Determination Of Multiple Analytes Research Articles

Determination of multiple analytes in urine using label-free SERS coupled with simple sample pretreatments

BackgroundA key restriction of label-free surface-enhanced Raman spectroscopy (SERS) in analysis of objects with complex composition (including with several target analytes) is the competition of mixture components for interaction with SERS-active surface. This leads to poor selectivity of the analysis of such mixtures (e.g., body fluids) and the need to use advanced sample pretreatment procedures such as HPLC or TLC. Therefore, this work aims to develop a set of simple and fast pretreatment steps (dilution, pH correction, etc.) to increase the sorption of the target analyte, reduce the sorption of admixtures, and prevent suppression of the target analyte SERS signal. ResultsWe have developed label-free SERS assay suitable for the determination of three analytes (methotrexate, cephalosporin antibiotic, and creatinine) in one real urine sample as a model matrix with complex and deviating composition. The choice of drugs is justified by the need to monitor their concentration in urine during joint drug treatment of cancer patients with concomitant bacterial infection, while monitoring creatinine concentration helps to evaluate kidney function of the patients. Additionally, three cephalosporin representatives were used in the study to maximize versatility of the assay. As a results, the optimized pretreatment steps enable to eliminate the negative influence of excess of interferences (including other analytes) and achieve precise (≤12% RSD) and accurate (88–111% recovery) determination of several analytes in the therapeutically relevant ranges: 300–3000 μg mL–1 for creatinine, 20–200 μg mL–1 for methotrexate and cephalosporins. SignificanceTherefore, in addition to reporting a new SERS assay for the analysis of body fluids, this study clearly demonstrates the importance of taking into account competitive adsorption processes on the SERS substrate surface. We suggest making this practice mandatory when developing any label-free SERS assay because it enables to maximize the selectivity and accuracy of the analysis as well as to simplify the analysis procedure.

Displacement Assay in a Polythiophene Sensor System Based on Supramacromolecuar Disassembly-Caused Emission Quenching.

Exploring new methodologies for simple and on-demand methods of manipulating the emission and sensing ability of fluorescence sensor devices with solid-state emission molecular systems is important for realizing on-site sensing platforms. In this regard, although conjugated polymers (CPs) are some of the best candidates for preparing molecular sensor devices owing to their luminescent and molecular recognition properties, the development of CP-based sensor devices is still in its early stages. In this study, we herein propose a novel strategy for preparing a chemical stimuli-responsive solid-state emission system based on supramacromolecular assembly-induced emission enhancement (SmAIEE). The system was spontaneously developed by mixing only the component polymers (i.e., polythiophene and a transient cross-linking polymer). The proposed strategy can be applied to the facile preparation of molecular sensor devices. The analyte-induced fluorescent response of polythiophene originated from the dynamic displacement of the transient cross-linker in the polythiophene ensemble and the generation of the polythiophene-analyte complex. Our successful demonstration of the spontaneous preparation of the fluorescence sensor system by mixing two component polymers could lead to the development of on-site molecular analyzers including the determination of multiple analytes.

Design of Experiments (DoE) Application in Two Cases of Study in Pharmaceutical Industries

This study illustrates the practical application of Design of Experiments (DoE) in two real-life scenarios within the pharmaceutical industry. The first case involved optimizing a chromatographic method for the determination of multiple analytes and their degradation products. The primary variable of interest was gradient time, and the most favorable outcomes were achieved at a pH value of 2. In the second case, we conducted a shelf-life study for a veterinary product, revealing that the vial filling variable exerted a statistically significant impact (p-value < 0.05). The incorporation of DoE in both cases played an important role in ensuring the attainment of dependable and statistically validated results.

Construction of a microcavity-based microfluidic chip with simultaneous SERS quantification of dual biomarkers for early diagnosis of Alzheimer's disease

Since there is no effective Alzheimer's disease (AD)-modifying therapy available currently, early analysis of AD core biomarkers has become one of great significance and common concern in clinical diagnosis. Herein, we designed an Au-plasmonic shell attached polystyrene (PS) microsphere in a microfluidic chip for simultaneous detection of Aβ1-42 and p-Tau181 protein. The corresponding Raman reporters were identified in femto gram level by ultrasensitive surface enhanced Raman spectroscopy (SERS). Both of Raman experimental data and finite-difference time-domain modeling demonstrates the synergetic coupling between PS microcavity with the optical confinement property and the localized surface plasmon resonance (LSPR) of AuNPs, so leading to highly amplified electromagnetic fields at the ‘hot spot’. Moreover, the microfluidic system is designed with multiplex testing and control channels in which the AD-related dual proteins were detected quantitatively with a lower limit of 100 fg mL−1. Thus, the proposed microcavity-based SERS strategy initiates a new way for accurately prediction of AD in human blood samples and provides the potential application for synchronous determination of multiple analytes in general disease assays.

Application of the QuEChERS Strategy as a Useful Sample Preparation Tool for the Multiresidue Determination of Pyrrolizidine Alkaloids in Food and Feed Samples: A Critical Overview

The identification of concerning high levels of pyrrolizidine alkaloids (PAs) in a wide variety of food products has raised the occurrence of these natural toxins as one of the main current issues of the food safety field. Consequently, a regulation with maximum concentration levels of these alkaloids has recently been published to monitor their occurrence in several foodstuffs. According to legislation, the analytical methodologies developed for their determination must include multiresidue extractions with high selectivity and sensitivity, as a set of 21 + 14 PAs should be simultaneously monitored. However, the multiresidue extraction of these alkaloids is a difficult task due to the high complexity of food and feed samples. Accordingly, although solid-phase extraction is still the technique most widely used for sample preparation, the QuEChERS method can be a suitable alternative for the simultaneous determination of multiple analytes, providing green extraction and clean-up of samples in a quick and cost-effective way. Hence, this review proposes an overview about the QuEChERS concept and its evolution through different modifications that have broadened its applicability over time, focusing mainly on its application regarding the determination of PAs in food and feed, including the revision of published works within the last 11 years.

An integrated plastic microchip for enhancing electrophoretic separation using tunable pressure-driven backflows.

Microfluidic CE (MCE) is an effective solution for rapid and sensitive determination of multiple analytes. Herein, a dynamic coated cyclic olefin copolymer microchip was developed having an on-chip micropump for fluid velocity adjusting in electrophoretic separations. This micropump was fabricated by constructing a polyacrylamide gel membrane at one channel terminal. Once applying electric field across the membrane, a pressure-driven flow generated automatically to balance the electroosmotic flow (EOF) mismatch at the channel-membrane interface. The influence of gel precursor concentration and operating voltages on the fluid velocity was carefully evaluated. Moreover, the highly integration of injection, separation, and pumping units of the MCE system minimized the dead volume and provides satisfied column efficiency. Experiments showed that by adjusting of pumping voltage reduced the fluid velocity by a factor of 6, resulting six- and threefold resolving power enhancements of rhodamine dye mixture and amino acid mixture, respectively. Furthermore, the developed MCE method was applied for rhodamines and amino acids quantitation in food and cosmetics, with standard addition recoveries of 87.3-106.9% and 89.9-117.4%, respectively. These results were also confirmed by standard HPLC method, revealing the application potential in fast and onsite analysis of complex samples.

Chemiluminescence microarray immunoassay for multiple aminoglycoside antibiotics based on carbon nanotube-assisted signal amplification.

The simultaneous determination of multiple analytes has been an urgent demand in screening of antibiotic residues in food products of animal origin due to its higher analysis efficiency. Five aminoglycoside antibiotics (AGAs) have been monitored in milk, including gentamicin (GEN), kanamycin (KAN), neomycin (NEO), and streptomycin/dihydrostreptomycin (STR/diSTR). A chemiluminescence microarray immunoassay (CLMIA) based on nitrocellulose membrane had been developed for the detection of multiple AGAs, which the LODs for STR, KAN, NEO, and GEN were 4.74ng/mL, 4.97ng/mL, 2.99ng/mL, and 4.42ng/mL respectively. To improve the sensitivity of immunoassay, single-well carbon tubes (SWCNTs) were utilized as solid support for loading horseradish peroxidase-labelled goat anti-mouse antibody to obtain the multi-enzyme particles. After the optimization of usage of multi-enzyme particles and antibodies, the enhanced CLMIA was established and evaluated. The LODs were 1.25ng/mL for STR, 0.64ng/mL for KAN, 0.38ng/mL for GEN, and 0.39ng/mL for NEO, which was improved by threefold, sevenfold, 11-fold, and sevenfold compared with the conventional CLMIA developed. These methods presented higher specificity and repeatability. Finally, the enhanced CLMIA based on CNT-assisted multi-enzyme particles was utilized to analyze twenty-five milk samples from local market and dairy farm, which all the results were below the LOD. The enhanced CLMIA showed the great application potential for the detection of multiple targets simultaneously and provided efficient tool for the screening of pollutants in food.

The novel sensitive and selective chemosensors for determination of multiple analytes

Novel chemosensors that are azo dyes bearing coumarin with indole and thiophene recognition moieties were synthesized. Sensor properties of the dyes were investigated within multiple basic environments that include various pH ranges, anions, and organic amines. Recognition of the basic media by the chemosensors that have acidic NH and NH2 groups, occurs via the deprotonation process which was supported with reversibility study and 1H NMR experiments. The Density Functional Theory calculations have employed to support the experimental results. Chemosensors 6a and 6b showed remarkable detection ability in a pH range of 7–11 and they have been also successfully utilized in the determination of CN− in semi-aqueous media.

Systematic Investigation of Molecular Recognition Ability in FET-Based Chemical Sensors Functionalized with a Mixed Self-Assembled Monolayer System

Exploring new strategies for simple and on-demand methods of manipulating the sensing ability of sensor devices functionalized with artificial receptors embedded in a molecular assembly is important to realizing high-throughput on-site sensing systems based on integrated and miniaturized devices such as field-effect transistors (FETs). Although FET-based chemical sensors can be used for rapid, quantitative, and simultaneous determination of various desired analytes, detectable targets in conventional FET sensors are currently restricted owing to the complicated processes used to prepare sensing materials. In this study, we investigated the relationship between the sensing features of FETs and the nanostructures of mixed self-assembled monolayers (mSAMs) for the detection of biomolecules. The FET devices were systematically functionalized using mixtures of benzenethiol derivatives (4-mercaptobenzoic acid and benzenethiol), which changed the nanostructure of the SAMs formed on gold sensing electrodes. The obtained cross-reactivity in the FETs modified with the mSAMs was derived from the multidimensional variations of the SAM characteristics. Our successful demonstration of continuous control of the molecular recognition ability in the FETs by applying the mSAM system could lead to the development of next-generation versatile analyzers, including chemical sensor arrays for the determination of multiple analytes anytime, anywhere.

ZnO/Ag nanorods as a prominent SERS substrate contributed by synergistic charge transfer effect for simultaneous detection of oral antidiabetic drugs pioglitazone and phenformin

It is popular and promising to explore a prominent surface-enhanced Raman scattering (SERS) substrate for simultaneous determination of multiple analytes in drug safety and quality monitoring. To achieve the goal, in this work, we constructed a plasmonic metal/semiconductor oxide heterostructure by depositing Ag nanoparticles (NPs) on ZnO nanorods. The ZnO/Ag substrate exhibited prominent SERS performance by using rhodamine 6 G as the probe molecule, including transferable nature onto various available substrate, high enhancement factor, stability and uniformity. The synergistic charge transfer effect was systematically studied, including the effects of the Ag loading amount and laser wavelength, photoluminescence, UV–vis spectra and XPS analysis. The ZnO/Ag substrate was developed as a SERS probe for separate and simultaneous detection of two oral antidiabetic drugs (OADs), pioglitazone (PIO) and phenformin (PHE) in human urine. The detection limits were as low as 1 and 5 nM for PIO and PHE (respectively) with good selectivity and satisfactory recoveries. This type of SERS substrate will be suitable for rapid and noninvasive determination of OADs in clinic applications.

Reactive Blue 4 as a Single Colorimetric Chemosensor for Sequential Determination of Multiple Analytes with Different Optical Responses in Aqueous Media: Cu2+-Cysteine Using a Metal Ion Displacement and Cu2+-Arginine Through the Host-Guest Interaction.

In the current study, we reported a novel label-free and facile colorimetric approach for the sequential detection of copper ion (Cu2+), L-arginine (Arg), and L-cysteine (Cys) in the H2O (10.0mmolL-1 HEPES buffer solution, pH7.0) using Reactive Blue 4 (RB4). First, the presence of Cu2+ led to a naked-eye color and spectral changes according to the binding site-signaling subunit approach. Then, the RB4-Cu2+ complex was successfully applied for Cys and Arg through different recognition pathways. The optical signals for Arg were observed due to its association involving the amino group, as well as the participation of the carboxylate group in a bidentate form to the complex, while selective behavior for Cys was explained by a metal displacement mechanism. The limits of detection for Cu2+, Arg, and Cys were calculated to be 1.96, 1.06, and 1.33μmolL-1, respectively. It could also be employed for the determination of three analytes in environmental, biological, and pharmaceutical samples. Importantly, the test strips based on RB4-Cu2+ complex could be used as a solid-state sensor for the detection of Cys and Arg. In addition, NAND and IMPLICATION molecular logic gates were obtained by using chemical inputs and UV-Vis absorbance signal as the output. Graphical Abstract.

Ionic polymer enhanced electrophoresis in plastic microchips for rapid and robust determination of rhodamine dyes

Electrophoresis on plastic microchips is an economic way for rapid separation and determination of multiple analytes, but its application is frequently limited by the unstable electroosmotic flow (EOF) and non-specific adsorption of certain analytes and sample matrices. In this work, cationic polymer poly (diallyldimethylammonium chloride) (PDDA) was used as the background electrolyte (BGE) to eliminate the non-specific adsorption, control the EOF, regulate the viscosity and improve the separation efficiency on cyclic olefin copolymer microchips with 75μm id microchannels. Three rhodamine dyes that are easily adsorbed on plastics were separated in the presence of organic solvents. Influences of the experimental conditions were systematically investigated and the performance was compared with anionic polymer polyacrylic acid or neutral polymer hydroxypropyl cellulose. The effectiveness of ionic polymers for the modulation of EOF was demonstrated. With PDDA as BGE, three rhodamine dyes were well separated and baseline separation could be achieved in 20s with a voltage as low as 400V using a microchannel of 1cm long and 75μm id. The determination of rhodamine dyes in foods and lipsticks was performed to verify the robustness and applicability of the proposed methods. Separation of peptides and proteins labeled by rhodamine B isothiocyanate was also performed with the proposed running medium.

Reagent-free simultaneous determination of glucose and cholesterol in whole blood by FTIR-ATR

Reagent-free determination of multiple analytes is an active and promising field of research in clinical analysis. In this work, the determination of glucose and cholesterol in whole blood using Fourier transform infrared (FTIR) spectroscopy equipped with an attenuated total reflectance (ATR) accessory was performed. A comprehensive sample selection rule in multi space based on SPXY was proposed, termed C-SPXY. The core idea is to make full use of different derivative spectra space to construct the calibration set which preserves the more effective information. On this basis, a partial least squares (PLS) regression fusion modeling method was also presented aiming at improving prediction accuracy of glucose and cholesterol concentration in whole blood samples. Compared with other methods based on single spectra space, the proposed fusion model based on multi spectra space C-SPXY method provides smaller RMSEP values. Experimental results demonstrate that the proposed method and model provides superior predictive power and holds a good application prospect in the field of clinical analysis.

Graphene Oxide-Based Homogeneous Fluorescence Sensor for Multiplex Determination of Various Targets by a Multifunctional Aptamer

A sensitive and selective fluorescent aptasensor for the determination of metal ions, small molecules, sequence-specific DNA, and proteins was fabricated. The analytical approach was based on target-induced conformational changes of the probe and the self-assembled probe-graphene oxide architecture. The probe employed fluorescence “on/off” switching in a single step in solution. This approach was simple to prepare and had little background with good sensitivity and rapid response. In the absence of targets, the probe adsorbed on the surface of graphene oxide through π–π stacking and quenched fluorescence of the probe. Upon the addition of analyte, the random coil of the probe bonded to the targets, reducing the interaction between the probe and graphene oxide, which disrupted the energy transfer from the probe to graphene oxide and therefore increased the fluorescence. This approach was employed for the determination of Ag+, Hg2+, cysteine, sequence-specific DNA, and thrombin. The results demonstrated that the probe-graphene oxide architecture was an excellent and versatile platform for the determination of multiple analytes.

Profiling intact steroid sulfates and unconjugated steroids in biological fluids by liquid chromatography-tandem mass spectrometry (LC-MS-MS)

Within the combined DFG research project "Sulfated Steroids in Reproduction" an analytical method was needed for determining sulfated and unconjugated steroids with highest specificity out of different biological matrices such as aqueous solution, cell lysate and serum. With regard to this analytical challenge, LC-MS-MS presents the technique of choice because it permits (1) analysis of the intact steroid conjugate, (2) allows for simultaneous determination of multiple analytes (profiling, targeted metabolomics approach) and (3) is independent of phenomena such as cross-reactivity. Sample work up consisted of incubation of sample with internal standards (deuterium labeled steroids) followed by solid phase extraction. Only serum samples required a protein precipitation step prior to solid phase extraction. The extract was divided in two parts: six steroid sulfates (E1S, E2S, AS, 16-OH-DHEAS, PREGS, DHEAS) were analyzed by C18aQ-ESI-MS-MS in negative ion mode and eleven unconjugated steroids (E3, 16-OH-DHEA, E1, E2, (4)A, DHEA, T, 17-OH-PREG, Prog, An, PREG) were analyzed by C18-APCI-MS-MS in positive ion mode. For steroid sulfates, we found high sensitivities with LoQ values ranging from 0.08 to 1 ng mL(-1). Unconjugated steroids showed LoQ values between 0.5 and 10 ng mL(-1). Calibration plots showed excellent linearity. Mean intra- and inter-assay CVs were 2.4% for steroid sulfates and 6.4% for unconjugated steroids. Accuracy - determined in a two-level spike experiment - showed mean relative errors of 5.9% for steroid sulfates and 6.1% for unconjugated steroids. In summary, we describe a novel LC-MS-MS procedure capable of profiling six steroid sulfates and eleven unconjugated steroids from various biological matrices.

Application of thermally reduced graphene oxide modified electrode in simultaneous determination of dihydroxybenzene isomers

Thermally reduced graphene oxide (TRGO) was used as an excellent electrocatalyst for the construction of electrochemical sensor for simultaneous determination of dihydroxybenzene isomers including hydroquinone (HQ), catechol (CC) and resorcinol (RC). The surface morphology and structure of TRGO were investigated by utilizing transmission electron microscope, scanning electron microscopy, atomic force microscope, Raman spectroscopy and X-ray diffraction. The electrode modified with TRGO shows excellent electrocatalytic activity toward the oxidation of HQ, CC and RC. As a result, the oxidation peaks of three isomers in pH 6.0 PBS can be clearly discriminated due to large peak potential separation among them. In addition, HQ and CC can be simultaneously determined by using differential pulse voltammetry without any separation step. Under the optimized condition, the detection limits of HQ and CC are 0.75 and 0.8 μM (S/N = 3) with linear ranges of 1–500 μM in the presence of 50 μM isomer, respectively. The proposed method was successfully applied to the simultaneous determination of CC and HQ in synthetic water sample with reliable recovery. The proposed sensor has some important advantages such as low cost, ease of preparation, good stability and high reproducibility. Therefore, the present work promises for the application of graphene modified electrode in the simultaneous determination of multiple analytes.

Development of an Evidence biochip array kit for the multiplex screening of more than 20 anthelmintic drugs

Anthelmintic drugs are used in clinical and veterinary practice for the treatment of infections caused by parasitic worms. Their extensive use in food-producing animals can cause the presence of residues in food. For consumer protection it is necessary to monitor the levels of anthelmintic residues to ensure that they remain within the legally permitted maximum acceptable concentrations. For this purpose, the use of multiplex screening methods is advantageous. Biochip array technology allows the simultaneous determination of multiple analytes from a single sample at a single point in time. This study reports the development of an Evidence biochip array for the multiplex screening of anthelmintic drugs. Simultaneous competitive chemiluminescent immunoassays are employed. The solid support and vessel is the biochip, which contains an array of discrete test sites. The assays were applied to the semiautomated bench-top analyser Evidence Investigator. The aminobenzimidazoles assay detected aminomebendazole, albendazole 2-aminosulphone and aminoflubendazole, the avermectins assay detected emamectin benzoate, eprinomectin, abamectin, ivermectin and doramectin, the benzimidazoles assay detected albendazole sulphone, albendazole, albendazole sulphoxide, oxibendazole, oxfendazole and flubendazole, the thiabendazole assay detected cambendazole, thiabendazole and 5-hydroxythiabendazole and the triclabendazole assay detected ketotriclabendazole, triclabendazole and triclabendazole sulphoxide. The limits of detection ranged from 0.3 ppb (aminobenzimidazoles) to 2.0 ppb (levamisole) in milk and from 0.15 ppb (aminobenzimidazoles) to 6.5 ppb (levamisole) in tissue. The average recovery range was 71-135 %. This multianalytical approach on a biochip platform is applicable to the screening of more than 20 anthelmintic drugs in different food matrices, leading to consolidation of tests and enhancement of the test result output.

Simultaneous Determination of Nitrite and Nitrate Ions by Air-Segmented Amplitude-Modulated Multiplexed Flow Analysis

The concept of amplitude-modulated multiplexed flow analysis has been extended to the simultaneous determination of multiple analytes in a sample. A sample solution containing nitrite and nitrate ions is delivered from two channels, but the flow rates are varied at different frequencies. One of the channels has a reduction column for converting nitrate ions to nitrite ions. Downstream, the absorbance of the diazo-coupling product is monitored after the merging of both solutions with a Griess reagent. The signal is analyzed by a fast Fourier transform (FFT) in real time. From the thus-obtained amplitude, a µmol dm(-3) level of the ions can be determined. The introduction of air bubbles is effective to reduce any axial dispersion, and hence to improve the sensitivity.

Amplitude modulated multiplexed flow analysis

We propose a new concept, amplitude modulated multiplexed flow analysis, for the simultaneous determination of multiple analytes from a single continuous analytical signal. The flow rates ( F S1 and F S2) of two sample solutions are independently varied in response to trigonometric voltage signals each at a different anharmonic frequency. With the total flow rate F T held constant, the solutions are merged with a reagent solution that is aspirated to the confluence point at the flow rate of F T − F S1 − F S2. Downstream, the analytical signal due to the mixed reacted solution, V d, is monitored. V d will show a complicated profile resulting from the trigonometrically varying flow rates of the sample solutions. The signal contains information on the multiple analytes. The signal can be demodulated to the contribution of each sample solution through fast Fourier transform (FFT). The amplitudes of the separated wave components are related to the concentration of the analytes in the sample solutions. By moving a window for FFT analysis with time, temporal profile of amplitudes can be obtained in real-time. We demonstrate the concept by applying it to the simultaneous determination of food dyes and to that of ferrous ions in two sample solutions.

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.