Simultaneous determination of ferulic acid and gastrodin of Tianshu Capsule in rat plasma by ultra‐fast liquid chromatography with tandem mass spectrometry and its application to a comparative pharmacokinetic study in normal and migraine rats

Tianshu Capsule, consisting of Ligusticum chuanxiong Hort and Gastrodia elata Blume, is a widely used Traditional Chinese Medicine preparation for the treatment of migraine. Ferulic acid and gastrodin are main active constituents in Ligusticum chuanxiong Hort and Gastrodia elata Blume, and have been used as marker components for quality control of Tianshu Capsule. In this study, a selective, sensitive, and reliable ultra‐fast liquid chromatography with tandem mass spectrometry method was developed for simultaneous determination of ferulic acid and gastrodin in rat plasma using geniposide as internal standard. The plasma samples were extracted by protein precipitation with methanol after acidification and separated on a Shim‐Pack XR‐ODS C₁₈ column (75 × 3.0 mm, 2.2 μm) using gradient elution with a mobile phase consisting of water (containing 0.1% formic acid) and acetonitrile at a flow rate of 0.6 mL/min. Detection was performed on 3200 QTRAP mass spectrometry equipped with turbo ion spray source in negative ionization mode. Validation parameters were within acceptable ranges. The validated method was applied to compare the pharmacokinetic profiles of ferulic acid and gastrodin in normal and migraine rats. Our results showed that there were remarkable differences in the pharmacokinetic properties of the analytes between the normal and migraine groups.     

Ultra‐fast liquid chromatography with tandem mass spectrometry determination of eight bioactive components of Kai‐Xin‐San in rat plasma and its application to a comparative pharmacokinetic study in normal and Alzheimer's disease rats

A method of ultra‐fast liquid chromatography with tandem mass spectrometry was developed and validated for the simultaneous quantitation of eight bioactive components, including polygalaxanthone III, sibiricaxanthone B, tenuifolin, sibiricose A5, sibiricose A6, tenuifoliside A, ginsenoside Re and ginsenoside Rb1 in rat plasma after oral administration of Kai‐Xin‐San. The plasma samples were extracted by liquid–liquid extraction using digoxin as an internal standard. Chromatographic separation was performed on a Venusil MP C₁₈ column (100 mm × 2.1 mm, 3 μm) with methanol and 0.05% acetic acid in water as mobile phase. The tandem mass spectrometric detection was performed in the multiple reaction monitoring with turbo ion spray source in the negative ionization. Validation parameters were within acceptable ranges. The established method has been successfully applied to compare the pharmacokinetic profiles of the analytes between normal and Alzheimer's disease rats. The results indicated that there were significant differences in pharmacokinetic parameters of some components between two groups, which may be due to the mechanisms of Alzheimer's disease and pharmacological effects of the analytes. The pharmacokinetic research in the pathological state might provide more useful information to guide the clinical usage of herbal medicine.     

Simultaneous high‐performance liquid chromatography with tandem mass spectrometry quantification of six bioactive components in rat plasma after oral administration of Yougui pill

Yougui pills are a classic Chinese medicine that shows significant effects on nerve regeneration and neuroprotection in modern pharmacological studies. With a complex formula, Yougui pills have faced significant challenges in the fields of bioanalysis and pharmacokinetics in animals and human studies. In the present study, a specific and accurate high‐performance liquid chromatography with tandem mass spectrometry method was developed and validated for the quantitative determination of the six bioactive components in rat plasma after oral administration of Yougui pills. Chromatographic separation was performed on a C18 column with a gradient elution system. Samples were analysed using positive ion mode with multiple reaction monitoring mode. The assay showed good linearity for all six bioactive components in the dynamic range of 0.50 to 50 ng/mL with acceptable intra‐ and inter‐batch accuracy and precision. The lower limits of quantification were 0.50 ng/mL for all six bioactive components. The method was successfully applied to rat pharmacokinetics after oral administration of Yougui pills. All six bioactive components were detected in rat plasma, including songorine, benzoylhypaconitine, benzoylmesaconitine, neoline, karacoline, and sweroside, while some other target compounds were not detected, such as rhmannioside A, loganin, and cornuside I. After oral administration of Yougui pills at a dose of 2500 mg/kg, all six bioactive components were rapidly absorbed, resulting in t_max values less than 1 h and relative lower C_max values. The t_1/2 values for songorine, benzoylhypaconitine, benzoylmesaconitine, neoline, karacoline, and sweroside were calculated to be 2.62 ± 0.67, 2.11 ± 0.45, 1.94 ± 0.35, 1.88 ± 0.31, 2.07 ± 0.44, and 1.59 ± 0.30 h, which indicated that Yougui pills should be taken in multiple oral doses over a relatively short period.     

Determination of the transformation of ginsenosides in Ginseng Radix et Rhizoma during decoction with water using ultra‐fast liquid chromatography coupled with tandem mass spectrometry

This study was conducted to determine the variations of ginsenosides in Ginseng Radix et Rhizoma when using different preparation solvents and explore the major factors for changes. With an established ultra‐fast liquid chromatography coupled with tandem mass spectrometry method which could quantify 52 ginsenosides, the extraction differences were characterized and compared using different solvents (water, 70% aqueous ethanol, and ethanol). Subsequently, a series of aqueous solutions with different pH were prepared to test the influence of pH to the changes of ginsenosides. Meanwhile, acetic acid and aspartic acid were used to verify whether the reaction had a relationship with the kind of acids. After refluxing with water, not only highly polar ginsenosides were extracted, some less polar ginsenosides such as ginsenoside Rg₃, Rg₅, Rk₁, and Rh₂ occurred or increased rapidly. Further experiments indicated that less polar ginsenosides were easier to generate at low pH values, and the reaction was only related to pH other than what kind of acids were used. It is the first time to elaborate the contents variation of 52 ginsenosides when using different extraction methods. The results indicated that decoction with water could enhance the transformation of highly polar ginsenosides to less polar ginsenosides and the process was pH dependent.     

Simultaneous determination of eight bioactive constituents of Zhi‐Zi‐Hou‐Po decoction in rat plasma by ultra high performance liquid chromatography with tandem mass spectrometry and its application to a pharmacokinetic study

Zhi‐Zi‐Hou‐Po Decoction, consisting of Gardenia jasminoides Ellis, Magnolia officinalis Rehd. et Wils., and Citrus aurantium L, is a classical Traditional Chinese Medicine formula for the treatment of depression. In order to make good and rational use of this formula in the future, a sensitive, selective, and reliable ultra high performance liquid chromatography with tandem mass spectrometry method was developed for simultaneous determination of two iridoid glycosides (geniposide and genipin gentiobioside), two lignans (honokiol and magnolol), four flavonoid glycosides (isonaringin, naringin, hesperidin, and neohesperidin), the major bioactive constituents of Zhi‐Zi‐Hou‐Po Decoction, in rat plasma using paeoniflorin as internal standard. Plasma samples were pretreated by a simple protein precipitation with acetonitrile. Chromatographic separation was performed on a shim‐pack XR‐ODS C₁₈ column (75 × 3.0 mm, 2.2 µm) using gradient elution with mobile phase consisting of 0.1% formic acid aqueous solution and acetonitrile at a flow rate of 0.5 mL/min. Mass spectrometric detection was conducted on a 3200 QTRAP mass spectrometry equipped with electrospray ionization source in negative ionization mode. Quantification was performed using multiple reactions monitoring mode. Calibration curves exhibited good linearity (r > 0.9947) over a wide concentration range for all analytes, and the lower limits of quantification were 10, 5, 1, 5, 1, 5, 1, and 5 ng/mL for geniposide, genipin gentiobioside, honokiol, magnolol, isonaringin, naringin, hesperidin, and neohesperidin, respectively. The intraday and interday precisions at three quality control levels were less than 12.3% and the accuracies ranged from ?11.2 to 10.7%. Extraction recovery, matrix effect, and stability were satisfactory in rat plasma. The validated method was successfully applied to a pharmacokinetic study of the eight analytes after oral administration of Zhi‐Zi‐Hou‐Po decoction to rats.     

Simultaneous determination of ferulic acid,paeoniflorin, and albiflorin in rat plasma by ultra‐high performance liquid chromatography with tandem mass spectrometry: Application to a pharmacokinetic study of Danggui‐Shaoyao‐San

A rapid, selective, and sensitive ultra‐high performance liquid chromatography‐tandem mass spectrometry method was developed for simultaneous determination of ferulic acid, paeoniflorin, and albiflorin, the major active constituents of Danggui‐Shaoyao‐San, in rat plasma using geniposide as the internal standard. The plasma samples were processed by protein precipitation with acetonitrile, and then separated on a Shim‐Pack XR‐ODS C₁₈ column (75 mm × 3.0 mm, 2.2 μm) using gradient elution program with a mobile phase consisting of 0.1% aqueous formic acid and acetonitrile at a flow rate of 0.4 mL/min. The detection was achieved on a 3200 QTRAP mass spectrometer equipped with electrospray ionization source in negative ionization mode. Quantification was performed using multiple reaction monitoring mode by monitoring the fragmentation of m/z 192.9→134.0 for ferulic acid, m/z 525.0→120.9 for paeoniflorin, m/z 525.2→121.0 for albiflorin, and m/z 433.1→225.1 for the internal standard, respectively. The calibration curve was linear in the range of 5–2500 ng/mL for all the three analytes (r ≥ 0.9972) with the lower limit of quantitation of 5 ng/mL. The intraday and interday precisions were below 12.1% for all the analytes in terms of relative standard deviation, and the accuracy was within ±11.5% in terms of relative error. The extraction recovery, matrix effect and stability were satisfactory in rat plasma. The validated method was successfully applied to a pharmacokinetic study of ferulic acid, paeoniflorin, and albiflorin after oral administration of Danggui‐Shaoyao‐San to rats.     

Simultaneous determination and pharmacokinetics study of four quinones in rat plasma by ultra high performance liquid chromatography with electrospray ionization tandem mass spectrometry after the oral administration of Qianzhi capsules

A sensitive, specific, and accurate ultra high‐performance liquid chromatography with electrospray ionization tandem mass spectrometry method was developed and validated for the simultaneous quantification of purpurin, munjistin, mollugin, and alizarin from Qianzhi capsules in rat plasma. Chromatographic separation was performed on an Agilent Eclipse Plus C₁₈ RRHD column with a mobile phase consisting of methanol and 5 mM ammonium acetate/water with gradient elution. The analytes were quantified on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring mode and switching the electrospray ion source polarity with positive electrospray ionization in a single run. Samples were pretreated by liquid–liquid extraction with cyclohexane. The intra‐ and interday precision and accuracy of the assay were within acceptable ranges. Matrix effects for all of the analytes were between 90.16 and 100.21%. The average recovery ranged from 75.38 to 88.96%. This method was successfully applied to study the pharmacokinetic parameters of the four compounds in rat plasma after oral administration of Qianzhi capsules. Four quinones could be rapidly absorbed into blood (t_max, 0.80–1.93 h) and eliminated relatively slowly (t_1/2, 8.07–11.97 h). The results might be helpful for guiding the clinical application of Qianzhi capsules in the future.     

Simultaneous determination of three alkaloids,four ginsenosides and limonin in the plasma of normal and headache rats after oral administration of Wu‐Zhu‐Yu decoction by a novel ultra fast liquid chromatography‐tandem mass spectrometry method: application to a comparative pharmacokinetics and ethological study

A novel, sensitive and reliable ultra fast liquid chromatography‐tandem mass spectrometry (UFLC‐MS/MS) method has been developed and validated for simultaneous quantitation of eight main active ingredients (evodiamine, rutaecarpine, dehydroevodiamine, limonin, ginsenoside Rb₁, Rd, Re and Rg₁) in rat plasma after oral administration of Wu‐Zhu‐Yu (WZY) decoction, which is a celebrated and widely used Traditional Chinese Medicine formula for the treatment of headache. The analytes and internal standard (IS) were separated on a SHIM‐PACK XR‐ODS II column, and the detection was performed on a UFLC‐MS/MS system with turbo ion spray source. The lower limits of quantification were 1.5, 0.5, 1.0, 2.0, 2.0, 1.0, 0.5 and 0.2 ng ml^?1 for evodiamine, rutaecarpine, dehydroevodiamine, limonin, gensenoside Rb₁, Rd, Re and Rg₁, respectively. Linearity, accuracy, precision and absolute recoveries of the eight analytes were all within satisfaction. The IS‐normalized matrix factor was adopted for assessing the matrix effect and accompanied with a satisfactory result. The validated method has been successfully applied to compare pharmacokinetic profiles of the eight active ingredients in rat plasma between normal and headache rats after administration. Exact pharmaceutical effect of WZY decoction on headache was demonstrated by the ethological response of headache rats induced by nitric oxide donor after administration. The results indicated that the absorption of evodiamine, rutaecarpine, gensenoside Rb₁, Re and Rg₁ in headache group were significantly higher than those in normal group with similar concentration–time curves while no significant differences existed in limonin and ginsenoside Rd between the two groups. Copyright © 2013 John Wiley & Sons, Ltd.     

An ultra high performance liquid chromatography with tandem mass spectrometry method for simultaneous determination of thirteen components extracted from Radix Puerariae in rat plasma and tissues: Application to pharmacokinetic and tissue distribution study

A rapid and sensitive ultra high performance liquid chromatography with tandem mass spectrometry method was established and validated for simultaneous determination of thirteen bioactive components (gallic acid, protocatechuic acid, puerarin, p‐hydroxycinnamic acid, daidzin, ononin, daidzein, naringenin, genistein, apigenin, formononetin, biochanin A, and β‐sitosterol) of Radix Puerariae extract in rat plasma and tissues. The plasma and tissues samples were pretreated by protein precipitation extraction, and umbelliferone and rutin were used as internal standards. Sample separation was performed on a ZORBAX RRHD Eclipse plus C₁₈ column (2.1 mm × 50 mm, 1.8 µm, Agilent) with a mobile phase consisting of methanol–water (containing 0.1% formic acid). The mass spectrometry analysis was conducted in positive and negative ionization modes with multiple reaction monitoring. The lower limit of quantitation range for the 13 analytes was 0.2?35 ng/mL. The intra‐ and inter‐day precision of all the analytes were less than 10.92%, with an accuracy ranging from ?13.10 to 11.96%. Both the recovery and matrix effect were within acceptable limits. This method was successfully applied to pharmacokinetic and tissue distribution study of the 13 bioactive components in rats after oral administration of R. Puerariae extract.     

Comparative analysis of main bio‐active components in the herb pair Danshen‐Honghua and its single herbs by ultra‐high performance liquid chromatography coupled to triple quadrupole tandem mass spectrometry

A sensitive, reliable, and powerful ultra‐high performance liquid chromatography coupled to triple quadrupole tandem mass spectrometry method was developed for simultaneous quantification of the 15 main bio‐active components including phenolic acids and flavonoids within 13 min for the first time. The proposed method was first reported and validated by good linearity (r ² > 0.9975), limit of detection (1.12–7.01 ng/mL), limit of quantification (3.73–23.37 ng/mL), intra‐ and inter‐day precisions (RSD ≤ 1.92%, RSD ≤ 2.45%), stability (RSD ≤ 5.63%), repeatability (RSD ≤ 4.34%), recovery (96.84–102.12%), and matrix effects (0.92–1.02). The established analytical methodology was successfully applied to comparative analysis of main bio‐active components in the herb pair Danshen‐Honghua and its single herbs. Compared to the single herb, the content of most flavonoid glycosides was remarkably increased in their herb pair, and main phenolic acids were decreased, conversely. The content changes of the main components in the herb pair supported the synergistic effects on promoting blood circulation and removing blood stasis. The results provide a scientific basis and reference for the quality control of Danshen‐Honghua herb pair and the drug interactions based on variation of bio‐active components in herb pairs.     

Simultaneous assay of sildenafil and desmethylsildenafil in neonatal plasma by ultra‐performance liquid chromatography–tandem mass spectrometry

Sildenafil is used to treat pulmonary hypertension in neonatal and pediatric patients. Pharmacokinetic studies in these patients are complicated by the limited sample volume. We present the validation results of an assay method to quantitate sildenafil and desmethylsildenafil simultaneously in 50 µL of plasma. Deuterated sildenafil was used as an internal standard. After liquid–liquid extraction, analytes were separated on an ultra‐performance liquid chromatography (UPLC)‐column and quantified via tandem mass spectrometry. The calibration range was linear, with acceptable accuracy and a precision of <15% for both compounds. The lower limits of quantification were 1 ng/mL. Matrix effects were present, but inter‐plasma batch variability was under 12%. The method was successfully applied to samples from a pharmacokinetic study into sildenafil pharmacokinetics in neonates, making maximum use of the limited number and amount of plasma samples available. Copyright © 2009 John Wiley & Sons, Ltd.     

Comparative pharmacokinetic study of the components of Jia‐Wei‐Kai‐Xin‐San in normal and vascular dementia rats by ultra‐fast liquid chromatography coupled with tandem mass spectrometry

A fast, sensitive, and reliable ultra‐high performance liquid chromatography coupled with tandem mass spectrometry method has been developed and validated for simultaneous quantification of geniposide, polygalaxanthone III, 3,6′‐disinapoyl sucrose, α‐asarone, β‐asarone, poricoic acid A, poricoic acid B, dehydrotumulosic acid, deoxyschizandrin, schizandrin B, and kaempferide in plasma after oral administration of extracts of Jia‐Wei‐Kai‐Xin‐San in normal and vascular dementia rats. The developed method was precise and accurate within the linearity range of the analytes. The lower limits of quantification were 1.04–2.68 ng/mL for all the analytes. Both intra‐ and inter day precision and accuracy of the analytes were all within accepted criteria. The mean extraction recoveries of the analytes and the internal standard from rat plasma were all >60.0%. The validated method had been successfully applied to compare pharmacokinetic profiles of the analytes in plasma of normal and vascular dementia rat treated with herbal extracts. Results indicated that differences existed between normal and vascular dementia model rats except dehydrotumulosic acid and kaempferide, which might be due to the pathology of vascular dementia and pharmacological effect of the analytes. These pharmacokinetic studies might benefit for the mechanism exploration and clinical use of traditional Chinese medicine formulae.     

Simultaneous determination and pharmacokinetic study of four phenol compounds in rat plasma by ultra‐high performance liquid chromatography with tandem mass spectrometry after oral administration of Echinacea purpurea extract

A rapid and sensitive assay based on ultra‐high performance liquid chromatography with electrospray ionization tandem mass spectrometry was established and validated for the simultaneous determination of cichoric acid, chlorogenic acid, quinic acid, and caffeic acid in rat plasma after oral administration of Echinacea purpurea extract using butylparaben as the internal standard. Samples were pretreated by liquid–liquid extraction with ethyl acetate. The separations for analytes were performed on an ACQUITY UPLC HSS C₁₈ column (1.8 μm 2.1 × 100 mm) using a gradient elution program with acetonitrile/10 mM ammonium acetate (pH 5.6) at a flow rate of 0.3 mL/min. The analytes were detected in multiple reaction monitoring mode with negative electrospray ionization. The lower limit of quantification of each analyte was not higher than 10.85 ng/mL. The relative standard deviation of the intraday and interday precisions was less than 14.69%. The relative errors of accuracies were in the range of –13.80 to 14.91%. The mean recoveries for extraction recovery and matrix effect were higher than 80.79 and 89.98%, respectively. The method validation results demonstrated that the proposed method was sensitive, specific, and reliable, which was successfully applied to the pharmacokinetic study of four components after oral administration of Echinacea purpurea extract.     

Simultaneous quantification of twelve active components in Yiqing granule by ultra-performance liquid chromatography: application to quality control study

An ultra‐performance liquid chromatography (UPLC) method coupled with photodiode array (PDA) detection has been developed and validated for simultaneous quantification of 12 active components (berberine, palmatine, jatrorrhizine, aloe‐emodin, rhein, emodin, chrysophanol, physcion, baicalin, baicalein, wogonoside and wogonin) in Yiqing granule. Optimum separation were achieved on a C₁₈ column (50 × 2.1 mm i.d., 1.7 µm particle) through a 7.5 min gradient delivery of a mixture of A (acetonitrile) and B (0.1% aqueous phosphoric acid containing 1.8 mmol/L sodium dodecyl sulfonate and 10% acetonitrile, v/v) at a flow rate of 0.3 mL/min at 30°C. Because of the different UV characteristics of these components, three detection wavelengths were used for quantitative analysis. All of the analytes showed good linearity (r of >0.999). The method was validated for repeatability, precision, stability, accuracy and selectivity. The validated method was applied to quality control of Yiqing granule from different production batches. Copyright © 2010 John Wiley & Sons, Ltd.     

Untargeted metabolomic study on the insomnia effect of Suan‐Zao‐Ren decoction in the rat serum and brain using ultra‐high‐performance liquid chromatography quadrupole time‐of‐flight mass spectrometry combined with data processing analysis

Insomnia is a common clinical disease that can seriously damage the normal lives of sufferers. Suan‐Zao‐Ren decoction has been used to treat insomnia for a long time. However, the underlying molecular mechanism of Suan‐Zao‐Ren decoction is still not clear. In this study, the nontargeted metabolomics based on high‐resolution mass spectrometry and multiple statistical approaches were initially used to investigate the changes of potential serum and brain biomarkers and metabolic pathways in the insomnia model rat. Principal component analysis‐discriminate analysis indicated that the Suan‐Zao‐Ren decoction treatment improved the metabolic phenotype insomnia. Moreover, the heatmap analysis identified the most important biomarkers involved in insomnia. According to the pathway analysis, phenylalanine metabolism, tryptophan metabolism, and so on were recognized as the most affected metabolic pathways associated with insomnia disease. These findings provided a comprehensive understanding of the regulative effects of Suan‐Zao‐Ren decoction on the host metabolic phenotype of the insomnia rats. Our work demonstrated that the metabolomics approach is a promising tool that could help us to conduct the exploration of the therapeutic effects and mechanism of traditional Chinese medicines.     

A simple liquid chromatography coupled with tandem mass spectrometry approach for the simultaneous quantification of thirteen compounds in rats following oral administration of raw and processed Fructus Xanthii: Application in a comparative pharmacokinetic study

A simple and sensitive analysis using ultra high performance liquid chromatography with a tandem mass spectrometric system operated in selected reaction monitoring mode was developed for the determination of 11 phenolic acids, atractyloside, and carboxyatractyloside in rat plasma. The two classes of analytes were then separated on a Waters ACQUITY? UPLC HSS T3 column (50 mm × 2.1 mm, 1.8 µm) using gradient elution with a mobile phase of 0.2% formic acid in water containing 10 mM ammonium acetate and methanol. Detection was accomplished by selected reaction monitoring scanning via an electrospray source operating in negative ionization mode. The calibration curve was linear (R² = 0.990) over a concentration range of 1.20–3500 ng/mL, while the validated lower limit of quantification was 1.20 ng/mL. The precision varied from 0.84 to 4.62%, and the accuracy varied within ±5%. The method proved robust with sample freezing and thawing and with short‐ and long‐term sample storage. The established method was used for simultaneous quantification and was successfully used for the first time for the pharmacokinetic evaluation of 13 compounds after the intragastric administration of raw and processed Fructus Xanthii in rats. The results indicated that processing affects the absorption and metabolism of Fructus Xanthii extract. Importantly, the results also indicated the importance of processing for the clinical application of traditional Chinese medicine.     

Identification and analysis of chemical constituents and rat serum metabolites in Suan‐Zao‐Ren granule using ultra high performance liquid chromatography quadrupole time‐of‐flight mass spectrometry combined with multiple data processing approaches

Suan‐Zao‐Ren granule is widely used to treat insomnia in China. However, because of the complexity and diversity of the chemical compositions in traditional Chinese medicine formula, the comprehensive analysis of constituents in vitro and in vivo is rather difficult. In our study, an ultra high performance liquid chromatography with quadrupole time‐of‐flight mass spectrometry and the PeakView® software, which uses multiple data processing approaches including product ion filter, neutral loss filter, and mass defect filter, method was developed to characterize the ingredients and rat serum metabolites in Suan‐Zao‐Ren granule. A total of 101 constituents were detected in vitro. Under the same analysis conditions, 68 constituents were characterized in rat serum, including 35 prototype components and 33 metabolites. The metabolic pathways of main components were also illustrated. Among them, the metabolic pathways of timosaponin AI were firstly revealed. The bioactive compounds mainly underwent the phase I metabolic pathways including hydroxylation, oxidation, hydrolysis, and phase II metabolic pathways including sulfate conjugation, glucuronide conjugation, cysteine conjugation, acetycysteine conjugation, and glutathione conjugation. In conclusion, our results showed that this analysis approach was extremely useful for the in‐depth pharmacological research of Suan‐Zao‐Ren granule and provided a chemical basis for its rational.     

Simultaneous detection of multiple hydroxylated polychlorinated biphenyls from biological samples using ultra‐high‐performance liquid chromatography with isotope dilution tandem mass spectrometry

We established a method for the separation and detection of nine hydroxylated polychlorinated biphenyls in whole blood and urine samples using ultra high performance liquid chromatography coupled with electrospray negative ionization tandem mass spectrometry. Clean‐up procedures involved a filtration step, and optimization involved a pretreatment step consisting of a simple liquid–liquid extraction using hydrated silica‐gel chromatography (5%). Nine hydroxylated polychlorinated biphenyls were separated on an ultra high performance liquid chromatography HSS T3 column using a gradient elution program of 2 mmol ammonium formate aqueous solution (A) and methanol (B). Recovery ranged from 84.0 to 105.4% for the nine different hydroxylated polychlorinated biphenyls in urine with three spiked levels of 0.1, 1, and 2 ng and from 73.5 to 98.6% for the blood with spiked levels of 0.2, 1, and 2 ng. The relative standard deviations were <8.7% (n = 6), and the limits of detection in urine and whole blood for the nine hydroxylated polychlorinated biphenyls were in the range of 1.5–4 and 20–100 pg/g, respectively. This analytical method may enable the simultaneous detection of various hydroxylated polychlorinated biphenyls from complex tissue matrices.     

Simultaneous determination of eight bioactive components in rat plasma after oral administration of Yan‐Ke‐Ning‐Tablet by liquid chromatography coupled with Q‐Exactive Orbitrap tandem mass spectrometry

A rapid, sensitive and reliable quantitative method based on ultra‐high performance liquid chromatography coupled with Q‐Exactive Orbitrap tandem mass spectrometry was developed for simultaneous determination of berberine, berberrubine, palmatine, jatrorrhizine, columbamine, baicalin, baicalein and wogonin in rat plasma after oral administration with Yan‐Ke‐Ning‐Tablet (YKNT). After precipitation with acetonitrile, the plasma samples were separated on a reverse‐phase C₁₈ column with 1 mm ammonium acetate containing 0.2% acetic acid–acetonitrile as mobile phase. Calibration curves showed good linearity (r > 0.9983) over the tested concentration ranges of 0.5–200 ng/mL for berberine, berberrubine, palmatine, jatrorrhizine and columbamine, and 1–300 ng/mL for baicalin, baicalein and wogonin. The precision (relative standard deviation) at three different concentration levels was <12.15% and the accuracy (relative error) ranged from ?8.24 to 10.85%. No matrix effects were observed with matrix effect value ranging from 89.23 to 107.68%. The extraction recovery was in the range of 82.34–92.31%. The validated assay was further successfully applied to the pharmacokinetic study of these components after oral administration of YKNT. The present study provides the pharmacokinetic profiles of major bioactive components found in YKNT, and provides valuable information regarding the chemical components that were absorbed into plasma, which will be helpful for understanding the therapeutic effects of YKNT.