Measured proton electromagnetic structure deviates from theoretical predictions.

The visible world is founded on the proton, the only composite building block of matter that is stable in nature. Consequently, understanding the formation of matter relies on explaining the dynamics and the properties of the proton's bound state. A fundamental property of the proton involves the response of the system to an external electromagnetic field. It is characterized by the electromagnetic polarizabilities1 that describe how easily the charge and magnetization distributions inside the system are distorted by the electromagnetic field. Moreover, the generalized polarizabilities2 map out the resulting deformation of the densities in a proton subject to an electromagnetic field. They disclose essential information about the underlying system dynamics and provide a key for decoding the proton structure in terms of the theory of the strong interaction that binds its elementary quark and gluon constituents. Of particular interest is a puzzle in the electric generalized polarizability of the proton that remains unresolved for two decades2. Here we report measurements of the proton's electromagnetic generalized polarizabilities at low four-momentum transfer squared. We show evidence of an anomaly to the behaviour of the proton's electric generalized polarizability that contradicts the predictions of nuclear theory and derive its signature in the spatial distribution of the induced polarization in the proton. The reported measurements suggest the presence of a new, not-yet-understood dynamical mechanism in the proton and present notable challenges to the nuclear theory.

First Concurrent Extraction of the Leading-Order Scalar and Spin Proton Polarizabilities.

We performed the first simultaneous extraction of the six leading-order proton polarizabilities. We reached this milestone thanks to both new high-quality experimental data and an innovative bootstrap-based fitting method. These new results provide a self-consistent and fundamental benchmark for all future theoretical and experimental polarizability estimates.

Light-front interpretation of proton generalized polarizabilities.

We extend the recently developed formalism to extract light-front quark charge densities from nucleon form factor data to the deformations of these quark charge densities when applying an external electric field. We show that the resulting induced polarizations can be extracted from proton generalized polarizabilities. The available data for the generalized electric polarizability of the proton yield a pronounced structure in its induced polarization at large transverse distances, which will be pinned down by forthcoming high precision virtual Compton scattering experiments.

Quality by design compliant strategy for the development of a liquid chromatography-tandem mass spectrometry method for the determination of selected polyphenols in Diospyros kaki.

Diospyros kaki fruits possess great beneficial properties for human health due to their strong antioxidant and antiradical activities related to the high level of bioactive compounds and particularly polyphenols. In this paper a rapid and efficient liquid chromatography-tandem mass spectrometry method for the determination of 38 polyphenolic compounds in Diospyros kaki flesh was developed. The optimization of the chromatographic method was performed applying a Quality by Design approach, which is unexplored in the field of food analysis. The Critical Method Attributes (CMAs) were the critical resolutions of some isobaric compounds and analysis time. The Critical Methods Parameters (CMPs) were related to the characteristics of both the mobile phase and the column: flow rate, temperature, starting organic phase concentration of the mobile phase, formic acid percentage in the eluents, type of organic solvent in the mobile phase and gradient of organic eluents. The effects of the CMPs on the CMAs were evaluated by experimental design, at first carrying out a screening phase by an asymmetric screening matrix and then applying Response Surface Methodology by a Doehlert Design. The quadratic polynomial models postulated to link the CMAs to CMPs were calculated and the Method Operable Design Region was identified with the aid of Monte Carlo simulations as the multidimensional combination of the CMPs that satisfied the requirements for the CMAs with a probability ≥90%. The developed method was applied to real samples obtained by the extraction of Diospyros kaki flesh from two different cultivars (Rojo Brillante and Kaki Tipo), making it possible to obtain extensive information on their polyphenolic profiles.

Transverse pion structure beyond leading twist in constituent models.

The understanding of the pion structure as described in terms of transverse-momentum-dependent parton distribution functions (TMDs) is of importance for the interpretation of currently ongoing Drell-Yan experiments with pion beams. In this work we discuss the description of pion TMDs beyond leading twist in a pion model formulated in the light-front constituent framework. For comparison, we also review and derive new results for pion TMDs in the bag and spectator model.

Enantioseparation and impurity determination of ambrisentan using cyclodextrin-modified micellar electrokinetic chromatography: Visualizing the design space within quality by design framework.

A capillary electrophoresis method for the simultaneous determination of the enantiomeric purity and of impurities of the chiral drug ambrisentan has been developed following the Quality by Design principles. The selected separation system consisted of a micellar pseudostationary phase made by sodium dodecyl sulphate with the addition of γ-cyclodextrin. The effects of critical process parameters (capillary length, temperature, voltage, borate concentration, pH, sodium dodecyl sulphate concentration, γ-cyclodextrin concentration) on enantioresolution of ambrisentan and analysis time were extensively investigated by multivariate strategies involving a screening phase and Response Surface Methodology. The Design Space was defined with a desired probability level π≥90%, and the working conditions, with the limits of the Design Space, corresponded to the following: capillary length, 64.5cm; temperature, 22°C; voltage, 30kV (26-30kV); background electrolyte, 100mM borate buffer pH 9.20 (8.80-9.60), 100mM sodium dodecyl sulphate, 50mM (43-50mM) γ-cyclodextrin. A Plackett-Burman design was applied for robustness testing, and a method control strategy was established. The method was fully validated according to the International Conference on Harmonisation guidelines and was applied to ambrisentan coated tablets.

A comprehensive strategy in the development of a cyclodextrin-modified microemulsion electrokinetic chromatographic method for the assay of diclofenac and its impurities: Mixture-process variable experiments and quality by design.

A comprehensive strategy involving the use of mixture-process variable (MPV) approach and Quality by Design principles has been applied in the development of a capillary electrophoresis method for the simultaneous determination of the anti-inflammatory drug diclofenac and its five related substances. The selected operative mode consisted in microemulsion electrokinetic chromatography with the addition of methyl-ß-cyclodextrin. The critical process parameters included both the mixture components (MCs) of the microemulsion and the process variables (PVs). The MPV approach allowed the simultaneous investigation of the effects of MCs and PVs on the critical resolution between diclofenac and its 2-deschloro-2-bromo analogue and on analysis time. MPV experiments were used both in the screening phase and in the Response Surface Methodology, making it possible to draw MCs and PVs contour plots and to find important interactions between MCs and PVs. Robustness testing was carried out by MPV experiments and validation was performed following International Conference on Harmonisation guidelines. The method was applied to a real sample of diclofenac gastro-resistant tablets.

Quality by design in the chiral separation strategy for the determination of enantiomeric impurities: development of a capillary electrophoresis method based on dual cyclodextrin systems for the analysis of levosulpiride.

Quality by design (QbD) concepts, in accordance with International Conference on Harmonisation Pharmaceutical Development guideline Q8(R2), represent an innovative strategy for the development of analytical methods. In this paper QbD principles have been comprehensively applied in the set-up of a capillary electrophoresis method aimed to quantify enantiomeric impurities. The test compound was the chiral drug substance levosulpiride (S-SUL) and the developed method was intended to be used for routine analysis of the pharmaceutical product. The target of analytical QbD approach is to establish a design space (DS) of critical process parameters (CPPs) where the critical quality attributes (CQAs) of the method have been assured to fulfil the desired requirements with a selected probability. QbD can improve the understanding of the enantioseparation process, including both the electrophoretic behavior of enantiomers and their separation, therefore enabling its control. The CQAs were represented by enantioresolution and analysis time. The scouting phase made it possible to select a separation system made by sulfated-ß-cyclodextrin and a neutral cyclodextrin, operating in reverse polarity mode. The type of neutral cyclodextrin was included among other CPPs, both instrumental and related to background electrolyte composition, which were evaluated in a screening phase by an asymmetric screening matrix. Response surface methodology was carried out by a Doehlert design and allowed the contour plots to be drawn, highlighting significant interactions between some of the CPPs. DS was defined by applying Monte-Carlo simulations, and corresponded to the following intervals: sulfated-ß-cyclodextrin concentration, 9-12 mM; methyl-ß-cyclodextrin concentration, 29-38 mM; Britton-Robinson buffer pH, 3.24-3.50; voltage, 12-14 kV. Robustness of the method was examined by a Plackett-Burman matrix and the obtained results, together with system repeatability data, led to define a method control strategy. The method was validated and was finally applied to determine the enantiomeric purity of S-SUL in pharmaceutical dosage forms.

An integrated quality by design and mixture-process variable approach in the development of a capillary electrophoresis method for the analysis of almotriptan and its impurities.

The development of a capillary electrophoresis (CE) method for the assay of almotriptan (ALM) and its main impurities using an integrated Quality by Design and mixture-process variable (MPV) approach is described. A scouting phase was initially carried out by evaluating different CE operative modes, including the addition of pseudostationary phases and additives to the background electrolyte, in order to approach the analytical target profile. This step made it possible to select normal polarity microemulsion electrokinetic chromatography (MEEKC) as operative mode, which allowed a good selectivity to be achieved in a low analysis time. On the basis of a general Ishikawa diagram for MEEKC methods, a screening asymmetric matrix was applied in order to screen the effects of the process variables (PVs) voltage, temperature, buffer concentration and buffer pH, on critical quality attributes (CQAs), represented by critical separation values and analysis time. A response surface study was then carried out considering all the critical process parameters, including both the PVs and the mixture components (MCs) of the microemulsion (borate buffer, n-heptane as oil, sodium dodecyl sulphate/n-butanol as surfactant/cosurfactant). The values of PVs and MCs were simultaneously changed in a MPV study, making it possible to find significant interaction effects. The design space (DS) was defined as the multidimensional combination of PVs and MCs where the probability for the different considered CQAs to be acceptable was higher than a quality level π=90%. DS was identified by risk of failure maps, which were drawn on the basis of Monte-Carlo simulations, and verification points spanning the design space were tested. Robustness testing of the method, performed by a D-optimal design, and system suitability criteria allowed a control strategy to be designed. The optimized method was validated following ICH Guideline Q2(R1) and was applied to a real sample of ALM coated tablets.

Quality by Design approach in the development of a solvent-modified micellar electrokinetic chromatography method: finding the design space for the determination of amitriptyline and its impurities.

A solvent-modified micellar electrokinetic chromatography method was set up for the simultaneous determination of the tricyclic antidepressant amitriptyline (AMI) and its main impurities. The method was developed following Quality by Design (QbD) principles according to ICH Guideline Q8(R2). QbD approach made it possible to find the design space (DS), where quality was assured. After a scouting phase, aimed at selecting a suitable capillary electrophoresis pseudostationary phase, risk assessment tools were employed to define the critical process parameters (CPPs) to be considered in a screening phase (applied voltage, concentration and pH of the background electrolyte, concentration of the surfactant sodium dodecyl sulphate, of the cosurfactant n-butanol and of the organic modifiers acetonitrile and urea). The effects of the seven selected CPPs on critical quality attributes (CQAs), namely resolution values between critical peak pairs and analysis time, were investigated throughout the knowledge space by means of a symmetric screening matrix. Response surface study was then carried out on four selected CPPs by applying a Doehlert Design. Monte-Carlo simulations were performed in order to estimate the probability of meeting the desired specifications on CQAs, and thus to define the DS by means of a risk of failure map. Additional points at the edges of the DS were tested in order to verify the requirements for CQAs to be fulfilled. A control strategy was implemented by defining system suitability tests. The developed method was validated following ICH Guideline Q2(R1), including robustness assessment by Plackett-Burman design, and was applied to the analysis of real samples of amitriptyline coated tablets.

Mixture-process variable approach to optimize a microemulsion electrokinetic chromatography method for the quality control of a nutraceutical based on coenzyme Q10.

In recent years, multivariate optimization has played an increasing role in analytical method development. ICH guidelines recommend using statistical design of experiments to identify the design space, in which multivariate combinations of composition variables and process variables have been demonstrated to provide quality results. Considering a microemulsion electrokinetic chromatography method (MEEKC), the performance of the electrophoretic run depends on the proportions of mixture components (MCs) of the microemulsion and on the values of process variables (PVs). In the present work, for the first time in the literature, a mixture-process variable (MPV) approach was applied to optimize a MEEKC method for the analysis of coenzyme Q10 (Q10), ascorbic acid (AA), and folic acid (FA) contained in nutraceuticals. The MCs (buffer, surfactant-cosurfactant, oil) and the PVs (voltage, buffer concentration, buffer pH) were simultaneously changed according to a MPV experimental design. A 62-run MPV design was generated using the I-optimality criterion, assuming a 46-term MPV model allowing for special-cubic blending of the MCs, quadratic effects of the PVs, and some MC-PV interactions. The obtained data were used to develop MPV models that express the performance of an electrophoretic run (measured as peak efficiencies of Q10, AA, and FA) in terms of the MCs and PVs. Contour and perturbation plots were drawn for each of the responses. Finally, the MPV models and criteria for the peak efficiencies were used to develop the design space and an optimal subregion (i.e., the settings of the mixture MCs and PVs that satisfy the respective criteria), as well as a unique optimal combination of MCs and PVs.