Ultra-performance liquid chromatography-tandem mass spectrometry determination and pharmacokinetic studies of five phenolic acids in rat plasma after intravenous administration of salvianolic acid for injection.

A simple, efficient, and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for simultaneous determination and pharmacokinetic study of salvianolic acid D, rosmarinic acid, lithospermicic acid, salvianolic acid B, and salvianolic acid Y in rat plasma after intravenous administration of salvianolic acid for injection. Three doses of administration, containing 10, 25, and 62.5 mg/kg, were investigated. Plasma samples were pretreated using protein precipitation with pre-cooled acetonitrile. As shown in S1, Chromatographic separation was achieved on a Waters Acquity UPLC® BEH C18 column (1.7 µm, 2.1 × 100 mm) with a mobile phase composed of acetonitrile-methanol-0.5% aqueous formic acid (10:30:60, v/v/v) at a flow rate of 0.3 ml/min. MS was detected by electrospray ion source negative ion mode and multiple reaction monitoring mode. The method was fully validated. The calibration curves for the five phenolic acids were linear in the given concentration ranges. The extraction recoveries, matrix effects, intra-day and inter-day precisions, and accuracies of the five analytes were all within acceptable limits. No significant difference of elimination half-life time (T1/2 ) of five analytes at three doses was observed. Area under the curve and peak concentration (Cmax ) of the five analytes demonstrated a linear increase in the doses with the linear correlation r of each analyte at three doses being greater than 0.915. It indicated that the pharmacokinetic behavior of is positively related to the dose in the range of 10-62.5 mg/kg.

Simultaneous determination and tissue distribution studies of four phenolic acids in rat tissue by UFLC-MS/MS after intravenous administration of salvianolic acid for injection.

A rapid, simple and sensitive ultra-fast liquid chromatography tandem mass spectrometric method was developed and validated for simultaneous determination and tissue distribution studies of rosmarinic acid, salvianolic acid D, lithospermic acid and salvianolic acid B in rats after intravenous administration of salvianolic acid for injection. The tissue homogenate samples were pretreated by protein precipitation with pre-cooled acetonitrile. Chromatographic separation was achieved on a Waters Cortecs UPLC C18 column (1.6 µm, 2.1 × 100 mm) with a mobile phase composed of 0.1% formic acid-water and 0.1% formic acid-acetonitrile. Analytes were detected by electrospray ionization mass spectrometry and quantitated using multiple reaction monitoring. The method was fully validated. The calibration curves for the four phenolic acids were linear in the given concentration ranges. The precisions (relative standard deviation) in the measurement of quality control samples were <10% and the accuracies (relative error) were in the range of 0.28-11.22%. The reliable method was successfully applied to the tissue distribution studies of the four phenolic acids. The results showed that rosmarinic acid, salvianolic acid D, lithospermic acid and salvianolic acid B were rapidly distributed in tissues with the major amount found in kidney, and little crossed the blood-brain barrier. The developed method and the results provide a basis for further studies.

Characterization of metabolites in rats after intravenous administration of salvianolic acid for injection by ultra-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry.

It is an essential requirement to clarify the metabolites of traditional Chinese medicine (TCM) injections, which contain numerous ingredients, to assess their safe and effective use in clinic. Salvianolic acid for injection (SAFI), made from hydrophilic phenolic acids in Salvia miltiorrhiza Bunge, has been widely used for the treatment of cerebrovascular diseases, but information on its metabolites in vivo is still lacking. In the present study, we aimed to holistically characterize the metabolites of the main active ingredients in rat plasma, bile, urine and feces following intravenous administration of SAFI. An ultra-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UPLC/Q-TOF-MS) method was developed. Combining information on retention behaviors, multistage mass spectra and literature data, a total of eight prototypes and 52 metabolites were tentatively characterized. Metabolites originated from rosmarinic acid and salvianolic acid B comprised the majority of identified compounds. Meanwhile, four metabolites derived from salvianolic acid D and five from salvianolic acid B are reported for the first time. This study revealed that methylation, sulfation and glucuronidation were the major metabolic pathways of phenolic acids in SAFI in vivo. Furthermore, the developed UPLC/Q-TOF-MS method could also benefit the metabolic investigation of extracts and preparations in TCM with hydrophilic ingredients. Copyright © 2016 John Wiley & Sons, Ltd.

[Construction of NIRS-based process analytical system for production of salvianolic acid for injection and relative discussion].

Currently, near infrared spectroscopy (NIRS) has been considered as an efficient tool for achieving process analytical technology(PAT) in the manufacture of traditional Chinese medicine (TCM) products. In this article, the NIRS based process analytical system for the production of salvianolic acid for injection was introduced. The design of the process analytical system was described in detail, including the selection of monitored processes and testing mode, and potential risks that should be avoided. Moreover, the development of relative technologies was also presented, which contained the establishment of the monitoring methods for the elution of polyamide resin and macroporous resin chromatography processes, as well as the rapid analysis method for finished products. Based on author's experience of research and work, several issues in the application of NIRS to the process monitoring and control in TCM production were then raised, and some potential solutions were also discussed. The issues include building the technical team for process analytical system, the design of the process analytical system in the manufacture of TCM products, standardization of the NIRS-based analytical methods, and improving the management of process analytical system. Finally, the prospect for the application of NIRS in the TCM industry was put forward.

Exploration of the Mechanism of Salvianolic Acid for Injection Against Ischemic Stroke: A Research Based on Computational Prediction and Experimental Validation.

Ischemic stroke (IS) is an acute neurological injury that occurs when a vessel supplying blood to the brain is obstructed, which is a leading cause of death and disability. Salvia miltiorrhiza has been used in the treatment of cardiovascular and cerebrovascular diseases for over thousands of years due to its effect activating blood circulation and dissipating blood stasis. However, the herbal preparation is chemically complex and the diversity of potential targets makes difficult to determine its mechanism of action. To gain insight into its mechanism of action, we analyzed "Salvianolic acid for injection" (SAFI), a traditional Chinese herbal medicine with anti-IS effects, using computational systems pharmacology. The potential targets of SAFI, obtained from literature mining and database searches, were compared with IS-associated genes, giving 38 common genes that were related with pathways involved in inflammatory response. This suggests that SAFI might function as an anti-inflammatory agent. Two genes associated with inflammation (PTGS1 and PTGS2), which were inhibited by SAFI, were preliminarily validated in vitro. The results showed that SAFI inhibited PTGS1 and PTGS2 activity in a dose-dependent manner and inhibited the production of prostaglandin E2 induced by lipopolysaccharide in RAW264.7 macrophages and BV-2 microglia. This approach reveals the possible pharmacological mechanism of SAFI acting on IS, and also provides a feasible way to elucidate the mechanism of traditional Chinese medicine (TCM).

Simultaneous determination and pharmacokinetic study of four phenolic acids in rat plasma using UFLC-MS/MS after intravenous administration of salvianolic acid for injection.

A simple, sensitive and selective ultra-fast liquid chromatography-tandem mass spectrometry (UFLC-MS/MS) method was established for simultaneous determination and pharmacokinetic study of rosmarinic acid (RA), salvianolic acid D (Sal D), lithospermic acid (LA) and salvianolic acid B (Sal B) in rat plasma after intravenous administration of salvianolic acid for injection (SAFI). Three doses of administration, containing 14, 28 and 56mg/kg, were investigated in this study. Plasma samples were pretreated using protein precipitation (PP) with pre-cooled acetonitrile. Chromatographic separation was achieved on a CORTECS™ UPLC C18 column (1.6µm, 2.1×100mm) with a mobile phase composed of 0.1% formic acid aqueous (V/V) and 0.1% formic acid acetonitrile (V/V). Analytes were detected using electrospray ionization (ESI) source in negative ionization mode and quantified in multiple reaction monitoring (MRM) mode. The validated method is stable and reliable. No significant difference of half lives (t1/2) of four analytes at three doses was observed. Area under the curve (AUC0-∞) and peak concentration (Cmax) of the four analytes demonstrated a linear increase in across the doses with the linear correlation r of each analyte at three doses were greater than 0.95. It indicated that the pharmacokinetic behavior of SAFI is positively related to dose at the range of 14-56mg/kg.