Neuroprotective effects of total saponins from Rubus parvifolius L. on cerebral ischemia/reperfusion injury in rats

This study examines the neuroprotective effects and mechanisms of action of total saponins from Rubus parvifolius L. （TSRP） on focal cerebral ischemia and reperfusion injury in rats. Focal cerebral ischemia and reperfusion injury was performed in rats using the suture method. The results indicate that intragastric injection of TSRP, at 5, 10 and 20 mg/kg, could decrease neurological impairment, reduce cerebral infarct volume, diminish pathological changes, and significantly inhibit the apoptosis of neurons surrounding the ischemic area. In addition, TSRP upregulated the expression of the anti-apoptotic factor Bcl-2, at the protein and mRNA levels, and it downregulated the expression of the pro-apoptotic factor Bax, at the protein and mRNA levels. These findings indicate that TSRP protects against cerebral ischemia/reperfusion injury, and that it may do so by regulating the expression of Bcl-2 and Bax.

Total saponins in Rubus parvifolius L.induce lymphoma cells apoptosis through upregulated Bax/Fas and downregulated Bcl-2 in vivo and in vitro

Purpose:To investigate the effect of total saponins of Rubus parvifolius L.(TSRP)on lymphoma Raji cells and further discuss its mechanism.Methods:The model of nude mice bearing Raji cells was established,the volume,weight and inhibition rate of the transplanted tumor were analyzed and compared after different concentrations of TSRP treatment.Cell apoptosis and expression of Bcl-2,Bax,Fas proteins were detected by TUNEL and immunohistochemiscal method respectively.Effects of TSRP on cell proliferation were tested with MTT assay in vitro.Cell apoptosis and expression of Caspase-9,Caspase-3,Bcl-2,Bax and Fas proteins were tested with DAPI staining and Western blot.Results:TSRP significantly reduced the volume and tumor weight of Raji subcutaneous transplanted tumor and induced the apoptosis of Raji cells in vivo.The tumor inhibition rate of high-dose(100 mg/kg)TSRP is 90.84%.The TUNEL test results show that the fluorescence intensity of the tumor issue treated with TSRP is significantly improved.Compared with the control group,the fluorescence intensity of high-concentration TSRP is 82.43 ± 7.81,which is significantly different(P<0.001).The results of immunohistochemistry test showed that the Bcl-2 expression of Raji cell treated with TSRP is obviously reduced,and Bax expression is obviously increased.Meanwhile,compared with that of control group,Fas expression is obviously reduced.MTT assay showed that TSRP can significantly inhibit proliferation of Raji cells with dose dependence.The inhibition rate of 400 μg/mL TSRP is 53.46 ± 4.90%(P<0.001).DAPI staining results showed that TSRP can significantly induce cell apoptosis.According to Western blot results,it is found that TSRP can significantly inhibit activity of Bcl-2 and increase Bax expression,and TSRP can also inhibit Fas expression.Meanwhile,expression of Caspase-9 and Caspase-3 is also increased.Conclusion:TSRP could inhibit the proliferation of lymphoma via induction of apoptosis in a time and dose-dependent manner.Apoptotic signaling induced by TSRP was characterized by up-regulating Bax,Fas and Caspase-8 protein expression,and down-regulating of Bcl-2 protein expression.

Total Saponins of Rubus Parvifolius L.Exhibited Anti-Leukemia Effect in vivo through STAT3 and eIF4E Signaling Pathways

Objective： To investigate the anti-leukemia effect of total saponins of Rubus parvifo/ius L. （TSRP） on K562 cell xenografts in nude mice and the mechanisms of action. Methods： The K562 cell xenografts in nude mice were established, and then randomly divided into 5 groups, the control group, the cytosine arabinoside group（Am-c） and 3 TSRP groups （20, 40 and 100 mg/kg）. The tumor volume and mass of each group of nude mice were measured and the anti-tumor rates of TSRP were calculated subsequently. The apoptosis status of tumor cells was detected by hematoxylin-eosin （HE） and terminal dexynucleotidyl transferase （TdT）-mediated dUTP nick end labeling （TUNEL） staining analysis. Finally, the activities of apoptosis related signaling of signal transducer and activator of transcription 3 （STAT3）, eukaryotic initiation factor 4E （eIF4E） and B-cell lymphoma-2 （bcl-2） were determined with immunohistochemistry tests. Results： Subcutaneous injection of K562 cells induced tumor formation in nude mice, and the TSRP treated group showed a significant inhibitory effect on tumor formation. The nude mice treated with TSRP showed a significant decrease in tumor growth rate and tumor weight in comparison to the control group （all P〈0.05）. The HE staining and TUNEL assay showed that TSRP induced cell death by apoptosis. The immunohistochemical assay showed down-regulation of the bcl-2 gene in the TSRP treated cells. The phosphorylation levels of elF4E and STAT3 were decreased obviously after the treatment of TSRP. Conclusion： TSRP had an excellent tumor-suppressing effect on K562 cells in the nude mice xenograft model, suggesting that TSPR can be developed as a promising anti-chronic myeloide leukemia drug.

Rubus Parvifolius L.Inhibited the Growth of Leukemia K562 Cells in Vitro and in Vivo

Objective：To determine the antiproliferative activity of Rubus parvifolius L.（RP）extract,its medicinal serum and RP total saponins（RPTS）against K562 cells in vitro and in vivo.Methods：Nude mice models bearing leukemia tumors were treated with different concentrations of RP extract.The size,weight and histopathological change of leukemic tumors were determined.Semi-solid agar culture and methylthiazolyl tetrazolium（MTT）assay were used to determine in vitro the inhibition of colony formation and proliferation of K562 cells respectively by different concentrations of RP medicinal serum and RPTS.Results：RP extract had a tumor inhibition rate of 84.8%when administered to mice at a dose of 1.0 g/day of crude RP root equivalent.Semi-solid agar culture of K562cells in the presence of 20%（v/v）of RP medicinal serum and 150 mg/L RPTS demonstrated a 50.8%and 100%inhibition of the colony forming unit（CFU）-K562,respectively.The same doses of RP medicinal serum and RPTS showed a proliferation inhibition of 31.4%and 86.3%,respectively against K562 cells in MTT assay.Conclusion：RP extract and RPTS show effective antiproliferative activity against myeloid leukemia cells in vitro and in vivo.

Evaluation of the hepatoprotective and antioxidant activities of Rubus parvifolius L.

The hepatoprotective and antioxidant activities of the n-butanol extract of Rubus parvifolius L. （RPL）, a widely used medicinal plant, were evaluated. Results demonstrated that RPL extract possessed pronounced hepatoprotective effects against carbon tetrachloride （CCI4）-induced hepatic injury in mice, which was at least partially attributed to its strong antioxidant capacity. Treatment with RPL extract markedly attenuated the increases in serum alanine ami- notransferase （ALT） and aspartate aminotransferase （AST） levels caused by CCI4 intoxication. It also significantly prevented the decrease in superoxide dismutase （SOD） activity and the increase in malondialdehyde （MDA） content of liver tissue. Meanwhile, histopathological changes of hepatic damage were also remarkably ameliorated. Phytochemical analysis based on high-performance liquid chromatography/tandem mass spectrometry （HPLC-MS/MS） revealed the presence of various phenolic compounds, including caffeic acid conjugates, ellagic acid glycosides, and flavonol glycosides, which might be responsible for the hepatoprotective and antioxidant activities of RPL.

红树莓(Rubus idaeus L.)雌雄配子体发育的研究

红树莓雄蕊多数,排成3轮、同一轮花药发育基本同步,各轮间发育不同步,自外向内顺序发育。最早的造孢细胞和小孢子母细胞于5月下旬产生。小孢子母细胞的胞质分裂为同时型,四分体呈四面体型,成熟花粉为2—细胞。雌蕊亦多数离生,整齐排列在膨大的花托上。每一子房内有2枚胚珠,但后期仅1枚正常发育。胚珠倒生、单珠被、厚珠心。胚囊发育属蓼型,但大孢子母细胞、功能大孢子常有异常现象。

Isolation and Crystal Structure of Ent-kaurane Diterpenes from Rubus corchorifolius L. f.

In this study, ent-Kaurane-3β,16β,17-trioi （1） and ent-kaurane-2a,16β,17-triol （2）, were isolated from the leaves ofRubus corchorifolius L. f. Their structures were elucidated based on extensive spectroscopic analysis. NMR experiments identified the two compounds and X-ray crystallographic analysis confirmed their crystal structures. The crystal of 1 belongs to monoclinic with space group P21 and a = 10.7641（3）, b = 7.2789（3）, c = 11.7862（4） A, β = 95.275（3）°, V = 919.55（6） A3, Z = 2, C20H34O4, Mr = 322.49 g/mol, Dc = 1.230 g/m3, F（000） = 376, 2 = 1.54178 A,μ = 0.661 mm-1, R = 0.0319, and wR = 0.0811 for 10889 observed reflections （I〉 2σ（I））. The crystal of 2 is classified as orthorhombic with space group P212121 and a = 10.7641（3）, b = 12.72224（19）, c = 21.3929（3） A, V= 1748.94（4） A3, Z= 4, C20H3404, Mr = 322.47, De = 1.225 Mg/m3 F（000） = 712, λ = 1.54184 A,μ = 0.625 mm-1, R = 0.0303 and wR = 0.0759 for 10470 observed reflections （1〉 2σ（I））. Meanwhile, the compound revealed low inhibitory activities toward HepG2, MCF-7, and SCG-7901 cells.

Phenol Compounds of Blackberry （Rubus caucasicus Focke and Rubus anatolicus L.） Fruit and Leaf

It is investigated of the effect of plant blackberry （Rubus caucasicus Focke and Rubus anatolicus L.） flower, fruit and leaf content of phenol compounds qualitative and quantitative HPLC （high-pressure liquid chromatography） and spectral methods. HPLC analysis revealed that the fetus was a major of both species, cyanidin-3-glucoside, and it reached full maturity period 1,493.54 ± 44.81 and 1,636.58 ±49.10 mg/kg （cyanidin-3-O-glucoside chloride equivalents） calculated on the dry weight. In mature fetus flavonols is （884.8±26.54）-（979.0 ± 29.37） mg/kg （rutin equivalents） calculated on the dry weight, catechins （21.2± 0.64）-（25.01 ± 0.7537） mg/kg （（＋） catechin equivalents） calculated on the dry weight, leucoantocyanins （48.8 ±1.46）-（52.1± 1.56） mg/kg （cyanidin equivalents） calculated on the dry weight. In this regard, there was a slight difference between the species. Most of the leaf flavonols were presented 3.7 ± 0.111 g/kg. Phenol carbonic acids--the leaves of 13.4 ± 0.402 mg/kg calculated on the dry weight, the flowers of 8.8± 0.264 mg/kg, calculated on the dry weight. Full maturity period, the number of phenol carbonic acids cent is （3,707.8 ± 111.23）-（4,287.5 ± 128.63） mg/kg, （caffeic acid equivalents） is calculated on the dry weight of the unit. Also been determined HPLC following the method of compounds is： cyanidin-3-glucoside, quercetin-3-rutinoside, quercetin-3-glucoside, ellagic acid, gallic acid and chlorogenic acid.

茅莓总皂苷联合化疗药通过Fas途径协同诱导白血病HL-60细胞凋亡研究

目的 :探讨茅莓总皂苷(TSRP)抑制HL-60细胞及分别与化疗药高三尖杉酯碱(Hom)、阿糖胞苷(Ara-c)的协同作用,并探讨其协同作用的可能机制。方法 :MTT法检测不同浓度的TSRP与化疗药联用对HL-60细胞增殖影响;DAPI染色法观察一定浓度下TSRP联合化疗药诱导HL-60细胞凋亡的形态变化;Annexin V-FITC/PI检测不同浓度的TSRP与化疗药联用对HL-60细胞凋亡情况;RT-PCR检测TSRP联合化疗药干预HL-60白血病细胞后Fas和Caspase-3 mRNA表达。以上实验均用药物相互作用指数(CDI)评价药物协同作用。结果:MTT显示在一定浓度范围内TSRP和化疗药有显著的协同作用;DAPI染色法和Annexin V-FITC/PI法检测到一定浓度范围内TSRP联合化疗药后显示明显的凋亡,CDI均<1,具有协同作用;RT-PCR检测到在一定浓度范围内TSRP联合化疗较单用TSRP的FasmRNA表达明显增加和Caspase-3 mRNA表达明显减少,CDI均<1,具有协同作用。结论:TSRP分别和化疗药Hom或Ara-c联用对HL-60细胞有明显的协同增殖抑制和诱导凋亡作用,可能是通过Fas-Caspase途径增强凋亡效应,从而增加化疗药物的敏感性,具有良好的临床应用前景。

茅莓总皂苷诱导HL-60白血病细胞凋亡机制研究

目的:明确茅莓总皂苷(TSRP)在体外对HL-60细胞抑制增殖和诱导凋亡作用。方法:MTT法检测不同浓度TSRP对HL-60细胞的增殖抑制;使用DAPI和Annexin V-FITC/PI法观察TSRP诱导凋亡作用;RT-PCR法检测TSRP对HL-60细胞Bcl-2、Bax、Caspase-3、Caspase-9和Fas mRNA表达的影响。结果:MTT显示TSRP在浓度200、400、800 mg/L对HL-60细胞生长抑制呈剂量依赖性,浓度800mg/L时吸光度为(0.232±0.030),与空白对照组比较差异显著(P<0.01)。DAPI显示TSRP在一定浓度内干预HL-60细胞后出现明显凋亡现象;Annexin V-FITC/PI凋亡检测显示经不同浓度TSRP处理后,HL-60细胞的早期凋亡率明显增加;RT-PCR检测到在一定浓度范围内,TSRP显著抑制细胞Bcl-2和Fas mRNA表达,同时Bax、Caspase-9和Caspase-3 mRNA表达明显增加,以上均呈剂量依赖性。结论:在体外TSRP是通过Bcl-2和Fas途径诱导HL-60细胞凋亡,具有较好的临床应用前景。

NaCl与NaHCO3单盐胁迫下秋萍红树莓的光合及生理指标响应

【目的】研究红树莓品种秋萍对盐碱化土壤栽培的适应力,为该品种的建园选址和生产提供参考。【方法】以秋萍红树莓组培苗为材料,盆栽培养一段时间后,用不同浓度的中性盐(NaCl)和碱性盐(NaHCO3)进行单盐胁迫试验,再测定其光合特性以及生理指标。【结果】低浓度(30~60 mmol/L)盐胁迫处理对秋萍生长有促进作用,主要表现为净光合速率和气孔导度增大、色素含量增加、落叶减少以及SOD和POD活性降低;中等浓度(90~120 mmol/L)盐胁迫下,秋萍植株虽然能生长,但呈现生理干旱、光合受到抑制、气孔导度减小、CAT活性升高和脯氨酸含量增加等现象;高浓度(150~180 mmol/L)胁迫下,秋萍的光合受到强烈抑制,叶片色素含量降低,保护酶活性降低,渗透调节物质含量减少,180 mmol/L NaCl胁迫下植株死亡。【结论】秋萍对NaHCO3的耐性较NaCl强,能够适应较大浓度范围的盐碱环境,且轻度盐碱胁迫可促进秋萍生长。

Total Phenolic Compounds, Carotenoids and <i>In Vitro</i>Antioxidant Activity of Three Traditional Indigenous Medicinal Plants of Saskatchewan, Canada

Medicinal plants are an important component in Indigenous cultures. Aralia nudicaulis L., Rubus idaeus L., and Rosa arkansana Porter were analyzed for total phenolic compounds, carotenoids and antioxidant activity by DPPH (2,2-diphenyl-1-picrylhydrazyl), FRAP (ferric-reducing antioxidant power), and ABTS (2,2-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid). The samples were harvested in Saskatchewan, Canada, with the help of an Indigenous Traditional Knowledge Keeper and the analyses were performed by spectrophotometry. The results showed that total phenolic compounds amount ranged from 0.08 to 0.88 mg GAE/mg d.w. and the total carotenoid contents ranged from 0.03 to 1.26 mg/g d.w. The in vitro antioxidant activity ranged from 0.02 to 0.71 μmol TE/mg d.w. by DPPH, from 0.05 to 2.53 μmol TE/mg d.w. by FRAP, and from 0.04 to 1.06 μmol TE/mg d.w. by ABTS. Rosa arkansana leaves stood out with higher amounts of total phenolic compounds (TPC) (0.88 ± 0.02 mg GAE/g d.w.), carotenoids (TC) (1.26 ± 0.03 mg/g d.w.) and antioxidant activity (TAA) by DPPH (0.71 ± 0.01 μmol TE/mg d.w.), ABTS (1.06 ± 0.01 μmol TE/mg d.w.) and FRAP (2.32 ± 0.01 μmol TE/mg d.w.), with the same amount of Rubus idaeus belowground (2.53 ± 0.01 μmol TE/mg d.w.) in last technique (2.32 ± 0.01 μmol TE/mg d.w.). The first principal component describes 83.88% of the total variance and all the variables have high influence on this component (factor loadings: T = 0.976, TC = 0.735, TAA by DPPH = 0.955, FRAP = 0.894 and ABTS = 0.996), demonstrating that these samples do not have large dissimilarity. The second principal component represents 13.64% of the total variance, and the TC is the dominant variable on the second principal component (0.658). Aralia nudicaulis, Rubus idaeus, and Rosa arkansana had interesting amounts of total phenolic compounds, total carotenoids and in vitro antioxidant activity. Rosa arkansana leaves and Rubus idaeus have the highest amount of total phenolic compounds and antioxidant activity in this study. Rosa arkansana leaves are also a good source of carotenoids, and so, they have great potential health benefits and use in industry as a source of bioactive compounds with high antioxidant activity. This study enriches the literature on medicinal plants used by Indigenous people of Saskatchewan and surrounding Canada. More studies are necessary to identify its applications, security and to assess which compounds generate the benefits reported by Traditional Knowledge Keepers.

中国西南地区悬钩子属植物资源研究

悬钩子属植物具有很高的经济价值,目前认为中国西南地区是悬钩子属植物的分布中心;然而,尚不清楚该区域悬钩子属野生植物资源的分布现状。本研究通过野外考察、查阅标本和文献的方法,对中国西南地区的悬钩子属植物资源进行调查,同时对西南地区悬钩子属植物资源的收集、保存及有药用价值的植物进行初步探讨。结果发现:中国西南地区有悬钩子属植物142种、69变种,占全国悬钩子属的68.27%,其中特有种47种、40变种;西南地区的悬钩子资源有草本8种、2变种,灌木134种、67变种;悬钩子果实颜色多为黄色、红色或黑色,以小核果集中生于花托上形成聚合果,多浆或干燥,果实多为近球形或球形,直径多为1cm左右。据报道有药用价值的有22种、1变种;适合在滇西北高海拔地区作为育种材料或引种驯化的有5种;这些悬钩子属植物资源分布在海拔100~4300 m。本文较为系统地介绍了中国西南地区悬钩子属植物资源的分布区、海拔、生活型、生境类型以及植物学特性和遗传多样性,为悬钩子属植物资源的开发和遗传育种提供了基础数据。

采后常温储存红树莓病原菌的分离与鉴定

红树莓采后常温贮藏过程中极易腐烂变质,严重影响红树莓果实的品质。本研究以云南种植的西班牙品种Adilita红树莓果实为试材,用常规组织分离法和回接试验,对采后常温贮藏过程中自然发病果实表面的病原菌进行分离与筛选,采用传统真菌形态学显微观察,结合r DNA-ITS序列分析方法鉴定,明确红树莓腐烂过程中的病原菌种类和病原菌的系统进化关系。结果表明:从自然发病腐烂的样本红树莓果实中分离得到6株病原真菌,分别为1株木霉属(Trichoderma sp.),2株镰刀菌属(Fusarium sp.),2株青霉属(Penicillium sp.)和1株曲霉属(Aspergillus sp.)。2株镰刀菌属(Fusarium sp.)和2株青霉属(Penicillium sp.)分别有两个不同种的病原菌。相关研究结果对红树莓采后病害和防治措施提供依据,为后期红树莓的保鲜奠定理论基础。

红树莓罐头加工工艺研究

红树莓是一种深受消费者喜爱的果实,但果实酸味突出,且部分消费者无法接受其口感。本文以红树莓果实为研究对象,制作红树莓罐头,通过单因素试验和正交试验对红树莓罐头加工工艺进行优化。结果表明,红树莓罐头最佳工艺为红树莓添加量27%、银耳添加量6%、糖水浓度10%、氯化钙溶液浓度0.7%、氯化钙溶液浸泡20 min,该条件下红树莓罐头的感官评分最高。

Isolation and Chemotaxonomic Significance of Chemical Constituents from Rubus parvifolius

Objective To study the chemical constituents from the roots of Rubus parvifolius. Methods The chemical constituents were extracted and purified by silica gel column chromatography. NMR spectra were used for structural identification. Results Phytochemical study on the roots of R. parvifolius led to the isolation of one ceramide （1）, two anthraquinones （2 and 3）, four triterpenoids （4-7）, two flavonoids （8 and 9）, one fatty acid ester （10）, and two sterols （1 1 and 12）. Conclusion Compound 1 is isolated from the plants of family Rosaceae for the first time, and compounds 2-5 are isolated from genus Rubus for the first time. Though R. parvifolius shares the same major chemical types （triterpenoid, flavonoid, and anthraquinone） with those of R. alceaefolius, a substituent of R. parvifolius, their individual constituents are different. In addition, R. parvifolius contains ceramide （1） with high concentration, while caffeoylquinic acid reported in R. alceaefolius has not been found in R. parvifolius. Furthermore, the results from our phytochemical study are consistent with the DNA phylogenic relationship between R. parvifolius and R. alceaefolius （two separated subgenera）, suggesting that the substitution of the former species with the latter one in folk medicine might not be suitable.

Triterpenoids from the roots of Rubus parvifolius

Two new oleanane-type triterpenoids, parvifolactone A(1) and rubuside P(2), together with 11 known triterpenoids, fupenzic acid(3), 18,19-seco,2α,3α-dihydroxyl-19-oxo-urs-11,13(18)-dien-28-oic acid(4), euscaphic acid(5), maslinic acid(6), 1β-hydroxyeuscaphic acid(7), 2α,3α,19α,23-tetrahydroxyolean-12-en-28-oic acid(8), 2α,3β,19α,23-tetrahydroxyurs-12-en-28-oic acid(9), glucosyl pinfaensate(10), rubuside J(11), 2α,3α,19α,23-tetrahydroxyurs-12-en-24,28-dioic acid(12), and 2α,3β,19α- trihydroxyurs-12-en-23,28-dioic acid(13), were isolated from the roots of Rubus parvifolius.

茅莓鉴别与空心莓组ITS序列分析

目的:通过对茅莓(Rubus parvifolius L.)的鉴别,为茅莓的开发利用,质量标准制定提供理论依据。方法:采用性状鉴定、显微鉴定、薄层色谱鉴定及分子鉴定进行研究,对空心组植物ITS序列进行分析。结果:茅莓根和茎均有草酸钙簇晶、草酸钙方晶和具缘纹孔导管;叶中有两种非腺毛,一种棒状腺毛。薄层色谱可以有效鉴别茅莓。内部转录间隔区(ITS)序列作为DNA条形码,可以将52种空心莓组植物区分,得到与传统分类不同的分组,遗传距离推测绒毛叶亚组和柔毛叶亚组可能为其他亚组的祖先。结论:该结果可为茅莓及其同组植物的进一步开发利用提供参考依据。

茅莓化学成分的分离鉴定

目的研究茅莓RubusparvifoliusL的化学成分。方法采用柱色谱和制备型高效液相色谱进行分离纯化,通过理化方法和光谱分析鉴定化合物结构。结果从茅莓的乙醇提取物中分得4个三萜成分和1个二糖成分,分别鉴定为suavissimosideR1(1),nigaichigosideF1(2),camelliageninA(3),camelliageninC(4)和sucrose(5)。结论化合物3～5均为首次从该植物中分得。

茅莓化学成分研究

目的:研究茅莓的化学成分。方法:利用硅胶柱层析色谱进行分离纯化,通过波谱分析鉴定化合物结构。结果:从茅莓中分离得到了7个化合物,分别是:β-谷甾醇(Ⅰ)、月桂酸(Ⅱ)、邻硝基苯酚(Ⅲ)、β-胡萝卜苷(Ⅳ)、蔷薇酸(Ⅴ)、3β,16α2,2α,28-四羟基齐墩果-12-烯(Ⅵ)和(+)-儿茶素(Ⅶ)。结论:其中,化合物Ⅲ、Ⅶ为首次从该植物中分离得到。