The Molecular Mechanism of FABP4 Inhibition Effects of GAS and 4-HBA in Gastrodia elata Blume Was Discussed Based on NMR and Molecular Docking.

To isolate gastrodin (GAS), 4-hydroxybenzyl alcohol (4-HBA), and phenolic compounds from Chinese medicine Gastrodia elata Blume, and to explore the binding mode of fatty acid binding protein 4 (FABP4/aP2) that is closely related to macrophage inflammation, we study their anti-inflammatory targets. After the ultrasonic extraction of the main active components with 70% ethanol, three resins and three eluents were selected, and eight phenolic monomers with similar structures, such as gastrodin and 4-hydroxybenzyl alcohol, were isolated from Gastrodia elata by AB-8 macroporous resin and silica gel column chromatography and eluted with the CHCl3-MeOH gradient. Their structures were identified by HPLC and nuclear magnetic resonance (NMR). The FABP4 protein was added to GAS and 4-HBA, and the NMR experiment was performed to observe ligand binding. Finally, according to the spectral information of STD-NMR and molecular docking technology, the interaction between ligands and protein was studied. The fluorescence competition experiment confirmed that both GAS and 4-HBA were in the binding cavity of FABP4. Moreover, 3-phenoxy-2-phenylbenzoic acid (PPA) is a possible inhibitor of FABP4, reducing macrophage-related inflammation and endoplasmic reticulum stress. This work provides a new basis for the anti-inflammatory mechanism of Gastrodia elata, paving the way for the research and development of FABP4 inhibitor drugs.

Design of a prodrug photocage for cancer cells detection and anticancer drug release.

Developing probes for simultaneous diagnosis and killing of cancer cells is crucial, yet challenging. This article presents the design and synthesis of a novel Rhodamine B fluorescence probe. The design strategy involves utilizing an anticancer drug (Melphalan) to bind with a fluorescent group (HRhod-OH), forming HRhod-MeL, which is non-fluorescent. However, when exposed to the high levels of reactive oxygen species (ROS) of cancer cells, HRhod-MeL transforms into a red-emitting Photocage (Rhod-MeL), and selectively accumulates in the mitochondria of cancer cells, where, when activated with green light (556 nm), anti-cancer drugs released. The Photocage improve the efficacy of anti-cancer drugs and enables the precise diagnosis and killing of cancer cells. Therefore, the prepared Photocage can detect cancer cells and release anticancer drugs in situ, which provides a new method for the development of prodrugs.

Visual monitoring of biocatalytic processes using small molecular fluorescent probes: strategies-mechanisms-applications.

Real-time monitoring of biocatalytic-based processes is significantly improved and simplified when they can be visualized. Visual monitoring can be achieved by integrating a fluorescent unit with the biocatalyst. Herein, we outline the design strategies of fluorescent probes for monitoring biocatalysis: (1) probes for monitoring biocatalytic transfer: γ-glutamine is linked to the fluorophore as both a recognition group and for intramolecular charge transfer (ICT) inhibition; the probe is initially in an off state and is activated via the transfer of the γ-glutamine group and the release of the free amino group, which results in restoration of the "Donor-π-Acceptor" (D-π-A) system and fluorescence recovery. (2) Probes for monitoring biocatalytic oxidation: a propylamine is connected to the fluorophore as a recognition group, which cages the hydroxyl group, leading to the inhibition of ICT; propylamine is oxidized and subsequently ß-elimination occurs, resulting in exposure of the hydroxyl group and fluorescence recovery. (3) Probes for monitoring biocatalytic reduction: a nitro group attached to a fluorophore as a fluorescence quenching group, this is converted to an amino group by catalytic reduction, resulting in fluorescence recovery. (4) Probes for monitoring biocatalytic hydrolysis: ß-D-galactopyranoside or phosphate acts as a recognition group attached to hydroxyl groups of the fluorophore; the subsequent biocatalytic hydrolysis reaction releases the hydroxyl group resulting in fluorescence recovery. Following these 4 mechanisms, fluorophores including cyanine, coumarin, rhodamine, and Nile-red, have been used to develop systems for monitoring biocatalytic reactions. We anticipate that these strategies will result in systems able to rapidly diagnose and facilitate the treatment of serious diseases.

Effect of Rhei Radix et Rhizoma and Eupolyphaga Steleophaga on liver protection mechanism based on pharmacokinetics and metabonomics.

Objective: Based on metabonomics technology of high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) and hydrogen nuclear magnetic resonance spectroscopy (1H NMR), the pharmacokinetic characteristics and therapeutic mechanism of Rhei Radix et Rhizoma (RhRR, Dahuang in Chinese), Eupolyphaga Steleophaga (EuS, Tubiechong in Chinese) combined with RhRR acting on acute liver injury were explored. Methods: Models of acute liver injury were established, and the pharmacokinetic methods of five components of RhRR-EuS in rats were found by HPLC-MS/MS. The liver tissues of different groups of mice were analyzed by 1H NMR spectroscopy combined with multivariate statistical analysis to investigate the metabolomics of RhRR-EuS and RhRR. Results: Pharmacokinetic results showed there were different levels of bimodal phenomenon in different groups, and the absorption of free anthraquinone in RhRR increased after compatibility with EuS. In addition, the pathological state of acute liver injury in rats can selectively promote the absorption of emodin, chrysophanol, physcion and aloe emodin. Through 15 differential metabolites in the liver tissue of acute liver injury mice, it was revealed that RhRR-EuS and RhRR could protect the liver injury by regulating the metabolism of glutamine and glutamic acid, alanine, aspartic acid and glutamic acid, and phosphoinositide. However, the regulation of RhRR was weaker than that of RhRR-EuS. Conclusion: For the first time, we studied the pharmacokinetics and metabolomics differences of RhRR-EuS and RhRR in rats and mice with acute liver injury, in order to provide theoretical reference for clinical treatment of liver disease by DHZCP.

Metabolic regulation mechanism of Aconiti Radix Cocta extract in rats based on 1H-NMR metabonomics.

Objective: To establish a metabonomics research technique based on the combination of 1H-NMR and multivariate statistical analysis, so as to explore the metabolic regulation mechanism of Aconiti Radix Cocta extract (ARCE) in rat tissues and serum. Methods: SD rats were randomly divided into blank group, female group and male group. The 1H-NMR technique was used to collect the information of rat tissues and serum samples in each group. The principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA) and other methods were used for data pattern recognition, so as to screen out potential differential metabolites and metabolic pathways, and then network analysis and KEGG database were used to analyze the relationship between metabolites, metabolic pathways and diseases. Results: The external features and 1H-NMR analysis showed that the sex of rats had no obvious effect on the drug action. A total of 15 potential differential metabolites and six metabolic pathways were screened out through data pattern recognition. Through network analysis and KEGG pathway analysis, three target diseases closely related to differential metabolites were found, and the metabolic pathway related to lung cancer was the central carbon metabolism of cancer. Conclusion: This study shows that Aconiti Radix Cocta (ARC) may regulate the energy metabolism of the body by influencing arginine synthesis, so as to play the roles of anti-inflammation, analgesia, anti-tumor and immune regulation.

Study on pharmacokinetics and tissue distribution of Polygonatum sibiricum polysaccharide in rats by fluorescence labeling.

To study the pharmacokinetics and tissue distribution characteristics of Polygonatum sibiricum (P. sibiricum) polysaccharide administered orally and intravenously in rats, the latest quantitative analysis method was established where P. sibiricum polysaccharide was labeled with fluorescein isothiocyanate (FITC) in plasma and tissues. Quantitative analysis method of P. sibiricum polysaccharide in rat plasma and tissues was established by fluorescence spectrophotometry with FITC as a highly sensitive fluorescent molecular probe. The results showed that P. sibiricum polysaccharide was successfully labeled with FITC, and the degree of substitution was 0.55 %. Pharmacokinetic characteristics showed that oral administration (ig) and intravenous injection (iv) were consistent with the characteristics of two-compartment model. PRP-TYR-FITC administered orally was poorly absorbed in rats with low bioavailability. After a single ig and iv administration in rats for 8 h, P. sibiricum polysaccharide can be distributed in most tissues. The analysis results showed that P. sibiricum polysaccharide was distributed mostly in lung, kidney and liver for both routes of administration. When taking orally, the distribution pattern was: lung > liver > kidney > small intestine > stomach > heart > spleen > brain. When taking intravenously, the distribution pattern was: liver > lung > kidney > small intestine > heart > stomach > spleen > brain. Fluorescence labeling of P. sibiricum polysaccharide by FITC was successfully realized. This method was proved to be suitable for the study of pharmacokinetics and tissue distribution of P. sibiricum polysaccharide in rats. The above research lays foundation for further elucidating the clinical pharmacological mechanism of polysaccharide in P. sibiricum.

Determination of polysaccharides composition in Polygonatum sibiricum and Polygonatum odoratum by HPLC-FLD with pre-column derivatization.

A high-performance liquid chromatography-fluorescence detection (HPLC-FLD) method was established for the determination of seven monosaccharides in Polygonatum sibiricum and Polygonatum odoratum. The polysaccharides were de-esterified, extracted, hydrolyzed and derivatized with p-aminobenzoic acid (PABA) to obtain fluorescently labeled monosaccharide compounds, which were finally detected by HPLC-FLD. Inertsil ODS-3, C18 chromatographic column (250 mm × 4.6 mm, 5 µm) was used for chromatography. The excitation wavelength (Ex) was 313 nm, and the emission wavelength (Em) was 358 nm. Ethyl acetate extraction reduced the peaks of chromatogram and improved the detection sensitivity than other agents. The established method had high sensitivity, strong specificity, good linear relationship and recovery efficiency. The results showed that the roots and fibrous roots of Polygonatum sibiricum and Polygonatum odoratum contained these seven monosaccharides, and the highest monosaccharide content was mannose. The method of PABA-HPLC-FLD for determination of monosaccharide content in Polygonatum sibiricum and Polygonatum odoratum was sensitive and accurate. The method established in this work provides a feasible analytical tool for the study of polysaccharides, and the findings on polysaccharides from Polygonatum sibiricum and Polygonatum odoratum can provide guidance for the natural medicine industry.

Simultaneous Determination of Six Alkaloids in Rat Plasma by SPE-HPLC-MS/MS and Their Pharmacokinetics after Oral Administration of Radix aconiti Preparata Extract.

Simultaneous determination of the content of six alkaloids (aconitine, hypoaconitine, mesaconitine, benzoylaconine, benzoylhypaconine, and benzoylmesaconine) in rat plasma is enabled by HPLC-MS/MS combined with microsolid phase extraction (micro-SPE). To study its pharmacokinetics in rat plasma, the extracted plasma sample was passed through a C18 extraction column and eluted with acetonitrile. The six alkaloids in the Radix aconiti Preparata extract can be completely separated as peaks with good shape. The six components in the plasma sample showed a good linear relationship within their respective linear ranges (R 2 > 0.997). The analysis of the six alkaloids can be completed within 20 min. This method has high intraday and interday precision, and the room temperature stability and freeze-thaw stability are good. The matrix effect of the plasma samples is between 86.4 and 114%. The metabolism of the six Aconitum alkaloids in plasma is analyzed using a two-compartment model, which is characterized by fast absorption, slow elimination, and good linear fit, R 2 > 0.99. The peak time (T max) for aconitine, hypaconitine, and neoaconitine ranged from 29.95 to 42.07 min, while the peak time (T max) for benzoaconitine, benzohypaconitine, and benzoxinaconitine ranged from 42.88 to 73.08 min. With the increased dosage, the bioavailability of Aconitum alkaloids decreased gradually. The method for the determination of Aconitum alkaloids in rat plasma by high performance liquid chromatography-tandem mass spectrometry is sensitive and accurate, which is suitable for rat plasma analysis. The results provide a scientific basis for metabolic study of Aconitum alkaloids in vivo, and pave the way for clinical use of Aconitum medicinal materials and extracts.

Metabolomics study of polysaccharide extracts from Polygonatum sibiricum in mice based on 1 H NMR technology.

BACKGROUND: Polygonatum sibiricum Liliaceae perennial herb, as a commonly used medicine and food homologous plant, has been widely used in clinical practice of Chinese medicine since ancient times, with a history of 2000 years. As the main active ingredient, P. sibiricum polysaccharides have important pharmacological effects in blood sugar reduction and antitumor, antioxidant and liver protection. RESULTS: Mouse models of P. sibiricum polysaccharides were used in combination with 1 H NMR to investigate the metabolic regulation mechanism in mouse tissue and blood. The metabolite maps of the control group and the drug group in the liver had significant changes. The main differential metabolites were glucose 6-phosphate, inositol, lactose, glutamylglycine, galactose, rhamnose, cis-aconitic acid and histidine, indicating that there was definite correlation between the metabolic detection based on 1 H NMR and the metabolic characteristics of P. sibiricum. The common differential metabolites obtained by overall metabolism analysis were 3-hydroxybutyric acid, d-ribose, adenosine phosphate, inositol, fructose 6-phosphate, histidine, aspartic acid and cis-aconitic acid. CONCLUSIONS: This work forms the basis for identification of metabolic states combined with metabolic pathways, which could be used as diagnostic and prognostic indicators, providing therapeutic targets for new diseases. © 2020 Society of Chemical Industry.

Signal Sensitivity Enhancement of High-Spatial-Resolution MR Imaging with a Concatenated Cylindrical Parametric RF-Resonator.

Phased array MRI coils can increase sensitivity of superficial tissues owing to their proximity to the detection region. Deep-lying tissues, on the other hand, do not benefit to the same degree. Here we investigate the use of a localized cylindrically symmetric quadruple frequency resonator concatenated with a double frequency resonator to increase the longitudinal field-of-view (FOV) without compromising the spatial-resolution and detection sensitivity. These concatenated array coils work on the principle of a parametric amplification to provide wireless amplification of the locally detected NMR signal prior to inductively coupling the coil to an external pick-up loop with connection to the system receiver. When both the detectors are activated together, the effective range of both overlay to create a larger FOV enabling better identification of detectable regions. Furthermore, the in-vivo test of the concatenated detector provides a worst-case 5-fold SNR gain in regions separated from the cylindrical surface larger than its own diameter. This proposed approach of concatenated detector realization can be individually activated and manipulated to enlarge the sensitivity-enhanced region without sacrificing their individual performance. Compared to double frequency detectors, quadruple frequency detectors offer more flexibility in the choice of detector dimension, enabling multi-element concatenation over an extended FOV.

Efficient, accurate and comprehensive evaluation of polysaccharides from Fritillaria and their inhibitory responses to mouse inflammation.

Fritillaria is a perennial herb with bulbs with multiple medicinal usages. Here, we optimized extraction and hydrolysis methods for different species and origins of Fritillaria to study their anti-inflammatory activities. Using ultrasound-assisted hot water extraction after hydrolysis by hydrochloric acid and trifluoroacetate (TFA), monosaccharide derived from 1-phenyl-3-methyl-5-pyrazolone (PMP) was separated by using HPLC. The optimal conditions for hot water extraction were found via single factor analysis and orthogonal experiments. The PMP derivatization HPLC method could accurately determine the Mannose (Man), Glucose (Glu), Galactose (Gal), Xylose (Xyl), and Fucose (Fuc) content values in Fritillaria. This pre-column derivatization method is simple and rapid, providing less variation, high sensitivity and good reproducibility for saccharide separation. To study Fritillaria's anti-inflammatory effects in a mouse swelling model, HE-staining was performed to observe morphological changes in liver and kidney samples. Fritillaria's active ingredients could significantly inhibit mouse ear swelling induced by xylene and inhibit toe swelling induced by egg white. They mainly inhibited the secretion of inflammatory cytokines inside the body and maintained the dynamic balance between pro-inflammatory and anti-inflammatory factors in the body. According to histopathological analysis, Fritillaria had good anti-inflammatory activity against LPS-induced inflammation, reducing the expression of inflammatory cytokines.

A rapid, accurate and sensitive method for determination of monosaccharides in different varieties of Osmanthus fragrans Lour by pre-column derivatization with HPLC-MS/MS.

A specific HPLC-MS/MS (High-Performance Liquid Chromatography with tandem Mass Spectrometry) method was developed and validated for simultaneous determination of several monosaccharides in three kinds Osmanthus fragrans Lour. After extraction, separation, protein removal, pigment removal and hydrolysis, monosaccharides was finally obtained from Osmanthus fragrans Lour. Positive ion mode detection and Multiple Reaction Monitoring (MRM) mode were used for quantitative analysis by PMP pre-column derivatization and Electrospray Ionization (ESI). Analysis and content determination of 6 monosaccharide components in 3 kinds of Osmanthus fragrans Lour. The HPLC separation was achieved on a Shim-pack VP-ODS6022748 (150 L × 2.0) with gradient elution at a flow rate of 0.2 ml/min in a run time of 40 min, and the mobile phase was acetonitrile-5 mmol/L ammonium acetate. PMP derivatization in HPLC-MS/MS can accurately measure Osmanthus fragrans Lour. mannose (Man), ribose (Rib), rhamnose (Rha), galacturonic acid (Gal UA), glucose (Glu), galactose (Gal), xylose (Xyl), fucose (Fuc). The results showed that HPLC-MS/MS pre-column derivatization method was simple and rapid, with small measurement error, but high sensitivity and good repeatability. The analysis of monosaccharide components in polysaccharide components has important practical significance.

A novel and sensitive method for determining vitamin B3 and B7 by pre-column derivatization and high-performance liquid chromatography method with fluorescence detection.

A new labeling reagent for vitamin analysis, 2-amino-10-ethyl acridine ketone (AEAO), has been synthesized and successfully applied to the analysis of vitamin B3 and vitamin B7 in different tea samples. The reaction of AEAO with vitamins could proceed easily and quickly in the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC) as condensing reagent within 45 min. The derivatives exhibited excellent fluorescence property with excitation and emission wavelengths of 290 nm and 430 nm, respectively. Response surface methodology (RSM) was applied to the optimization of pre-column derivatization. Solid phase extraction with HLB cartridges was used for the extraction and purification of water-soluble vitamins in tea samples. The LODs for vitamin B3 and vitamin B7 were 2.56 and 2.22 ng mL-1, respectively. The proposed method was successfully applied to the analysis of vitamin B3 and vitamin B7 in different tea samples. The study provided a highly sensitive method for accurate analysis of trace vitamins from natural products.

Accurate Analysis and Evaluation of Acidic Plant Growth Regulators in Transgenic and Nontransgenic Edible Oils with Facile Microwave-Assisted Extraction-Derivatization.

Determination of plant growth regulators (PGRs) in a signal transduction system (STS) is significant for transgenic food safety, but may be challenged by poor accuracy and analyte instability. In this work, a microwave-assisted extraction-derivatization (MAED) method is developed for six acidic PGRs in oil samples, allowing an efficient (<1.5 h) and facile (one step) pretreatment. Accuracies are greatly improved, particularly for gibberellin A3 (-2.72 to -0.65%) as compared with those reported (-22 to -2%). Excellent selectivity and quite low detection limits (0.37-1.36 ng mL(-1)) are enabled by fluorescence detection-mass spectrum monitoring. Results show the significant differences in acidic PGRs between transgenic and nontransgenic oils, particularly 1-naphthaleneacetic acid (1-NAA), implying the PGRs induced variations of components and genes. This study provides, for the first time, an accurate and efficient determination for labile PGRs involved in STS and a promising concept for objectively evaluating the safety of transgenic foods.

Sensitive determination of taurine, γ-aminobutyric acid and ornithine in wolfberry fruit and cortex lycii by HPLC with fluorescence detection and online mass spectrometry identification.

A new, simple and highly sensitive method for the determination of taurine, γ-aminobutyric acid and ornithine using high-performance liquid chromatography (HPLC) with fluorescence detection is described. Three non-protein amino acids were derivatized by a novel precolumn derivatization reagent 2-[2-(dibenzocarbazol)-ethoxy]ethyl chloroformate before injected. Optimum derivatization was obtained at 40°C for 5 min in the presence of sodium borate buffer (pH 9.0). Derivatives were sufficiently stable to be efficiently analyzed by HPLC without pretreatment. On a reversed-phase Hypersil BDS C8 column, the amino acids were separated in conjunction with a gradient elution with a good baseline resolution. The identification of derivatives was carried out by online postcolumn mass spectrometry with an electrospray ionization source in positive ion mode. Excellent linear responses were observed with the correlation coefficients of >0.9996, and instrument detection limits (at a signal to noise of 3 : 1) were in the range of 0.30-0.33 nmol/L. The proposed method is sensitive and reproducible for the precise determination of the amino acids from wolfberry fruit and cortex lycii samples.