Analysis of terpenes fromGinkgo biloba L. extracts by reversed phase high-performance liquid chromatography

Summary The main terpenes ofGinkgo biloba L. extracts (bilobalide, ginkgolide A, ginkgolide B and ginkgolide C) have been separated by isocratic elution on a 3 μm C₁₈ Spherical column using 2-propanol:water (10∶90) as eluent.     

THE APPLICATION OF A NOVEL ADSORBENT IN RAPID SAMPLE CLEAN-UP OF GINKGOLIDES AND BILOBALIDE IN EXTRACTS OF GINKGO BILOBA LEAVES

1 INTRODUCTIONGinkgolides, known as specific antagonist of platelet factor (PAF), possess uniquepharmaceutical properties and are main active constituent of many phytopharrnaceuticpreparations based on ginkgo bilioba leaves extracts II]. Another active terpenoid, namelybilobalide, is reported recently to show potent therapeutic efficacy to the disease related to nervesystem 121. They are all terpene lactones, which are highly oxidized as illustrated in Figure 1.The first attempts to ana…     

Simultaneous determination of ginkgolides A,B, C and bilobalide by LC‐MS/MS and its application to a pharmacokinetic study in rats

The study of pharmacokinetics of Ginkgo biloba extracts in Traditional Chinese Medicine was relatively recent. In this study, a simple, quick and sensitive LC‐MS/MS analytical method was developed for the determination of ginkgolides A, B, C and bilobalide in rat plasma. The analytes were completely separated from the endogenous compounds on an Agilent Zorbax Eclipse plus C₁₈ column (50 mm × 3.0 mm, 1.8 µm) using an isocratic elution. The single‐run analysis time was as short as 5.0 min. Sample preparation for protein removal was accomplished used a simple methanol precipitation method, after SPE showing a simultaneous extraction and cleanup of extracts allowing for a direct analysis. Extraction recoveries in rat plasma for ginkgolides A, B, C and bilobalide ranged from 75.6% to 89.0%. The calibration curves were determined over the ranges 0.5–20,000 ng/mL for ginkgolides A, B, C and bilobalide respectively. The lower limits of quantification (LLOQ) of the analytes were 0.5 ng/mL. Inter‐day and intra‐day precision and accuracy were below 15% and between 85 and 115%, respectively. Finally, the developed method was successfully applied to a pharmacokinetic study following oral administration of the Ginkgo biloba extracts to the male ICR rats. Copyright © 2015 John Wiley & Sons, Ltd.     

LC-ESI-MS Determination of Bilobalide and Ginkgolides in Canine Plasma

A sensitive and selective method using liquid chromatography with electrospray ionization mass spectrometric detection was developed for the quantification of bilobalide and ginkgolides in canine plasma. The analytes were extracted with diethyl ether-dichloromethane-isopropanol (6:3:1, v/v) after spiking the samples with daidzein (internal standard). The lower limit of quantification (LLOQ) of the method was 2.5 μg L⁻¹ for ginkgolide B and 10.0 μg L⁻¹ for bilabolide, ginkgolide A and ginkgolide C. The accuracy of the method was within 15% of the actual values over a wide range of plasma concentrations. The intra-day and inter-day precision was better than 15% (R.S.D.). Finally, the LC-ESI-MS method was successfully applied to study the pharmacokinetics of ginkgolides and bilabolide after administration of Ginkgo biloba extracts to dogs.     

THE APPLICATION OF A NOVEL ADSORBENT IN RAPID SAMPLE CLEAN—UP OF GINKGOLIDES AND BILOBALIDE IN EXTRACTS OF GINKGO BILOBA LEAVES

A rapid method has been developed for rapid sample clean-up in the determination of the pharmacologically active terpenoid including ginkgolide A,B,C and bilobalide in ginkgo biloba leaves extracts (GBE).The extracts are dissolved in 7% of ethanol aqueous solution and then purified by a highly selective polyeric adsorbent solid-phase chromatographic column.After being concentrated,the separated terpenoids with no phenolic distrubance are determined by highperformance liquid chromatorgraphy on a Nova-Pak C18 column with methanol-water(30:70)as effluent and refractive index detection.The recovery of the method is about 95% and the new method saves more time than the conventional two-column purification method.     

A new sesquiterpene trilactone from the roots of Ginkgo biloba

A new sesquiterpene trilactone, named bilobanol (1), along with four known terpene trilactones (ginkgolide A, B, C and bilobalide) were isolated from the roots of Ginkgo biloba collected in Jiangsu Province, China. The structure elucidation was accomplished by 1D and 2D NMR methods, HR-ESI-MS, and CD spectrum.     

Development and validation of a gas chromatographic-mass spectrometric method for simultaneous identification and quantification of marker compounds including bilobalide,ginkgolides and flavonoids in Ginkgo biloba L. extract and pharmaceutical preparations

A gas chromatography-mass spectrometry (GC-MS) method was developed and validated for the simultaneous determination of seven major chemical markers (bilobalide, ginkgolides A, B, C, kaempferol, quercetin and isorhamnetin) in phytopharmaceuticals of Ginkgo biloba L. The intra-day relative standard deviations (RSD) and inter-day RSD's were based on the analysis of the standardized Ginkgo biloba L. extract on the same day and on the following 3 consecutive days. The intra-day RSD's ranged from 1.21% (bilobalide) to 6.20% (kaempferol). The inter-day RSD's ranged from 2.10% (bilobalide) to 10.42% (isorhamnetin). Mean recoveries ranged from a low of 63.0 +/- 5.3% (isorhamnetin) to a maximum of 103.5 +/- 6.0% (ginkgolide A). Calibration curves were linear in ranges between 2.73 and 36.36 microg/ml for the markers. Limits of detection ranged from a low of 0.5 microg/ml (bilobalide) to a high of 2.5 microg/ml (quercetin). The limits of quantitation were a low of 1.1 microg/ml (gingkolides A, B, C) to a high of 7.5 microg/ml (kaempferol). The method was applied to a standard extract (>6% total terpenoids and >24% total flavonoids) and six ginkgo capsule phytopharmaceuticals.     

Complete 1H NMR spectral analysis of ten chemical markers of Ginkgo biloba

The complete and unambiguous ¹H NMR assignments of ten marker constituents of Ginkgo biloba are described. The comprehensive ¹H NMR profiles (fingerprints) of ginkgolide A, ginkgolide B, ginkgolide C, ginkgolide J, bilobalide, quercetin, kaempferol, isorhamnetin, isoquercetin, and rutin in DMSO‐d₆ were obtained through the examination of 1D ¹H NMR and 2D ¹H,¹H‐COSY data, in combination with ¹H iterative full spin analysis (HiFSA). The computational analysis of discrete spin systems allowed a detailed characterization of all the ¹H NMR signals in terms of chemical shifts (δ_H) and spin‐spin coupling constants (J_HH), regardless of signal overlap and higher order coupling effects. The capability of the HiFSA‐generated ¹H fingerprints to reproduce experimental ¹H NMR spectra at different field strengths was also evaluated. As a result of this analysis, a revised set of ¹H NMR parameters for all ten phytoconstituents was assembled. Furthermore, precise ¹H NMR assignments of the sugar moieties of isoquercetin and rutin are reported for the first time. Copyright © 2012 John Wiley & Sons, Ltd.     

Identification and the retention index of bilobalide and ginkgolides using capillary gas chromatography

Ginkgo biloba L. is known to contain the unique terpene trilactone compounds bilobalide and ginkgolides. Capillary gas chromatography is used for the quantitative identification of bilobalide and the main ginkgolides (ginkgolide A, B, and C). The retention indices of these compounds are also determined. Retention indices of bilobalide and ginkgolide A, B, and C substitute the use of their standards at their routine identification. Our procedure does not require temperature-programmed operation.     

Liquid chromatography/electrospray tandem mass spectrometry of terpenoid lactones in Ginkgo biloba

Ginkgo biloba (ginkgo) is one of most frequently used botanical dietary supplements. The bioactive constituents include the terpenoid lactones consisting of bilobalide and the ginkgolides A, B, C and J. A new assay based on high-performance liquid chromatography/electrospray tandem mass spectrometry (LC/MS/MS) was developed for the measurement of the terpenoid lactones in ginkgo products such as leaf powder and extracts. Initially, the MS/MS fragmentation pathways of ginkgolides were investigated to identify abundant fragment ions that might be useful for the sensitive and selective detection of ginkgolides and bilobalide during LC/MS/MS. Then, sample preparation and clean-up procedures were streamlined to maximize throughput by taking advantage of the selectivity of LC/MS/MS detection. Analyte recoveries exceeded 90%, the intra-assay and inter-assay relative standard deviations were <5%, the relative error was <8% and the limits of detection and quantification were 3.6-120 and 11-350 fmol, depending on the analyte that was injected on to the LC column. Therefore, this LC/MS/MS assay facilitated the rapid quantitative analysis of ginkgolides A, B, C and J and bilobalide in ginkgo dietary supplements with excellent recovery, reproducibity, accuracy and sensitivity.     

Liquid chromatography-electrospray mass spectrometric studies of ginkgolides and bilobalide using simultaneous monitoring of proton,ammonium and sodium adducts

A liquid chromatographic-electrospray mass spectrometric method was developed for the determination of ginkgolides and bilobalide and was applied to the analysis of commercial products of Ginkgo biloba leaf extracts. Adducts of these compounds with ammonium, proton and sodium were detected and their relative abundance depended on the electrospray fragmentor voltage. The relative standard deviation (RSD) was improved from > 17% to < 6%, when three adduct ions of (M + H)+, (M + NH4)+ and (M + Na)+ were used for quantification compared with single ion monitoring. The characteristic mass spectra of bilobalide were different from those of ginkgolides; in particular, dimers of (2M + Na)+ were observed for bilobalide only. Analysis of 26 commercial ginkgo products revealed large variations in the composition and concentrations of ginkgolides and bilobalide in herbal products.     

Assay of 44 compounds in the cortex and xylem from roots and branches of Ginkgo biloba L. by ultra high performance liquid chromatography coupled with tandem mass spectrometry and chemometric analysis

The leaves of Ginkgo biloba L. have received much attention, whereas there has been little systematic analysis of the cortex and xylem from roots and branches. A comprehensive evaluation of the 44 compounds in the cortex and xylem would thus be of value to fully understand the potential medicinal properties of roots and branches. An assay of amino acids, terpene lactones, flavones, and phenolic acids was accomplished using ultra high performance liquid chromatography with tandem mass spectrometry. All of the calibration curves showed good linear regression (R² > 0.9902) within the tested ranges. The intra‐ and interday precision was less than 4.9% and the accuracy was within ±6.8%. The amount of terpene lactones in the cortex was 1.75–2.07‐fold higher than that in the leaves. The amount of glutamine (360 μg/g) in the taproot xylem was 2.64‐fold higher than that in the leaves (136 μg/g). Principal component analysis decreased in the order leaves > taproot cortex > rootlet > laterals cortex > branch cortex > stem cortex > taproot xylem > branch xylem > laterals xylem > stem xylem. The taproot of G. biloba might provide a supplementary source of terpene lactones, especially ginkgolide A and C, and of glutamine.     

Determination of carbendazim,thiabendazole, and<Emphasis Type="Italic"> o-</Emphasis>phenylphenol residues in lemons by HPLC following sample clean-up by ion-pairing

An efficient analytical method is presented involving effective sample clean-up with solid-phase extraction and HPLC-UV analysis for the simultaneous determination of carbendazim, thiabendazole, and o-phenylphenol residues in lemons. Sample preparation involves extraction with acetonitrile acidified with trifluoroacetic acid and an ethyl acetate/petroleum ether mixture. Purification of the crude extract was carried out with liquid–liquid partitioning after addition of an aqueous ammonia solution. Final clean-up was performed on polymeric reversed-phase cartridges pretreated with sodium dodecyl sulfate. Chromatographic analysis was performed on a reversed-phase HPLC column isocratically eluted with an acetonitrile/water/ammonia mixture and UV detection at 254 nm. The chromatographic method is repeatable, reproducible, and sensitive. Fungicide recoveries from lemon samples fortified at levels of 5 and 1 mg kg^–1 were 81–85% for carbendazim, 96–98% for thiabendazole, and 81–106% for o-phenylphenol with coefficients of variation of 2.5–7.4%. Detection limits for carbendazim, thiabendazole, and o-phenylphenol in lemons were 0.21, 0.27, and 0.51 mg kg^–1, respectively.     

HPLC-ELSD法测定银杏酮酯软胶囊中萜类内酯的含量

建立银杏酮酯软胶囊中萜类内酯含量的测定方法。采用HPLC—ELSD法,色谱条件为Diamonsil C18柱（250mm×4．6mm,5μm）,正丙醇-四氢呋喃-水（体积比l：25：74）为流动相,ELSD漂移管温度107％2,空气流速3．0L／rain。银杏内酯A、银杏内酯B、银杏内酯C、白果内酯分别在4．0088-20．0440,3．9284-19．6420,3．8348～19．1740,3．7888～30.3104gg范围内呈良好的线性关系,线性相关系数分别为0．9992,0．9999,0．9995,0．9998,回收率分别为98．48％,97．62％,97．54％,97．24％,测定结果的相对标准偏差分别为0．99％,1．13％,1．17％,0．83％（n=5）。该方法准确可靠,可用于银杏酮酯软胶囊的质量控制。     

Investigation of 2-[(<Emphasis Type="Italic">E</Emphasis>)-2-(4-diethylaminophenyl)-1-ethenyl]-1,3,3-trimethyl-3H-indolium as a new highly sensitive reagent for the spectrophotometric determination of nitrophenols

A new, simple, rapid, and sensitive spectrophotometric method has been developed for the determination of nitrophenols [picric acid (PA); dinitrophenols (DNP)] in wastewater samples. The method is based on the reaction of nitrophenols with 2-[(E)-2-(4-diethylaminophenyl)-1-ethenyl]-1,3,3-trimethyl-3 H-indolium chloride reagent to form the colored ion associates, which are extracted by organic solvents. The molar absorptivity of the ion associates of PA with the investigated reagent ranges from 8.3×10⁴ to 11.3×10⁴ L mol^–1 cm^–1, depending on the extractant. Because only PA is extracted in an acidic medium with the investigated reagent, but both PA and DNP are extracted in an alkaline medium, it is possible to determine both substances in a mixture. Appropriate reaction conditions have been established. The absorbance of the colored extracts obeys Beers law in the range of 0.04–4.58 mg L^–1 PA, 1.0–18.4 mg L^–1 2,4-DNP and 1.2–14.7 mg L^–1 2,6-DNP, respectively. The limit of detections, calculated from a blank test (n=10; P=0.95), are 0.05 mg L^–1 PA, 0.9 mg L^–1 (2,4-DNP), and 1.1 mg L^–1 (2,6-DNP), respectively.     

Bromokomplexe von Co(II) und Ni(II) in Acetonitril,Propandiol-1,2-carbonat und Trimethylphosphat

Zusammenfassung Die Bildung von Bromokomplexen von Co(II) und Ni(II) wird in Acetonitril (AN), Propandiol-1,2-carbonat (PDC) und Trimethylphosphat (TMP) auf spektrophotometrischem, potentiometrischem und konduktometrischem Wege untersucht. Folgende Koordinationsformen dürften vorliegen: [CoBr]⁺ (oktaedrisch inAN undTMP), CoBr₂ (tetraedrisch inAN, oktaedrisch inTMP), [CoBr₃]^– (tetraedrisch inAN, oktaedrisch inTMP), [CoBr₄]^2– (tetraedrisch inAN undPDC, oktaedrisch inTMP), [NiBr]⁺ (oktaedrisch inAN), NiBr₂ (tetraedrisch inAN, oktaedrisch inTMP), [NiBr₃]^– (tetraedrisch inAN), [NiBr₄]^2– (tetraedrisch inAN undPDC, oktaedrisch inTMP).

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The formation of bromo complexes of Co(II) and Ni(II) is investigated in acetonitrile (AN), propanediol-1,2-carbonate (PDC) and trimethylphosphate (TMP) by spectrophotometric, potentiometric and conductometric methods. The following coordination forms are reported: [CoBr]⁺ (octahedral inAN andTMP), CoBr₂ (tetrahedral inAN, octahedral inTMP), [CoBr₃]^– tetrahedral inAN, octahedral inTMP), [CoBr₄]^2– (tetrahedral inAN andPDC, octahedral inTMP), [NiBr]⁺ (octahedral inAN), NiBr₂ (tetrahedral inAN, octahedral inTMP), [NiBr₃]^– (tetrahedral inAN), [NiBr₄]^2– (tetrahedral inAN andPDC, octahedral inTMP).

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Mit 8 Abbildungen     

Bromokomplexe von Mn(II) und V(III) in Acetonitril,Propandiol-1,2-carbonat und Trimethylphosphat

Zusammenfassung Die Bromokomplexe von Mn(II) und V(III) inAN, PDC undTMP wurden auf spektrophotometrischem, potentiometrischem und konduktometrischem Wege untersucht. Folgende Koordinationsformen dürften vorliegen: [MnBr]⁺ (inAN), MnBr₂ (tetraedrisch inAN), [MnBr₃]^– (tetraedrisch inAN), [MnBr₄]^2– (tetraedrisch inAN undPDC); [VBr]²⁺ (oktaedrisch inTMP), [VBr₂]⁺ (oktaedrisch inAN), VBr₃ (oktaedrisch inAN), [VBr₄]^– (oktaedrisch inAN undPDC).

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Bromocomplexes of Mn(II) and V(III) were investigated inAN, PDC andTMP by spectrophotometric, potentiometric and conductometric methods. The presence of the following species is indicated: [MnBr]⁺ (inAN), MnBr₂ (tetrahedral inAN), MnBr₃]^– (tetrahedral inAN), [MnBr₄]^2– (tetrahedral inAN) andPDC); [VBr]²⁺ (octahedral inTMP), [VBr₂]⁺ (octahedral inAN), VBr₃ (octahedral inAN), [VBr₄]^– (octahedral inAN andPDC).

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Mit 5 Abbildungen     

HPLC Separation of O-Acylated Flavonoids and Biflavones from Some Species of Gymnospermae

Flavonoids present in the extracts from leaves of Pseudotsuga menziesii (Pinaceae), Ginkgo biloba (Ginkgoaceae) and Podocarpus dacrydioides (Podocarpaceae) were separated by use of the reversed phase HPLC method. The analysed compounds belong to different groups of flavonoids – biflavones (amentoflavone, bilobetin, 5–methoxybilobetin, podocarpusflavone A, sequoiaflavone, podocarpusflavone B, ginkgetin, isoginkgetin, sciadopitysin, kayaflavone, hinokiflavone, 2,3–dihydrosciadopitysin, 2,3–dihydroisoginkgetin), O–acylated flavonol glycosides (daglesiosides I, II, III, IV, trans–tiliroside, trans–ditiliroside), flavonol O–glycosides (astragalin, isoquercetin) and flavonol aglycones (kaempferol, quercetin, isorhamnetin). The conditions for flavonoid separation were optimized using various RP–18 columns. The chromatographic resolution was performed with isocratic or gradient elution – optimized by Drylab program or by traditional trial-and-error method, depending on the composition of flavonoid complex.     

Vibronic interactions in {6} and {18}hetero(A,B)annulenes

The vibronic (vibrational–electronic) interactions and the Jahn–Teller effects in the monoanions and trianions of {6}hetero(B,N), (C,N), and (B,O)annulenes and {18}hetero(B,N), (C,N), and (B,O)annulenes are discussed. All the heteroannulenes have threefold axis of symmetry and the twofold degenerate lowest unoccupied molecular orbital (LUMOs), and the E^′ or E vibrational modes can cause Jahn–Teller distortions in their monoanions and trianions. State vibronic coupling constants of the monoanions and trianions and orbital vibronic coupling constants concerning the LUMOs are calculated for each Jahn–Teller active vibrational mode at the B3LYP/6-31G* level of theory. Vibrational modes near 1500 cm⁻¹ of the {6}hetero(A,B)annulenes and low-frequency modes (<500 cm⁻¹) of the {18}hetero(A,B)annulenes give large coupling constants, and therefore, these modes are essential in the Jahn–Teller distortions and the vibronic interactions. The coupling constants are qualitatively analyzed by looking at the nuclear motions of the Jahn–Teller active modes and the shapes of the LUMOs on the basis of one-electron approximation.