Characterization of in vitro metabolites of irisflorentin by rat liver microsomes using high-performance liquid chromatography coupled with tandem mass spectrometry

Belamcanda chinensis has been extensively used as antibechic, expectorant and anti-inflammatory agent in traditional medicine. Irisflorentin is one of the major active ingredients. However, little is known about the metabolism of irisflorentin so far. In this work, rat liver microsomes (RLMs) were used to investigate the metabolism of this compound for the first time. Seven metabolites were detected. Five of them were identified as 6,7-dihydroxy-5,3,4,5-tetramethoxy isoflavone (M1), irigenin (M2), 5,7,4-trihydroxy-6,3,5-trimethoxy isoflavone (M3), 6,7,4-trihydroxy-5,3,5-trimethoxy isoflavone (M4) and 6,7,5-trihydroxy-5,3,4-trimethoxy isoflavone (M5) by means of NMR and/or HPLC-ESI-MS. The structures of M6 and M7 were not elucidated because they produced no MS signals. The predominant metabolite M1 was noted to be a new compound. Interestingly, it was found to possess anticancer activity much higher than the parent compound. The enzymatic kinetic parameters of M1 revealed a sigmoidal profile, with V-max = 12.02 m/mg protein/min, K-m = 37.24 m, CLint = 0.32 L/mg protein/min and h = 1.48, indicating the positive cooperation. For the first time in this work, a new metabolite of irisflorentin was found to demonstrate a much higher biological activity than its parent compound, suggesting a new avenue for the development of drugs from B. chinensis, which was also applicable for other herbal plants. Copyright (c) 2016 John Wiley & Sons, Ltd

Quantification of mycotoxins in vegetable oil by UPLC-MS/MS after magnetic solid-phase extraction

The detection of mycotoxin contamination in foodstuffs is highly significant for public health. Herein we report an analytical method based on magnetic solid-phase extraction (MSPE) and UPLC-MS/MS for the simultaneous determination of mycotoxins, including fumonisins B-1 (FB1), zearalenone (ZON) and ochratoxin A (OTA), in vegetable oil. Magnetic nanoparticles coated with double layers of silicon dioxide were synthesised and found to be an effective MSPE adsorbent for mycotoxins. The proposed MSPE procedure serves not only for sample clean-up but also for mycotoxin enrichment that enhances greatly the assay's sensitivity. Under the selected MSPE conditions, linear matrix-matched calibration curves were obtained for mycotoxins in a concentration range from 0.178 to 625gkg(-1). The limits of detection were 0.210gkg(-1) for FB1, 0.0800gkg(-1) for OTA and 1.03gkg(-1) for ZON. The proposed MSPE UPLC-MS/MS method was applied for the determination of mycotoxins in vegetable oil samples, including maize oil, rapeseed oil and soybean oil. ZON was detected in a maize oil at 101gkg(-1), which is below the European Union limit of 200gkg(-1) in foodstuffs

Magnetism-assisted modification of screen printed electrode with magnetic multi-walled carbon nanotubes for electrochemical determination of dopamine

A simple and sensitive dopamine (DA) electrochemical sensor was fabricated based on magnetism-assisted modification of screen printed electrode (SPE) with magnetic multi-walled carbon nanotubes (mMWCNTs). The mMWCNTs modified electrodes (mMWCNTs/SPE) combines the advantages of SPE and the simultaneous contribution of magnetic nanoparticles (MNPs) and MWCNTs, increasing sensitivity and selectivity of DA detection. The linearity was found between 5 mu M to 180 mu M, with the limit of detection (LOD) of 0.43 mu M. In the mean time, this modified electrode exhibited excellent selectivity for DA detection with almost no interference from ascorbic acid (AA), which co-exists with DA in many bio-samples and causes common interference. Finally, this novel electrode has been applied to determine DA concentration in spiked human blood serum and satisfactory recovery was found in the range of 97.43-102.94% with the RSDs of less than 2.27%. This work developed a sensitive and reliable electrochemical analytical method based on mMWCNTs/SPE, which exhibits great potential for diagnosis of the diseases related to DA. (C) 2017 Elsevier B.V. All rights reserved

Functionalized carbon quantum dots with dopamine for tyrosinase activity analysis

Tyrosinase (TYR) is a key enzyme in melanin biosynthesis and its activity is an important biomarker for dermatological disorders, such as vitiligo, melanoma and actinic damages. Sensitive assay for TYR activity is significant for basic and clinical research. In this work, a facile fluorescent assay for TYR activity based on dopamine functionalized carbon quantum dots (CQDs-Dopa) has been developed. Dopamine (Dopa) was covalently bond to CQDs through a simple one-pot hydrothermal method, and the prepared CQDs-Dopa exhibited a fluorescence emission at 499 nm under exciting wavelength at 310 nm with a quantum yield of approximately 2.1%. When TYR was mixed with CODs-Dopa, the dopamine moiety in CQDs-Dopa conjugate was oxidized to O-dopaquinone, and an intra-particle photo-induced electron transfer (PET) process consequently occurred between CQDs and O-dopaquinone to quench the fluorescence of CQDs-Dopa. TYR activity can be determined based on the fluorescence quenching degree of CQDs-Dopa. This assay covered two broad linear ranges: 44.4-711.1 U L-1 and 711.1-2925.4 U L-1, with detection limit of 17.7 U L-1. The proposed fluorescent assay was applied to TYR activity measurement in human serum samples. It showed promising potential for TYR activity assay in clinical applications. (C) 2017 Elsevier B.V. All rights reserved

PEGylation of magnetic multi-walled carbon nanotubes for enhanced selectivity of dispersive solid phase extraction

Carbon nanotubes (CNTs) possess large potential as extraction absorbents in solid phase extraction. They have been widely applied in biomedicine research, while very rare application in natural product chemistry has been reported. In this work, methoxypolyethylene glycol amine (mPEG-NH2) is covalently coupled to CNTsmagnetic nanoparticles (CNTs-MNP) to prepare a novel magnetic nanocomposite (PEG-CNTs-MNP) for use as dispersive solid-phase extraction (DSPE) absorbent. The average particle size was 86 nm, and the saturation magnetization was 52.30 emu/g. This nanocomposite exhibits excellent dispersibility in aqueous systems, high selectivity and fast binding kinetics when used for extraction of Z-ligustilide, the characteristic bioactive compound from two popular Asian herbal plants, R chuanxiong and R. ligusticum. HPLC quantification of Z-ligustilide extracted from the standard sample solution showed a high recovery of 98.9%, and the extraction rate"from the extracts of the above two herbs are both around 70.0%. To our knowledge, this is the first report on using PEG-CNTs-MNP as DSPE nanosorbents for selective extraction of natural products. This nano-material has promising application in isolation and enrichment of targeted components from complex matrices. (C) 2016 Elsevier B.V. All rights reserved

Covalent immobilization of porcine pancreatic lipase on carboxyl-activated magnetic nanoparticles: Characterization and application for enzymatic inhibition assays

Using carboxyl functionalized silica-coated magnetic nanoparticles (MNPs) as carrier, a novel immobilized porcine pancreatic lipase (PPL) was prepared through the 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) coupling reaction. Transmission electron microscopic images showed that the synthesized nanoparticles (Fe3O4-SiO2) possessed three dimensional core-shell structures with an average diameter of similar to 20 nm. The effective enzyme immobilization onto the nanocomposite was confirmed by atomic force microscopic (AFM) analysis. Results from Fourier-transform infrared spectroscopy (FT-IR), Bradford protein assay, and thermo-gravimetric analysis (TGA) indicated that PPL was covalently attached to the surface of magnetic nanoparticles with a PPL immobilization yield of 50 mg enzyme/g MNPs. Vibrating sample magnetometer (VSM) analysis revealed that the MNPs-PPL nanocomposite had a high saturation magnetization of 4225 emu.g(-1). The properties of the immobilized PPL were investigated in comparison with the free enzyme counterpart. Enzymatic activity, reusability, thermo-stability, and storage stability of the immobilized PPL were found significantly superior to those of the free one. The K-m and the V-max values (0.02 mM, 6.40 U.mg(-1) enzyme) indicated the enhanced activity of the immobilized PPL compared to those of the free enzyme (0.29 mM, 3.16 U.mg(-1) enzyme). Furthermore, at an elevated temperature of 70 degrees C, immobilized PPL retained 60% of its initial activity. The PPL-MNPs nanocomposite was applied in the enzyme inhibition assays using orlistat, and two natural products isolated from oolong tea (i.e., EGCG and EGC) as the test compounds. (C) 2014 Elsevier B.V. All rights reserved

Quick and selective extraction of Z-ligustilide from Angelica sinensis using magnetic multiwalled carbon nanotubes

A facile and highly efficient magnetic solid-phase extraction method has been developed for Z-ligustilide, the major therapeutic agent in Angelica sinensis. The solid-phase adsorbent material used was prepared by conjugating carbon nanotubes with magnetic Fe3O4 nanoparticles via a hydrothermal reaction. The magnetic material showed a high affinity toward Z-ligustilide due to the pi-pi stacking interaction between the carbon nanotubes and Z-ligustilide, allowing a quick and selective exaction of Z-ligustilide from complex sample matrices. Factors influencing the magnetic solid-phase extraction such as the amount of the added adsorbent, adsorption and desorption time, and desorption solvent, were investigated. Due to its high extraction efficiency, this method was proved highly useful for sample cleanup/enrichment in quantitative high-performance liquid chromatography analysis. The proposed method had a linear calibration curve (R-2 = 0.9983) over the concentration between 4 ng/mL and 200 mu g/mL Z-ligustilide. The accuracy of the method was determined by the recovery, which was from 92.07 to 104.02%, with the relative standard deviations >4.51%

Screening of lipase inhibitors from Scutellaria baicalensis extract using lipase immobilized on magnetic nanoparticles and study on the inhibitory mechanism

Scutellaria baicalensis is a traditional Chinese medicinal plant possessing a wide variety of biological activities. In this work, lipase immobilized on magnetic nanoparticles (LMNPs) was used as solid phase extract absorbent for screening of lipase inhibitors from this plant. Three flavonoids were found to bind to LMNPs and were identified as baicalin, wogonin, and oroxylin A by liquid chromatography-mass spectrometry (HPLC-MS). Their IC50 values were determined to be 229.22 +/- 12.67, 153.71 +/- 9.21, and 56.07 +/- 4.90 mu M, respectively. Fluorescence spectroscopy and molecular docking were used to probe the interactions between these flavonoids and lipase. All the flavonoids quenched the fluorescence of lipase statically by forming new complexes, implying their affinities with the enzyme. The thermodynamic analysis suggested that van der Waals force and hydrogen bond were the main forces between wogonin and lipase, while hydrophobic force was the main force for the other two flavonoids. The results from a molecular docking study further revealed that all of them could insert into the pocket of lipase binding to a couple of amino acid residues