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IntroductionTrifolium pratense L. has anti-inflammatory, antioxidant, cardiovascular disease prevention, and estrogen-like effects. The existing method for the assay of effective components is commonly based on a spectrophotometer, which could not meet the requirement of quality control. Furthermore, although there have been many studies on the anti-inflammation effect of red clover, a few have been reported on the regulatory effect of red clover isoflavones (RCI) on lipopolysaccharide (LPS)-induced inflammatory response in porcine alveolar macrophages (3D4/2 cells), and its mechanism of action is still unclear.MethodsThe main components of RCI including daidzein, genistein, and biochanin A were accurately quantified by high-performance liquid chromatography coupled with diode array detection (HPLC-DAD) after optimizing the extraction process through response surface methodology. The anti-inflammatory potential of RCI was carried out by detecting the level of inflammatory cytokines and mRNA expression of related genes. Furthermore, its anti-inflammatory mechanism was explored by investigating two signaling pathways (NF-κB and MAPK).ResultsThe optimal extraction conditions of RCI were as follows: the concentration of ethanol is 86% and the solid–liquid ratio is 1:29, with the herb particle size of 40 mesh sieve. Under the optimal conditions, the total extraction of target components of RCI was 2,641.469 μg/g. The RCI could significantly suppress the production and expression of many pro-inflammatory cytokines. The results of the Western blot revealed that RCI dramatically reduced the expression of p65, p-p65, IκB-α, p38, and p-p38. These results are associated with the suppression of the signal pathway of p38 MAPK, and on the contrary, activating the NF-κB pathway. Collectively, our data demonstrated that RCI reversed the transcription of inflammatory factors and inhibited the expression of p65, p-p65, IκB-α, and p38, indicating that RCI had excellent anti-inflammatory properties through disturbing the activation of p38 MAPK and NF-κB pathways.ConclusionThe extraction conditions of RCI were optimized by HPLC-DAD combined with response surface methodology, which will contribute to the quality control of RCI. RCI had anti-inflammatory effects on the LPS-induced 3D4/2 cells. Its mechanism is to control the activation of NF-κB and p38 MAPK pathways, thereby reducing the expression of inflammatory-related genes and suppressing the release of cytokines.

IntroductionThe pharmacokinetic profile and residue depletion of eugenol in carp (Cyprinus carpio) tissues and plasma were performed by a convenient and reliable high-performance liquid chromatography (HPLC) method.MethodsThe eugenol in carp tissues and plasma was extracted with a mixed solution of acetonitrile and methanol. N-hexane was used to remove lipid impurities. The method was successfully applied to the pharmacokinetic and residue elimination of eugenol in carp after the carp was administered a medicated bath.ResultsThe average recoveries of eugenol in tissues and plasma fortified with four concentration levels were 69.0–106.6% and 80.0–86.7%, respectively. The relative standard deviations were < 8.9%. The limit of detection (LOD) was 0.01 μg/g in tissue and 0.008 μg/ml in plasma, respectively. The pharmacokinetic parameter of Cmax for eugenol in plasma at the concentrations of 20, 35, and 75 mg/L were 10.86, 17.21, and 37.32 mg/L, respectively. The t1/2 values were 3.68, 4.22, and 9.31 h. After the investigation of the anesthetic effect, 35 mg/L of eugenol was the optimal concentration for anesthesia. The highest accumulation concentration of eugenol in carp is in the liver and the lowest is in the muscle. In addition, the eugenol in tissue was eliminated rapidly and at a lower level than the LOD at 48 h. According to the residue elimination, the withdrawal time of eugenol was suggested at 5.2 days.DiscussionThese results indicate that the developed method had good linearity and accuracy, and is sensitive enough for the monitoring of eugenol residue in carp. The half-life of eugenol decreased with the increase in drug concentration and the eugenol was eliminated rapidly in carp tissues. 35 mg/L eugenol was recommended as an anesthetic in carp due to its favorable anesthetic effect and no mortality. This study will contribute to the establishment of MRL regulation and setting a withdrawal period.

The rise of novel mcr mobile resistance genes seriously threatens the use of colistin as a last resort antibiotic for treatment of multidrug-resistant Gram-negative bacterial infections in humans. Large quantities of colistin are released annually into the environment through animal feces. This leads to environmental toxicity and promotes horizontal transmission of the mcr gene in aqueous environments. We examined colistin degradation catalyzed by the presence of strong oxidant Fe (VI). We found almost complete colistin degradation (>95%) by Fe (VI) at initial colistin levels of 30 μM at a molar ratio of Fe (VI): colistin of 30 using an initial pH 7.0 at 25°C for 60 min. The presence of humic acid did not alter the degradation rate and had no significant impact on the removal of colistin by Fe (VI). Quantitative microbiological assays of Fe (VI)-treated colistin solutions using Escherichia coli, Staphylococcus aureus, and Bacillus subtilis indicated that the residual antibacterial activity was effectively eliminated by Fe (VI) oxidation. Luminescent bacteria toxicity tests using Vibrio fischeri indicated that both colistin and its degradation products in water were of low toxicity and the products showed decreased toxicity compared to the parent drug. Therefore, Fe (VI) oxidation is a highly effective and environment-friendly strategy to degrade colistin in water.

Since antimicrobials were banned as feed additives, coccidiostats with favorable anticoccidial action and growth promotion have been widely used in the breeding industry. The monitoring of coccidiostats in feed is necessary, while the current methods based on mass-spectrometer analysis have limited applicability and matrix effects could interfere with the results. Accordingly, in the present paper, a rapid analytical strategy for the simultaneous determination of six synthetic coccidiostats in feed using high-performance liquid chromatography coupled with diode-array detection was developed. Coccidiostats in chicken feeds were extracted with the trichloroacetic acid–acetonitrile solution. The cleanup was performed by dispersive solid-phase extraction after the optimization of the response surface methodology. The method exhibited good linearity for target coccidiostats within the range of 0.05~20 µg/mL. Recoveries for six compounds in fortified feed samples were from 67.2% to 107.2% with relative standard deviations less than 9.6%. The limit of detection was 0.2~0.3 mg/kg. The successful application of the method in commercial feed verified that it is effective and sensitive for the rapid determination of multiple coccidiostats in chicken feeds.

Lipoxygenases (LOXs; EC1.13.11.12) are widely involved in plant growth, development, maturation, senescence and stress resistance. LOXs have been studied in many crop species but not in Tartary buckwheat [Fagopyrum tataricum (L.) Gaertn.]. This study aimed to systematically identify and analyze LOX genes in Tartary buckwheat. Eight LOX genes, which were unevenly distributed on five chromosomes, were identified in the Pinu1 Genome database. The data indicated that during long evolution, FtLOX genes were subjected to purifying selection. The conservation of motifs within the 9-LOX and 13-LOX subfamilies supported the results of phylogenetic analyses. All FtLOX proteins contained the LOX family-specific His-(X)4-His-(X)4-His-(X)17-His-(X)8-His motif, suggesting that this motif is essential for protein function. Analyses of physical and chemical properties showed that FtLOX6 might function under both acidic and alkaline conditions. The FtLOX gene promoters contained many light-responsive elements, and transcriptome data further suggested their roles in growth regulation and the response to light. Quantitative real-time polymerase chain reaction analysis of expression differences in plants exposed to different wavelengths of light confirmed that FtLOX1 and FtLOX4–7 could respond to light. Genes co-expressed with FtLOX genes were involved in the photoreaction-related pathway. This study provides a basis for further functional research in order to better understand the biological role of individual LOX gene in Tartary buckwheat.

Under solvent-free conditions, syntheses of 2-aryl-4-(4- β-D-allopyranosyloxyphenyl)-4,6,7,8tetrahydroquinolin-5(1 H)-one derivatives were carried out from chalcone ( 2a-2e), cyclohexane-1,3-dione ( 3), and NHOAc in excellent yield without using any catalysts. The structure of the new compounds were characterized by H NMR, IR, and HR-MS spectroscopy. The preliminary bioassay tests of 4a-4j indicated that compounds 4b, 4e, and 4f exhibited potent sedative and hypnotic activity. [ABSTRACT FROM AUTHOR]