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PubMed2005

Determination and pharmacokinetic analysis of salvianolic acid B in rat blood and bile by microdialysis and liquid chromatography.

Salvianolic acid B is an herbal ingredient isolated from Salvia miltiorrhiza. An in vivo microdialysis sampling method coupled to high-performance liquid chromatography has been developed for continuous monitoring of protein-unbound salvianolic acid B in rat blood and bile. Microdialysis probes were inserted into the jugular vein/right atrium and bile duct of Sprague-Dawley rats, and a dose of 100 mg/kg salvianolic acid B was then administered via the femoral vein. Dialysates were collected and directly injected into a liquid chromatographic system. Salvianolic acid B was eluted using a microbore reversed-phase ODS 5 microm (150 mm x 1 mm I.D.) column. Isocratic elution of salvianolic acid B was achieved within 10 min using the liquid chromatographic system. The chromatographic mobile phase consisted of acetonitrile-methanol-20 mM monosodium phosphoric acid (pH 3.5) (10:30:60, v/v/v) containing 0.1 mM 1-octanesulfonic acid with 0.05 ml/min. The wavelength of the UV detector was set at 290 nm. Salvianolic acid B in both blood and bile dialysates was adequately determined using the liquid chromatographic conditions described, although the blank bile pattern was more complex. The retention times of salvianolic acid B in rat blood and bile dialysates were found to be 7.2 min. Peak-areas of salvianolic acid B were linear (r2 > 0.995) over a concentration range of 0.1-50 microg/ml. In vivo recoveries of microdialysis probes of salvianolic acid B in rat blood and bile averaged 22 +/- 2% and 41 +/- 1%, respectively. This study indicates that salvianolic acid B undergoes hepatobiliary excretion.

Animals↗
PubMed2006

Purification of salvianolic acid B from the crude extract of Salvia miltiorrhiza with hydrophilic organic/salt-containing aqueous two-phase system by counter-current chromatography.

Establishment of hydrophilic organic/salt-containing aqueous two-phase system and purification of salvianolic acid B from crude extract of S. miltiorrhiza by counter-current chromatography with said system were studied. Ethanol and n-propanol were selected to constitute biphasic systems with ammonia sulphate, sodium chloride and phosphate separately, and related system characteristics including phase diagrams, phase ratio, separation time were tested. The partition coefficient of crude salvianolic acid B was also tested in above systems and further finely adjusted by altering the constitution of phosphate in n-propanol/phosphate system. Salvianolic acid B was purified to 95.5% purity by counter-current chromatography in 36% (w/w) n-propanol/8% (w/w) phosphate system with the ratio between dipotassium hydrogen phosphate and sodium dihydrogen phosphate of 94:6. One hundred and eight milligrams of salvianolic acid B was purified from 285 mg crude extract with the recovery of 89%.

Benzofurans↗
PubMed2006

Automated post-collection concentration for purified preparative fractions via solid phase extraction.

Recent advancements in preparative HPLC have improved and streamlined compound purification. However, fraction evaporation remains a bottleneck within the process. An alternative to fraction evaporation is to remove the water and reduce the overall volume of the collection by trapping the fraction onto a solid phase extraction (SPE) cartridge. This method (as opposed to analytical applications involving SPE) works by collecting and then diluting the fraction(s), passing the fraction(s) through a SPE, drying the SPE with nitrogen and ultimately eluting the concentrated fraction(s) in a small amount of 100% organic solvent. An appreciable breakthrough is not observed using this method. In addition, recovery from the SPE for the tested compounds rosmarinic acid and carvacrol, two naturally occurring antioxidants in oregano, was found to be 95-98% for a 100mg injection via preparative HPLC purification at 50mg/compound.

Antioxidants↗
PubMed2006

One-step separation and purification of 3,4-dihydroxyphenyllactic acid, salvianolic acid B and protocatechualdehyde from Salvia miltiorrhiza Bunge by high-speed counter-current chromatography.

Three kinds of polyphenols of Salvia miltiorrhiza Bunge, 3,4-dihydroxyphenyllactic acid, salvianolic acid B and protocatechualdehyde, were separated and purified in one step with solvent system n-hexane-ethyl acetate-methanol-acetic acid-water (1:6:1.5:1.5:8) by high-speed counter-current chromatography. Acetic acid was successfully used to increase the partition of high polar target compounds in organic phase to modify partition coefficient value. 3,4-Dihydroxyphenyllactic acid, salvianolic acid B and protocatechualdehyde were purified from 100mg water extracted crude sample of Salvia miltiorrhiza Bunge at purity of 97.6%, 94.2% and 98.2% and at yield of 98.6%, 73.6% and 90.2%. High-speed counter-current chromatography together with organic/aqueous solvent system supplied an efficient method to purify water-soluble compounds directly from crude samples of traditional Chinese medicines.

Benzaldehydes↗
PubMed2006

Activation of the iberiotoxin-sensitive BKCa channels by salvianolic acid B of the porcine coronary artery smooth muscle cells.

In this study, we examined the effects of Salvia miltiorrhiza (Danshen) crude extract, some of its lipid-soluble components (tanshinone I, tanshinone II(A), cryptotanshinone, dihydroisotanshinone I) and the water-soluble compounds (danshensu and salvianolic acid B) on the K(+) channels such as the iberiotoxin-sensitive Ca(2+)-activated K(+) (BK(Ca)) channels and the glibenclamide-sensitive ATP-dependent K(+) (IK(ATP)) channels of the porcine left anterior descending coronary artery smooth muscle cells. Cumulative application of salvianolic acid B (30-300 microM) caused a l-NNA (100 microM)-insensitive, potentiation of the outward BK(Ca) current amplitude with no apparent effect on the IK(ATP) channels opening. Salvianolic acid B (300 microM) caused an ODQ (10 microM, a guanylate cyclase inhibitor)-sensitive enhancement of the outward BK(Ca) current amplitude. In contrast, none of the other isolated chemical constituents of S. miltiorrhiza modified the openings of the two types of K(+) channels studied. In conclusion, our results suggest that salvianolic acid B, a major hydrophilic constituent found in Radix S. miltiorrhiza, activated the opening of the BK(Ca) channels of the porcine coronary artery smooth muscle cells through the activation of guanylate cyclase without the involvement of the nitric oxide synthase activation.

Animals↗
PubMed2006

Salvianolic acid B, an aqueous component of danshen (Salvia miltiorrhiza), relaxes rat coronary artery by inhibition of calcium channels.

In this study, we have investigated the mechanism of action through which salvianolic acid, a constituent of the medicinal herb danshen (Salvia miltiorrhiza), causes relaxation of isolated coronary artery rings precontracted with 1 microM 5-hydroxytryptamine (5-HT). The vasorelaxant effects of salvianolic acid B were also compared with the aqueous extract of danshen. Extraction of the water-soluble fraction from danshen provided a yield of 17.5%. The amount of salvianolic acid B determined in this aqueous extract was 3.9%, and the extract was 6.3 times less potent than salvianolic acid B in relaxing 5-HT-precontracted coronary artery rings; IC(50) values were 930.3+/-133.5 microg/ml and 147.9+/-17.4 microg/ml, respectively. Removal of the endothelium did not significantly affect their vasodilator potencies; IC(50) values were 842.1+/-123.8 mictog/ml and 160.3+/-25.9 microg/ml. On the other hand, a potassium channel inhibitor tetraethylammonium (TEA, 10 mM) shifted their concentration-response curves by 1.7 and 2.9 folds. The possible involvement of Ca(2+) channels was investigated in artery rings incubated with Ca(2+)-free buffer and primed with 1 microM 5-HT or 60 mM KCl for 5 min prior to adding CaCl(2) to elicit contraction. In 5-HT-primed preparations, 2 mg/ml danshen aqueous extract and 0.72 mg/ml salvianolic acid B abolished the CaCl(2)-induced vasoconstriction, whereas, in KCl-primed preparations, 10 mg/ml danshen aqueous extract and 1.44 mg/ml salvianolic acid B produced 90% inhibition. These findings suggest the vasorelaxant effects of danshen aqueous extract and salvianolic acid B were produced by inhibition of Ca(2+) influx in the vascular smooth muscle cells. The opening of K(+) channels had a minor contribution to their effects, but endothelium-dependent mechanisms were not involved. The high vasodilator potency and the quantity of salvianolic acid B contained in danshen aqueous extract suggest it is an important vasodilator constituent in this medicinal herb.

Animals↗
PubMed2004

Rosmarinic acid from Zataria multiflora tops and in vitro cultures.

Rosmarinic acid (RA) was obtained from Zataria multiflora tops' extract and its structure was confirmed by spectroscopic methods. Various in vitro cultures were established on Murashige and Skoog (MS) or Modified Tobacco (MT) medium containing growth hormones. The results indicated that cultures of Z. multiflora biosynthesize RA (55-355 mg/100 g dry wt.) and the highest accumulation were reached on MT media containing NAA 2 mg/l.

Cells, Cultured↗
PubMed2005

Rosmarinic acid content and RAPD analysis of in vitro regenerated basil (Ocimum americanum) plants.

Various in vitro cultures were established from shoot tips of Ocimum americanum seedlings. Rosmarinic acid content of the in vitro produced plants as well as parent plant were determined by HPLC analysis and subjected to RAPD analysis. MS medium with BA at a concentration of 1 mg/l and 0.25 mg/l IAA supports maximum rosmarinic acid production in plants produced from cultures grown on that medium. RAPD analysis revealed 64 scorable bands from four primers, including six polymorphic bands. The band pattern revealed differences between the parent plant and the in vitro regenerated plants. Certain band changes were found in O. americanum plants regenerated in vitro, suggesting the existence of genetic variation that might affect the biochemical synthesis of plants derived from tissue culture.

Cell Culture Techniques↗
PubMed2005

Cytoprotectivity of Prunella vulgaris on doxorubicin-treated rat cardiomyocytes.

The cardioprotective effect of Prunella vulgaris ethylacetate fraction (PVEF) and its constituent rosmarinic acid (RA) was evaluated on isolated rat cardiomyocytes subjected to doxorubicin-induced oxidative stress. The cytoprotectivities of PVEF and RA were concentration-dependent in the range of 0.005 to 0.05 mg/ml and the effect of PVEF correlated with the RA content. Dexrazoxan (DE), used as positive control, was less effective than PVEF or RA.

Animals↗
PubMed2006

Bioavailability of salvianolic acid B in conscious and freely moving rats.

Salvianolic acid B is an herbal ingredient isolated from Salvia miltiorrhiza. The aim of this study was to apply an automated blood sampling system coupled to a simple liquid chromatographic system to determine the bioavailability of salvianolic acid B in stress-free rats. The plasma sample (25 microl) was vortex-mixed with 50 microl of internal standard solution (chloramphenicol 10 microg/ml in acetonitrile) to achieve protein precipitation. Salvianolic acid B in the rat plasma was separated using a reversed-phase C18 column (250 mm x 4.6 mm, 5 microm) with a mobile phase of acetonitrile-methanol-20mM NaH(2)PO(4) (adjusted to pH 3.5 with H(3)PO(4)) (20:10:70 v/v/v) containing 0.1mM 1-octanesulfonic acid, and the flow-rate of 1 ml/min. The UV detection wavelength was 286 nm. The concentration-response relationship from the present method indicated linearity over a concentration range of 0.5-200 microg/ml. Intra- and inter-assay precision and accuracy of salvianolic acid B fell well within the predefined limits of acceptability (<15%). The plasma sample of salvianolic acid B was further identified by LC-MS/MS in the negative ion mode using mass transition m/z 358.2 to the product ion m/z 196.9. After salvianolic acid B (100mg/kg, i.v.; 500 mg/kg, p.o.) was given in conscious and freely moving rats, the AUC were 5030+/-565 and 582+/-222 min microg/ml for intravenous (100 mg/kg) and oral (500 mg/kg) doses, respectively. The oral bioavailability of salvianolic acid B in freely moving rats was calculated to be 2.3%.

Animals↗
PubMed2005

Aqueous extract of Salvia miltiorrhiza attenuates increased endothelial permeability induced by tumor necrosis factor-alpha.

Salvia miltiorrhiza Bunge, a traditional Chinese herbal medicine, is often used for prevention and treatment of cardiovascular disorders such as atherosclerosis. To understand its mechanism of pharmacological action, its effects on endothelial monolayer permeability are studied. The present study demonstrated that extract of S. miltiorrhiza (ESM) and its major ingredients, Danshensu (DSS) and salvianolic acid B (Sal B), inhibited tumor necrosis factor (TNF-alpha) induced endothelial permeability, whereas the other major ingredient, protocatechualdehyde, was ineffective. ESM, DSS and Sal B also repressed expression of vascular endothelial growth factor (VEGF) and extracellular signal-regulated kinase (ERK) activation in TNF-alpha induced HUVEC cells. Furthermore, it was found that ESM attenuated the disorganization of vascular endothelial (VE)-cadherin induced by TNF-alpha. The effect of ESM on TNF-alpha induced endothelial permeability and redistribution of VE-cadherin is likely due to a reduction of VEGF protein expression as a result of modulation of the ERK signaling pathway. Endothelial cell hyperpermeability is implicated in inflammation and subsequent ischemic reperfusion injury and atherosclerosis. Data from this study suggest that one of the mechanisms S. miltiorrhiza exerts its pharmacological effect is through its modulation of endothelial cell permeability.

Benzaldehydes↗
PubMed2006

Complement inhibitors selectively attenuate injury following administration of cobra venom factor to rats.

Systemic activation of complement is a pathophysiological response common to severe disturbances such as hemorrhagic shock, major burn injury and sepsis. Intravenous infusion of cobra venom factor (CVF) has been used as an animal model of acute respiratory distress syndrome (ARDS), and reliably and selectively induces rapid intravascular activation of the complement system, leading to acute organ damage. In the present study, we have used different complement inhibitors to investigate the roles of complement products in CVF-induced responses in the rat. Rats were treated with either a C5a receptor antagonist (C5aRA, AcF-[OP(d-Cha)WR], 1 mg/kg i.v. or 10 mg/kg p.o.), a C3a receptor antagonist (C3aRA, N(2)-[(2,2-diphenylethoxy)acetyl]-l-arginine, 0.1 mg/kg i.v.) or a convertase inhibitor, rosmarinic acid (RMA, 10 mg/kg i.v.), prior to CVF-induced complement challenge. Intravenous CVF resulted in hallmark events evident in the development of ARDS, including systemic neutropenia followed by neutrophil migration to the lung and bronchoalveolar vascular leakage, blood pressure alterations, and an increase in TNFalpha levels in both serum and bronchoalveolar lavage fluid. These hemodynamic changes were differentially inhibited by antagonism of C5a receptors, C3a receptors or by inhibition of the entire complement cascade using RMA. This evidence strongly implicates complement factors in the development of lung injury associated with systemic complement activation and identifies complement inhibition as a potential therapeutic target for acute syndromes such as ARDS and other severe systemic shock states mediated by activation of complement.

Administration, Oral↗
PubMed2005

Simultaneous determination of six phenolic constituents of danshen in human serum using liquid chromatography/tandem mass spectrometry.

The six phenolic constituents are water-soluble components extracted from the Chinese medical herb danshen, the dried roots of Salvia miltiorrhiza Bunge (Labiatae). An liquid chromatography/tandem mass spectrometry (LC/MS/MS)-based method has been developed for the simultaneous quantification of six phenolic constituents of danshen (magnesium lithospermate B (MLB), rosmarinic acid (RA) and lithospermic acid (LA), caffeic acid (CAA), protocatechuic aldehyde (3,4-dihydroxybenzaldehyde, Pal), 3,4-dihydroxyphenyllactic acid (danshensu)) in human serum with chloramphenicol as internal standard. The serum samples were treated by special liquid-liquid extraction, and the analytes were determined using electrospray negative ionization mass spectrometry in the multiple reaction monitoring (MRM) mode, with sufficient sensitivity to allow analysis of human serum samples generated following administration of a clinically relevant dose. Good linearity over the range 8-2048 ng/mL for six phenolic constituents was observed. The intra- and inter-day precisions (CV) of analysis were <13%, and the accuracy ranged from 88 to 116%. This quantitation method was successfully applied to a pharmacokinetic study of i.v. drip infusion of Danshen injection fluid in human.

Benzaldehydes↗
PubMed2005

In vitro protective effects of salvianolic acid B on primary hepatocytes and hepatic stellate cells.

The production of reactive oxygen species (ROS) is believed to be involved in liver injury and hepatic fibrosis. Activation of hepatic stellate cells (HSCs) is a key feature of liver fibrosis. Salvia miltiorrhiza is a traditional Chinese herb used in the treatment of cardiovascular and liver diseases to resolve stasis. The effects of salvianolic acid B (Sal B), a major component of Salvia miltiorrhiza, on oxidative damage include free radical DPPH scavenging, malondialdehyde (MDA) formation and ROS generation in primary rat hepatocytes and HSCs, and on alpha-SMA, and collagen expression in transforming growth factor-beta1 (TGF-beta1)-stimulated HSCs were examined. Results indicated that Sal B scavenged DPPH potently with an IC50 2.2+/-0.2 microg/ml (3.06+/-0.3 microM), inhibited lipid peroxidation and eliminated ROS accumulation in a concentration-dependent manner on primary rat hepatocytes and HSCs. Sal B also reduced alpha-SMA and collagen synthesis and deposition in HSCs, and had no direct cytotoxicity on both hepatocytes and HSCs. Our results suggest that Sal B ameliorated oxidative damage and eliminated ROS accumulation in hepatocytes, and attenuated HSC activation, potentially conferring hepatoprotective and anti-fibrogenic effects.

Actins↗
PubMed2026

Targeting pancreatic cancer progression: The formononetin and salvianolic acid B combination suppresses JAK/STAT signaling via MBOAT2 downregulation.

OBJECTIVE: Formononetin and salvianolic acid B (FcS) are the primary bioactive components of the Astragalus mongholicus-Salvia miltiorrhiza herbal pair, a classic combination for treating pancreatic cancer associated with qi deficiency and blood stasis. This study elucidates the therapeutic potential and mechanisms of FcS in the treatment of pancreatic cancer. METHODS: A zebrafish xenograft model was used to screen bioactive combinations derived from A. mongholicus and S. miltiorrhiza, identifying FcS as a candidate with antitumor activity. Its efficacy was evaluated in vivo using the zebrafish model, orthotopic LSL-KrasG12D/+, LSL-Trp53R172H/+ and Pdx-1-Cre (KPC) mice, and subcutaneous xenograft models. Cell viability and proliferation were assessed using cell counting kit-8, 5-ethynyl-2'-deoxyuridine and colony formation assays, and migration and invasion were evaluated by wound healing and transwell assays. Membrane-bound O-acyltransferase 2 (MBOAT2) was identified as a potential target through a molecular docking study and the Cancer Genome Atlas (TCGA) analysis. MBOAT2 knockdown cells were used to explore its roles and the Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway in FcS-mediated inhibition. RESULTS: In the zebrafish model, FcS strongly inhibited pancreatic tumor growth. FcS reduced tumor volume, the expression of proliferation marker Ki-67, and proliferating cell nuclear antigen in KPC mice. In vitro, FcS inhibited pancreatic cancer cell viability, proliferation, migration and invasion, which was accompanied by downregulation of MBOAT2 expression. TCGA analysis linked high MBOAT2 expression to aggressive phenotypes. MBOAT2 knockdown reduced the survival, proliferation and invasion of BxPC-3 cells. Rescue experiments revealed that MBOAT2 knockdown attenuated the antitumor effects of FcS, possibly through modulation of the JAK/STAT signaling pathway. FcS also inhibited tumor proliferation in xenograft models, and MBOAT2 expression was elevated in tumor tissues from pancreatic cancer patients. CONCLUSION: FcS suppresses pancreatic cancer progression via MBOAT2 downregulation and JAK/STAT pathway inhibition, which highlights MBOAT2 as a potential therapeutic target. Please cite this article as: Xu Y, Xu CS, Jin HB, Gu WG, Shen HZ, Lu L, Chen Y, Xu DC, Zhang XF, Yang JF, Wang Y. Targeting pancreatic cancer progression: The formononetin and salvianolic acid B combination suppresses JAK/STAT signaling via MBOAT2 downregulation. J Integr Med. 2026; 24(5):725-741.

Animals↗
PubMed2005

Determination of active components in rosemary by capillary electrophoresis with electrochemical detection.

Rosemary (Rosmarinus officinalis L.) is a spice and medicinal herb widely used around the world. Among natural antioxidants, rosemary has been widely accepted as one of the spices with the highest antioxidant activity. A capillary electrophoresis method for the determination of its active components using electrochemical detection was developed. Effects of several important factors were investigated to acquire the optimum conditions. The detection electrode was a 300 microm carbon disc electrode at a working potential of +0.90 V (versus SCE). The analytes can be well separated with 25 min in a 75 cm length fused-silica capillary at a separation voltage of 16 kV in an 80 mmol/l borate buffer (pH 9.0). The current response was linear over about three orders of magnitude with detection limits (S/N=3) ranging from 2 x 10(-7) to 1 x 10(-6)g/ml for all the analytes. The method was successfully used in the analysis of rosemary with relatively simple extraction procedures, and the assay results were satisfactory.

Antioxidants↗
PubMed2006

Determination of phenolic compounds in fennel by HPLC and HPLC-MS using a monolithic reversed-phase column.

A reversed-phase high-performance liquid chromatography (HPLC) method for analyzing phenolic compounds in fennel (Foeniculum vulgare) has been developed. The use of a monolithic column with short dimensions in combination with optimized chromatographic conditions allows over 100 samples per day to be analyzed. Chromatographic parameters such as column temperature and injection volume, were found to be crucial in obtaining adequate selectivity and resolution, consequently allowing short run times. The method was validated for the major phenolic compounds present in fennel plant material: 3-O-caffeoylquinic acid (3-CQA), chlorogenic acid, 4-O-caffeoylquinic acid (4-CQA), eriocitrin, rutin, miquelianin, 1,3-O-dicaffeoylquinic acid (1,3-diCQA), 1,5-O-dicaffeoylquinic acid (1,5-diCQA), 1,4-O-dicaffeoylquinic acid (1,4-diCQA) and rosmarinic acid. The limits of detection (LOD) and the limits of quantitation (LOQ) ranged from 0.05 to 1.0 microg/mL and from 0.15 to 2.5 microg/mL, respectively. With some adaptation, the extraction procedure could be even less invasive, which is useful in screening work.

Chlorogenic Acid↗
PubMed2006

Hydrolytic kinetics of lithospermic acid B extracted from roots of Salvia miltiorrhiza.

The hydrolytic kinetics of lithospermic acid B (LAB) extracted from the roots of Salvia miltiorrhiza (Chinese herb: danshen) was investigated by using reversed-phase high-performance liquid chromatography (HPLC) with UV-vis detection. The influences of initial drug concentration, pH and temperature on hydrolysis of LAB were studied in aqueous solutions. The results showed that initial concentration of LAB has no effect on the degradation rate at pH 2.0. The hydrolysis followed pseudo-first-order kinetics at 90 degrees C. The log k(obs)-pH profile indicated that the optimal stability range was at pH 2.0-5.0. The rate constant of overall hydrolysis as a function of temperature under the given conditions obeyed the Arrhenius equation. Analysis of the acid-induced degraded solution of LAB by liquid chromatography-mass spectrometry (LC-MS) revealed at least four degradation products [M-H](-) ion at m/z 197, 137, 537 and 537, respectively. Three of these degradation products, i.e. danshensu (DSU), protocatechuic aldehyde (PRO), and lithospermic acid, were further identified by comparing the retention times with standard samples. According to the structure of LAB and its hydrolysis behavior in solution, the other product was proposed to be the isomer of lithospermic acid.

Benzaldehydes↗