status of material use in products in society. Our methodology could be an alternative method to derive the average number of times material is used, which could be used for allocation of environmental burdens of virgin material. Results and Discussions The methodology was applied to hypothetical examples of material flows, in which a virgin material was produced and used in products, recycled and finally landfilled. In some cases, closed loop and open loop recycling of materials existed. The transition probability matrix was created for each material flow, and how many times a virgin material is used in products until all of the elements are ultimately landfilled. Conclusions This methodology is applicable to a complicated material flow if the status of residence of a material and its flow in a society can be figured out. All the necessary data are the amount of virgin material production, amount of the material used in products, recycling rate of the material at the end of life of each product, the amount of scrap of the material that are used for products. In Part 2 of this paper, case studies for steel were conducted.     

The role of phase separation and feed cycle length in leach beds coupled to methanogenic reactors for digestion of a solid substrate (Part 1): Optimisation of reactors' performance

A two-phase system composed by a leach bed and a methanogenic reactor was modified for the first time to improve volumetric substrate degradation and methane yields from a complex substrate (maize; Zeamays). The system, which was operated for consecutive feed cycles of different durations for 120 days, was highly flexible and its performance improved by altering operational conditions. Daily substrate degradation was higher the shorter the feed cycle, reaching 8.5 g TS_destroyed d⁻¹ (7-day feed cycle) but the overall substrate degradation was higher by up to 55% when longer feed cycles (14 and 28 days) were applied. The same occurred with volumetric methane yields, reaching 0.839 m³ (m³)⁻¹ d⁻¹. The system performed better than others on specific methane yields, reaching 0.434 m³ kg⁻¹ TS_added, in the 14-day and 28-day systems. The UASB and AF designs performed similarly as second stage reactors on methane yields, SCOD and VFA removal efficiencies.