Yidi Yin

Ultra‐fast LC–ESI‐MS/MS method for the simultaneous determination of six highly toxic Aconitum alkaloids from Aconiti kusnezoffii radix in rat plasma and its application to a pharmacokinetic study

A fast, sensitive, and efficient ultra-fast LC-ESI-MS/MS method was developed for the simultaneous quantitation of six highly toxic Aconitum alkaloids, that is, aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, and benzoylhypaconine, in rat plasma after oral administration of crude ethanol extracts from Aconiti kusnezoffii radix by ultrasonic extraction, reflux extraction for 1 h, and reflux extraction for 3 h, respectively. The separation of six Aconitum alkaloids and aminopyrine (internal standard) was performed on an InertSustain® C18 column, and the quantification of the analytes was performed on a 4000Q ultra-fast LC-MS/MS system with turbo ion spray source in the positive ion and multiple-reaction monitoring mode. Absolute recoveries ranged within 65.06-85.1% for plasma samples. The intra- and interday precision and accuracy of analytes were satisfactory. The methods were validated with sensitivity reaching the lower LOQ for aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, and benzoylhypaconine, which were 0.025, 0.025, 0.050, 0.025, 0.025, and 0.100 ng/mL, respectively. The method was successfully applied to a pharmacokinetic study of six Aconitum alkaloids in rat plasma after oral administration of crude ethanol extracts from the raw root of Aconitum kusnezoffii Reichb. by three different extraction processes.

Determination of catecholamines and their metabolites in rat urine by ultra‐performance liquid chromatography–tandem mass spectrometry for the study of identifying potential markers for Alzheimer's disease

In order to investigate the potential links between catecholamines (CAs) and Alzheimers disease (AD), rapid and sensitive ultra-performance liquid chromatography (UPLC)-tandem mass spectrometry (MS/MS) methods in different ionization modes for the quantification of 14 CAs and their metabolites in rat urine without derivatization or complex sample pre-treatments were developed. After addition of the internal standard, isoproterenol, the urine samples were extracted by protein precipitation and separated on an Inertsil ODS-EP column (Shimadzu, Japan) at a flow of 1.0 ml min(-1). Tandem mass spectrometric detection was performed on a 4000Q UPLC-MS/MS in the multiple reaction monitoring mode with turbo ion spray source. Tyrosine, dopamine, noradrenaline, epinephrine, 3-methoxytyramine, normetanephrine and metanephrine were determined in positive mode, while 3,4-dihyroxy-L-phenylalanine (DOPA), 3,4-dihydroxyphenylacetic acid, DL-3,4-dihydroxymandelic acid, DL-3,4-dihydroxyphenyl glycol, homovanillic acid, DL-4-hydroxy-3-methoxymandelic acid and 4-hydroxy-3-methoxy-phenylglycol were determined in negative mode. The methods were examined and were found to be precise and accurate within the linearity range of the assays. The intra-day and inter-day precision and accuracy of the analytes were well within acceptance criteria (±15%). The mean extraction recoveries of analytes and internal standard were all more than 60%. The validated methods have been successfully applied to compare CAs profiles in normal and AD rats. The results indicated the urine levels of DL-3,4-dihydroxyphenyl glycol and 4-hydroxy-3-methoxy-phenylglycol in AD rats were significantly higher than those in the normal group, and the other CAs have an opposite performance. These may attribute to the difference of some enzyme activity between rats with AD and normal. Furthermore, this may be helpful in clinical diagnostics and monitor the efficacy of AD treatment.

Enrichment and purification of six Aconitum alkaloids from Aconiti kusnezoffii radix by macroporous resins and quantification by HPLC–MS

Aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine and benzoylhypaconine are six main Aconitum alkaloids from traditional Chinese medicine, Aconiti kusnezoffii radix, which possess highly bioactive as well as highly toxic character for medicinal use. In the present study, for the purpose of better utilizing the toxic herbal material, the performance characteristics of NKA-II, D101, X-5, AB-8, S-8, HPD722 and HPD750 macroporous resins for the enrichment and purification of these six Aconitum alkaloids were critically evaluated. Results showed that NKA-II offered the best adsorption and desorption capacities for six Aconitum alkaloids among the seven macroporous resins tested, which were affected significantly by the pH value. Subsequently, dynamic adsorption and desorption experiments had been carried out with the column packed by NKA-II resin to optimize the separation process of six Aconitum alkaloids. After one run treatment with NKA-II resin, the content of total six Aconitum alkaloids were increased from 5.87% to 60.3%, the recovery was 75.8%. Meanwhile, a validated HPLC-MS method had been developed to qualitative and quantitative these six Aconitum alkaloids. This method would provide scientific references to the large-scale production of six Aconitum alkaloids from Aconiti kusnezoffii radix or other plants and might also expand the secure application of these highly toxic components for pharmacy.

A UFLC-MS/MS method with a switching ionization mode for simultaneous quantitation of polygalaxanthone III, four ginsenosides and tumulosic acid in rat plasma: application to a comparative pharmacokinetic study in normal and Alzheimer's disease rats

A fast, sensitive and reliable ultra fast liquid chromatography-tandem mass spectrometry (UFLC-MS/MS) method has been developed and validated for simultaneous quantitation of polygalaxanthone III (POL), ginsenoside Rb1 (GRb1), ginsenoside Rd (GRd), ginsenoside Re (GRe), ginsenoside Rg1 (GRg1) and tumulosic acid (TUM) in rat plasma after oral administration of Kai-Xin-San, which plays an important role for the treatment of Alzheimers disease (AD). The plasma samples were extracted by liquid-liquid extraction using ethyl acetate-isopropanol (1:1, v/v) with salidrdoside as internal standard (IS). Good chromatographic separation was achieved using gradient elution with the mobile phase consisting of methanol and 0.01% acetic acid in water. The tandem mass spectrometric detection was performed in multiple reaction monitoring mode on 4000Q UFLC-MS/MS system with turbo ion spray source in a negative and positive switching ionization mode. The lower limits of quantification were 0.2-1.5 ng/ml for all the analytes. Both intra-day and inter-day precision and accuracy of analytes were well within acceptance criteria (±15%). The mean absolute extraction recoveries of analytes and IS from rat plasma were all more than 60.0%. The validated method has been successfully applied to comparing pharmacokinetic profiles of analytes in normal and AD rat plasma. The results indicated that no significant differences in pharmacokinetic parameters of GRe, GRg1 and TUM were observed between the two groups, while the absorption of POL and GRd in AD group were significantly higher than those in normal group; moreover, the GRb1 absorbed more rapidly in model group. The different characters of pharmacokinetics might be caused by pharmacological effects of the analytes.

Simultaneous quantitation of polygalaxanthone III and four ginsenosides by ultra‐fast liquid chromatography with tandem mass spectrometry in rat and beagle dog plasma after oral administration of Kai‐Xin‐San: Application to a comparative pharmacokinetic study

A fast, selective, and quantitative ultra-fast liquid chromatography with tandem mass spectrometry method has been developed and validated for the simultaneous quantitation of polygalaxanthone III, ginsenoside Rb1, ginsenoside Rd, ginsenoside Re, and ginsenoside Rg1 in the plasma of rat and beagle dog after oral administration of Kai-Xin-San. After addition of the internal standard, salidroside, the plasma samples were extracted by liquid-liquid extraction and separated on a Venusil MP C18 column with methanol/0.01% acetic acid water as mobile phase. The tandem mass spectrometric detection was performed in the multiple reaction monitoring with turbo ion spray source in a switching ionization mode. The method was examined, and found to be precise and accurate with the linearity range of the compounds. The intra- and interday precision and accuracy of the analytes were well within acceptance criteria (±15%). The mean extraction recoveries of analytes and internal standard were all >75.0%. The validated method has been successfully applied to comparing pharmacokinetic profiles of analytes in rat and beagle dog plasma. The results indicated that no significant differences were observed in pharmacokinetic parameters of ginsenoside Rg1, while the others had significant differences, which may due to the different mechanisms of absorption and metabolism.

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.

Combinative method using multi-components quantitation by single reference standard and HPLC fingerprint for comprehensive evaluation of Rhodiola crenulata H.Ohba

A combinative method using HPLC-DAD quantitative analysis and fingerprints were developed to evaluate the quality of Rhodiola crenulata H.Ohba (R. crenulata) from different origins. In this study, three phenolic compounds were first simultaneously determined by the single standard to determine multi-components (SSDMC) method. The chromatographic separation was performed on a Merck Purospher STAR RP-18 column (4.6 × 250 mm, 5 μm) with a gradient-elution program within 28 min at 275 nm wavelength. The method has achieved desired specificity, linearity (r2 ≥ 0.9999), precision, accuracy (95–105%), stability and robustness. Compared with the normal external standard method (ESM), this alternative method can be used for simultaneous determination of multiple indices effectively and accurately, and can resolve the problem of inadequate chemical standards. Moreover, the herbal chromatographic fingerprint was calculated using a similarity evaluation system and SPSS 19.0 software as a result of analysing all of the R. crenulata samples. The obtained data showed good repeatability and stability of the chromatographic fingerprint. The 15 R. crenulata samples from different origins were classified by hierarchical clustering analysis (HCA) and principal components analysis (PCA) according to the characteristic of common peaks. Moreover, similarity values were all more than 0.90. This study demonstrated that a combination of the chromatographic quantitative analysis and fingerprint offers an efficient way to quality consistency evaluation of R. crenulata.

Plasma N-acetylputrescine, Cadaverine and 1,3-diaminopropane: Potential Biomarkers of Lung Cancer Used to Evaluate the Efficacy of Anticancer Drugs

Polyamines have been widely investigated as potential biomarkers for various types of cancers, including lung cancer, which is one of the most common causes of death from cancer worldwide. This study was carried out to evaluate the value of polyamines that serve as early diagnostic and cancer progression markers as well as drug evaluation for lung cancer (squamous cell carcinoma of lung, SCCL). SCCL was induced in Wistar rats by intratracheal instillation of 3-methylcholanthrene and treated with three different anti-cancer drugs, Aidi injections, fluorouracil, and a combination of them. After carcinogenesis for 28, 70 and 98 days and therapy for 28 and 56 days, the polyamine levels in plasma of SCCL, healthy and treated rats were determined using a UHPLC-MS/MS assay base on the means of targeted metabolomics. Results showed that increased N-acetylputrescine, cadaverine and 1,3-diaminopropane levels were associated with progression of SCCL. The levels of cadaverine and 1,3-diaminopropane returned to normal after administration of the three different kinds of anticancer drug. In addition, the suitability of using N-acetylputrescine, cadaverine and 1,3-diaminopropane as biomarkers was confirmed by PLS-DA and ROC analysis. It can provide an innovative and effective way for the clinical diagnosis, prevention and treatment of lung cancer, and stimulate a theoretical basis for the design and development of new anticancer drugs. At the same time, this increased the clinical options for polyamines as cancer biomarkers.Polyamines have been widely investigated as potential biomarkers for various types of cancers, including lung cancer, which is one of the most common causes of death from cancer worldwide. This study was carried out to evaluate the value of polyamines that serve as early diagnostic and cancer progression markers as well as drug evaluation for lung cancer (squamous cell carcinoma of lung, SCCL). SCCL was induced in Wistar rats by intratracheal instillation of 3-methylcholanthrene and treated with three different anti-cancer drugs, Aidi injections, fluorouracil, and a combination of them. After carcinogenesis for 28, 70 and 98 days and therapy for 28 and 56 days, the polyamine levels in plasma of SCCL, healthy and treated rats were determined using a UHPLC-MS/MS assay base on the means of targeted metabolomics. Results showed that increased N-acetylputrescine, cadaverine and 1,3-diaminopropane levels were associated with progression of SCCL. The levels of cadaverine and 1,3-diaminopropane returned to normal after administration of the three different kinds of anticancer drug. In addition, the suitability of using N-acetylputrescine, cadaverine and 1,3-diaminopropane as biomarkers was confirmed by PLS-DA and ROC analysis. It can provide an innovative and effective way for the clinical diagnosis, prevention and treatment of lung cancer, and stimulate a theoretical basis for the design and development of new anticancer drugs. At the same time, this increased the clinical options for polyamines as cancer biomarkers.

Quantitation of eleven active compounds of Aidi injection in rat plasma and its application to comparative pharmacokinetic study.

Aidi injection has been widely used for the treatment of colorectal cancer. The purpose of this study was to develop a sensitive and reliable method for simultaneous quantitation of 11 main active ingredients in Aidi injection and to compare the pharmacokinetics of these ingredients in normal and colorectal model cancer rats after tail vein injection. After being extracted by isopropanol-ethyl acetate (1:1, v/v), the plasma samples were analyzed with domperidone as internal standard. Then the analytes were separated on a Venusil MP C18 column with 0.15% formic acid and methanol. The detection was performed on HPLC-MS/MS system with turbo ion spray source in the positive ion and multiple reaction-monitoring mode. The assay was shown to be linear over the range of 0.004-4.0μgmL(-1) of syringin B, astragaloside II and isofraxidin; 0.01-10.0μgmL(-1) of calycosin-7-O-β-d-glucoside and astragaloside IV; 0.02-20.0μgmL(-1) of ginsenoside Rg1, Rb1, Rc and Rd; 0.04-40.0μgmL(-1) of syringin E; 0.06-60.0μgmL(-1) of ginsenoside Re. And the validated method has been successfully applied to compare pharmacokinetic profiles of the 11 ingredients in plasma. The pharmacokinetic results showed here were significant differences in pharmacokinetic parameters for eight analytes between two groups after injection, while no significant differences for astragaloside II, astragaloside IV and ginsenoside Rc. The present study has the advantages of short analysis time and easy sample preparation, which could more comprehensively reflect the quality of Aidi injection in single run. The method proposed could be of great use for pharmacokinetics, bioavailability or bioequivalence studies of Aidi injection in biological samples.

Simultaneous determination of benzoylmesaconine and piperine in rat plasma after oral administration of Naru-3 by an ultra fast liquid chromatography-tandem mass spectrometry method and its application in a comparative pharmacokinetic study

A rapid, sensitive and reliable ultra fast liquid chromatography-tandem mass spectrometry (UFLC-MS/MS) method has been developed for the simultaneous quantitation of benzoylmesaconine and piperine in rat plasma after oral administration of Naru-3. Naru-3 is a well-known traditional Mongolian medical analgesic formula, which contains three Chinese herb powders – Aconiti Kusnezoffii Radix Cocta, Piperis Longi Fructus and Chebulae Fructus. After addition of brucine (internal standard, IS), 100 μL of plasma was extracted by liquid–liquid extraction with methyl tert-butyl ether, and then the two analytes and IS were separated on a Venusil MP C18 column (100 mm × 2.1 mm, 3.0 μm) at 30 °C with a gradient program of 0.1% formic acid–methanol in water as the mobile phase. The UFLC-MS/MS system coupled with an electrospray ionization source was performed in multiple reaction monitoring mode. The linear range was 0.075–15 ng mL−1 for benzoylmesaconine and 5–1000 ng mL−1 for piperine. The linearity, accuracy, precision, recovery and matrix effect of the two analytes were all within satisfaction. The validated method was successfully applied to compare the pharmacokinetic profiles of the analytes in rat plasma after oral administration of three compatibility-composition suspensions designed according to the Naru-3 pill. The pharmacokinetic data obtained by this method indicated that some ingredients in the Naru-3 pill could have an influence on the pharmacokinetic behavior of the two analytes.