Simultaneous Determination and Pharmacokinetics Study of Three Triterpenes from Sanguisorba officinalis L. in Rats by UHPLC-MS/MS.

A selective and rapid ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was established and validated for the determination of ziyuglycoside I, 3ß,19α-dihydroxyurs-12-en-28-oic-acid 28-ß-d-glucopyranosyl ester, and pomolic acid in rats after the oral administration of ziyuglycoside I, 3ß,19α-dihydroxyurs-12-en-28-oic-acid 28-ß-d-glucopyranosyl ester, pomolic acid, and Sanguisorba officinalis L. extract. The separation was carried out on an ACQUITY UPLC®HSS T3 column (2.1 mm × 100 mm, 1.8 µm), using methanol and 5 mmol/L ammonium acetate water as the mobile phase. The three compounds were quantified using the multiple reaction monitoring mode with the electrospray ion source in both the positive and negative mode. Liquid-liquid extraction was applied to the plasma sample preparation. Bifendate was selected as the internal standard. The intra-day and inter-day precision and the accuracy of the method were all within receivable ranges. The lower limit of quantification of ziyuglycoside I, 3ß,19α-dihydroxyurs-12-en-28-oic-acid 28-ß-d-glucopyranosyl ester, and pomolic acid were 6.50, 5.75, and 2.63 ng/mL, respectively. The extraction recoveries of analytes in rat plasma ranged from 83 to 94%. The three components could be rapidly absorbed into the blood (Tmax, 1.4-1.6 h) both in the single-administration group or S. officinalis extract group, but the first peak of PA occurred at 0.5 h and the second peak at 4-5 h in the S. officinalis extract. Three compounds were eliminated relatively slowly (t1/2, 7.3-11 h). The research was to establish a rapid, sensible, and sensitive UHPLC-MS/MS method using the multi-ion mode for multi-channel simultaneous mensuration pharmacokinetics parameters of three compounds in rats after oral administration of S. officinalis extract. This study found, for the first time, differences in the pharmacokinetic parameters of the three compounds in the monomer compounds and S. officinalis extract administration, which preliminarily revealed the transformation and metabolism of the three compounds in vivo.

Standardized extract of Sanguisorba minor attenuates injury in aging rat model via the Nrf2/HO­1 pathway.

Aging promotes damage to vulnerable organs like brain and liver. Sanguisorba minor has been traditionally used to cure various ailments. Few studies have reported pharmacological activities of this medicinal plant. This research aimed to investigate the effects of Sanguisorba minor extract (SME) on brain and liver injury in aging rats and identify the underlying mechanisms. The aging model was developed by subcutaneously injecting D­galactose and simultaneously treating them with SME. After biochemical and pathological assessments, mRNA expression levels of nuclear factor­erythroid factor 2­related factor 2 (Nrf2) and Nrf2­ regulated gene, heme oxygenase­1 (HO­1), in the brain and liver tissues were determined. As a result, malondialdehyde and acetylcholinesterase levels were elevated while total thiol content and superoxide dismutase were reduced in the aging rats. Treatment with the extract remarkably attenuated oxidative injury and pathological changes in liver and brain tissues. Concomitantly, the extract up­regulated Nrf2 and HO­1 genes. Our findings exhibited SME may improve the aging­related brain and liver damage through the Nrf2­HO­1 pathway.

Sanguisorbae Radix Suppresses Colorectal Tumor Growth Through PD-1/PD-L1 Blockade and Synergistic Effect With Pembrolizumab in a Humanized PD-L1-Expressing Colorectal Cancer Mouse Model.

Immune checkpoints such as programmed death-1 (PD-1) have been proven as antitumor targets by enhancing cytotoxic T cell activity. All immune checkpoint blockades are antibody therapeutics that have large size and high affinity, as well as known immune-related side effects and low responses. To overcome the limitation of antibody therapeutics, we have explored PD-1/PD-L1 (programmed death-ligand 1) blockades in traditional oriental medicine, which has a long history but has not yet studied PD-1/PD-L1 blockades. Sanguisorbae Radix extract (SRE) blocked PD-1 and PD-L1 binding in competitive ELISA. SRE effectively inhibited the PD-1/PD-L1 interaction, thereby improving T cell receptor (TCR) signaling and the NFAT-mediated luciferase activity of T cells. SRE treatment reduced tumor growth in the humanized PD-L1 MC38 cell allograft humanized PD-1 mouse model. Additionally, the combination of SRE and pembrolizumab (anti-PD-1 antibody) suppressed tumor growth and increased infiltrated cytotoxic T cells to a greater extent did either agent alone. This study showed that SRE alone has anticancer effects via PD-1/PD-L1 blockade and that the combination therapy of SRE and pembrolizumab has enhanced immuno-oncologic effects.

Determination of Triterpenoids and Phenolic Acids from Sanguisorba officinalis L. by HPLC-ELSD and Its Application.

A novel analytical method involving high-performance liquid chromatography with evaporative light scattering detection (HPLC-ELSD) was developed for simultaneous determination of 11 phenolic acids and 12 triterpenes in Sanguisorba officinalis L. Chromatographic separation was conducted with gradient elution mode by using a DiamonsilTM C18 column (250 mm × 4.6 mm, 5 µm) with the mobile phase of 0.1% acetic acid water (A) and methanol (B). The drift tube temperature of ELSD was set at 70 °C and the nitrogen cumulative flow rate was 1.6 L/min. The method was fully validated to be linear over a wide concentration range (R2 ≥ 0.9991). The precisions (RSD) were less than 3.0% and the recoveries were between 97.7% and 101.4% for all compounds. The results indicated that this method is accurate and effective for the determination of 23 functional components in Sanguisorba officinalis L. and could also be successfully applied to study the influence of processing method on those functional components in Sanguisorba officinalis L.

A Bioactive Compound from Sanguisorba officinalis L. Inhibits Cell Proliferation and Induces Cell Death in 5-Fluorouracil-Sensitive/Resistant Colorectal Cancer Cells.

Colorectal cancer (CRC) is one of the most common cancer in the world. The first line chemotherapeutic agent, 5-fluorouracil (5-FU), plays a predominant role in the clinical treatment of CRC. However, with the wide use of 5-FU, more and more CRC patients have been obtaining drug resistance to 5-FU, which leads to a large amount of treatment failures. One of the effective strategies to overcome this obstacle is to find bioactive natural products from traditional medicine. In our previous work, Sanguisorba officinalis L. was found to exert a strong anti-proliferative activity against 5-FU-senstive/resistant CRC cells. Therefore, several compounds were isolated from this herb and screened for their anti-CRC effects to find promising compounds. Among them, a triterpenoid compound named 3ß-[(α-l-arabinopyranosyl) oxy]-urs-12,18(19)-dien-28-oic acid ß-d-glucopyranosyl ester (AGE), showed strong activity against both 5-FU-senstive and resistant CRC cells. In order to further study the mechanism of AGE on CRC cells, flow cytometer analysis, mitochondrial membrane potential (MMP) measurement, Western blotting, and RT-PCR assays were performed. Results demonstrated that AGE induced cell death by apoptosis pathway and autophagy, and inhibited cell proliferation via cell cycle arrest in G0-G1 phase mediated by Wnt signaling pathway. Therefore, AGE may be a potential bioactive compound for CRC treatment in clinic.

Ziyuglycoside II inhibits the growth of digestive system cancer cells through multiple mechanisms.

Digestive system cancers, including liver, gastric, colon, esophageal and pancreatic cancers, are the leading cause of cancers with high morbidity and mortality, and the question of their clinical treatment is still open. Previous studies have indicated that Ziyuglycoside II (ZYG II), the major bioactive ingredient extract from Sanguisorba officinalis L., significantly inhibits the growth of various cancer cells. However, the selective anti-tumor effects of ZYG II against digestive system cancers are not systemically investigated. In this study, we reported the anti-cancer effect of ZYG II on esophageal cancer cells (OE21), cholangiocarcinoma cells (HuCCT1), gastric cancer cells (BGC-823), liver cancer cells (HepG2), human colonic cancer cells (HCT116), and pancreatic cancer cells (PANC-1). We also found that ZYG II induced cell cycle arrest, oxidative stress and mitochondrial apoptosis. Network pharmacology analysis suggested that UBC, EGFR and IKBKG are predicted targets of ZYG II. EGFR signaling was suggested as the critical pathway underlying the anti-cancer effects of ZYG II and both docking simulation and western blot analysis demonstrated that ZYG II was a potential EGFR inhibitor. Furthermore, our results showed synergistic inhibitory effects of ZYG II and chemotherapy 5-FU on the growth of cancer cells. In summary, ZYG II are effective anti-tumor agents against digestive cancers. Further systemic evaluation of the anti-cancer activities in vitro and in vivo and characterization of underlying mechanism will promote the development of novel supplementary therapeutic strategies based on ZYG II for the treatment of digestive system cancers.

The Oral Administration of Sanguisorba officinalis Extract Improves Physical Performance through LDHA Modulation.

Muscle fatigue is induced by an acute or chronic physical performance inability after excessive physical activity often associated with lactate accumulation, the end-product of glycolysis. In this study, the water-extracted roots of Sanguisorba officinalis L., a herbal medicine traditionally used for inflammation and diarrhea, reduced the activities of lactate dehydrogenase A (LDHA) in in vitro enzyme assay myoblast C2C12 cells and murine muscle tissue. Physical performance measured by a treadmill test was improved in the S. officinalis-administrated group. The analysis of mouse serum and tissues showed significant changes in lactate levels. Among the proteins related to energy metabolism-related physical performance, phosphorylated-AMP-activated protein kinase alpha (AMPKα) and peroxisome proliferator-activated receptor-coactivator-1 alpha (PGC-1α) levels were enhanced, whereas the amount of LDHA was suppressed. Therefore, S. officinalis might be a candidate for improving physical performance via inhibiting LDHA and glycolysis.

Whole transcriptome sequencing and integrated network analysis elucidates the effects of 3,8-Di-O-methylellagic acid 2-O-glucoside derived from Sanguisorba offcinalis L., a novel differentiation inducer on erythroleukemia cells.

Acute erythroid leukemia (AEL) is a rare and aggressive hematologic malignancy with no specific treatment. Sanguisorba officinalis L. (S. officinalis), a well-known traditional Chinese medicine, possesses potent anticancer activity. However, the active components of S. officinalis against AEL and the associated molecular mechanisms remain unknown. In this study, we predicted the anti-AML effect of S. officinalis based on network pharmacology. Through the identification of active components of S. officinalis, we found that 3,8-Di-O-methylellagic acid 2-O-glucoside (DMAG) not only significantly inhibited the proliferation of erythroleukemic cell line HEL, but also induced their differentiation to megakaryocytes. Furthermore, we demonstrated that DMAG could prolong the survival of AEL mice model. Whole-transcriptome sequencing was performed to elucidate the underlying molecular mechanisms associated with anti-AEL effect of DMAG. The results showed that the total of 68 miRNAs, 595 lncRNAs, 4030 mRNAs and 35 circRNAs were significantly differentially expressed during DMAG induced proliferation inhibition and differentiation of HEL cells. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed that the differentially expressed miRNAs, lncRNAs, mRNAs and circRNAs were mainly involved in metabolic, HIF-1, MAPK, Notch pathway and apoptosis. The co-expression networks showed that miR-23a-5p, miR-92a-1-5p, miR-146b and miR-760 regulatory networks were crucial for megakaryocyte differentiation induced by DMAG. In conclusion, our results suggest that DMAG, derived from S. officinalis might be a potent differentiation inducer of AEL cells and provide important information on the underlying mechanisms associated with its anti-AEL activity.

A new triterpenoid and other constituents with cytotoxic activity from the roots of Sanguisorba officinalis L.

Sanguisorba officinalis L. is a traditional herbal plant that belongs to the genus Sanguisorba and the family Rosaceae. A new ursane-type triterpenoid, 3-oxo-urs-11, 13(18)-dien-19, 28-olide (1), two known ursane-type triterpenoids (3 - 4) and three known oleanane-type triterpenoids (2, 5 - 6) were isolated from the roots of S. officinalis by silica gel column and MPLC. Their structures were identified by interpretation of spectroscopic data (1 D NMR, 2 D NMR, HR-ESI-MS) and comparison with those reported in the literature. Compound 2 was isolated from the Rosaceae family, compounds 3-5 were obtained from the genus Sanguisorba, and compound 6 was obtained from the S. officinalis for the first time. Additionally, all of the isolated compounds were evaluated for their cytotoxic activity against three human cancer cells. Compound 3 showed better cytotoxic activity against A549, HeLa, SK-Hep1 cells than the other compounds with IC50 values of 48.58 ± 1.88, 47.84 ± 2.01, 42.31 ± 2.43 µM, respectively.

Antibacterial activity and synergy of antibiotics with sanguisorbigenin isolated from Sanguisorba officinalis L. against methicillin-resistant Staphylococcus aureus.

The present study evaluated the antibacterial activity and the synergy of the sanguisorbigenin (SGB) from the dried root of Sanguisorba officinalis L. combined with ß-lactam antibiotics against methicillin-resistant Staphylococcus aureus. A total of six strains of reference strain and clinical isolates were used to determine the antibacterial activity using a broth microdilution assay, and the synergistic effects were determined using a checkerboard assay. To analyse the mechanism of synergy, we conducted the level of penicillin-binding protein 2a by western blot. In addition, quantitative RT-PCR was performed to analyse the mecA gene expression. The minimal inhibitory concentration values of SGB against six strains of S. aureus were in the range of 12·5-50 µg ml-1 , and there were synergy, or partial synergy effects when SGB was combined with antibiotics. Furthermore, when treated with SGB, the level of penicillin-binding protein 2a and the expression of the mecA gene was reduced significantly. In conclusion, this study demonstrated that SGB is a potential natural antibacterial agent against methicillin-resistant S. aureus that represents a considerable burden on the healthcare system worldwide, and may an exceptionally modulator of ß-lactam antibiotics.

Endotoxin Contamination and Reaction Interfering Substances in the Plant Extract Library.

Endotoxin is an unintentional contaminant that has numerous activities and can affect various biological experiments using cells. In this study, we measured the endotoxin activity of samples from a plant extract library (PEL) and determined their degrees of contamination. Endotoxin was detected in approx. 48% (n = 139) and approx. 4% (n = 5) of field-collected and crude drug samples, respectively, and in concentrations >5.0 EU/mL in some samples. The concentrations of endotoxin that affect cells in vitro vary depending on the target cell type. Although the degree of contamination varied in the present study, it was considered to have little effect on the cell experiments. More than 150 PEL samples had problems with reaction courses or recovery rates of Limulus amoebocyte lysate (LAL) tests. In the LAL tests, using three plant extracts [Sanguisorba officinalis L. (Rosaceae), Oenothera biennis L. (Onagraceae), and Lythrum salicaria L. (Lythraceae)], the polyphenolic compounds in the plant extracts affected LAL test and their effects differed depending on the plant species. When the 16 single polyphenol compounds were added to the LAL tests, the compounds with caffeoyl and pyrogallol moieties were found to affect the LAL reaction course and recovery rate. Furthermore, none of the compounds had any effects at concentrations of 1 µM. Because the plant extracts contained analogs of various polyphenolic compounds, they were presumed to actually act synergistically. Our findings demonstrated that attention must be paid to the recovery rate and reaction process of LAL tests with samples containing polyphenolic compounds.

Comparison of the pharmacokinetic profiles of 13 phenolic acids and 6 triterpenes in normal and leukopenia rats after oral administration of Sanguisorba officinalis L. extract by LC-MS/MS.

A selective, accurate, and efficient liquid chromatography-tandem mass spectrometry method was developed for the simultaneous determination of 13 phenolic acids. Additionally, for more comprehensively determining the chemical constituents in Sanguisorba officinalis L. extract, a previously developed method was employed for the simultaneous determination of six triterpenes. Thus, two methods were used to ensure the comprehensiveness and reliability of this study. Based on these methods, the pharmacokinetic profiles of the 13 phenolic acids and 6 triterpenes in normal and leukopenia rats after oral administration of S. officinalis L. extract were compared for the first time in the present study. Quantitative detection of the 13 phenolic acids and 6 triterpenes was performed using the multiple reaction monitoring mode with the electrospray ion source in negative and positive electrospray ionization, respectively. Chromatographic separation was performed on an Agilent Eclipse Plus C18 RRHD column (50 × 2.1 mm, 1.8 µm) using gradient elution with a mobile phase composed of methanol-0.1% aqueous formic acid. The pharmacokinetic results demonstrated that the pharmacokinetic characteristics of the 19 analytes in leukopenia rats differed significantly from those determined in normal rats, which could provide a helpful reference for the clinical application of S. officinalis L. in the prevention and treatment of leucopenia.

Changes in cell membrane properties and phospholipid fatty acids of bacillus subtilis induced by polyphenolic extract of Sanguisorba officinalis L.

Sanguisorba officinalis L. (family Rosaceae, subfamily Rosoideae) is a plant found throughout Southern Europe, Northern Africa, and Eastern Asia. This study demonstrated the antibacterial activity of a purified polyphenolic extract (PPE) from S. officinalis L. against Bacillus subtilis using growth inhibitory and apoptosis assays, and investigated the antibacterial mechanism responsible for changes in cell membrane properties. Fourier transform infrared spectroscopy suggested that PPE altered the cell wall and membrane properties of B. subtilis. Further determination of cell membrane integrity and permeability revealed that B. subtilis membrane integrity was more severely damaged by PPE at the minimum inhibitory concentration (MIC) than at the minimum bactericidal concentrati on (MBC). Instead, PPE at the MBC reduced cell membrane fluidity by significantly decreasing the proportion of anteiso- and iso-branched phospholipid fatty acids (PLFAs) from 64.17 ± 0.28% and 27.23 ± 0.03% in the control to 5.57 ± 1.06% and 6.00 ± 1.40%, respectively (P < 0.001). Scanning electron microscopy revealed different effects of PPE on cell morphology, demonstrating that, at the MIC and MBC, PPE exerted antibacterial activity by disrupting the cell membrane and reducing cell membrane fluidity, respectively. Consequently, this study elucidated changes in the bacterial membrane due to exposure to PPE and its potential use as an antimicrobial agent. PRACTICAL APPLICATION: The abuse of synthetic chemical preservatives raises food safety concerns; however, plant-derived polyphenolic compounds may be a safe and effective alternative. This study demonstrated the strong antibacterial activity of a purified polyphenolic extract (PPE) of Sanguisorba officinalis L. and revealed its antibacterial mechanism against Bacillus subtilis, suggesting that it may provide a useful antimicrobial agent in food industry applications.

Sanguisorba officinalis L. derived from herbal medicine prevents intestinal inflammation by inducing autophagy in macrophages.

Disturbed activation of autophagy is implicated in the pathogenesis of inflammatory bowel disease. Accordingly, several autophagy-related genes have been identified as Crohn's disease susceptibility genes. We screened the autophagy activators from a library including 3,922 natural extracts using a high-throughput assay system. The extracts identified as autophagy activators were administered to mice with 2% dextran sodium sulfate (DSS). Among the autophagy inducers, Sanguisorba officinalis L. (SO) suppressed DSS-induced colitis. To identify the mechanism by which SO ameliorates colitis, epithelial cell and innate myeloid cells-specific Atg7-deficient mice (Villin-cre; Atg7f/f and LysM-cre; Atg7f/f mice, respectively) were analyzed. SO-mediated inhibition of colitis was observed in Villin-cre; Atg7f/f mice. However, SO and a mixture of its components including catechin acid, ellagic acid, gallic acid, and ziyuglycoside II (Mix4) did not suppressed colitis in LysM-cre; Atg7f/f mice. In large intestinal macrophages (Mφ) of Atg7f/f mice, SO and Mix4 upregulated the expression of marker genes of anti-inflammatory Mφ including Arg1, Cd206, and Relma. However, these alterations were not induced in LysM-cre; Atg7f/f mice. These findings indicate that SO and its active components ameliorate DSS-induced colitis by providing intestinal Mφ with anti-inflammatory profiles via promotion of Atg7-dependent autophagy.

Bioactive properties of Sanguisorba minor L. cultivated in central Greece under different fertilization regimes.

In this study, the chemical characterization and bioactive properties of S. minor cultivated under different fertilization rates (control, half rate and full rate) were evaluated. Twenty-two phenolic compounds were identified, including five phenolic acids, seven flavonoids and ten tannins. Hydrolysable tannins were prevalent, namely Sanguiin H-10, especially in leaves without fertilization (control). Roots of full-rate fertilizer (660 Kg/ha) presented the highest flavonoid content, mainly catechin and its isomers, whereas half-rate fertilizer (330 Kg/ha), presented the highest content of total phenolic compounds, due to the higher amount of ellagitannins (lambertianin C: 84 ± 1 mg/g of dry extract). Antimicrobial activities were also promising, especially against Salmonella typhimurium (MBC = 0.44 mg/mL). Moreover, root samples revealed activity against all tested cell lines regardless of fertilization rate, whereas leaves were effective only against HeLa cell line. In conclusion, S. minor could be a source of natural bioactive compounds, while fertilization could increase phenolic compounds content.

Positive effects of platelet-rich plasma (PRP) and a Sanguisorba officinalis polysaccharide on the proliferation and differentiation of anterior cruciate ligament (ACL) fibroblasts in vitro.

Context: Sanguisorba officinalis L. (Rosaceae), a famous traditional Chinese medicine. It was recently reported that its polysaccharide could facilitate collagen production.Objectives: We investigated the mechanism by which S. officinalis polysaccharide (SOWPa) and/or platelet-rich plasma (PRP) promote regenerative potential of anterior cruciate ligament (ACL) in vitro.Materials and methods: ACL fibroblasts were treated with SOWPa (25 and 100 mg/kg), PRP, PRP + SOWPa (25 and 100 mg/kg) or vehicle alone for 24, 48, or 72 h. Cell viability, migration ability and apoptosis were evaluated by MTT, transwell and flow cytometry, respectively. Western blot analysis was performed to assess associated protein expression.Results: PRP, SOWPa (100 mg/kg) or PRP + SOWPa (100 mg/kg) treatment for 72 h significantly improved the cell viability of ACL fibroblasts from 100 ± 7.5% (control) to 156.85 ± 12.82%, 188.08 ± 15.92%, and 223.67 ± 18.82%, respectively, which was evidenced by individual decreased apoptosis rate from 31.26 ± 2.35% (control) to 20.80 ± 1.89%, 18.01 ± 1.55% and 9.33 ± 0.78%. Furthermore, the motility of ACL fibroblasts was significantly improved with increased migrated cell number per field from 5 for control to 26 for PRP, 36 for SOWPa and 44 for PRP + SOWPa, respectively. Moreover, the protein expression of differentiation markers (RUNX2, ALP, BMP2 and Col I) and TLR-4 and phosphorylated p65 (p-p65) was inhibited by the above treatment.Discussion and conclusions: Data suggested that the addition of SOWPa to PRP increased the regenerative ability of ACL fibroblasts by blocking the TLR-4/NF-κB pathway.

Characterization of Secondary Metabolites in Flowers of Sanguisorba officinalis L. by HPLC-DAD-MSn and GC/MS.

The investigations reported here focus on an in-depth characterization of the secondary metabolite profile of Sanguisorba officinalis flowers. For this purpose, fresh flowers were extracted with MeOH/H2 O and EtOH/H2 O and the resulting crude extracts fractionated using CH2 Cl2 , AcOEt, and BuOH. Individual compounds were characterized by high performance liquid chromatography and gas chromatography coupled with mass spectrometric detection (HPLC-DAD-MSn and GC/MS). MeOH/H2 O extraction and LC/MSn investigations revealed the occurrence of flavonoid glycosides (quercetin, kaempferol), ellagitannin glycosides and four anthocyanins. Among the latter, two components, i. e., cyanidin-malonyl-glucose and cyanidin-galloyl-hexose, have not been reported for S. officinalis so far. Furthermore, phenylethylamine was characterized for the first time in Sanguisorba by pH value dependent extraction with CH2 Cl2 . In addition, AcOEt and BuOH extracts were analyzed by GC/MS both prior to and after acid hydrolysis of secondary metabolites. For this purpose, the extracts were treated with 1 n HCl solution (105 °C, 1 h) and derivatized with BSTFA. Analyses revealed the occurrence of several classes of phenolic compounds, such as gallic acid, hydroxybenzoic acid, hydroxycinnamic acid and ellagic acid derivatives. Additionally, the most prominent ursane-type triterpenoid (ziyu-glycoside I) from Sanguisorba and its corresponding aglycone isomers were detected and assigned based on their characteristic fragmentation patterns.

A rapid and sensitive UHPLC-MS/MS method for the determination of ziyuglycoside I and its application in a preliminary pharmacokinetic study in healthy and leukopenic rats.

Ziyuglycoside I (ZgI), one of the main active ingredients in the popular Diyushengbai tablet made from Sanguisorba officinalis L., has been proven to relieve leukopenia clinically. However, to our knowledge, no studies have investigated the pharmacokinetics of either Diyushengbai tablet or ZgI in leukopenic vs. healthy individuals. In the present study, a rapid and sensitive UHPLC-MS/MS method was developed for detecting ZgI. On using this method on a novel cyclophosphamide-induced leukopenia model, we investigated differences in the pharmacokinetic characteristics of ZgI between leukopenic and normal rats. Chromatographic separation of ZgI and glycyrrhetinic acid (IS) was achieved via gradient elution in 0.5 min, and the total run time lasted for 5 min. Methodological validation results presented a good accuracy (102.6 %-110.8 %) and precision (% RSD ≤ 13.8) with a limit of quantitation of 0.5 ng/mL. Pharmacokinetic results showed a significantly shortened peak time (Tmax) (0.93 vs. 0.33 h) while a remarkably decreased maximum concentration (Cmax) (7.96 vs. 3.40 ng/L) in the 20 mg/kg leukopenia group in comparison with those in the 20 mg/kg normal group. In addition, a prolonged elimination half-life (t1/2ß) was observed in the 20 mg/kg leukopenia group (5.02 vs. 18.51 h). We observed similar trends in the 5 mg/kg oral dosing treatment and control groups, except for Cmax, which did not differ between the groups. We did not find pharmacokinetic differences in ZgI between the two leukopenia groups. Thus, the pharmacokinetic parameters of ZgI (e.g., Tmax, Cmax, and T1/2ß) changed based on the presence of a leukopenic state. This study may provide guidance for the development of ZgI as an agent for the treatment of leukopenia.

Anti-inflammatory Effects of Sanguisorbae Radix on Contact Dermatitis Induced by Dinitrofluorobenzene in Mice.

OBJECTIVE: To investigate the anti-inflflammatory effects of Sanguisorbae Radix on contact dermatitis (CD). METHODS: Mice were sensitized by painting 30 µL of 1-fluoro-2,4-dinitrofluorobenzene (DNFB) onto each ear for 3 days. Four days later, mice were challenged by painting with 50 µL of DNFB onto the shaved dorsum every 2 days. Sanguisorbae Radix methanol extract (MESR) was applied onto the shaved dorsum every 2 days. The effects of MESR on skin thickness, skin weights, histopathological changes, skin lesions and cytokine production in DNFB-induced CD mice were investigated, as well as its effects on body weights and spleen/body weight ratio. RESULTS: Topical application of MESR effectively inhibited enlargement of skin thickness and weight (P<0.05). MESR treatment also inhibited hyperplasia, spongiosis and immune cell infiltration induced by DNFB in inflamed tissues and improved lesions on dorsum skin in CD mice. Moreover, treatment with MESR suppressed the increase in the levels of tumor necrosis factor α (TNF-α,P<0.01) and interferon γ (IFN-γ,P<0.05), respectively. Finally, MESR had no effect on body weight gain or spleen/body weight ratio. CONCLUSION: These data suggest that MESR acts as an anti-inflflammatory agent that decreases the production of TNF-α and IFN-γ, resulting in reductions of skin lesions and histopathological changes in inflamed skin tissues.

Structural characterization and in vitro-in vivo evaluation of effect of a polysaccharide from Sanguisorba officinalis on acute kidney injury.

We report here an acidic polysaccharide, namely RSP-3, which ameliorates acute kidney injury and is obtained from Sanguisorba officinalis. We extracted and purified two polysaccharides from this herb based on the acidity and screened them for their effect in regulating the immunological activity of macrophages. Among them, RSP-3 exhibited significant anti-inflammatory activity against lipopolysaccharide (LPS)-stimulated macrophages by decreasing TNF-α and IL-6 levels. Subsequently, we found that RSP-3 suppressed ER stress, reduced ROS production and blocked NF-κBp65 translocation. After fully characterizing RSP-3 with a series of analytical technologies, we tested its anti-acute kidney injury (AKI) effect in vivo. In a murine AKI model induced by LPS, treatment with RSP-3 effectively ameliorated renal function. Besides, it decreased the levels of TNF-α and IL-6 in serum and reduced macrophage infiltration in injured kidney tissue. In sum, RSP-3, with a significant protective effect against AKI by showing anti-inflammatory activity, may become a meaningful drug candidate for treatment of AKI.