Improved LC Method for the Simultaneous Determination of Five Active Components in Danshen and Its Preparations

A novel LC method for the simultaneous determination of tanshinone IIA, tanshinone I, cryptotanshinone, dihydrotanshinone, and salvianolic acid B in Danshen and its preparations was developed on a 3 μm particle-sized ODS short column. The five analytes were well separated within 45 min with good linearities (r ² > 0.9999), precises (<4.0%) and recoveries (97.9–105.3%) by a gradient elution of acetonitrile-0.1% aqueous trifluoroacetic acid (v/v). In addition, ultrasonic extraction procedures for sample preparation were optimized using an orthogonal array design. Quantitative determinations on nine herb samples and four preparations demonstrated the developed approaches were suitable for standardization of this herb and its preparations.

     

Application of solid-phase extraction for determination of phenolic compounds in barrique wines

A fast, selective and sensitive chromatographic method has been developed for determination of gallic, protocatechuic, p-hydroxybenzoic, vanillic, caffeic, syringic, p-coumaric, benzoic, ferulic, sinapic, cinnamic, and ellagic acids and p-hydroxybenzaldehyde, vanillin, syringaldehyde, 2-furfural, 5-methylfurfural, and 5-methoxyfurfural. The compounds from untreated wine samples were pre-concentrated and cleaned using solid-phase extraction on RP-105 polymeric sorbent. The cartridge was conditioned with methanol and water. Co-extracted ballast substances were rinsed from the sorbent with 0.1 mol L^–1 hydrochloric acid–methanol, 1:4 (v/v). Retained phenolic compounds were selectively eluted with diethyl ether. A linear mobile phase gradient containing 0.3% acetic acid and methanol was used for final baseline chromatographic separation on a Hypersil BDS C18 column. Limits of detection (LOD=3s_bl) in the range 5.2 to 181.2 g L^–1, resolution (R) better than 1.7, and repeatability of 2.7–5.1% (RSD for real samples) were achieved. The method was applied for quantification of individual phenolic compounds in barrique wines.     

Pharmacokinetics of 13 active components in a rat model of middle cerebral artery occlusion after intravenous injection of Radix Salviae miltiorrhizae‐Lignum dalbergiae odoriferae prescription

As a representative formulation of Radix Salviae miltiorrhizae (Danshen)‐Lignum Dalbergiae odoriferae (Jiangxiang), Xiangdan injection is widely prescribed for cardio‐ and cerebrovascular diseases in practice. This necessitates a pharmacokinetic investigation of this formulation to make it safer and more broadly applicable. We developed and validated a sensitive, selective, and reliable high‐performance liquid chromatography with tandem mass spectrometry method for the simultaneous determination of 11 phenolic compounds including danshensu plus two diterpenoid quinones like cryptotanshinone and tanshinone IIA in rat. We applied this method for the pharmacokinetic studies of the 13 compounds in a rat model of middle cerebral artery occlusion after intravenous injection of Xiangdan injection or Danshen injection. In sham‐operated rats, the animals taking Xiangdan injection exhibited significant growth of the area under the curve for danshensu, protocatechuic aldehyde, and tanshinone IIA compared with the changes seen in the data of those administrated with Danshen injection. Such a pattern was also observed in middle cerebral artery occlusion rats, whereas increased the area under the curve values were observed for danshensu, protocatechuic aldehyde, caffeic acid, rosmarinic acid, and tanshinone IIA. These results demonstrated that synergistic interactions occurred between the components of Danshen and the active compounds of Jiangxiang both in sham‐operated and middle cerebral artery occlusion rats, increasing the bioavailability of Danshen. The results presented herein can be used to determine a reference dose for the clinical application of Xiangdan injection, and to elucidate the synergistic mechanism of Danshen and Jiangxiang.     

Isolation and Simultaneous Determination of Coumarin Compounds in Radix Angelica dahurica

For the first time simple, rapid, and systematic methods have been established for preparative isolation and purification of coumarin compounds in an important traditional Chinese Medicine, Radix Angelica dahurica, and for simultaneous determination of several of the compounds in the medicine. Bergapten, imperatorin, and cnidilin, three of the biologically active coumarin compounds, were isolated from the chloroform-soluble fraction of the ethanol extract of Radix Angelica dahurica. After further purification by open column ODS chromatography the purified components were simultaneously determined, with two other coumarins (osthole and isoimperatorin), by reversed phase high-performance liquid chromatography (RP-HPLC) on a C₁₈ column, with methanol–water, 66:34 (v/v), as mobile phase at a flow rate of 0.8 mL min⁻¹. The compounds were detected by UV absorption at 310 nm. Calibration plots for all the coumarins had correlation coefficients close to unity. Limits of detection (S/N = 3) were <92 ng mL⁻¹ and limits of quantification (S/N = 10) were <259 ng mL⁻¹. Mean recovery of the coumarins was in the range 96.7–101.9% and the intra-day and inter-day precision, as relative standard deviation, was <2.3 and <2.9%, respectively. This simple, sensitive, and reproducible method can be used for quality control of Radix Angelica dahurica.     

Simultaneous Determination of Six Major Constituents in the Stems of Kadsura heteroclita by LC-DAD

The stem of Kadsura heteroclita is a traditional Chinese medicine with a variety of biological activities. For efficient monitoring of the quality of the herb, a simple, rapid, and accurate HPLC method has been developed for simultaneous determination of five cyclolanostane triterpenoid compounds (schisanlactone E, nigranoic acid, schisandronic acid, and heteroclitalactones A and D) and one tetrahydrofuran lignan (d-epigalbacin) in the stems of K. heteroclita. These six compounds were separated on a C₁₈ column by elution with a mobile-phase gradient prepared from acetonitrile and 0.05% (v/v) aqueous phosphoric acid. The flow rate was 1.0 mL min⁻¹ and the detection wavelength was 210 nm. The recovery of the method was in the range 95.03–102.19% and a good linear relationship (r ² > 0.9999) was obtained between response and compound concentration over a relatively wide range. The method was successfully applied to analysis of the herb collected at different stages of growth.     

Determination of Apigenin by LC with Electrochemical Detection

A rapid, sensitive, and specific method for quantification of olmesartan, the prodrug of olmesartan medoxomil, in human plasma, using zidovudine as internal standard, is described. Sample preparation involved a simple solid-phase extraction procedure. The extract was analyzed by high-performance liquid chromatography coupled to electrospray tandem mass spectrometry (LC–MS–MS). Chromatography was performed isocratically on a 5 μm C₁₈ analytical column (50 mm × 4.6 mm i.d.) with water–acetonitrile–formic acid 20:80:0.1 (v/v) as mobile phase. The response to olmesartan was a linear function of concentration over the range 4.82–1,928 ng mL⁻¹. The lower limit of quantification in plasma was 4.82 ng mL⁻¹. The method was successfully applied in a bioequivalence study of an olmesartan formulation after administration as a single oral dose.     

Extraction of <Emphasis Type="Italic">p</Emphasis>-hydroxyacetophenone and catechin from Norway spruce needles. Comparison of different extraction solvents

The phenolic compounds p-hydroxyacetophenone and catechin have been extracted from Norway spruce needles with pure methanol, 80 and 50% (v/v) aqueous methanol, pure acetonitrile, 80% (v/v) aqueous acetonitrile, and pure water. Extraction efficiency of the individual solvents was compared. Although 80% aqueous methanol is the solvent most frequently used for extraction of soluble phenolic compounds from needles, it was found that pure methanol is a more suitable extraction solvent. Surprisingly, a two-step procedure based on the extraction of crushed needles with water then re-extraction with methanol proved a good alternative to direct extraction with methanol. Extraction of uncrushed spruce needles might indicate that relatively more p-hydroxyacetophenone than catechin was located in the surface layer of the needle.     

Determination and Pharmacokinetic Study of 10-O-Dimethylaminoethylginkgolide B in Rat Plasma by LC–MS

To evaluate the pharmacokinetics of a novel analogue of ginkgolide B, 10-O-dimethylaminoethylginkgolide B (XQ-1) in rat plasma in pre-clinical studies, a sensitive and specific liquid chromatographic method with electrospray ionization mass spectrometry detection (LC–ESI–MS) was developed and validated. After a simple extraction with ethyl acetate, XQ-1 was analyzed on a Shim-pack C18 column with a mobile phase of a mixture of 1 μmol L⁻¹ ammonium acetate containing 0.02% formic acid and methanol (55:45, v/v) at a flowrate of 0.3 mL min⁻¹. Detection was performed in selected ion monitoring (SIM) mode using target ions at [M + H]⁺ m/z 496.05 for XQ-1 and m/z 432.10 for the internal standard (lafutidine). Linearity was established for the concentration range from 2 to 1,000 ng mL⁻¹ . The extraction recoveries ranged from 86.0 to 89.9% in plasma at concentrations of 5, 50, and 500 ng mL⁻¹. The lower limit of quantification was 2 ng mL⁻¹ with 100 μL plasma. The validated method was successfully applied to a pharmacokinetic study after intragastic administration of XQ-1 mesylate in rats at a dose of 20 mg kg⁻¹.     

Determination of 6-Benzyladenine in Bean Sprout by LC–ESI-IT-MS–MS

A rapid and sensitive method has been developed and validated to determine 6-benzyladenine (6-BA) residues in bean sprout using high-performance liquid chromatography coupled with electrospray ionization ion trap tandem mass spectrometry. Liquid–liquid extraction, using acidified methanol, was used to isolate 6-BA from the sample matrix. The separation was carried out on a Zorbax SB C₁₈ column (150 mm × 4.6 mm i.d., 5 μm) with 0.1% acetic acid/methanol (25:75 v/v) as mobile phase in isocratic mode. The quantitation of 6-BA was based on fragmentation of the molecular ion at m/z 226.1 to a product ion at m/z 91.1. In addition, the recoveries of three extraction solvents by ultrasound extraction and conventional extraction method were compared, and it was found that acidified methanol as the extraction solvent was the best. The calibration curve was linear (r ² > 0.997) in the concentration range of 0.04–10.0 ng mL⁻¹ with a lower limit of quantification of 0.5 ng g⁻¹ for 6-BA in bean sprout. Intra-day and inter-day relative standard deviations (RSDs) were less than 7.1 and 9.8%. Recoveries of 6-BA were between 85.0 and 88.5%. This assay has been successfully applied to the determination of trace 6-BA residues in bean sprout and monitoring the market in Ningbo, China.     

A Simple and Validated LC Method for the Simultaneous Determination of Three Compounds in Mikania laevigata Extracts

The Mikania genus is widely known as guaco and is used to treat fever, rheumatism, influenza and respiratory diseases. This article deals with the simultaneous quantification of three commercially available phenolic markers (o-coumaric acid, coumarin and syringaldehyde) in M. laevigata extracts, through LC-PDA. The validation data show that the method is specific, accurate, precise and robust, and also indicative of the stability of guaco extract. The method was linear, over a range of 1.25–20.0 μg mL⁻¹ for o-coumaric acid, 2.5–40.0 μg mL⁻¹ for coumarin, and 0.25–4.0 μg mL⁻¹ for syringaldehyde. The range of recovery was 94.3–96.4% for all the components, at a level of 100%.     

SPE of Tanshinones from Salvia miltiorrhiza Bunge by using Imprinted Functionalized Ionic Liquid-Modified Silica

Ionic liquid-modified silica, with functional groups based on imidazole as the cation, was obtained. A molecular imprinting technique was introduced to form the order of functional groups. The selectivity of the obtained ionic liquid-modified silica was successfully used as a special imprinted sorbent in the solid-phase extraction to isolate cryptotanshinone, tanshinone I and tanshinone IIA from Salvia miltiorrhiza Bunge. Several washing and elution solvents with different polarities were evaluated. The ionic liquid-modified silica as the sorbent exhibited a higher selectivity than blank ionic liquid-modified silica, traditional silica and C₁₈ cartridges. A quantitative analysis was conducted by liquid chromatography with a C₁₈ column and methanol/water (75:25, v/v, containing 0.5% acetic acid) as the mobile phase. A good linearity was obtained from 0.5 × 10^?4 to 0.1 mg mL^?1 (r ²  > 0.99) with relative standard deviations that were less than 4.6%.     

Quantification of cis-Abienol in Oriental Tobacco Leaves by LC

A rapid and accurate method for the quantification of cis-abienol in oriental tobacco leaves by normal phase liquid chromatography was developed. Freeze-dried tobacco samples were sonicated in methylene chloride for 10 min. The supernatant was purified using a silica gel solid phase extraction cartridge. Ten milliliter of the resulting methylene chloride eluate was collected, then separated on a 250 × 4.6 mm, 5 μm particle-size CN column with n-hexane: ethyl acetate, 100:2 (v/v) at a flow rate of 1 mL min⁻¹. cis-Abienol was detected by UV absorption at 254 nm. The linear range was from 2.14 × 10⁻⁴ to 4.28 × 10⁻² mg mL⁻¹ and the correlation coefficient was 1.000. The average recovery was 98.7, 105.2 and 103.1% in five replicated sets of tobacco samples spiked with 0.2856, 0.7140 and 1.904 mg cis-abienol. The relative standard deviations (RSDs) were 1.04, 0.63 and 1.25%, respectively (n = 5). Limit of detection (S/N = 3) was 21.84 μg g⁻¹ and limit of quantification (S/N = 10) was 72.80 μg g⁻¹. The method was found to be suitable for determination of cis-abienol in oriental tobacco leaves. Furthermore, pure cis-abienol used for method validation was obtained by preparative reversed phase high-performance liquid chromatography. Identification was performed by UV detection, nuclear magnetic resonance and mass spectrometry.     

LC Method for Determination of Ginkgolic Acids in Mice Plasma and Its Application to a Pharmacokinetic Study

A sensitive and simple LC method for the quantification of ginkgolic acids in mice plasma has been developed. Following acetonitrile deproteinization, samples were separated on a SinoChrom ODS-AP C₁₈ column. The mobile phase was 3% (v/v) acetic acid water solution–methanol (8:92, v/v) at a flow rate of 1.0 mL min⁻¹. Detection was at 310 nm. Calibration curve was linear over the range of 0.25–50 μg mL⁻¹ with intra- and inter-day precisions (RSD%) of less than 9.5%. The extraction recovery ranged from 87.0 to 90.2% (RSD 2.4–6.4%) for ginkgolic acids. The method was successfully applied to the pharmacokinetic study of ginkgolic acids in mice after oral dosing of 1.0 g kg⁻¹.

     

HPLC Determination and Pharmacokinetic Study of Valdecoxib in Human Plasma

A sensitive, simple, and accurate high-performance liquid chromatographic method has been developed for determination of valdecoxib and the internal standard rofecoxib in human plasma. Protein was precipitated from plasma samples by addition of perchloric acid (HClO₄); the drug was then extracted with diethyl ether. Separation was performed on a Cosmosil C₁₈ column (150 mm × 4.6 mm i.d., 5 m particles) with ammonium acetate buffer-acetonitrile, 60:40 (v/v), containing 0.1% TEA, pH 6.5, as mobile phase. Detection and quantification were performed by UV-visible detection at 239 nm. Detection and quantification limits were 3 and 5 ng mL^–1, respectively. The linear concentration range for valdecoxib was 5–400 ng mL^–1. The validated RP HPLC method was used for determination of the pharmacokinetic data for the drug in humans.     

Pharmacokinetic comparison of five tanshinones in normal and arthritic rats after oral administration of Huo Luo Xiao Ling Dan or its single herb extract by UPLC‐MS/MS

A fast, sensitive and reliable ultra performance liquid chromatography–tandem mass spectrometry (UPLC‐MS/MS) method has been developed and validated for simultaneous quantitation and pharmacokinetic study of five tanshinones (tanshinone I, tanshinone IIA, tanshinone IIB, dihydrotanshinone I, cryptotanshinone), the bio‐active ingredients of Huo Luo Xiao Ling Dan (HLXLD) in rat plasma. After liquid–liquid extraction, chromatographic separation was accomplished on a Shim‐pack XR‐ODS column (75 × 3.0 mm, 2.2 µm particles) and eluted with a mobile phase consisting of acetonitrile–0.05% formic acid aqueous solution (80:20, v/v) at a flow rate of 0.4 mL/min, and the total run time was 7.0 min. The detection was performed on a triple quadrupole tandem mass spectrometry equipped with an electrospray ionization source in positive ionization and multiple reaction monitoring mode. The lower limits of quantification were 0.050–0.400 ng/mL for all the analytes. Linearity, precision and accuracy, the mean extraction recoveries and matrix effects all satisfied criteria for acceptance. This validated method was successfully applied to a comparative pharmacokinetic study of five bio‐active components in rat plasma after oral administration of HLXLD or Salvia miltiorrhiza extract in normal and arthritic rats. The results showed that there were different pharmacokinetic characteristics among different groups. Copyright © 2016 John Wiley & Sons, Ltd.     

MSPD procedure for determining buprofezin,tetradifon, vinclozolin,and bifenthrin residues in propolis by gas chromatography–mass spectrometry

A simple and effective extraction method based on matrix solid-phase dispersion (MSPD) was developed to determine bifenthrin, buprofezin, tetradifon, and vinclozolin in propolis using gas chromatography–mass spectrometry in selected ion monitoring mode (GC–MS, SIM). Different method conditions were evaluated, for example type of solid phase (C₁₈, alumina, silica, and Florisil), the amount of solid phase and eluent (n-hexane, dichloromethane, dichloromethane–n-hexane (8:2 and 1:1, v/v) and dichloromethane–ethyl acetate (9:1, 8:2 and 7:3, v/v)). The best results were obtained using 0.5 g propolis, 1.0 g silica as dispersant sorbent, 1.0 g Florisil as clean-up sorbent, and dichloromethane–ethyl acetate (9:1, v/v) as eluting solvent. The method was validated by analysis of propolis samples fortified at different concentration levels (0.25 to 1.0 mg kg⁻¹). Average recoveries (four replicates) ranged from 67% to 175% with relative standard deviation between 5.6% and 12.1%. Detection and quantification limits ranged from 0.05 to 0.10 mg kg⁻¹ and 0.15 to 0.25 mg kg⁻¹ propolis, respectively.     

Quantitation and identity confirmation of residues of quinolones in tilapia fillets by LC-ESI-MS-MS QToF

A method for simultaneous determination of flumequine (FLM), oxolinic acid (OXO), sarafloxacin (SAR), danofloxacin (DAN), enrofloxacin (ENR), and ciprofloxacin (CIP) in tilapia (Orechromis niloticus) fillets, using liquid chromatography-tandem mass spectrometry (LC-ESI-MS-MS QToF) is presented. The quinolones were extracted from the food matrix with a solution of 10% trichloroacetic acid-methanol (80:20 v/v) with ultrasonic assistance. Clean-up of the extract solution was performed by using polymeric solid-phase extraction cartridges. The LC separation was carried out on an octadecyl hybrid silica column (C₁₈, 150 mm × 3 mm, 5 μm). The column temperature was set at 30 °C, and gradient elution (0.2 mL min⁻¹) was performed using water and acetonitrile, both containing 0.1% of acetic acid, as mobile phase components. The analytes were ionized using electrospray in the positive polarity mode. The following analytical results were obtained: linearity was about 0.99 for all the quinolones; intra and inter-assay precision (RSD%) were lower than 12.7 and 20%, respectively; and recoveries were from 89 to 112%. The quantitation limits were below the maximum residue limits established for the analytes. The method is suitable for the determination of quinolone residues in fish fillets and the QToF technique made it possible to obtain m/z ratios with less than 10 ppm of error for each analyte.     

Simultaneous Determination of Four Active Components in Tobacco Wastes by LC

A liquid chromatographic method was developed for the simultaneous quantification of four major active components in tobacco (Nicotiana tobaccum L.) wastes. Samples were extracted with 70% v/v aqueous methanol, four compounds including chlorogenic acid, cryptochlorogenic acid, neochlorogenic acid and caffeic acid were identified and determined by using LC coupled to electrospray tandem mass spectrometry and LC–UV method, respectively. Separation in LC–UV was on an Alltima C₁₈ column (250 mm × 4.6 mm i.d.; 5 μm) with a mobile phase consisting acetonitrile: ammonium acetate buffer (pH 4.5) (5:95 v/v), at a flow rate of 1.0 mL min⁻¹, detected at 327 nm. Four regression equations showed good linear relationships (r ² > 0.999) between the peak area of each marker and concentration. The method has good repeatability and precision, the intra-day and inter-day RSD for both retention time and peak area was less than 1.0%. The recoveries, measured at three concentration levels, varied from 96.33 to 101.10%. The LOD (S/N = 3) and LOQ (S/N = 6) were less than 0.010 and 0.795 μg·mL⁻¹, respectively. This assay was successfully applied to the determination of four active compounds in ten samples. The results indicated that the developed assay method was rapid, accurate, reliable and could be readily utilized as a quantitative analysis method for various of tobacco wastes.

     

A UFLC–MS/MS method for the simultaneous determination of eight bioactive constituents from red wine and dealcoholized red wine in rat plasma: Application to a comparative pharmacokinetic study

To explore whether alcohol has an effect on the pharmacokinetic behavior of phenolic acids, the main bioactive constituents in red wine, a highly sensitive and simple ultra‐fast liquid chromatography coupled with triple quadrupole mass spectrometry (UFLC–MS/MS) method was developed for simultaneous quantitation of eight phenolic acids in plasma samples. Plasma samples were extracted by liquid–liquid extraction and the chromatographic separation was achieved on a Zorbax SB‐C₁₈ column within 7.0 min. Results of the validated method revealed that all of the calibration curves displayed good linear regression (r > 0.99). The intra‐ and inter‐day precisions of the analytes were <14.0% and accuracies ranged from ?8.5 to 7.3%. The extraction recoveries of the analytes were from 71.2 to 110.2% and the matrix effects ranged from 86.2 to 105.5%. The stability of these compounds under various conditions satisfied the requirements of biological sample measurement. The method was successfully applied to a comparative pharmacokinetic study of phenolic acids in rat plasma. For gallic acid and gentisic acid, the parameters AUC_0–t and AUC_0–∞ increased remarkably (p < 0.05) after oral administration of red wine, which suggested that alcohol might enhance their absorption. This is the first report to compare the pharmacokinetic behavior of phenolic acids in red wine and dealcoholized red wine.     

Chemical Fingerprint and Quantitative Analysis of Cirsium setosum by LC

A reverse phase liquid chromatography method with diode array detection was developed to evaluate the quality of Cirsium setosum through establishing chromatographic fingerprint and simultaneous determination of six phenolic compounds, namely chlorogenic acid, caffeic acid, rutin, linarin, luteolin and apigenin. The chromatographic separation was performed on an Agilent SB-C₁₈ column (250 × 4.6 mm, 5.0 μm) with a gradient elution program using a mixture of acetonitrile and 0.5% aqueous acetic acid (v/v) as mobile phase within 25 min at 326 nm wavelength. The correlation coefficients of similarity were determined from the LC fingerprints, and they shared a close similarity. The LC with electrospray ionization mass spectrometry experiment was performed to further confirm the identity of phenolic compounds. The six phenolic compounds showed good regression (R ² > 0.9995) within test ranges and the recovery of the method was in the range of 95.8–102.8%. In addition, the content of those six phenolic compounds in C. setosum growing in different locations of China was determined to establish the effectiveness of the method. The results indicated that the developed method by having a combination of chromatographic fingerprint and quantification analysis could be readily utilized as a quality control method for C. setosum and its related traditional Chinese medicinal preparations.