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Constipation and depression are tightly related and often co-occur and coexist in clinic. Yet, the relationships and the underlying mechanisms are still unclear. Fecal metabolomics and network pharmacology were, for the first time, applied to investigate the potential correlations from multiple levels including classic behaviors, metabolomics, and gene targets. The behavioral indicators were analyzed, providing behavioral correlations at a macrolevel. Besides, fecal samples were analyzed by nuclear magnetic resonance spectroscopy to screen the shared and the unique metabolites and pathways, revealing correlations from a metabolic perspective. Finally, the disease targets and the functional pathways were obtained via network pharmacology, demonstrating correlations at the molecular level. The correlations between constipation and depression were demonstrated and supported by four-level evidence: (1) general behaviors, (2) gastrointestinal functions, (3) fecal metabolites and pathways, and (4) common gene targets and functional pathways. Especially, the correlations of behaviors and common metabolites showed that metabolites, including choline, betaine, and glycine, were significantly associated with constipation and depression. Besides, inflammation and immune abnormalities and energy metabolism were significantly involved in the mechanisms. The current findings prove the correlations between constipation and depression, and provide a basis for deeply understanding the comorbidities of constipation and depression.

miR-205-5p plays a vital role in the inflammation of allergic rhinitis (AR). The study is designed to investigate the effects and mechanism of miR-205-5p in AR in vivo and in vitro. An OVA-induced mice model and anti-DNP IgE-induced RBL-2H3 cell model were established. The pathological alterations in the nasal mucosa were evaluated by hematoxylin-eosin (HE) staining. IgE and histamine levels were detected by corresponding kits and the expressions of PEBP1, High mobility group box-1 (HMGB1) and Toll-like receptor 4 (TLR4) were detected by western blot. The association of miR-205-5p and PEBP1 was determined by dual-luciferase reported assay. β-hexosaminidase activity was to evaluate the degranulation of RBL-2H3 cell. The pathological injury of nasal mucosa was significantly improved by miR-205-5p inhibition compared to AR mice. Following the treatment of miR-205-5p inhibitor, the levels of helper T cell (Th1) cytokines, interleukin (IL)-2 and interferon-γ (IFN-γ) were increased, while the levels of Th2 cytokines, IL-4 and IL-13, as well as the levels of IgE and histamine were markedly decreased in AR mice. We further found that miR-205-5P inhibition induced increased expression of PEBP1 and decreased expressions of HMGB1and TLR4. In vitro, miR-205-5P was verified to bind to PEBP1. PEBP1 silencing led to the reverse of miR-205-5p effects on decreasing the levels of β-hexosaminidase activity and histamine, as well as the expressions of HMGB1 and TLR4 on anti-DNP IgE-induced RBL-2H3 cells. Our results indicate that miR-205-5P inhibition may ameliorate pathological injury via PEBP1. MiR-205-5P/ PEBP1 could be potential drug targets in AR.

This study aimed to investigate the presence of eight virulence genes (ace, asa1, esp, efaA, gelE, cylA, agg, fsr) in Enterococcus from a variety of animals and to explore the drug resistance and pathogenicity. This could provide a theoretical basis for clinical treatment of Enterococcus infections. Anal swabs from pigs, chickens, cattle, and dogs in farms and pet hospitals were collected for Enterococcus isolation and identification. Eight virulence genes were detected (PCR method), and drug resistance was assessed (drug-sensitive paper method). The strains containing different virulence genes were then divided into EV1, EV2, and EV3 groups. The LD

Aging is a complex physiological process associated with degenerative disorders of metabolism and intestinal flora. Constipation is an age-dependent gastrointestinal disease, which is involved in several diseases, such as cardiovascular and cerebrovascular diseases, colon cancer, etc. Gut microbiota is involved in both aging and constipation, which is probably through metabolism. Yet, the underlying mechanisms are still unclear. Our findings showed significant changes in the cecum microbiome compositions across aging and constipation, and the community composition is similar in both. A cluster analysis demonstrated that the key gut bacteria associated with aging and the key gut bacteria associated with constipation were clustered together. Function analysis explored the associations between both via specific gut bacteria's metabolism pathways, involving in 8 metabolic pathways. Associated networks showed that not only gut microbial metabolites are important signaling molecules associated with aging and constipation, but also the occurrence and the development of senescence and constipation are affected at multiple levels and by multiple factors. In this study, both macro and micro indexes, and traditional and modern indexes are combined to prove the important regulatory roles of intestinal microorganisms in aging and constipation, and systematically elaborated the fact that constipation and aging are mutual causation and mutual influences.

Background Intranasal corticosteroids are the most efficacious anti-inflammatory medications for allergic rhinitis (AR). However, the efficacy and safety of intranasal corticosteroids in children have not yet been subject to specific research in China. The aim of this study was to investigate the efficacy and safety of fluticasone furoate nasal spray (FFNS) in a Chinese pediatric population. Methods In this phase 4 randomized, double-blind, placebo-controlled, multicenter study, pediatric AR patients aged 2–12 years were randomized 1:1:1, receiving either FFNS 55 µg or 110 µg or placebo. Electronic diary cards were completed to record symptoms, rescue medication use, and treatment compliance. Anterior rhinoscopy and overall response to therapy were evaluated and recorded. Results Patients treated with FFNS at either dose experienced a significantly greater reduction in daily reflective total nasal symptom score compared with placebo. This was maintained in a younger subset of patients (2–6 years). Drug-related adverse events occurred in Conclusions This study demonstrates favorable efficacy and safety profiles for FFNS 55 µg or 110 µg in Chinese pediatric populations (2–12 years), supporting its use in clinical treatment for AR children, including younger children aged 2–6 years. Impact The aim of this study was to investigate the efficacy and safety of intranasal fluticasone furoate in Chinese pediatric allergic rhinitis. This research not only addresses the deficiency in efficacy and safety data for intranasal corticosteroids in very young patients (aged 2–6 years) worldwide but also demonstrates that fluticasone furoate nasal spray shows a favorable benefit/risk profile at different dose levels. Our data will be of interest to the broad readership of Pediatric Research and will positively contribute to the dialog regarding the treatment of allergic rhinitis in children aged 2–6 years.

Guilingji (GLJ), a traditional Chinese medicine, is of wide concern because of its remarkable antiaging effect with a long application history. It mainly consists of traditional Chinese herbs, that is, Ginseng radix et rhizoma rubra. This study focused on the anti-aging effects of GLJ on natural aging rats and its underlying mechanisms. Morris water maze was used to determine the learning and memory ability of rats. The levels of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), malondialdehyde (MDA), acetylcholine (ACh), and acetylcholinesterase (AChE) in serum were determined. Furthermore, a 1H-NMR-based serum metabolomics coupled with multivariate data analysis was used to identify potential biomarkers and corresponding metabolic pathways. The results showed that GLJ significantly improved the learning and memorial dysfunctions of natural aging rats. The mechanisms of the anti-aging and memory ameliorative effects of GLJ related to balancing oxidative stress, improving cholinergic system. Its specific mechanism of action may be through regulating pyruvate metabolism and arginine and proline metabolism.

Long noncoding RNA nuclear enriched abundant transcript 1 (lnc-NEAT1) and its target microRNA-125a (miR-125a) are reported to regulate immune and inflammation process in allergic rhinitis (AR). Hence, this study intended to investigate the correlation between lnc-NEAT1 and miR-125a expressions, as well as their clinical values in pediatric AR patients.Peripheral blood mononuclear cell samples from 80 pediatric AR patients, 40 disease controls (DCs), and 40 healthy controls (HCs) were collected to detect lnc-NEAT1 and miR-125a expressions by reverse transcription-quantitative polymerase chain reaction. For pediatric AR patients only, serum interferon-gamma (IFN-γ) and interleukin (IL)-10 were measured by enzyme linked immunosorbent assay; meanwhile, T helper (Th) 1 and Th2 cells in CD4Lnc-NEAT1 was overexpressed, while miR-125a downregulated in pediatric AR patients compared to DCs and HCs (all p 0.001). Moreover, lnc-NEAT1 expression negatively correlated with miR-125a expression in pediatric AR patients (p = 0.002), but not in DCs (p = 0.226) or HCs (p = 0.237). Furthermore, in pediatric AR patients, lnc-NEAT1 expression positively associated with TNSS (p 0.001), sneezing score (p = 0.006), and congestion score (p = 0.008); miR-125a expression was negatively related to TNSS (p 0.001), itching score (p = 0.040), and sneezing score (p = 0.005). Additionally, lnc-NEAT1 expression positively, while miR-125a expression negatively correlated with Th2 cells and IL-10 (all p 0.05), but they were not correlated with Th1 cells or IFN-γ in pediatric AR patients.Circulating lnc-NEAT1 and miR-125a are aberrantly expressed and linked with Th2 cells and symptom severity in pediatric allergic rhinitis.

Objectives: Although miR-653-5p has been validated to participate in the progression of multiple types of cancer, the functional role of exosomal miR-653-5p derived from Mesenchymal Stem Cells (MSCs) in Laryngeal Papilloma (LP) has still remained elusive. Hence, this study aimed to investigate the role of MSCs-derived exosomal miR-653-5p in LP. Methods: LP tissues (n = 15) and adjacent normal tissues (n = 10) were collected to examine the expression level of miR-653-5p. The expression level of miR-653-5p in LP cells and normal cells was also detected. Then, miR-653-5p was overexpressed or silenced to explore its effects on the proliferation, migration, invasion, and apoptosis of LP cells. Thereafter, the effects of exosomal miR-653-5p derived from MSCs on LP cell progression and the potential regulatory mechanism of miR-653-5p were assessed. Results: It was revealed that the expression level of miR-653-5p was downregulated in LP tissues and cells. In addition, miR-653-5p suppressed the proliferation, migration, invasion, and apoptosis of LP cells. Exosomes derived from MSCs played a suppressive role in LP development and mediated the transmission of miR-653-5p to LP cells. Further exploration identified Basic leucine Zipper and W2 domains 2 (BZW2) as the target of miR-653-5p. More importantly, the rescue experiments revealed that MSCs-secreted exosomal miR-653-5p efficiently inhibited the aggressive phenotypes of LP cells, which could be significantly reversed by BZW2 overexpression in LP cells. Conclusion: MSCs-derived exosomal miR-653-5p exerted inhibitory effects on LP progression through targeting BZW2, which provided a novel idea for the therapy of LP. Clinical Trial registration number: chictr-ior-17011021.

Three kinds of immunochromatographic assays (ICAs) are proposed for the highly sensitive and rapid determination of tylosin (TYL) and tilmicosin (TIM) in eggs based on colloidal gold (CG), latex microsphere (LM), and time-resolved fluorescent microsphere (TRFM). Three types of ICAs could tolerate the egg matrix via simple sample pretreatment and demonstrated high sensitivity for TYL and TIM with cut-off values of 6/6/3 μg/kg and 14/14/6 μg/kg, respectively. Furthermore, in a single-blind parallel study 20 egg samples were analyzed by the three developed ICAs and confirmed by LC-MS/MS. The results showed good consistency, and there were no false positive and false negative results in our three ICAs. Consequently, the proposed three ICAs offered rapid, highly sensitive, reliable, and selectable testing platforms for screening veterinary medicine or other small molecule contaminants.

Guilingji (GLJ), which has been used to treat male diseases in China for centuries, contains 28 Chinese herbs and was previously established as an effective treatment for male sexual dysfunction. However, its mechanism of action remains unclear.To explore the efficacy and mechanism of action of GLJ in improving senile sexual dysfunction (SSD) in aging rats.An aging rat model of SSD was induced by the subcutaneous injection of d-galactose (300 mg⋅kgAfter treatment with GLJ, the mount and ejaculation latency levels were increased in the treatment group than those in model group (P 0.05), moreover, the testicular morphology was improved. Gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) levels in rats were also improved significant (P 0.05) compared with those in the model group. Furthermore, the metabonomics results in the testicular and brain tissue showed that GLJ improved SSD by adjusting amino acid and lipid metabolism.This study integrated the complementary metabolic profiles of the target tissues. GLJ might affect SSD rats by regulating amino acid and lipid metabolism and may modulate sensitivity to the signaling pathway in the HPG axis. This study provides an essential basis for the broad clinical application of GLJ.

To explore the effect of the caregiver's knowledge, behaviour and attitude on the prevention of foreign bodies in children's respiratory tracts to provide a basis for publicity and education on foreign bodies in children's respiratory tracts.A case-control method was used to investigate the knowledge, behaviour and attitude of the caregivers of 900 paediatric patients at Hunan Children's Hospital from June 2017 to February 2019. Caregivers of 300 children with foreign bodies removed by rigid bronchoscopy were selected as the case group. Caregivers of 600 children with other conditions were selected as the control group. The knowledge, behaviour and attitude of the caregivers towards foreign bodies in children's respiratory tracts and the demographic characteristics of the two groups were analyzed and compared.The results showed significant differences for the children in terms of their gender, age and living conditions between the two groups (all p 0.05) and for the caregivers in terms of their age, educational level, relationship with the child and the number of children in the family (all p 0.05). There were statistically significant differences in the caregivers' knowledge of foreign bodies in the respiratory tract between the two groups (p 0.05). In addition, a significant difference was also found between the groups in the caregivers' understanding of the prevention of foreign bodies in the respiratory tract, such as whether they were aware of the specific first-aid measures needed to treat the condition (p 0.05).The caregiver's relationship to the child, their age, their level of education and the number of children in the family affected the occurrence of the condition, while the caregiver's knowledge, behaviour and attitude regarding foreign bodies in the respiratory tract also had some influence. However, in the future, national multiple-centre courses could be extremely useful in educating caregivers.

In this study, an enhancing immunochromatography assay (ICA) based on colloidal gold-decorated polydopamine (CG@PDA) was firstly developed for gentamicin in milk, muscle, liver and kidney simultaneously. CG@PDA was synthesized by one-pot method with better signal intensity, colloidal stability, and antibody coupling efficiency than traditional colloidal gold. The detection limit of the developed ICA was about 1.50 μg/kg for the four animal-derived food, which was up to 92-fold more sensitive than the reported ICA based on colloidal gold. The recovery rates were ranged from 86.0% to 114.0% with coefficient of variation between 1.6% and 13.1%. Parallel analysis of 40 samples by commercial ELISA kits was confirmed the reliability. Our research indicated that polydopamine decorated can chemically modify the surface of colloidal gold and can thus remarkably improve the detection performances of ICA.

Ethnopharmacological relevance The quality control of Traditional Chinese medicine (TCM) is a scientific problem and an industrial issue, which hampers the development of evidence based TCM. The concept of quality markers (Q-markers) is proposed and has been applied to the quality evaluation of TCM based on its clinical efficacy. However, more specific methods are needed to put this idea into practice. The standard decoction is a representative of decoction used in clinical practice and it can be used for the discovery of Q-markers related to the clinical efficacy of TCM. Aim of the study In this study, a systemic strategy was established to discover Q-markers related to the clinical efficacy of TCM Ephedrae Herba (EH), dried stem of Ephedra sinica Stapf. The different processed materials of EH have different clinical applications, though originating from the same medicinal herb. Materials and methods The standard decoction of each of the processed materials was prepared and a 1HNMR metabolomics approach and total polysaccharide analysis were used to identify potential Q-markers related to the different clinical applications of EH. Correlation analysis was made of the measured biological activity and the holistic chemical profile. Results The results showed that total polysaccharides and alkaloids were Q-markers for EH preparations. Conclusion This study demonstrates that the standard decoction is a reasonable research objective to explore chemical markers that correlate with the clinical efficacy of TCM.

Idiopathic normal pressure hydrocephalus (iNPH) is a neurological disorder that occurs in about 1% of individuals over age 60 and is characterized by enlarged cerebral ventricles, gait difficulty, incontinence, and cognitive decline. The cause and pathophysiology of iNPH are largely unknown. We performed whole exome sequencing of DNA obtained from 53 unrelated iNPH patients. Two recurrent heterozygous loss of function deletions in CWH43 were observed in 15% of iNPH patients and were significantly enriched 6.6‐fold and 2.7‐fold, respectively, when compared to the general population. Cwh43 modifies the lipid anchor of glycosylphosphatidylinositol‐anchored proteins. Mice heterozygous for CWH43 deletion appeared grossly normal but displayed hydrocephalus, gait and balance abnormalities, decreased numbers of ependymal cilia, and decreased localization of glycosylphosphatidylinositol‐anchored proteins to the apical surfaces of choroid plexus and ependymal cells. Our findings provide novel mechanistic insights into the origins of iNPH and demonstrate that it represents a distinct disease entity.

Traditional Chinese medicines (TCMs) have attracted more attentions in the treatment of depression. Xiaoyaosan (XYS), a classic anti-depression TCM prescription, contains eight herbs. However, the compatibility effects of XYS in modern pharmacology need to be investigated in depth. In this study, the chronic unpredictable mild stress (CUMS) depression-like model was constructed. Afterwards, XYS was divided into the Shugan and the Jianpi groups according to the research strategy ofefficacy groups. Meanwhile, a proton nuclear magnetic resonance spectrometry (1H NMR) based serum metabolomics was applied. XYS and its efficacy groups significantly regulated the abnormal levels of differential metabolites related to depression, but to different degrees. Metabolic profiling by orthogonal partial least squares discriminant analysis showed that XYS at high dose (XH) exhibited the strongest effects than other treatment groups. Ten metabolites related to depression were identified as differential metabolites. Besides, relative distance (Rd) was calculated to quantitatively evaluate the effects. We found that XH group had the highest Rd value. Moreover, among the five metabolic pathways of depression, XYS and Jianpi groups significantly regulated all pathways while Shugan group regulated four pathways. These findings lay a solid foundation for comprehensively and deeply understanding the compatibility effects of XYS against depression.

Ethnopharmacological relevance Xiaoyaosan (XYS), a classic description, has a history of thousands of years for treating depression through invigorating the liver and strengthening the spleen, which have been verified both clinically and experimentally. However, explanation of its underlying mechanisms remains a great challenge. Aim of the study The mechanisms of XYS in treating depression were investigated, with emphasis on the important biomarkers, targets and pathways. Materials and methods In this study, taking the targeted organ of depression, hippocampus, as the object, a combination of GC-MS based metabolomics and network pharmacology was established to illustrate the abnormality of metabolic characteristics of hippocampus of depression rats and to demonstrate the antidepressant mechanisms of XYS. Hippocampal metabolomics demonstrated potential metabolites involving in the antidepressant effects of XYS, as well as the corresponding metabolic pathways. Network pharmacology screened the potential ingredients and the targets of XYS against depression. Results Metabolomics revealed that XYS significantly regulated the abnormal levels of lactic acid, glycerol, glutamine, glutamic acid, hypoxanthine, myo-inositol and cholesterol, which involved in the D-glutamine and D-glutamate metabolism, arginine biosynthesis and alanine, aspartate and glutamate metabolism. Network pharmacology showed that XYS exhibited anti-depression effects through paeoniflorin, quercetin, licochalcone a, naringenin, β-sitosterol, formononetin and kaempferol acting on interleukin-6 (IL6), mitogen-activated protein kinase 1 (MAPK1), signal transducer and activator of transcription 3 (STAT3) and transcription factor AP-1 (JUN). Conclusion Based on hippocampal metabolomics and network pharmacology, this study proved that the actions of XYS in treating depression depend on multi-components, multi-targets and multi-pathways, the unique characteristics of TCMs.

Background Mast cell (MC)-mediated inflammation is essential for allergic rhinitis, and nuclear factor E2 related factor (NRF2) is found to inhibit inflammation. This study investigated whether NRF2 could inhibit MC inflammation and its molecular mechanisms concerning SIRT4. Methods Real-time quantitative PCR (RT-qPCR) and western blot were used to detect gene expression, and Elisa kit was used to detect the content of histamine and inflammatory cytokines in the medium of MCs, and Seahorse XF instrument was used to measure the mitochondrial metabolism of MCs. Knockdown SIRT4 and establish SIRT4 overexpression of HMC-1 cells to study the function of SIRT4. Results As an activator of NRF2, 4-Octyl Itaconate increases not only NRF2 expression but also increases SIRT4 expression. Although 4-Octyl Itaconate could reduce the histamine release and degranulation of MCs, which was induced by compound 48/80, SIRT4 knockdown decreased the inhibition of 4-Octyl Itaconate. Similarly,4-Octyl Itaconate inhibited the secretion of inflammatory cytokines (TNF-α, IL-1β, IL6, and IL-8) by MCs, which was induced by LPS, but SIRT4 knockdown decreases the inhibition of 4-Octyl Itaconate. Also, the up-regulation of SIRT4 significantly inhibited mitochondrial metabolism in MCs and inhibited SIRT1 and P-p65 protein expression after inducing by 100 ng/mL LPS for 1 hour. Conclusions NRF2 inhibits MC degranulation and MC-mediated inflammation by promoting SIRT4, and SIRT4 overexpression inhibits the mitochondrial metabolism of MCs.

A time-resolved fluorescent immunochromatographic assay (TRFICA) was successfully developed for the sensitive, simultaneous, and quantitative detection of aflatoxin B1 (AFB1) and zearalenone (ZEN) ...

Six aflatoxins (AFs; AF B1, B2, G1, G2, M1 and M2) and six zearalenone (ZEN) analogs (ZEN, zearalanone, α-zeralanol, β-zeralanol, α-zearalenol, and β-zearalenol) were simultaneously extracted from edible and medicinal herbs using a group-specific immunoaffinity column (IAC) and then identified by ultra-high-performance liquid chromatography tandem mass spectrometry (UPLC–MS/MS). The IAC was prepared by coupling N-hydroxysuccinimide-activated Sepharose 4B Fast Flow gel with two group-specific monoclonal antibodies. The column capacities to six AFs and six ZEN analogs ranged from 100.2 ng to 167.1 ng and from 59.5 ng to 244.4 ng, respectively. The IAC–UPLC–MS/MS method was developed and validated with three different matrices (Chinese yam [Dioscorea polystachya], Platycodon grandiflorum and coix seed [Semen Coicis]). Recoveries of twelve analytes from edible and medicinal herbs were in the range of 64.7%–112.1%, with relative standard deviations below 13.7%. The limits of quantification were in the range from 0.08 μg kg−1 to 0.2 μg kg−1. The method was proven to be sensitive and accurate, and suitable for the determination of real samples.

Antibodies are the most important reagents for the development of highly sensitive and specific immunoassays to quantify analytes of interest in food and environmental samples. While immunoglobulin G (IgG)-derived antibodies from rabbit and mouse are traditionally employed in immunoassays, recent findings suggest that chicken egg yolk antibody (immunoglobulin Y (IgY)) provides several advantages over mammalian IgG. However, limited studies to date have examined the possibility of replacing IgG with IgY in immunoassays. In the current investigation, the performance of chicken IgY and IgG derived from rabbit and mouse was systematically compared in terms of sensitivity, specificity, and matrix effect under parallel conditions with three typical assay formats, specifically, indirect competitive enzyme-linked immunosorbent assay (icELISA), fluorescence polarization immunoassay (FPIA), and colloidal gold immunochromatographic assay (GICA), for detection of sulfamethazine (SMZ) as the reference molecule. We evaluated and discussed the influence of different coating antigens, tracers, and physicochemical factors on the performance of IgY and IgG in the immunoassays. Under optimized conditions, the sensitivities of icELISA (IC50 values of 6.70, 4.76, and 1.66 ng mL−1 with recoveries of 86.1–131.8% and precision of

To increase information content per screen, a dual-wavelength, homologous and high-throughput fluorescence polarization immunoassay (DWFPIA) for simultaneous detection of total aflatoxins (AFs) and family zearalenones (ZENs) was developed. Aflatoxin B1 (AFB1) and zearalanone (ZAN) were labeled with different fluoresceins, and then combined with corresponding broad-specific antibodies to develop the DWFPIA. Under optimal conditions, the half maximal inhibitory concentrations of the FPIA in buffer were 2.68 μg/L and 4.08 μg/L for total four AFs (the weight ratio of AFB1: Aflatoxin B2: Aflatoxin G1: Aflatoxin G2 was 1: 1: 1: 1) and six ZEN analogs (the weight ratio of ZEN: ZAN: α-zearalenone: β-zearalenone: α-zearalanone: β-zearalanone was 1: 1: 1: 1: 1: 1) by mixing equal volumes (1: 1, v/v), respectively. The corresponding limit of detection values in maize flour samples were 4.98 μg/kg and 11.03 μg/kg, and the mean recoveries ranged from 78.6 to 103.6% with the coefficients of variation below 19.2%. The whole detection procedure, including sample preparation, took less than 30 min. Finally, the DWFPIA was applied to screen naturally contaminated maize flour samples, and the detection data were similar to those with HPLC-MS/MS.

Three immunizing haptens of bisphenol A (BPA), including two new haptens, were used to produce highly sensitive and specific polyclonal antibodies. The spacer arms of haptens for coupling to the protein carrier were located at different positions in BPA, and different length spacer arms were tested. Highly sensitive polyclonal antibodies were obtained and characterized using indirect competitive enzyme-linked immunosorbent assay (icELISA). Under optimized conditions, the half maximal inhibitory concentration (IC50) value of the best polyclonal antibody was 2.1 mg·L−1, based on coating heterogeneous antigens, and this optimal polyclonal antibody was highly sensitive toward BPA and displayed negligible cross-reactivity with bisphenol B and bisphenol E. A sensitive icELISA method utilizing the polyclonal antibody was developed for the determination of BPA in milk. In spiked samples (5, 10 and 20 mg·L−1), the recovery ranged from 80% to 102% with a coefficient of variation (CV) value below 15.8%. The limit of detection of icELISA was 1.95 mg·L−1. These results indicate that the icELISA method is suitable for the detection of BPA in milk.

The present study analyzed the prevalence and antimicrobial resistance of Salmonella along an integrated broiler chicken supply chain. A total of 172 Salmonella isolates were recovered from 1148 samples collected from four sample sources (breeder farms, broiler farms, abattoir, and retail markets), representing nine production stages. These Salmonella isolates were examined for antimicrobial susceptibility to 12 different antimicrobial agents using a disk diffusion assay. Among them, 168 were identified as six different serotypes of Salmonella enterica. The predominant serotype was S. Enteritidis (n = 116), followed by S. Infantis (n = 18), S. Gueuletapee (n = 16), S. Derby (n = 12), S. Meleagridis (n = 4), and S. London (n = 2). The remaining four isolates were serogroup-untypeable. A majority of the 172 isolates (96.51%) was resistant to one or more antibiotics and 61.05% of the Salmonella isolates showed a multidrug resistance phenotype. Statistical analysis indicated the one risk product stage for Salmonella contamination occurred in the sample source at the abattoir, specifically the stage of Carcasses after chilling. The majority of S. Enteritidis isolates shared the same pulsed-field gel electrophoresis (PFGE) cluster, suggesting that the S. Enteritidis strain might spread along the broiler chicken supply chain. The prevalence and antimicrobial resistance of Salmonella in different production stages suggest the importance of controlling Salmonella in the broiler chicken supply chain for public health, underlying the need for improved measures of reducing carcass contamination in abattoirs and the appropriate use of antimicrobials in broiler flocks.

Context: Huangqi injection (HQI), extracted from Astragali Radix, which has capability on treating the leucopenia. However, the potential metabolic mechanism is poorly understood. Objective: To investigate the effect of HQI on cyclophosphamide (Cy)-induced leucopenia in mice, the nuclear magnetic resonance (NMR)-based metabolomic profiling technique coupled with multivariate statistical analysis was applied. Materials and methods: NMR analysis was used to identify the various compounds of HQI, and high-performance liquid chromatography was applied to determine the contents of major compounds. A experimental mice model of leucopenia induced by Cy and NMR-based metabolomic approach was used to evaluate the pharmacological effect of HQI and to investigate its probable acting mechanism on leucopenia. Results: HQI increased body weight and elevated the white blood cell (WBC), monocytes (MO), neutrophils (NE), and lymphocyte (LY) levels of Cy-treated mice. In addition, the levels of most perturbed endogenous metabolites could be reversed after HQI treatment. Correlations between WBC, MO, NE, LY, and altered metabolite profiles in spleen were greater than that in serum, and the correlation in MO was more evident than those for WBC, NE, and LY. Discussion and conclusion: HQI showed obvious efficacy on the mice model of leucopenia. And the drug action of HQI on leucopenia was probably related with regulating metabolic pathways of energy metabolism, amino acids metabolism, oxidative stress, and choline metabolism. However, various compounds were present in the HQI, and the bioactive compounds responsible for the drug actions should be further investigated.

Ethnopharmacological relevance Xiaoyaosan (XYS), a represent and classic TCM prescription, consists of Radix Bupleuri, Radix Angelicae Sinensis, Radix Paeoniae Alba, Rhizoma Atractylodis Macrocephalae, Poria, Herba Menthae, Rhizoma Zingiberis Recens and Radix Glycyrrhizae. XYS can sooth the liver and strengthen the spleen through improving the circulation of qi and nourishing blood according to the TCM theory, therefore exhibiting anti-depression effects. Aim of the study This study was conducted to investigate the compatibility rule of antidepressant effect of XYS by using both the “Efficacy Compositions” research strategy and fecal metabolomics approach. Materials and methods XYS was divided into two efficacy groups, i.e. the Shugan (SG) and the Jianpi (JP) groups, according to the efficacies of both XYS and the eight herbs recorded in the TCM theory and the research strategy of “Efficacy Compositions”. A CUMS-induced depression model was constructed, where rats were randomly divided into 5 groups: negative control (NC), CUMS model (MS), XYS, SG, and JP. Multivariate data analysis including Principal Components Analysis (PCA) and Orthogonal Partial Least Squares-Discriminate Analysis (OPLS-DA) was utilized. Efficacy Index (EI) was calculated. Results Metabolic profiling by PCA showed that XYS exhibited the strongest effect than the two efficacy groups, locating closest to the control group. OPLS-DA showed 10 metabolites were identified as potential biomarkers for the CUMS-induced depression. 8 potential biomarkers were significantly reversed by XYS while 5 and 4 biomarkers were reversed by SG and JP, respectively. The results of regulatory degrees showed that XYS had the highest EI than SG and JP. Concerning metabolic pathways, XYS regulated all the seven metabolic pathways associated with CUMS-induced depression, while SG and JP groups regulated six and three pathways, respectively. Conclusions The antidepressant effect of XYS was stronger than that of SG and JP. The combined effects of SG and JP brought the integrated antidepressant effect of XYS. This study suggests that a combination of “Efficacy Compositions” strategy and metabolomics approach has great potentials in comprehensively and deeply understanding the scientific connotation of the compatibility rule of TCM prescriptions.

A multiplex lateral flow immunoassay sensor based on highly luminescent green-emitting Au nanoclusters (AuNCs-MLFIA sensor) was successfully established for the simultaneous and quantitative determination of clenbuterol (Clen) and ractopamine (RAC) in swine urine. The antigens of Clen and RAC were dispersed on a nitrocellulose membrane as two test lines, and the Au nanoclusters were synthesized from 6-aza-2-thiothymine and l -arginine to obtain highly green luminescence and ultra-small nanoparticles (Arg/ATT/AuNCs). Free carboxyl groups on Arg/ATT/AuNCs enabled conjugation with biomolecules to afford an indicator for the biosensor. The AuNCs-MLFIA sensor is based on the indirect competition assay and could successfully detect samples within 18 min without sample pretreatment, qualitative results can be obtained by visual inspection under a UV lamp. The limits of detection of Clen and RAC by the naked eye were both 0.25 μg L−1. In addition, the AuNCs-MLFIA sensor allowed quantitative detection combined with a portable fluorescence reader. The half-maximal inhibitory concentrations of Clen and RAC were 0.06 and 0.32 μg L−1, respectively, with detection limits of 0.003 and 0.023 μg L−1. Thirty blind-spiked swine urine samples were analyzed by the AuNCs-MLFIA sensor and liquid chromatography–tandem mass spectrometry, and the results of the two methods showed a significant correlation. The newly developed AuNCs-MLFIA sensor overcomes several limitations of conventional LFIA sensors, including their low sensitivity, limitation to quantify analytes, and single-analyte detection.

Organic/inorganic hybrid nanoflowers were synthesized from calcium phosphate and protein modified fluorescent gold nanoclusters and antigens. These nanoflowers are shown to be well suited labels for bioassay because they fulfill the functions of biological recognition and signal output. A fluorometric immunoassay was developed that was combined with immunomagnetic separation. In the detection system, the red fluorescence of the supernatant (measured at excitation/emission wavelengths of 360/640 nm) is found to be proportional to the clenbuterol (Clen) concentration after two immunomagnetic separations. The assay has a linear response in the 0.5 μg L−1 to 40 μg L−1 Clen concentration range, and 0.167 μg L−1 limit of detection. This makes it well suited for food safety monitoring. The average recoveries from spiked samples range from 92.7 to 109.1% (intra-assay) and 101.2 to 125.7% (inter-assay) with relative standard deviations of

In this study, 10 fluorescein-labeled ractopamine (RAC) derivatives (tracers) were synthesized and characterized to develop a rapid fluorescence polarization immunoassay (FPIA) for the detection of RAC in pork, using previously produced RAC polyclonal antibodies. The effect of the tracer structure on the sensitivity of the FPIA was investigated. The specificity of the FPIA was evaluated with 70 β-agonists and β-blockers. The FPIA showed a limit of detection of 0.56 μg kg−1 for RAC in pork, with recoveries ranging from 74.8% to 86.6% in spiked samples. The total analysis time, including sample pretreatment, was less than 1 h. The FPIA was used to screen 150 commercial pork samples for RAC residues and the results were consistent with those of an enzyme-linked immunosorbent assay (ELISA) and ultra-high-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS). Our results demonstrate that the FPIA developed here is a rapid, accurate, and sensitive screening method for RAC residues in pork.

Ethnopharmacological relevance Guilingji (GLJ), a famous and classical traditional Chinese medicine (TCM) prescription, has been used to extend the lifespan and improve the life qualities of the elderly for hundreds of years in China. Aim of the study: We aimed to explore the protective effects of GLJ on the testicular dysfunction of aging rats, as well as the regulating effects of GLJ on the metabolic disturbance and metabolite changes in natural aging rats. Materials and methods Forty 23-month-old rats were divided randomly into four groups, including the old control group and three groups of GLJ treatment at 37.5, 75, and 150 mg/kg doses, respectively. Additionally, 10 four-month rats were included as the youth control group. Testicular dysfunction was first evaluated by measuring the changes in the wet weights of the testicles, concentration of serum testosterone (T), and morphologic changes of the testis. Subsequently, an 1H NMR-based metabolomics approach coupled with multivariate analysis, including partial least squares discriminant analysis (PLS-DA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) was applied to monitor the metabolite changes. Results Compared with the old control group, the wet weights of the testicles and T concentration were significantly increased, while the morphologic abnormality of testicular tissues was improved by a 4-week treatment course with GLJ. Furthermore, compared with the old control group, the urinary levels of alanine, pantothenate, phenylalanine, β-hydroxybutyrate and pyruvate were significantly decreased after a 4-week treatment course with GLJ. Additionally, we found that amino acid metabolism and pyruvate metabolism were significantly involved in the regulatory effect of GLJ. Conclusions The current findings provided, for the first time, sound evidence of the protective effects of GLJ on testicular dysfunction from both biochemical and metabolomics perspectives. The mechanisms of GLJ could be related to regulating amino acid metabolism and pyruvate metabolism. The current study lays an important foundation for further research and for the broad clinical application of GLJ.

An immunoaffinity chromatographic method was developed for the purification of 9 sulfonamides (sulfamethazine, sulfadimethoxine, sulfamerazine, sulfathiazole, sulfamethoxazole, sulfamethizole, sulfadiazine, sulfamonomethoxine, and sulfapyridine) from chicken tissue (muscle and liver) samples. Two monoclonal antibodies (antisulfamethazine and antisulfamethoxazole) were simultaneously covalently coupled to CNBr-activated Sepharose 4B for the preparation of a re-usable immunoaffinity column. After extraction with methanolwater and purification by immunoaffinity column, the sulfonamides were determined by reversed-phase liquid chromatography and UV detection at 270 nm. The recoveries for each drug at fortification levels of 1050 ng/g ranged from 74.1 to 108.9 with relative standard deviations from 1.9 to 11.5. The limits of detection were 2 ng/g for each drug.

A method has been developed for simultaneous analysis of salinomycin and narasin in chicken muscle. Muscle samples were extracted with acetonitrile. Clean-up of the extracts on an immunoaffinity chromatography column was followed by liquid chromatography with postcolumn derivatisation and UV–visible detection at 520 nm. The immunoaffinity columns were prepared by coupling the anti-salinomycin monoclonal antibody to CNBr-activated Sepharose 4B. When chicken muscle fortified at 5, 25, and 50 ng g−1 was analyzed, intra-assay mean recoveries of salinomycin and narasin were in the ranges 87.5–93.1 and 86.2–94.3%, respectively, with relative standard deviation (RSD) of 4.7–6.2 and 2.4–5.7%. Inter-assay mean recoveries were 86.0–93.0 and 86.0–92.1%, respectively, with RSD of 4.8–6.5 and 5.8–7.4%. The limit of detection of the method was 2.5 ng g−1 for both drugs in chicken muscle.

A simple, rapid and sensitive liquid chromatographic method with programmable fluorescence and ultraviolet detection was developed and validated for simultaneous determination of seven fluoroquinolones (marbofloxacin, ofloxacin, ciprofloxacin, lomefloxacin, danofloxacin, enrofloxacin and difloxacin) and four sulfonamides (sulfadiazine, sulfapyridine, sulfathiazole and sulfadimidine) in chicken muscle in a single run. The tissue sample was extracted with phosphate buffer (pH 6.0) and cleaned-up with a solid phase extraction cartridge. The mean recoveries for each drug in chicken muscle ranged from 78.0 to 105.2% with a relative standard deviation below 9.3% at 0.2–400 ng g−1 fortification levels. The limit of quantification was 0.2–4.0 ng g−1 for fluoroquinolones and 15.0 ng g−1 for sulfonamides.

Eprinomectin is a novel and potent antiparasitic animal health drug. An analytical procedure for the determination of EPR in bovine urine and feces has been developed. The urine sample was centrifuged and alkalized with ammonia following solid phase extraction. The fecal sample was extracted with acetonitrile, defatted with hexane, cleaned-up using C18 cartridge. All samples were analyzed by high performance liquid chromatography with fluorescence detection after derivatization with N-methylimidazole. The limits of detection are 0.5 ng mL−1 and 0.5 ng g−1, respectively. Fortified at 2, 10, 50, and 100 ng mL−1(ng g−1), inter-assay recoveries of EPR in cattle urine and feces were in the range of 87.9–91.5% and 78.6–86.3%, with coefficients of variation of 5.4–10.2% and 1.4–7.2%, respectively. Intra-assay mean recoveries of the analytes were 82.2–86.5% and 79.6–87.3%, with coefficients of variation of 7.8–11.5% and 6.3–7.8%, respectively. The method was used to study the excretion of eprinomectin in bovine urine and feces after subcutaneous administration at a dose of 0.5 mg kg−1.

A rapid and robust multiresidue method was developed for the analysis of seven quinolones (ciprofloxacin, danofloxacin, enrofloxacin, sarafloxacin, difloxacin, oxolinic acid, and flumequine) in animal muscle tissues by HPLC-FLD. The samples were extracted with phosphate aqueous solution and cleaned with HLB-SPE. The analytes were separated using formic acid aqueous solution-acetontrile system as mobile phase with a linear gradient elution program, and determined by programmable fluorescence detection. The linear range was 0.3–1000 μg l −1 with correlation coefficients ( r ) greater than 0.9989. The limits of detection were 0.1–0.3 μg kg −1 , and limits of quantification were 0.3–1.0 μg kg −1 . The mean recoveries for each analyte in chicken and pig muscles ranged from 70.4% to 105.8% with relative standard deviation below 9.3% at 1–100 μg kg −1 fortification levels. The method is convenient, fast, safe, sensitive, and accurate for determining residual quinolones in actual samples.

A simple multiresidue method, HPLC with programmable fluorescence detection and gradient elution, has been developed for analysis of nine (fluoro)quinolones (FQs)—norfloxacin, ciprofloxacin, lomefloxacin, danofloxacin, enrofloxacin, sarafloxacin, difloxacin, oxolinic acid, and flumequine in chicken muscle. The samples were extracted with phosphate-buffered saline (PBS, 0.01 mol L−1, pH 7.0) and cleaned by SPE. Cleaned extracts reconstituted in different solvents were tested to determine which gave the maximum fluorescence response for each drug. PBS (0.01 mol L−1, pH 7.0) was used as reconstitution solvent, because the sensitivity for FQs dissolved in PBS was 1.8–3.2 times greater than when dissolved in the mobile phase. Under the optimum conditions excellent linearity was obtained, with satisfactory correlation coefficients (r > 0.9994) for PBS. The “matrix effect” was eliminated. Limits of quantification for each drug were in the range 0.3–1.0 ng g−1. In fortification studies recoveries of the analytes were in the range 71.8–102.1% for 1–100 ng g−1 concentrations. Inter- and intra-day coefficients of variation were from 0.5 to 5.2% and from 1.7 to 9.0%, respectively. Short-term stability in PBS was also determined.