Pharmacometabolomics and mass spectrometry imaging approach to reveal the neurochemical mechanisms of Polygala tenuifolia

Polygala tenuifolia,commonly known as Yuanzhi(YZ)in Chinese,has been shown to possess antiinsomnia properties.However,the material basis and the mechanism underlying its sedative-hypnotic effects remain unclear.Herein,we investigated the active components and neurochemical mechanism of YZ extracts using liquid chromatography tandem mass spectrometry(LC-MS/MS)-based pharmacometabolomics and mass spectrometry imaging(MSI)-based spatial resolved metabolomics.According to the results,17 prototypes out of 101 ingredients in the YZ extract were detected in both the plasma and brain,which might be the major components contributing to the sedative-hypnotic effects.Network pharmacology analysis revealed that these prototypes may exert their effects through neuroactive ligand-receptor interaction,serotonergic synapse,dopaminergic synapse,and dopaminergic synapse,among other pathways.LC-MS/MS-based targeted metabolomics and Western blot(WB)revealed that tryptophan-serotonin-melatonin(Trp-5-HT-Mel)and tyrosine-norepinephrine-adrenaline(Tyr-Ne-Ad)are the key regulated pathways.Dopa decarboxylase(DDC)upregulation and phenylethanolamine Nmethyltransferase(PNMT)downregulation further confirmed these pathways.Furthermore,MSI-based spatially resolved metabolomics revealed notable alterations in 5-HT in the pineal gland(PG),and Ad in the brainstem,including the middle brain(MB),pons(PN),and hypothalamus(HY).In summary,this study illustrates the efficacy of an integrated multidimensional metabolomics approach in unraveling the sedative-hypnotic effects and neurochemical mechanisms of a Chinese herbal medicine,YZ.

Antidepressant effects of Yuanzhi (Polygalae Radix) extract on chronic unpredictable mild stress-induced depression in rats: modulation of the NLRP3 inflammasome and NF-κB pathway

Objective To investigate the antidepressant effects of Yuanzhi(Polygalae Radix;PR)aqueous extract on chronic unpredictable mild stress(CUMS)-induced depression rat models and the underlying mechanisms.Methods A total of 40 male Sprague Dawley(SD)rats were randomly divided into control;model;low dose of PR(PR-L;0.5 g/kg);high dose of PR(PR-H;1 g/kg);and fluoxetine(10 mg/kg)groups;with 8 rats in each group.Except for the rats in control group;those in the other four groups underwent CUMS-induced depression modeling.PR and fluoxetine were administered intragastrically once daily;30 min prior to the CUMS procedure;for 14 consecu-tive days until the behavioral tests were performed.After CUMS modeling;the sucrose prefer-ence test(SPT);open field test(OFT);novelty-suppressed feeding test(NSFT);forced swim test(FST);and tail suspension test(TST)were employed to assess the pharmacological ef-fects of PR on the mitigation of depressive-like behaviors in rat models.Additionally;the en-zyme-linked immunosorbent assay(ELISA)was utilized to quantify the serum levels of tumor necrosis factor(TNF)-α;interleukin(IL)-6;and IL-1βin the rats.Western blot analysis was al-so conducted to evaluate the protein expression levels of nuclear factor kappa-B(NF-κB);in-ducible nitric oxide synthase(iNOS);cyclooxygenase-2(COX-2);nucleotide-binding oligomerization domain(NOD)-like receptor family pyrin domain containing 3(NLRP3);apoptosis-associated speck-like protein containing caspase recruitment domain(ASC);and caspase-1 in the hippocampal tissues of the rats.Immunofluorescence staining was per-formed to observe the morphological changes in ionized calcium-binding adapter molecule 1 positive(Iba-1+)cells in the dentate gyrus(DG)of rats with CUMS-induced depression.Results(i)Treatment with PR-H and fluoxetine resulted in significant enhancements in both the total distance and time the rats moved during tests(P<0.01 and P<0.05;respectively).Post-administration of PR-H and fluoxetine also led to statistically significant increase in su-crose preference among rats(P<0.05).Besides;PR-L;PR-H;and fluoxetine treatment markedly decreased the latency of ingestion(P<0.05;P<0.05;and P<0.01;respectively).As observed from the FST;PR-L;PR-H;and fluoxetine presented antidepressant effects on rats with CUMS-induced depression;leading to the reduction in time of their immobility(P<0.05;P<0.01;and P<0.01;respectively).The results of TST indicated reduced immobility time in rats receiving PR-H and fluoxetine treatment as well(P<0.01).(ii)Rats in model group showed an increase in the levels of Iba-1+microglia in their left and right brains in compari-son with control group(P<0.01).However;such increase was negated post PR treatment(P<0.01).Treatment with PR-L;PR-H;and fluoxetine considerably reduced the levels of inflam-matory factors(TNF-α;IL-1β;and IL-6;P<0.01).In addition;treatment of PR-L and PR-H ef-fectively counteracted the elevated levels of NLRP3;ASC;and caspase-1;and markedly down-regulated the expression levels of phosphorylated p65(p-p65);COX-2;and iNOS in rats’hip-pocampus(P<0.01).Conclusion Collectively;these findings indicate that PR exerts an antidepressant effect on rats with CUMS-induced depression partially through the modulation of the NLRP3 and NF-κB signaling pathways.

Effects of Polygala fallax Hemsl Water Extract on a Mouse Model of Gastric Motility Disorders

[Objectives]To explore the effects of Polygona fallax Hemsl water extract on gastrointestinal motility in normal mice and gastric motility disorder model mice and approximate mechanism.[Methods]Using normal mice and mice with gastric motility disorders(modeled with atropine),the effects of different mass concentration groups of P.fallax Hemsl water extract(0.125,0.250,0.500 g/mL)and domperidone groups on gastric residual rate,small intestine propulsion rate,serum motilin(MLT),vasoactive intestinal peptide(VIP),and tissue morphology were studied.[Results]There was a highly significant difference(P<0.01)in the small intestine propulsion rate of liquid in normal mice among the different concentration groups of P.fallax Hemsl water extract compared to the blank group.The small intestine propulsion rate and gastric residue rate of semi-solid paste were statistically significant compared to the blank group(P<0.05).Among them,there was a highly significant difference between the high concentration group(67.75%±7.65%,46.5%±10.62%)and the medium concentration group(60.90%±5.87%,59.27%±7.82%)(P<0.01).There was statistical significance in normal mouse serum MLT content in the high concentration group(P<0.05).There was no effect on serum VIP levels in normal mice;no effect on the morphology of stomach and intestinal tissues of normal mice.The small intestine propulsion rate and gastric residue rate of liquid and semi-solid paste in mice with gastric motility disorders were statistically significant compared to the atropine group,with extremely significant differences(P<0.01).[Conclusions]P.fallax Hemsl water extract has a promoting effect on gastrointestinal motility.One of the specific mechanisms by which P.fallax Hemsl promotes gastrointestinal motility in normal mice may be related to the content of MLT in mouse serum.The mechanism of action in atropine induced gastric paresis mice may be related to the reactivation of M receptors,and the action mechanism of P.fallax Hemsl does not change the original histological basis.It can be inferred that P.fallax Hemsl water extract has a synergistic effect on promoting gastrointestinal motility through other mechanisms,but it is not fully understood and further in-depth research is needed.

Chemical constituents from the roots of Polygala sibirica L.

To investigate the chemical constituents of the roots ofPolygala sibirica L. The separation and purification were performed by solvent extraction and repeated chromatography with silica gel, Sephadex LH-20, ODS columns, and semiprep. HPLC. The structures were elucidated by spectral analysis. Twelve known compounds were isolated and identified as tenuifoliside A （1）, tenuifoliside B （2）, glomeratose A （3）, 3＇,6-disinapoyl sucrose （4）, sibiricose A5 （5）, sibiricose A6 （6）, sibiricose A1 （7）, sibiricose A2 （8）, polygalatenoside E （9）, 1-O-L-arabinopyranosyl-O-（6→1）-β-D-glucopyranosyl-salicylate （10）, canthoside A （11）, and methyl- 3,4,5-trimethoxycinnamate （12）. Compound 11 was obtained from genus Polygala for the first time, and compounds 2, 9, 10 and 12 were isolated from this plant for the first time.

黄花倒水莲(Polygala fallax)多糖含量的测定

目的建立黄花倒水莲多糖含量的测定方法.方法以葡萄糖为对照品,采用蒽酮-硫酸法在625nm处测定多糖含量.结果葡萄糖对照品溶液在0.02mg/mL-0.12mg/mL范围内呈良好曲线关系,线性回归方程为:A=11.8926C-0.1363,R2=0.9986,平均加样回收率为100.4%,RSD为1.69%.结论该方法操作方便、快速、重现性好,可用于黄花倒水莲多糖含量的测定.

黄花倒水莲(Polygala fallax Hemsl.)对^(60)Coγ射线辐射损伤小鼠组织DNA的防护作用

目的:探讨黄花倒水莲对辐射损伤小鼠组织DNA的防护作用.方法:实验设计分照射前给药和照射后给药2个模块,每模块各5组.复制60Coγ射线(辐射强度为6Gy)对雄性昆明种小鼠辐射损伤的病理模型,黄花倒水莲提取液2、4、8 g/(kg.d)连续灌胃7天,正常对照组、照射模型组给予等量生理盐水,用彗星试验检测小鼠多种组织细胞DNA损伤情况.结果:黄花倒水莲照射前给药和照射后给药,均可明显降低小鼠肺、肝、脾、睾丸、淋巴细胞DNA的尾部DNA百分率和尾矩(P<0.01),呈一定的量效关系.结论:黄花倒水莲可以减轻60Coγ射线所致小鼠组织DNA的损伤,具有辐射防护作用.

黄花倒水莲(Polygala fallax Hemsl)组培快繁技术研究

以当年生黄花倒水莲幼嫩带腋芽茎段作为外植体,开展组织培养试验研究。结果表明:最佳外植体诱导培养基为1/2 MS+BA 2.0 mg/L+NAA 0.1 mg/L,诱导率可达95.1%;最佳增殖培养基为WPM+BA 1.5 mg/L+NAA 0.1 mg/L,增殖系数为6.12,周期为25 d,不定芽生长状况好;最佳生根培养基为1/2 WPM+IBA0.1 mg/L+ABT0.4 mg/L,生根率为95.67%;以泥炭土∶黄泥土∶珍珠岩(2∶1∶1)为移栽基质,移栽成活率可达92.6%,苗木长势好,叶色绿。

Three New Xanthones from Polygala crotalarioides

Three new xanthones, 3, 6-dihydroxy-1, 2, 7-trimethoxyxanthone 1, 1, 3, 6-trihydroxy-2, 7, 8-trimethoxyxanthone 2, and 3, 6-dihydroxy-1, 2, 7, 8-tetramethoxyxanthone 3, were isolated from the roots of Polygala crotalarioides. Their structures were elucidated by spectral and chemical methods.

Two New Xanthone Glycosides from Polygala tenuifolia

Two new xanthone glycosides, polygalaxanthone IV and V were isolated from the roots of Polygala tenuifolia Willd. Their structures were established as 6-O-[a-L-rhamnopyranosyl- (12)-b-D-glucopyranosyl]-1-hydroxy-3, 7-dimethoxyxanthone (polygalaxanthone IV), and 6-O- [a-L-rhamnopyranosyl-(12)-b-D-glucopyranosyl]-1, 3-dihydroxy-7-methoxyxanthone (polyga- laxanthone V), respectively, on the basis of chemical and spectral evidence.

A new glycoside from Polygala tenuifolia Willd

A new tdterpenoid glycoside, 3-β-O-β-D-glucopyranosyl presenegenin 28-O-α-L-arabipyranosyl（1 → 3）-6-β- xylopyranosyl（1 → 4）-[β-D-apiofuranosyl（1 → 3）]-Ct-L-rhamnopyranosyl（1 → 2）-[α-L-rhamnopyranosyl（1 → 3）]-β-D-fucopyranosyl ester （1） was isolated from the Polygala tenuifolia Willd., together with two known saponins, including polygalasaponinXXIV （2） and polygalasaponinXXVIII （3）. The structure of new compound was elucidated by spectroscopic methods.

Two new triterpenoid saponins from Polygala crotalarioides

Two new oleanane-type saponins,crotalarioside A（1） and crotalarioside B（2）,were isolated from the roots of Polygala crotalarioides.Their structures were elucidated on the basis of spectroscopic and chemical evidence.

Studies on chemical constituents of Polygala japonica Houtt.

Objective:To investigate the chemical constituents of Polygala japonica Houtt.. Methods:The constituents were separated and purified by column chromatographies with silical gel, RP-silical gel and Sephadex LH-20. Their structures were elucidated on the basis of spectral data (IR, MS, 1H-NMR, 13C-NMR, HMQC and HMBC). Results: Totally 15 compounds were obtained from Polygala japonica Houtt. And their structures were identified as castaprenol-12 (Ⅰ), β-sitosterol (Ⅱ), stigmasterol (Ⅲ),stigmasta-7,22-di-ene-3-one (Ⅳ), n-dotriacontanol ( Ⅴ ), n-hexadecanol ( Ⅵ ), arachidic acid (Ⅶ), erogosterol (Ⅷ), kaempferol (Ⅸ), quercetin (Ⅹ), daucosterol (Ⅺ), p-hydroxybenzoic acid (Ⅻ),coumaric acid (ⅩⅢ), rhamnocitrin 3-O-β-D-galactopyranoside (ⅪⅤ), quercetin 3-O-β-D-glucopyranoside (ⅩⅤ). Conclusion: Compounds Ⅰ , Ⅲ , Ⅳ, Ⅴ , Ⅵ, Ⅶ, Ⅷ, Ⅻ , ⅩⅢ and ⅩⅣ were obtained from this plant for the first time.

Dioxyxanthones from Polygala hongkongensis and Their Cytotoxicity

Two new dioxyxanthones, polyhongkongenoxanthones A and B（1 and 2） were isolated from the herbs of Polygala hongkongensis, together with six known xanthones. Their structures were elucidated on the basis of chemical and spectroscopic evidence. The isolates were tested for their cytotoxicity against three tumor cell lines（HepG2, GLC-82 and MCF-7, HepG2=human hepatocellular carcinoma cells; GLC-82=human lung carcinoma cells; MCF-7= human breast carcinoma cells） by MTT assay, among which polyhongkongenoxanthone B（2）, 1,7-dihydroxy-2,3- methylenedioxyxanthone（3） and 1,7-dihydroxy-3,4,8-trimethoxyxanthone（6） are potential antitumor candidate due to their significant cytotoxic effects on the three cell lines..

A new benzophenone C-glycoside from Polygala telephioides Willd

A new benzophenone C-glucoside,3’-C-[4-O-(5-hydroxyferuloyl)-β-D-glucopyranosyl]-2′,4′,6′-trihydroxy-3,4-dimethoxy-benzophenone, named telephenone D,was isolated from the whole plants of Polygala telephioides,and its structure was determined by analysis of spectroscopic data.

Research on the Effects of the Extract of Polygala fallax Hemsl on Sex Hormones andβ-EP in Perimenopausal Rat Models

Objective:To study the effects of the ethnic medicine Polygala fallax Hemsl with Guangxi characteristics on the sex hormones andβ-EP in research objective perimenopausal rat models.Methods:40 female SPF rats were randomly divided into 4 groups,including the normal,model,high-dose and lowdose groups.Rats of three groups except for the normal one were treated with perimenopausal modelling through the method of subcutaneous injection of compound 4-VCD for 15 consecutive days.Rats of the normal and model group were normally fed without any treatment.Rats of the high-dose and low-dose groups were administered by high-and low-dose intragastric administration of the extract of Polygala fallax Hemsl.According to the menstrual cycle of the vaginal smear of the rat,each menstrual cycle is a course of treatment and 6 consecutive courses of treatment would be given.The indexes of serum sex hormones(E2,FSH,LH)andβ-EP of rats in each group were observed after treatment.Results:After the treatment of 6 cycles,for the levels ofβ-EP and E2,the model group was lowest(P<0.05),the normal group was highest(P<0.05);and the high-dose group was higher than the low-dose group;For the levels of FSH and LH,the normal group was lowest(P<0.05),the model group was highest(P<0.05),and the high-dose group was lower than the low-dose group.Conclusion:Guangxi characteristic national medicine Polygala fallax Hemsl can effectively improve the levels of serum sex hormones andβ-EP in perimenopausal rat models and relieve the related symptoms with a certain dose-effect relationship.

New Xanthones from Polygala crotalarioides

Introduction Xanthones are secondary metabolites commonly occurring in a few higher plant families, fungi and lichen. Their high taxonomic value in such families and their pharmacological properties, including in vitro cytotoxicity, in vivo antitumour activity, antifungal, antibacterial, antiinflammatory properties and aldose reductase inhibitory activity have provoked great interest. The discovery of the antidiabetic activity of some axnthones＇ from Swertiaceae has prompted us to undertake a more broadly search for novel xanthones from plants of Polygalaceae. The phytochemical investigation has not been performed on this plant up to now. Herein we report the isolation and structure elucidation of two new xanthones （ compounds 1 and 2 ） and three known xanthones （compound 3, 4 and 5 ）.

Effects of Different Storage Conditions on Physiological and Biochemical Characteristics of Polygala tenuifolia Willd. Seeds

[Objectives] Taking Polygala tenuifolia Willd. seeds as experimental material, the changes of physiological and biochemical characteristics of seeds under different storage temperatures and seed moisture contents were studied. [Methods] The seeds of P. tenuifolia Willd. with moisture contents of 3%, 5% and 15% were stored at-10 ℃, 0 ℃, 20 ℃ and 36 ℃, respectively, and the storage time was one year. The physiological and biochemical characteristics of the seeds, such as conductivity, SOD activity, POD activity, and soluble sugar content were studied.[Results] After one-year storage, as the storage temperature and seed moisture content increased, the relative conductivity, MDA content and soluble sugar content of P. tenuifolia Willd. seeds increased, while the SOD activity, POD activity, and protein content declined. This phenomenon was most obvious in the P. tenuifolia Willd. seeds with moisture content of 15%. [Conclusions] This study will provide reference for seed storage of medicinal plants.

Onjisaponin B derived from Radix Polygalae enhances autophagy and accelerates the degradation of mutant α-Synuclein and Huntingtin in PC-12 cells

OBJECTIVE To investigate the autophagic effect of the compounds from the Chinese medicinal herbs,Radix polygalae as a potential neuroprotective agent that enhances the clearance of mutant Huntingtin andα-synuclein in PC-12 cells.METHODS Radix polygalae was extracted with 75%ethanol using refluxing method,and its quality was assayed by UHPLC-TOF-MS.The autophagic effect of Radix polygalae extract,and its major components including polygalacic acid,senegenin and onjisaponin B were investigated using the green fluorescent protein-light chain 3(GFP-LC3)autophagy detection and Western blot platforms for detecting the autophagic markers,GFP-LC3 puncta formation and LC3Ⅱ expression.The degradation of A53Tα-synuclein and the inhibition of α-synuclein oligo merization related to the Parkinson disease were assayed using Western blot and flow cytometer analysis,respectively.While the cytotoxicity and the degradation of the mutant Huntingtin associated with the Huntingtin disease were investigated using MTT method and flow cytometer analysis.RESULTS Radix polygalae ethanol extract and onjisaponin B improved the GFP-LC3 puncta formation and expression of LC3Ⅱ with time and dose manner,and this induction was activated via AMPK-m TOR and Atg 7 gene pathway.Furthermore,the clearance ofα-synuclein and mutant Huntingtin was enhanced via autophagy induction with the treatment of Radix polygalae ethanol extract and onjisaponin B.CONCLUSION Findings in the current study provide detailed insights into the protective mechanism of a novel autophagy inducer,onjisaponin B,which is valuable for further investigation as a new candidate agent for modulating neurodegenerative disorders through the reduction of toxicity and clearance of mutant proteins in the cellular level.

Analysis of Volatile Components in Different Parts of Polygala japonica Houtt.

[Objectives] To analyze the volatile components in different parts of Polygala japonica Houtt. and compare the differences of these volatile components. [Methods] The volatile components in different parts of P. japonica Houtt. were analyzed by the headspace solid-phase microextraction( HS-SPME) combined with GC-MS,and the relative percentage of each component was determined by peak area normalization. [Results] Thirty kinds of volatile components were identified from the leaves and rhizomes of P. japonica Houtt.,mainly including olefins,aromatic hydrocarbons,alkanes and esters. [Conclusions] The volatile components in different parts of P. japonica Houtt. were different,and there may be difference in the medicinal value of volatile components in different parts,thus it is required to take an overall consideration of these differences in the development and utilization of P. japonica Houtt.

Effects of Polysaccharide from Polygala fallax on Apoptosis of B16 Melanoma

[Objectives]The paper was to study the effects of polysaccharide from Polygala fallax on apoptosis of B16 melanoma and its mechanism.[Methods]P.fallax polysaccharide was prepared by water extraction and alcohol precipitation method combined with Sevag method.The effects of different concentrations of P.fallax polysaccharide on B16 cell proliferation were detected by MTT assay.Apoptosis was detected by Hoechst 33258 staining assay and flow cytometry.The expressions of Bcl-2,Bax,Cleaved Caspase-3 mRNA were detected by Real-time PCR assay.The expressions of Bcl-2,Bax,Caspase-3,Caspase-8,Caspase-9,Cleaved Caspase-3,Cleaved Caspase-9,ERK,p-ERK,JNK,p-JNK proteins were detected by Western Blot assay.[Results]P.fallax polysaccharide inhibited the cell proliferation of B16 melanoma in a concentration-dependent manner(24 h IC 50=0.2133 mg/L;48 h IC 50=0.08489μg/mL).P.fallax polysaccharide up-regulated the expressions of Bax,Cleaved Caspase-3 mRNA and protein,down-regulated the expressions of Bcl-2 mRNA and protein,activated Caspase-3,Caspase-8 and Caspase-9,and decreased the levels of p-ERK/ERK and p-JNK/JNK.[Conclusions]P.fallax polysaccharide can inhibit the cell proliferation of B16 melanoma and induce their apoptosis,probably attributed to the regulation of MAPK,Bcl-2 and Caspase family signaling pathways.