High-performance Liquid Chromatography-fluorescence Detection Research Articles

High-Performance Liquid Chromatography–Fluorescence Detection Method for Ochratoxin A Quantification in Small Mice Sample Volumes: Versatile Application across Diverse Matrices Relevant for Neurodegeneration Research

Ochratoxin A (OTA) is a mycotoxin commonly found in various food products, which poses potential health risks to humans and animals. Recently, more attention has been directed towards its potential neurodegenerative effects. However, there are currently no fully validated HPLC analytical methods established for its quantification in mice, the primary animal model in this field, that include pivotal tissues in this area of research, such as the intestine and brain. To address this gap, we developed and validated a highly sensitive, rapid, and simple method using HPLC-FLD for OTA determination in mice tissues (kidney, liver, brain, and intestine) as well as plasma samples. The method was rigorously validated for selectivity, linearity, accuracy, precision, recovery, dilution integrity, carry-over effect, stability, and robustness, meeting the validation criteria outlined by FDA and EMA guidelines. Furthermore, the described method enables the quantification of OTA in each individual sample using minimal tissue mass while maintaining excellent recovery values. The applicability of the method was demonstrated in a repeated low-dose OTA study in Balb/c mice, which, together with the inclusion of relevant and less common tissues in the validation process, underscore its suitability for neurodegeneration-related research.

Establishment of the HPLC fluorescence detection method for plasma trace ivermectin and its pharmacokinetics in Bactrian camel.

Ivermectin (IVM), a widely used veterinary anthelmintic, lacks recommended doses for Bactrian camels. This study aims to establish its pharmacokinetics in Bactrian camels, comparing with other livestock. A method for high-performance liquid chromatography fluorescence detection of IVM in plasma was developed. IVM exhibited linear scaling (y=0.6946x+0.0088, R2=0.9988) within 0.025-5ng/mL, with a lower limit of quantification of 25.00pg/mL, high recovery (>70%) and low RSD (<7%). In Bactrian camels, IVM injection showed a low Cmax, extended Tmax and apparent secondary absorption compared to cattle and sheep. Slow absorption and widespread distribution were observed, with peak concentration and area under the curve correlating positively with the dose. This study provides insights into IVM pharmacokinetics in Bactrian camels, informing dose determination and highlighting potential metabolic differences compared to other livestock.

An integrated method for rapid extraction of fluorescent whitening agents from selected cosmetics by in situ formation of deep eutectic solvent

A novel extraction methodology was developed based on deep eutectic solvent (DES) in situ formation. The solvent was synthesized from a binary mixture of L-cysteine, functioning as a hydrogen bond acceptor, and 2-methylphenol, serving as a hydrogen bond donor. The DES’s inherent molecular structures and extensive hydrogen bonding networks were characterized using the Fourier-transform infrared spectroscopy (FT-IR), while its hydrophobic nature was assessed by determining molecular polarity. Its extraction efficiency was demonstrated through the extraction of fluorescent whitening agents (FWAs) in cosmetic products, acting as a model analytical application. This was accomplished using vortex-assisted emulsification solid–liquid extraction (SLE) method, followed by high-performance liquid chromatography-fluorescence detection. Response surface methodology was employed to optimize extraction conditions in a multivariate experimental design. This approach streamlined the multi-operation extraction process via ingeniously integrating extraction and clean-up into a single step, resulting in significant reductions in extraction time required and high sample throughputs of approximately 40 samples per hour. The methodology was validated and exhibited excellent linearity in concentration ranges of 0.01–24 mg/L for five FWAs and 0.01–40 mg/L for other three FWAs, with correlation coefficients (r) ranging from 0.9985 to 1.000. The limits of detection and quantification were in the range of 0.03–0.26 mg kg−1 and 0.10–0.87 mg kg−1, respectively. The reliability and practicality of the proposed method were affirmed by quantifying eight FWAs in ninety-four commercially available facial masks and whitening creams, yielding recoveries values between 90.0 % and 106.0 % with relative standard deviations (RSDs) of 1.0–4.3 %. Inorganic ions and organic compounds present in the samples had minimal impact on FWAs detection following sample pre-treatment. This procedure showcased exceptional extraction capabilities towards a diverse array of FWAs with the aid of multiple interactions, suggesting significant potential for increasing sample throughput while reducing organic solvent and material consumptions compared with conventional SLE methods. The low-toxicity DES is an excellent alternative to traditional extractants typically applied to SLE techniques.

In situ extraction and enrichment of multiple amino acids by integrated mechanochemical extraction and aqueous two-phase system

A highly efficient and environment-friendly extraction technology was established to extract and enrich multiple amino acids (AAs) in situ by deep eutectic solvent (DES)-assisted mechanochemical extraction (MCE) and aqueous two-phase system (ATPS). Firstly, the DES was prepared from betaine and ethylene glycol in a 1:2 M ratio. Then, a series of conditions were evaluated by single-factor and multifactor experiments to select the optimum experimental parameters. The highest extraction yields of AAs were obtained by DES-assisted MCE and ATPS enrichment, including that the amount of DES was 0.97 mL, the milling time was 4.88 min, the amount of salt was 8 g, and the solid-to-liquid ratio was 0.055 g mL−1. After extracting, the diluted AA extracts were analyzed by a high-performance liquid chromatography-fluorescence detector after pre-column derivatization with 6-aminoquinolyl-N-hydroxysccinimidyl carbamate. The methodological validation demonstrated the proposed method exhibited satisfactory limits of detection ranged from 0.11 to 0.82 ng mL−1, reliable interday precision (%RSD) of 0.014–0.067 % for retention time and 1.29–3.34 % for the peak area, acceptable reproducibility with %RSD below 4.1 %, and good %recovery between 90.3 % and 122.5 % at three spiked concentrations. The surface morphology of raw, only ground, and DES-ground sample powders was tested by scanning electron microscopy to further explore the changes in the physicochemical properties of samples. By comparing with other AAs extraction methods, the developed method is not only eco-friendly, but greatly improves the yield of free AAs extracted from chrysanthemum.

Determination of ochratoxin A in brown rice through a rapid, straightforward strategy based on FaTEx coupled with high-performance liquid chromatography–fluorescence detection

Ochratoxin A is a mycotoxin commonly found in cereals. In this study, a rapid and straightforward method was developed for measuring trace concentrations of ochratoxin A in brown rice; the method combines the fast toxin extraction (FaTEx) technique with high-performance liquid chromatography–fluorescence detection. The proposed method was validated and exhibited high performance and satisfactory accuracy (91.6%–99.2%), precision (relative standard deviation < 3%), linearity (R2 > 0.9999), sensitivity (limit of quantification = 1.0 ng/g), and matrix effects (magnitude = −2%). The proposed method is simple, accurate, inexpensive, reliable, and environmental friendly and requires minimal sample preparation. It was successfully used to analyze brown rice samples from markets and rice shops in Taiwan. To our knowledge, no study has reported the use of FaTEx for the extraction of ochratoxin A from cereals. The proposed method overcomes the shortcomings of traditional column cleanup and satisfies current food safety and analysis demands.

Determination of bisphenols in beeswax based on sugaring out–assisted liquid–liquid extraction: Method development and application in survey, recycling and degradation studies

The methodology of sugaring out–assisted liquid–liquid extraction (SULLE) coupled with high-performance liquid chromatography–fluorescence detection was devised for quantifying bisphenol A (BPA) and bisphenol B (BPB) in beeswax. The effectiveness of SULLE was methodically explored and proved superior to the salting out–assisted liquid–liquid extraction approach for beeswax sample preparation. The analytical performance underwent comprehensive validation, revealing detection limits of 10 μg/kg for BPA and 20 μg/kg for BPB. The method developed was employed to analyse commercial beeswax (n = 15), beeswax foundation (n = 15) and wild-build comb wax (n = 26) samples. The analysis revealed BPA presence in four commercial beeswax samples and three beeswax foundation samples, with the highest detected residue content being 88 ± 7 μg/kg. For BPB, two beeswax foundation samples were positive, with concentrations below the limits of quantification and 85 ± 4 μg/kg, respectively. No bisphenols were detected in wild-build comb wax. Furthermore, the bisphenol removal efficacy of two recycling methods—boiling in water and methanol extraction—was assessed. The findings indicated that after four recycling cycles using water boiling, 9.6% of BPA and 29.2% of BPB remained in the beeswax. Whereas methanol extraction resulted in approximately 7% residual after one recycling process. A long-term study over 210 days revealed the slow degradation of bisphenols in comb beeswax. This degradation fitted well with a first-order model, indicating half-lives (DT50) of 139 days for BPA and 151 days for BPB, respectively. This research provides the first report on bisphenol contamination in beeswax. The low removal rate during the recycling process and the gradual degradation in beeswax underscore the significance of bisphenol contamination and migration in bee hives along with their potential risk to pollinators warranting concern. Furthermore, the developed SULLE method shows promise in preparing beeswax samples to analyse other analytes.

Development of a QuEChERS-HPLC-FLD Procedure for the Simultaneous Detection of Residues of Florfenicol, Its Metabolite Florfenicol Amine, and Three Fluoroquinolones in Eggs.

A method utilizing high-performance liquid chromatography-fluorescence detection (HPLC-FLD) has been developed and refined for the simultaneous detection of florfenicol (FF) and its metabolite florfenicol amine (FFA) along with three fluoroquinolone (ciprofloxacin (CIP), enrofloxacin (ENR), and sarafloxacin (SAR)) residues in different parts of eggs (whole egg, egg yolk, and egg albumen). The QuEChERS ("Quick, easy, cheap, effective, rugged, and safe") procedure utilized 0.1 M disodium EDTA solution, water, and acetonitrile as extractants; sodium sulfate, sodium chloride, and trisodium citrate as dehydrating salts; and N-propylethylenediamine and C18 as adsorbents. A dual-channel FLD method was utilized to analyze the target compounds using an XBridge BEH C18 chromatographic column (4.6 mm × 150 mm, 5 μm). The mobile phase was employed isocratically using a solution of 0.01 M sodium dihydrogen phosphate, 0.005 M sodium dodecyl sulfate, and 0.1% triethylamine (pH 4.8) in combination with acetonitrile at a ratio of 65:35 (V/V). The limits of detection (LOD) and quantification (LOQ) of the analytes ranged from 0.03 to 1.5 µg/kg and from 0.1 to 5.0 µg/kg, respectively. The recoveries of the analytes in the blank egg samples ranged from 71.9% to 94.8% when reference standard concentrations of the LOQ, half of the maximum residual limit (MRL), MRL, and twice the MRL were added. The parameters of the presented protocol were validated and subsequently applied to the analysis of real samples, demonstrating the applicability and reliability of the method.

A novel method for evaluating pseudoallergy based on β-hexosaminidase and its application for traditional Chinese medicine injections.

Pseudoallergy is a typical and common adverse drug reaction to injections, especially in traditional Chinese medicine injections (TCMIs). At present, the evaluation methods for pseudoallergy include cell methods in vitro and animal methods in vivo. The mast cell evaluation method based on the β-hexosaminidase (β-Hex)-catalyzed substrate, 4-nitrophenyl-β-N-acetyl-D-glucosaminide (4-NPG), is an important method for the evaluation of drug-induced pseudoallergy, but it is prone to false positive results and has insufficient sensitivity. In this study, a novel β-Hex evaluation system with rat basophilic leukemia-2H3 cells based on high-performance liquid chromatography-fluorescence detection (HPLC-FLD) was established, which effectively increased the sensitivity and avoided false positive results. Cell viabilities were measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide assay. In addition, a method for the determination of histamine, which is another indicator in the development of pseudoallergy, was established to validate the above method. The results of this novel method indicated that two TCMIs (Shuxuening injection and Shenqi Fuzheng injection), which were considered to be pseudoallergenic using 4-NPG, were not pseudoallergenic. Overall, the novel β-Hex/HPLC-FLD evaluation system using Rat basophilic leukemia-2H3 cells established was effective and precise. It could be used for the evaluation of pseudoallergic reactions caused by TCMIs and other injections.

Adaptive polarity of graphene oxide anchored silica doped with C18 for effective enrichment of aflatoxins from foodstuff

Selecting adsorbent with appropriate polarity is a practicable way to improve the enrichment efficiency for aflatoxins (AFs) during the solid phase extraction in sample pretreatment. In this study, graphene oxide anchored silica was prepared and applied to dop with C18 with suitable polarity for effective pre-concentration and purification of AFs from foodstuff. With systematic optimizing, GO anchored silica/C18 (1/1, w/w) exhibited excellent extraction performance for AFs. As the medium, a facile and efficient pipette-tip solid-phase extraction (PT-SPE) technique was developed for enrichment of AFs. Coupling with high-performance liquid chromatography fluorescence detection (HPLC-FLD), the developed analysis method illustrated the lower limits of detection of 0.075 ng/g and 0.17 ng/g for AFG2, AFB2 and AFG1, AFB1 (R2 > 0.99). The recoveries were calculated as 71.27 ∼ 94.52 % for the spiking level of 1, 5, and 20 ng/g with the intra-day and inter-day precision of 1.30 to 4.01 and 3.56 to 5.78, respectively. Hence, the developed PT-SPE-HPLC-FLD determination method was sensitive, accurate, and convenient for quantitative analysis of AFs at trace contents in foodstuff.

A novel deep eutectic solvent as an extractant for the determination of bisphenol compounds in edible oils by high-performance liquid-phase fluorescence detection

In this study, an efficient, low-cost and environmentally friendly deep eutectic solvent (DES) was prepared using methanol and betaine, which was then employed as an extractant to detect BPA and BPF in several edible oils by high performance liquid chromatography-fluorescence detection (HPLC-FLD) with good results. The electrostatic potentials of methanol, betaine, water and BPs were analyzed by quantum chemical computational techniques to explain the mechanism of DES extraction. Several important parameters affecting the extraction efficiency, composition, volume and extraction time of DES were optimized. The optimum conditions for the use of DES were the molar ratio of 1:30 (betaine: methanol), the volume of DES of 200 μL and the extraction time of 1.5 min. The results of the method evaluation showed a wide linear range (0.01–0.8 μg g−1) and a high linear correlation coefficient (R2 ≥0.994). The limit of detection (LOD) and limit of quantification (LOQ) values ranged from 1.65 to 2.92 ng g−1 and 5.45–9.64 ng g−1, respectively. The greenness of the assay was evaluated by applying the green analytical procedure index method, and the results showed that the experimental method has the advantages of simple sample pre-treatment and low amount of waste generated during the assay. The experimental method has potential applications in the field of the detection of bisphenol compounds in edible oils.

Smartphone as a fluorescence detector for high-performance liquid chromatography

BackgroundFluorescence detection is employed in high-performance liquid chromatography (HPLC) due to its high specificity and sensitivity. However, it is often limited by expensive components and bulkiness. Recently, advances in technology and electronics have led to the development of smartphones that can serve as portable recording, analysis, and monitoring tools. Smartphone-based detection provides advantages of cost effectiveness, rapid signal/data processing, and the display of results on a handhold monitor. The combination of smartphone-based detection with HPLC can offer unique features that are beneficial in overcoming limitations of commercial fluorescence detectors. (90) ResultsA miniaturized and low-cost HPLC fluorescence detector based on a smartphone is introduced for the detection of six fluorescent molecules. The smartphone is able to capture emitted fluorescence in video format while MATLAB code is used for data processing to provide chromatograms based on different detection channels. A custom designed double-channel flow cell was utilized to enable simultaneous detection of fluorescent compounds with different excitation wavelengths. The detector consists of a lab-made flow cell, monochromatic LEDs as the light source, 3D printed housing and connector box, fiber optic cables, and a smartphone. The effects of flow cell geometry, channel width and light slit diameter, as well as a comparison of different flow cell manufacturing techniques, are studied and discussed. The validated system was successfully applied to samples from diverse water sources, yielding spiking recoveries within the range of 91.7% and 109.7%. (141) SignificanceThis study introduces the first smartphone-based fluorescence detector for HPLC with cost-effective and customizable flow cells, allowing for the simultaneous detection of fluorescent compounds with different excitation wavelengths and offering a potential solution for the analysis of co-eluting compounds. Beyond its user-friendly interface and low-cost, smartphone detection in HPLC provides tremendous opportunities in further miniaturizing chromatographic instrumentation while offering high sensitivity and can be expanded to other mechanisms of detection. (70)

High-performance liquid chromatography with fluorescence detection method for measuring colistin in human serum and its application in critically ill patients treated with colistin methanesulfonate sodium

Colistin, in the form of colistin methanesulfonate sodium (CMS), has a narrow therapeutic window, and colistin levels must be monitored during treatment. The aim of this study was to develop and validate a high-performance liquid chromatography-fluorescence detection (HPLC-FLD) method for measuring colistin in human serum. Colistin was extracted from human serum using trichloroacetic acid and methanol for protein precipitation, followed by in-solid phase extraction derivatization with 9-fluorenylmethyl chloroformate. HPLC-FLD system using a reverse-phase HPLC C18 column and a mobile phase consisting of acetonitrile, tetrahydrofuran, and water (80%:4%:16%) were used for the quantification of colistin A, colistin B, and polymyxin B (internal standard) in human serum. The extraction recoveries were between 71% and 103%. Linear calibration curves were obtained for colistin in concentrations from 0.3 to 8.0 μg/mL, with good fit (r2 = 0.9993). The lower limit of quantitation was 0.3 μg/mL. The intra- and inter-day precision of the assay was 0.5–5% and 3.5–9.4%, respectively. The accuracy ranged from 98% to 100%. This validated method was successfully applied to the analysis of serum-receiving CMS in critically ill patients. This method will be useful to assist colistin dosage adjustment based on individual blood colistin levels for optimization of colistin therapy.

Highly sensitive zero-order emission system - fluorescence detection-liquid chromatographic method for simultaneous estimation of two banned phenyl xanthene dyes in bottled chili sauce

ABSTRACT. A zero-order emission system was applied to address the variation in emission wavelengths between analytes. Also, it assists to compensate for the drop in emission intensity resulted from the changes in analyte excitation wavelength. So, a high performance liquid chromatography-fluorescence detector (HPLC-FLD) with zero order emission mode was used for the simultaneous estimation of rhodamine B (RhB) and eosin Y (EsY) based on their native fluorescence. The presented method is considered the first fluorescence method for simultaneous determination of RhB and EsY. The efficiency of the developed method was tested by determining the two dyes in a high matrix sample, chili sauce. Under the optimum condition, a linear calibration graphs for RhB and EsY in range of 0.5 -300 ng/mL with r = 0.9994 and 0.9991, respectively, were acquired. The limits of detection for RhB and EsY were 0.344 ng/mL (0.355 ng/g) and 0.215 ng/mL (0.222 ng/g), respectively. The recovery percentage and relative standard deviation of two analytes were obtained in the range of 98.38-101.86% and 0.83-1.66%, respectively. RhB was detected only in one chili sauce sample at a concentration of 0.388 ng/mL while EsY was not found in any sample.&#x0D; KEY WORDS: Rhodamine B, Eosin Y, HPLC-FLD, Zero-order emission mode, Solid phase extraction&#x0D; Bull. Chem. Soc. Ethiop. 2023, 37(6), 1315-1323. DOI: https://dx.doi.org/10.4314/bcse.v37i6.2

Construction of magnetic hydroxyl group-enriched hyper cross-linked polymers with functional triazine as the core for efficient enrichment of plant growth regulators

The determination of trace plant growth regulators (PGRs) residues in water and food samples make it crucial to develop novel sample pretreatment methods for the enrichment of PGRs. Herein, a novel magnetic hyper cross-linked polymer (M-CTT-9OH-HCP) was constructed and served as a magnetic adsorbent for the efficient extraction of some PGRs from water, watermelon, tomatoes, and milk samples for the first time. Combined with high performance liquid chromatography-fluorescence detection (HPLC-FLD), the established method presented a good linearity (0.03–60.0 ng g−1 (ng mL−1), (r) ≥ 0.9973), satisfactory accuracy with method recoveries (83.0%-119%) and acceptable precision with the intra-day and inter-day variations (expressed as the relative standard deviations (RSDs) ≤ 9.8%). The limits of detection (LODs) and limits of quantitation (LOQs) were in the range of 0.01–1.50 and 0.03–5.00 ng g−1/ ng mL−1. The results show that the established method is sensitive and efficient for the determination of PGRs in real samples.

Simultaneous determination of six α-dicarbonyl compounds in traditional Chinese medicines using high-performance liquid chromatography-fluorescence detector with pre-column derivatization.

A reliable method for determination of six α-dicarbonyl compounds in traditional Chinese medicines was first developed and validated by high-performance liquid chromatography-fluorescence detector with pre-column derivatization. α-Dicarbonyl compounds in traditional Chinese medicines were extracted and derivatized with 2,3-diaminaphthalene. The derivatization procedure of six α-dicarbonyl compounds was confirmed by high-resolution mass spectrometry. The limits of quantitation for six α-dicarbonyl compounds ranged from 3.70×10-3 to 2.21×10-2 μM. The established method showed good linearity (regression coefficient>0.9990), precision (relative standard deviation<3.37%), and high recovery (97.8%∼113.1%). The developed method was successfully applied to detect the six α-dicarbonyl compounds in traditional Chinese medicines. The result exhibited six α-dicarbonyl compounds was found in the 15 kinds of traditional Chinese medicines, which suggested us that the determination of α-dicarbonyl compounds should be paid more attention in the quality control of traditional Chinese medicines.

Study on the effectiveness of a multi-toxin immunoaffinity clean-up for reliable cost-effective HPLC-FLD analysis of mycotoxins in corn-based foods

A different, reliable, and cost-effective strategy for the analysis of aflatoxins, ochratoxin A, and zearalenone in corn-based foods was proposed, including one multi-toxin immunoaffinity column (IAC) sample preparation and three different high-performance liquid chromatography fluorescence detection methods. The analytical procedures were tested and verified, keeping in mind their occurrence at trace levels in corn-based foods. With the validation of the proposed multi-toxin IAC methodology and comparison of the performance characteristics with methods using a single-toxin IAC, we confirmed the reliability of the multi-toxin IAC procedure versus the single-toxin IAC. The methods were validated by revealing satisfactory performance characteristics; for example, the obtained values of limit of detection were significantly lower than the maximum limits for all mycotoxins of concern. In addition, the recovery values were between 70.9 % and 106.1 % for all mycotoxins of interest, with precision values lower than 10.5 %.  

Vitamins E and A increase the passing of the P-gp substrate ivermectin into the brain in mice.

This study aimed to determine the effect of administration of oral vitamins A and E at different doses on plasma and brain concentrations of ivermectin in mice. The study was carried out on 174 Swiss Albino male mice aged 8-10 weeks. After leaving six mice for method validation, the remaining mice were randomly divided into seven groups with equal numbers of animals. Mice received ivermectin (0.2mg/kg, subcutaneous) alone and in combination with low (vitamin A: 4000IU/kg; vitamin E: 35mg/kg) and high (vitamin A: 30 000IU/kg; vitamin E: 500mg/kg) oral doses of vitamins A and E. The plasma and brain concentrations of ivermectin were measured using high-performance liquid chromatography-fluorescence detector. We determined that high doses of vitamins A and E and their combinations increased the passing ratio of ivermectin into the brain significantly. The high-dose vitamin E and the combination of high-concentration vitamins E and A significantly increased the plasma concentration of ivermectin (P<0.05). The high-dose vitamins E and A and their high-dose combination increased the brain concentration of ivermectin by 3, 2, and 2.7 times, respectively. This research is the first in vivo study to determine the interaction between P-gp substrates and vitamins E and A.

A Rapid Quantification of Hydroxyproline in Leather using High-Performance Liquid Chromatography - Fluorescence Detection (HPLC-FLD) Method

In the area of leather development research, determining collagen content through measurement of 4-Hydroxyproline (Hyp) has been major challenging task due to the interference of various leather-processing chemicals. To overcome this problem, for the first time we have used a High-Performance Liquid Chromatography coupled with Fluorescence Detection (HPLC-FLD) method for accurate determination of Hyp in leather samples through chemical derivatization of Hyp with 7-chloro 4 nitro benzofuran (NBD-Cl) reagent. The HPLC-FLD analysis was performed on a PICO TAG column with an isocratic mobile phase (80 % of 0.1M, pH 7.2 sodium acetate and 20% of acetonitrile, v/v) at a flow rate of 0.7mL/min. The detection was carried out at an excitation wavelength of 465 nm and its emission at 535nm. The retention time of Hyp was found to be ~3.5 minutes and the total run time was about 10 minutes. The method validation indicated that this analytical method is precise (3-12%RSD), accurate (90-100%), the limit of detection 0.01μg/ml, the limit of quantification - 0.03 μg/ml and linear (R2 -0.9995) over the concentration range of 0.1 -2.0 μg/ml. The obtained result indicated that the assay linear range was acceptable for repeated analysis and suitable for the complete range of hydroxyproline levels present in leather samples. Compared to the traditional method (IUC 17:1980), this analytical method demonstrates higher simplicity, specificity, reproducibility, and it could be useful for certifying leather products as well as inspecting international trade in leather and hides.

Dissipation Dynamic, Residue Distribution and Risk Assessment of Emamectin Benzoate in Longan by High-Performance Liquid Chromatography with Fluorescence Detection.

A derivatization method combined with high-performance liquid chromatography-fluorescence detection (HPLC-FLD) was used to evaluate the dissipation, residue distribution and risk assessment of emamectin benzoate in whole longan and pulp. The average recoveries were 82-111% with relative standard deviation (RSD) less than 11%. The limit of quantification (LOQ) was 0.001 mg/kg in longan and pulp. The half-lives were 3.3-4.2 days. The terminal residues in whole longan were <0.001-0.025 mg/kg applied two and three times at two levels of dosage with PHIs of 10, 14, and 21 days. The residues in whole longan had a higher quantity than those in the pulp, and the terminal residues of pulp were all lower than LOQ (0.001 mg/kg). The chronic risk of emamectin benzoate was not negligible to humans depending on ADI% value, which was higher than 1; and the acute risk was acceptable to the consumer. This study could provide guidance for the safe use of emamectin benzoate in longan and serve as a reference for the establishment of maximum residue limits (MRLs) in China.

Methacrylate bonded covalent organic framework monolithic column online coupling with high-performance liquid chromatography for analysis of trace estrogens in food

Covalent organic frameworks (COFs) are a burgeoning class of crystalline porous materials with unique properties and have been considered as a promising functional extraction medium in sample pretreatment. In this study, a new methacrylate-bonded COF (TpTh-MA) was well designed and synthesized via the aldehyde-amine condensation reaction, and the TpTh-MA was incorporated into poly (ethylene dimethacrylate) porous monolith by a facile polymerization reaction inside capillary to prepare a novel TpTh-MA monolithic column. The fabricated TpTh-MA monolithic column was characterized with scanning electron microscope, Fourier transform infrared spectrometer, X-ray diffraction, and N2 adsorption–desorption experiments. Then, the homogeneous porous structure, good permeability and high mechanical stability of TpTh-MA monolithic column was used as separation and enrichment media of capillary microextraction, which was coupled with high-performance liquid chromatography fluorescence detection for online enrichment and analysis of trace estrogens. The main experimental parameters influencing the extraction efficiency were systematically investigated. The adsorption mechanism for three estrogens was also explored and discussed based on hydrophobic effect, π-π affinity and hydrogen bonding interaction, which contributed to its strong recognition affinity to target compounds. The enrichment factors of the TpTh-MA monolithic column micro extraction method for the three estrogens were 107–114, indicating a significant preconcentration ability. Under optimal conditions, a new online analysis method was developed and exhibited good sensitivity and wide linearity range of 0.25–100.0 µg·L-1 with a coefficient of determination (R2) higher than 0.9990 and a low limit of detection with 0.05–0.07 µg·L-1. The method was successfully applied for online analysis of three estrogens of milk and shrimp samples and the recoveries obtained from spiking experiments were in range of 81.4–113% and 77.9–111%, with the relative standard deviations of 2.6–7.9% and 2.1–8.3% (n = 5), respectively. The results revealed the great potential for the application of the COFs-bonded monolithic column in the field of sample pretreatment.