Chiral separation principles in chromatographic and electromigration techniques

Almost half of the drugs in use today are chiral. It is well established that the pharmacological activity is mostly restricted to one of the enantiomers (eutomer). There can be qualitative and quantitative differences in the activity of the enantiomers. In many cases, the inactive enantiomer (distomer) shows unwanted side effects or even toxic effects. Even if the side effects are not that drastic, the distomer has to be metabolized and this represents an unnecessary burden for the organism. Therefore, the development of methods for the separation of enantiomers, both on analytical and preparative scale, has become increasingly important. Chromatographic techniques such as thin layer chromatography (TLC), gas chromatography (GC), supercritical fluid chromatography (SFC), and above all high-performance liquid chromatography (HPLC) have been used for enantiomer separation for about two decades. More recently, electromigration techniques, such as capillary electrophoresis and capillary electrochromatography, have been shown to be powerful alternatives to chromatographic methods. This review gives a short overview of different chiral separation principles and their application. Several new developments are discussed.     

Revisiting the agronomic benefits of manure: a correct assessment and exploitation of its fertilizer value spares the environment

The ongoing disintegration of agriculture into specialized farms, specialized regions and even specialized countries, has disrupted local nutrient cycles. This process is promoted and sustained by the use of mineral fertilizers. Ample availability of mineral fertilizers has changed farmers' perception and appreciation of manure. By now, manure is looked upon as 'waste' in some regions. It is a contradiction in terms, however, to regard biological by-products as 'waste'. Apart from this cultural aspect, economical and environmental considerations per se, justify a rehabilitation of manure. First of all a correct assessment of the fertilizer value is required. The present paper explains how to determine the nitrogen fertilizer value (NFV) of manures in a unbiased manner. Subsequently, recommendations are made on how to exploit the potential NFV as good as possible. Undeniably, it is easier to manage mineral fertilizers than manure. However, when proper attention is given to the composition of manure and decisions on rates, timing and placement are made correspondingly, the NFV can be enhanced. This should lead to a drastic reduction of mineral fertilizer use, nutrient surpluses and environmental pollution. Under such conditions, 'manure', 'precision farming' and 'environment' can become reconciled again.     

Do early branching lineages signify ancestral traits?

A reverence for ancestors that has pre-occupied humans since time immemorial persists to the present. Reconstructing ancestry is the focus of many biological studies but failure to distinguish between present-day descendants and long-dead ancestors has led to incorrect interpretation of phylogenetic trees. This has resulted in erroneous reconstruction of traits such as morphology and ancestral areas. Misinterpretation becomes evident when authors use the terms 'basal' or 'early diverging' to refer to extant taxa. Here, we discuss the correct interpretation of trees and methods for reconstructing the ancestral features of organisms using recently developed statistical models. These models can be inaccurate unless they use information that is independent of phylogenies, such as genetics, molecular and developmental biology, functional morphology, geological and climatic processes, and the fossil record.     

Considerations for the use of cellular electrophysiology models within cardiac tissue simulations

The use of mathematical models to study cardiac electrophysiology has a long history, and numerous cellular scale models are now available, covering a range of species and cell types. Their use to study emergent properties in tissue is also widespread, typically using the monodomain or bidomain equations coupled to one or more cell models. Despite the relative maturity of this field, little has been written looking in detail at the interface between the cellular and tissue-level models. Mathematically this is relatively straightforward and well-defined. There are however many details and potential inconsistencies that need to be addressed, in order to ensure correct operation of a cellular model within a tissue simulation. This paper will describe these issues and how to address them.Simply having models available in a common format such as CellML is still of limited utility, with significant manual effort being required to integrate these models within a tissue simulation. We will thus also discuss the facilities available for automating this in a consistent fashion within Chaste, our robust and high-performance cardiac electrophysiology simulator.It will be seen that a common theme arising is the need to go beyond a representation of the model mathematics in a standard language, to include additional semantic information required in determining the model’s interface, and hence to enhance interoperability. Such information can be added as metadata, but agreement is needed on the terms to use, including development of appropriate ontologies, if reliable automated use of CellML models is to become common.     

Efficient and accurate experimental design for enzyme kinetics: Bayesian studies reveal a systematic approach

In areas such as drug development, clinical diagnosis and biotechnology research, acquiring details about the kinetic parameters of enzymes is crucial. The correct design of an experiment is critical to collecting data suitable for analysis, modelling and deriving the correct information. As classical design methods are not targeted to the more complex kinetics being frequently studied, attention is needed to estimate parameters of such models with low variance. We demonstrate that a Bayesian approach (the use of prior knowledge) can produce major gains quantifiable in terms of information, productivity and accuracy of each experiment. Developing the use of Bayesian Utility functions, we have used a systematic method to identify the optimum experimental designs for a number of kinetic model data sets. This has enabled the identification of trends between kinetic model types, sets of design rules and the key conclusion that such designs should be based on some prior knowledge of K(M) and/or the kinetic model. We suggest an optimal and iterative method for selecting features of the design such as the substrate range, number of measurements and choice of intermediate points. The final design collects data suitable for accurate modelling and analysis and minimises the error in the parameters estimated.     

Enantioseparation of Racemic Flurbiprofen by Aqueous Two‐Phase Extraction With Binary Chiral Selectors of L‐dioctyl Tartrate and L‐tryptophan

A novel method for chiral separation of flurbiprofen enantiomers was developed using aqueous two‐phase extraction (ATPE) coupled with biphasic recognition chiral extraction (BRCE). An aqueous two‐phase system (ATPS) was used as an extracting solvent which was composed of ethanol (35.0% w/w) and ammonium sulfate (18.0% w/w). The chiral selectors in ATPS for BRCE consideration were L‐dioctyl tartrate and L‐tryptophan, which were screened from amino acids, β‐cyclodextrin derivatives, and L‐tartrate esters. Factors such as the amounts of L‐dioctyl tartrate and L‐tryptophan, pH, flurbiprofen concentration, and the operation temperature were investigated in terms of chiral separation of flurbiprofen enantiomers. The optimum conditions were as follows: L‐dioctyl tartrate, 80 mg; L‐tryptophan, 40 mg; pH, 4.0; flurbiprofen concentration, 0.10 mmol/L; and temperature, 25 °C. The maximum separation factor α for flurbiprofen enantiomers could reach 2.34. The mechanism of chiral separation of flurbiprofen enantiomers is discussed and studied. The results showed that synergistic extraction has been established by L‐dioctyl tartrate and L‐tryptophan, which enantioselectively recognized R‐ and S‐enantiomers in top and bottom phases, respectively. Compared to conventional liquid–liquid extraction, ATPE coupled with BRCE possessed higher separation efficiency and enantioselectivity without the use of any other organic solvents. The proposed method is a potential and powerful alternative to conventional extraction for separation of various enantiomers. Chirality 27:650–657, 2015. © 2015 Wiley Periodicals, Inc.     

Improved molecular dynamics simulations for the determination of peptide structures

In this article a few methods or modifications proven to be useful in the conformational examination of peptides and related molecules by molecular dynamics are illustrated. The first is the explicit use of organic solvents in the simulations. For many cases such solvents are appropriate since the nmr measurements (or other experimental observations) were carried out in the same solvent. Here, the use of dimethylsulfoxide and chloroform in molecular dynamics is described, with some advantages of the use of these solvents highlighted. A constant allowing for the scaling of the nonbonded interactions of the force field, an idea previously employed in distance geometry and simulated annealing, has been implemented. The usefulness of this method is that when the nonbonded term is turned to zero, atoms can pass through each other, while the connectivity of the molecule is maintained. It will be shown that such simulations, if a sufficient driving force is present (i.e., nuclear Overhauser effects restraints), can produce the correct stereoconfiguration (i.e., chiral center) as well as configurational isomer (i.e., cis/trans isomers). Lastly, a penalty term for coupling constants directly related to the Karplus curve has been plemented into the potential energy force field. The advantages of this method over the commonly used dihedral angle restraining are discussed. In particular, it is shown that with more than one coupling constant about a dihedral angle a great reduction of the allowed conformational space is obtained. © 1993 John Wiley & Sons, Inc.     

Network-based auto-probit modeling for protein function prediction

Predicting the functional roles of proteins based on various genome-wide data, such as protein-protein association networks, has become a canonical problem in computational biology. Approaching this task as a binary classification problem, we develop a network-based extension of the spatial auto-probit model. In particular, we develop a hierarchical Bayesian probit-based framework for modeling binary network-indexed processes, with a latent multivariate conditional autoregressive Gaussian process. The latter allows for the easy incorporation of protein-protein association network topologies-either binary or weighted-in modeling protein functional similarity. We use this framework to predict protein functions, for functions defined as terms in the Gene Ontology (GO) database, a popular rigorous vocabulary for biological functionality. Furthermore, we show how a natural extension of this framework can be used to model and correct for the high percentage of false negative labels in training data derived from GO, a serious shortcoming endemic to biological databases of this type. Our method performance is evaluated and compared with standard algorithms on weighted yeast protein-protein association networks, extracted from a recently developed integrative database called Search Tool for the Retrieval of INteracting Genes/proteins (STRING). Results show that our basic method is competitive with these other methods, and that the extended method-incorporating the uncertainty in negative labels among the training data-can yield nontrivial improvements in predictive accuracy.     

Probability rule for chiral recognition

Molecular Chirality is of central interest in biological studies because enantiomeric compounds, while indistinguishable by most inanimate systems, show profoundly different properties in biochemical environments. Enantioselective separation methods, based on the differential recognition of two optical isomers by a chiral selector, have been amply documented. Also, great effort has been directed towards a theoretical understanding of the fundamental mechanisms underlying the chiral recognition process. Here we report a comprehensive data examination of enantio separation measurements for over 72000 chiral selector-select and pairs from the chiral selection compendium CHIRBASE. The distribution of alpha = k'(D)/k'(L) values was found to follow a power law, equivalent to an exponential decay for chiral differential free energies. This observation is experimentally relevant in terms of the number of different individual or combinatorial selectors that need to be screened in order to observe alpha values higher than a preset minimum. A string model for enantiorecognition (SMED) formalism is proposed to account for this observation on the basis of an extended Ogston three-point interaction model. Partially overlapping molecular interaction domains are analyzed in terms of a string complementarity model for ligand-receptor complementarity. The results suggest that chiral selection statistics may be interpreted in terms of more general concepts related to biomolecular recognition.     

Reviving the superorganism

Individuals become functionally organized to survive and reproduce in their environments by the process of natural selection. The question of whether larger units such as groups and communities can possess similar properties of functional organization, and therefore be regarded as "superorganisms", has a long history in biological thought. Modern evolutionary biology has rejected the concept of superorganisms, explaining virtually all adaptations at the individual or gene level. We criticize the modern literature on three counts. First, individual selection in its strong form is founded on a logical contradiction, in which genes-in-individuals are treated differently than individuals-in-groups or species-in-communities. Imposing consistency clearly shows that groups and communities can be organisms in the same sense that individuals are. Furthermore, superorganisms are more than just a theoretical possibility and actually exist in nature. Second, the view that genes are the "ultimate" unit of selection is irrelevant to the question of functional organization. Third, modern evolutionary biology includes numerous conceptual frameworks for analyzing evolution in structured populations. These frameworks should be regarded as different ways of analyzing a common process which, to be correct, must converge on the same conclusions. Unfortunately, evolutionists frequently regard them as competing theories that invoke different mechanisms, such that if one is "right" the others must be "wrong". The problem of multiple frameworks is aggravated by the fact that major terms, such as "units of selection", are defined differently within each framework, yet many evolutionists who use one framework to argue against another assume shared meanings. We suggest that focusing on the concept of organism will help dispell this fog of semantic confusion, allowing all frameworks to converge on the same conclusions regarding units of functional organization.     

The Definition and Measurement of Individual Condition in Evolutionary Studies

Over the past four decades, as the use of the term condition has become more frequent, the meaning of the term has become increasingly vague. This is especially true in evolutionary theory where condition is now equated with reproductive value, genetic quality, or defined as the ‘total pool of resources available for reproduction.’ Condition with the latter meaning is essentially impossible to measure empirically because it is associated with multiple attributes, such as nutritional state, health, experience, foraging success, ability to cope with environmental pressures, and social status, that collectively affect individual fitness. The addition of qualifying terms that often precede condition (e.g., phenotypic, energetic, and nutritional) and the usage of terms that describe an individual's state (e.g., physiological state, energetic state, and nutritional state) add confusion to the issue. It is therefore important to evaluate the meaning of condition, the limits of its usefulness, and how it can be best measured. We suggest using a more narrow definition of condition, amenable to empirical study, would benefit evolutionary and behavioral studies.     

Physicochemical evaluation of protein folds predicted by threading

Protein structure prediction remains an unsolved problem. Since prediction of the native structure seems very difficult, one usually tries to predict the correct fold of a protein. Here the "fold" is defined by the approximate backbone structure of the protein. However, physicochemical factors that determine the correct fold are not well understood. It has recently been reported that molecular mechanics energy functions combined with effective solvent terms can discriminate the native structures from misfolded ones. Using such a physicochemical energy function, we studied the factors necessary for discrimination of correct and incorrect folds. We first selected correct and incorrect folds by a conventional threading method. Then, all-atom models of those folds were constructed by simply minimizing the atomic overlaps. The constructed correct model representing the native fold has almost the same backbone structure as the native structure but differs in side-chain packing. Finally, the energy values of the constructed models were compared with that of the experimentally determined native structure. The correct model as well as the native structure showed lower energy than misfolded models. However, a large energy gap was found between the native structure and the correct model. By decomposing the energy values into their components, it was found that solvent effects such as the hydrophobic interaction or solvent shielding and the Born energy stabilized the correct model rather than the native structure. The large energetic stabilization of the native structure was attained by specific side-chain packing. The stabilization by solvent effects is small compared to that by side-chain packing. Therefore, it is suggested that in order to confidently predict the correct fold of a protein, it is also necessary to predict correct side-chain packing.     

Isothermal Titration Calorimetry of Chiral Polymeric Nanoparticles

Chiral polymeric nanoparticles are of prime importance, mainly due to their enantioselective potential, for many applications such as catalysis and chiral separation in chromatography. In this article we report on the preparation of chiral polymeric nanoparticles by miniemulsion polymerization. In addition, we describe the use of isothermal titration calorimetry (ITC) to measure the chiral interactions and the energetics of the adsorption of enantiomers from aqueous solutions onto chiral polymeric nanoparticles. The characterization of chirality in nano‐systems is a very challenging task; here, we demonstrate that ITC can be used to accurately determine the thermodynamic parameters associated with the chiral interactions of nanoparticles. The use of ITC to measure the energetics of chiral interactions and recognition at the surfaces of chiral nanoparticles can be applied to other nanoscale chiral systems and can provide further insight into the chiral discrimination processes of nanomaterials. Chirality 27:613–618, 2015. © 2015 Wiley Periodicals, Inc.     

Different terms used to describe "red tides"

An event characterized by sudden increase in phytoplankton population, in the sea or aqueous environment, is often designated by different Spanish terms that attempt to describe the nature, aspect, characteristics, and/or properties of such phenomena. In this communication, we discuss the convenience of reaching an agreement among the Spanish-speaking scientific community to use a simple Spanish term that could be much more informative and accurate when referring to Harmful Algal Blooms (HABs), in general. Summarizing the different Spanish terms historically employed to describe the proliferation of noxious phytoplankton in the sea, we propose "Proliferación Microalgal Nociva" (PMN = HAB) as a term that, on the basis of its etymological meaning could be considered correct. Its use could help to avoid the prevailing confusion in our language caused by different misleading terms now employed when referring to a Harmful Algal Bloom event.     

Rapid chiral separation methods development by cyclodextrin-mediated capillary electrophoresis for acidic and basic compounds

Chiral separation methods development using conventional techniques such as GC or HPLC requires a lot of experience, effort, and expense, due to the wide diversity of the optically active solutes and their possible chiral selectors. Capillary electrophoresis has received increased attention as an alternative technique for chiral separation due to its inherent high efficiencies and ease of methods development. However, due to the wide variety of chiral selectors available in CE, the benefits of this technique might be diminished without an appropriate methods development scheme. In this paper detailed examples are shown for fast, efficient, and predictable chiral capillary electrophoresis separation methods development based on a new and systematic theory. Optimized separations and their parameters are presented for several enantiomeric acids and bases. All the three possible cases, such as the use of low and high pH, as well as pH = pK buffer systems are thoroughly discussed. © 1995 Wiley-Liss, Inc.     

Use of a new pirkle-type chiral stationary phase in analytical and preparative subcritical fluid chromatography of pharmaceutical compounds

Good results have been obtained with use of the new bonded chiral stationary phase Whelk-O 1 in analytical and preparative subcritical fluid chromatography. A wide variety of enantiomeric pairs of compounds with different functional groups that are of pharmaceutical and biological interest have been resolved. This Pirkle-concept CSP appears to be more rugged than cellulosic phases (e.g., Chiralcel) with regards to solvents and pressure. In comparing the usefulness of the column for SFC versus HPLC chiral analysis, we have observed a clear superiority of SFC in terms of higher speed and efficiency of analysis, and faster method development. This is consistent with our experience with Chiralcel CSPs. With the Whelk-O 1 we have shown that the effects of temperature and modifier on SFC separations are similar to what has been reported for most other CSPs. We also observed a unique selectivity advantage of SFC for verapamil. We had good success with using a 1-in. diameter column packed with Whelk-O 1 to perform preparative SFC separations of a number of enantiomeric mixtures. The advantages of preparative SFC over preparative HPLC will be discussed. The feasibility of preparative SFC is dependent on how well we meet the practical challenges such as sample introduction issues, special hardware requirements due to the high pressure, and fraction collection issues. © 1994 Wiley-Liss, Inc.     

Determination of enantiomeric composition by negative-ion electrospray ionization-mass spectrometry using deprotonated N-(3,5-dinitrobenzoyl)amino acids as chiral selectors

The ability to use mixtures of deprotonated N-(3,5-dinitrobenzoyl)amino acids as chiral selectors for the determination of enantiomeric composition by electrospray ionization-mass spectrometry is demonstrated. For each experiment, two N-(3,5-dinitrobenzoyl)amino acids were chosen such that each would have opposite selectivity for the enantiomers of the analyte. Electrospray ionization-mass spectrometry, monitored in the negative ion mode, of solutions containing the two N-(3,5-dinitrobenzoyl)amino acids, sodium hydroxide, and the analyte, in a one-to-one mixture of methanol and water, afford peaks in the mass spectrum that correspond to the deprotonated 1:1 analyte-selector complexes. The ratio of the intensities of the complexes in the mass spectrum can be related to the enantiomeric composition of the analyte. Additionally, the sense and extent of chiral recognition is consistent with chromatographic observations, using chiral stationary phases derived from N-(3,5-dinitrobenzoyl)amino acids. Each analysis of enantiomeric composition requires less than 10 s to complete, indicating that this method has great potential for the development of fast-/high-throughput chiral analyses.     

Separation of drug enantiomers by liquid chromatography and capillary electrophoresis, using immobilized proteins as chiral selectors

Proteins display interesting chiral discrimination properties owing to multiple possibilities of intermolecular interactions with chiral compounds. This review deals with proteins which have been used as immobilized chiral selectors for the enantioseparation of drugs in liquid chromatography and capillary electrophoresis. The main procedures allowing the immobilization of proteins onto matrices, such as silica and zirconia particles, membranes and capillaries are first presented. Then the factors affecting the enantioseparation of drugs in liquid chromatography, using various protein-based chiral stationary phases (CSPs), are reviewed and discussed. Last, chiral separations already achieved using immobilized protein selectors in affinity capillary electrochromatography (ACEC) are presented and compared in terms of efficiency, stability and reproducibility.     

Fluorescence determination of enantiomeric composition of pharmaceuticals via use of ionic liquid that serves as both solvent and chiral selector

A new method has been developed for the sensitive and accurate determination of enantiomeric compositions of a variety of drugs, including propranolol, naproxen, and warfarin. The method is based on the use of the fluorescence technique to measure diastereomeric interactions between both enantiomeric forms of a drug with an optically active room temperature ionic liquid (RTIL) followed by partial least squares analysis of the data. The chiral RTIL used in this study, S-[(3-chloro-2-hydroxypropyl) trimethylammonium] [bis((trifluoromethyl)sulfonyl)amide] (S-[CHTA](+) [Tf(2)N](-)), is a novel chiral RTIL that has been synthesized successfully recently in our laboratory in optically pure form using a simple one-step reaction with commercially available reagents. The high solubility power and strong enantiomeric recognition ability make it possible to use this chiral RTIL to solubilize a drug and to induce diastereomeric interactions for the determination of enantiomeric purity, that is, to use it as both solvent and chiral selector. Enantiomeric compositions of a variety of pharmaceutical products with different shapes, sizes, and functional groups can be determined sensitively (microgram concentration) and accurately (enantiomeric excess as low as 0.30% and enantiomeric impurity as low as 0.08%) by use of this method.     

Chiral Polymerization in Open Systems From Chiral-Selective Reaction Rates

We investigate the possibility that prebiotic homochirality can be achieved exclusively through chiral-selective reaction rate parameters without any other explicit mechanism for chiral bias. Specifically, we examine an open network of polymerization reactions, where the reaction rates can have chiral-selective values. The reactions are neither autocatalytic nor do they contain explicit enantiomeric cross-inhibition terms. We are thus investigating how rare a set of chiral-selective reaction rates needs to be in order to generate a reasonable amount of chiral bias. We quantify our results adopting a statistical approach: varying both the mean value and the rms dispersion of the relevant reaction rates, we show that moderate to high levels of chiral excess can be achieved with fairly small chiral bias, below 10%. Considering the various unknowns related to prebiotic chemical networks in early Earth and the dependence of reaction rates to environmental properties such as temperature and pressure variations, we argue that homochirality could have been achieved from moderate amounts of chiral selectivity in the reaction rates.