An Indirect Competitive Enzyme-linked Immunosorbent Assay for Simultaneous Determination of Florfenicol and Thiamphenicol in Animal Meat and Urine

A high-affinity polyclonal antibody was prepared by immunizing animals with haptens FFD and FFM. Under the optimal combination of coating antigen and antibody, an indirect competitive enzyme-linked immunosorbent assay (icELISA) for simultaneous detection of florfenicol and thiamphenicol residues in animal meat and urine samples was developed. The icELISA showed an IC₅₀ value of 1.32 ng mL^?1 for florfenicol and 2.13 ng mL^?1 for thiamphenicol, respectively. The linear ranges were from 0.31 to 5.61 ng mL^?1 with a limit of detection of 0.12 ng mL^?1 for florfenicol, and 0.41 to 11.2 ng mL^?1 with a limit of detection of 0.15 ng mL^?1 for thiamphenicol, respectively. The average recoveries of florfenicol and thiamphenicol in spiked samples ranged from 77.2% to 116.0% with a relative standard deviation of less than 15%. Therefore, this proposed icELISA provided a valid detection method for florfenicol and thiamphenicol residues in animal tissue and urine samples.     

Postcolumn Iodine–Azide Reaction as a Detection System in HPLC for the Determination of Methylthiouracil in Urine

Abstract

This article describes a system for determining methylthiouracil in urine based on the sensitizing induction of methylthiouracil on the reaction between iodine and azide ions and combined techniques of high-performance liquid chromatography and simple postcolumn detection. The optimal conditions for iodine–azide reaction and high-performance liquid chromatographic separation were determined. The reproducibility, linearity, and recovery were evaluated under the optimal conditions. The methylthiouracil standards added to normal urine show that the response of the detector, set at 350 nm (corresponding to unreacted iodine in the postcolumn iodine–azide reaction), is linear within the concentration range of 0.4–5.0 nmol mL^?1 urine. The relative standard deviation values for precision and recovery within the calibration range varied from 0.9 to 4.8% and from 94 to 105%, respectively. Limits of detection (LOD) and quantitation (LOQ) were 0.3 and 0.4 nmol mL^?1 urine, respectively.     

Comparative Evaluation of CE and HPLC for Determination of Cotinine in Human Urine

Two rapid and popular methods—capillary electrophoresis (CE) and high-performance liquid chromatography (HPLC) have been compared for analysis of cotinine in human urine. Cotinine was analyzed in less than 7 min, with detection limits of 5 and 3.2 ng mL⁻¹ for CE and HPLC, respectively. The performance of the methods was evaluated in terms of sensitivity, specificity, precision, accuracy, and limits of detection and quantification. Calibration plots were linear in the range 50–4,000 ng mL⁻¹, at least, and mean recoveries were satisfactory for both techniques. The methods were successfully used for quantification of cotinine in urine.     

Development of colloidal gold-based immumochromatographic assay for the rapid detection of medroxyprogesterone acetate residues in biological materials

A competitive colloidal gold-based immunoassay in lateral-flow format for the rapid detection of medroxyprogesterone acetate (MPA) in biological materials was developed. A nitro-cellulose membrane strip was separately coated with goat anti-rabbit IgG (control line) and MPA hapten-OVA conjugate (test line). Anti-MPA polyclonal antibody labelled with colloidal gold particles was first incubated with MPA. The limit of detection for lateral flow was 5?ng?g^?1 for detecting an MPA standard solution, and the limit of detection was 10?ng?mL^?1 for detecting the MPA spiked in pig urine and 10?ng?g^?1 for spiked in pig liver. The results were confirmed by high-performance liquid chromatography tandem mass spectrometry (HPLC/MS/MS) and indicated that there was a good agreement between both methods (R ²?=?0.976). The assay time for the test was less than 5?min, suitable for rapid testing on site.     

Multi‐residue analysis of eight thioamphetamine designer drugs in human urine by liquid chromatography/tandem mass spectrometry

An analytical procedure for the simultaneous determination in human urine of several thioamphetamine designer drugs (2C‐T and ALEPH series) is reported. The quantitative analysis was performed by liquid chromatography/tandem mass spectrometry and has been fully validated. The mass spectrometer was operated in positive‐ion, selected reaction monitoring (SRM) mode. In order to minimize interferences with matrix components and to preconcentrate target analytes, solid‐phase extraction was introduced in the method as a clean‐up step. The entire method was validated for selectivity, linearity, precision and accuracy. The method turned out to be specific, sensitive, and reliable for the analysis of amphetamine derivatives in urine samples. The calibration curves were linear over the concentration range of 1 to 100 ng mL^?1 for all drugs with correlation coefficients that exceeded 0.996. The lower limits of detection (LODs) and quantification (LOQs) ranged from 1.2 to 4.9 ng mL^?1 and from 3.2 to 9.6 ng mL^?1, respectively. Copyright © 2009 John Wiley & Sons, Ltd.     

Simultaneous chiral separation and determination of carvedilol and 5′-hydroxyphenyl carvedilol enantiomers from human urine by high performance liquid chromatography coupled with fluorescent detection

A sensitive and specific high performance liquid chromatography coupled with fluorescent detection (HPLC-FL) and tandem mass spectrometry detection (HPLC-MS/MS) methods for separation and determination of carvedilol (CAR) enantiomers and 5′-hydroxyphenyl carvedilol (5′-HCAR) enantiomers has been developed and validated. The analysed compounds were extracted from human urine by solid phase extraction. Good enantioseparation of the studied enantiomers was achieved on CHIRALCEL® OD-RH column using 0.05% trifluoroacetic acid and 0.05% diethylamine in water and acetonitrile in a gradient elution. The mass spectrometric data were acquired using the multiple reaction monitoring mode by positive electrospray ionisation. The method was validated over the concentration range from 25.0 ng mL^?1 to 200 ng mL^?1 for the analysed compounds. The limit of quantification varied from 14.2 ng mL^?1 to 24.2 ng mL^?1. Both the repeatability and inter-day precisions were below 10.0%, and the accuracy varied from ?13.2% to 3.77%. The extraction recoveries ranged from 79.2% to 108%. The present paper reports the method for the simultaneous determination of CAR enantiomers and their metabolite enantiomers (5′-HCAR) in human urine samples. This newly developed method was successfully used to analyse the aforementioned analytes in human urine samples obtained from patients suffering from cardiovascular disease.      

Development of Immunoaffinity Sample-Purification for GC–MS Analysis of Ractopamine in Swine Tissue

A method combining immunoaffinity chromatography with gas chromatography–mass spectrometry (GC–MS) has been established for determination of ractopamine residues in swine liver and urine. After clean-up on an immunoaffinity chromatography column, GC–MS analysis revealed recovery from blank swine liver and urine fortified at 2.5–20 ng g^?1 (ng mL^?1 for urine), respectively, was 68.2–78.6 and 76.2–83.1%. The limits of detection and quantification were 0.5 ng g^?1 (or ng mL^?1) and 2.0 ng g^?1 (or ng mL^?1), respectively. The procedure was used for analysis of ractopamine residues in samples of swine liver and urine in which the levels were unknown. The amounts detected were 9–216 ng g^?1 (ng mL^?1).     

Optimisation of an in-tube solid phase microextraction method coupled with HPLC for determination of some oestrogens in environmental liquid samples using different capillary columns

A simple on-line method for simultaneous determination of some oestrogens including oestriol (E3), norethisterone (NORE), ethynylestradiol (EE2), D-norgestrel (NORG) and bisphenol A (BPA), in environmental liquid samples was developed by coupling in-tube solid phase microextraction (in-tube SPME) to high-performance liquid chromatography with diode array (DAD) and fluorescence (FLD) detectors. Two capillary chromatographic columns (Supel-Q? and Carboxen? 1006 porous layer open tubular) were selected to develop this method. To achieve optimum extraction performance, several parameters were investigated including number of draw/eject cycles and the sample volume for each of the columns. Reproducibility was satisfactory for inter- and intra-day precision, yielding % RSDs of less than 10% and 7.6%, respectively. Limits of detection (LODs) and quantification (LOQs) for the proposed method using a DAD detector were achieved in the ranges of 0.04–0.63?ng?mL^?1 and 0.12–1.9?ng?mL^?1, depending of the capillary column used. Fluorescence detection improved these parameters for E3, BPA and EE2, obtaining LODs of 0.005–0.03?ng?mL^?1 and LOQs of 0.015–0.08?ng?mL^?1 using Supel-Q and LODs of 0.01–0.015?ng?mL^?1 and LOQs of 0.025–0.04?ng?mL^?1 using Carboxen. The proposed method was successfully applied to spiked environmental waters obtaining recoveries greater than 80%.     

Electrochemical Enzyme‐Linked Immunoassay for the Determination of Estriol Using Methyl Red as Substrate

Abstract

A new electrochemical substrate for horseradish peroxidase, methyl red, is reported. In this reaction system, horseradish peroxidase can catalyze the redox reaction of methyl red and H₂O₂. Methyl red exhibits a sensitive voltammetric peak at?0.51 V vs. Ag/AgCl reference electrode, the decrease of the peak current of methyl red is in proportion to the concentration of horseradish peroxidase (HRP). The linear range for determination of horseradish peroxidase is 5.0×10^?8～5.0×10^?7 g mL^?1 and the detection limit is 1.8×10^?8 g mL^?1. The relative standard deviation is 3.3% when 2.0×10^?7 g mL^?1 HRP was sequentially determined 11 times. A voltammetric enzyme‐linked immunoassay method for the determination of estriol was developed, based on this electrochemical system. The linear range for determination of estriol is 1.0～1000.0 ng mL^?1, and the detection limit is 0.33 ng mL^?1. The relative standard deviation for 11 parallel determinations with 200 ng mL^?1 estriol is 4.8%. Some pregnancy serum samples were analyzed with satisfactory results.     

Selective extraction of catecholamines by packed fiber solid‐phase using composite nanofibers composing of polymeric crown ether with polystyrene

For the first time, electrospun composite nanofibers comprising polymeric crown ether with polystyrene (PCE‐PS) have been used for the selective extraction of catecholamines – dopamine (DA), norepinephrine (NE) and epinephrine (E) – prior to their analysis by high‐performance liquid chromatography–electrochemical detection. Using a minicartridge packed with PCE‐PS composite nanofibers, the target compounds were extracted effectively from urine samples to which diphenylborinic acid 2‐aminoethyl ester was added as a complexing reagent. The extracted catecholamines could be liberated from the fiber by the addition of acetic acid. A good linearity was observed for catecholamines in the range of 2.0–200 ng mL^?1 (NE, E and DA). The detection limits of catecholamines (signal‐to‐noise ratio = 3) were 0.5 ng mL^?1 (NE), 0.2 ng mL^?1 (E) and 0.2 ng mL^?1 (DA), respectively. Under the optimized conditions, the absolute recoveries of the above three catecholamines were 90.6% (NE), 88.5% (E) and 94.5% (DA). The repeatability of extraction performance was from 5.4 to 9.2% (expressed as relative standard deviation). Our results indicate that the proposed method could be used for the determination of NE, E and DA in urine. Copyright © 2014 John Wiley & Sons, Ltd.     

Flow Injection Spectrophotometric Determination of the Antibacterial Levofloxacin in Tablets and Human Urine

Abstract

A sensitive and fast flow‐injection (FI) spectrophotometric method for the determination of levofloxacin based on the formation of a colored product upon oxidation with N‐bromosuccinimide (NBS) in acidic medium is proposed. Optimization of chemical and FI variables has been made. Under the optimized conditions, the sampling rate was over 90 h^?1, the calibration curve obtained was linear over the range 10–300 µg·mL^?1, and the detection limit was 3 µg·mL^?1. The proposed method was successfully applied to the determination of levofloxacin in pharmaceuticals and human urine samples. It was also found that the excipients in the commercial tablet preparation did not interfere with the assay. Results are precise (RSD<2.7%; n =10) and in agreement with those found by the reference high pressure liquid chromatography (HPLC) procedure.     

Chromatographic separation of thulium from erbium for neutron capture cross section measurements—Part I: Trace scale optimization of ion chromatography method with various complexing agents

The impact of various ion chromatography parameters on the separation of trace amounts of thulium from erbium was examined to address the need for the preparation of a ¹⁷¹Tm target for neutron capture cross section measurements. The following optimal operation parameters for analytical scale separations with cation exchange resin were established based on a modified separation resolution: 0.046 M α-HIB^? as eluent with a flow rate of 1.2 mL min^?1 at 25 °C. Different carboxylic acids with varying pH were also investigated, which reaffirmed the use of α-hydroxyisobutyrate as the most suitable complexant for the separation of these neighboring lanthanides.     

Efficient HPLC method for determination of cephalosporin residues on spiked stainless-steel plates and human plasma: application of a worst-case product for Cosa®CIP

ABSTRACT

The main objective of the cleaning validation procedure is to verify the effectiveness of the cleaning procedure for removal and minimising the risk of cross-contamination, a topic that has become more important regarding the development of the medicines. Furthermore, if a product is found to be the worst among many of the products, one cleaning validation procedure of the worst-case product can cover the validation of the remaining ones, thus saving time and money. A novel, reproducible and efficient high-performance liquid chromatography (HPLC) method was optimised and validated for the detection of the following cephalosporin residues: cephalexin (CPH), cefaclor (CFC), cefixime (CFX), cefdinir (CFR) and ceftazidime (CFZ) in human spiked plasma and in production machines using rinse and swab sampling collected from surfaces and application to Cosa®CIP Detergent. Isocratic chromatographic system was performed at ambient temperature using mobile phase consisting of acetonitrile: 40% tetrabutylammonium hydroxide adjusted to pH 7.0 ± 0.1 with 10% phosphoric acid (72.5:27.5, v/v) on Ultrasphere ion pair column (250 mm × 4.6 mm, 5.0 μm particle size) at a flow rate 1.0 mL/min, injection volume 10 μL and UV detection at 265 nm. The chromatographic run time was less than 20 min for the mixture. Linear relationships were obtained over the concentration ranges 0.5–25 ng mL^?1 for CPH, 1.5–30 ng mL^?1 for CFC, 2–33 ng mL^?1 for CFX, 3–35 ng mL^?1 for CFR and 4–40 ng mL^?1 for CFZ with correlation coefficients >0.998. Analytical and bioanalytical validation methods were carried out following terms of linearity, specificity, LOQ, LOD, accuracy and precision for determination of cephalosporin residues in production machines and in human spiked plasma according to FDA guidelines.     

Determination of Metal Ions by High Performance Liquid Chromatography

The mixture of metal ions [Bi(III), Fe(III), Fe(II), Cu(II), Zn(II), Ni(II), Co(II), Pb(II), Cd(II), Mn(II)] were separated in the bonded-phase strong cation exchange column (Vydac-401 SA) and monitored at 540 nm after a postcolumn reaction with 4-(2-pyridylazo)-resorcinol (PAR). Citrate, tartrate, lactate and α-hydroxyisobutyrate buffer were used as eluent and it has been found that the elution order of some metal ions were changed with different eluents. The detection limits and the calibration curves of metal ions were also studied.     

A Highly Sensitive Method for Determination of Copper (II) in Water Samples for Field Screening

Abstract

A new method has been developed for field screening of copper (II) in water samples, which is based on an enzyme inhibition reaction between copper (II) and nitrate reductase. The concentration of copper (II) was acquired by indirect determination the reaction product (nitrite) with a mini optical reflection sensor. Under the optimum conditions, the calibration graph was linear in the range of 5.0–50 ng mL^?1. The limit of detection was 0.5 ng mL^?1. This method has been used for the field screening of copper (II) with satisfactory results.     

Simple High-Performance Liquid Chromatographic Assay, with Post-Column Derivatization, for Simultaneous Determination of Mycophenolic Acid and its Glucuronide Metabolite in Human Plasma and Urine

Two simple high-performance liquid chromatographic (HPLC) methods have been established for simultaneous determination of mycophenolic acid (MPA) and its glucuronide metabolite (MPAG) in human urine, and of their total and unbound forms in human plasma. For total MPA and MPAG analysis sample preparation entailed precipitation of protein with acetonitrile and isolation of the free analytes from the plasma by ultrafiltration. For urine samples, fivefold dilution with water was used. MPAG was determined by UV detection whereas MPA was quantified by fluorescence detection after post-column derivatization with 0.2 M sodium hydroxide solution. For plasma, response was found to be linearly dependent on concentration over the ranges 0.1–40 μg mL^-1 and 0.01–1 μg mL^-1 for total and free MPA, respectively, and 10–200 μg mL^-1 and 2.5–100 μg mL^-1 for total and free MPAG, respectively. For urine, linearity was observed from 0.1 to 50 μg mL^-1 for MPA and 10 to 500 μg mL^-1 MPAG in the urine before dilution. The methods reported were found to be accurate and reproducible for quantifying the level of MPA and MPAG and can thus be used for clinical pharmacokinetic studies and for therapeutic drug monitoring. Contributed equally to this work An erratum to this article is available at .     

Application of dispersive liquid–liquid microextraction for the determination of donepezil in urine by HPLC using a core–shell column

A rapid and novel method combining dispersive liquid–liquid microextraction and high-performance liquid chromatography coupled with fluorescence detection was developed for the determination of donepezil in human urine. Parameters affecting extraction efficiency and chromatographic determination, such as the type and volume of the extraction and disperser solvent, pH of sample for dispersive liquid–liquid microextraction, mobile-phase composition, pH, column oven temperature, and flow rate for chromatographic determination, were evaluated and optimized. Using a C₁₈ core–shell column (7.5 × 4.6?mm, 2.7?μm), the determination of donepezil was accomplished within 5?min. Under optimum conditions, developed method was linear in the range of 0.5–25?ng?mL^?1 with the correlation coefficient >0.99. Limit of detection was 0.15?ng?mL^?1. The relative standard deviation at three concentration levels (2, 12.5, and 20?ng?mL^?1) was less than 11% with accuracy in the range of 96.9–102.8%. The results of this study demonstrate that the use of dispersive liquid–liquid microextraction and core–shell column can be considered as a powerful tool for the analysis of donepezil in human urine.     

Simple and rapid surface-enhanced Raman Spectroscopy assay for safranine T and its application in highly sensitive determination of mercury (Ⅱ)

A simple and sensitive surface-enhanced Raman spectroscopy (SERS) method for the detection of safranine T (ST) and Hg²⁺ using silver nanoparticles (AgNPs) as substrate was developed. ST can absorb on the surface of AgNPs through electrostatic interaction, the electromagnetic effect combined with chemical adsorption effect give a notable Raman enhancement for ST. The presence of Hg²⁺ well decreased the absorbed ST molecules on AgNPs, leading to a significant decrease of SERS signals thus enabling to detect Hg²⁺. The determination conditions for SERS, including the amount of AgNPs, the concentration of NaCl, the concentration of HCl, the concentration of ST and the reaction time, were optimised. Under the optimised experimental conditions, good linear responses were obtained for ST and Hg²⁺ in the concentration ranges of 0.01–4.0 μmol L^?1 (3.5–1403.4 ng mL^?1) and 0.01–2.0 μmol L^?1 (2.0–401.2 ng mL^?1), the limit of detection were 3.0 nmol L^?1 (1.1 ng mL^?1) and 2.0 nmol L^?1 (0.4 ng mL^?1), respectively. The present method was subsequently applied to the determination of ST in tomato sauces and Hg²⁺ in environmental waters, the recoveries of ST and Hg²⁺ in spiked samples are 95.5–107.8% and 91.4–110.8 %, respectively.     

Concurrent detection of cabozantinib as an anticancer agent and its major metabolites in human serum using fluorescence-coupled micellar liquid chromatography

A novel, highly sensitive, simple, and rapid strategy was designed and developed for simultaneous determination of cabozantinib (CBZ) as an anticancer agent and its main metabolites including monohydroxy sulfate (EXEL-1646), N-oxide (EXEL-5162(, amide cleavage product (EXEL-5366), and 6-desmethyl amide cleavage product sulfate) EXEL-1644). Measurements were done through a micellar liquid chromatography (MLC) method coupled with fluorescence detection. The high-performance liquid chromatography (HPLC) was performed using a Kinetex C18 100 Å column as well as acetonitrile, cetyltrimethylammonium bromide (CTAB; 0.2 mol.L^?1), and tris buffer (pH 8.5) solutions as the mobile phase at a 40:50:10 (v/v) ratio. The method’s linearity (20 to 700 ng.mL^?1), limit of detection (LOD; 2.11 to 3.69 ng.mL^?1), limit of quantification (LOQ; 20 to 30 ng.mL^?1), intra- and inter-day precisions (RSD < 4.00%), selectivity, recovery, and robustness were fully evaluated. According to the obtained results, the developed method can be used for simple and rapid (~35 min) quantification of CBZ as an anticancer drug and its major metabolites in human serum samples with high sensitivity and low cost.     

In-situ ionic liquid-dispersive liquid-liquid microextraction method to determine endocrine disrupting phenols in seawaters and industrial effluents

We have evaluated an in-situ ionic liquid-dispersive liquid-liquid microextraction procedure for the determination of six endocrine disrupting phenols in seawaters and industrial effluents using HPLC. The optimized method requires 38???L of the water-soluble ionic liquid 1-butyl-3-methylimidazolium chloride, and 5?mL of seawater or industrial effluent. After appropriate work-up, a drop (~10???L) of an ionic liquid is formed that contains the analytes of interest. It is diluted with acetonitrile and injected into the HPLC system. This procedure is accomplished without heating or cooling the solutions. The method is characterized by (a) average relative recoveries of 90.2%, (b) enrichment factors ranging from 140 to 989, and (c) precisions (expressed as relative standard deviations) of less than 11% when using a spiking level of 10?ng?mL^?1. The limits of detection range from 0.8?ng?mL^?1 for 4-cumylphenol to 4.8?ng?mL^?1 for bisphenol-A.