Fast separation of flavonoids by supercritical fluid chromatography using a column packed with a sub‐2 μm particle stationary phase

The purpose of this study was to compare the effects of different chromatographic columns for the separation of seven flavonoids. Four different stationary phases are available, including bridged ethyl hybrid, BEH and the same hybrid phase modified with 2‐ethylpyridine, CSH fluorophenyl, and HSS C₁₈ SB. The analytes included calycosin, genistein, medicarpin, calycosin‐7‐O‐β‐d ‐glucoside, formononetin, formononetin‐7‐O‐β‐d ‐glucoside, and liquiritigenin. The CSH fluorophenyl column was determined to be the most suitable and provided the fastest separation within 17 min using gradient elution with carbon dioxide as the mobile phase and methanol as the co‐solvent. Good peak shapes were obtained, and the values of the peak asymmetry were close to 1.0 for all of the flavonoids. The resolution was more than 1.41 for all of the separated peaks. Baseline separation on the optimal columns was achieved by changing the co‐solvent type and adjusting the temperature and pressure. Quantitative performance was evaluated under optimized conditions, and method validation was accomplished. The validation parameters, such as linearity, sensitivity, precision, and accuracy, were satisfactory. Good repeatability of both peak area (relative standard deviation <1.02%) and retention time (relative standard deviation <0.88%) was observed. The optimized chromatographic methods were successfully used for the determination of seven flavonoids in Radix astragali . The sensitivity was sufficient for the analysis of real samples.     

Analysis and retention behavior of isoflavone glycosides and aglycones in Radix Astragali by HPLC with hydroxypropyl‐β‐cyclodextrin as a mobile phase additive

In order to determine isoflavone glycosides (calycosin‐7‐O‐β‐d ‐glucoside and formononetin‐7‐O‐β‐d ‐glucoside) and aglycones (calycosin and formononetin), a simple HPLC method with isocratic elution employing hydroxypropyl‐β‐cyclodextrin (HP‐β‐CD) as a mobile phase additive was developed. Various factors affecting the retention of isoflavone glycosides and aglycones in the C₁₈ reversed‐phase column, such as the nature of cyclodextrins, HP‐β‐CD concentration, and methanol concentration, were systematically studied. The results show that HP‐β‐CD, as a very effective mobile phase additive, can markedly reduce the retention of isoflavone glycosides and aglycones, and the decrease magnitudes of isoflavone aglycones are more than those of their glycosides. The role of HP‐β‐CD in the developed HPLC method is attributed to the formation of the inclusion complexes between isoflavone glycosides (or aglycones) and HP‐β‐CD. So, the apparent formation constants of the isoflavone glycosides (or aglycones)/HP‐β‐CD inclusion complexes also were investigated. Isoflavone glycosides (and aglycones) form the 1:1 inclusion complexes with HP‐β‐CD, and the isoflavone aglycones/HP‐β‐CD complexes are more stable than the isoflavone glycosides/HP‐β‐CD complexes. Finally, the optimized method was successfully applied for the determination of isoflavone glycosides and aglycones in Radix Astragali samples.     

Simultaneous determination of calycosin‐7‐O‐β‐d‐glucoside,calycosin, formononetin,astragaloside IV and schisandrin in rat plasma by LC‐MS/MS: application to a pharmacokinetic study after oral administration of Shenqi Wuwei chewable tablets

A rapid, sensitive and reliable high‐performance liquid chromatography–mass spectrometry (LC‐MS/MS) method was developed and validated for simultaneous quantification of the five main bioactive components, calycosin, calycosin‐7‐O‐β‐d ‐glucoside, formononetin, astragaloside IV and schisandrin in rat plasma after oral administration of Shenqi Wuwei chewable tablets. Plasma samples were extracted using solid‐phase extraction separated on a CEC₁₈ column and detected by MS with an electrospray ionization interface in multiple‐reaction monitoring mode. Calibration curves offered linear ranges of two orders of magnitude with r > 0.995. The method had a lower limit of quantitation of 0.1, 0.02, 0.1, 1 and 0.1 ng/mL for calycosin, calycosin‐7‐O‐β‐d ‐glucoside, formononetin, astragaloside IV and schisandrin, respectively. Intra‐ and inter‐day precisions (relative standard deviation) for all analytes ranged from 0.97 to 7.63% and from 3.45 to 10.89%, respectively. This method was successfully applied to the pharmacokinetic study of the five compounds in rats after oral administration of Shenqi Wuwei chewable tablets. Copyright © 2014 John Wiley & Sons, Ltd.     

Rapid characterization and determination of multiple components in Bu‐Shen‐Yi‐Qi‐Fang by high‐performance liquid chromatography coupled to electrospray ionization and quadrupole time‐of‐flight mass spectrometry

In this study, a qualitative and quantitative analysis using high‐performance liquid chromatography coupled to electrospray ionization and quadrupole time‐of‐flight mass spectrometry was performed for the quality control of Bu‐Shen‐Yi‐Qi‐Fang, a traditional Chinese formula used for asthma. Thirty‐four compounds, including flavonoids, isoflavonoids, triterpenoid saponins, and iridoid glycosides were identified or tentatively characterized by comparing their retention times and mass spectra with those of authentic standards or literature data. Sixteen components were considered as the main bioactive constituents of Bu‐Shen‐Yi‐Qi‐Fang and they were chosen as the chemical markers in quantitative analysis, including catalpol, leonuride, calycosin‐7‐O‐β‐d ‐glucoside, hyperoside, acteoside, formononetin‐7‐O‐β‐d ‐glucoside, epimedin A, calycosin, icariin, epimedin B, epimedin C, formononetin, astragaloside IV, astragaloside II, baohuoside‐I, and astragaloside I. The total run time was 20 min. It was found that the calibration curves for all analytes showed good linearity (R² > 0.99) within the test ranges. The relative standard deviations for intra‐ and inter‐day precisions were below 3.9 and 11.7%, respectively. The accuracy was evaluated by the recovery test within the range of 89.20–110.71% with the relative standard deviation < 4.8%. The sample was stable for at least 48 h at 4°C. The results showed that the new approach was effective for the quality control of Bu‐Shen‐Yi‐Qi‐Fang.     

Development of a sensitive ultra high performance liquid chromatography with tandem mass spectrometry method for the simultaneous quantification of nine active compounds in rat plasma and its application to a pharmacokinetic study after administration of Viscum coloratum extracts

A simple, specific, and sensitive ultra high performance liquid chromatography with tandem mass spectrometry method was developed and validated for the simultaneous quantification of nine compounds including a new compound, rhamnazin‐3‐Ο‐β‐d ‐(6″‐β‐hydroxy‐β‐methyglutaryl)‐β‐d ‐glucoside‐4′‐Ο‐β‐d ‐glucoside, in rat plasma using baicalin as an internal standard. The plasma samples were pretreated and extracted by protein precipitation with 0.2% formic acid in acetonitrile. The analytes were separated on a Thermo Syncronis C₁₈ column by gradient elution with a mobile phase consisting of acetonitrile and 0.1% aqueous formic acid at a flow rate of 0.25 mL/min. The detection of the analytes was performed on an electrospray ionization interface operating in positive‐ion and multiple reaction monitoring acquisition modes. The calibration curves of these analytes showed good linearity (r > 0.99) within the test ranges. The lower limit of quantification ranged from 0.4 to 20.1 ng/mL for the analytes. The intra‐ and interday precision and accuracy were all within ±15%, and the recoveries were higher than 80.0%. The validated method was successfully applied to a pharmacokinetic study of the nine flavonoids after administration of the Viscum coloratum extracts by intravenous injection.     

Photoirradiation surface molecularly imprinted polymers for the separation of 6‐O‐α‐d‐maltosyl‐β‐cyclodextrin

Photoirradiation surface molecularly imprinted polymers for the separation of 6‐O‐α‐d ‐maltosyl‐β‐cyclodextrin were synthesized using functionalized silica as a matrix, 4‐(phenyldiazenyl)phenol as a light‐sensitive monomer, and 6‐O‐α‐d ‐maltosyl‐β‐cyclodextrin as a template. Fourier transform infrared spectroscopy results indicated that 4‐(phenyldiazenyl)phenol was grafted onto the surface of functionalized silica. The obtained imprinted polymers exhibited specific recognition toward 6‐O‐α‐d ‐maltosyl‐β‐cyclodextrin. Equilibrium binding experiments showed that the photoirradiation surface molecularly imprinted polymers obtained the maximum adsorption amount of 6‐O‐α‐d ‐maltosyl‐β‐cyclodextrin at 20.5 mg/g. In binding kinetic experiments, the adsorption reached saturation within 2 h with binding capacity of 72.8%. The experimental results showed that the adsorption capacity and selectivity of imprinted polymers were effective for the separation of 6‐O‐α‐d ‐maltosyl‐β‐cyclodextrin, indicating that imprinted polymers could be used to isolate 6‐O‐α‐d ‐maltosyl‐β‐cyclodextrin from a conversion mixture containing β‐cyclodextrin and maltose. The results showed that the imprinted polymers prepared by this method were very promising for the selective separation of 6‐O‐α‐d ‐maltosyl‐β‐cyclodextrin.     

Simultaneous quantification of triterpenoid saponins in rat plasma by UHPLC–MS/MS and its application to a pharmacokinetic study after oral total saponin of Aralia elata leaves

A rapid, sensitive, and reliable analytical ultra performance liquid chromatography with tandem mass spectrometry method was developed for the simultaneous determination of Aralia‐saponin IV, 3‐O‐β‐d ‐glucopyranosyl‐(1→3)‐β‐d ‐glucopyranosyl‐(1→3)‐β‐d ‐glucopyranosyl oleanolic acid 28‐O‐β‐d ‐glucopyranoside, Aralia‐saponin A and Aralia‐saponin B after the oral administration of total saponin of Aralia elata leaves in rat plasma. Plasma samples were pretreated by protein precipitation with methanol. The analysis was performed on an ACQUITY UPLC HSS T3 column. The detection was performed on a triple quadrupole tandem mass spectrometer in multiple reaction monitoring mode using an electrospray ionization source with negative ionization mode. Under the experimental conditions, the calibration curves of four analytes had good linearity values (r > 0.991). The intra‐ and inter‐day precision values of the four analytes were ≤ 11.6%, and the accuracy was between –6.2 and 4.2%.The extraction recoveries of four triterpenoid saponins were in the range of 84.06–91.66% (RSD < 10.5%), and all values of the matrix effect were more than 90.30%. The developed analytical method was successfully applied to pharmacokinetic study on simultaneous determination of the four triterpenoid saponins in rat plasma after oral administration of total saponin of Aralia elata leaves, which helps guiding clinical usage of Aralia elata leaves.     

Conformational analysis of the disaccharide methyl α‐d‐mannopyranosyl‐(1→3)‐2‐O‐acetyl‐β‐d‐mannopyranoside monohydrate

The crystal structure of methyl α‐d ‐mannopyranosyl‐(1→3)‐2‐O‐acetyl‐β‐d ‐mannopyranoside monohydrate, C₁₅H₂₆O₁₂·H₂O, ( II ), has been determined and the structural parameters for its constituent α‐d ‐mannopyranosyl residue compared with those for methyl α‐d ‐mannopyranoside. Mono‐O‐acetylation appears to promote the crystallization of ( II ), inferred from the difficulty in crystallizing methyl α‐d ‐mannopyranosyl‐(1→3)‐β‐d ‐mannopyranoside despite repeated attempts. The conformational properties of the O‐acetyl side chain in ( II ) are similar to those observed in recent studies of peracetylated mannose‐containing oligosaccharides, having a preferred geometry in which the C2—H2 bond eclipses the C=O bond of the acetyl group. The C2—O2 bond in ( II ) elongates by ～0.02 Å upon O‐acetylation. The phi (?) and psi (ψ) torsion angles that dictate the conformation of the internal O‐glycosidic linkage in ( II ) are similar to those determined recently in aqueous solution by NMR spectroscopy for unacetylated ( II ) using the statistical program MA′AT, with a greater disparity found for ψ (Δ = ～16°) than for ? (Δ = ～6°).     

Simultaneous determination of major components of Huangqi–Honghua extract in rat plasma using LC–MS/MS and application to a pharmacokinetic study

A sensitive and reliable LC–MS/MS method was developed and validated for simultaneous quantification of the major components of Huangqi–Honghua extact in rat plasma, including hydroxysafflor yellow A (HSYA), astragaloside IV (ASIV), calycosin‐7‐O‐β‐d ‐glucoside (CAG), calycosin, calycosin‐3′‐O‐glucuronide (C‐3′‐G) and calycosin‐3′‐O‐sulfate (C‐3′‐S). After extraction by protein precipitation with acetonitrile and methanol from plasma, the analytes were separated on a Hypersil BDS C₁₈ column by gradient elution with acetonitrile and 5 mM ammonium acetate. The detection was carried out on a triple quadrupole tandem mass spectrometer equipped with electrospray ionization source switched between negative and positive modes. HSYA was monitored in negative ionization mode from 0 to 4.9 min, and ASIV, CAG, calycosin, C‐3′‐G and C‐3′‐S were determined in positive ionization mode from 4.9 to 10 min. The lower limits of quantification of the analytes were 6.25 ng/mL for HSYA, 0.781 ng/mL for CAG and 1.56 ng/mL for ASIV and calycosin. The intra‐ and inter‐assay precision (RSD) values were within 13.43%, and accuracy (RE) ranged from ?8.75 to 9.92%. The validated method was then applied to the pharmacokinetic study of HSYA, ASIV, CAG, calycosin, C‐3′‐G and C‐3′‐S in rat after an oral administration of Huangqi–Honghua extract.     

2,3,3′,6‐Tetra‐O‐acetyl‐4,1′,6′‐trichloro‐4,1′,4′,6′‐tetradeoxygalactosucrose

At 160 K, one of the Cl atoms in the furanoid moiety of 3‐O‐acetyl‐1,6‐di­chloro‐1,4,6‐tri­deoxy‐β‐d ‐fructo­furan­osyl 2,3,6‐tri‐O‐acetyl‐4‐chloro‐4‐deoxy‐α‐d ‐galacto­pyran­oside, C₂₀H₂₇­Cl₃O₁₁, is disordered over two orientations, which differ by a rotation of about 107° about the parent C—C bond. The conformation of the core of the mol­ecule is very similar to that of 3‐O‐acetyl‐1,4,6‐tri­chloro‐1,4,6‐tri­deoxy‐β‐d ‐tagato­furanos­yl 2,3,6‐tri‐O‐acetyl‐4‐chloro‐4‐deoxy‐α‐d ‐galacto­pyran­oside, particularly with regard to the conformation about the glycosidic linkage.     

Simultaneous quantification of multiple components in rat plasma by UPLC‐MS/MS and pharmacokinetic study after oral administration of Huangqi decoction

A rapid, sensitive and accurate UPLC‐MS/MS method was developed for the simultaneous quantification of components of Huangqi decoction (HQD), such as calycosin‐7‐O‐β‐d ‐glucoside, calycosin‐glucuronide, liquiritin, formononetin‐glucuronide, isoliquiritin, liquiritigenin, ononin, calycosin, isoliquiritigenin, formononetin, glycyrrhizic acid, astragaloside IV, cycloastragenol, and glycyrrhetinic acid, in rat plasma. After plasma samples were extracted by protein precipitation, chromatographic separation was performed with a C₁₈ column, using a gradient of methanol and 0.05% acetic acid containing 4mm ammonium acetate as the mobile phase. Multiple reaction monitoring scanning was performed to quantify the analytes, and the electrospray ion source polarity was switched between positive and negative modes in a single run of 10 min. Method validation showed that specificity, linearity, accuracy, precision, extraction recovery, matrix effect and stability for 14 components met the requirements for their quantitation in biological samples. The established method was successfully applied to the pharmacokinetic study of multiple components in rats after intragastric administration of HQD. The results clarified the pharmacokinetic characteristics of multiple components found in HQD. This research provides useful information for understanding the relation between the chemical components of HQD and their therapeutic effects.     

Determination and pharmacokinetic study of four lignans in rat plasma after oral administration of an extract of Valeriana amurensis by ultra‐high performance liquid chromatography with tandem mass spectrometry

A selective and sensitive ultra‐high performance liquid chromatography with tandem mass spectrometry method was developed and validated for the determination and pharmacokinetic study of (+)‐8‐hydroxypinoresinol‐4’‐O‐β ‐D‐glucopyranoside, prinsepiol‐4‐O‐β‐D‐glucopyranoside, (+)‐pinoresinol‐4,4’‐di‐O‐β‐D‐glucopyranoside, and (−)‐massoniresinol 3α‐O‐β‐D‐glucopyranoside in rat plasma after the oral administration of a Valeriana amurensis extract. The analytes and ethyl 4‐hydroxybenzoate (internal standard) were separated on a Waters ACQUITY UPLC HSS T3 chromatographic column. The detection was performed on a triple quadrupole tandem mass spectrometer in multiple reaction monitoring mode using an electrospray ionization source operating in negative ionization mode. The linear ranges (ng/mL) of the standard curves were 0.39–154.00, 0.62–244.70, 0.50–198.60, and 0.34–134.50 for (+)‐8‐hydroxypinoresinol‐4’‐O‐β‐D‐glucopyranoside, prinsepiol‐4‐O‐β‐D‐glucopyranoside, (+)‐pinoresinol‐4,4’‐di‐O‐β‐D‐glucopyranoside, and (−)‐massoniresinol 3α‐O‐β‐D‐glucopyranoside, respectively. The inter‐ and intra‐day precisions were less than 11.0%, the accuracies were between −5.9 and 7.7%, and the extraction recoveries of the four analytes were > 81.2% from rat plasma. The method was successfully applied to a pharmacokinetic study of the four analytes after oral administration of a Valeriana amurensis extract to rats. The developed method has the potential for pharmacokinetic analysis and to provide additional information in the clinical application of Valeriana amurensis.     

Quantitative determination of seven chemical constituents and chemo‐type differentiation of chamomiles using high‐performance thin‐layer chromatography

A simple and rapid high‐performance thin‐layer chromatographic method was developed for the separation and determination of six flavonoids (rutin, luteolin‐7‐O‐β‐glucoside, chamaemeloside, apigenin‐7‐O‐β‐glucoside, luteolin, apigenin) and one coumarin, umbelliferone from chamomile plant samples and dietary supplements. The separation was achieved on amino silica stationary phase using dichloromethane/acetonitrile/ethyl formate/glacial acetic acid/formic acid (11:2.5:3:1.25:1.25 v/v/v/v/v) as the mobile phase. The quantitation of each compound was carried out using densitometric reflection/absorption mode at their respective absorbance maxima after postchromatographic derivatization using natural products reagent (1% w/v methanolic solution of diphenylboric acid‐β‐ethylamino ester). The method was validated for specificity, limits of detection and quantification, precision (intra‐ and interday) and accuracy. The limits of detection and quantification were found to be in the range from 6–18 and 16–55 ng/band for six flavonoids and one coumarin, respectively. The intra‐ and interday precision was found to be <5% RSD and recovery of all the compounds was >90%. The data acquired from high‐performance thin‐layer chromatography was processed by principal component analysis using XLSTAT statistical software. Application of principal component analysis and agglomerative hierarchial clustering was successfully able to differentiate two chamomiles (German and Roman) and Chrysanthemum.     

Separation of three anthraquinone glycosides including two isomers by preparative high‐performance liquid chromatography and high‐speed countercurrent chromatography from Rheum tanguticum Maxim. ex Balf

Anthraquinone glycosides, such as chrysophanol 1‐O‐β‐d‐ glucoside, chrysophanol 8‐O‐β‐d‐ glucoside, and physion 8‐O‐β‐d‐ glucoside, are the accepted important active components of Rheum tanguticum Maxim. ex Balf. due to their pharmacological properties: antifungal, antimicrobial, cytotoxic, and antioxidant activities. However, an effective method for the separation of the above‐mentioned anthraquinone glycosides from this herb is not currently available. Especially, greater difficulty existed in the separation of the two isomers chrysophanol 1‐O‐β‐d‐ glucoside and chrysophanol 8‐O‐β‐d‐ glucoside. This study demonstrated an efficient strategy based on preparative high‐performance liquid chromatography and high‐speed countercurrent chromatography for the separation of the above‐mentioned anthraquinone glycosides from Rheum tanguticum Maxim.ex Balf.     

Polar Constituents from Sageretia Thea Leaf Characterized by HPLC‐SPE‐NMR Assisted Approaches

Nine glycosides ( 1–9 ) were characterized from the n‐butanol‐soluble fraction of the ethanolic extract of the leaves of Sageretia thea by the general approach. Among these, Compounds 6 and 7 were identified as a mixture. Application of HPLC‐SPE‐NMR in two selected fractions led to the separation of this mixture and the characterization of three additional minors ( 10–12 ). Among these, 7‐O‐methylmyricetin 3‐O‐α‐l ‐arabinofuranoside ( 8 ) is a new natural product and eight compounds, i.e. glucofragulin A ( 1 ), quercetin‐3‐O‐α‐l ‐arabinopyranoside ( 5 ), 3‐O‐β‐d ‐galactopyranoside ( 6 ), 3‐O‐β‐d ‐glucopyranoside ( 7 ), and 3‐O‐α‐l ‐arabinofuranoside ( 11 ), myricetin‐3‐O‐α‐l ‐arabinofuranoside ( 9 ) and 3‐O‐β‐d‐glucopyranoside ( 10 ), and quercetrin ( 12 ), are found for the first time from the title plant.     

Synthesis of 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(4′′‐hydroxyphenyl)‐thiazoles and their O‐glucosides

In continuation of our work, we synthesized 2‐(sulfamoylphenyl)‐4′‐amino‐4‐(4″‐hydroxyphenyl)‐thiazole ( 3a ), which were reacted with various (aryl/hetroaryl) aldehyde to form 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(4″‐hydroxyphenyl)‐thiazoles ( 4a , 4b , 4c , 4d , 4e , 4f ). Glucosylation of compounds ( 4a , 4b , 4c , 4d , 4e , 4f ) have been done by using acetobromoglucose as a glucosyl donor to afford 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(2,3,4,6‐tetra‐O‐acetyl‐4″‐O‐β‐D ‐glucosidoxyphenyl)‐thiazoles ( 5a , 5b , 5c , 5d , 5e , 5f ), further on deacetylation to produce 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(4″‐O‐β‐D ‐glucosidoxyphenyl)‐thiazoles ( 6a , 6b , 6c , 6d , 6e , 6f ). The compounds are confirmed by FTIR, ¹H‐NMR, ¹³C‐NMR, and ES‐Mass spectral analysis. J. Heterocyclic Chem., (2011).     

Development of a high‐performance liquid chromatography procedure to identify known and detect novel C‐13 oligosaccharide moieties in diterpene glycosides from Stevia rebaudiana (Bertoni) Bertoni (Asteraceae): Structure elucidation of rebaudiosides V and W

As an aid for structure elucidation of new steviol glycosides, reversed‐phase C18 high‐performance liquid chromatography method was developed with several previously characterized diterpene glycosides, to identify known and detect novel aglycone‐C13 oligosaccharide moieties and indirectly identify C‐19 interlinkages. Elution order of several diterpene glycosides and their aglycone‐C13 oligosaccharide substituted with different sugar arrangements were also summarized. Comparison of the retention time of a product obtained after alkaline hydrolysis with the aglycone‐C‐13 portions of known compounds reported herein allowed us to deduce the exact positions of the sugars in the C‐13 oligosaccharide portion. The elution position of several steviol glycosides with an ent‐kaurene skeleton was helpful to describe an identification key. Two previously uncharacterized diterpene glycosides together with two known compounds were isolated from a commercial Stevia rebaudiana leaf extract. One was found to be 13‐[(2‐O‐β‐d ‐xylopyranosyl‐ 3‐O‐β‐d ‐glucopyranosyl‐β‐d ‐glucopyranosyl)oxy]ent‐kaur‐16‐en‐19‐oic acid‐(2‐O‐β‐d ‐glucopyranosyl‐β‐d ‐glucopyranosyl) ester (rebaudioside V), whereas the other was determined to be 13‐[(2‐O‐β‐d ‐xylopyranosyl‐ 3‐O‐β‐d ‐glucopyranosyl‐β‐d‐ glucopyranosyl)oxy]ent‐kaur‐16‐en‐19‐oic acid‐(2‐O‐α‐l ‐rhamnopyranosyl‐3‐O‐β‐d ‐glucopyranosyl‐β‐d ‐glucopyranosyl) ester (rebaudioside W). Previously reported compounds were isolated in gram quantities and identified as rebaudioside J and rebaudioside H. In addition, a C‐19 sugar‐free derivative was also prepared from rebaudioside H to afford rebaudioside H₁. Chemical structures were partially determined by the high‐performance liquid chromatography method and unambiguously characterized by using one‐dimensional and two‐dimensional nuclear magnetic resonance experiments.     

O‐Ethyl and O‐methyl N‐(2,3,4,6‐tetra‐O‐acetyl‐β‐d‐glucopyranosyl)thiocarbamate

In both the title structures, O‐ethyl N‐(2,3,4,6‐tetra‐O‐acetyl‐β‐d ‐gluco­pyran­osyl)­thio­carbam­ate, C₁₇H₂₅NO₁₀S, and O‐methyl N‐(2,3,4,6‐tetra‐O‐acetyl‐β‐d ‐gluco­pyran­osyl)­thiocar­bam­ate, C₁₆H₂₃NO₁₀S, the hexo­pyran­osyl ring adopts the ⁴C₁ conformation. All the ring substituents are in equatorial positions. The acetoxy­methyl group is in a gauche–gauche conformation. The S atom is in a synperi­planar conformation, while the C—N—C—O linkage is antiperiplanar. N—H?O intermolecular hydrogen bonds link the mol­ecules into infinite chains and these are connected by C—H?O interactions.     

Determination of paeoniflorin,calycosin‐7‐O‐β‐d‐glucoside,ononin, calycosin and formononetin in rat plasma after oral administration of Buyang Huanwu decoction for their pharmacokinetic study by liquid chromatography–mass spectrometry

The purpose of this study was to simultaneously investigate the pharmacokinetics of five bioactive compounds in rat plasma after oral administration of Buyang Huanwu decoction (BYHWD) using high‐performance liquid chromatography coupled with mass spectrometry (HPLC‐MS). The separations were performed on a Thermo Hypersil Gold C₁₈ analytical column (50 × 2.1 mm, 3 µm) with the column temperature kept at 30°C. The quantitative analysis was performed using a quadrupole mass spectrometer detector operated under selected ion monitoring mode. A linear gradient elution of A (0.1% formic acid solution) and B (100% acetonitrile) was used at a flow rate of 0.2 mL/min. The method was validated within the concentration ranges 1.8–450, 6.0–1500, 2.0–500, 1.2–300 and 1.2–150 ng/mL for paeoniflorin, calycosin‐7‐O‐β‐d ‐glucoside, ononin, calycosin and formononetin, respectively. The calibration curves were linear with correlation coefficients > 0.99. The lower limits of quantitations were < 6.0 ng/mL. The method was further applied to assess the pharmacokinetics of the five bioactive constituents of BYHWD in rat plasma. Copyright © 2010 John Wiley & Sons, Ltd.     

7‐Iodo‐8‐aza‐7‐deaza‐2′‐deoxy­adenosine and 7‐bromo‐8‐aza‐7‐deaza‐2′‐deoxyadenosine

The isomorphous structures of the title molecules, 4‐amino‐1‐(2‐deoxy‐β‐d ‐erythro‐pento­furan­osyl)‐3‐iodo‐1H‐pyrazolo‐[3,4‐d]pyrimidine, (I), C₁₀H₁₂IN₅O₃, and 4‐amino‐3‐bromo‐1‐(2‐deoxy‐β‐d ‐erythro‐pento­furan­osyl)‐1H‐pyrazolo[3,4‐d]­pyrimidine, (II), C₁₀H₁₂BrN₅O₃, have been determined. The sugar puckering of both compounds is C1′‐endo (^1′E). The N‐­glycosidic bond torsion angle χ¹ is in the high‐anti range [?73.2 (4)° for (I) and ?74.1 (4)° for (II)] and the crystal structure is stabilized by hydrogen bonds.