Characteristic identification of Sophorae Flavescentis Radix and Sophorae Tonkinensis Radix et Rhizoma based on HPTLC-ESI-MS / 药学学报

Sophorae Flavescentis Radix is the dried root of Sophora flavescens Ait. and Sophorae Tonkinensis Radix et Rhizoma is the dried root and rhizome of Sophora tonkinensis Gagnep. The two drugs are both from the same genus Sophora, having similar and different compositions and efficacies, however, their differences are not fully demonstrated in current standard. In this study, the high-performance thin-layer chromatography with multi-dimensional and multi-level features combined with electric spray mass spectrometry (HPTLC-ESI-MS) was used to discover and identify the characteristic zones in extracts of Sophorae Flavescentis Radix and Sophorae Tonkinensis Radix et Rhizoma, after optimizing the preparation method of the test solution and chromatographic parameters. As a result, 17 main characteristic zones were found on HPTLC chromatograms of Sophorae Flavescentis Radix and Sophorae Tonkinensis Radix et Rhizoma, among them, besides 3 known chemicals, another 12 unknown components were identified by HPTLC-ESI-MS, they are 1 alkaloid and 11 flavonoids. The identification results were verified by the reference standards partially and nuclear magnetic resonance spectra after guided-isolation. Finally, a unified HPTLC specific identification method with different markers was established to identify Sophorae Flavescentis Radix and Sophorae Tonkinensis Radix et Rhizoma simultaneously. Thanks to abundant chemical information provided when using diverse polarity mobile phases and derivatization reagents, the HPTLC technology offers a convenient strategy for discovery, quality evaluation, and identification of target chemicals when connecting with mass spectrometry.

Quantitative determination of multi-class bioactive constituents for quality control of Yiqi Jiangzhi Granules / 中草药·英文版

Objective: To establish a reliable and sensitive method for evaluating quality of Yiqi Jiangzhi Granules (YQJZG). Methods: Ultra performance liquid chromatography electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) was employed for simultaneous determination of eight marker components. Separation was performed on an AQUITY UPLC® HSS T3 column, the mobile phase consisted of acetonitrile as the organic phase and 0.1% (volume percentage) formic acid as the aqueous. Eight marker components, ginsenoside Rg1 (GRg1), ginsenoside Re (GRe), ginsenoside Rb1 (Gb1), typhaneoside (TEO), isorhamnetin-3-O-neohespeidoside (IN), hesperidin (HPD), aurantio-obtusin-6-O-β-D-glucoside (AG) and curcumin (CCM), were detected by multiple reaction monitoring (MRM) mode. The Chinese Pharmacopoeia (2020 edition) was regarded as the guidance document for this method validation. Results: The method showed good linearity (R

New insights on the utilization of ultrasonicated mustard seed cake: chemical composition and antagonistic potential for root-knot nematode, / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

This study focused, for the first time, on the effect of ultrasonic features on the extraction efficiency of secondary metabolites in mustard seed cake (MSC). The nematostatic potential of sonicated seed cake was examined against the second-stage juveniles (J2s) of root-knot nematode,

Novel neuroprotective peptides in the venom of the solitary scoliid wasp Scolia decorata ventralis

Background Solitary wasp venoms may be a rich source of neuroactive substances, since their venoms are used for paralyzing preys. We have been exploring bioactive constituents of solitary wasp venoms and, in this study, the component profile of the venom from a solitary scoliid wasp, Scolia decorata ventralis, was investigated through a comprehensive analysis using LC-MS. Two peptides were synthesized, and their neuroprotective properties were evaluated. Methods A reverse-phase HPLC connected to ESI-MS was used for LC-MS analyses. Online mass fingerprinting was performed from TIC, and data-dependent tandem mass spectrometry gave the MS/MS spectra. The sequences of two major peptide components were determined by MALDI-TOF/TOF MS analysis, confirmed by solid phase synthesis. Using the synthetic peptides, biological activities were assessed. Cell integrity tests and neuroprotection analyzes using H2O2 as an oxidative stress inducer were performed for both peptides. Results Online mass fingerprinting revealed that the venom contains 123 components, and the MS/MS analysis resulted in 33 full sequences of peptide components. The two main peptides, α-scoliidine (DYVTVKGFSPLR) and β-scoliidine (DYVTVKGFSPLRKA), present homology with the bradykinin C-terminal. Despite this, both peptides did not behave as substrates or inhibitors of ACE, indicating that they do not interact with this metallopeptidase. In further studies, β-scoliidine, but not α -scoliidine, showed protective effects against oxidative stress-induced neurotoxicity in PC12 cells through integrity and metabolism cell assays. Interestingly, β-scoliidine has the extension of the KA dipeptide at the C-terminal in comparison with α-scoliidine. Conclusion Comprehensive LC-MS and MS/MS analyses from the Scolia decorata ventralis venom displayed the component profile of this venom. β-scoliidine showed an effective cytoprotective effect, probably due to the observed increase in the number of cells. This is the first report of solitary wasp venom peptides showing neuroprotective activity.(AU)

Hypolipidemic, hipoglycemiant and antioxidant effects of Myrcia splendens (Sw) DC in an animal type 2 diabetes model induced by streptozotocin / Efectos hipolipemiante, hipoglucemiante y antioxidante de Myrcia splendens (Sw) DC en un modelo animal de diabetes tipo 2 inducido por estreptozotocina

We investigated the effects of dichloromethane extract (DME) from Myrcia splendenson alterations caused by type 2 diabetes in the blood and kidney of rats, in order to reduce side effects caused by synthetic drugs. Rats received streptozotocin (60 mg/kg),15 minutes after nicotinamide (120 mg/kg) or water. After 72 hours, the glycemic levels were evaluated to confirm diabetes and the animals received (15 days) DME (25, 50, 100 or 150 mg/Kg) or water. DME partially reversed hyperglycemia and (100 and 150 mg/kg) reversed hypertriglyceridemia. Histopathological findings elucidated that DME reduced damage to pancreatic islets. DME 150 mg/kgreversed the increases in TBA-RS, the reduction in the sulfhydryl content, 100 and 150 mg/kg increased CAT, reversed the decrease in GSH-Px and increased it activity in the blood. DME 150 mg/kg reversed CAT and GSH-Px reductions in the kidney. We believe that DME effects might be dependent on the presence of phenolic compounds.

Investigamos los efectos del extracto de diclorometano (DME)de Myrcia splendens sobre las alteraciones causadas por la diabetes tipo 2 en la sangre y los riñones de las ratas, para reducir los efectos secundarios causados por las drogas sintéticas. Las ratas recibieron estreptozotocina (60 mg/kg), 15 minutos después de la nicotinamida (120 mg/kg) o agua. Después de 72 horas, se confirmo la diabetes y los animales recibieron (15 días) DME (25, 50, 100 o 150 mg/Kg) o agua. DME revierte parcialmente la hiperglucemia y revierte la hipertrigliceridemia. DME redujo el daño a los islotes pancreáticos. DME revirtió los aumentos en TBA-RS, la reducción en el contenido de sulfhidrilo, aumentó la CAT, revirtió la disminución en GSH-Px y aumentó su actividad en la sangre. Además, DME revirtió las reducciones de CAT y GSH-Px en el riñón. Creemos que los efectos provocados por DME pueden depender de la presencia de compuestos fenólicos.

Simultaneous determination of fourteen components of Gumiganghwal-tang tablet in human plasma by UPLC-ESI-MS/MS and its application to pharmacokinetic study / 药物分析学报

Gumiganghwal-tang is a traditional herbal medicine widely used for its anti-inflammatory,analgesic,and antipyretic effects.However,the safety and efficacy of its active ingredients based on an in vivo pharmacokinetic (PK) study have yet been investigated.We have established a sensitive and accurate UPLC-ESI-MS/MS method and conducted a PK study on 14 constituents of Gumiganghwal-tang through human plasma analysis.Analytical conditions were optimized according to the physicochemical prop-erties of the 14 compounds to facilitate efficient separation and eliminate overlap or interference be-tween peaks.KINETEX-C18 and lnertsil-C8 columns were used as UPLC stationary phases,and acetonitrile and aqueous formic acid were used as mobile phases.All the analytes were quantified with a triple quadrupole mass spectrometer using electrospray ionization in multiple reaction monitoring mode.The chromatograms of 14 bioactive compounds showed excellent elution and sensitivity,and each peak was selectively separated and quantified without interference with each other or impurities.The established analytical method was based on international guidelines and was successfully used to perform PK studies of 14 herbal ingredients in humans after oral administration with Gumiganghwal-tang tablets.The oral absorption of most active components of Gumiganghwal-tang was relatively rapid and remained considerably long in the body to be quantified in plasma up to 48 h after administration.

Pharmacokinetic comparison with different assays for simultaneous determination of cis-,trans-cefprozil diastereomers in human plasma / 药物分析学报

The purpose of this study was to compare pharmacokinetic (PK) parameters obtained using two newly developed assays,HPLC-UV and UPLC-ESI-MS/MS.Selection of assay and results obtained therefrom are very important in PK studies and can have a major impact on the PK-based clinical dose and usage settings.For this study,we developed two new methods that are most commonly used in biosample analysis and focused on PK parameters obtained from them.By HPLC-UV equipped with a Luna-C8 column using UV detector,cefprozil diastereomers were separated using water containing 2％ (V/V) acetic acid and acetonitrile as a mobile phase.By UPLC-ESI-MS/MS equipped with a HALO-C18column,cefprozil diastereomers were separated using 0.5％ (V/V) aqueous formic acid containing 5 mM ammonium-formate buffer and methanol as a mobile phase.Chromatograms showed high resolution,sensitivity,and selectivity without interference by plasma constituents.Both intra-and inter-day precisions (CV,％)were within 8.88％ for HPLC-UV and UPLC-ESI-MS/MS.Accuracy of both methods was 95.67％-107.50％.These two analytical methods satisfied the criteria of international guidance and could be successfully applied to PK study.Comparison of PK parameters between two assays confirmed that there is a dif-ference in the predicted minimum plasma concentrations at steady state,which may affect clinical dose and usage settings.Furthermore,we confirmed possible correlation between PK parameters and various biochemical parameters after oral administration of 1000 mg cefprozil to humans.

Systematic analysis on chemical constituents of Mori Cortex, mulberry root bark and its phellem layer based on HPLC-ESI-MS / 中国中药杂志

In this study, HPLC-ESI-MS and HPLC methods were established to explore the differences in the main chemical components and content of Mori Cortex with(mulberry root bark) and without(Mori Cortex) the phellem layer from both qualitative and quantitative aspects. The HPLC-ESI-MS method was used for quality analysis in positive and negative ion modes, and 33 compounds were identified in mulberry root bark, 22 compounds in Mori Cortex, and 26 compounds in phellem layer; mulberry root bark and Mori Cortex shared 22 components, and mulberry root bark has 11 unique compounds; Mori Cortex and its phellem layer shared 15 components, while Mori Cortex has 7 unique compounds. HPLC method was used to simultaneously determine 7 major constituents, including mulberroside A, chlorogenic acid, dihydromorin, oxyresveratrol, moracin O, kuwanon G, and kuwanon H, and the developed method showed good linearity(r>0.998 9) within the concentration range and the recoveries varied from 99.88% to 103.0%, and the RSD was 1.7%-2.9%. The HPLC results showed that the contents of the 7 compounds have great differences in 13 batches samples, compared with mulberry root bark, the contents of mulberroside A, chlorogenic acid, dihydromorin and moracin O of Mori Cortex were increased, while the contents of oxyresveratrol, kuwanon G and kuwanon H were decreased after peeling process. These results can provide a basis for the rationality and quality control of Mori Cortex required to remove the phellem layer.

Phytochemicals and antioxidant activity of alcoholic/hydroalcoholic extract of Trifolium pratense / 中草药·英文版

Objective: Trifolium pratense has many healing properties, including fewer complications of menopause, cancer cell suppression, reducing blood glucose and lipids, as well as cardiovascular beneficial effects. The purpose of this study was to identify the phytochemical and mineral composition of T. pratense. Methods: Plant aerial parts were harvested and dried, and then hydroalcoholic and alcoholic extracts were prepared. Gas chromatography–mass spectrometry (GC–MS) analytical method was used to identify volatile compounds then liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) was used to identify polyphenols and the mineral elements were identify by inductively coupled plasma atomic emission spectrometer/ICP-AES and scanning electron microscope-energy-dispersive X-ray spectroscopy (SEM-EDS) methods. Total phenolic content (TPC) was determined based on colorimetric method, and total flavonoid content (TFC) was established based on the folin-chiocalteau reagent. Furthermore, two assays (DPPH and FRAP) were used to measure the antioxidant capacity of T. pratense ethanolic extract. Results: A total of 37 polyphenols and 107 peaks were identified by LC-ESI-MS analysis, and the GC/MS method also detected 21 volatile compounds, the most important of which were methylcyclopentane, dimethylpentanal and hexadecanol. A total of 18 mineral elements, including K, Mg, Al, Si, Zn, Ni, Cu, Se, Co, Fe, Mn, and Ca in the plant, were identified ICP-AES and SEM-EDS analysis. Conclusion: T. pratense has many therapeutic compounds such as polyphenol (isoflavone and flavonoids), volatile compounds, and essential mineral elements, which can be formulated purely and used in the pharmaceutical and traditional medicine industries.

Characterization of Anthocyanins in Sweet Potato Leaves Grown in Various Stages and Conditions

Phytochemical-enriched edible greens, sweet potato leaves (Ipomoea batatas L.), have become popular due to potential health benefits. However, the phytochemical contents in sweet potato leaves and their subsequent change over harvest stages and growth condition are mostly unknown. In this study, the anthocyanin profile and content in leaves of four sweet potato cultivars, i.e., white-skinned and white-fleshed Bonita, red-skinned and orange-fleshed Beauregard, red-skinned and white-fleshed Murasaki and purple-skinned and purple-fleshed P40, were evaluated. Fourteen anthocyanins were isolated and identified by HPLC-MSI/MS. The most abundant was cyanidin 3-caffeoyl-p-hydroxybenzoyl sophoroside-5-glucoside, which comprised up to 20% of the total anthocyanins. Of the young leaves (1st and 2nd slip cuttings), Bonita contained the highest anthocyanin content followed by P40. Of the mature leaves (vine stage), Beauregard had the greatest anthocyanin (592.5 ± 86.4 mg/kg DW) and total phenolic (52.2 ± 3 mg GAE/g DW). It should be noted that the lowest anthocyanin and total phenolic content of shoots were found in P40, while tubers of P40 contain the highest content of each. Furthermore, the increase in leaf anthocyanin content over the growth stages that was observed in three of the cultivars but not in P40. No significant difference of anthocyanin content was found in Beauregard leaves grown in the high tunnels when compared with that in the open field. This study demonstrated for the first time that anthocyanin levels were significantly changed in response to various growth stages but not high tunnel condition, indicating that the effect of anthocyanin biosynthesis in sweet potato leaves is highly variable and genotype specific.

Development And Validation Of Sensitive Lc-Esi-Ms/Ms Method For The Simultaneous Estimation Of Dapagliflozin And Saxagliptin In Human Plasma

Objective: To develop and validate a sensitive liquid chromatography-electrospray ionization-tandem mass spectrometric (LC-ESI-MS/MS) technique for the quantification of dapagliflozin and saxagliptin in plasma by linagliptin as internal standard. Methods: Chromatography was achieved on hypersil C18 (50 mmx4 mm) 5 µ analytical column with 0.1% formic acid and acetonitrile (25:75 V/V) as mobile phase at 0.7 ml/min flow rate. Dapagliflozin, saxagliptin, and linagliptin were detected at m/z 409.14/135.0, m/z 316.2/180.13 and m/z 472.54/456.21 respectively. Drugs and internal standard were extracted by LLE (liquid-liquid extraction). Results: Developed technique was validated over 0.5-1500.0 ng/ml linear concentration range for dapagliflozin and 2.00-2000.0 ng/ml for saxagliptin. This method established with intra-batch and inter-batch precision within 2.44-8.12% and 1.25-7.14 % for dapagliflozin and 1.84-7.5 % and 1.02–6.00 % for saxagliptin. This method established with intra-batch and inter-batch accuracy for dapagliflozin within 98.86-103% and 96.98-102 % respectively and for saxagliptin within 98.05-109.06 % and 97.00-104.00 % respectively. Conclusion: Both dapagliflozin and saxagliptin were stable during three freeze-thaw cycles, long term and bench-top stability studies. The developed method was useful for the routine analysis of dapagliflozin and saxagliptin simultaneously in plasma samples.

Characterization of hepatoprotective metabolites from Artemisia annua and Cleome droserifolia using HPLC/PDA/ESI/MS-MS

ABSTRACT The hepatoprotective activities of two traditionally used plants, Cleome droserifolia (Forssk.) Delile, Cleomaceae, and Artemisia annua L., Asteraceae, were recently reported. However, the biologically active metabolites responsible for this activity were not identified. The aqueous extract of C. droserifolia aerial parts, and the polar fraction of A. annua leaves were screened for their antioxidant activities using the 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) assay. The in vitro viability of HepG-2 cells treated with CCl4 and the extracts were assessed by MTT assay. The effects of the extracts on the liver enzymes and the total soluble protein in CCl4-intoxicated HepG-2 cells were investigated. An HPLC/PDA/ESI/MS-MS based analysis was carried out for extract of C. droserifolia and polar fraction of A. annua. Both exhibited pronounced free radical scavenging activities (86 and 83%, respectively). Both showed a significant increase in cell viability: 86.43% for the extract of C. droserifolia and 79.32% for polar fraction of A. annua. Only the extract of C. droserifolia (39.6 ± 5.41 and 20.4 ± 6.91 IU/dl, respectively) and polar fraction of A. annua (40.8 ± 2.14 and 24.5 ± 3.11 IU/dl, respectively) restored the levels of liver enzymes (aspartate transaminase and alanine transaminase, respectively) compared to the CCl4 intoxicated group (87.5 ± 4.34 and 34.1 ± 8.12 IU/dl, respectively) and other herbal extracts. More than fifty phenolic secondary metabolites were identified in the extracts under investigation. The significant hepatoprotective activities of both extracts seemed to be strongly connected to their content of hydroxycinnamoyl quinic acids and flavonoids.

Comparative analysis of aconitine alkaloids in aconite tube using direct ionization and wooden tip spray mass spectrometry / 药学学报

In this manuscript, a rapid and simple analysis method for aconitine alkaloids was established. The method was based on the use of direct ionization and wooden tip spray ionization technology to detect the aconite. The aconite tuber slices were wrapped with wet filter paper overnight, cut into triangles, and extracted with a few solvents for direct ionization and wooden tip spray mass spectrometry. The results showed that alkaloids in aconite tuber can be rapid detected by two mass spectrometric methods without tedious sample pretreatment. Both methods are superior to that of traditional capillary electrospray ionization mass spectrometry (ESI-MS). The direct ionization MS is better than the wooden tip spray MS for analysis of aconite except under the condition of methylene chloride as extract or spray solvent. Different types of alkaloids in aconite tuber can be selectively detected when different solvents are used. The experiments provide a rapid and no pretreatment MS spectrometric method for analysis of alkaloids in aconite. These sample methods are important for research on aspects of plant varieties, storage, prescription compatibility, and quality control of aconite.

Simultaneous determination of nine bioactive components in Bazhen Yimu Pills by HPLC-ESI-MS/MS / 中草药

Objective To develop and validate a high performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC-ESI-MS/MS) method for simultaneously qualitative and quantitative determination of nine major bioactive components (stachydrine hydrochloride, leonurine hydrochloride, paeoniflorin, ferulic acid, liquiritin, lobetyolin, verbascoside, atracylenolide I, and polyporenic acid C) in Bazhen Yimu Pills (BYP). Methods The chromatographic separation was performed on an Waters Atlantis T3 column (150 mm × 4.6 mm, 3.0 μm) with a gradient elution of methanol and 0.1% formic acid in water at a flow rate of 0.5 mL/min, and the injection volume was 10 μL. The nine major bioactive components were detected using an electrospray ionization source in negative ionization mode (ESI-) and quantified by multiple reaction monitor (MRM) scanning at the same time. Results The linear ranges of stachydrine hydrochloride, leonurine hydrochloride, paeoniflorin, ferulic acid, liquiritin, lobetyolin, verbascoside, atracylenolide I, and polyporenic acid C were 0.04-40.00 μg/mL (r = 0.999 2), 0.04-40.00 μg/mL (r = 0.999 3), 1.0-100.0 μg/mL (r = 0.999 1), 0.2-20.0 μg/mL (r = 0.999 6), 0.2-20.0 μg/mL (r = 0.997 5), 0.05-5.00 μg/mL (r = 0.999 4), 0.1-10.0 μg/mL (r = 0.999 4), 0.1-10.0 μg/mL (r = 0.999 2), 0.1-10.0 μg/mL (r = 0.999 2), and the average recoveries were 99.7% (RSD = 0.52%), 98.1% (RSD = 0.64%), 98.5% (RSD = 1.08%), 101.5% (RSD = 1.12%), 99.5% (RSD = 0.39%), 98.4% (RSD = 0.74%), 99.1% (RSD = 0.91%), 101.2% (RSD = 0.54%), and 100.1% (RSD = 0.47%), respectively. The content of nine batches of the nine major bioactive components were 0.423-0.752, 0.505-0.722, 0.613-1.300, 0.102-0.184, 0.195-0.255, 0.021-0.035, 0.034-0.072, 0.039-0.063, and 0.051-0.095 mg/g, respectively. Conclusion The developed method is simple, specific, and sensitive, and it can be applied for the determination of nine major bioactive components and the quality control of BYP collected from different production batches.

Antioxidant activities in vitro and components analysis of methanol extract of Actinidia arguta by HPLC-DAD-ESI-MS/MS / 中草药

Objective: To investigate and analyze antioxidant activities in vitro and the active ingredients of the methanol extract of Actinidia arguta. Methods: In this study, antioxidant activities of extract of A. arguta were carried out by using DPPH and ABTS radical scavenging assays in vitro. Qualitative analysis of major active components was performed by HPLC-DAD-ESI-MS/MS. Results: Extract of A. arguta had good scavenging effect on DPPH and ABTS free radical in vitro, and the EC50 values of scavenging effect of extract of A. arguta on DPPH and ABTS free radical were (26.275 ± 1.464) and (29.826 ± 1.309) mg/mL, respectively. On the basis of UV and mass spectral analysis, a total of 19 chemical compositions were preliminarily identified. Conclusion: extract of A. arguta has good antioxidant activity in vitro, and polyhydroxyl and unsaturated double bonds are the main active constituents.

Chemical constituents from Rhamnus heterophylla / 中草药

Objective: To investigate the chemical constituents from Rhamnus heterophylla. Methods: Multiple chromatographic separation techniques including silica gel, Sephadex LH-20 gel and MCI gel were employed to isolate and purify the compounds. Their structures were identified by means of the nuclear magnetic resonance (NMR) and physicochemical properties. The chemical compositions were quickly analyzed by HPLC-DAD-ESI-MS/MS and scanned in negative ions. Results: These compounds were isolated and determined as cis-aloeemodin bianthrone (1), trans-aloeemodin bianthrone (2), emodin-8-O-rhamnoside (3), citreorosein (4), emodin (5), chrysophanol (6), physcion (7), kaempferol (8), luteolin (9), and quercetin (10). In addition to compounds 5, 8, the other compounds are isolated from R. heterophylla for the first time, and the compound 3 is isolated from the genus Rhamnus for the first time. 28 Compounds were speculated via comparing MS data (detected mass and MS2 fragment ions) and coupled with related literature, including three hydroxybenzoic acids, eight anthraquinones and 17 flavonoids. Conclusion: The analysis of chemical constituents from R. heterophylla could provide the references for the study on pharmacodynamic material basis and reasonable application and development.

Composition analysis and antioxidant activities of the Rhus typhina L. stem / 药物分析学报

The present investigation reports the chemical composition of the Rhus typhina L. stem identified via mass spectrometry and NMR as gallic acid, 1-O-galloyl-β-D-glucose, tryptophan, scopolin, methyl gallate, fustin, quercetin, rutin, and 1,2,3,4,6-penta-O-galloyl-β-D-glucose. The antioxidant properties and the chemical composition contents of the R. typhina L. stem grown in different regions in China were de-termined. To determine the antioxidant activity, a total phenolic content analysis, 2, 2-diphenyl-1-pi-crylhydrazyl radical scavenging activity assay, ferric reducing antioxidant power assay, andβ-carotene linoleic acid model system were conducted. The results showed that the Rhus typhina L. stem possessed high antioxidant capacities due to its high phenolic content. The contents of the nine isolated compounds were determined by UPLC-ESI-MS/MS. The calibration curves of the nine isolated compounds were linear within the concentration range and the average recoveries were high. The result showed that 1-O-galloyl-β-D-glucose, gallic acid, methyl gallate, and 1,2,3,4,6-penta-O-galloyl-β-D-glucose could be the compounds mainly responsible for the antioxidant capacity of the R. typhina L. stem. This reveals that the R. typhina L. stem is a good source of antioxidants.

Fermentation optimization and enzyme characterization of a new ι-Carrageenase from Pseudoalteromonas carrageenovora ASY5

Background: A new ι-carrageenase-producing strain was screened from mangroves and authenticated as Pseudoalteromonas carrageenovora ASY5 in our laboratory. The potential application of this new strain was evaluated. Results: Medium compositions and culturing conditions in shaking flask fermentation were firstly optimized by single-factor experiment. ι-Carrageenase activity increased from 0.34 U/mL to 1.08 U/mL after test optimization. Optimal fermentation conditions were 20°C, pH 7.0, incubation time of 40 h, 15 g/L NaCl, 1.5% (w/v) yeast extract as nitrogen source, and 0.9% (w/v) ι-carrageenan as carbon source. Then, the crude ι-carrageenase was characterized. The optimum temperature and pH of the ι-carrageenase were 40°C and 8.0, respectively. The enzymatic activity at 35­40°C for 45 min retained more than 40% of the maximum activity. Meanwhile, The ι-carrageenase was inhibited by the addition of 1 mmol/L Cd2+ and Fe3+ but increased by the addition of 1 mmol/L Ag+, Ba2+, Ca2+, Co2+, Mn2+, Zn2+, Fe2+, and Al3+. The structure of oligosaccharides derived from ι-carrageenan was detected using electrospray ionization mass spectrometry (ESI-MS). The ι-carrageenase degraded ι-carrageenan, yielding disaccharides and tetrasaccharides as main products. Conclusions: The discovery and study of new ι-carrageenases are beneficial not only for the production of ι-carrageenan oligosaccharides but also for the further utilization in industrial production.

Mass spectrometry guided strategy based on feature fragment ions for guided-separation on quinoline alkaloids from root barks of Dictamnus dasycarpus / 中国中药杂志

The root bark of Dictamnus dasycarpus is one of common traditional Chinese medicines (TCMs). Quinoline alkaloids are one of the main active substances in this TCM and possess many biological activities including anti-titumor, anti-inflammation, anti-bacteria, anti-oxidation, and anti-platelet aggregation activities. In this study, eight quinoline alkaloids 1-8 were firstly separated from the root barks of D. dasycarpus. It was difficult to isolate more quinoline alkaloids from the remaining fraction 8 in D. dasycarpus by this conventional chemical separation, so the target analysis method combined LC-MS guided-separation of quinoline alkaloids from fraction 8 was established. MS/MS fragmentation patterns of eight quinoline alkaloids reference standard compounds 1-8 were studied by ultra-performance liquid chromatography-electrospary ionization-mass spectrometry (UPLC-ESI-MS/MS). Based on the feature fragment ion 200, the parent ion scan mode was established for the target analysis of quinoline alkaloids in fraction 8. Finally, 8-methoxyflindersine (9) and N-metilatanina (10) were discovered and isolated quickly from fraction 8 guided by LC-MS, and their structures were identified by NMR and MS. Among them, compound 10 was isolated from the genus Dictamnus for the first time. These results indicated that this method is not only quick and sensitive for analyzing the quinoline alkaloids, but also to effectively guided-separate this kind of alkaloids in the root barks of D. dasycarpus.

Simultaneous determination of nine bioactive components in Sishen Pills by HPLC-ESI-MS/MS / 中草药

Objective To develop and validate an high performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC-ESI-MS/MS) method for simultaneously qualitative and quantitative determination of nine major bioactive components (deoxyschizandrin, γ-schizandrin, schizandrin, schizandrol B, schisantherin A, psoralen, isopsoralen, evodiamine, and rutaecarpine) in Sishen Pills. Methods The chromatographic separation was performed on an Agilent Zorbax Eclipse Plus C18 column (100 mm × 4.6mm, 3.5 μm) with a gradient elution of methanol and 0.1% formic acid in water at a flow rate of 0.4 mL/min, and the injection volume was 20 μL. The nine major bioactive components were detected using an electrospray ionization source in positive ionization mode (ESI+) and quantified by multiple reaction monitor (MRM) scanning at the same time. Results The linear ranges of deoxyschizandrin, γ-schizandrin, schizandrin, schizandrol B, schisantherin A, psoralen, isopsoralen, evodiamine, and rutaecarpine were 8.50-850.00 ng/mL (r = 0.999 7), 1.32-132.00 ng/mL (r = 0.997 4), 9.60-960.00 ng/mL (r = 0.999 8), 12.00-1 200.00 ng/mL (r = 0.999 3), 11.50-1 150.00 ng/mL (r = 0.997 9), 21.70-2 170.00 ng/mL (r = 0.999 7), 23.80-2 380.00 ng/mL (r = 0.999 6), 10.70-1 070.00 ng/mL (r = 0.999 5), 8.54-854.00 ng/mL (r = 0.998 0), and the average recoveries were 98.3% (RSD = 2.21%), 100.3% (RSD = 1.78%), 99.2% (RSD = 2.19%), 100.4% (RSD = 2.23%), 99.1% (RSD = 2.18%), 97.7% (RSD = 3.03%), 99.0% (RSD = 2.51%), 98.9% (RSD = 2.72%), and 100.3% (RSD = 2.10%), respectively. The contents of eight batches of the nine major bioactive components were 67.6-425.6, 0-131.5, 2.1-258.0, 0-71.2, 23.2-678.8, 806.4-1310.8, 718.5-1293.7, 11.5-123.2, and 10.9-62.4 μg/g, respectively. Conclusion The developed method is simple, specific, and sensitive, and it can be applied for the determination of nine major bioactive components and the quality control of Sishen Pills collected from different production batches.