Simultaneous and dynamic measurement of Schisandrol A changes in rat blood and brain and its comparative pharmacokinetic study in control and Parkinson's disease rats by dual-probe in vivo microdialysis.

Schisandrol A (SchA) is the main active ingredient of Schisandra chinensis (Turcz.) Baill., which is a famous traditional Chinese herbal medicine. SchA can penetrate the blood-brain barrier and has a significant neuroprotective effect. A group of multiplexed stable isotope mass tags (MSIMTs, m/z 332, 338, 346, 349, 351, 354, 360, 363, 374 and 377) were synthesized to perform multiplexed stable isotope labeling derivatization (MSILD) of SchA in rat microdialysates and standards. A new magnetic molecularly imprinted polymer was prepared using MSIMT-375-SchA as dummy template. All the 10-plexed derivatives of MSIMTs-SchA can be efficiently and selectively enriched and purified using this adsorbent by magnetic dispersive solid phase extraction (MDSPE) before ultra high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) analysis. It should be pointed out that the MSIMT-346-SchA standard derivative was used as internal standard in the process of MDSPE and UHPLC-MS/MS. On these bases, 9 different rat microdialysate samples can be determined by UHPLC-MS/MS in a single run. The utilization of MSIMTs significantly increased the sensitivity, accuracy, selectivity and analysis throughput. Under the optimized conditions, satisfactory linearity (R2> 0.987), limit of detection (LODs, 0.15-0.26 pg/mL) and lower limit of quantitative (LLOQ, 0.8-2.0 pg/mL) were obtained. Intra- and inter-day precisions were in the range of 2.2% -12.5%, and recoveries 94.2% -106.2%. The matrix effects were very low, and the average derivatization efficiency of 10-plex MSIMTs to SchA was as high as 97.8%. Using the developed dual-probe in vivo microdialysis sampling technique, the proposed analytical method has been applied for comparative pharmacokinetics of SchA in the brain and blood of control and Parkinson's disease (PD) rats.

A fluorescence strategy for monitoring α-glucosidase activity and screening its inhibitors from Chinese herbal medicines based on Cu nanoclusters with aggregation-induced emission.

Herein, the self-assembly of 1-dodecanethiol-capped Cu nanoclusters (DT-Cu NCs) is obtained by annealing of dibenzyl ether solution of nanoclusters. These aggregates are composed of small clusters and emit a high level of aggregation-induced emission (AIE) in water. Based on the quenching effect of 4-nitrophenol (4-NP) on DT-Cu NCs, a fluorescence strategy is developed to monitor α-glucosidase (α-Glu) activity and screen its inhibitors from Chinese herbal medicines. 4-Nitrophenyl-α-D-glucopyranoside (NGP) is selected as the substrate, which is further hydrolyzed to yield 4-NP through the catalysis of α-Glu. The quenching efficiency is positively correlated to the concentration of α-Glu. Furthermore, the inhibitory effects of the extracts from four Chinese herbal medicines (i.e., the rind of Punica granatum L., Momordica grosvenorii Swingle., Crataegus pinnatifida Bge., and Lycium barbarum L.) on the α-Glu activity have been studied. The IC50 values of extracts from the rind of Punica granatum L. and Momordica grosvenorii Swingle are 0.23 and 0.37 g/L, respectively, so they show obvious inhibitory effects on α-Glu. The extracts of Crataegus pinnatifida Bge. and Lycium barbarum L. exhibit relatively weak inhibitory effects. Hence, the proposed strategy can be applicable for screening α-Glu inhibitors from Chinese herbal medicines. Last but not the least, by immobilizing DT-Cu NCs into agarose hydrogels in polyethylene tubes, a visual device is fabricated to screen α-Glu inhibitors with high throughput and sensitivity.

Simultaneous Determination of Oleanolic Acid and Ursolic Acid by in Vivo Microdialysis via UHPLC-MS/MS Using Magnetic Dispersive Solid Phase Extraction Coupling with Microwave-Assisted Derivatization and Its Application to a Pharmacokinetic Study of Arctiumlappa L. Root Extract in Rats.

Simultaneous detection of oleanolic acid and ursolic acid in rat blood by in vivo microdialysis can provide important pharmacokinetics information. Microwave-assisted derivatization coupled with magnetic dispersive solid phase extraction was established for the determination of oleanolic acid and ursolic acid by liquid chromatography tandem mass spectrometry. 2'-Carbonyl-piperazine rhodamine B was first designed and synthesized as the derivatization reagent, which was easily adsorbed onto the surface of Fe3O4/graphene oxide. Simultaneous derivatization and extraction of oleanolic acid and ursolic acid were performed on Fe3O4/graphene oxide. The permanent positive charge of the derivatization reagent significantly improved the ionization efficiencies. The limits of detection were 0.025 and 0.020 ng/mL for oleanolic acid and ursolic acid, respectively. The validated method was shown to be promising for sensitive, accurate, and simultaneous determination of oleanolic acid and ursolic acid. It was used for their pharmacokinetics study in rat blood after oral administration of Arctiumlappa L. root extract.

A simple and novel colorimetric assay for tyrosinase and inhibitor screening using 3,3',5,5'-tetramethylbenzidine as a chromogenic probe.

A novel colorimetric method for the detection of tyrosinase (TYR) and its inhibitor by taking utilization of Ag+-3,3',5,5'-tetramethylbenzidine (TMB) detection system has been proposed. Ag+ could oxidize TMB to oxidized TMB (oxTMB) and induce a blue color solution corresponding to an absorption peak centered at 652nm. The addition of dopamine (DA) could cause the reduction of oxTMB which resulted in the fading of the blue color and a decrease of the absorbance at 652nm. However, in the presence of TYR, DA could be oxidized to dopaquinone, which inhibited the reduction of oxTMB by DA, resulting in a blue color recovery and an increase of the absorbance at 652nm. Based on this finding, we propose a method to quantitatively detect TYR activity with the help of UV-vis spectroscopy. The developed assay is highly sensitive with a low detection limit of 0.010U/mL. More importantly, this method is fairly simple and inexpensive without the use of complicated nanomaterials. In addition, it constructs a useful platform for TYR inhibitor screening.

Rapid and sensitive determination of phytosterols in functional foods and medicinal herbs by using UHPLC-MS/MS with microwave-assisted derivatization combined with dual ultrasound-assisted dispersive liquid-liquid microextraction.

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-ultrasound-assisted dispersive liquid-liquid microextraction for the determination of neurotransmitters in Parkinson's rat brain microdialysates by ultra high performance liquid chromatography-tandem mass spectrometry.

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, 50µL 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 30µL of microdialysates, 150µL of acetonitrile containing LRSC, 50µL of bromobenzene and 800µL of NaHCO3-Na2CO3 buffer (pH 10.5) for 3.0min 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.

[Determination of L-dopa and dopamine in rat brain microdialysate by ultra high performance liquid chromatography-tandem mass spectrometry using stable isotope-coded derivatization coupled with dispersive liquid-liquid microextraction].

The sensitive detection method of levodopa (L-DOPA) and dopamine (DA) in rat brain microdialysate of Parkinson's disease (PD) is an essential tool for the clinical study and attenuated synergistic drug screening for L-DOPA from traditional Chinese medicines. Using d0/d3-10-methyl-acridone-2-sulfonyl chloride (d0/d3-MASC) as stable isotope derivatization reagent, a novel ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for L-DOPA and DA by stable isotope- coded derivatization coupled with ultrasonic-assisted dispersive liquid-liquid microextraction (UA-DLLME). d3-MASC (light) and d3-MASC (heavy) were used as derivatization reagents for microdialysate samples and standards, respectively. Mixtures of the two solutions were prepared by UA-DLLME for UHPLC-MS/MS analysis with multiple reaction monitoring (MRM) mode. With d3-MASC heavy derivatives as internal standards for corresponding light derivatives from samples, the stable isotope internal standard quantification for L-DOPA and DA was carried out. The stable derivatives were obtained in aqueous acetonitrile (pH 10.8 sodium carbonate-sodium bicarbonate buffer) at 37 °C for 3.0 min, and then were separated within 2.0 min using gradient elution. Linear range was 0.20-1500.0 nmol/L (R > 0.994). LODs were 0.005 and 0.009 nmol/L for DA and L-DOPA (S/N = 3), respectively. This method was validated, and it showed obvious advantages in comparing with the reported methods in terms of sensitivity, analysis speed and anti-matrix interference. This method has been successfully applied to the study of effect of Shouwu Fang on L-DOPA and DA concentration fluctuations in PD rat brain microdialysate.

A combination of ultrasonic-assisted extraction with RRLC-QQQ method for the determination of artemisinin in the Chinese herb Artemisia annua L.

INTRODUCTION: Artemisinin, the primary active ingredient of the Chinese herb Artemisia annua L., is known to have considerable anti-malaria properties. However, rapid, sensitive and selective method for the determination of artemisinin in it is not currently available. OBJECTIVE: To develop and validate an efficient method for extraction and analysis of artemisinin from the plant samples of Artemisia annua L. by rapid resolution liquid chromatography triple quadrupole mass spectrometry (RRLC-QQQ). METHODOLOGY: Following ultrasound-assisted extraction (USE), RRLC-QQQ was utilised to separate and determine artemisinin from the plant sample of Artemisia annua L. The LC separation, QQQ-MS detection and multiple reaction monitoring (MRM) mode were optimised, and the method validation concluding selectivity, calibration, accuracy and precision, and recovery were also evaluated. RESULTS: LC separation was performed with an isocratic elution of 20% of methanol-water (10 mmol/L ammonium acetate, pH 4.0) on a C(18) column. The triple quadrupole MS detection was carried out under MRM mode of precursor ion [M + H]+ → fragment ions m/z 265.1 and m/z 247.2. The limits of detection and quantitation of artemisinin were 0.20 and 0.75 ng/mL, respectively. The intra- and inter-day precisions did not exceed 3.71%, and the deviation of the intra- and inter-day mean values did not exceed ±7.50. The average recoveries for artemisinin ranged from 92.45 to 103.8% with an RSD from 2.47 to 2.79%. CONCLUSION: The developed RRLC-QQQ assay is an efficient method for separation and determination of artemisinin from the plant samples of Artemisia annua L.