A new CSLM-based method for determination of the phase behaviour of aqueous mixtures of biopolymers

On mixing different types of high molecular weight (bio)polymers in an aqueous solution, phase separation often occurs. In some cases, the occurrence of phase separation may be readily observed, because due to density differences the heavier of the two phases is accumulated at the bottom of the vessel in which the mixture is contained. By using classical techniques, the composition of the two phases may then be determined. In the case where the density differences are not so large, and the viscosity of the system is high, the two phases remain intimately mixed. An alternative route to determine the phase behaviour of these systems might be a microscopic technique (Confocal Scanning Laser Microscopy, CSLM), using the fluorescence intensity of labelled biopolymers to quantify their concentration and phase volume in the system. Experiments were performed with several mixtures of sodium alginate, labelled with fluorescein, and sodium caseinate, fluorescently labelled with Texas Red. The viscosity of the mixtures studied was low enough to allow bulk phase separation of the phases by using an ultracentrifuge. Results of the phase volumes, and the composition of the phases, obtained independently by applying the two different methods (CSLM, or analysis of the separate phases after centrifugation) were compared and found to be in reasonable agreement.     

Towards a methodology for taking physical degradation of ecosystems into account in LCA

An analytic procedure has been followed to select adequate methods to express ecosystem degradation in LCA. This procedure consisted of problem definition, identification of relevant issues, of a quantitative expression for ecosystem degradation and of possible nature value indicators and building a framework of criteria for selecting adequate methods. With the selection framework a first screening of methods was performed. For full quantification the following formula is proposed: ED = L (N_r- N_a), with land use I. = A t and nature value change (N_r- N_a) Degradation due to an activity appears difficult to operationalise, but ecosystem suppression by activities can well be assessed. N_r is then the natural background or would-be natural situation. N_a can best be described by the actual state during the activity, unless hard data on restoration is available. N_a and N_r can be expressed in the biomass production indicator NPP - NCR Biodiversity and/or erosion may be added to include irreversible effects.     

Controlled clinical trial of L-dopa and nafoxidine in advanced breast cancer: an E.O.R.T.C. study.

L-Dopa lowers plasma prolactin levels, and there have been reports that patients with advanced breast cancer have been successfully treated with L-dopa. To test the potential value of L-dopa in this disease a randomized clinical trial of L-dopa and nafoxidine (as the reference compound) was conducted in postmenopausal women with advanced breast cancer. Objective remissions were obtained in sever out of 36 patients (19%) treated with nafoxidine but in none out of 40 patients treated with L-dopa. L-Dopa in the dose schedule used seems to be ineffective in advanced breast cancer.     

Estimating Breeding Values With Molecular Relatedness and Reconstructed Pedigrees in Natural Mating Populations of Common Sole,Solea Solea

Captive populations where natural mating in groups is used to obtain offspring typically yield unbalanced population structures with highly skewed parental contributions and unknown pedigrees. Consequently, for genetic parameter estimation, relationships need to be reconstructed or estimated using DNA marker data. With missing parents and natural mating groups, commonly used pedigree reconstruction methods are not accurate and lead to loss of data. Relatedness estimators, however, infer relationships between all animals sampled. In this study, we compared a pedigree relatedness method and a relatedness estimator (“molecular relatedness”) method using accuracy of estimated breeding values. A commercial data set of common sole, Solea solea, with 51 parents and 1953 offspring (“full data set”) was used. Due to missing parents, for 1338 offspring, a pedigree could be reconstructed with 10 microsatellite markers (“reduced data set”). Cross-validation of both methods using the reduced data set showed an accuracy of estimated breeding values of 0.54 with pedigree reconstruction and 0.55 with molecular relatedness. Accuracy of estimated breeding values increased to 0.60 when applying molecular relatedness to the full data set. Our results indicate that pedigree reconstruction and molecular relatedness predict breeding values equally well in a population with skewed contributions to families. This is probably due to the presence of few large full-sib families. However, unlike methods with pedigree reconstruction, molecular relatedness methods ensure availability of all genotyped selection candidates, which results in higher accuracy of breeding value estimation.To estimate genetic parameters, additive genetic relationships between individuals are inferred from known pedigrees (Falconer and Mackay 1996; Lynch and Walsh 1997). However, in natural populations (Ritland 2000; Thomas et al. 2002) and in captive species where natural mating in groups is used to obtain offspring (Brown et al. 2005; Fessehaye et al. 2006; Blonk et al. 2009) pedigrees are reconstructed. In these populations there is no control on mating structure, and typically unbalanced population structures with highly skewed parental contributions are obtained (Bekkevold et al. 2002; Brown et al. 2005; Fessehaye et al. 2006; Blonk et al. 2009). To reconstruct pedigrees, parental allocation methods are often used (Marshall et al. 1998; Avise et al. 2002; Duchesne et al. 2002). These methods require that all parents be known. For situations where parental information is not available, numerous DNA-marker-based methods for estimating molecular relatedness have been developed (Lynch 1988; Queller and Goodnight 1989; Ritland 2000; Toro et al. 2002). These relatedness estimators determine relationship values between individuals on a continuous scale. Evaluation of relatedness estimators within real and simulated data in both plants and animals (e.g., see Van de Casteele et al. 2001 ; Milligan 2003; Oliehoek et al. 2006; Rodríguez-Ramilo et al. 2007; Bink et al. 2008) has generally focused on bias and sampling error of estimated genetic variances or relatedness values. Relatively little attention has been paid to their efficiency for estimation of breeding values.Two types of relatedness estimators are currently available: method-of-moments estimators and maximum-likelihood estimators. Method-of-moments estimators (e.g., Queller and Goodnight 1989; Li et al. 1993; Ritland 1996; Lynch and Ritland 1999; Toro et al. 2002) determine relationships while calculating sharing of alleles between pairs in different ways. A variant of method-of-moments estimators is the transformation of continuous relatedness values to categorical genealogical relationships using “explicit pedigree reconstruction” (Fernández and Toro 2006) or thresholds (Rodríguez-Ramilo et al. 2007). However, correlations of transformed coancestries with known genealogical coancestries are low (Rodríguez-Ramilo et al. 2007). Several studies have compared different method-of-moments estimators but none revealed one single best estimator (Van de Casteele et al. 2001; Oliehoek et al. 2006; Rodríguez-Ramilo et al. 2007; Bink et al. 2008).Maximum-likelihood (ML) approaches classify animals into a limited number of relationship classes (Mousseau et al. 1998; Thomas et al. 2002; Wang 2004; Herbinger et al. 2006; Anderson and Weir 2007). For each pair a likelihood to fall into a possible relatedness class (e.g., full sib vs. unrelated) is calculated given its genotype and phenotype. ML techniques combined with a Markov chain Monte Carlo approach reconstruct groups with specific relationships jointly and are therefore more efficient than other ML approaches. To minimize standard errors, all discussed ML methods require balanced population structures, large sibling groups, and a large variance of relatedness (Thomas et al. 2002; Wang 2004; Anderson and Weir 2007). Therefore, these methods may not be suitable for natural mating systems.Unlike parental allocation methods, a benefit from relatedness estimators is that essentially all selection candidates are maintained for breeding value estimation, even with missing parents. The question is, however, whether such relatedness estimators also give accurate breeding values to perform selection.In this study, we test suitability of a relatedness estimator to obtain breeding values in a population of common sole, Solea solea (n = 1953) obtained by natural mating. First, we estimate breeding values using pedigree relatedness of animals for which a pedigree could be reconstructed (using parental allocation). This data set (n = 1338) is further referred to as “reduced data set.” We compare results with estimated breeding values using a simple but robust method-of-moments relatedness estimator: “molecular relatedness” (Toro et al. 2002, 2003). Next, we estimate breeding values using molecular relatedness in the full data set (n = 1953). Results show that accuracies of estimated breeding values obtained with molecular relatedness and pedigree relatedness are comparable. Accuracy increases when breeding values are estimated with molecular relatedness in the full data set. This implies that a molecular relatedness estimator can be used to estimate breeding values in captive natural mating populations.     

A framework for actualising normalisation data in LCA: Experiences in the Netherlands

In LCA, normalisation is applied to quantify the relative size of the impact scores. Several sets of normalisation data exist in the Netherlands, which all have a certain degree of unreliability. The purpose of this study is to actualise Dutch normalisation data and to make a framework for deriving these data. In this study normalisation data are calculated for three different levels in order to give the LCA practitioner a more extended basis for preparing the interpretation process. The first level of normalisation contains all impacts relating to activities that take place within the Dutch territory. The second level is based on the Dutch final consumption, which means that import and export are taken into account. The third level is an attempt to estimate impacts in Europe based on European data if possible, and otherwise based on extrapolation from the Dutch situation.     

Just eating healthier is not enough: studying the environmental impact of different diet scenarios for Dutch women (31–50 years old) by linear programming

Purpose

Eating healthier or vegetarian and vegan diets are suggested options to reduce the environmental impact of the current diet. In this paper, we demonstrate a method that is able to identify diets with reduced environmental impact and are more similar to the current diet than predetermined scenarios. All diets were adequate and consisted of commonly available foods.

Methods

We used linear programming to find solutions that are as close as possible to the current diet of an average woman with age 31–50, first without any food groups’ constraints and later by imposing constraints on meat, fish, dairy, and eggs. Finally, we use a similar technique to search for the closest diet that achieves the same environmental reduction as the most restricted option (no meat, fish, dairy, or eggs), without restrictions on products. In the optimization algorithm, we incorporate popularity of food products in order to design menus which are feasible for the Dutch population.

Results and discussion

The results show that simply eating according to guidelines does not guarantee a diet with an improved environmental profile. Removing meat and fish from the diet reduces the environmental impact by about 21 %. A healthy vegan diet reaches 30 % environmental impact reduction, but leads to a diet with many changes in comparison to a typical Dutch diet and without meeting one of the health constraints (EPA?+?DHA—Eicosapentaenoic acid?+?Docosahexaenoic acid). We show that it is possible to find less restrictive solutions than vegetarian or vegan diets that still satisfy all nutritional requirements and have less environmental impact than the current one.

Conclusions

Just eating healthier is not enough in order to reduce environmental impact. However, designing a diet that meets dietary requirements must be a prerequisite for sustainable diets. Simply removing products from a diet can have as consequence that other products have to be added to compensate for the nutritional imbalances. We show, by using linear programming, that it is possible to reach 30 % reduction in the environmental impact with a diet which is relatively similar to the current one and could be more likely to be accepted.

     

Vinculin potentiates E-cadherin mechanosensing and is recruited to actin-anchored sites within adherens junctions in a myosin II–dependent manner

Cell surface receptors integrate chemical and mechanical cues to regulate a wide range of biological processes. Integrin complexes are the mechanotransducers between the extracellular matrix and the actomyosin cytoskeleton. By analogy, cadherin complexes may function as mechanosensors at cell–cell junctions, but this capacity of cadherins has not been directly demonstrated. Furthermore, the molecular composition of the link between E-cadherin and actin, which is needed to sustain such a function, is unresolved. In this study, we describe nanomechanical measurements demonstrating that E-cadherin complexes are functional mechanosensors that transmit force between F-actin and E-cadherin. Imaging experiments reveal that intercellular forces coincide with vinculin accumulation at actin-anchored cadherin adhesions, and nanomechanical measurements show that vinculin potentiates the E-cadherin mechanosensory response. These investigations directly demonstrate the mechanosensory capacity of the E-cadherin complex and identify a novel function for vinculin at cell–cell junctions. These findings have implications for barrier function, morphogenesis, cell migration, and invasion and may extend to all soft tissues in which classical cadherins regulate cell–cell adhesion.     

Comparing apples with oranges: on the functional equivalence of food products for comparative LCAs

Purpose

In this article, we present an innovative way of deriving comparable functional systems for comparative life cycle assessments (LCAs) of food products. We define the functional unit as the contribution of one or more foods to the nutrient composition of a weekly diet and, after a product substitution, employ a product system expansion approach to search for an alternative set of products which provides an equivalent nutritional composition.

Methods

Replacement is regarded within the context of a weekly diet. The comparable diet is a solution to a linear problem which finds the diet that is most similar to the starting one, subject to nutritional and/or other constraints that guarantee a minimum dietary quality. The formulation gives priority to selecting food products according to popularity.

Results

We illustrate our method with two examples. We show that a baseline diet containing 3.6 servings of apples a week is equivalent to a similar diet in which the apples are replaced with 3.6 servings of oranges and servings of strawberry and kiwi are removed. These changes are necessary mainly because of differences in the vitamin C content between apples and oranges. The second example is a replacement of all meat in a weekly diet by a soy-based meat substitute. In this case, additional fish products need to be consumed to make up for a lack of selenium and essential amino acids.

Conclusions

We present an innovative and objective way to overcome the challenge of comparing two or more food products in a comparative LCA. Our approach is systematic and finds the alternative diet that best meets the nutritional criteria as well as reflecting the food preferences of the population. The method selects products according to the role they play in the dietary pattern. Moreover, the method is flexible enough to allow for different selection criteria and other nutritional and non-nutritional constraints.     

Quantification of levetiracetam in plasma of neonates by ultra performance liquid chromatography–tandem mass spectrometry

A sensitive and specific method using ultra performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) was developed for the determination of levetiracetam (LEV) in plasma of neonates. A plasma aliquot of 50 μl was deproteinized by addition of 500 μl methanol which contained 5 μg/ml UCB 17025 as an internal standard. After centrifugation, 50 μl of supernatant was diluted with 1000 μl of 0.1% formic acid–10 mM ammonium formate in water (pH 3.5) (mobile phase solution A) and 2 μl was injected onto the UPLC-system. Compounds were separated on a Acquity UPLC BEH C₁₈ 2.1 mm × 100 mm column using gradient elution with mobile phase solution A and 0.1% formic acid in methanol (mobile phase solution B) with a flow rate of 0.4 ml/min and a total runtime of 4.0 min. LEV and the internal standard were detected using positive ion electrospray ionization followed by tandem mass spectrometry (ESI-MS/MS). The assay allowed quantification of LEV plasma concentrations in the range from 0.5 μg/ml to 150 μg/ml. Inter-assay inaccuracy was within ±2.7% and inter-assay precision was less than 4.5%. Matrix effects were minor: the recovery of LEV was between 97.7% and 100%. The developed method required minimal sample preparation and less plasma sample volume compared to earlier published LC–MS/MS methods. The method was successfully applied in a clinical pharmacokinetic study in which neonates received intravenous administrations of LEV for the treatment of neonatal seizures.