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<br>Display Omitted<br>• 4′-Carbonyl chloride rosamine was synthesized and used for NTs by UHPLC-MS/MS. • <b>In situ</b> UA-DDLLME was reported for the simultaneous determination of AANTs and MANTs. • The method was sensitive, selective, low matrix effect, speedy and eco-friendly. • A new analytical tool in diagnosis of AD-related disease.<br>Neurotransmitters (NTs) may play an important role in neurodegenerative disorders such as Alzheimer’s disease (AD). In order to investigate the potential links, a new simple, fast, accurate and sensitive analytical method, based on <b>in situ</b> ultrasound-assisted derivatization dispersive liquid-liquid microextraction (<b>in situ</b> UA-DDLLME) coupled with ultra high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS), has been developed and validated. The quantitation of amino acid neurotransmitters (AANTs) and monoamine neurotransmitters (MANTs) in urine of AD rats were performed in this work. The <b>in situ</b> UA-DDLLME procedure involved the rapid injection of the mixture of low toxic 4-bromoanisole (extractant) and acetonitrile (dispersant), which containing the new designed and synthesized 4′-carbonyl chloride rosamine (CCR) as derivatization reagent, into the aqueous phase of real sample and buffer. Under the selected conditions, the derivatization and microextraction of analytes were simultaneously completed within 1 min. Good linearity for each analyte (R > 0.992) was observed with low limit of detections (LODs, S/N > 3). Moreover, the proposed method was compared with direct detection or other reported methods, and the results showed that low matrix effects and good recoveries results were obtained in this work. Taken together, <b>in situ</b> UA-DDLLME coupled with UHPLC-MS/MS analysis was demonstrated to be a good method for sensitive, accurate and simultaneous monitoring of AANTs and MANTs. This method would be expected to be highly useful in AD diseases’ clinical diagnostics and may have potential value in monitoring the efficacy of treatment.
This study presents an efficient strategy based on liquid-liquid extraction, high-speed counter-current chromatography, and preparative HPLC for the rapid enrichment, separation, and purification of four anthraquinones from Rheum tanguticum. A new solvent system composed of petroleum ether/ethyl acetate/water (4:2:1, v/v/v) was developed for the liquid-liquid extraction of the crude extract from R. tanguticum. As a result, emodin, aloe-emodin, physcion, and chrysophanol were greatly enriched in the organic layer. In addition, an efficient method was successfully established to separate and purify the above anthraquinones by high-speed counter-current chromatography and preparative HPLC. This study supplies a new alternative method for the rapid enrichment, separation, and purification of emodin, aloe-emodin, physcione, and chrysophanol.
In this paper, an efficient method was successfully established by the combination of macroporous resin (MR) and high-speed counter-current chromatography (HSCCC) for rapid enrichment and separation of aloe-emodin 8-O-β-D-glucoside, emodin 1-O-β-D-glucoside, emodin 8-O-β-D-glucoside and piceatannol 4'-O-β-D-(6″-O-gallate)-glucoside. Six kinds of macroporous resins were investigated in the first step and X-5 macroporous resin was selected for the enrichment of the target compounds. The recoveries of the target compounds reached 89.0, 85.9, 82.3 and 84.9% respectively after 40% ethanol elution. In the second step, the target compounds were separated by HSCCC with a two-phase solvent system composed of chloroform/ethyl acetate/methanol/water (8:1:6:5, v/v). The established method will be helpful for further characterization and utilization of Rheum tanguticum. The results demonstrate that MR coupled with HSCCC is a powerful technique for separation of bioactive compounds from natural products.
Zuota is regarded as the king of Tibetan medicine. However, due to the confidentiality of this precious medicine, the scientific characterization of Zuota is very scarce, which limits the pharmacology and biosafety studies of Zuota. Herein, we collected four different Zuota samples from Tibet, Qinghai, Gansu, and Sichuan and characterized them by multiple techniques. Our results showed that Zuota was mainly an inorganic mixture of HgS, sulfur, and graphite. Morphologically, Zuota samples were composed of nanoparticles, which further aggregated into microsized particles. Chemically, the majorities of Zuota were S and Hg (in the forms of HgS and pure sulfur). All samples contained pure sulfur with orthorhombic crystalline. Zuota from Qinghai province had different HgS crystalline, namely, hexagonal crystalline. The others were all face-centered cubic crystalline. Carbon in Zuota NPs was in the form of graphite. The implication to future studies of Zuota was discussed.
Four novel lactams, colletotrilactam A-D (1-4), along with six known compounds (5-10) were isolated from the culture broth of Colletotrichum gloeosporioides GT-7, a fungal endophyte of Uncaria rhynchophylla. The structures of these compounds were elucidated by comprehensive NMR spectroscopy. Isolates were tested for monoamine oxidase (MAO) inhibitory activity and compound 9 showed potent MAO inhibitory activity with IC50 value of 8.93±0.34μg/mL, when the IC50 value of iproniazid as a standard was 1.80±0.5μg/mL.
A simple and sensitive high-performance liquid chromatographic (HPLC) method with fluorescence detection and mass spectrometric identification has been developed for analysis of 30 long-chain and short-chain free fatty acids (FFAs). The fatty acids were derivatized to their esters with 1-[2-(<i>p</i>-toluenesulfonate)ethyl]-2-phenylimidazole-[4,5-<i>f</i>]-9,10-phenanthrene (TSPP) in <i>N</i>,<i>N</i>-dimethylformamide (DMF) at 90 °C with anhydrous K<sub>2</sub>CO<sub>3</sub> as catalyst. A mixture of C<sub>1</sub>-C<sub>30</sub> fatty acids was completely separated within 60 min by gradient elution on a reversed-phase C<sub>8</sub> column. Qualitative identification of the acids was performed by atmospheric-pressure chemical ionization mass spectrometry (APCI-MS) in positive-ion mode. The fluorescence excitation and emission wavelengths were 260 and 380 nm, respectively. Quantitative determination of the 30 acids in two Tibetan medicines <i>Gentiana straminea</i> and <i>G. dahurica</i> was performed. The results indicated that the medicines contained many FFAs. Linear correlation coefficients for the FFA derivatives were >0.9991. Relative standard deviations (RSDs, <i>n</i> = 6) for the fatty acid derivatives were <3%. Detection limits (at a signal-to-noise ratio of 3:1) were 3.1-38 fmol. When the fatty acid derivatives were determined in the two real samples results were satisfactory and the sensitivity and reproducibility of the method were good.
• A new low toxic dual-UADLLME coupled with microwave-assisted derivatization was proposed. • 4′-Carboxy-substituted rosamine was firstly used as derivatization reagent. • Simultaneous determination of PPD and PPT in rat plasma was achieved by UHPLC-MS/MS. • This method was successfully applied to pharmacokinetics study.<br>This paper, for the first time, reported a speedy hyphenated technique of low toxic dual ultrasonic-assisted dispersive liquid-liquid microextraction (dual-UADLLME) coupled with microwave-assisted derivatization (MAD) for the simultaneous determination of 20(<b>S</b>)-protopanaxadiol (PPD) and 20(<b>S</b>)-protopanaxatriol (PPT). The developed method was based on ultra high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) detection using multiple-reaction monitoring (MRM) mode. A mass spectrometry sensitizing reagent, 4′-carboxy-substituted rosamine (CSR) with high reaction activity and ionization efficiency was synthesized and firstly used as derivatization reagent. Parameters of dual-UADLLME, MAD and UHPLC-MS/MS conditions were all optimized in detail. Low toxic brominated solvents were used as extractant instead of traditional chlorinated solvents. Satisfactory linearity, recovery, repeatability, accuracy and precision, absence of matrix effect and extremely low limits of detection (LODs, 0.010 and 0.015 ng/mL for PPD and PPT, respectively) were achieved. The main advantages were rapid, sensitive and environmentally friendly, and exhibited high selectivity, accuracy and good matrix effect results. The proposed method was successfully applied to pharmacokinetics of PPD and PPT in rat plasma.
Concentrations of 20 free amino aicds (FAAs) in a famous Tibetan medicine Gentiana dahurica was first investigated using 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC) as the pre-column fluorescence derivatization reagent by reversed-phase high performance liquid chromatography (RP-LC). 20 amino acid derivatives (AAD) were separated on a Hypersil BDS C<sub>18</sub> column with a good baseline resolution within 65 min. Identification of 20 AAD was by online post-column mass spectrometry with an electrospray ionization (ESI) source. The validation of the method was examined by linearity, repeatability, and detection limits. Most linear correlation coefficients for AAD were >0.9990, and detection limits (at signal-to-noise of 3:1) were 6.5-178.2 fmol. There were 18 FAAs found in G. dahurica, of which seven FAAs were necessary to the people's health and related to the treatment of liver and gall disease. Variation of concentrations of the 20 FAAs showed geographical distribution difference among populations. Meanwhile a stable genetic diversity of FAAs composition of G. dahurica was also revealed at the species level. Results of the present study proved that the established method was rapid and reproducible for further separation and determination of FAAs in more medicinal plants.
In this work, a hyphenated technique of dual ultrasound-assisted dispersive liquid-liquid microextraction combined with microwave-assisted derivatization followed by ultra high performance liquid chromatography tandem mass spectrometry has been developed for the determination of phytosterols in functional foods and medicinal herbs. Multiple reaction monitoring mode was used for the tandem mass spectrometry detection. A mass spectrometry sensitive reagent, 4'-carboxy-substituted rosamine, has been used as the derivatization reagent for five phytosterols, and internal standard diosgenin was used for the first time. Parameters for the dual microextraction, microwave-assisted derivatization, and ultra high performance liquid chromatography tandem mass spectrometry were all optimized in detail. Satisfactory linearity, recovery, repeatability, accuracy and precision, absence of matrix effect, extremely low limits of detection (0.005-0.015 ng/mL) and limits of quantification (0.030-0.10 ng/mL) were achieved. The proposed method was compared with previously reported methods. It showed better sensitivity, selectivity, and accuracy. The matrix effect was also significantly reduced. The proposed method was successfully applied to the determination of five phytosterols in vegetable oil (sunflower oil, olive oil, corn oil, peanut oil), milk and orange juice (soymilk, peanut milk, orange juice), and medicinal herbs (Ginseng, Ganoderma lucidum, Cordyceps, Polygonum multiflorum) for the quality control of functional foods and medicinal herbs.
• In situ derivatization-UADLLME was firstly reported for NTs in rat brain microdialysates. • Lissamine rhodamine B sulfonyl chloride was firstly used as derivatization reagent. • The method was simple, rapid, green, efficient, sensitive and low matrix effect. • This method was successfully applied for Parkinson’s rat brain microdialysates.<br>Simultaneous monitoring of several neurotransmitters (NTs) linked to Parkinson’s disease (PD) has important scientific significance for PD related pathology, pharmacology and drug screening. A new simple, fast and sensitive analytical method, based on in situ derivatization-ultrasound-assisted dispersive liquid-liquid microextraction (in situ DUADLLME) in a single step, has been proposed for the quantitative determination of catecholamines and their biosynthesis precursors and metabolites in rat brain microdialysates. The method involved the rapid injection of the mixture of low toxic bromobenzene (extractant) and acetonitrile (dispersant), which containing commercial Lissamine rhodamine B sulfonyl chloride (LRSC) as derivatization reagent, into the aqueous phase of sample and buffer, and the following in situ DUADLLME procedure. After centrifugation, 50 μL of the sedimented phase (bromobenzene) was directly injected for ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) detection in multiple reaction monitoring (MRM) mode. This interesting combination brought the advantages of speediness, simpleness, low matrix effects and high sensitivity in an effective way. Parameters of in situ DUADLLME and UHPLC-MS/MS conditions were all optimized in detail. The optimum conditions of in situ DUADLLME were found to be 30 μL of microdialysates, 150 μL of acetonitrile containing LRSC, 50 μL of bromobenzene and 800 μL of NaHCO3-Na2CO3 buffer (pH 10.5) for 3.0 min at 37 °C. Under the optimized conditions, good linearity was observed with LODs (S/N > 3) and LOQs (S/N > 10) of LRSC derivatized-NTs in the range of 0.002-0.004 and 0.007-0.015 nmol/L, respectively. It also brought good precision (3.2-12.8%, peak area CVs%), accuracy (94.2-108.6%), recovery (94.5-105.5%) and stability (3.8-8.1%, peak area CVs%) results. Moreover, LRSC derivatization significantly improved chromatographic resolution and MS detection sensitivity of NTs when compared with the reported studies through the introduction of a permanent charged moiety from LRSC into NTs. Taken together, this in situ DUADLLME method was successfully applied for the simultaneous determination of six NTs in biological samples.