Analysis of Chlorpyrifos in Water,Fruit Juice,and Honeybee Extract by Chemiluminescent Elisa

Abstract

The suitability of competitive enzyme-linked immunosorbent assays (ELISAs) with chemiluminescent detection-based immobilized antigen (indirect assay) for rapid and accurate determination of chlorpyrifos in various food matrices was tested. The limit of detection (LOD) values were 1–1.75 ng mL^?1, the standard curve midpoint (IC₅₀) was 3.5 ng mL^?1, and the assay duration was 1.5 h. Assay application to the analysis of honeybee extract resulted in chlorpyrifos recoveries varying between 62 and 83% in 5–15 ng mL^?1 herbicide concentration range.     

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.     

Catalytic Square-Wave Voltammetric Determination of Acrylamide in Potato Chips

Abstract

Square-wave voltammetry is a fast technique used for determination of trace amounts of acrylamide. When cobalt(II) ions were added to the acrylamide solution, a catalytic peak at about ?1.35 V vs. Ag/AgCl was observed, which was proportional to acrylamide concentration. The calibration curve showed good linearity in the range of 200–800 ng mL^?1 of acrylamide with a regression coefficient of 0.9989. The limit of detection of the method was 3.52 ng mL^?1, and the relative standard deviations for concentrations of 300 ng mL^?1 and 700 ng mL^?1 were 99.8% × 10^?2 and 79.7% × 10^?2, respectively.     

Determination of Danofloxacin and Marbofloxacin in Milk Samples by Micellar Liquid Chromatography with Fluorescence Detection

Abstract

A simple, rapid, and sensitive analytical method has been developed for the determination of two fluoroquinolones, danofloxacin and marbofloxacin, in bovine milk samples. Separation and quantification were performed by micellar liquid chromatography with fluorescence detection (MLC?FD), using sodium dodecyl sulfate (SDS) as a surfactant. The influence of the principal factors, namely, the micelle concentration, the amount of organic modifier, tail‐reducing agents, the pH, and the temperature were studied. The suitable condition was found to be 75 mM SDS?10 mM phosphate buffer–18 mM tetrabutylammonium bromide/3% (v/v) 1‐propanol at pH 3.0 for the separation of marbofloxacin, danofloxacin, and tosufloxacin (internal standard) in about 20 min. The linear concentration range of application was 1.8–30.0 ng · mL^?1 for danofloxacin and 16–120 ng · mL^?1 for marbofloxacin, and the relative standard deviation ranged between 4.9 and 2.7%. The limit of detection found for danofloxacin was 0.5 ng · mL^?1 and 5 ng · mL^?1 for marbofloxacin. These values were lower than the maximum residue limits (MRLs) established by the European Union for these compounds in bovine milk. It was applied to check the eventual existence of these compounds above these limits on commercial milk samples. The validation method was completed with spiked milk samples. Recovery levels obtained were 90.3–108.2%.     

Sensitivity enhancement using nanometer silica particles on the surface of a quartz cell in mercury and selenium determination by vapor generation atomic absorption spectrometry

The sensitivity of on-line vapor generation atomic absorption spectrometry of mercury and selenium was improved by using a new atom trap technology. The inner wall of a T-shaped quartz tube was coated with nanometer SiO₂ so increase the residence time of the analyte atoms in the light path. The linear range of the calibration plots thus was increased to a range from 5.0 to 150 ng mL^?1 for mercury, and from 4.5 to 100 ng mL^?1 for selenium. The detection limits are 0.9 ng mL^?1 for Hg and 1.0 ng mL^?1 for Se which is a 2-fold improvement. The technique was applied to the determination of Hg and Se in herbs and hair.     

Rabbit Monoclonal Antibody-Based Lateral Flow Immunoassay Platform for Sensitive Quantitation of Four Sulfonamide Residues in Milk and Swine Urine

Based on the available rabbit monoclonal antibody (RabMAb), a rapid and sensitive lateral flow immunoassay (LFA) platform has been developed for quantitative detection of four sulfonamide residues(SRs) of sulfadiazine (SD), sulfathiazole (STZ), sulfapyridine (SP), and sulfamethoxazole (SMX).Within the designed LFA competitive format assay, which was based on antigen-antibody properties, the hapten conjugate N¹-[4-(carboxymethyl)-2-thiazolyl] sulfanilamide linked to protein ovalbumin (TS-OVA) and goat anti-rabbit antibody were sprayed as capture and control reagents, respectively, and then the antibody was conjugated to colloidal gold particles as the detection reagent. With quantitative assessment aided by a colorimetric strip reader, the sensitivities of the established LFA method for SD, STZ, SP, and SMX were 0.91 ng mL^?1, 0.10ng mL^?1,0.12ng mL^?1, and 2.13ng mL^?1, and the half-maximum inhibition concentrations (IC₅₀) were 5.19 ng mL^?1, 1.25 ng mL^?1, 0.66 ng mL^?1, and 24.14 ng mL^?1, respectively. The recoveries at three spiked levels (5, 20, 50 ng mL^?1for SD, STZ, and SP; 20, 50, 100 ng mL^?1 for SMX) were in the range of 78.02–135.10% and 76.40–137.16% for milk and swine urine, respectively. More importantly, the detection performance of the established platform was consistent with that of in-parallel LC-MS/MS analysis. In conclusion, the proposed LFA platform has showed the potential for fast, sensitive and relatively accurate quantification of four sulfonamide residues in practical uses.     

Development of an Indirect Chemiluminescent Competitive ELISA to Detect Danofloxacin Residues in Milk

In this study, a sensitive and specific indirect chemiluminescent enzyme-linked immunosorbance assay method (CL-ELISA) was developed to detect danofloxacin residue in milk. The influence of experimental factors on the results of the method was investigated, including the concentration of coating antigen and primary antibody, the dilution factor of the IgG-HRP antibody, as well as the type of microtiter plate. Under the optimized condition, the linear working range and IC₅₀ value were determined to be 0.025–0.5 ng · mL^?1 and 0.1 ng · mL^?1, respectively. Compared with traditional colorimetric ELISA, the CL-ELISA developed in this research demonstrated significant improvement in terms of sensitivity (20 times improvement) and linear range (25 times wider). The developed method was applied in detection of danofloxacin residue in milk and satisfied recovery rates of 92.6–105.2% and coefficient variations of 2.4–9.5% were obtained, demonstrating that the proposed method was accurate and reliable. Moreover, there was almost no any meaningful cross-reactivity with nine other (fluoro)quinolones, indicating its high specificity.     

Fluorimetric Determination of Gatifloxacin in Aqueous,Pure and Pharmaceutical Formulations

Abstract

A spectrofluorimetric method was developed for the determination of gatifloxacin. The emission peak for gatifloxacin was recorded at 495 nm upon excitation at 291 nm. The fluorescence process was pH dependent. The dynamic range for the method was 16–80 ng ml^?1with detection limit of 3.97 ng ml^?1. A linear relationship between the fluorescence intensity and the concentration of gatifloxacin solution was obtained with r ² of 0.9968. The method has successfully applied to the determination of gatifloxacin in pure, authentic and aqueous samples.     

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.     

A Novel Fluorescence Immunoassay Based on Two-time Amplified Fluorescence Signal by Hemin and Magnetic Nanoparticles for the Detection of Antigen

Abstract

A novel, selective, and sensitive magnetic-mimetic enzyme fluorescence immunoassay method for antigen detection has been developed by taking advantage of a magnetic separation process and the amplification feature of the hemin label. This method is based on a twice amplified fluorescence signal. The signal is first amplified due to the ultrasmall size and the high surface-to-volume ratio of the silica-coated magnetite nanoparticles, which enable the nanoparticles to carry much more antibodies. Second, the mimetic enzyme (hemin) as a labeling reagent catalyzes the reaction of p-hydroxyphenyl acetic acid and H₂O₂ can further amplify the fluorescence signal. This protocol was also evaluated for a sandwich-type immunoassay of human IgG, and the calibration graph for human IgG was linear over the range of 0–100 ng mL^?1 with a detection limit of 9.8 ng mL^?1. This method can easily separate magnetic nanoparticles from the solution, which simplified the process and played a promising role for various applications in immunoassay.     

Study on the Effect of Moderate Exercise on Lactic Acid Content in Breast Milk by Indirect CE with Amperometric Detection

An indirect high-performance capillary electrophoresis with amperometric detection (CE-AD) method has been developed for determination of lactic acid (LA) in body fluids of lactating postpartum women. Several important factors such as the running buffer additive and concentration, the working electrode potential, the pH value and concentration of the running buffer, the separation voltage and injection time were investigated. Under the optimum conditions, LA could be well separated with co-existing interferences including uric acid (UA) in real samples in a 90-cm-length capillary at separation voltage of 12 kV in 4.0 × 10^?6 g mL^?1 3,4-dihydroxybenzylamine (DHBA)/40 mmol L^?1 H₃BO₃?CNa₂B₄O₇ buffer (pH 7.8). The linearity between peak current and concentration of LA was over three orders of magnitude with detection limit of 5.00 × 10^?7 g mL^?1 (S/N = 3). This proposed method has been successfully used to study the effects of moderate exercise on LA content in breast milk and urine samples of lactating postpartum women, and assay results showed that LA content in breast milk can return to normal level through 60 min rest without decreasing acceptance by breast-feeding infants, although the LA level did increase by 4?C6 times in both breast milk and urine samples at 10 min after moderate exercise.     

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.     

Simultaneous derivatization and extraction of iodine from milk samples by hollow fiber liquid-phase microextraction followed by gas chromatography-electron capture detection

A simple, low-cost and sensitive method is demonstrated for derivatization and extraction of iodine from milk samples using hollow fiber liquid-phase microextraction (HF-LPME) and gas chromatography-electron capture detection. Iodide ions are converted to iodine under acidic medium and in the presence of an oxidant. The generated iodine reacted with 3-pentanone in extraction vial to give 2-iodo-3-pentanone and was extracted into 4 μL of 1-octanol located in the lumen of a hollow fiber. Organic solvent was selected using one variable at a time optimization method and the other main factors affecting derivatization and HF-LPME procedures were evaluated using a Taguchi’s L₁₆ (4⁵) orthogonal array. Under optimal conditions, the method showed low limit of detection (0.5 ng mL^?1), wide linear range (1–2,000 ng mL^?1) with good correlation coefficient (0.9997) and acceptable relative standard deviation (4.6 %, n = 5). Finally, the developed method was successfully applied for determination of iodide in real samples including infant milk formulas and cow milk with reasonable relative recoveries (99.8–110.5 %).     

Quantification of Thiram in Honeybees: Development of a Chemiluminescent ELISA

Abstract

A Chemiluminescence Enzyme‐Linked Immuno‐Sorbent Assay (CL‐ELISA) for determination and quantification of the fungicide thiram in honeybees was developed in an indirect competitive format. The assay was optimized by determining: the optimal coating conjugate concentration and anti‐thiram antiserum dilution, the effect of the incubation time on the competitive step, the tolerance to organic solvents. The IC₅₀ and the limit of detection (LOD) values were 60 ng mL^?1 and 9 ng mL^?1, respectively, similar to those of colorimetric ELISA with a calibration range of 9–15,000 ng mL^?1. Cross reactivity of some related compounds such as some dithiocarbamates, a thiocarbamate, the ethylenethiourea and the tetramethylthiourea were tested. The assay was then applied to honeybees sample extracts obtained by using the liquid‐liquid extraction or the graphitized carbon‐based solid phase extraction.

The calibration curves in honeybee extracts from liquid‐liquid procedure gave an IC₅₀ of 141 ng mL^?1 and a LOD of 17 ng mL^?1. In case of extracts obtained by SPE these values were 139 ng mL^?1 and 15 ng mL^?1, respectively. The average recovery value from honeybee extracts spiked with 75 ng mL^?1 of thiram was 72% for SPE, higher than for liquid‐liquid extraction (60%). On the opposite, when the honeybees were directly spiked with 2 and 10 ppm the average recovery was higher for liquid‐liquid extraction (54%), than for SPE (31%). Finally, the assay was applied to honeybee samples collected during monitoring activities in Italy and Russia.     

Mixed immunoassay design for multiple chemical residues detection

In this research, a mixed immunoassay design for multiple chemical residues detection based on combined reverse competitive enzyme-linked immunosorbent assay (ELISA) procedure was developed. This method integrated two reverse ELISA reactions in one assay by labeling horseradish peroxidase to deoxynivalenol (DON) and orbifloxacin. Within this method, IC50 of the two mAbs for each analyte we produced ranged from 23?～?68 ng?mL^?1 for DONs and 4.1?～?49 ng?mL^?1 for quinolones (QNs). The limit of detection measured by IC10 was achieved at 0.45–1.3 ng?mL^?1 for DONs and 0.59–6.9 ng?mL^?1 for QNs, which was lower than the maximum residue levels. Recoveries in negative samples spiked at concentrations of 100, 200, and 500 ng?mL^?1 ranged from 91.3 to 102.2 % for DONs and 88.7–98.05 % for QNs with relative standard deviation less than 9.88 and 12.67 %. The results demonstrated that this developed immunoassay was suitable for screening of low molecular weight contaminants.

Optimization of Continuous Flow Hydride Generation Inductively Coupled Plasma Optical Emission Spectrometry for Sensitivity Improvement of Bismuth

Experimental variables in continuous flow hydride generation inductively coupled plasma-optical emission spectrometry (CF-HG-ICP-OES) were optimized for determination of bismuth. Concentrations of NaBH₄, HCl, and NaOH, flow rates of NaBH₄, sample solution, waste and carrier argon, radio frequency power, lengths of reaction, and stripping coils were optimized to obtain lower detection limits. Under optimum conditions, the detection limit was calculated as 0.16 ng mL^?1, and the calibration plot was linear between 1.0–50.0 ng mL^?1. An improvement in detection limit of 5.75 times by CF-HG-ICP-OES was reached vs. ICP-OES. Relative standard deviation (RSD) for ten replicate measurements of 10.0 ng mL^?1 Bi was calculated as 3.9%. Effect of possible interferic ions on Bi signal was evaluated. Accuracy of method was verified by using a standard reference material, SRM 1643e. Results found for Bi were in satisfactory agreement with certified values. The proposed method was then employed to determine trace concentration of Bi in milk samples. Bi amounts in samples were found in the range from lower than the quantitation limit to 14.5 ng mL^?1, whereas Bi concentrations were lower than the detection limit in three samples.     

Indirect Competitive Enzyme-Linked Immunosorbent Assay for Detection of Tylosin in Milk and Water Samples

A specific polyclonal antibody was prepared based on a new hapten with stable structure. Based on this, an indirect competitive enzyme-linked immunosorbent assay (icELISA) was established for determination of tylosin residues in food and environmental samples. The experimental conditions were optimized as follows: the coating antigen and antibody were respectively diluted by 6000 times and 2000 times, and the competitive reaction time was 40 min. Under the optimized conditions, the method showed a detection limit of 0.07 ng mL^?1, an IC₅₀ of 1.39 ng mL^?1 and a linear range of 0.17–11.0 ng mL^?1. The recoveries of spiked raw milk and water samples were ranged from 78.4% to 105.6%, with the RSDs of less than 15%. Good correlation between icELISA and HPLC method was obtained for spiked samples (R² = 0.97). This method was suitable for the determination of tylosin residues in milk and water samples.     

Nonextractive Trace Level Simultaneous Determination of Mercury(II) and Zinc(II) in Environmental Samples with 2‐(5‐Bromo‐2‐pyridylazo)‐5‐diethylaminophenol and Cetylpyridinium Chloride

Abstract

A simple, rapid, selective, and sensitive method for the derivative spectrophotometric determination of Hg(II) and its simultaneous determination in the presence of Zn(II) using 2‐(5‐bromo‐2‐pyridylazo)‐5‐diethylaminophenol in the presence of cetylpyridinium chloride, a cationic surfactant, has been developed. The molar absorption coefficient and analytical sensitivity of the 1∶1 Hg(II) complex at 558 nm (λ_max) are 5.78×10⁴ L mol^?1 cm^?1 and 0.67 ng mL^?1, respectively. The detection limit of Hg(II) is 1.40×10^?2 ng mL^?1, and Beer's law is valid in the concentration range 0.05–2.40 µg mL^?1. Overlapping spectral profiles of Hg(II) and Zn(II) complexes in zero‐order mode interfere in their simultaneous determination. However, 0.10–2.00 µg mL^?1 of Hg(II) and 0.065–0.650 µg mL^?1 of Zn(II), when present together, can be simultaneously determined at zero cross point of the derivative spectrum, without any prior separation. The relative standard deviation for six replicate measurements of solutions containing 0.134 µg mL^?1 of Hg(II) and 0.620 µg mL^?1 of Zn(II) is 1.72 and 1.47%, respectively. The proposed method has successfully been evaluated for trace level simultaneous determination of Hg(II) and Zn(II) in environmental samples.     

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.     

Immunofluorescent Analysis with Magnetic Nanoparticles for Simultaneous Determination of Antibiotic Residues in Milk

An analytical system for simultaneous detection of three antibiotic residues in milk (penicillin, tetracycline, and sulfadimethoxine sodium salt) was developed. The method was based on a fluorescence immunoassay. Magnetic nanoparticles (MNPs) were prepared and then coated with 3-(aminopropyl) triethoxysilane (APTES). Antibodies, against penicillin, tetracycline, and sulfadimethoxine, were immobilized on APTES-MNPs. Two immobilization methods were used: a random and an oriented method by protein A. The fluorochrome-antigen conjugates, containing FITC, ATTO 590, and ATTO 630 were obtained. Three separate immunofluorescent analyses were investigated for penicillin, tetracycline, and sulfadimethoxine, first in buffer and then in milk. Linearity values of standard curves in milk were: for penicillin 4–15 ng mL^?1, for tetracycline 50–500 ng mL^?1 (using random immobilization) and 100–500 ng mL^?1(using oriented immobilization), and for sulfadimethoxine 100–500 ng mL^?1. The linearity and sensitivity of calibration curves of determination of penicillin, tetracycline and sulfadimethoxine in milk were very close to the obtained results for separate determination of three antibiotics. It was shown that the multiple immunofluorescence analysis for antibiotic residues in milk without preliminary treatment is effective and rapid.