A novel triple-signal biosensor based on ZrFe-MOF@PtNPs for ultrasensitive aflatoxins detection.

The development of more sensitive, stable, and portable biosensors is crucial for meeting the growing demands of diverse and complex detection environments. MOF-based nanozymes have emerged as excellent optical reporters, making them ideal signal donors for constructing multi-signal lateral flow immunoassays (LFIA). In this study, a ZrFe-MOF@PtNPs nanocomposite was synthesized by uniformly depositing platinum nanoparticles (PtNPs) onto the surface of ZrFe-MOFs using an impregnation-reduction method. The ZrFe-MOF@PtNPs exhibited broad absorption spectra, excellent peroxidase-like activity (SA = 21.77 U/mg), high solvent stability, and efficient antibody binding capability. A portable LFIA platform was developed based on ZrFe-MOF@PtNPs and a smartphone for the targeted detection of carcinogenic aflatoxins. This method enabled the readout of colorimetric, fluorescent, and catalytic signals, significantly enhancing detection sensitivity, ensuring result accuracy, and expanding the dynamic detection range. For aflatoxin M1, the calculation of the detection limit of the three signal modes reached as low as 0.0062 ng/mL, which is two orders of magnitude more sensitive than AuNPs-LFIA (0.1839 ng/mL). This study provides effective guidance for multifunctional modification of MOFs and serves as a reference for the application of MOF-based nanozymes in point-of-care biosensors.

Label-free direct detection of melamine using functionalized gold nanoparticles-based dual-fluorescence colorimetric nanoswitch sensing platform.

Developing a simple, economical, sensitive, and selective method for label-free direct detection analytes is attractive, especially the strategies that could achieve signal amplification without complicated operations. Herein, a dual-fluorescence colorimetric nanoswitch sensing platform for label-free direct melamine (MEL) detection was established. We first explored the relationship between MEL-induced aggregation of gold nanoparticles (AuNPs) and size and determined the optimal size to be 37 nm. Using surfactant Triton X-100 to modify AuNPs and clarify possible interaction mechanisms to improve detection performance. The dynamic changes of surface plasmon resonance absorption peaks in the dispersed and aggregated states of AuNPs were skillfully utilized to match the emission of multicolor gold nanoclusters to trigger the multi-inner filter effect. Accompanied by the addition of MEL-induced AuNPs to change from dispersed to aggregated state, the fluorescence of green-emitting and red-emitting gradually turned on and turned off, respectively. The fluorescence turn-on mode detection limit was 10 times higher than the colorimetric method and as low as 5.5 ng/mL; the detection took only 10 min. The sensor detected MEL in spiked milk samples with a good recovery in the range of 81.2-111.0 % with a coefficient of variation less than 11.4 % and achieved a good correlation with commercial kits. The proposed sensor integrates numerous merits of label-free, multi-signal readout, self-calibration, simple operations, and economical, which provides a promising tool for convenient on-site detection of MEL.

Four birds with one stone: Aggregation-induced emission-type zeolitic imidazolate framework-8 based bionic nanoreactor for portable detection of olaquindox in environmental water and swine urine by smartphone.

The abuse of olaquindox (OLA) as both an antimicrobial agent and a growth promoter poses significant threats to the environment and human health. While nanoreactors have proven effective in hazard detection, their widespread adoption has been hindered by tedious chemical processes and limited functionality. In this study, we introduce a novel green self-assembly strategy utilizing invertase, horseradish peroxidase, antibodies, and gold nanoclusters to form an aggregation-induced emission-type zeolitic imidazolate framework-8 nanoreactor. The results demonstrate that the lateral flow immunoassay not only allows for qualitative naked eye detection but also enables optical analysis through the fluorescence generated by aggregated gold nanoclusters and enzyme-catalyzed enhancement of visible colorimetric signals. To accommodate more detection scenarios, the photothermal effects and redox reactions of the nanoreactor can fulfill the requirements of thermal sensing and electrochemical analysis for smartphone applications. Remarkably, the proposed approach achieves a detection limit 17 times lower than conventional methods. Besides, the maximum linear range spans from 0.25 to 5 µg/L with high specificity, and the recovery is 85.2-112.9% in environmental water and swine urine. The application of this high-performance nanoreactor opens up avenues for the construction of multifunctional biosensors with great potential in monitoring hazardous materials.

Highly efficient dual-mode detection of AFB1 based on the inner filter effect: Donor-acceptor selection and application.

BACKGROUND: The utilization of inner filter effect (IFE) brings more opportunities for construction of fluorescence immunoassays but remains a great challenge, especially how to select best donor in the face of extensive fluorescent nanomaterials. Aflatoxin B1 possesses high toxicity among mycotoxins and is frequently found in agricultural products that may significantly threaten to human health. Therefore, with the help of signal transduction mechanism of IFE to develop a convenient and sensitive approach for AFB1 detection is of great significance in ensuring food safety. RESULTS: Herein, the classical alkaline phosphatase (ALP) catalyzes hydrolysis of p-nitrophenylphosphate to produce p-nitrophenol (PNP) was employed as a model reaction, which intends to explore tunable multicolor fluorescence of gold nanoclusters (AuNCs) for matching PNP to maximize IFE efficiency. The luminescent green-emitting AuNCs were selected as an optimal donor in terms of excellent spectral overlap, high photoluminescence, and adequate system adaptability, thus achieving a 22-fold increase in sensitivity improvement compared to colorimetric method for ALP detection. The fluorescence quenching mechanism between PNP and AuNCs was validated as IFE by studying ultraviolet absorption, zeta potentials and fluorescence lifetime. In light of this, we integrated a highly specific antibody-antigen recognition system, efficient enzymatic reaction and excellent optical characteristics of AuNCs to develop dual-mode immunoassay for AFB1 monitoring. The sensitivity of fluorometric immunoassay was lower to 0.06 ng/mL, which obtained a 3.5-fold improvement compared to "gold standard" ELISA. Their practicability and applicability were confirmed in the tap water, corn, wheat and peanuts samples. SIGNIFICANCE: This work provides an easy-to-understand screening procedure to select optimal donor-acceptor pairs in IFE analysis. Furthermore, we expect that integration of IFE-based signal conversion strategy into mature immunoassay not only extends the signal types, simplifies signal amplification steps, and reduces the false-positive/false-negative rates, but also provides a simple, convenient, and versatile strategy for monitoring of trace other contaminants.

Fe3O4-Incorporated Metal-Organic Framework for Chemo/Ferroptosis Synergistic Anti-Tumor via the Enhanced Chemodynamic Therapy.

Metal-organic framework (MOF)-based drug delivery nanomaterials for cancer therapy have attracted increasing attention in recent years. Here, an enhanced chemodynamic anti-tumor therapy strategy by promoting the Fenton reaction by using core-shell zeolitic imidazolate framework-8 (ZIF-8)@Fe3O4 as a therapeutic platform is proposed. Carboxymethyl cellulose (CMC) is used as a stabilizer of Fe3O4, which is then decorated on the surface of ZIF-8 via the electrostatic interaction and serves as an efficient Fenton reaction trigger. Meanwhile, the pH-responsive ZIF-8 scaffold acts as a container to encapsulate the chemotherapeutic drug doxorubicin (DOX). The obtained DOX-ZIF-8@Fe3O4/CMC (DZFC) nanoparticles concomitantly accelerate DOX release and generate more hydroxyl radicals by targeting the lysosomes in cancer cells. In vitro and in vivo studies verify that the DZFC nanoparticles trigger glutathione peroxidase 4 (GPX4)-dependent ferroptosis via the activation of the c-Jun N-terminal kinases (JNK) signaling pathway, following to achieve the chemo/ferroptosis synergistic anti-tumor efficacy. No marked toxic effects are detected during DZFC treatment in a tumor-bearing mouse model. This composite nanoparticle remarkably suppresses the tumor growth with minimized systemic toxicity, opening new horizons for the next generation of theragnostic nanomedicines.

Novel Label-Free Nanocrystalline Gold Interdigitated Microelectrode Immunosensor for the Rapid and Ultrasensitive Detection of SARS-CoV-2.

Waves of COVID-19 outbreaks have dragged down the global economy and endangered human life. There is an urgent need for timeliness and sensitive SARS-CoV-2 detection techniques to complement the existing PCR assay. Herein, the controllable growth of gold crystalline grains was achieved by applying the reverse current during pulse electrochemical deposition (PED) interval. The proposed method validates the effects of pulse reverse current (PRC) on the atomic arrangement, crystal structures, orientations, and film characteristics in Au PED. The gap between the gold grains on the surface of the nanocrystalline gold interdigitated microelectrodes (NG-IDME) fabricated by the PED+PRC process matches the size of the antiviral antibody. Immunosensors are prepared by binding a large number of antiviral antibodies on the surface of NG-IDME. The NG-IDME immunosensor has a high specific capture ability for SARS-CoV-2 nucleocapsid protein (SARS-CoV-2/N-Pro) and completes ultrasensitive and quantification of SARS-CoV-2/N-Pro in humans and pets within 5 min (the LOQ as low as 75 fg/mL). The specificity, accuracy, stability, and actual blind sample tests show that the NG-IDME immunosensor is suitable for the detection of SARS-CoV-2 in humans and animals. This approach assists in monitoring the transmission of SARS-CoV-2-infected animals to humans.

MC-PRPA-HLFIA Cascade Detection System for Point-of-Care Testing Pan-Drug-Resistant Genes in Urinary Tract Infection Samples.

Recently, urinary tract infection (UTI) triggered by bacteria carrying pan-drug-resistant genes, including carbapenem resistance gene blaNDM and blaKPC, colistin resistance gene mcr-1, and tet(X) for tigecycline resistance, have been reported, posing a serious challenge to the treatment of clinical UTI. Therefore, point-of-care (POC) detection of these genes in UTI samples without the need for pre-culturing is urgently needed. Based on PEG 200-enhanced recombinase polymerase amplification (RPA) and a refined Chelex-100 lysis method with HRP-catalyzed lateral flow immunoassay (LFIA), we developed an MCL-PRPA-HLFIA cascade assay system for detecting these genes in UTI samples. The refined Chelex-100 lysis method extracts target DNA from UTI samples in 20 min without high-speed centrifugation or pre-incubation of urine samples. Following optimization, the cascade detection system achieved an LOD of 102 CFU/mL with satisfactory specificity and could detect these genes in both simulated and actual UTI samples. It takes less than an hour to complete the process without the use of high-speed centrifuges or other specialized equipment, such as PCR amplifiers. The MCL-PRPA-HLFIA cascade assay system provides new ideas for the construction of rapid detection methods for pan-drug-resistant genes in clinical UTI samples and provides the necessary medication guidance for UTI treatment.

The effects of UiO-66 ultrafine particles on the rapid detection of sulfonamides in milk: Adsorption performance and mechanism.

Nanoscale MOFs particles possess both excellent adsorption and dispersion properties. In this study, ultrafine particles UiO-66 (UP/UiO-66) with a particle size below 50 nm were synthesised by a template-controlled method. UP/UiO-66 was able to achieve a maximum adsorption capacity of 139.64 mg/g for 5 methoxylated sulfonamides. Adsorption studies showed that UP/UiO-66 adsorption of sulfonamides can be classified as a pseudo-secondary kinetic adsorption model for single molecular layer adsorption. ELISA (validated by Raman and molecular docking) showed that the sulfonamide molecule was still immunoreactive with antibodies after adsorption by UP/UiO-66. In 15 min, UP/UiO-66 could be used directly in the ELISA test for sulfonamides in milk without elution and separation. The LOQ (IC20) of UP/UiO-66-ELISA for sulfonamides in milk was 0.21-2.05 ng/mL. The ultrafine particle strategy of UiO-66 is expected to be applied to other MOFs and used as a general pretreatment material for residue monitoring in complex matrices.

Aptasensor-based assay for dual-readout determination of aflatoxin B1 in corn and wheat via an electrostatic force-mediated FRET strategy.

A rapid and sensitive aptasensor was established for the dual-readout determination of aflatoxin B1 (AFB1) utilizing an electrostatically mediated fluorescence resonance energy transfer (FRET) signal amplification strategy. In the presence of AFB1, the aptamer preferentially bound to AFB1, resulting in the aggregation of bare gold nanoparticles (AuNPs) induced by NaCl, accompanied by a change of AuNP solution from wine-red to purple. This color change was used for colorimetric channel analysis. Then, the positively charged quantum dots were introduced into reaction system and interacted with negatively charged AuNPs, which successfully converted the color signal into a more sensitive fluorescence signal through FRET. The fluorescence quenching efficiency decreased with increasing concentrations of AFB1, and the fluorescence of aptasensor gradually recovered. The variation of fluorescence intensity was employed for fluorometric channel analysis. Under the optimal conditions, the color and fluorescence signals exhibited excellent response to AFB1 concentration within the ranges 10-320 ng·mL-1 and 3-320 ng·mL-1, respectively, and the limit of detection was as low as 7.32 ng·mL-1 and 1.48 ng·mL-1, respectively. The proposed aptasensor exhibited favorable selectivity, good recovery (85.3-113.4% in spiked corn and wheat samples), stable reproducibility (RSD<13.3%), and satisfactory correlation with commercial kits (R2=0.998). The aptasensor developed integrates advantages of modification-free, dual-readout, self-calibration, easy operation, and cost-effectiveness, while providing a simple and universal strategy for rapid and sensitive detection of mycotoxins in foodstuffs.

Rapid On-Site Detection of Extensively Drug-Resistant Genes in Enterobacteriaceae via Enhanced Recombinase Polymerase Amplification and Lateral Flow Biosensor.

The widespread emergence of transferable extensively drug-resistant (XDR) genes, including blaNDM and blaKPC for carbapenem resistance, mcr-1 for colistin resistance, and tet(X4) and tet(X6) for tigecycline resistance, in Enterobacteriaceae poses a major threat to public health. Thus, rapid on-site detection of these XDR genes is urgently needed. We developed a cascade system with a unitary polyethylene glycol (PEG) 200-enhanced recombinase polymerase amplification (RPA) as the core, combined with a modified Chelex-100 lysis method and a horseradish peroxidase (HRP)-catalyzed lateral flow immunoassay (LFIA) biosensor, to accurately detect these genes in Enterobacteriaceae. The conventional Chelex-100 lysis method was modified to allow in situ extraction of bacterial DNA in 20 min without requiring bulky high-speed centrifuges. Using PEG 200 increased the amplification efficiency of the RPA by 13%, and the HRP-catalyzed LFIA biosensor intensified the colorimetric signal of the test line. Following optimization, the sensitivity of the cascade system was <10 copies/µL with satisfactory specificity, allowing for highly sensitive detection of these XDR genes in Enterobacteriaceae. The complete detection procedure can be completed in less than 1 h without using large-scale instruments. This assay is conducive to rapid on-site visual detection of these XDR genes in Enterobacteriaceae in practical applications, thus providing better technical support for clinical surveillance of these genes and better treatment of XDR pathogens. IMPORTANCE Carbapenem, colistin, and tigecycline are considered the last resorts for treating severe bacterial infections caused by extensively drug-resistant (XDR) pathogens. A major threat to public health is the emergence and prevalence of transferable XDR genes in Enterobacteriaceae, such as blaNDM and blaKPC for carbapenem resistance, mcr-1 for colistin resistance, and tet(X4) and tet(X6) for tigecycline resistance. Therefore, it is imperative to develop rapid on-site methods to detect these XDR genes. In this study, we constructed a cascade system for detecting these genes based on PEG 200-enhanced recombinase polymerase amplification combined with a modified Chelex-100 lysis method and HRP-catalyzed lateral flow immunoassay. The current method is capable of detecting the above-mentioned XDR genes in situ with satisfactory specificity and sensitivity, which could provide technical support for the surveillance of these genes and provide medication recommendations for the treatment of relevant clinical infections.

Targeted preparation and recognition mechanism of broad-spectrum antibody specific to Aconitum alkaloids based on molecular modeling and its application in immunoassay.

Ester-type Aconitum alkaloids (AAs), the main medicinal ingredients of Aconitum L. herbs, could cause brain and heart damage in humans and animals and have raised concerns worldwide. In the present study, we aimed to produce a high-performance and broad-spectrum antibody and establish an immunoassay method of ester-type AAs, 3-succinyl aconitine (ACO-HS) was selected as an optimal hapten from five designed haptens comparing the similarity of stereo structure, electronic distribution, and physicochemical properties using the computer-aided molecular modeling technology. The monoclonal antibody (mAb) 1A9 exhibited broad-spectrum recognition specificity of 15 ester-type AAs was obtained and had a high sensitivity with the binding affinity (half-maximum inhibition concentration, IC50) of 0.73-130.36 µg L-1. Through molecular docking, it was found that mAb 1A9 and ester-type AAs showed a semi-enveloped structure through hydrogen bonds and hydrophobicity interaction. The amino acid residues that responsible for recognition were ARG107, GLU55, PRO113, VAL36, and SER64, and the critical structures to be recognized of AAs were acetyl group, benzoyl group, and N-linked carbon chains. The developed indirect competitive enzyme-linked immunosorbent assay (icELISA) based on mAb 1A9 allowed a sensitive determination of 15 ester-type AAs with the limit of detection (LOD) of 0.21-43.72 µg L-1, and it was suitable for the analysis of ester-type AAs in various Aconitum L. samples. These results provided an effective strategy for the preparation of targeted broad-spectrum antibodies of small molecules and proposed an icELISA method available for rapid, sensitive, and high-throughput detection of toxic ester-type AAs in Aconitum L. herbs.

Magnetic Fluorescent Quantum Dots Nanocomposites in Food Contaminants Analysis: Current Challenges and Opportunities.

The presence of food contaminants can cause foodborne illnesses, posing a severe threat to human health. Therefore, a rapid, sensitive, and convenient method for monitoring food contaminants is eagerly needed. The complex matrix interferences of food samples and poor performance of existing sensing probes bring significant challenges to improving detection performances. Nanocomposites with multifunctional features provide a solution to these problems. The combination of the superior characteristics of magnetic nanoparticles (MNPs) and quantum dots (QDs) to fabricate magnetic fluorescent quantum dots (MNPs@QDs) nanocomposites are regarded as an ideal multifunctional probe for food contaminants analysis. The high-efficiency pretreatment and rapid fluorescence detection are concurrently integrated into one sensing platform using MNPs@QDs nanocomposites. In this review, the contemporary synthetic strategies to fabricate MNPs@QDs, including hetero-crystalline growth, template embedding, layer-by-layer assembly, microemulsion technique, and one-pot method, are described in detail, and their advantages and limitations are discussed. The recent advances of MNPs@QDs nanocomposites in detecting metal ions, foodborne pathogens, toxins, pesticides, antibiotics, and illegal additives are comprehensively introduced from the perspectives of modes and detection performances. The review ends with current challenges and opportunities in practical applications and prospects in food contaminants analysis, aiming to promote the enthusiasm for multifunctional sensing platform research.

Development of a sensitive and rapid fluorescence polarization immunoassay for high throughput screening eight glucocorticoids in beef.

In this study, a sensitive, rapid, homogeneous, and high-throughput fluorescence polarization immunoassay (FPIA) for the rapid screening of eight glucocorticoids (GCs) in beef samples was successfully established. Two tracers including 5-aminofluorescein-labeled dexamethasone (5-AF-DMS) and fluorescein isothiocyanate-labeled dexamethasone (FITC-DMS) were studied to select appropriate antibody-tracer pairs using four previously produced broad-specific monoclonal antibodies. An optimal combination of the antibody 12D9 and the tracer FITC-DMS was selected. Under optimal detection conditions, the half inhibitory concentrations of dexamethasone (DMS), betamethasone (BMS), prednisolone (PNS), hydrocortisone (HCS), beclomethasone (BCMS), cortisone (CS), 6-α-methylprednisone (6-α-MPNS), fludrocortisone acetate (HFCS) were 1.00, 2.17, 3.49, 12.45, 1.20, 5.66, 6.85 and 3.45 ng/mL, respectively. The average recoveries of the proposed method in beef samples ranged from 77.3-91.7% with the coefficient of variation less than 12%. The developed FPIA was time-saving that could be completed within 10 min. The FPIA was applied to beef samples and showed a good correlation with liquid chromatography-tandem mass spectrometry (LC-MS/MS) (R2 = 0.9894). Thus, the proposed method provides a rapid, reliable, sensitive, and high-throughput screening tool for the simultaneous screening of eight GCs in beef, which shows great potential in the food safety analysis.

Dual-readout fluorescence quenching immunochromatographic test strips for highly sensitive simultaneous detection of chloramphenicol and amantadine based on gold nanoparticle-triggered photoluminescent nanoswitch control.

Herein, a novel fluorescence quenching immunochromatographic test strip (FQICTS) for simultaneous detection of chloramphenicol (CAP) and amantadine (AMD) was developed on the basis of inner filter effect (IFE), with the combination of gold nanoparticles (AuNPs) and highly luminescent green-emitting gold nanoclusters (AuNCs) as the IFE quencher/donor pair. The AuNPs could quench the excitation light and emission light of AuNCs and achieve a high IFE efficiency due to dual spectral overlapping. Under optimal conditions, the "turn-on" mode of the AuNCs-based dual-readout FQICTS showed good linearity for CAP detection in chicken samples from 0.05 ng/g to 10 ng/g, with a limit of detection (LOD) of 0.043 ng/g. The linear range of AMD is 0.5-50 ng/g, with LOD of 0.45 ng/g. The visual LODs of CAP and AMD in "turn-on" mode were 200 and 10 times lower than that in "turn-off" mode, respectively. The "turn-on" mode of FQICTS showed high recovery for detecting CAP (82.5-94.5%) and AMD (81.9-110.7%) spiked into chicken samples. The performance and practicability of the established method were verified with commercial enzyme-immunoassay kits, and good correlations were observed. Overall, the newly developed AuNCs-based dual-readout FQICTS is a promising on-site screening tool for rapid, high-sensitivity detection of multiple food contaminants in practical applications.

Adsorption and convenient ELISA detection of sulfamethazine in milk based on MOFs pretreatment.

Metal-organic frameworks (MOFs) has excellent adsorption performance, herein, three kinds of common MOFs were used for the adsorption of sulfamethazine (SM2) in milk, then enzyme-linked immunosorbent assay (MOF-ELISA) was established. Firstly, NH2-UiO-66, NH2-MIL-101, and ZIF-8 were successfully prepared and their adsorption characteristics for SM2 were investigated. The kinetic models of the three MOFs were more in line with the pseudo-second-order adsorption kinetics, and the saturated adsorption capacity of NH2-UiO-66, NH2-MIL-101, and ZIF-8 for SM2 at 298 K were 139.64, 29.98, and 36.5 mg/g, respectively. Using three different MOFs as adsorbents, the pretreatment of milk samples could be completed within 1 h, the half inhibitory concentrations (IC50) of MOF-ELISA were 1.26, 1.86 and 2.74 ng/mL, the limit of detections (LOD) were 0.05, 0.12, and 0.19 ng/mL and the recovery rate were from 82.30% to 105.62% with the intra-day coefficient of variations (CVs) below 5.81% and inter-day CVs below 7.21%. Detection results showed good correlations with LC-MS/MS (R2 > 0.99), indicated that MOFs could effectively eliminate the interference of sample matrix, and has the potential to become a general pretreatment method for the detection of various matrices residues in food safety monitoring.

Toxic Prediction of Pyrrolizidine Alkaloids and Structure-Dependent Induction of Apoptosis in HepaRG Cells.

Pyrrolizidine alkaloids (PAs) are common phytotoxins and could cause liver genotoxicity/carcinogenicity following metabolic activation. However, the toxicity of different structures remains unclear due to the wide variety of PAs. In this study, the absorption, distribution, metabolism, excretion, and toxicity (ADMET) of 40 PAs were analyzed, and their toxicity was predicted by Komputer Assisted Technology (TOPKAT) using Discovery Studio software. The in silico results showed that all PAs except retronecine had good intestinal absorption, and all PAs were predicted to have different toxicity ranges. To verify the predictive results, 4 PAs were selected to investigate cell injury and possible mechanisms of the differentiation in HepaRG cells, including retronecine type of twelve-membered cyclic diester (retrorsine), eleven-membered cyclic diester (monocrotaline), noncyclic diester (retronecine), and platynecine type (platyphylline). After 24 h exposure, retronecine-type PAs exhibited concentration-dependent cytotoxicity. The high-content screening assay showed that cell oxidative stress, mitochondrial damage, endoplasmic reticulum stress, and the concentration of calcium ions increased, and neutral lipid metabolism was changed notably in HepaRG cells. Induced apoptosis by PAs was indicated by cell cycle arrest in the G2/M phase, disrupting the mitochondrial membrane potential. Overall, our study revealed structure-dependent cytotoxicity and apoptosis after PA exposure, suggesting that the prediction results of in silico have certain reference values for compound toxicity. A 1,2-membered cyclic diester seems to be a more potent apoptosis inducer than other PAs.

Indirect competitive enzyme-linked immunosorbent assay based on a broad-spectrum monoclonal antibody for tropane alkaloids detection in pig urine, pork and cereal flours.

In this study, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) based on a broad-spectrum monoclonal antibody for tropane alkaloids (TAs) was established for the rapid screening of atropine, scopolamine, homatropine, apoatropine, anisodamine, anisodine and L-hyoscyamine residues in pig urine, pork and cereal flour samples through a simple sample preparation procedure. The half inhibitory concentrations of atropine, homatropine, L-hyoscyamine, apoatropine, scopolamine, anisodamine and anisodine were 0.05, 0.07, 0.14, 0.14, 0.24, 5.30 and 10.15 ng mL-1, respectivelyThe detection and quantitative limits of this method for TAs in samples were 0.18-73.18 and 0.44-74.77 µg kg-1. The spiked recoveries ranged from 69.88% to 147.93%, and the coefficient of variations were less than 14%. Good correlation (R2 = 0.9929) between the results of the ic-ELISA and the high performance liquid chromatography-tandem mass spectrometry support the reliability of the developed ic-ELISA method.

Pharmacokinetics and bioavailability of tildipirosin in rabbits following single-dose intravenous and intramuscular administration.

The objective of this study was to determine the pharmacokinetics of tildipirosin in rabbits after a single intravenous (i.v.) and intramuscular (i.m.) injection at a dose of 4 mg/kg. Twelve white New Zealand rabbits were assigned to a randomized, parallel trial design. Blood samples were collected prior to administration and up to 14 days postadministration. Plasma concentrations of tildipirosin were quantified using a validated ultra-high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method. The pharmacokinetic parameters were calculated using a noncompartmental model in WinNonlin 5.2 software. Following i.v. and i.m. administration, the elimination half-life (T1/2λ ) was 81.17 ± 9.28 and 96.68 ± 15.37 hr, respectively, and the mean residence time (MRTlast ) was 65.44 ± 10.89 and 67.06 ± 10.49 hr, respectively. After i.v. injection, the plasma clearance rate (Cl) and volume of distribution at steady state (Vdss ) were 0.28 ± 0.10 L kg-1  h-1 and 17.78 ± 5.15 L/kg, respectively. The maximum plasma concentration (Cmax ) and time to reach maximum plasma concentration (Tmax ) after i.m. administration were 836.2 ± 117.9 ng/ml and 0.33 ± 0.17 hr, respectively. The absolute bioavailability of i.m. administration was 105.4%. Tildipirosin shows favorable pharmacokinetic characteristics in rabbits, with fast absorption, extensive distribution, and high bioavailability. These findings suggest that tildipirosin might be a potential drug for the prevention and treatment of respiratory diseases in rabbits.

Tilmicosin enteric granules and premix to pigs: Antimicrobial susceptibility testing and comparative pharmacokinetics.

The purpose of this study was to compare the pharmacokinetics and relative bioavailability of tilmicosin enteric granules and premix after oral administration at a dose of 40 mg/kg in pigs. Three kinds of different respiratory pathogens were selected for determination of minimal inhibitory concentration (MIC) to tilmicosin. Eight healthy pigs were assigned to a two-period, randomized crossover design. A modified rapid, sensitive HPLC method was used for determining the concentrations of tilmicosin in plasma. Pharmacokinetic parameters were calculated by using WinNonlin 5.2 software. The MIC90 of tilmicosin against Haemophilus parasuis, Actinbacillus pleuropneumoniae, and Pasteurella multocida were all 8 µg/ml. These results indicated that these common pig respiratory bacteria are sensitive to tilmicosin. The main parameters of time to reach maximum plasma concentration (Tmax ), elimination half-life (t1/2ß ), mean residence time (MRT), and apparent volume of distribution (VF ) were 2.03 ± 0.37 hr, 29.31 ± 5.56 hr, 25.22 ± 2.57 hr, 4.06 ± 1.04 L/kg, and 3.05 ± 0.08 hr, 17.06 ± 1.77 hr, 15.55 ± 1.37 hr, 2.95 ± 0.62 L/kg after the orally administrated tilmicosin enteric granules and premix. The relative bioavailability of tilmicosin enteric granules to premix was 114.97 ± 7.19%, according to the AUC0-t values. These results demonstrated that tilmicosin enteric granules produced faster tilmicosin absorption, slower elimination, larger tissue distribution, and higher bioavailability compared to the tilmicosin premix. The present study results manifest that tilmicosin enteric granules can be used as a therapeutic alternative to premix in clinical treatment.

Bioequivalence evaluation of two 5% ceftiofur hydrochloride sterile suspension in pigs.

The purpose of this study was to evaluate the bioequivalence of 5% ceftiofur hydrochloride sterile suspension in two formulations, a test formulation (Saifukang 5% CEF, Hvsen) and a reference formulation (Excenel®RTU 5% CEF, Pfizer). Twenty-four healthy pigs were assigned to a two-period, two-treatment crossover parallel trial, and both formulations were administered at a single intramuscular dose of 5 mg/kg weight, with a 7-day washout period. Blood samples were collected consecutively for up to 144 hr after administration. The concentrations of ceftiofur- and desfuroylceftiofur-related metabolites in the plasma were determined by high-performance liquid chromatography. In addition, the major pharmacokinetic parameters (Cmax, AUC0-t and AUC0-∞) were computed and compared via analysis of variance, with 90% confidence intervals. Bioequivalence evaluation of Tmax was statistically analyzed with the nonparametric test. The comparison values between test and reference formulation for AUC0-t, AUC0-∞, Cmax, and Tmax were 376.7 ± 75.3 µg·hr/ml, 390.5 ± 78.6 µg·hr/ml, 385.9 ± 79.2 µg·hr/ml, 402.7 ± 80.4 µg·hr/ml, 34.6 ± 5.5 µg/ml, 36.1 ± 6.2 µg/ml, 1.27 ± 0.18 hr, and 1.26 ± 0.21 hr, respectively, and we observed no significant differences between the two formulations. The 90% CI values were within the recommended range of 80-125% (P>0.05), and the relative bioavailability of the test product was 96.47 ± 10.92% according to AUC0-t values. Based on our results, the two formulations exhibit comparable pharmacokinetic profiles, and the test product is bioequivalent to the reference formulation.