Detection and chemical profiling of medicine counterfeits by Raman spectroscopy and chemometrics

Raman spectroscopy combined with chemometrics has recently become a widespread technique for the analysis of pharmaceutical solid forms. The application presented in this paper is the investigation of counterfeit medicines. This increasingly serious issue involves networks that are an integral part of industrialized organized crime. Efficient analytical tools are consequently required to fight against it. Quick and reliable authentication means are needed to allow the deployment of measures from the company and the authorities. For this purpose a method in two steps has been implemented here. The first step enables the identification of pharmaceutical tablets and capsules and the detection of their counterfeits. A nonlinear classification method, the Support Vector Machines (SVM), is computed together with a correlation with the database and the detection of Active Pharmaceutical Ingredient (API) peaks in the suspect product. If a counterfeit is detected, the second step allows its chemical profiling among former counterfeits in a forensic intelligence perspective. For this second step a classification based on Principal Component Analysis (PCA) and correlation distance measurements is applied to the Raman spectra of the counterfeits.

Diagnostic and prognostic value of cerebral 31P magnetic resonance spectroscopy in neonates with perinatal asphyxia.

The impact of depressed neonatal cerebral oxidative phosphorylation for diagnosing the severity of perinatal asphyxia was estimated by correlating the concentrations of phosphocreatine (PCr) and ATP as determined by magnetic resonance spectroscopy with the degree of hypoxic-ischemic encephalopathy (HIE) in 23 asphyxiated term neonates. Ten healthy age-matched neonates served as controls. In patients, the mean concentrations +/- SD of PCr and ATP were 0.99 +/- 0.46 mmol/L (1.6 +/- 0.2 mmol/L) and 0.99 +/- 0.35 mmol/L (1.7 +/- 0.2 mmol/L), respectively (normal values in parentheses). [PCr] and [ATP] correlated significantly with the severity of HIE (r = 0.85 and 0.9, respectively, p < 0.001), indicating that the neonatal encephalopathy is the clinical manifestation of a marred brain energy metabolism. Neurodevelopmental outcome was evaluated in 21 children at 3, 9, and 18 mo. Seven infants had multiple impairments, five were moderately handicapped, five had only mild symptoms, and four were normal. There was a significant correlation between the cerebral concentrations of PCr or ATP at birth and outcome (r = 0.8, p < 0.001) and between the degree of neonatal neurologic depression and outcome (r = 0.7). More important, the outcome of neonates with moderate HIE could better be predicted with information from quantitative 31P magnetic resonance spectroscopy than from neurologic examinations. In general, the accuracy of outcome predictability could significantly be increased by adding results from 31P magnetic resonance spectroscopy to the neonatal neurologic score, but not vice versa. No correlation with outcome was found for other perinatal risk factors, including Apgar score.

Self-Organising Maps and Correlation Analysis as a Tool to Explore Patterns in Excitation-Emission Matrix Data Sets and to Discriminate Dissolved Organic Matter Fluorescence Components

Dissolved organic matter (DOM) is a complex mixture of organic compounds, ubiquitous in marine and freshwater systems. Fluorescence spectroscopy, by means of Excitation-Emission Matrices (EEM), has become an indispensable tool to study DOM sources, transport and fate in aquatic ecosystems. However the statistical treatment of large and heterogeneous EEM data sets still represents an important challenge for biogeochemists. Recently, Self-Organising Maps (SOM) has been proposed as a tool to explore patterns in large EEM data sets. SOM is a pattern recognition method which clusterizes and reduces the dimensionality of input EEMs without relying on any assumption about the data structure. In this paper, we show how SOM, coupled with a correlation analysis of the component planes, can be used both to explore patterns among samples, as well as to identify individual fluorescence components. We analysed a large and heterogeneous EEM data set, including samples from a river catchment collected under a range of hydrological conditions, along a 60-km downstream gradient, and under the influence of different degrees of anthropogenic impact. According to our results, chemical industry effluents appeared to have unique and distinctive spectral characteristics. On the other hand, river samples collected under flash flood conditions showed homogeneous EEM shapes. The correlation analysis of the component planes suggested the presence of four fluorescence components, consistent with DOM components previously described in the literature. A remarkable strength of this methodology was that outlier samples appeared naturally integrated in the analysis. We conclude that SOM coupled with a correlation analysis procedure is a promising tool for studying large and heterogeneous EEM data sets.

Self-Organising Maps and Correlation Analysis as a Tool to Explore Patterns in Excitation-Emission Matrix Data Sets and to Discriminate Dissolved Organic Matter Fluorescence Components

Dissolved organic matter (DOM) is a complex mixture of organic compounds, ubiquitous in marine and freshwater systems. Fluorescence spectroscopy, by means of Excitation-Emission Matrices (EEM), has become an indispensable tool to study DOM sources, transport and fate in aquatic ecosystems. However the statistical treatment of large and heterogeneous EEM data sets still represents an important challenge for biogeochemists. Recently, Self-Organising Maps (SOM) has been proposed as a tool to explore patterns in large EEM data sets. SOM is a pattern recognition method which clusterizes and reduces the dimensionality of input EEMs without relying on any assumption about the data structure. In this paper, we show how SOM, coupled with a correlation analysis of the component planes, can be used both to explore patterns among samples, as well as to identify individual fluorescence components. We analysed a large and heterogeneous EEM data set, including samples from a river catchment collected under a range of hydrological conditions, along a 60-km downstream gradient, and under the influence of different degrees of anthropogenic impact. According to our results, chemical industry effluents appeared to have unique and distinctive spectral characteristics. On the other hand, river samples collected under flash flood conditions showed homogeneous EEM shapes. The correlation analysis of the component planes suggested the presence of four fluorescence components, consistent with DOM components previously described in the literature. A remarkable strength of this methodology was that outlier samples appeared naturally integrated in the analysis. We conclude that SOM coupled with a correlation analysis procedure is a promising tool for studying large and heterogeneous EEM data sets.

Determination of hydrochlorothiazide in pharmaceutical formulations by diffuse reflectance spectroscopy

This paper describes a method for quantitative spot test analysis of hydrochlorothiazide using diffuse reflectance spectroscopy. The reflectance measurements were performed analyzing the colored compound (l = 585 nm) produced from the reaction between hydrochlorothiazide and p-dimethylaminocinnamaldehyde (PDAC) in acid medium. This reaction occurred on filter paper after heating to 80ºC for 8 minutes. Factorial designs allowed varying multiple reaction factors simultaneously in order to obtain the best reaction conditions. These factors included heating temperature, heating time, acid volume and PDAC volume. The linearity was studied in the range of 3.36x10-2 to 1.01x10-1 mol L-1 with a correlation coefficient of 0.998. The limit of detection was estimated to be 1.32x10² mol L-1. Commercial samples were analyzed using the proposed method and the results were favorably compared with those of the United States Pharmacopeia method, showing that quantitative spot test analysis by diffuse reflectance could be successfully used to determine hydrochlorothiazide in medicines.

White mold detection in common beans through leaf reflectance spectroscopy

ABSTRACT This study aimed to identify wavelengths based on leaf reflectance (400-1050 nm) to estimate white mold severity in common beans at different seasons. Two experiments were carried out, one during fall and another in winter. Partial Least Squares (PLS) regression was used to establish a set of wavelengths that better estimates the disease severity at a specific date. Therefore, observations were previously divided in two sub-groups. The first one (calibration) was used for model building and the second subgroup for model testing. Error measurements and correlation between measured and predicted values of disease severity index were employed to provide the best wavelengths in both seasons. The average indexes of each experiment were of 5.8% and 7.4%, which is considered low. Spectral bands ranged between blue and green, green and red, and red and infrared, being most sensitive for disease estimation. Beyond the transition ranges, other spectral regions also presented wavelengths with potential to determine the disease severity, such as red, green, and near infrared.

Proton magnetic resonance spectroscopy of the frontal, cingulate and perirolandic cortices and its relationship to skin conductance in patients with schizophrenia

The aim of the present study was to determine whether specific subgroups of schizophrenic patients, grouped according to electrodermal characteristics, show differences in the N-acetylaspartate/creatine plus choline (NAA / (Cr + Cho)) ratios in the frontal, cingulate and perirolandic cortices. Skin conductance levels (SCL) and skin conductance responses to auditory stimulation were measured in 38 patients with schizophrenia and in the same number of matched healthy volunteers (control). All subjects were submitted to multivoxel proton magnetic resonance spectroscopic imaging. When compared to the control group, patients presented significantly lower NAA / (Cr + Cho) ratios in the right dorsolateral prefrontal cortex (schizophrenia = 0.95 ± 0.03; control = 1.12 ± 0.04) and in the right (schizophrenia = 0.88 ± 0.02; control = 0.94 ± 0.03) and left (schizophrenia = 0.84 ± 0.03; control = 0.94 ± 0.03) cingulates. These ratios did not differ between electrodermally responsive and non-responsive patients. When patients were divided into two groups: lower SCL (less than the mean SCL of the control group minus two standard deviations) and normal SCL (similar to the control group), the subgroup with a lower level of SCL showed a lower NAA / (Cr + Cho) ratio in the left cingulate (0.78 ± 0.05) than the controls (0.95 ± 0.02, P < 0.05) and the subgroup with normal SCL (0.88 ± 0.03, P < 0.05). There was a negative correlation between the NAA / (Cr + Cho) ratio in the left cingulate of patients with schizophrenia and the duration of the disease and years under medication. These data suggest the existence of a schizophrenic subgroup characterized by low SCL that could be a consequence of the lower neuronal viability observed in the left cingulate of these patients.

Studies on Laser Induced and RF Sputtered Plasma using Optical Emission Spectroscopy and Langmuir Probe

The main objective of the present study is to understand different mechanisms involved in the production and evolution of plasma by the pulsed laser ablation and radio frequency magnetron sputtering. These two methods are of particular interest, as these are well accomplished methods used for surface coatings, nanostructure fabrications and other thin film devices fabrications. Material science researchers all over the world are involved in the development of devices based on transparent conducting oxide (TCO) thin films. Our laboratory has been involved in the development of TCO devices like thin film diodes using zinc oxide (ZnO) and zinc magnesium oxide (ZnMgO), thin film transistors (TFT's) using zinc indium oxide and zinc indium tin oxide, and some electroluminescent (EL) devices by pulsed laser ablation and RF magnetron sputtering.In contrast to the extensive literature relating to pure ZnO and other thin films produced by various deposition techniques, there appears to have been relatively little effort directed towards the characterization of plasmas from which such films are produced. The knowledge of plasma dynamics corresponding to the variations in the input parameters of ablation and sputtering, with the kind of laser/magnetron used for the generation of plasma, is limited. To improve the quality of the deposited films for desired application, a sound understanding of the plume dynamics, physical and chemical properties of the species in the plume is required. Generally, there is a correlation between the plume dynamics and the structural properties of the films deposited. Thus the study of the characteristics of the plume contributes to a better understanding and control of the deposition process itself. The hydrodynamic expansion of the plume, the composition, and SIze distribution of clusters depend not only on initial conditions of plasma production but also on the ambient gas composition and pressure. The growth and deposition of the films are detennined by the thermodynamic parameters of the target material and initial conditions such as electron temperature and density of the plasma.For optimizing the deposition parameters of various films (stoichiometric or otherwise), in-situ or ex-situ monitoring of plasma plume dynamics become necessary for the purpose of repeatability and reliability. With this in mind, the plume dynamics and compositions of laser ablated and RF magnetron sputtered zinc oxide plasmas have been investigated. The plasmas studied were produced at conditions employed typically for the deposition of ZnO films by both methods. Apart from this two component ZnO plasma, a multi-component material (lead zirconium titanate) was ablated and plasma was characterized.

Size dependent fluorescence spectroscopy of nanocolloids of ZnO

In this article we present size dependent spectroscopic observations of nanocolloids of ZnO. ZnO is reported to show two emission bands, an ultraviolet (UV) emission band and another in the green region. Apart from the known band gap 380 nm and impurity 530 nm emissions, we have found some peculiar features in the fluorescence spectra that are consistent with the nanoparticle size distribution. Results show that additional emissions at 420 and 490 nm are developed with particle size. The origin of the visible band emission is discussed. The mechanism of the luminescence suggests that UV luminescence of ZnO colloid is related to the transition from conduction band edge to valence band, and visible luminescence is caused by the transition from deep donor level to valence band due to oxygen vacancies and by the transition from conduction band to deep acceptor level due to impurities and defect states. A correlation analysis between the particle size and spectroscopic observations is also discussed.

Electron energy loss spectroscopy assessment of cationic migration in La2/3Ca1/3MnO3 thin films

Màster en Nanociència i Nanotecnologia curs 2006-2007. Directors: Francesca Peiró i Martínez and Jordi Arbiol i Cobos

Quantitative analysis of mannitol polymorphs: FT-Raman spectroscopy

Mannitol is a polymorphic excipient which is usually used in pharmaceutical products as the beta form, although other polymorphs (alpha and delta) are common contaminants. Binary mixtures containing beta and delta mannitol were prepared to quantify the concentration of the beta form using FT-Raman spectroscopy. Spectral regions characteristic of each form were selected and peak intensity ratios of beta peaks to delta peaks were calculated. Using these ratios, a correlation curve was established which was then validated by analysing further samples of known composition. The results indicate that levels down to 2% beta could be quantified using this novel, non-destructive approach. Potential errors associated with quantitative studies using FT-Raman spectroscopy were also researched. The principal source of variability arose from inhomogeneities on mixing of the samples; a significant reduction of these errors was observed by reducing and controlling the particle size range. The results show that FT-Raman spectroscopy can be used to rapidly and accurately quantitate polymorphic mixtures.

Self-assembling of phenothiazine compounds investigated by small-angle X-ray scattering and electron paramagnetic resonance spectroscopy

Small-angle X-ray scattering (SAXS) and electron paramagnetic resonance (EPR) have been carried out to investigate the structure of the self-aggregates of two phenothiazine drugs, chlorpromazine (CPZ) and trifluoperazine (TFP), in aqueous solution. In the SAXS studies, drug solutions of 20 and 60 mM, at pH 4.0 and 7.0, were investigated and the best data fittings were achieved assuming several different particle form factors with a homogeneous electron density distribution in respect to the water environment. Because of the limitation of scattering intensity in the q range above 0.15 angstrom(-1), precise determination of the aggregate shape was not possible and all of the tested models for ellipsoids, cylinders, or parallelepipeds fitted the experimental data equally well. The SAXS data allows inferring, however, that CPZ molecules might self-assemble in a basis set of an orthorhombic cell, remaining as nanocrystallites in solution. Such nanocrystals are composed of a small number of unit cells (up to 10, in c-direction), with CPZ aggregation numbers of 60-80. EPR spectra of 5- and 16-doxyl stearic acids bound to the aggregates were analyzed through simulation, and the dynamic and magnetic parameters were obtained. The phenothiazine concentration in EPR experiments was in the range of 5-60 mM. Critical aggregation concentration of TFP is lower than that for CPZ, consistent with a higher hydrophobicity of TFP. At acidic pH 4.0 a significant residual motion of the nitroxide relative to the aggregate is observed, and the EPR spectra and corresponding parameters are similar to those reported for aqueous surfactant micelles. However, at pH 6.5 a significant motional restriction is observed, and the nitroxide rotational correlation times correlate very well with those estimated for the whole aggregated particle from SAXS data. This implies that the aggregate is densely packed at this pH and that the nitroxide is tightly bound to it producing a strongly immobilized EPR spectrum. Besides that, at pH 6.5 the differences in motional restriction observed between 5- and 16-DSA are small, which is different from that observed for aqueous surfactant micelles.

Electric field gradients in (111)In-doped (Hf/Zr)(3)Al(2) and (Hf/Zr)(4)Al(3) mixed compounds: ab initio calculations, perturbed angular correlation measurements and site preference

The quadrupolar hyperfine interactions of in-diffused (111)In -> (111)Cd probes in polycrystalline isostructural Zr(4)Al(3) and Hf(4)Al(3) samples containing small admixtures of the phases (Zr/Hf)(3)Al(2) were investigated. A strong preference of (111)In solutes for the contaminant (Zr/Hf)(3)Al(2) minority phases was observed. Detailed calculations of the electric field gradient (EFG) at the Cd nucleus using the full-potential augmented plane wave + local orbital formalism allowed us to assign the observed EFG fractions to the various lattice sites in the (Zr/Hf)(3)Al(2) compounds and to understand the preferential site occupation of the minority phases by the (111)In atoms. The effects of the size of the supercell and relaxation around the oversized In and Cd probe atoms were investigated in detail.

Direct Observation of Millisecond to Second Motions in Proteins by Dipolar CODEX NMR Spectroscopy

We present a site-resolved study of stow (ms to s) motions in a protein in the solid (microcrystalline) state performed with the use of a modified version of the centerband-only detection of exchange (CODEX) NMR experiment. CODEX was originally based on measuring changes in molecular orientation by means of the chemical shift anisotropy (CSA) tensor, and in our modification, angular reorientations of internuclear vectors are observed. The experiment was applied to the study of stow (15)N-(1)H motions of the SH3 domain of chicken a-spectrin. The protein was perdeuterated with partial back-exchange of protons at labile sites. This allowed indirect (proton) detection of (15)N nuclei and thus a significant enhancement of sensitivity. The diluted proton system also made negligible proton-driven spin diffusion between (15)N nuclei, which interferes with the molecular exchange (motion) and hampers the acquisition of dynamic parameters. The experiment has shown that approximately half of the peaks in the 2D (15)N-(1)H correlation spectrum exhibit exchange in a different extent. The correlation time of the slow motion for most peaks is 1 to 3 s. This is the first NMR study of the internal dynamics of proteins in the solid state on the millisecond to second time scale with site-specific spectral resolution that provides both time-scale and geometry information about molecular motions.

The Mobility of Chondroitin Sulfate in Articular and Artificial Cartilage Characterized by (13)C Magic-Angle Spinning NMR Spectroscopy

We have studied the molecular dynamics of one of the major macromolecules in articular cartilage, chondroitin sulfate. Applying (13)C high-resolution magic-angle spinning NMR techniques, the NMR signals of all rigid macromolecules in cartilage can be suppressed, allowing the exclusive detection of the highly mobile chondroitin sulfate. The technique is also used to detect the chondroitin sulfate in artificial tissue-engineered cartilage. The tissue-engineered material that is based on matrix producing chondrocytes cultured in a collagen gel should provide properties as close as possible to those of the natural cartilage. Nuclear relaxation times of the chondroitin sulfate were determined for both tissues. Although T(1) relaxation times are rather similar, the T(2) relaxation in tissue-engineered cartilage is significantly shorter. This suggests that the motions of chondroitin sulfate in data:rat and artificial cartilage different. The nuclear relaxation times of chondroitin sulfate in natural and tissue-engineered cartilage were modeled using a broad distribution function for the motional correlation times. Although the description of the microscopic molecular dynamics of the chondroitin sulfate in natural and artificial cartilage required the identical broad distribution functions for the correlation times of motion, significant differences in the correlation times of motion that are extracted from the model indicate that the artificial tissue does not fully meet the standards of the natural ideal. This could also be confirmed by macroscopic biomechanical elasticity measurements. Nevertheless, these results suggest that NMR is a useful tool for the investigation of the quality of artificially engineered tissue. (C) 2010 Wiley Periodicals, Inc. Biopolymers 93: 520-532, 2010.