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1.
The detailed metabolite profiling of Laguncularia racemosa was accomplished by high-performance countercurrent chromatography (HPCCC) using the three-phase system n-hexane–tert-butyl methyl ether–acetonitrile–water 2:3:3:2 (v/v/v/v) in step-gradient elution mode. The gradient elution was adjusted to the chemical complexity of the L. racemosa ethyl acetate partition and strongly improved the polarity range of chromatography. The three-phase solvent system was chosen for the gradient to avoid equilibrium problems when changing mobile phase compositions encountered between the gradient steps. The tentative recognition of metabolites including the identification of novel ones was possible due to the off-line injection of fractions to electrospray ionization mass spectrometry (ESI-MS/MS) in the sequence of recovery. The off-line hyphenation profiling experiment of HPCCC and ESI-MS projected the preparative elution by selected single ion traces in the negative ionization mode. Co-elution effects were monitored and MS/MS fragmentation data of more than 100 substances were used for structural characterization and identification. The metabolite profile in the L. racemosa extract comprised flavonoids, hydrolysable tannins, condensed tannins and low molecular weight polyphenols.  相似文献   

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3.
Radix Ophiopogonis is a widely used traditional Chinese medicine. The quality of Radix Ophiopogonis available in the market varies, and some confusing or fake herbs exist. In order to improve the quality control of Radix Ophiopogonis, a novel fingerprinting method was established using HPLC coupled with UV and evaporative light-scattering detectors (ELSDs). Extraction with methanol and liquid-liquid extraction with water-saturated n-butanol were employed for the preparation of the sample solution. Chromatographic separation was performed on a Lichrospher C(18) column (250x4.6 mm id, 5.0 microm particle size) with a linear gradient elution program. UV detection at 280 nm and evaporative light-scattering detection were utilized to obtain two subfingerprinting chromatograms. A novel protocol for data processing was proposed, in order to identify and remove redundant data obtained by the two detectors, and balance the weight of the two subfingerprints on the similarity values. The method was validated and applied to quality evaluation of 16 samples of Radix Ophiopogonis and related herbs.  相似文献   

4.

HPTLC is a useful and practical analytical tool to characterize plant compositions. This study was focused on exploring the results of high-performance thin-layer chromatography (HPTLC) analysis, particularly as a useful tool for the authentication of Allanblackia parviflora seed and kernel cakes. Bulked samples from sixteen different Ghanaian communities were analysed by HPTLC and their fingerprints were compared. The optimum experimental conditions were established: sample weight of 2.0 g, methanol:water (80:20 v/v) as extraction solvent, 30 min extraction time and twice extraction, ethyl acetate:methanol:water (100:16.5:13.5 v/v) as mobile phase, vanillic acid as derivatisation agent and 7 min of plate heating time after derivatisation. The HPTLC profile generated from extracts across 16 communities and 157 trees was very reproducible and demonstrates the robustness of the technique in characterising the profile.

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5.
Ultra-performance liquid chromatography (UPLC) in reversed-phase (RP), ion pair (IP) and hydrophilic interaction chromatography (HILIC) has been investigated for the separation of imidazolium-based ionic liquid (IL) cations. Among the three stationary phases (i.e., C18, C8 and phenyl) studied under RP conditions the phenyl phase provided much stronger retention for the IL cations. Four acids (hydrochloric, methanesulfonic, perchloric and trifluoroacetic) as mobile phase additives were compared in light of their effects on the retention of IL cations. It was shown that the retention of all IL cations decreased upon acidification of the mobile phase, possibly due to suppression of residual silanol ionization. Very fast (~3 min) and efficient RP-UPLC separation of six cations was achieved by gradient elution with acetonitrile?Cwater mobile phase containing 2.5 mmol L?1 perchloric acid. In IP-UPLC all solutes were well resolved in about 4 min by gradient elution with acetonitrile?Cwater mobile phase containing 1 mmol L?1 sodium 1-octanesulfonate as ion pairing reagent. Finally, under HILIC conditions by using isocratic elution with acetonitrile?Cwater (85:15, v/v) mobile phase containing 5 mmol L?1 ammonium formate (pH 3.2) the separation time was reduced to less than 2 min while maintaining excellent peak shapes and sufficient resolution. Compared to current LC systems UPLC allowed considerably faster separations with better peak shapes.  相似文献   

6.
Monitoring and quantification of active pharmaceutical ingredients (APIs) in the environment constitute important and challenging tasks, as they are directly associated with human health. Three commonly used proton pump inhibitors (PPIs), namely, omeprazole sodium (OMP), pantoprazole sodium (PNT), and lansoprazole sodium (LNZ) are well separated and quantified using ultra-performance liquid chromatography (UPLC) in pharmaceutical industrial wastewater. The separation of the studied drugs was performed on a stationary phase with a WatersTM column (100 × 2.1 mm, 1.7 µm). The mobile phase was composed of methanol:0.05 M potassium dihydrogen phosphate buffer (adjusted to pH 7.5 using NaOH) (50:50, v/v). The elution process was done in gradient mode by changing the relative proportions of the mobile phase components with time to get an optimum separation pattern. The flow rate of the developing system was adjusted to 0.8 mL/minute. Detection of the separated drugs was performed at 230 nm. The studied drugs were quantified in the concentration range of 10–200 ng/mL for all drugs. The cited method was fully validated according to the international conference on harmonization (ICH-Q2B) guidelines, then it was applied successfully for quantification of the studied PPIs in real wastewater samples after their solid phase extraction (SPE).  相似文献   

7.
A reversed-phase liquid chromatographic method was established for the simultaneous determination of imperatorin and isoimperatorin in eleven kinds of traditional Chinese medicinal preparations (TCMPs) containing Radix Angelicae dahuricae (Chinese herbal name: Baizhi). Imperatorin and isoimperatorin were successfully separated on an Ultimate XB-C18 column (150 mm × 4.6 mm i.d., 5 μm). The mobile phase was a mixture of acetonitrile and 50 mmol L?1 sodium acetate (pH 3.5) buffer (45:55, v/v), employing isocratic elution at a flow rate of 1.0 mL min?1. Detection was accomplished at 302 nm. Regression equations revealed good linear relationship between the peak areas of the constituents and their concentrations (correlation coefficients: 0.9995 for imperatorin, 0.9991 for isoimperatorin). The recoveries were between 97.38% and 103.78%. The proposed method has been successfully applied to the simultaneous determination of imperatorin and isoimperatorin in eleven kinds of TCMPs containing Baizhi.  相似文献   

8.
A simple and fast method was developed for simultaneous quantitative determination of three biologically active phenolic compounds i.e. quercetin, rutin and coumaric acid in flowers of Rhododendron arboreum using high-performance thin-layer chromatography (HPTLC). The separation was performed on TLC aluminium plates precoated with silica gel RP-18 F254S. Good separation was achieved in the mobile phase of methanol-water-formic acid (40:57:3, v/v/v) and densitometric determination of these compounds was carried out at 280 nm in reflectance/absorbance mode. The linear regression data for the calibration plots showed a good linear relationship with r = 0.9971, 0.9953 and 0.9960 for quercetin, rutin and coumaric acid, respectively. Accuracy of the method was checked by recovery study conducted at two different levels with the average recovery of 99.90%, 99.02% and 99.16% for quercetin, rutin and coumaric acid, respectively. The present method is being reported first time and may be used for routine quality control of the flowers of R. arboreum.  相似文献   

9.
A sensitive LC‐MS/MS method was developed and validated for simultaneous quantification of 11 constituents, ginsenoside Rg1, Re, Rf, Rg2, Rb1, Rd, Rc, ophiopogonin D, schisandrin, schisandrol B and schizandrin B, in rat serum using digoxin as the internal standard (IS). The serum samples were pretreated and extracted with a two‐step liquid–liquid extraction. Chromatographic separation was achieved on a C18 analytical column with a proper gradient elution using 0.02% acetic acid aqueous solution and 0.02% acetic acid–acetonitrile as mobile phase at a flow rate of 0.5 mL/min. MS detection was performed using multiple reaction monitoring via an electrospray ionization source. Good linearity was observed in the validated concentration range for every analyte (r2 ≥0.9929), and the lower limits of quantitation of the analytes were in the range of 0.044–1.190 ng/mL in rat serum. Intra‐ and inter‐day precisions were <14.2%. The accuracy expressed as recovery was within the range of 85.1–112.8%. The extraction recoveries were >75.8%.The validated method was successfully applied to a pharmacokinetic study of all analytes in rats after single intravenous administration of Shengmai injection. Copyright © 2014 John Wiley & Sons, Ltd.  相似文献   

10.
In order to differentiate two species of Radix Puerariae (Radix Puerariae lobatae and Radix Puerariae thomsonii) and to determine major isoflavonoids (puerarin, daidzin, daidzein and genistein) in the samples, a simple high performance liquid chromatography (HPLC) method with isocratic elution employing cyclodextrins (CDs) as mobile phase additives was developed. Various factors affecting the retention of isoflavonoids in the C18 reversed-phase column, such as the nature of CDs, the concentration of hydroxypropyl-β-cyclodextrin (HP-β-CD) and the methanol percentage in the mobile phase, were studied. Experimental results confirmed that HP-β-CD, as a very effective mobile phase additive, could markedly reduce the retention of isoflavonoids, especially daidzein and genistein. The elution of four isoflavonoids could be achieved on a Kromasil® C18 column within 56 min by using the methanol–water contained 5 mM HP-β-CD (25/75, v/v) mixture as the mobile phase. The formation of the inclusion complexes between isoflavonoids and HP-β-CD explained the modification of the retention of analytes. The apparent formation constants determined by HPLC confirmed that the stoichiometry of HP-β-CD-isoflavonoid complexes was 1:1, and the stability of the complexes depended on the size and property of isoflavonoids. The optimized method was successfully applied for the simultaneous determination of major isoflavonoids in P. lobatae and P. thomsonii samples. This work provides a useful method for the analysis of traditional Chinese herbs.  相似文献   

11.
Toddalia asiatica (L.) Lam. (Rutaceae) has shown a broad spectrum of biological properties, such as anti-inflammatory, antioxidant, antimicrobial, anti-HIV, and anticancer properties. The present study is concerned with the separation of the main components with broad partition coefficients (KD values) from T. asiatica, using linear gradient high-speed counter-current chromatography (LGCCC) combined with an off-line two-dimensional (2D) mode. Similar to the binary gradient HPLC, the LGCCC mode is operated by the adjustment of the proportion between the mobile phase of 5:5:1:9 (v/v) (pump A) and 5:5:4.5:5.5 (v/v) (pump B) in an n-hexane/ethyl acetate/methanol/water solvent system. The off-line 2D-CCC mode was used in this study for the secondary separation of two similar KD value compounds with n-hexane/ethyl acetate/methanol/water (5:5:4:6, v/v). Notably, six coumarins, namely, tomentin (1), toddalolactone (2), 5,7,8-trimethoxycoumarin (3), mexoticin (4), isopimpinellin (5), and toddanone (6), were efficiently separated. The structures of the pure compounds were elucidated by spectral techniques and compared with the literature.  相似文献   

12.
A novel, validated, reversed-phase (RP), chiral high performance liquid chromatography (HPLC) method was developed for the enantiopurity control analysis of naproxen, a frequently used non-steroidal anti-inflammatory agent using polysaccharide-type chiral stationary phase (CSP). In the screening phase of method development, seven columns were tested in polar organic (PO) mode using mobile phases consisting of 0.1% acetic acid in methanol, ethanol, 2-propanol, and acetonitrile. Enantiorecognition was observed only in five cases. The best enantioseparation was observed on a Lux Amylose-1 column with 0.1% (v/v) acetic acid in ethanol with a resolution (Rs) of 1.24. The enantiomer elution order was unfavorable, as the distomer eluted after the eutomer. When the ethanolic mobile phase was supplemented with water, enantiomer elution order reversal was observed, indicating a difference in the enantiorecognition mechanism upon switching from PO to RP mode. Furthermore, by changing ethanol to methanol, not only lower backpressure, but also higher resolution was obtained. Subsequent method optimization was performed using a face-centered central composite design (FCCD) to achieve higher chiral resolution in a shorter analysis time. Optimized parameters offering baseline separation were as follows: Lux Amylose-1 stationary phase, thermostated at 40 °C, and a mobile phase consisting of methanol:water:acetic acid 85:15:0.1 (v/v/v), delivered with 0.65 mL/min flow rate. Using these optimized parameters, a Rs = 3.21 ± 0.03 was achieved within seven minutes. The optimized method was validated according to the ICH guidelines and successfully applied for the analysis of different pharmaceutical preparations, such as film-coated tablets and gel, as well as fixed-dose combination tablets, containing both naproxen and esomeprazole.  相似文献   

13.
Muehlenbeckia volcanica (Benth.) Endl. (M. volcanica), native to South America, is a traditional Peruvian medicinal plant that has multi-therapeutic properties; however, no phytochemicals have been identified from it yet. In this study, a five-step polarity-stepwise elution counter-current chromatography (CCC) was developed using methanol/water (1:5, v/v) as the stationary phase and different ratios of n-hexane, ethyl acetate, and n-butanol as mobile phases to separate the compounds from the 70% methanol extract of M. volcanica, by which six compounds with a wide range of polarities were separated in a single run of CCC and were identified as gallic acid, protocatechuic acid, 4,4′-dihydroxy-3,3′-imino-di-benzoic acid, rutin, quercitrin, and quercetin. Then, two compounds from the fractions of stepwise elution CCC were separated using conventional high-speed CCC, pH-zone-refining CCC, and preparative high-performance liquid chromatography, and identified as shikimic acid and miquelianin. These compounds are reported from M. volcanica for the first time. Notably, except for shikimic acid, all other compounds showed anti-diabetic potentials via antioxidant, antiglycation, and aldose reductase inhibition. The results suggest that the polarity-stepwise elution CCC can be used to efficiently separate or fractionate compounds with a wide range of polarities from natural products. Moreover, M. volcanica and its bioactive compounds are potent anti-diabetic agents.  相似文献   

14.

The lipase inhibitory activities of four main components from the rhizomes of Alisma orientale (Sam.) Juz. were evaluated by an in situ high-performance thin-layer chromatography (HPTLC)‒bioautographic assay taking orlistat as control standard. The order of relative activity was alisol B 23-acetate > alisol B > alisol A > alisol C 23-acetate. With that, an accurate, efficient and sustainable HPTLC method was developed to simultaneously determine the four lipase inhibitors from the methanolic extracts of Alismatis Rhizoma (AR). The method was carried out on HPTLC glassed plates (20 × 10 cm) coated with silica gel 60 F254 (0.2 mm thickness) using a mixture of cyclohexane and ethyl acetate (1:1, V/V) as the mobile phase. The RF values found for alisol B 23-acetate, alisol C 23-acetate, alisol B and alisol A were 0.62, 0.42, 0.28 and 0.09, respectively. The method was validated for specificity, linear range, precision, stability, and recovery. The results determined by scanning densitometry showed no significant difference to the results obtained by HPLC. The developed method was verified to be trustworthy for the evaluation of quality markers in AR.

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15.
Closantel is an antiparasitic drug marketed in a racemic form with one chiral center. It is meaningful to develop a method for separating and analyzing the closantel enantiomers. In this work, two enantiomeric separation methods of closantel were explored by normal-phase high-performance liquid chromatography. The influences of the chiral stationary phase (CSP) structure, the mobile phase composition, the nature and proportion of different mobile phase modifiers (alcohols and acids), and the column temperature on the enantiomeric separation of closantel were investigated in detail. The two enantiomers were successfully separated on the novel CSP of isopropyl derivatives of cyclofructan 6 and n-hexane-isopropanol-trifluoroacetic acid (97:3:0.1, v/v/v) as a mobile phase with a resolution (Rs) of about 2.48. The enantiomers were also well separated on the CSP of tris-carbamates of amylose with a higher Rs (about 3.79) when a mixture of n-hexane-isopropanol-trifluoroacetic acid (55:45:0.1, v/v/v) was used as mobile phase. Thus, the proposed separation methods can facilitate molecular pharmacological and biological research on closantel and its enantiomers.  相似文献   

16.
HPTLC-densitometric and HPLC–UV techniques were used for qualitative and quantitative determination of luteolin-7-O-glucuronide, lithospermic acid, rosmarinic acid and caffeic acid in several herbal drugs from the Lamiaceae family: Thymi herba, Serpylli herba, Majoranae herba and Menthae piperitae folium. Unmodified silica gel (HPTLC Si60) and silica gel chemically modified with aminopropyl groups (HPTLC NH2) were used during the investigation process. Among HPTLC methods the best resolution and selectivity was achieved with mobile phases: diisopropyl ether–acetone–formic acid–water (50:30:10:10, v/v/v/v) and acetone–formic acid (85:15, v/v), respectively. Plates were densitometrically evaluated. Contents of analyzed compounds in the studied aqueous extracts prepared from herbal drugs were established using both techniques. The results from the HPTLC-densitometric analysis have been compared with those from HPLC–UV on a C18 column with acetonitrile–water–formic acid as a mobile phase. The chromatographic methods were validated for linearity, LOD, LOQ, repeatability, intermediate precision and recovery. An analysis of variance showed that the HPTLC-densitometric and HPLC–UV methods are equivalent and sufficiently precise for the estimation of polyphenolic compounds mentioned above, in investigated herbal drugs. All of the suggested methods (HPTLC NH2, HPTLC Si60 and HPLC RP18) give results with good agreement.  相似文献   

17.
Pesticides are widely used in rice cultivation, often resulting in detection of their residues in rice grains. So far, no analytical method has been available for the simultaneous determination of most rice pesticides in rice grains. This paper reports the development and validation of such a method for the determination of eight rice pesticides (penoxsulam tricyclazole, propanil, azoxystrobin, molinate, profoxydim, cyhalofop-butyl, deltamethrin) and 3,4-dichloroaniline, the main metabolite of propanil. Pesticide extraction and clean-up was performed by an optimized matrix solid-phase dispersion (MSPD) protocol on neutral alumina (5 g) using acetonitrile as the elution solvent. Samples were analyzed in a high-performance liquid chromatography–diode array detection (HPLC-DAD) system. Pesticide separation was achieved with a mobile phase of acetonitrile/water in a linear elution gradient from 30:70% (v/v) to 100:0% (v/v) in 14 min at a flow rate of 0.8 mL min?1. Method validation was performed by means of linearity, intra-day accuracy, inter-day precision and sensitivity. Linear regression coefficients (R 2) were always above 0.9948. Limits of detection (LOD) and quantification (LOQ) varied from 0.002 to 0.200 mg kg?1 and 0.006 to 0.600 mg kg?1, respectively. Recoveries were investigated at three fortification levels and were found to be acceptable (74–127%) with relative standard deviations (RSD) below 12%. Application of the method for the analysis of five commercial rice grain samples showed that the pesticide levels were below the LOD. Overall, the method developed is suitable for the determination of residues of most rice pesticides in rice grains at levels below the established MRLs.  相似文献   

18.
Deng‐Zhan‐Sheng‐Mai capsules are a well‐known traditional Chinese patent medicine that was developed in China for the treatment of ischemic stroke. Its quality control focuses on Erigerontis Herba but ignores the contributions of Ginseng Radix et Rhizoma, Schisandrae Chinensis Fructus, and Ophiopogonis Radix. To improve the quality standards for this medicine, this work reports the application of a systematic ultra high performance liquid chromatography tandem mass spectrometric method coupled with chemometrics. Three qualitative and quantitative parameters are established for the evaluation of quality: chemical profiling, the relationship between the contents of 18 compounds and the antioxidant activity, and chemometric analysis. A total of 55 compounds, including 20 phenolic acids, 10 flavonoids, 15 saponins, and 10 lignans, were identified. The method for the quantitative determination of the aforementioned 18 compounds was validated. The limit of quantification ranged from 0.13 to 9.60 ng/mL. The overall recoveries ranged from 95.31 to 103.54%. Hierarchical cluster analysis and principal component analysis were applied to the data of 18 components in ten batches of samples. Nine compounds, including scutellarin, 3,5‐O‐dicaffeoylquinic acid, 4,5‐O‐dicaffeoylquinic acid, ginsenoside Rb1, ginsenoside Re, ginsenoside Rg1, ophiopogonin D, schisandrin, and schisandrol B, are suggested as chemical markers for evaluating the quality.  相似文献   

19.
An effective method for simultaneous separation of four flavonoids and two astragalosides was developed by semi-preparative high-performance liquid chromatography in this work. A crude sample was firstly cleaned-up from ethanol aqueous extract of Radix Astragali by LS-300B resin-based column chromatography and was further isolated by semi-preparative LC with a gradient elution of water–methanol. Six compounds including calycosin (3.5 mg), formononetin (2.6 mg), (6aR,11aR)-10-hydroxy-3,9-dimethoxypterocarpan (2.6 mg), (3R)-7,2′-dihydroxy-3′,4′-dimethoxyisoflavan) (2.0 mg), astragaloside I (5.4 mg) and astragaloside II (9.8 mg) were obtained with a recovery of 67.8, 77.2, 88.4, 85.3, 62.1 and 57.9%, respectively. Their chemical structures were confirmed by MS and NMR analysis. This study developed an effective and rapid method for simultaneous separation of multiple components from Radix astragali.  相似文献   

20.
In this study, we focused on the studying of taurine complexes with phenol and sodium hypochlorite, and of taurine with sodium hypobromite by spectrometry, reverse phase chromatography and ion-exchange chromatography. The formed complexes were studied under various conditions such as temperature (10, 20, 30, 40, 50 and 60 °C), and/or time of interaction (0, 5, 10, 15, 20, 25 and 30 min). In addition, we optimized high performance liquid chromatography coupled with UV detector for detection of taurine and its complexes with the acids. Taurine–phenol–hypochlorite complex was effectively separated under isocratic elution, mobile phase water:methanol 30:70 %, v:v, flow rate 1 mL min?1 and 55 °C. Taurine-bromamine complex was isolated under the following optimized conditions as isocratic elution, mobile phase water:methanol 85:15 % v:v, flow rate 1 mL min?1 and 55 °C. The limits of detection (3 S/N) were estimated as 1 μM for both types of complexes, i.e. for taurine. Further, we estimated recovery in one sample of urine (male 25 years), commercially achieved energy drink and tea leaves and varied from 79 to 86 %. Further, we aimed our attention at investigating the ability of the above characterized taurine and taurine complexes to scavenge reactive oxygen species. For this purpose, an ion-exchange liquid chromatography with post-column derivatization with ninhydrin and VIS detector was used. It clearly follows from the results obtained that taurine itself reacts with peroxide more intensely than in a bound form, which can be associated with the highest signal decrease. Complexes stabilized structure taurine against peroxide radicals, resulting in slower decreasing of peak heights. The most stable was taurine complexes with phenol and hypobromite.  相似文献   

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