Multiresidue screening of pesticides in Panax Ginseng C. A. Meyer by ultra-high-performance liquid chromatography with quadrupole time-of-flight mass spectrometry.

In this study, an efficient screening method based on a modified quick, easy, cheap, effective, rugged, and safe extraction method combined with ultra-high-performance liquid chromatography coupled to tandem quadrupole time-of-flight mass spectrometry was established for the determination of 90 pesticides residues in Panax Ginseng. The accuracy of the method was then verified by analyzing the false positive rate and the screening detection limit in Ginseng. The results revealed that the screening detection limit of 33 of 90 pesticide residues were 0.01 mg·kg-1 , 22 species were 0.05 mg·kg-1 , 11 species were 0.10 mg·kg-1 , 8 species were 0.20 mg·kg-1 , and another 16 species were greater than 0.20 mg·kg-1 . A total of 73 pesticides were ultimately suitable to be practically applied for rapid analysis of pesticide residues in Ginseng. Finally, the established method was used to analyze the pesticide residues in 35 Ginseng samples available on the market. And the residual of dimethomorph, azoxystrobin, tebuconazole, and pyraclostrobin was relatively severe in Ginseng samples. This work expanded the range of pesticides detected and provided a rapid, effective method for pesticides screening in Ginseng.

Comprehensive characterization of in vivo metabolic profile of Polygalae radix based on ultra-high-performance liquid chromatography-tandem mass spectrometry.

In this study, a novel analysis strategy for progressively targeted screening and characterization of drug ingredients from in vitro to in vivo was proposed based on ultra-high performance liquid chromatography-tandem mass spectrometry for comprehensive characterization of in vivo metabolic profile of Polygalae radix (PR). First, an in vitro chemical profile of PR was described with the assistance of UNIFI™ software. The characteristic neutral small molecule losses were summarized to distinguish different chemical structures in the PR extract. Second, the in vitro intestinal microflora metabolism model was applied to describe an in vitro metabolic profile of the main ingredients of PR. The metabolic rule and metabolites were integrated for subsequent targeted screening of metabolites in vivo. Finally, an integrated strategy was established and applied to screen and characterize the major absorbed components in vivo, including blood, urine, brain, feces, and liver, based on the prototypes and metabolic rules obtained in vitro. As a result, in vitro and in vivo metabolic profiles of PR were effectively depicted. A total of 136 compounds were isolated and identified from the crude extract in vitro, and 12 compounds were reported for the first time based on the proposed fragmentations. A total of 13, 32, and 3 compounds were identified and characterized in the dosed plasma, liver, and brain, respectively. A total of 40 and 73 compounds were identified in urine and feces, respectively. This strategy not only provided a comprehensive insight into the chemical and metabolic profiles of PR but also presented a new perspective for the discovery of new drugs for medicinal application.

A targeted strategy for analyzing untargeted mass spectral data to identify lanostane-type triterpene acids in Poria cocos by integrating a scientific information system and liquid chromatography-tandem mass spectrometry combined with ion mobility spectrometry.

Rapid structural identification of natural compounds in the crude extract of traditional Chinese medicine by conventional liquid chromatography-mass spectrometry is complex and challenging. In particular, it is difficult to distinguish and identify structural isomers. In this work, we proposed a novel strategy that combines a typical ultrahigh-performance liquid chromatography-multidimensional mass spectrometry approach and the post-processing UNIFI scientific information system to rapidly identify lanostane analogs and isomers in Poria cocos. First, this strategy requires setting up a high-resolution key MS database and an in-house compound library. Then, ultrahigh-performance liquid chromatography coupled with high-resolution tandem data-independent mass spectrometry and ion mobility mass spectrometry was used to acquire untargeted multidimensional mass spectral data. Finally, a new and reliable multidimensional MS analytical workflow was developed to targeted filter the acquired data based on an in-house compound library via the UNIFI™ software. As result, a total of 121 lanostane-type triterpene acids were identified by high-resolution molecular mass, fragment ions, and collision cross-section values. Eight triterpene acids were unambiguously identified by comparing the retention time and MS/MS data with those of reference compounds. Three compounds were detected and reported for the first time based on their neutral losses, characteristic ions, and fragmentation pathways compared with those of known compounds. We anticipate that such an analytical approach can be extended to rapidly screen and characterize other herbal medicine compounds with multiple isomers.

Effect of type 2 diabetes mellitus on flavonoid pharmacokinetics and tissue distribution after oral administration of Radix Scutellaria extract in rats.

Radix Scutellaria is widely applied to the treatment of diabetes mellitus in China. Its main bioactive constituents contain baicalin, wogonoside, oroxyloside, and their aglycones. To investigate the effect of type 2 diabetes mellitus on both pharmacokinetics and tissue distribution of these flavonoid compounds, the six flavonoids in plasma and tissues from the normal and type 2 diabetic rats after oral administration of Radix Scutellaria extract were simultaneously measured by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method. The results showed that baicalin, wogonoside, and oroxyloside had higher Cmax and AUC values (P < 0.05) in type 2 diabetic rats than that in normal rats and the tissue-distribution behaviors of the six flavonoid compounds in hearts, livers, spleens, lungs, kidneys, brains, pancreas, fat and muscle of the type 2 diabetic rats showed obviously differences from the normal rats (P < 0.05). In conclusion, the differences in the pharmacokinetics of oroxyloside and tissue distribution of the six flavanoids in Radix Scutellaria extract between diabetic and normal rats were found for the first time. The results from the present study provided a crucial basis for a better understanding of in vivo anti-diabetic mechanism of action of the six flavonoids from Radix Scutellaria.

Studies on the chemical and intestinal metabolic profiles of Polygalae Radix by using UHPLC-IT-MSn and UHPLC-Q-TOF-MS method coupled with intestinal bacteria incubation model in vitro.

Polygala Radix (PR) is one of the most commonly used traditional Chinese medicines (TCM), but its effective materials are still not very clear. In order to solve the existing problems, the chemical and intestinal metabolic profiles of PR were analyzed via intestinal bacteria incubation model. An integrated analysis method combined ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF MS) method and ultra-high performance liquid chromatography with ion-trap multi-stage tandem mass spectrometry (UHPLC-IT-MSn) method was established. As a result, a total of 115 polygala compounds were identified from the PR extract. Moreover, 44 metabolites via intestinal microflora were characterized, of which three compounds were formed from xanthone C-glycosides, 25 compounds were formed from polygala saponins and 16 compounds were formed from oligosaccharide multi-esters. To our knowledge, the metabolites of the PR via intestinal microflora metabolism, especially polygala saponins, are the first reported in vitro. The obtained results would provide the methodological support for further studying the effective materials of the PR in vivo.

[Application of modern mass spectrometry in metabolism and pharmacokinetics of traditional Chinese medicine].

Modern mass spectrometry plays an important role in the study of metabolism and pharmacokinetics of traditional Chinese medicine due to its high sensitivity, high analysis speed and good stability. First, this article gives a brief introduction to the development of modern mass spectrometry techniques. Second, the application of modern mass spectrometry is summarized in the study of metabolism and pharmacokinetics of traditional Chinese medicine including in vitro and in vivo studies. In vitro research is reviewed with a focus on metabolism of intestinal bacteria and cytochrome P450 enzymes; in vivo research is covered mainly with a focuse on the application of microdialysis-MS, GC/LC-MS, new ambient mass spectrometry, and imaging mass spectrometry in metabolism and pharmacokinetics.

[Study on Therapeutic Effects of Ermiao Pill and Ermiao Pill Categorized Formula in Hyperuricemic Rats Using Spectroscopic Methods].

A stable mouse model of hyperuricemia was established by intraperitoneal injection of xanthine and oxonate, comparing the water extracts (containing crude drug 360 mg · mL(-1)) of Ermiao pill categorized formula (The ratio of atractylodes lancea to cortex phellodendri was 1:1, 1:2 and 2:1, respectively) and Ermiao pill (360 mg · mL(-1)) administered to different groups of animals continuously for two weeks and assessing the protection or treatment of drug on hyperuricemia. The xanthine oxidase (XOD) activities in serum and liver were detected by ultraviolet-visible spectroscopy at 570 nm wavelength, The results showed that as compared with each group, the XOD activity of the model group was significantly increased in serum (p < 0.01), XOD activity showed no significant difference in liver (p > 0.05), but compared with the model group, the XOD activity of each treatment group was significantly lower in serum and liver (p < 0.01), especially for the group treated with Ermiao pill categorized formula with the ratio of Atractylodes lancea to Cortex Phellodendri being 1:2. The morphological changes of glomerular and tubular interstitial fibrosis were measured by Hematoxylin-eosin staining (HE staining) and Masson trichrome staining (Masson staining)for kidney paraffin sections. The results showed that the glomerular atrophy, vascular loops confusion, a certain degree of inflammatory cell infiltration, interstitial fibrosis and other phenomena appeared in the model group. Compared to model group, these pathological phenomena of the treatment groups were significantly improved. The area showed that compared with each group, the fibrosis of the model group was significantly increased (p < 0.01 or p < 0.05), but compared with Ermiao pill categorized formula, the differences for the group of Ermiao pill was lower (p < 0.01), especially for the group treated with Ermiao pill categorized formula with the ratio of Atractylodes lancea to Cortex Phellodendri being 1 : 2. In this experiment, the damage of kidney and XOD activity serve as the index to evaluate the protection or treatment of drug on hyperuricemia, providing a scientific basis for the development of Ermiao pill categorized formula.

[Investigation on Inhibitory Capacities of Seventeen Herbal Extracts on Oxidative Stress using Ultraviolet and Fluorescence Spectroscopy].

Diabetic patients usually suffer from complications and the long-term secondary complications are the main cause of morbidity and mortality. The hyperglycemia-induced oxidative stress is one of the important pathogenesis of diabetic complications, while the oxidative stress is associated with the lipid peroxidation reaction and the formation of advanced glycation end products (AGEs). Our study was focus on the pathogenesis of diabetic complications and based on the oxidative stress reaction. In this research, the oxidative stress inhibiting effects of seventeen herbal extracts were studied based on spectroscopic methodology. The capacities of herbal extracts against the lipid peroxidation reaction of rat liver in vitro were investigated using spectrophotometric method. It showed that the inhibitory activity of Radix Scutellariae and Flos Sophorae Immaturus were better than other herbal extracts. Additionally, the herbal extracts rich in flavonoids, alkaloids and lignanoids showed good inhibitory activities on the lipid peroxidation reaction. On the contrary, the saponin-rich herbal extracts possessed weak inhibitory effects. We applied the BSA/glucose (fructose) system combined with fluorescent spectroscopy to determine the inhibitory activities of herbal extracts in glycation model reactions. The results showed that the AGEs formation inhibitory activity of Flos Sophorae Immaturus, Radix Scutellariae and Rhizoma Anemarrhenae were better than others in the BSA/glucose (fructose) system by fluorescene analysis. The results demonstrated that the herbal extracts rich in flavonoids were found to be more effective than that of those herbal extracts as alkaloids and terpenoids class in inhibiting oxidative stress, while the saponin-rich herbal extracts showed weak inhibitory activities against oxidative stress. The Flos Sophorae Immaturus and Radix Scutellariae extracts had better inhibitory activity to the oxidative stress, so their pharmacological activity could be explored in further investigations. These results demonstrated in this assay could provide a reference for the study of pharmacological activity, and thus lays the foundation for the further study of the application of natural products in the prevention and treatment of diabetic complications.

[High performance preparation and structural confirmation of lignans from Schisandrae chinensis fructus by using HSCCC combined with ESI-MSn method].

High-speed counter-current chromatography (HSCCC) was used to high performance separate and prepare lignans from Schisandrae chinensis fructus. The solvent system is composed of n-hexane-ethyl acetate-methanol-water (9 : 1 : 5 : 5) and n-hexane-ethyl acetate-methanol-water (9 : 1 : 9 : 5), speed is at 900 r.min-1, and flow rate is at 2.0 mL.min-1. Five fractions from Schisandrae chinensis fructus extract were separated and prepared with one HSCCC process. They were identified as schisandrin, gomisin J, schisandrol B, schisantherin A and deoxyschizandrin by electrospray ionization-multiple tandem mass spectrometry (ESI-MSn), respectively. Their contents were obtained in 98.74%, 94.32%, 99.53%, 94.23% and 98.68% by ultra high performance liquid chromatography (UPLC), separately. The rapid and simple method can be applied for the preparation of lignans from Schisandrae chinensis fructus.

[The metabolic fingerprint of the compatibility of Radix Aconite and Radix Paeoniae Alba and its effect on CYP450 enzymes].

Using a UPLC-MS/MS (MRM) and cocktail probe substrates method, the metabolic fingerprint of the compatibility of Radix Aconite (RA) and Radix Paeoniae Alba (RPA) and its effect on CYP450 enzymes were investigated. These main CYP isoforms include CYP 1A2, CYP 2C, CYP 2E1, CYP 2D and CYP 3A. Compared with the inhibition effect of RA decoctions on CYP450 isoforms, their co-decoctions of RA and RPA with different proportions can decrease RA' inhibition on CYP3A, CYP2D, CYP2C and CYP1A2, but can not reduce RA' effect on CYP2E1. The metabolic fingerprints of RA decoction and co-decoctions with different proportions of RPA in CYP450 of rat liver were analyzed by UPLC-MS. Compared with the metabolic fingerprints of RA decoction, the intensity of diester-diterpenoid aconitum alkaloids decreased significantly, while the intensity of monoester-diterpenoid alkaloids significantly increased in the metabolic fingerprints of co-decoctions of RA and RPA. The results suggest that RA coadministration with RPA increased the degradation of toxic alkaloid and show the effect of toxicity reducing and efficacy enhancing.

[Effects of six kinds of Chinese herb extracts on the activities of rat liver microsomes in vitro].

Effects of six kinds of Chinese herb extracts, including Folium Crataegi extract, Herba Epimedii extract, Folium Acanthopanacis Senticosi extract, Trifolium pratense L. extract, Folium Ginkgo extract and Radix Puerariae extract, on the activities of CYP450 isozymes (CYP1A2, CYP2C, CYP2E1, CYP2D, CYP3A) in rat hepatic microsomals were studied by using a UPLC-MS/MS (MRM) and cocktail probe substrates method. The results showed that effects of six kinds of Chinese herb extracts on each CYP450 isozyme activity were inhibitory. The IC50 of Folium Crataegi extract for the inhibition of rat microsomal CYP2D activity was only for 4.04 microg x mL(-1), which showed the highest inhibition; Trifolium pratense L. extract had strong inhibitory action to CYP2D, the IC50 value was 5.73 microg x mL(-1); Folium Crataegi extract also had strong inhibitory action on CYP2E1, the IC50 value was 10.91 microg x mL(-1). Furthermore, the IC50 of Folium Ginkgo extract for the inhibition of rat microsomal CYP3A, 2D, 2E1 activities were 45.12, 35.45 and 22.41 microg x mL(-1), respectively, and the IC50 of Folium Acanthopanacis Senticosi extract on the inhibition of rat microsomal CYP2E1 activity was 32.89 microg x mL(-1). In addition, mechanism of inhibition experimental results showed that the inhibiting abilities of Folium Crataegi extract and Radix Puerariae extract on each CYP450 isozyme increased with the increasing of the preincubation time, therefore, the inhibitory effects were a mechanism-based inhibition.

[Metabolites and metabolic pathways of mesaconitine in rat liver microsomal investigated by using UPLC-MS/MS method in vitro].

Mesaconitine was incubated with rat liver microsomes in vitro. The metabolites of mesaconitine in rat liver microsomes were identified by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method with high resolution power. A typical reaction mixture of 100 mol L-1 Tris-HCI buffer (pH 7.4) containing 0.5 gL-1 microsomal protein and 50 micro molL-1 mesaconitine was prepared. The above reaction mixture was divided into six groups, and the volume of each group was 200 micro L. The incubation mixture was pre-incubated at 37 degrees C for 2 min and the reactions were initiated by adding NADPH generating system. After 90 min incubation at 37 degrees C, 200 micro L of acetonitrile was added to each group to stop the reaction. The metabolites of mesaconitine were investigated by UPLC-MS/MS method. Mesaconitine and 6 metabolites M1-M6 were found in the incubation system. The structures were characterized according to the data from MS/MS spectra and literatures. The metabolic reactions of mesaconitine in rat liver microsomes included the demethylation, deacetylation, dehydrogenation and hydroxylation. The major metabolic pathways of mesaconitine in rat liver microsomes were determined by UPLC-MS/MS on multiple reaction monitoring (MRM) mode combined with specific inhibitors of cytochrome P450 (CYP) isoforms, including alpha-naphthoflavone (CYP1A2), quinine (CYP2D), diethyldithiocarbamate (CYP2E1), ketoconazole (CYP3A) and sulfaphenazole (CYP2C), separately. Mesaconitine was mainly metabolized by CYP3A. CYP2C and CYP2D were also more important CYP isoforms for the metabolism reactions of mesaconitine, but CYP1A2 and CYP2E1 haven't any contribution to MA metabolism in rat liver microsomes.

Studies on structures and activities of initial Maillard reaction products by electrospray ionisation mass spectrometry combined with liquid chromatography in processing of red ginseng.

Electrospray ionisation-mass spectrometry (ESI-MS) was applied to analyse the water-soluble extract of red ginseng (RG). Several new compounds were produced from the Maillard reaction during the steaming and drying process for preparing RG. Both the tandem electrospray ionisation (ESI-MS(n)) and Fourier transform ion cyclotron resonant mass spectrometric (FT-ICR-MS) data of these products proved that they were the initial Maillard reaction products (MRPs) of maltose with glutamic acid/aspartic acid, which were specific components in RG. In addition, their anti-diabetic and antioxidant activities were examined in vitro. The anti-diabetic activities were evaluated by studying the α-glucosidase inhibition using ultrafiltration LC-MS/MS techniques, while the antioxidant activities were investigated by UPLC-ESI-MS method. The results demonstrated that four initial MRPs in RG were identified as α-glucosidase inhibitors, and showed marked scavenging effect on the hydroxyl radical ((·)OH). Based on these studies, the processing method of RG was improved to generate more active compounds.

Formation of tungstate-containing cluster ions by polyoxotungstate anions under matrix-assisted laser desorption/ionization conditions in the gas phase.

The gas-phase studies of transition-metal oxides continue to attract interest as such oxides are being used as catalysts in various oxidation processes. In this paper, singly negatively charged heteropolyoxotungstate and isopolyoxotungstate ion clusters were produced from Keggin-type polyoxotungstates by matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FTICR MS). It was found that the ion series [(PO(3))(WO(3))(n)](-), [(WO(3))(n)](-) and [(OH)(WO(3))(n)](-) were the main fragment ions in the mass spectra and the matrix greatly influenced the resulting cluster ion abundances. [(PO(3))(WO(3))(3)](-), [(WO(3))(3)](-) and [(OH)(WO(3))(4)](-) were the most intense ions in each series when 2-(4-hydroxyphenylazo)benzoic acid was the matrix, whereas [(PO(3))(WO(3))(4)](-), [(WO(3))(6)](-) and [(OH)(WO(3))(4)](-) were the most intense when dithranol (DIT) was the matrix. In addition, a new kind of hybrid ion [W(2)C(14)H(7)O(8)](-) was produced through the reaction of DIT and [(OH)(WO(3))](-) in the plume of the gas phase. These results highlight the utility of the MALDI-FT method for obtaining novel ion clusters and also show the stability of these clusters.

[Determination of content changes of three diester-diterpenoid alkaloids in compatibility of radix aconiti laterlis preparata and Rhizoma pinelliae, Fructus trichosanthis, Bulbus fritillariae, Radix ampelopsis, Rhizoma bletillae by HPLC].

OBJECTIVE: To analyze the three diester-alkaloids content in the decoctions before and after Radix aconiti lateralis preparata combined with Rhizoma pinelliae, Fructus trichosanthis, Bulbus fritillariae, Radix ampelopsis, Rhizoma bletillae, respectively. METHODS: HPLC analysis was performed on a Agilent Extend C18 column,eluted with a mobile phase consisted of acetonitrile/methanol - 35 mmol/L ammonium acetate and gradient elution,with a flow rate of 0.6 ml/min and the detection wavelength of 235 nm. RESULTS: The contents of three diester-alkaloids in the co-decoctions of Radix aconiti laterlis preparata combined with Rhizoma pinelliae, Fructus trichosanthis, Bulbus fritillariae, Rhizoma bletillae were more than that of single Radix aconiti laterlis preparata decoction, expect the Bulbus fritillariae cirrhosae,Radix ampelopsis and Rhizoma pinelliae praeparatum. From the result,we can found that the content of three diester-alkaloids in decoctions was correlated with the decoction pH. CONCLUSION: The pH of decoction is one of the most important factors to relate the three diester-alkaloids content in the decoctions.

[Change of the content of chemical constituents and anti-oxidative action of the decoction of radix ginseng combined with Flos Lonicerae, radix Polygoni Multiflori and Radix Astragali].

Ginsenosides in the decoction of Radix Ginseng, Radix Ginseng with Flos Lonicerae, Radix Polygoni Multiflori or Radix Astragali have been investigated by high performance liquid chromatography (HPLC) and electrospray ionization mass spectrometric method (ESI-MS). Change of the content of ginsenosides was nonlinear in diverse combinative proportion of Radix Ginseng with Flos Lonicerae, while the stripping of ginsenosides was promoted by a small amount of Radix Polygoni Multiflori. In the combinative decoction of Radix Ginseng with Radix Astragali, ginsenosides contents were increased compared to single decoction of Radix Ginseng. Besides, ferric reducing antioxidant power (FRAP) method was developed for determination of the total antioxidative activity of n-butanol and water-soluble extracts from the decoction. The experimental results showed that antioxidative activity was better in the combinative decoction than that in single decoction, and the FRAP values of n-butanol extract were also greater compared with that of water extract.

[Lignans from different processed products of Schisandra chinensis fruits].

To study the lignan components of different processed products of Schisandra chinensis fruits, the study was conducted with electrospray ionization mass spectrometry (ESI-MS) and high performance liquid chromatography (HPLC). There was no new lignan components detected from the products obtained with different processed methods, the difference was in the content of lignan components. The method is accurate and effective, which can be used to evaluate the quality of different processed products of Schisandra chinensis fruits.