Validation of an analytical method to determine sulfamides in kidney by HPLC-DAD and PARAFAC2 with first-order derivative chromatograms

Six sulfamides were extracted from kidney and analysed by high-performance liquid chromatography with diode array detection (HPLC-DAD): sulfadiazine, sulfamethazine, sulfamethoxypyridazine, sulfamethoxazole, sulfadimethoxine and sulfaquinoxaline.Two main difficulties arose in identifying and quantifying the analytes. Firstly, the chromatographic peaks of the matrix interferences overlapped with those of the analytes. The uniqueness property of PARAFAC2 solved this problem. Secondly, the gradient elution caused a baseline drift. The first-order derivative of the chromatograms minimized its effect.The analytical method was validated. As the performance criteria detailed in the European Decision 2002/657/EC are based on specific signals, this paper generalizes those criteria for higher-order and non-specific signals. In this sense the proposed methodology is general and can be applied to any chromatographic method (HPLC or GC) with a detector that provide a multivariate signal (MS, DAD, EC, etc.).     

Determination of phenolic compounds and authentication of PDO Lambrusco wines by HPLC-DAD and chemometric techniques

This work proposes a fast and simple method for detection and quantification of phenolic compounds in PDO Lambrusco wines using HPLC-DAD and chemometric techniques. Samples belonging to three different varieties of Lambrusco (Grasparossa, Salamino and Sorbara) were analyzed to provide a methodology appropriate for routine analysis. Given the high complexity of the sample and the coelution among chromatographic peaks, the use of chemometric techniques to extract the information of the individual eluting compounds was needed. Multivariate curve resolution-alternating least squares (MCR-ALS) allowed the resolution of the chromatographic peaks obtained and the use of this information for the quantification of the phenolic analytes in the presence of interferences. Use of multiset analysis and local rank/selectivity information was proven to be crucial for the correct resolution and quantification of compounds. The quantitative data provided by MCR-ALS about the phenolic targets and additional compounds present in the samples analyzed provided wine composition profiles, which were afterwards used to distinguish among wine varieties. Principal component analysis applied to the wine profiles allowed characterizing the wines according to their varieties.     

Signal suppression/enhancement in high-performance liquid chromatography tandem mass spectrometry

The review discusses the pitfalls of the matrix effect in mass spectrometry detection hyphenated to liquid chromatography separation. Matrix effect heavily influences both qualitative and quantitative analyses, giving rise to suppression or enhancement of the signal. As generally recognised, the predominant cause is the presence of undesired components that co-elute in the chromatographic separation and alter the ionisation process. The interfering species can be components of the sample, compounds released during the pre-treatment/extraction process or reagents added to the mobile phase to improve chromatographic resolution. The different mechanisms proposed in literature to explain the suppression or the enhancement of the signal both in electrospray and atmospheric pressure chemical ionisations are presented and the results observed in the different experimental conditions are compared and discussed. All data together lead to conclude that the chemical properties of the target analyte, the kind of matrix, the matrix to analyte concentration ratio, the extraction process, the chromatographic conditions as well as the kind of the mass spectrometry instrumentation and the ionisation conditions can play a role. Likely all these potential causes act in a synergic way and the final effect observed is hardly due to only one of them. Depending on an unpredictable combination of conditions, signal suppression or enhancement can be observed. The review discusses the matrix effects observed in HPLC–MS and HPLC–MS/MS analysis proposes hypotheses to explain the observed behaviours and proposes methods and strategies to overcome the matrix effects.     

直观推导式演进特征投影法分辨枸杞类胡萝卜素的异构体

采用高效液相色谱-二极管阵列检测方法,初步分离了枸杞子中的类胡萝卜素。以乙腈-二氯甲烷(体积比为60∶40)为流动相,流速为1.0 mL/min,用C18柱从枸杞类胡萝卜素中分离出7个峰。但由于类胡萝卜素的结构多样性,尤其是顺反异构体的出现使得分离并不完全。利用所得三维数据提供的信息和化学计量学方法直观推导式演进特征投影法(HELP),对其中某些在二维色谱中被定性为单组分的峰进行解析,结果发现原来在二维色谱中被定性为单组分的峰大多是一些多组分峰。以其中第4个峰为例对其进行解析,得到了该类胡萝卜素的同分异构体色谱流出曲线及紫外光谱信息。该方法表明,化学计量学方法与现代分析手段有机结合,大大降低了此类复杂体系对色谱分离的要求,对同分异构体的分析具有一定的借鉴和指导意义。     

Spectral correlation of high-performance liquid chromatography-diode array detection data from two independent chromatographic runs peak tracking in pharmaceutical impurity profiling

A novel qualitative analytical method for peak tracking in impurity profiling control by the correlation of spectra was established. Two-dimensional (2D) standard spectrochromatographic data produced by high-performance liquid chromatography with diode array detection (HPLC-DAD) were compared with sample data to develop two-dimensional chromatographic spectral correlative maps. Taking full advantage of separation efficiency of HPLC and spectral specificity of the analytes, the method was successfully used to recognize impurities in quinolone antibacterials, when in combination with relative retention times (RRTs). For the comparison of spectra was expanded to three-dimensional space, simultaneous identification of the chromatographic peaks can be obtained rapidly without preparation and injection of a reference solution, even when the mobile phase changed or the peaks of multi-component samples overlapped.     

直观推导式演进特征投影法测定不同方法提取的国产血竭中龙血素B的含量

利用HPLC-DAD产生的二维数据和化学计量学方法-直观推导式演进特征投影法（HELP），解析了不同提取方法得到的国产血竭中的重叠色谱峰，并对其中的指标性成分-龙血素B进行了含量测定，结果满意。这表明这化学计量学方法与现代分析手段有机结合，将为中药等复杂体系的分析提供一条新途径。     

Data Preprocessing for Chromatographic Fingerprint of Herbal Medicine with Chemometric Approaches

Abstract

Recently, the fingerprinting approach using chromatography has become one of the most potent tools for quality assessment of herbal medicine. Due to the complexity of the chromatographic fingerprint and the irreproducibility of chromatographic instruments and experimental conditions, several chemometric approaches such as variance analysis, peak alignment, correlation analysis, and pattern recognition were employed to deal with the chromatographic fingerprint in this work. To facilitate the data preprocessing, a software named Computer Aided Similarity Evaluation (CASE) was also developed. All programs of chemometric algorithms for CASE were coded in MATLAB5.3 based on Windows. Data loading, removing, cutting, smoothing, compressing, background and retention time shift correction, normalization, peak identification and matching, variation determination of common peaks/regions, similarity comparison, sample classification, and other data processes associated with the chromatographic fingerprint were investigated in this software. The case study of high pressure liquid chromatographic HPLC fingerprints of 50 Rhizoma chuanxiong samples from different sources demonstrated that the chemometric approaches investigated in this work were reliable and user friendly for data preprocessing of chromatographic fingerprints of herbal medicines for quality assessment.     

Correction of retention time shifts for chromatographic fingerprints of herbal medicines

In this study, the combination of chemometric resolution and cubic spline data interpolation was investigated as a method to correct the retention time shifts for chromatographic fingerprints of herbal medicines obtained by high-performance liquid chromatography-diode array detection (HPLC-DAD). With the help of the resolution approaches in chemometrics, it was easy to identify the purity of chromatographic peak clusters and then resolve the two-dimensional response matrix into chromatograms and spectra of pure chemical components so as to select multiple mark compounds involved in chromatographic fingerprints. With these mark components determined, the retention time shifts of chromatographic fingerprints might be then corrected effectively. After this correction, the cubic spline interpolation technique was then used to reconstruct new chromatographic fingerprints. The results in this work showed that, the purity identification of the chromatographic peak clusters together with the resolution of overlapping peaks into pure chromatograms and spectra by means of chemometric approaches could provide the sufficient chromatographic and spectral information for selecting multiple mark compounds to correct the retention time shifts. The cubic spline data interpolation technique was user-friendly to the reconstruction of new chromatographic fingerprints with correction. The successful application to the simulated and real chromatographic fingerprints of two Cortex cinnamomi, fifty Rhizoma chuanxiong, ten Radix angelicae and seventeen Herba menthae samples from different sources demonstrated the reliability and applicability of the approach investigated in this work. Pattern recognition based on principal component analysis for identifying inhomogenity in chromatographic fingerprints from real herbal medicines could further interpret it.     

An absorbance-based micro-fluidic sensor for diffusion coefficient and molar mass determinations

The H-Sensor reported herein is a micro-fluidic device compatible with flow injection analysis (FIA) and high performance liquid chromatography (HPLC). The device detects analytes at two separate off-chip absorbance flow cells, providing two simultaneous absorbance measurements. The ratio of these two absorbance signals contains analyte diffusion coefficient information. A theoretical model for the sensing mechanism is presented. The model relates the signal Ratio to analyte diffusion coefficient. The model is qualitatively evaluated by comparing theoretical and experimental signal Ratio values. Experimental signal Ratios were collected via FIA for a variety of analytes, including sodium azide, benzoic acid, amino acids, peptides, and proteins. Measuring absorbance at multiple wavelengths provides higher order data allowing the analyte signals from mixtures to be deconvolved via classical least squares (CLS). As a result of the H-Sensor providing two simultaneous signals as a function of time for each sample injection, two simulated second-order HPLC chromatograms were generated using experimental H-Sensor data. The chemometric deconvolution method referred to as the generalized rank annihilation method (GRAM) was used to demonstrate chromatographic and spectroscopic deconvolution. GRAM also provides the signal Ratio value, therefore simultaneously obtaining the analyte diffusion coefficient information during deconvolution. The two chromatograms successfully serve as the standard and unknown for the GRAM deconvolution. GRAM was evaluated on chromatograms at various chromatographic resolutions. GRAM was found to function to a chromatographic resolution at and above 0.25 with a percent quantitative error of less then 10%.     

Simultaneous correction for drift and non-spectroscopic matrix effect in the measurement of geological samples by inductively coupled plasma-mass spectrometry using common analyte internal standardization chemometric technique

The common analyte internal standardization (CAIS) chemometric technique is extended to correct for drift in signal intensity in inductively coupled plasma-mass spectrometry. The CAIS technique is used because, unlike the conventional internal reference method, it allows for the analyte to behave differently from the internal reference under the influence of drift. Thus, only one internal reference element is sufficient to correct for drift effect for all the analytes irrespective of the difference of their mass and ionization potential from that of the internal reference element. Experimental test with 15 analytes, as a representative of the whole mass range, in different geological matrices, using only indium as internal reference element, demonstrates that the developed drift correction method is efficient in correcting up to 25% drift error and is easy to use. Further, a CAIS scheme was developed to correct for both drift and non-spectroscopic matrix effects simultaneously in a single run, using standard containing no matrix. This scheme is based on the use of two internal reference elements for simultaneous monitoring and correction of drift and matrix effects for all the analytes irrespective of their difference in mass and ionization potential from the two internal reference elements. The developed scheme was validated using river water samples containing appreciable matrix concentration and was measured under the influence of drift. The results of the experimental validation indicate that the proposed scheme is capable of removing error as large as 43% that arises from drift and non-spectroscopic matrix effects.     

Alternative calibration approaches for LC-MS quantitative determination of coeluted compounds in complex environmental mixtures using multivariate curve resolution

Different calibration approaches including external calibration, standard addition and internal standard are evaluated for quantification of coeluted compounds in liquid chromatography with MS spectrometry detection in scan mode and using multivariate curve resolution. These different calibration approaches are proposed to cope with sensitivity changes and matrix effects encountered in the analysis of complex natural environmental samples. By using them, multivariate curve resolution analysis of MS data in scan mode gave similar quantitative results to those obtained by LC-MS in selected ion monitoring (SIM) mode (in both cases errors were below 16% for internal standard combined with standard addition strategy), and it provided at the same time a means of analyte confirmation via their resolved pure MS spectra, and a means to gather a larger amount of information about the whole chromatographic process and to facilitate the simultaneous determination of multiple analytes in the same chromatographic run using the same experimental and instrumental conditions.     

Gas-chromatographic retention indices in dependence on the ratio of analytes to reference components

One of the main factors determining the interlaboratory reproducibility of gas-chromatographic retention indices is their linear dependence on the logarithms of the ratios of the amounts of analytes and reference components, which is rarely taken into account. It is found that the effect of this factor is much stronger for packed columns in comparison to that for more efficient capillary columns; it decreases with the sample dilution and with the increase in the temperature of gas-chromatographic separation. It was shown that the effect of relative amounts of components on their retention indices agrees with the main principles of the chromatographic plate theory and is associated with neither the overloading of chromatographic columns nor the nonlinearity of the adsorption isotherms. To improve the reproducibility of indices, it is recommended to use retention parameters measured at certain points in the front edges of the chromatographic peaks corresponding to similar intensities of the analytical signal rather than at the peak maxima.     

三维(二阶)算法在液相色谱分析中的应用

使用"垂线法"、"切线法"或"三角形法"等传统方法对液相色谱重叠峰分辨时经常会遇到误差过大的情况,而使用三维(二阶)算法对重叠和拖尾峰分辨可以最大限度地降低这种因几何分割而人为产生的误差。这样改进的色谱解析方法具有自动化程度高、抗干扰能力强、对重叠/拖尾峰定量准确等优点,甚至可以减少样品前处理和色谱条件优化。该方法的核心是基于化学计量学三维(二阶)算法抽取有效信息和建模的思想,三维色谱数据按照对三线性模型的符合程度有"三线性数据"和"非三线性数据"的区别,相应地将三维(二阶)算法分为"三线性算法"和"非三线性算法"。本文综述了近10年来国内外三维(二阶)算法在复杂体系液相色谱分析中的应用进展,侧重于样品前处理、辅助算法、校正算法间的联用和对比等问题。     

New temperature-assisted ionic liquid-based dispersive liquid–liquid microextraction method for the determination of glyphosate and aminomethylphosphonic acid in water samples

A new analytical temperature-assisted ionic liquid-based dispersive liquid–liquid microextraction (TA-IL-DLLME) method was developed for glyphosate and aminomethylphosphonic acid determination in water samples. Extracted analytes were derivatized using 9-fluoroenylmethylchloroformate and quantified by liquid chromatography with fluorescence detection. For the TA-IL-DLLME method, two strategies for phase solubilization were evaluated; in approach 1, the ionic liquid and aqueous matrix sample were mixed and then heated, while in approach 2, the aqueous sample was first heated and then the ionic liquid was injected. For both approaches, optimization included parameters that significantly affect extraction efficiency: ionic liquid type and volume, solubilization temperature and time, cooling and centrifugation time. Among the evaluated ionic liquids, 1-decyl-3-methylimidazolium tetrafluoroborate showed the best performance for TA-IL-DLLME and was selected for the two solubilization approaches; with approach 2, slightly better results were obtained. Thus, sample analyses were performed using a procedure based on approach 2. An important matrix effect, attributed to the presence of salts and metals in real water samples was observed. Sample acidification before derivatization allowed this problem to diminish, with recoveries ranging from 75 and 99%, and enrichment factors between 57 and 76 for target analytes.     

A review of multivariate calibration methods applied to biomedical analysis

The determination of the contents of therapeutic drugs, metabolites and other important biomedical analytes in biological samples is usually performed by using high-performance liquid chromatography (HPLC). Modern multivariate calibration methods constitute an attractive alternative, even when they are applied to intrinsically unselective spectroscopic or electrochemical signals. First-order (i.e., vectorized) data are conveniently analyzed with classical chemometric tools such as partial least-squares (PLS). Certain analytical problems require more sophisticated models, such as artificial neural networks (ANNs), which are especially able to cope with non-linearities in the data structure. Finally, models based on the acquisition and processing of second- or higher-order data (i.e., matrices or higher dimensional data arrays) present the phenomenon known as “second-order advantage”, which permits quantitation of calibrated analytes in the presence of interferents. The latter models show immense potentialities in the field of biomedical analysis. Pertinent literature examples are reviewed.     

Cross-sections of spectrochromatograms for the resolution of overlapping peaks in diode-array high-performance liquid-chromatography

Multi-wavelength detectors offer improved detection capabilities for liquid chromatographic methods, but require multivariate approaches to utilise all the available information. The photodiode-array detector in high-performance liquid chromatography (HPLC) generates a three-dimensional data matrix, which is conventionally presented as an isometric projection or a contour plot. In this work, a new graphical technique is described for improving the quantitative results obtained from HPLC, using the available spectrochromatographic information in both the time and wavelength domains. The technique consists of performing cross-sections through the data matrix to obtain the maximum analytical information for each of the analytes. Hence, the resolution of overlapping peaks and the sensitivity in the determination are optimised. In order to demonstrate the validity and simplicity of the approach, the method has been applied to the resolution of synthetic mixtures of iprodione, procymidone and chlorothalonil. Also, the method has been satisfactorily applied to the simultaneous determination of the pesticides in environmental groundwater samples.     

Turbulent flow chromatography in bioanalysis: a review

With advances in fast chromatography techniques, and highly sensitive and selective detection methods such as tandem mass spectrometry, very high-throughput bioanalytical methods can now be easily developed. The bottleneck of the analytical process then becomes the sample preparation, which it is now realized is crucial to the robust operation of the analytical system, especially for quantitative assays. Turbulent flow liquid chromatography was developed in the late 1990s, and combines 'size exclusion' and traditional stationary phase column chemistry to separate macromolecules, such as proteins, from smaller molecules and analytes of interest in biological fluids. By definition, the process is very rapid, and the instrumentation and software have been developed for fully automated, on-line extraction of neat biological fluids. This work aims to review the chromatographic theory of turbulent flow chromatography and illustrate, using examples from recent literature, the application of this technique to a range of analytes from a number of different biological matrices.     

Établissement du profil chromatographique liquide non aqueux des métabolites phytochimiques apolaires des phytomédicaments

Chromatographic profiling of plant metabolites is therefore a good tool for quality control of such herbal medicinal products. Our objective was to propose a protocol for sample preparation and liquid chromatographic profiling of non-polar metabolites for quality assessment of African herbal medicinal products. The methodology is based on the chemometric assessment of liquid chromatographic profiles of non-polar metabolites issued from several batches of leaves of Combretum micranthum and Mitracarpus scaber. Metabolic profiling is carried out by non-aqueous liquid chromatography on porous carbon graphite, coupled with mass spectrometry, after extraction with dichloromethane and removal of chlorophyll. Our method using liquid chromatography, coupled to mass spectrometry can detect non-polar metabolites already identified in the two herbal drugs. Chemometric data analysis of chromatographic profiles using the PLS-discriminant analysis with or without orthogonal signal correction, allowed a distinction between the two herbal drugs.     

Simultaneous determination of potassium guaiacolsulfonate, guaifenesin, diphenhydramine HCl and carbetapentane citrate in syrups by using HPLC-DAD coupled with partial least squares multivariate calibration

A simple and rapid analytical procedure was proposed for the determination of chromatographic peaks by means of partial least squares multivariate calibration (PLS) of high-performance liquid chromatography with diode array detection (HPLC-DAD). The method is exemplified with analysis of quaternary mixtures of potassium guaiacolsulfonate (PG), guaifenesin (GU), diphenhydramine HCI (DP) and carbetapentane citrate (CP) in syrup preparations. In this method, the area does not need to be directly measured and predictions are more accurate. Though the chromatographic and spectral peaks of the analytes were heavily overlapped and interferents coeluted with the compounds studied, good recoveries of analytes could be obtained with HPLC-DAD coupled with PLS calibration. This method was tested by analyzing the synthetic mixture of PG, GU, DP and CP. As a comparison method, a classsical HPLC method was used. The proposed methods were applied to syrups samples containing four drugs and the obtained results were statistically compared with each other. Finally, the main advantage of HPLC-PLS method over the classical HPLC method tried to emphasized as the using of simple mobile phase, shorter analysis time and no use of internal standard and gradient elution.     

Outlier detection for the Generalized Rank Annihilation Method in HPLC-DAD analysis

The Generalized Rank Annihilation Method (GRAM) is a second-order calibration method that is used in chromatography to quantify analytes that coelute with interferences. For a correct quantification, the peak of the analyte in the standard and in the test sample must be aligned and have the same shape (i.e., have a trilinear structure). Variations in retention time and shape between the two peaks may cause the test sample to behave as an outlier and produce an incorrect prediction. This situation cannot be detected by checking the coincidence of the recovered spectrum with the known spectrum of the analyte because the spectral domain is not affected. It cannot be detected either by checking if the recovered profile is correct (i.e., unimodal and positive). Several plots are presented to detect such outliers. The first plot compares the particular elution profiles in the standard and in the test sample that are recovered by least-squares regression from the spectra estimated by GRAM. The calculated elution profiles from both peaks should coincide. A second plot uses the elution profiles and spectra calculated by GRAM to define the vector space spanned by the interferences. The measured peaks in the standard and in the test sample are projected onto the space that is orthogonal to the space spanned by the interferences. These projections are proportional (up to the noise) if data are trilinear. The proportionality is checked graphically from the first singular vector of the projected peaks, or from the plot of the orthogonal signal versus the net sensitivity. The use of these graphs is shown for simulated data and for the determination of 4-nitrophenol in river water samples with liquid chromatography/UV-Vis detection.