Long-lived force patterns and deformation waves at repulsive epithelial boundaries.

For an organism to develop and maintain homeostasis, cell types with distinct functions must often be separated by physical boundaries. The formation and maintenance of such boundaries are commonly attributed to mechanisms restricted to the cells lining the boundary. Here we show that, besides these local subcellular mechanisms, the formation and maintenance of tissue boundaries involves long-lived, long-ranged mechanical events. Following contact between two epithelial monolayers expressing, respectively, EphB2 and its ligand ephrinB1, both monolayers exhibit oscillatory patterns of traction forces and intercellular stresses that tend to pull cell-matrix adhesions away from the boundary. With time, monolayers jam, accompanied by the emergence of deformation waves that propagate away from the boundary. This phenomenon is not specific to EphB2/ephrinB1 repulsion but is also present during the formation of boundaries with an inert interface and during fusion of homotypic epithelial layers. Our findings thus unveil a global physical mechanism that sustains tissue separation independently of the biochemical and mechanical features of the local tissue boundary.

Direct Separation of Pregabalin Enantiomers Using a Zwitterionic Chiral Selector by High Performance Liquid Chromatography Coupled to Mass Spectrometry and Ultraviolet Detection.

The chromatographic resolution of pregabalin enantiomers has been often achieved by derivatization of the molecule, in order to reach enough sensitivity at low concentrations of the minor enantiomer present in the active principle. In the present article, the development and optimization of two liquid chromatographic methods are presented for the direct resolution of pregabalin enantiomers on a chiral stationary phase (CSP) containing a zwitterionic selector derived from cinchona alkaloid and sulfonic acid (CHIRALPAK ZWIX). The key parameters for the separation as well as the compatibility of chromatographic conditions with different detection modes (ultraviolet and mass spectrometry) were investigated. The resulting methods were found to be selective, of high performance and low limits of detection (2 µg/mL by UV and 1 ng/mL by MS, respectively) and quantification (6 µg/mL by UV and 5 ng/mL by MS, respectively) for the minor enantiomer which is considered as a chiral impurity.

Multigram synthesis and in vivo efficacy studies of a novel multitarget anti-Alzheimer's compound.

We describe the multigram synthesis and in vivo efficacy studies of a donepezil‒huprine hybrid that has been found to display a promising in vitro multitarget profile of interest for the treatment of Alzheimer's disease (AD). Its synthesis features as the key step a novel multigram preparative chromatographic resolution of intermediate racemic huprine Y by chiral HPLC. Administration of this compound to transgenic CL4176 and CL2006 Caenorhabditis elegans strains expressing human Aß42, here used as simplified animal models of AD, led to a significant protection from the toxicity induced by Aß42. However, this protective effect was not accompanied, in CL2006 worms, by a reduction of amyloid deposits. Oral administration for 3 months to transgenic APPSL mice, a well-established animal model of AD, improved short-term memory, but did not alter brain levels of Aß peptides nor cortical and hippocampal amyloid plaque load. Despite the clear protective and cognitive effects of AVCRI104P4, the lack of Aß lowering effect in vivo might be related to its lower in vitro potency toward Aß aggregation and formation as compared with its higher anticholinesterase activities. Further lead optimization in this series should thus focus on improving the anti-amyloid/anticholinesterase activity ratio.

Zwitterionic chiral stationary phases based on cinchona and chiral sulfonic acids for the direct stereoselective separation of amino acids and other amphoteric compounds.

An extensive series of free amino acids and analogs were directly resolved into enantiomers (and stereoisomers where appropriate) by HPLC on zwitterionic chiral stationary phases (Chiralpak ZWIX(+) and Chiralpak ZWIX(-)). The interaction and chiral recognition mechanisms were based on the synergistic double ion-paring process between the analyte and the chiral selectors. The chiral separation and elution order were found to be predictable for primary α-amino acids with apolar aliphatic side chains. A systematic investigation was undertaken to gain an insight into the influence of the structural features on the enantiorecognition. The presence of polar and/or aromatic groups in the analyte structure is believed to tune the double ion-paring equilibrium by the involvement of the secondary interaction forces such as hydrogen bonding, Van der Waals forces and π-π stacking in concert with steric parameters. The ZWIX chiral columns were able to separate enantiomers and stereoisomers of various amphoteric compounds with no need for precolumn derivatization. Column switching between ZWIX(+) and ZWIX(-) is believed to be an instrumental tool to reverse or control the enantiomers elution order, due to the complementarity of the applied chiral selectors.

Investigation of retention behavior of natural cannabinoids on differently substituted polysaccharide-based chiral stationary phases under reversed-phase liquid chromatographic conditions.

The growing popularity of cannabis products and recent legalization of cannabis for recreational purposes have contributed to the increase of the demand for analytical methods able to give a detailed characterization of cannabis samples and derivatives. In this context, one of the aspects that is strongly emerging is about the hazardous potential of uncharacterised minor cannabinoids, including chiral ones, for which achiral potency testing methods currently employed do not give any information. For this reason, there is a growing interest towards the development of liquid chromatographic methods for the enantioseparation of cannabinoids. Much work is needed in this field where one of the major limitations is the lack of optically pure standards. This manuscript reports about the chromatographic behavior of five popular cannabinoids (including the cannabichromene racemate, CBC) on nine immobilised polysaccharide-based chiral stationary phases (CSPs) differently substituted, under reversed phase conditions. Results showed that chemo-selectivity of CSPs is not affected by changes in mobile phase composition, in the range of mobile phase investigated. In addition, the presence of electron withdrawing groups on the CSPs systematically leads to shorter retention times compared to when electron donating groups are present. An application of separation of cannabinoids from a real hemp extract on two of the chiral columns employed in this work revealed the presence of both CBC enantiomers in the sample.

Separation of regioisomers and enantiomers of underivatized saturated and unsaturated fatty acid monoacylglycerols using enantioselective HPLC.

The challenging separation of regioisomers and enantiomers of monoacylglycerols (MAGs) of fatty acids has been achieved using a single chiral HPLC column (Chiralpak® IC or Chiralpak® IA) and hexane/2-propanol eluent mixtures, without previous derivatization of the samples. The efficacy of the method is independent of the chain length and the degree of unsaturation of the fatty acids of the MAG. In addition, the separation method allows monitoring the time course for the loss of enantiomeric purity of the fatty acid-derived MAGs in polar hydroxylic solvents.

[Relationship between parathormone concentration during surgery and the post-operative outcome of primary hyperparathyroidism]. / Relación entre concentración intraoperatoria de parathormona y evolución postoperatoria del hiperparatiroidismo primario.

INTRODUCTION: The relationship between the intra-operative concentration of parathyroid hormone (IOPTH) and the long-term outcome of patients intervened due to primary hyperparathyroidism (PHPT). PATIENTS AND METHODS: A prospective observational study was performed with 120 patients. Three determinations were made of PTH in blood: baseline, when the diseases gland was located, and 10 minutes after its extirpation. The calcium, PTH and vitamin D (25-OH-D3) levels were measured during follow up. RESULTS: A decrease in IOPTH > 50% was observed in 96 (80%) patients, and the post-extirpation value returned to the normal range (Group I), in 18 (15%) a decrease of > 50% but the final value remained higher than normal (Group II) and in 6 (5%) the decrease was<50% (Group III). Persistent PHPT was detected during follow up in 6 patients (5%): one in Group I (1%), 3 (16.7%) in II and 2 (33.3%) in group III (P<.001). The risk of persistent PHPT was higher in Group II (odds ratio: 19; 95% CI: 1.85-194) and in Group III (odds ratio: 47; 95% CI: 3.53-639). There were no cases of recurrent PHPT. A normal calcium with an increased PTH was detected in 20 patients of Group I (20.8%), 11 (61.1%) in II and 3 (50%) in III (P<.001). These patients had a lower concentration of post-operative vitamin D (17 ng/ml, range: 24; compared to 28 ng/ml, range: 21) (P=.008) and higher frequency of hypovitaminosis D (70.6% compared to 26.2%) (P>.001). CONCLUSION: The risk of persistent PHPT is higher when the IOPTH decreases more than 50% but still remains high.

New approaches of LC-MS compatible method development on α(1)-acid glycoprotein-based stationary phase for resolution of enantiomers by HPLC.

Protein-based chiral stationary phases, in particular α(1)-acid glycoprotein, are chromatographic materials with a very broad application range. The chiral recognition ability of this selector allows the resolution of more than 90% of the molecules tested. A step-by-step description of the screening approach that we have developed is described in this article with two objectives in mind: obtaining a high success rate with a straightforward methodology that can be generally applied and allowing LC-MS compatibility. Second, the screening strategy is required to take into account the main functional nature of the racemic compound to be resolved (neutral, acidic or basic) for method optimisation step, but should be independent of other structural features, in order to allow the routine application of screening sequences and the application to the largest number of samples.

Enantiomer resolution screening strategy using multiple immobilised polysaccharide-based chiral stationary phases.

CHIRALPAK IA, CHIRALPAK IB and CHIRALPAK IC are a new generation of chiral packing materials for resolution of enantiomers by chromatography. They are all prepared by chemical immobilisation of the respective polysaccharide derivatives onto a supporting silica matrix. The present article aims to establish an understanding of the global enantioselective profiles of this series of chiral stationary phases (CSPs). When used in combination, high success rates for enantiomer resolution could be achieved. These three CSPs exhibit an exceptional level of complementary chiral recognition characteristics by applying common screening and method optimisation strategies.

Common approaches for efficient method development with immobilised polysaccharide-derived chiral stationary phases.

Immobilised polysaccharide-based chiral stationary phases (CSPs) are a new generation of chromatographic materials combining the remarkable enantioselective performance of the polysaccharide derivatives and solvent versatility for enantiomeric resolution. Based on extensive experimental work, this article will focus on the approach to efficient method development with these immobilised CSPs (CHIRALPAK IA, CHIRALPAK IB and CHIRALPAK IC) by applying a limited number of mobile phases. The manuscript will review the development of screening strategies, either by liquid and supercritical chromatography (LC and SFC), for the separation of enantiomers. The rational combination of both modes leads to efficient approaches to get enantiomeric resolutions in reasonable time frames and with high success rates.

Cellulose tris(3,5-dichlorophenylcarbamate) immobilised on silica: a novel chiral stationary phase for resolution of enantiomers.

CHIRALPAK IC is a new chiral stationary phase (CSP) made by immobilising cellulose tris(3,5-dichlorophenylcarbamate) on silica gel. The chiral selector is distinct from any other commercially available polysaccharide-based CSPs. Apart from its compatibility with the whole series of solvents; this CSP is able to operate under various chromatographic conditions and bring about new characteristics in enantiomeric recognition. It can afford many large and specific enantiomeric separations. It exhibits complementary properties with regard to the existing immobilised chiral packing materials of the same category.

Optimization of the chiral separation of a Ca-sensitizing drug on an immobilized polysaccharide-based chiral stationary phase: case study with a preparative perspective.

Sample solubility in the mobile phase and enantioselectivity are key factors in chiral preparative chromatography. In the search for a high throughput process for production of pure enantiomers, the rational design of the mobile phase and the selection of a suitable chiral stationary phase (CSP) are essential. However, one may sometimes be faced with the incompatibility between the CSP and the preferential eluent for sample solubility. Such a limitation may be circumvented by using an immobilized CSP such as CHIRALPAK IA. In this manuscript, the chiral separation of a Ca-sensitizing drug (EMD 53986) is optimized on CHIRALPAK IA in terms of sample solubility, enantioselectivity and preparative productivity. The approaches for method optimization and the impact of sample solubility on productivity are discussed. The preparative potential of CHIRALPAK IA is also demonstrated.

Solvent versatility of immobilized 3,5-dimethylphenylcarbamate of amylose in enantiomeric separations by HPLC.

CHIRALPAK IA is a new chiral stationary phase containing amylose 3.5-dimethylphenylcarbamate immobilized onto silica gel. It is compatible with the whole range of organic solvents. Its solvent versatility has been thoroughly investigated. The option to use a wide range of solvents, especially the "non-standards" ones, in the mobile phase enables the enhancement of chiral separation methods in terms of enantioselectivity, resolution, analysis time, sample injection and sample solubility. Parameters such as the mobile phase type, the nature of modifier and eluting strength of various solvents are examined and discussed. A guideline for method development and optimization on CHIRALPAK IA is also proposed.

Method development and optimization on cinchona and chiral sulfonic acid-based zwitterionic stationary phases for enantiomer separations of free amino acids by high-performance liquid chromatography.

CHIRALPAK ZWIX(+) and ZWIX(-) are cinchona alkaloid-derived zwitterionic chiral stationary phases (CSPs) containing a chiral sulfonic acid motif which serves as negatively charged interaction site. They are versatile for direct enantiomer resolution of amino acids and many other ampholytic compounds by HPLC. The synergistic double ion-pairing between the zwittrionic chiral selector and the ampholyte is the basis for interaction and chiral recognition mechanisms. ZWIX(+) and ZWIX(-) type CSPs or columns behave pseudo-enantiomerically and provide the feature of reversing enantiomer elution order by column switching. In the current study, extensive experimental work was carried out with the aim of developing schemes for an efficient generic screening and proposing straightforward approaches for method optimization on these ZWIX columns. Various chromatographic parameters were investigated using a large series of diverse amino acids and analogues for the purpose. The role of methanol (MeOH) as the protic solvent in the mobile phase is confirmed to be essential. The presence of water in a low percentage is beneficial for peak shape, resolution, analysis speed, sample solubility and MS detection performance. The involvement of acetonitrile (ACN) or tetrahydrofuran (THF) can help for adjusting retention time and selectivity. Incorporation of a suitable pair of acidic-basic additives at a right ratio in the mobile phase is determinant as well for the double ion-pairing mechanism. 50 mM formic acid+25 mM diethylamine (or ammonium hydroxide) in MeOH/ACN/H2O and in MeOH/THF/H2O at 49:49:2 (by volume) are recommended as the starting mobile phases for method development. Some other parameters are also considered in the proposed scheme to achieve successful enantioselective or stereoselective separation of the ampholytes.

On the method development of immobilized polysaccharide chiral stationary phases in supercritical fluid chromatography using an extended range of modifiers.

Polysaccharide-derived selectors are often used in the separation of enantiomers by supercritical fluid chromatography (SFC). Their recognition patterns are normally investigated with alcohols and acetonitrile as modifiers. The present paper describes the results of a research program designed by Pfizer and Chiral Technologies Inc. to explore the potential of other solvents (i.e. ethyl acetate, tetrahydrofuran, dichloromethane) in SFC by using a series of polysaccharide-derived supports with broad solvent versatility (CHIRALPAK IA, IB, IC, ID, IE and IF). The contribution of such extended solvent range to the overall success rate, as well as to overcome racemization, solubility and stability issues was confirmed by using standard non-proprietary samples and research molecules. Elution patterns with such lower polarity solvents, compared to alcohols, and the role of the different additives were also investigated.

Common screening approaches for efficient analytical method development in LC and SFC on columns packed with immobilized polysaccharide-derived chiral stationary phases.

Owing to their remarkable enantioselectivity, versatility, and stability, immobilized polysaccharide-based chiral stationary phases (CSPs) have been successfully integrated into the tool box of many research and industry groups for the separation of enantiomers or stereoisomers by liquid and supercritical fluid chromatography. Due to the structurally diverse range of compounds available, efficient method development for chiral separations utilizing such CSPs is a challenging subject. In this chapter, we will discuss simplified screening protocols and straightforward approaches to achieve chiral separations in HPLC and SFC using the column series CHIRALPAK™ IA, IB, IC, and ID in reasonable time frame and with limited experimental work and a high success rate.

Sexual function, satisfaction with life and menopausal symptoms in middle-aged women.

PURPOSE: To assess sexual function, satisfaction with life (SWL), and menopause-related symptoms among mid-aged Spanish women. MATERIALS AND METHODS: Cross-sectional study of 260 women, aged 40-59, attending the public gynecology consultations completed the 14-item Changes in Sexual Functioning Questionnaire (CSFQ-14), the SWL Scale (SWLS), the Menopause Rating Scale (MRS), and a socio-demographic questionnaire. RESULTS: Median [inter quartile range] age was 47 [8.0] years, 87.7% had a stable partner, 27.0% were postmenopausal, and 53.9% had increased body mass index (BMI). The prevalence of sexual dysfunction was 46.5% (CSFQ-14 score≤41). Postmenopausal status was associated with lower CSFQ-14 scores (worse sexual function) and severe menopausal symptoms whereas there were not significant differences in SWLS scores. CSFQ-14 scores correlated with SWLS (p<0.04), and inversely correlated with menopausal symptoms (p<0.02). Multiple linear regression analysis model predicted 26.6% of the total CSFQ-14 score variance, and higher scores (better sexual function) were correlated with better SWL, and inversely correlated to female age and worse menopausal symptoms. A second model predicted 38.4% of the SWLS score variance. The SWLS score correlated with the total CSFQ-14 score and BMI, and inversely correlated with economical problems, female tobacco use, lack of healthiness, menopausal symptoms, not having a partner, and partner's lack of healthiness. CONCLUSIONS: Lower sexual function was related to low SWL, age and menopausal symptoms while low SWLS score was related with economical problems, smoking, menopausal symptoms, and partner factors.

Complementary enantiorecognition patterns and specific method optimization aspects on immobilized polysaccharide-derived chiral stationary phases.

The immobilized polysaccharide-derived chiral stationary phases (CSPs) combine the benefits of broad application scope and high preparative potential with CSP robustness and universal solvent compatibility. Strategies for efficient and straightforward method development with these CSPs were previously overviewed. In the current study, six CSPs with different structural features in the immobilized series were examined for their complementary properties in separation of enantiomers. Some method development aspects related to polysaccharide-derived CSPs in general and certain specific features observed on these immobilized supports, such as the effects of mobile phase, sample solvent and mobile phase additive on chiral separation, are also discussed.

Evolution of target organ damage by different values of self-blood pressure measurement in untreated hypertensive patients.

BACKGROUND: To determine the prognostic value of various self-blood pressure (BP) monitoring (SBPM) cutoff at the time of diagnosis. METHODS: Cohort of 466 newly diagnosed and never-treated hypertensive patients. At baseline and at 1 year, the patients underwent a physical examination, clinic BP (CBP), SBPM, and ambulatory BP monitoring (ABPM), fasting blood and urine analysis, electrocardiogram (ECG), and retinography. The diagnosis of hypertension was made based on CBP average of two readings, separated by 2 min, taken over three different days, with results ≥ 140/90 mm Hg. At 1-year follow-up, target organ damage (TOD) evolution was classified as favorable or unfavorable. RESULTS: Mean age was 57.4 years, 56.8% were men. Adjusted multivariate analysis showed that hypertensive patients with baseline SBPM <135/85 mm Hg had a more favorable evolution of left ventricular hypertrophy (LVH) (odds ratio (OR): 1.9; 95% confidence interval (CI): 1.5-2.5), high urinary albumin excretion rate (UAER) (OR: 6.9; 95% CI: 3.4-14.4), and more favorable amount of TOD evolution (OR: 1.7; 95% CI: 1.4-2.0) than those with baseline SBPM ≥ 135/85 mm Hg. Patients with baseline SBPM <130/80 mm Hg, or <125/80 mm Hg had a more favorable evolution of the amount of TOD (OR: 2.7; 95% CI: 2.0-3.6, and OR: 2.9; 95% CI: 2.1-4.1, respectively) at 1 year than those with baseline SBPM <135/85 mm Hg. CONCLUSIONS: Baseline SBPM values <130/80 mm Hg is associated with better evolution of amount of TOD than SBPM values <135/85 mm Hg. These results would support a clinical trial to test a SBPM threshold <130/80 as an optimal pressure not needing pharmacological treatment among those with CBP ≥ 140/90.

Reversed-phase screening strategies for liquid chromatography on polysaccharide-derived chiral stationary phases.

Immobilised polysaccharide-based chiral stationary phases (CSPs) are chromatographic materials that combine the remarkable enantioselective performance of the polysaccharide derivatives in addition to solvent versatility for enantiomeric resolution. Their behaviour under normal phase conditions and polar organic mode has been quite extensively discussed in several scientific communications. This article will focus on an approach to developing efficient chiral analytical methods with these immobilised CSPs (CHIRALPAK IA, CHIRALPAK IB and CHIRALPAK IC) by applying a limited number of mobile phases under reversed-phase conditions. The manuscript will review the development of screening strategies by liquid chromatography for the separation of enantiomers in combination with applications compatible with LC-MS. The rational combination of this technique and the different supports will allow the identification of enantiomeric resolutions in reasonable time frames and with high success rates.