Miren Ostra

Study of a fluorometric-enzymatic method for bilirubin based on chemically modified bilirubin-oxidase and multivariate calibration

The chemical derivatization of bilirubin oxidase (BOx) with a fluorescein derivative (FS) yields a chemically modified enzyme (BOx-FS), with excitation and emission maxima at 487 and 520 nm, respectively. During the oxygen oxidation reaction of bilirubins, in the presence of the modified enzyme, the change in the fluorescence of the modified enzyme depends on the concentration and type of bilirubin. This effect can be used for analytical purposes. Firstly, a theoretical-experimental study of the analytical system was carried out. The mechanism responsible for the fluorescence variation was clarified, a mathematical model developed and the variables affecting the fluorescence changes optimized. The concentration ranges in which the model can be applied (up to 12 mg bilirubin l(-1)), and the precision of the measurement (about 4%) were established for the three bilirubins. The application of the methodology to the simultaneous determination of direct and total bilirubins were studied by applying multivariate calibration methods to the whole kinetic profiles. A reduced calibration matrix (derived from a 5(3) base matrix) is proposed for calibration and different numerical methods were tested: Principal Components Regression (PCR), Partial Least Squares Regression (PLS) and Artificial Neural Networks (ANN). The simultaneous determination of direct and total bilirubin (average validation errors of about 9 and 10%, respectively) can be carried out from a single run. Furthermore, a semi-quantitative speciation of the three bilirubins (free, conjugated and albumin-bonded bilirubin) may be simultaneously obtained.

Linear and non-linear multivariate calibration methods for multicomponent kinetic determinations in cases of severe non-linearity

Both p-aminophenol (p-AP) and p-phenylenediamine (p-PDA) undergo oxidative coupling reactions in the presence of octacyanomolibdate(V) as oxidant. The kinetics of these reactions can be followed spectrophotometrically. When they happen to be in the same reaction mixture, relevant characteristics are (1) related to reaction conditions; the reagent must be in deficit and (2) related to the reaction products; ‘cross-coupling’ reactions are also involved. All this gives rise to systems with severe non-linearities. This is the reason why they have been used here to test the predictive ability of several linear and non-linear algorithms in cases of special complexity. Principal component regression (PCR), partial least squares regression (PLS) and three different forms of applying artificial neural networks (ANNs), with and without reduction of variables, have been tested. All the models were applied to the same calibration and validation sets. Typical prediction errors ranged between 2 and 5%. Not very different results were found when independent accuracy and precision studies were accomplished, regardless of whether linear or non-linear calibration methods were used. This is probably due to differences in the spectroscopic as well as the kinetic behaviour of both analytes. These differences allowed the construction of ANN calibration models with only 30 original variables (kinetic profiles of 10 data points at three selected different wavelengths).

Optimization and validation of a nonaqueous micellar electrokinetic chromatography method for determination of polycyclic musks in perfumes

A nonaqueous micellar electrokinetic chromatography method was developed for determination of Tonalide®, Galaxolide®, and Traseolide® polycyclic musks (PCMs). These compounds are widely used as fragrance ingredients in cosmetics. The method was optimized by using a three variable Box-Behnken experimental design and response surface methodology. A modified chromatographic response function was defined in order to adequately weigh the terms in the response function. After optimization of experimental conditions, an electrolyte solution of 195 mM SDS and 40 mM NaH(2) PO(4) in formamide was selected for the separation of the three PCMs, and the applied voltage was fixed at 30 kV. The nonaqueous MEKC method was then checked in terms of linearity, limits of detection and quantification, repeatability, intermediate precision and accuracy, providing appropriate values (i.e. RSD values for precision never exceeding 7%, and accuracy 96-107%). Nonaqueous MEKC for determination of the selected compounds was successfully applied to the analysis of commercial perfume samples.

New way of application of the bromate–bromide mixture in kinetic analysis☆

Abstract The bromine generated in situ by the bromate–bromide mixture has usually been used in kinetic analysis to determine either individual species that give reaction with bromine or the total concentration of mixtures of those analytes. In this paper the bromate–bromide mixture is proposed as a reagent for multicomponent kinetic determinations. Mixtures of acetylsalicylic acid (ASA) and paracetamol (AAP) have been used to test the discriminating ability of reactions with bromine. The reaction between bromine and AAP is fast and the reaction between bromine and ASA is negligible. However, ASA slowly hydrolyzes to yield salicylic acid (SA) and acetic acid, and the reaction between bromine and SA can be considered fast in the reaction conditions. This complex behaviour contains enough information to resolve mixtures of ASA and AAP after applying partial least squares regression (PLS) to the kinetic photometric data. After optimization, errors around 5–6% were found for the determination of ASA and AAP. Quimiometric considerations on the calibration models are made. The method has been successfully applied to the determination of ASA and AAP in pharmaceuticals.

Image analysis for maintenance of coating quality in nickel electroplating baths--real time control.

The aim of this paper is to show how it is possible to extract analytical information from images acquired with a flatbed scanner and make use of this information for real time control of a nickel plating process. Digital images of plated steel sheets in a nickel bath are used to follow the process under degradation of specific additives. Dedicated software has been developed for making the obtained results accessible to process operators. This includes obtaining the RGB image, to select the red channel data exclusively, to calculate the histogram of the red channel data and to calculate the mean colour value (MCV) and the standard deviation of the red channel data. MCV is then used by the software to determine the concentration of the additives Supreme Plus Brightner (SPB) and SA-1 (for confidentiality reasons, the chemical contents cannot be further detailed) present in the bath (these two additives degrade and their concentration changes during the process). Finally, the software informs the operator when the bath is generating unsuitable quality plating and suggests the amount of SPB and SA-1 to be added in order to recover the original plating quality.

Additive Determination in an Electrolytic Zinc Bath by UV-Visible Spectroscopy and Multivariate Calibration

Benzoic acid (BA) and benzylidene acetone (BDA), together with polyethylene glycol, are used as additives in electroplating zinc baths. The simultaneous BA and BDA determination is proposed by applying partial least-squares regression to the ultraviolet (UV)-visible spectra between 216 and 350 nm. Concentrations between 1.02 and 10.2 g L -1 BA and 0.100 and 0.550 g L -1 BDA can be determined. Several proposals to estimate the multivariate limit of detection (LOD) have been applied, including the use of the net analytical signal, found vs added plots, and the use of multivariate standard deviation. Depending on the method used, the values of LOD found the range between 0.1 and 0.4 g L -1 for BA and 0.01 and 0.03 g L -1 for BDA. The multivariate method has been applied to follow BA and BDA concentrations along a Zn bath life. BA concentration remains practically unchanged, but BDA concentration continuously diminishes by degradation on electrodes and by evaporation. There is a direct relation between BDA concentration and brightness of the plated piece. Thus, the proposed method can be related to brightness.

Flatbed scanners as a source of imaging. Brightness assessment and additives determination in a nickel electroplating bath

Desktop flatbed scanners are very well-known devices that can provide digitized information of flat surfaces. They are practically present in most laboratories as a part of the computer support. Several quality levels can be found in the market, but all of them can be considered as tools with a high performance and low cost. The present paper shows how the information obtained with a scanner, from a flat surface, can be used with fine results for exploratory and quantitative purposes through image analysis. It provides cheap analytical measurements for assessment of quality parameters of coated metallic surfaces and monitoring of electrochemical coating bath lives. The samples used were steel sheets nickel-plated in an electrodeposition bath. The quality of the final deposit depends on the bath conditions and, especially, on the concentration of the additives in the bath. Some additives become degraded with the bath life and so is the quality of the plate finish. Analysis of the scanner images can be used to follow the evolution of the metal deposit and the concentration of additives in the bath. Principal component analysis (PCA) is applied to find significant differences in the coating of sheets, to find directions of maximum variability and to identify odd samples. The results found are favorably compared with those obtained by means of specular reflectance (SR), which is here used as a reference technique. Also the concentration of additives SPB and SA-1 along a nickel bath life can be followed using image data handled with algorithms such as partial least squares (PLS) regression and support vector regression (SVR). The quantitative results obtained with these and other algorithms are compared. All this opens new qualitative and quantitative possibilities to flatbed scanners.

Quantitative determination of additives in a commercial electroplating nickel bath by spectrophotometry and multivariate analysis

An electroplating nickel bath is usually composed of a number of organic additives to improve the plating process as well as to preserve its durability. Supreme Plus Brightener (SPB) and A-5(2X), used in a commercial electroplating nickel bath show highly overlapped UV-Vis spectra. These two additives are the only ones that present absorbance in the UV-Vis wavelength range. Therefore, a mixture of them can be resolved using multivariate calibration methods of UV-Vis measurements. In this work, Partial Least Squares (PLS) regression and Classical Least Squares (CLS) have been used to quantify both additives during the whole duration of an electroplating nickel bath. It was found that PLS regression provided the best results. To avoid negative influence of baseline drifts, first derivative spectra were used. Between 0.14 and 1.40 mL of the commercial product SPB per L of nickel bath can be determined with mean errors of about 6%. Between 4 and 24 mL of the commercial product A-5(2X) per L of nickel bath can be determined with mean errors of about 8%. The limits of detection (LOD) found for SPB and A-5(2X) were 0.11 mL L−1 and 3 mL L−1 respectively. The calibration models proved to be valid for at least eight months, including a change of spectrophotometer. The SPB concentration decays along the bath life according to a first order rate law, but A-5(2X) remains unchanged. Independent brightness measurements showed that it was intimately related to the SPB concentration, in such a way that any of them can be deduced from the other one. This is of prime importance to keep the bath conditions under control.

Multicomponent determinations by partial least-squares regression analysis of fast-reaction multiwavelength profiles obtained by continuous addition of a reagent

Partial least-squares regression (PLS) analysis of multiwavelength reaction profiles, obtained by continuous addition of a reagent to a binary mixture of analytes, giving fast reactions, is presented. The methodology has proved to be useful for the simultaneous determination of the mixture components. Octacyanomolibdate(V) (Mo(V)) has been used as a reagent for mixtures of hydroquinone (HQ) and pyrogallol (PG) in acidic medium. When the reaction profiles are followed spectrophotometrically at several wavelengths, and they are handled with PLS, the mixture can be resolved. The final spectrum of the mixture, after reaction with Mo(V), is enough to predict the HQ concentration, but in the case of PG successive spectra along the reaction are necessary to predict properly the concentration. The concentrations for HQ in the calibration set ranged between 1.8x10(-6) and 1.7x10(-5) M and for PG between 2.5x10(-6) and 2.5x10(-5) M. The concentration ratio, [HQ]:[PG], in the validation set ranged between 0.17 and 4.10. Mean validation errors of 1.6% for HQ and 4.2% for PG were found. Satisfactory results were obtained when independent studies of accuracy and precision were accomplished. The effect of p-phenylenediamine and aminophenols was studied as interferences and, in general, hydroquinone is more affected than pyrogallol. The tolerated concentration of interferences ranged between 10(-6) and 10(-4) M.

Gas chromatography with flame ionization detection for determination of additives in an electrolytic Zn bath

The monitoring of additives in electrolytic baths is a fundamental task for proper coatings. Among the additives used in zinc baths, benzylideneacetone (BDA), benzoic acid (BA) and polyethylene glycol 400 (PEG400) are easily found. This paper deals with the possibilities of handling the bath sample before it is taken to a gas chromatograph (GC) in order to follow the additives concentration along the bath life. The applied techniques include solid phase extraction (SPE), solvent extraction (SE), static headspace (SHS), direct injection (DI) and headspace solid phase microextraction (HS-SPME) recoveries up to 6:1, 14:1, 9:1, 1:1 and 80:1 respectively have been obtained. Internal standards have been found for every case. Advantages and disadvantages of the techniques are collected. In this paper DI and HS-SPME have finally been applied, though none of them is able for PEG400 determination. DI provides quantitative information for BA (limit of detection, LOD, 1.6 gL(-1)) and BDA (LOD, 0.09 gL(-1)); HS-SPME only provides quantitative information for BDA (LOD, 0.05 gL(-1)). Taking into account that PEG400 and BA do not practically change with the use of the bath, that DI is very quick and simple and that no significant differences with spectrophotometric results have been found, DI is recommended for the monitoring of BA and BDA along the zinc bath life. This should be considered a technique for process analysis for these additives.