Thin Layer Identification of Jiedu Shengxue Granules and Determination of Notoginsenoside R1 Content

[Objectives]To establish the quality standard of hospital preparation Jiedu Shengxue granules.[Methods]Scleromitrion diffusum and Prunella vulgaris in Jiedu Shengxue granules were qualitatively identified by thin layer chromatography(TLC).A high performance liquid chromatography(HPLC)was established to determine the content of notoginsenoside R1 in the granule.[Results]The traditional Chinese medicinal materials in Jiedu Shengxue granules could be identified by TLC,and the characteristic spots were stable and clear.Notoginsenoside R1 had a good linear relationship in the range of 10.45-104.5μg/mL,with an average recovery of 98.52%and RSD=2.36%.[Conclusions]TLC and HPLC,as the quality control methods of Jiedu Shengxue granules,have high accuracy and good repeatability,which lays a foundation for the quality control of this mixture.

Notoginsenoside as an environmentally friendly shale inhibitor in water-based drilling fluid

The demand for non-toxic and biodegradable shale inhibitors is growing in the drilling industry.In this paper,the effect of notoginsenoside(NS)as a new,environmentally friendly inhibitor of shale hydration is systematically studied for the first time.The inhibition performance of NS was evaluated via inhibition evaluation tests,including mud ball immersion tests,linear expansion tests,shale rolling recovery tests,and compressive strength tests.The inhibition mechanism of NS was analyzed using Fourier transform infrared spectroscopy(FTIR),contact angle measurements,particle size distribution determination,thermogravimetric analysis(TGA),and scanning electron microscopy(SEM).The experimental results demonstrate that NS is able to adhere to the clay surface,forming a hydrophobic film that prevents the entry of water molecules and inhibiting the hydration dispersion of the clay.Because of this,NS can maintain the original state of bentonite pellets in water,which can effectively reduce the swelling rate of bentonite,increase the recovery rate of shale drill cuttings,maintain the strength of the shale,and therefore maintain the stability of the borehole wall during drilling.In addition,NS is non-toxic,degradable,and compatible with water-based drilling fluids.The above advantages make NS a promising candidate for use as an environmentally friendly shale inhibitor.

Role of Notoginsenoside Rg1 in Improving Spatial Cognitive Ability and Lowering Phosphorylation Level of Tau Protein in AD Model Rats

[Objectives] To study the effects and mechanism of notoginsenoside Rg1 on the spatial learning and memory and phosphorylated tau protein in the AD( Alzheimer's Disease) model rat. [Methods]The AD model rat was replicated by injection of Aβ_(25-35) in the left lateral ventricles of SD rats. The low dose( 25 mg/kg),middle dose( 50 mg/kg) and high dose( 100 mg/kg) notoginsenoside Rg1 was used for intragastric administration,respectively,two times every day. After 4 weeks,the Morris water maze test was done to detect the learning and memory capacity,and the immunoblotting,immunohistochemical methods were used to detect the changes in the phosphorylation level and distribution of tau protein in hippocampus of the rats. [Results] After the intracerebroventricular injection of Aβ_(25-35),the learning and memory capacity of the model rats was significantly lower than the learning and memory capacity of the normal control rats. The immunoblotting test results showed that the phosphorylation level of tau protein threonine 231 site( Thr231) in hippocampus was significantly increased,and the nonphosphorylation level was significantly decreased. The morphological testing results showed that the phosphorylation level of tau protein Thr231 of AD model rats was increased markedly in region of DG,CA1 and CA3 of the hippocampus. The intervention of the middle dose notoginsenoside Rg1 could significantly improve the learning and memory capacity of the model rats in Morris water maze. The notoginsenoside Rg1 in three different doses could all reduce the phosphorylation level of tau protein Thr231 in the hippocampal DG,CA1,CA3 regions,and there were no significant differences among the three doses. [Conclusions]The notoginsenoside Rg1 could improve Aβ_(25-35)-induced spatial learning and memory impairment of the AD model rats,and decreased the phosphorylation level of tau protein in hippocampus.

Different Concentrations of Notoginsenoside Rg1 Attenuate Hypoxic and Hypercapnia Pulmonary Hypertension by Reducing the Expression of ERK in Rat PASMCs

Pulmonary arterial hypertension (PAH) is a serious disease which is characterized by increased vascular resistance and pressure. We have previously hypothesized that panax notoginseng saponins (PNS) might attenuate pulmonary vasoconstriction under hypoxia and hypercapnia condition. This study aims to investigate the effect of notoginsenoside Rg1, a main ingredient of PNS, with various concentrations (8, 40, 100 mg/L, respectively) on extracellular signal regulated kinase (ERK1/2) signaling pathway in pulmonary arterial smooth muscle cells (PASMCs). In addition, PASMCs were randomly divided into six groups: SD rat under normoxic condition as control group (N group), hypoxia hypercapnia group (H group), DMSO control group (HD group), Rg1-treatment groups (RgLRgM and RgH group). Western-blot and RT-PCR were used to test the expression of p-ERK protein and the expression of ERK1 mRNA and ERK2 mRNA. This study provided the evidence that the expression of p-ERK protein and the expression of ERK1 mRNA and ERK2 mRNA in HD group and H group were obviously higher than that in N group (P < 0.01), Whereas the level of ERK1/2 mRNA in Rg1-treatment groups was significantly lower than that in HD group and H group (P < 0.01), and the proper concentration of Rg1 is 40 mg/L. These results suggested that notoginsenoside Rg1 can attenuate pulmonary vasoconstriction which may lead to HHPV through reducing the expression of ERK1/2.

Notoginsenoside R₁Attenuates Hypoxia and Hypercapnia-Induced Vasoconstriction <i>In Vitro</i>by Reducing the Expression of p38

Notoginsenoside R1, the main active ingredient of Panax notoginseng saponins (PNS), has been proposed to play fatal roles in the development of hypoxic hypercapnia-induced pulmonary vasoconstriction (HHPV). Subsequently, pulmonary arterial smooth muscle cells (PASMCs) lead to pulmonary vascular system remodeling and chronic pulmonary disease in the development of HHPV. Despite considerable studies have contributed to pulmonary disease, the mechanism of how Notoginsenoside R1 affects HHPV remains unclear. In this view, we will discuss the effect of notoginsenoside R1 by investigating the expression of p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway in PASMCs under hypoxia and hypercapnia condition. The third order PASMCs of Sprague Dawley (SD) rats were cultured with various concentrations (8, 40, 100 mg/L, respectively) of Notoginsenoside R1. Our data showed that the protein and mRNA expression levels of p-38 MAPK were higher in hypoxic hypercapnia group compared with hypoxic DMSO and normoxia control groups (p 1 treatment groups, the level of p-p38 MAPK protein and p38 MAPK mRNA were significantly decreased with different degrees (p 1 treatment may contribute to attenuate HHPV via decreasing the protein and mRNA expression levels of p-38 MAPK.

Notoginsenoside Ft1 inhibits colorectal cancer growth by increasing CD8^(+)T cell proportion in tumor-bearing mice through the USP9X signaling pathway

The management of colorectal cancer(CRC)poses a significant challenge,necessitating the development of innovative and effective therapeutics.Our research has shown that notoginsenoside Ft1(Ng-Ft1),a small molecule,markedly inhibits subcutaneous tumor formation in CRC and enhances the proportion of CD8^(+)T cells in tumor-bearing mice,thus restraining tumor growth.Investigation into the mechanism revealed that Ng-Ft1 selectively targets the deubiquitination enzyme USP9X,undermining its role in shieldingβ-catenin.This leads to a reduction in the expression of downstream effectors in the Wnt signaling pathway.These findings indicate that Ng-Ft1 could be a promising small-molecule treatment for CRC,working by blocking tumor progression via the Wnt signaling pathway and augmenting CD8^(+)T cell prevalence within the tumor environment.

Pharmacological properties and mechanisms of Notoginsenoside R1 in ischemia-reperfusion injury

Panax notoginseng is an ancient Chinese medicinal plant that has great clinical value in regulating cardiovascular disease in China.As a single component of panax notoginosides,notoginsenoside R1(NGR1)belongs to the panaxatriol group.Many reports have demonstrated that NGR1 exerts multiple pharmacological effects in ischemic stroke,myocardial infarction,acute renal injury,and intestinal injury.Here,we outline the available reports on the pharmacological effects of NGR1 in ischemia-reperfusion(I/R)injury.We also discuss the chemistry,composition and molecular mechanism underlying the anti-I/R injury effects of NGR1.NGR1 had significant effects on reducing cerebral infarct size and neurological deficits in cerebral I/R injury,ameliorating the impaired mitochondrial morphology in myocardial I/R injury,decreasing kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin in renal I/R injury and attenuating jejunal mucosal epithelium injury in intestinal I/R injury.The various organ anti-I/R injury effects of NGR1 are mainly through the suppression of oxidative stress,apoptosis,inflammation,endoplasmic reticulum stress and promotion of angiogenesis and neurogenesis.These findings provide a reference basis for future research of NGR1 on I/R injury.

达玛烷皂苷Ginsenoside Re与Notoginsenoside R_1的全合成

采用汇聚式合成的方式,充分利用原人参三醇各羟基活性的差异,采用高效的区域选择性保护策略,并以Au(I)催化的糖苷化反应对达玛烷20位大位阻酸敏感羟基和6位羟基及6-O-Glc'糖基2位羟基连续三次糖苷化,以最长线性13步分别以5.1%和4.5%的收率合成了天然双糖基达玛烷皂苷Ginsenoside Re(1)和Notoginsenoside R1(2).

Notoginsenoside Ft1 acts as a TGR5 agonist but FXR antagonist to alleviate high fat diet-induced obesity and insulin resistance in mice

Obesity and its associated complications are highly related to a current public health crisis around the world.A growing body of evidence has indicated that G-protein coupled bile acid(BA) receptor TGR5(also known as Gpbar-1) is a potential drug target to treat obesity and associated metabolic disorders.We have identified notoginsenoside Ftl(Ftl) from Panax notoginseng as an agonist of TGR5 in vitro.However,the pharmacological effects of Ftl on diet-induced obese(DIO) mice and the underlying mechanisms are still elusive.Here we show that Ftl(100 mg/100 diet) increased adipose lipolysis,promoted fat browning in inguinal adipose tissue and induced glucagon-like peptide-1(GLP-1) secretion in the ileum of wild type but not Tgr5^(-/-) obese mice.In addition,Ftl elevated serum free and taurineconjugated bile acids(BAs) by antagonizing Fxr transcriptional activities in the ileum to activate Tgr5 in the adipose tissues.The metabolic benefits of Ftl were abolished in Cyp27 al^(-/-) mice which have much lower BA levels.These results identify Ftl as a single compound with opposite activities on two key BA receptors to alleviate high fat diet-induced obesity and insulin resistance in mice.

Notoginsenoside R1 attenuates breast cancer progression by targeting CCND2 and YBX3

Background:Breast cancer(BC)is a common malignancy with highly female incidence.So far the function of notoginsenoside R1(NGR1),the extract from Panax notoginseng,has not been clearly elucidated in BC.Methods:Optimal culture concentration and time of NGR1 were investigated by cell counting kit-8 assay.Cell proliferation ability was measured by colony formation assays.Transwell assay was used to detect the effect of NGR1 on cell migration and invasion.The apoptosis rate of cells between each group was measured by TUNEL assay.Results:NGR1 treatment has an inhibitory efect on proliferation,migration,invasion,and angiogenesis and a stimulating effect on cell cycle arrest and apoptosis of Michigan Cancer Foundation-7(MCF-7)cells.The 50%growth inhibitory concentration for MCF-7 cells at 24 h was 148.9 mmol/L.The proportions of MCF-7 cells arrested in the G0/G1 phase were 36.94+6.78%,45.06士5.60%,and 59.46+5.60%in the control group,75,and 150 mmol/L groups,respectively.Furthermore,we revealed that NGR1 treatment attenuates BC progression by targeted downregulating CCND2 and YBX3 genes.Additionally,YBX3 activates phosphatidylinositol 3-phosphate kinase(PI3K)/protein kinase B(Akt)signaling pathway by activating kirsten rat sarcoma viral oncogene,which is an activator of the PI3K/Akt signaling pathway.Conclusion:These results suggest that NGR1 can act as an efficacious drug candidate that targets the YBX3/PI3K/Akt axis in patients with BC.

Pro-angiogenic Activity of Notoginsenoside R1 in Human Umbilical Vein Endothelial Cells in vitro and in a Chemical-Induced Blood Vessel Loss Model of Zebrafish in vivo

Objective： This study aimed at investigating whether notoginsenoside R1 （R1）, a unique saponin found in Panax notoginseng could promote angiogenic activity on human umbilical vein endothelial cells （HUVECs） and elucidate their potential molecular mechanisms. In addition, vascular restorative activities of R1 was assessed in a chemically-induced blood vessel loss model in zebrafish. Methods： The in vitro angiogenic effect of R1 was compared with other previously reported angiogenic saponins Rgl and Re. The HUVECs proliferation in the presence of R1 was determined by cell proliferation kit lI （XTI＇） assay. R1, Rgl and Re-induced HUVECs invasion across polycarbonate membrane was stained with Hoechst-33342 and quantified microscopically. Tube formation assay using matrigel- coated wells was performed to evaluate the pro-angiogenic actions of RI. In order to understand the mechanism underlying the pro-angiogenic effect, various pathway inhibitors such as SU5416, wortmannin （wort） or L-N （o -nitro- L-arginine methyl ester hydrochloride （L-NAME）, SH-6 were used to probe the possible involvement of signaling pathway in the R1 mediated HUVECs proliferation. In in vivo assays, zebrafish embryos at 21 hpf were pre-treated with vascular endothelial growth factor （VEGF） receptor kinase inhibitor ]1 （VRI） for 3 h only and subsequently post-treated with R1 for 48 h, respectively. The intersegmental vessels （ISVs） in zebrafish were assessed for the restorative effect of R1 on defective blood vessels. Results： R1 could stimulate the proliferation of HUVECs. In the chemoinvasion assay, R1 significantly increased the number of cross-membrane HUVECs. In addition, R1 markedly enhanced the tube formation ability of HUVECs. The proliferative effects of these saponins on HUVECs were effectively blocked by the addition of SU5416 （a VEGF-KDR/FIk-1 inhibitor）. Similarly, pre-treatment with wort [a phosphatidylinositol 3-kinase （PI3K）-kinase inhibitor], L-NAME [an endothelial nitric oxide synthase （eNOS） inhibitor] or SH-6 （an Akt pathway inhibitor） significantly abrogated the R1 induced proliferation of HUVECs. In chemically- induced blood vessel loss model in zebrafish, R1 significantly rescue the damaged ISVs. Conclusion： R1, similar to Rgl and Re, had been showed pro-angiogenic action, possibly via the activation of the VEGF-KDR/FIk-1 and PI3K- Akt-eNOS signaling pathways. Our findings also shed light on intriguing pro-angiogenic effect of R1 under deficient angiogenesis condition in a pharmacologic-induced blood vessels loss model in zebrafish. The present study in vivo and in vitro provided scientific evidence to explain the ethnomedical use of Panax notoginseng in the treatment of cardiovascular diseases, traumatic injuries and wound healing.

Pharmacokinetics and brain distribution differences of baicalin in rat underlying the effect of Panax notoginsenosides after intravenous administration

Baicalin(BA) is the most well-known flavonoid present in Radix Scutellariae. The aim of this study was to explore whether the pharmacokinetic behavior of BA in rat brain can be affected by Panax notoginsenosides(PNS), and to assess the possible mechanism for the observed effects. Specific HPLC and HPLC/MS/MS methods were developed and validated for the determination of BA in the rat plasma and brain using carbamazepine as an internal standard. BA was found to enter rat brain quickly after a single intravenous dose. When co-administered with PNS, clearance(CL) of BA from rat plasma decreased by 50.00%, while the area under the curve AUC0-t and AUC0-∞ increased 94.69% and 87.68%, respectively. On the other hand, some pharmacokinetic parameters of BA in rat brain had obvious differences after PNS was administered, such as an increase in Tmax from 5 min to 15 min, an increase in AUC0-t and AUC0-∞ by 42.75% and 29.39%, respectively, as well as a decrease in CL by 27.95%. Together, these results indicate that PNS can decrease the elimination rate of BA from rat plasma, promote the penetration of BA into rat brain, increase the concentration and slow down the elimination of BA from rat brain. The data provide important information that compatibility with PNS can promote the consequent effects of BA for the treatment of encephalopathy.

Appropriate choice of collision-induced dissociation energy for qualitative analysis of notoginsenosides based on liquid chromatography hybrid ion trap time-of-flight mass spectrometry

Liquid chromatography hybrid ion trap/time-of-flight mass spectrometry possesses both the MS^n ability of ion trap and the excellent resolution of a time-of-flight and has been widely used to identify drug metabolites and determine trace multi-components in natural products. Collision energy, one of the most important factors in acquiring MS^n information, could be set freely in the range of 10%–400%. Herein, notoginsenosides were chosen as model compounds to build a novel methodology for the collision energy optimization. Firstly, the fragmental patterns of the representatives for the authentic standards of protopanaxadiol-type and protopanaxatriol-type notoginsenosides were obtained based on accurate MS^2 and MS^3 measurements via liquid chromatography hybrid ion trap/time-of-flight mass spectrometry. The extracted ion chromatograms of characteristic product ions of notoginsenosides in Panax Notoginseng Extract were produced under a series of collision energies and compared to screen the optimum collision energies values for MS^2 and MS^3. The results demonstrated that the qualitative capability of liquid chromatography hybrid ion trap/time-of-flight mass spectrometry was greatly influenced by collision energies, and 50% of MS^2 collision energy was found to produce the highest collision-induced dissociation efficiency for notoginsenosides. Addtionally, the highest collision-induced dissociation efficiency appeared when the collision energy was set at 75% in the MS^3 stage.

LC-MS/MS同时定量测定大鼠血浆中复方血栓通胶囊的8种入血成分

目的：建立复方血栓通胶囊中8种有效成分（三七皂苷R1、人参皂苷Rg1、人参皂苷Rb1、人参皂苷Re、人参皂苷Rd、丹参酮Ⅰ、哈巴俄苷、黄芪甲苷）在大鼠血浆中的液相色谱-质谱/质谱定量测定方法。方法色谱柱：Thermo Hypersil GOLD（2.1 mm×100 mm,5μm）；柱温：30℃；流动相：A为1%甲酸,B为乙腈；梯度洗脱：0～10 min 25%～55%B,10～20 min 55%～70%B；流速：0.2 mL/min；进样量：10μL。离子源：ESI源,正离子模式,SRM扫描。结果以盐酸巴马汀为内标,三七皂苷R1、人参皂苷Rg1、人参皂苷Re、人参皂苷Rb1、人参皂苷Rd、哈巴俄苷、黄芪甲苷、丹参酮Ⅰ的检测范围分别为1.00～800 ng/mL、0.950～760 ng/mL、1.44～1440 ng/mL、1.33～1330 ng/mL、9.90～990 ng/mL、1.01～1010 ng/mL、1.16～928 ng/mL、10.0～800 ng/mL。准确度分别在85%～115%之间,绝对回收率达到50%～70%,日内精密度与日间精密度RSD≤15%。结论该方法快速、灵敏、重复性好,可用于复方血栓通胶囊8种有效成分在大鼠体内的相关药代动力学研究。

A Novel Hexanordammarane Glycoside from the Roots of Panax notoginseng

Notoginsenoside R-111, a novel hexanordammarane glycoside was isolated from the roots of Panax nototginseng, as a minor constituent. Its structure was determined as 6-O-(beta -D-glucopyranosyl)-3 beta ,6 alpha ,12 beta -trihydroxy-22,23,24,25,26,27-hexanordammaran-20-one(1), by means of spectroscopic methods.

Two New Dammarane Glycosides from the Acid Hydrolysis Product of Panax Notoginseng

Two new dammarane glycosides named notoginsenoside T-1 and T-2 were isolated from the mild acid hydrolysis products of the root saponins of Panax notoginseng. On the basis of spectroscopic evidences, their structures were elucidated to be 6-O-beta -D-glucopyranosyl-24(25)-epoxy-3 beta ,6 alpha ,12 beta ,23 xi -tetrahydroxydammar-20(22)(E)-ene 1 and 6-O-beta -D-glucopyranosyl-24(25)-epoxy-23 xi -methoxyl-3 beta ,6 alpha ,12 beta -trihydroxydamm-ar-20(22)(E)-ene 2, respectively.

Dammarane-type saponins from steamed leaves of Panax notoginseng

Four new dammarane-type triterpenoid saponins,namely notoginsenosides SFt1-SFt4(1-4)were isolated from the steamed leaves of Panax notoginseng(Burk.)F.H.Chen(Araliaceae),together with 17 known saponins.Their structures were established on the basis of detailed spectroscopic analyses and acidic hydrolysis.The known ginsenosides Rk2 and Rh3 were obtained from P.notoginseng for the first time.All of these new saponins showed no in vitro cytotoxicity against five human cancer cell lines(HL-60,SMMC-7712,A-549,MCF-7,and SW480).

Stability of Xuesaitong capsules in gastrointestinal lavage fluid

Objective:To obtain a formulation with high bioavailability through evaluation of the stability of three types of Xuesaitong capsules in the stomachs and intestines of rats.We compared the stability of the Panax notoginsenoside R1 as well as the ginsenosides Rg1,Rb1,Re,and Rd in different formulations.Methods:Artificial stomach fluid(ASF)and artificial intestinal fluid(AIF)were prepared.Stability of three types of Xuesaitong capsules was examined for 4 h in stomachs and 24 h in intestines.Samples were analyzed at different times by high-performance liquid chromatography.Percent content of NGR1,GRg1,GRb1,GRe,and GRd at different times was calculated.Results:Hard capsules incubated in ASF disintegrated within 2e3 min,whereas soft capsules disintegrated within 7e8 min.Components in hard capsules were dissolved rapidly in water,with content of each compound reaching 90%in 5 min,and degradation of each compound reaching 30e50%after incubation for 240 min.Dissolution and degradation of each component in soft capsules with a water-soluble base tended to balance at 30 e90 min.Contents in soft capsules with a lipid-soluble base showed slow dissolution after ASF incubation for 120 min.Five saponins in identical types of capsules incubated in ASF had similar stability curves.Contents of hard capsules and soft capsules with a watersoluble base degraded rapidly within 30 min and reached a plateau when Xuesaitong capsules were incubated in AIF.

Water Extraction Process of Chinese Herbal Compound Man Gan Ning

[Objectives]To optimize the water extraction process of Chinese Herbal Compound Man Gan Ning and establish a method for its extraction and content determination.[Methods]The L9(34)orthogonal test was carried out,the comprehensive scores of ginsenoside Rb1 content,notoginsenoside R1 content,tanshinone I content and saikosaponin A content in the compound extract were used as evaluation indicators to explore the effects of water addition,the water addition,decoction time and decoction times on the water extraction process,and verification tests were carried out.[Results]The optimized water extraction process was as follows:adding 9 times of water in the first extraction,and extracting for 90 min;adding 7 times of water in the second extraction,and extracting for 60 min;adding 7 times of water in the third extraction,and extracting for 60 min.[Conclusions]The optimized water extraction process is stable and feasible,and can be used for the extraction of various herbs in Compound Man Gan Ning.It can also provide experimental basis for the formulation development of Compound Man Gan Ning sustained release tablet.

A rapid method for chemical fingerprint analysis of Panax notoginseng powders by ultra performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry

A method coupling ultra-performance liquid chromatography(UPLC) with quadrupole time-of-flight mass spectrometer(Qtof MS) using the electrospray ionization(ESI) source was developed for the identification of the major saponins from Panax notoginseng powder(PNP). Ten different PNP samples were analyzed and evaluated for their quality by similarity evaluation and principle component analysis(PCA). Based on the accurate mass, summarized characteristic fragmentation behaviors, retention times of different types of saponins, related botanical biogenesis, and reported chromatographic behavior of saponins, fifty-one common peaks were effectively separated and identified, including 28 protopanaxadiol saponins and 18 protopanaxatriol saponins. Simultaneously, 15 significant discrepancy compounds were identified from the disqualified PNP samples. The established UPLC/Qtof MS fingerprint method was successfully applied for profiling and identifying the major saponins of PNP, providing a fast quality evaluation tool for distinguishing the authentic PNP and the adulterated products.