Total Flavonoid Content, the Antioxidant Capacity, Fingerprinting and Quantitative Analysis of Fupenzi (<i>Rubus chingii</i>Hu.)

Objective: In this study, one of the objectives was to investigate the total flavonoid contents of Fupenzi (R. chingii Hu.) obtained from different regions of China and to evaluate their anatioxidant activities. And the second objective of this study was to develop a validated HPLC method for chromatographic fingerprints of the samples extracts of Fupenzi. Method: The total flavonoid contents were determined by a colorimetric method and the antioxidant activity was determined spectrophotometrically by DPPH and ABTS radical scavenging assays. The chromatographic fingerprint was developed by high-performance liquid chromatography coupled with diode array detection for the control of Fupenzi. Results: A significant correlation between antioxidant activity and the total flavonoid content was observed for the DPPH assay (r2 = 0.758, ρ = 0.004) and the ABTS assay (r2 = 0.788, ρ = 0.002). Under the optimized chromatographic conditions, the validated method was successfully applied to assessment of chemical fingerprinting of 12 batches of FPZ collected from different regions of China. Comparisons of the chromatograms showed that 15 characteristic peaks could be selected as markers for identification and evaluation of Fupenzi. In addition, the proposed method was also successfully applied to simultaneous determination of five compounds (including puerarin, rutin, hyperin, quercetin and kaempferol) in these samples. Conclusions: The qualitative and quantitative analysis described in this paper could be used for identification and evaluation of Fupenzi.

Chemical composition,pharmacological activity and development strategies of Rubus chingii:A review

Raspberries are used for both food and medicine,but it has not yet attracted widespread attention.In this paper,the chemical constituen of the original plant raspberry.R.chingii is one of the new“Zhe Bawei”medicinal materials selected in 2017.“Zhe Bawei”refers to eight kinds of genuine medicinal materials in Zhejiang Province.The chemical constituents,pharmacological effects,processing,and application of Rubus chingii Hu were reviewed to provide a reference for its further development.Relevant literature in recent years was collected in databases such as China Knowledge Network,Web of Science,Elsevier,PubMed,and X-Mol,using“raspberry”,“Rubus chingii”,“traditional use”,“chemical composition”,“pharmacology”,etc.as keywords individually or in combination.The summary of pharmacological activities shows that the relationship between the pharmacological activities of raspberry is still not deep enough.More in-depth research should be carried out in this direction to explore the mechanism of action of its active ingredients and provide effective reference for the further development of the raspberry industry.In the future,with the participation of more researchers,it is expected to develop innovative drugs based on raspberry for the treatment of diseases.

掌叶覆盆子HMGR基因家族的鉴定及表达分析

3-羟基-3-甲基戊二酰辅酶A还原酶(HMGR)是药用植物萜类产物骨架生物合成的重要酶之一。为了明确掌叶覆盆子HMGR蛋白的功能,使用生物信息学手段对RcHMGR家族成员的分类、基因结构、染色体定位、启动子信息等进行了系统分析,并比较了不同组织、果实不同成熟阶段及MeJA处理下RcHMGR基因的表达量。结果表明,RcHMGR基因家族共有7个成员,分布于3条染色体上,编码444~601 aa蛋白序列。RcHMGR蛋白分为4个亚家族,亚细胞定位于内质网,均含有HMGR保守功能域。RcHMGR家族基因的启动子上包含激素响应、光响应和低温响应等多种顺式元件。RcHMGR家族基因在掌叶覆盆子不同组织、果实不同成熟阶段和MeJA处理下的表达量均存在显著差异,其中RcHMGR5在果实中高丰度表达,并受到MeJA强烈诱导,与类胡萝卜素在掌叶覆盆子果实中的积累规律相一致,推测是类胡萝卜素合成途径的关键酶。上述结果为后续研究RcHMGR基因家族在萜类生物合成中的功能奠定了基础。

基于高通量测序分析覆盆子酵素自然发酵过程中的微生物多样性

为了解覆盆子酵素自然发酵过程中微生物结构组成变化。采用Illumina MiSeq高通量测序分析覆盆子酵素自然发酵前期(6 d),中期(16 d),后期(46 d)中细菌和真菌的群落结构组成及多样性差异。覆盆子酵素自然发酵过程的优势细菌门是变形菌门(Proteobacteria),葡萄糖杆菌属(Gluconobacter)是优势细菌属,相对丰度为95.9%、94.6%和28.2%。子囊菌门(Ascomycota)是优势真菌门,优势真菌属为汉逊酵母属(Hanseniaspora)和裂殖酵母属(Schizosaccharomyces),相对丰度为32.7%~52.4%和30.4%~44.8%。毕赤酵母属(Pichia)在发酵过程中相对丰度逐渐增加,发酵后期相对丰度达到31.2%。在覆盆子酵素自然发酵过程中,细菌的丰富度和多样性逐渐增加,真菌的丰富度逐渐减少,多样性逐渐增加。

覆盆子单宁富集组分消化稳定性及对肠道菌群的调节作用

单宁类化合物是覆盆子的主要活性成分,具有降血糖、降血脂、抗炎、延缓衰老及预防心脑血管疾病等功能。本实验从覆盆子中提取富集单宁,通过体外模拟消化模型探究覆盆子单宁富集组分消化前后单宁含量及抗氧化活性、α-葡萄糖苷酶抑制活性变化,通过体外厌氧发酵模型分析模拟消化对其肠道菌群调节作用的影响,最后采用超高效液相色谱-四极杆飞行时间串联质谱法(ultra-high performance liquid chromatography-quadrupole time offlight-tandem mass spectrometry,UPLC-ESI-QTOF-MS/MS)比较体外消化和酵解产物中单宁组成及其代谢产物含量变化情况。结果表明,在胃、肠消化过程中水解单宁含量均呈先增加后降低趋势,胃肠消化提升了覆盆子单宁富集组分的1,1-二苯基-2-三硝基苯肼自由基清除能力(半抑制质量浓度为29.15~39.65µg/mL),但对2,2’-联氮基-双(3-乙基苯并噻唑啉-6-磺酸)阳离子自由基清除能力无显著性影响(P>0.05),经肠道模拟消化后,样品的α-葡萄糖苷酶抑制能力提升了7.16~7.86倍。体外结肠发酵实验结果表明,覆盆子单宁显著增加了粪球菌属(Coprococcus)、丁酸弧菌(Anaerostipes)、瘤胃球菌属(Ruminococcus)、粪便杆菌属(Faecalibaterium)等有益菌的相对丰度,胃肠模拟消化降低其对双歧杆菌等有益菌的改善效果。UPLC-ESI-QTOF-MS/MS分析结果表明,胃、肠消化过程中,单宁代谢物含量均呈现先下降后上升的趋势,结肠发酵后样品中的尿石素A和尿石素B含量明显下降,而尿石素C和尿石素D未检测到。本研究可为覆盆子单宁稳态递送体系的构建,提高其体内的生物利用率提供理论依据。

基于UPLC-Q/TOF-MS研究广西甜茶在大鼠体内的入血成分

目的:采用超高效液相色谱-四级杆-飞行时间质谱(UPLC-Q/TOF-MS)法分析大鼠口服广西甜茶后的入血成分,为阐明其活性成分在机体转化和代谢情况提供基础。方法:超声法提取药物;大鼠灌胃给予广西甜茶提取物后,采用眼球后静脉丛取血;收集不同时间段的血浆,通过超高效液相色谱-四级杆-飞行时间质谱(UPLC-Q/TOF-MS)法,比较广西甜茶提取物、大鼠空白血浆、灌胃给予广西甜茶的大鼠血浆,初步鉴定和推测灌胃给予广西甜茶后大鼠血浆中的入血成分。结果:本实验共鉴定出50个原型入血成分,主要包括10个多酚类成分,13个黄酮类成分,20个二萜类成分,7个三萜类成分。结论:本实验初步阐明了广西甜茶中的入血成分,为其活性成分在机体转化和代谢情况提供基础,且为广西甜茶的后续开发利用提供了理论依据。

覆盆子叶化学成分的研究

目的研究覆盆子Rubus chingii Hu叶的化学成分。方法覆盆子叶水提取物采用硅胶、C 18中压反向柱、ODS、半制备HPLC进行分离纯化,根据理化性质及波谱数据鉴定所得化合物的结构。结果从中分离得到22个化合物,分别鉴定为异槲皮苷(1)、槲皮素-3-O-β-D-吡喃葡萄糖苷(2)、金丝桃苷(3)、(+)-异落叶松树脂醇(4)、山柰酚-3-O-β-D-吡喃葡萄糖苷(5)、山柰酚-3-O-β-D-吡喃半乳糖苷(6)、2,3-dihydro-7-methoxy-2-(4′-hydroxy-3′-methoxyphenyl)-3a-O-β-D-xylopyranosyloxymethyl-5-benzofuranpropanol(7)、鞣花酸(8)、山柰酚-3-O-β-D-半乳糖苷(9)、山柰酚-3-O-α-D-阿拉伯糖苷(10)、没食子酸(11)、15,18-二-O-β-D-吡喃葡萄糖-13(E)-对映-劳丹烷-7(8),13(14)-二烯-3β,15,18-三醇(12)、15,18-二-O-β-D-吡喃葡萄糖-13(E)-对映-劳丹烷-8(9),13(14)-二烯-3β,15,18-三醇(13)、覆盆子苷A(14)、goshonoside F5(15)、goshonoside F6(16)、dihydrobuddlenol(17)、(S)-3-hydroxy-β-ionone(18)、13(E)-对映-劳丹烷-8(17),13-二烯-3β,15,18-三醇(19)、icariol A2(20)、(+)-松脂醇(21)、gomojoside J(22)。结论化合物4、7、10、17~18、20~22为首次从覆盆子中分离得到,化合物1~3、5~6、8~9、11~14为首次从覆盆子叶中分离得到。

覆盆子乙醇提取物促进皮肤屏障修护的研究

使用乙醇对覆盆子进行了提取,乙酸乙酯萃取后,得到覆盆子水层提取物(样品号:ZS29)55.4 g,收率13.85%,多糖含量17.67%。使用得到的覆盆子水层提取物对角质形成细胞迁移、角蛋白K1和兜甲蛋白(LOR)表达进行了研究,结果表明,覆盆子水层提取物对角质形成细胞迁移没有明显促进作用,但对K1和LOR的基因表达有显著促进作用。覆盆子水层提取物可通过提升LOR、K1基因的表达量,促进细胞代谢,达到修护皮肤屏障功效。

不同产地覆盆子药材的指纹图谱研究

目的建立不同产地覆盆子药材指纹图谱评价体系。方法采用Agilent 5TC-C18色谱柱,柱温25℃,流动相为0.1%磷酸溶液-乙腈,梯度洗脱,流速1.0 mL/min,检测波长360 nm;采用中药色谱指纹图谱相似度评价系统(2012年版)计算相似度。结果9批覆盆子样品的指纹图谱共标定了13个共有色谱峰,相似度在0.933~0.992之间。结论建立的不同产地覆盆子高效液相色谱(HPLC)指纹图谱符合技术要求,为覆盆子的质量控制与资源开发提供了参考。

Protein tyrosine phosphatase 1B inhibitory activities of ursane-type triterpenes from Chinese raspberry, fruits of Rubus chingii

Protein tyrosine phosphatase 1 B(PTP1 B) has led to an intense interest in developing its inhibitors as anti-diabetes, anti-obesity and anti-cancer agents. The fruits of Rubus chingii(Chinese raspberry) were used as a kind of dietary traditional Chinese medicine. The methanolic extract of R. chingii fruits exhibited significant PTP1 B inhibitory activity. Further bioactivity-guided fractionation resulted in the isolation of three PTP1 B inhibitory ursane-type triterpenes: ursolic acid(1), 2-oxopomolic acid(2), and 2α, 19α-dihydroxy-3-oxo-urs-12-en-28-oic acid(3). Kinetics analyses revealed that 1 was a non-competitive PTP1 B inhibitor, and 2 and 3 were mixed type PTP1 B inhibitors. Compounds 1-3 and structurally related triterpenes(4-8) were further analyzed the structure-activity relationship, and were evaluated the inhibitory selectivity against four homologous protein tyrosine phosphatases(TCPTP, VHR, SHP-1 and SHP-2). Molecular docking simulations were also carried out, and the result indicated that 1, 3-acetoxyurs-12-ene-28-oic acid(5), and pomolic acid-3β-acetate(6) bound at the allosteric site including α3, α6, and α7 helix of PTP1 B.

Rubus chingii Hu: an overview of botany, traditional uses,phytochemistry, and pharmacology

Rubus chingii Hu, a member of the rosaceae family, is extensively distributed in China and Japan. Its unripe fruits(Fupenzi in Chinese) have a long history of use as an herbal tonic in traditional Chinese medicine for treating various diseases commonly associated with kidney deficiency, and they are still in use today. Phytochemical investigations on the fruits and leaves of R. chingii indicate the presence of terpenoids, flavonoids, steroids, alkaloids, phenylpropanoids, phenolics, and organic acids. Extracts or active substances from this plant are reported to have various pharmacological properties, including antioxidant, anti-inflammatory, antitumor, antifungal, antithrombotic, antiosteoporotic, hypoglycemic, and central nervous system-regulating effects. This review provides up-to-date information on the botanical characterizations, traditional usages, chemical constituents, pharmacological activities, toxicity,and quality control of R. chingii. Possible directions for future research are also briefly proposed. This review aims to supply fundamental data for the further study of R. chingii and contribute to the development of its clinical use.

覆盆子及其易混品桉叶悬钩子的鉴别

目的探究覆盆子及其易混品桉叶悬钩子的生药特征,为有效区分两者提供依据。方法采用体式显微镜对两者聚合果和小核果的构造特征进行观察,并通过显微镜对两者横切面和粉末显微特征展开比较。结果覆盆子宿萼短于聚合果,小核果顶端密被茸毛表面有横纹,非腺毛短而平直;桉叶悬钩子无宿萼基部中心深凹,小核果表面无横纹全身密被茸毛,非腺毛长而弯曲。结论本法可用于鉴别覆盆子与桉叶悬钩子,有效避免市场混用误用。

响应面优化覆盆子多糖滴丸的制备工艺研究

为优化保健食品覆盆子多糖滴丸的最佳制备工艺条件,并对产品质量进行评价,本文在单因素实验的基础上通过响应面法设计考察了基料比、料液温度、滴距3个因素对覆盆子多糖滴丸感官评价的影响,以获得滴丸最佳的制备工艺。结果表明,在基质与多糖质量比为2.6∶1(g∶g),料液温度为82℃,滴距为10cm时,得到的3批滴丸感官评分分别为92分、93分和92分,重量差异及溶散时限均符合相关标准,说明该制备工艺操作简便且制得产品的感官品质良好。

Simultaneous Determination of Eight Constituents in Fruits of Rubus chingii by UPLC

Objective To develop a simple, efficient, and reliable method for routine quantitative analysis of main constituents presented in the fruits of Rubus chingii, which is widely used in Chinese materia medica （CMM）, known as Fupenzi （FPZ） in Chinese. Methods An ultra performance liquid chromatography-photo diode array （UPLC-PDA） system was employed for simultaneous quantification of eight compounds, i. e. adenosine, gallic acid, brevifolin carboxylic acid, ethyl gallate, ellagic acid, kaempferol-3-O-rutinoside, kaempferol-3-O-^-D-glucopyranoside, and tiliroside. The chromatographic analysis was performed on a C18 column using a gradient elution of acetonitrile -0.1% formic acid aqueous solution within a runtime of 25 min. Results All calibration curves were linear （R2〉 0.9997） over the tested ranges. The intra- and inter-day precisions as determined from sample solutions were both less than 2.45% and 2.78%, respectively. The average recoveries for the eight constituents ranged from 94.77% to 101.35% with RSD ≤ 4.41%. The newly-developed method was applied to the quality assessment of various R. chingii samples, including both ripe and unripe fruits of R. ching/i from different habitats. Conclusion The relative levels of the investigated compounds vary remarkably in the fruits of R. chingiicollected from different habitats. As only two of the eight compounds, adenosine and ellagic acid, are determined in the ripe fruits of R. chingii, the results may explain the reason why only the unripe fruits can be used in CMM.

响应面法优化盐覆盆子有效成分鞣花酸的提取工艺

目的:采用星点设计-响应面法优化盐覆盆子有效成分鞣花酸的提取工艺。方法:建立HPLC测定鞣花酸含量,流动相为乙腈-0.2%磷酸溶液(15:85);检测波长为254 nm,流速为1.0 ml/min,柱温为30℃;采用星点设计-响应面法对甲醇浓度、提取时间、液料比3个因素进行考察,优选盐覆盆子有效成分鞣花酸的提取工艺。结果:优选所得的盐覆盆子中鞣花酸提取工艺为甲醇浓度73%、提取时间63 min、液料比60∶1,此条件下鞣花酸含量为0.353%。结论:本试验优选所得的提取工艺高效稳定、操作简易、可用于盐覆盆子有效成分鞣花酸的提取。

掌叶覆盆子的化学成分研究

目的对蔷薇科悬钩子属植物掌叶覆盆子Rubuschingii的果实进行化学成分研究。方法通过反复硅胶柱色谱、SephadexLH20和重结晶的方法分离纯化,根据化合物的理化性质和波谱数据鉴定结构。结果分离得到11个已知化合物,分别为齐墩果酸(Ⅰ),乌苏酸(Ⅱ),2α羟基齐墩果酸(Ⅲ),2α羟基乌苏酸(Ⅳ),arjunicacid(Ⅴ),hexacosylpcoumarate(Ⅵ),4′,5,7三羟基黄酮醇3OβD(6″对羟基桂皮酰基)葡萄糖苷(tiliroside)(Ⅶ),硬脂酸(Ⅷ),三十二烷酸(Ⅸ),β谷甾醇(Ⅹ),胡萝卜苷(Ⅺ)。结论除了化合物Ⅶ和Ⅹ,其他化合物均为首次从本植物中分离得到。

覆盆子的化学成分研究

从覆盆子中分离得到13个化合物,通过波谱数据和理化性质分别鉴定为:β-谷甾醇(1)、7α-羟基-3β-谷甾醇(2)、胡萝卜苷(3)、熊果酸(4)、对羟基苯乙酸(5)、水杨酸(6)、腺苷(7)、1-氧-1,2-二氢异喹啉-4-羧酸(8)、香橙素(9)、金丝桃苷(10)、ligballinol(11)、阿魏酸二十六醇酯(12)、香豆酸二十四醇酯(13)。化合物10为首次从该植物中分离得到,化合物2、5、7、9、12、13为首次从悬钩子属植物中分离得到。

掌叶覆盆子的化学成分研究

目的对蔷薇科悬钩子属植物掌叶覆盆子(Rubus chingii Hu)的果实进行化学成分研究。方法对掌叶覆盆子果实的甲醇提取物进行柱色谱,根据化合物的理化性质和波谱数据鉴定结构。结果分离得到7个已知化合物,分别为山柰酚(Ⅰ)、槲皮素(Ⅱ)、山柰酚-3-O-β-D-吡喃葡萄糖苷(Ⅲ)、槲皮素-3-O-β-D-吡喃葡萄糖苷(Ⅳ)、山柰酚-3-O-β-D-吡喃葡萄糖醛酸甲酯(Ⅴ)、对羟基苯甲酸(Ⅵ)、三十一烷(Ⅶ)。结论除了化合物Ⅵ,其他化合物均为首次从本植物中分离得到。

湖北掌叶覆盆子叶降血脂作用的动物实验和人群研究

目的:探讨湖北掌叶覆盆子叶对高脂模型动物和高血脂人群的降血脂作用。方法:采用预防性大鼠脂质代谢紊乱模型,50只大鼠根据血清总胆固醇(TC)、甘油三脂(TG)、高密度脂蛋白胆固醇(HDL-C)水平随机分成对照组、高脂模型组和掌叶覆盆子叶0.5、1.0、2.0g/kgbw剂量组,对照组饲喂基础饲粮,高脂模型组和掌叶覆盆子叶组喂饲高脂饲粮,掌叶覆盆子叶各剂量组灌胃给予水提取物,对照组和模型组给予蒸馏水,连续30d后,取血,分离血清,检测TC、TG和HDL-C水平。选择112高血脂症人群,随机盲法按血脂水平、年龄、性别、饮食等因素均衡分为试验组和对照组,试验组饮用掌叶覆盆子叶袋泡茶每日3次,每次2g,对照组不给予任何干预措施,不改变日常生活和饮食习惯,连续30d,取血,分离血清,检测TC、TG和HDL-C水平。结果:动物实验中,覆盆子叶1.0、2.0g/kgbw剂量组大鼠血清TC、TG显著低于高脂模型组(p<0.05)。人群试验中,试验组试验结束时的TC、TG与试验前以及对照比较,差异均有显著性(p<0.05);试验组降血脂的总有效率为80.4%。结论:湖北掌叶覆盆子叶有显著的降血脂作用。

施肥对掌叶覆盆子叶绿素荧光参数和生理特性的影响

【目的】探讨掌叶覆盆子Rubus chingii的需肥特性,为其优质高产栽培提供理论参考和指导。【方法】以掌叶覆盆子为材料,通过盆栽试验研究不同肥料的低(L)、中(M)和高(H)水平处理对掌叶覆盆子的叶绿素、株高、叶绿素荧光参数及相关生理指标的影响。【结果】随着施肥量的增加,掌叶覆盆子的叶绿素含量、株高、叶绿素荧光参数和生理指标均呈上升趋势。NPK肥L、M和H条件下叶绿素含量增幅稳定,依次增加36.3%、41.3%和53.4%,且各水平间差异显著;复合肥能迅速提高叶绿素含量,但各施肥水平无显著性差异;有机肥在M与H水平,叶绿素含量差异不明显。NPK肥与株高存在极显著相关性,且长势最优,L、M和H水平依次增高40.5%、50.2%和64.4%;有机肥次之,依次增高18.3%、44.0%和56.2%;复合肥稍弱,依次增高16.1%、20.3%和40.7%。NPK肥在各施肥量下,F_v/F_m、Φ_(ps)Ⅱ、q P均为最高水平,复合肥和有机肥与4个叶绿素荧光参数存在显著相关性,但复合肥处理下,q P在3种肥料中处于最低水平,有机肥处理下的F_v/F_m和Φ_(ps)Ⅱ最低。NPK肥与4种生理指标存在显著相关性,各施肥处理下可溶性蛋白、可溶性糖、脯氨酸的含量达最高水平,丙二醛含量最低;复合肥在H水平,可溶性蛋白、可溶性糖含量增幅分别为8.4%和2.1%,处于最低水平;有机肥在H水平处理下丙二醛含量增幅36.8%,达到最高水平。【结论】随着施肥量的递增,NPK肥处理下的掌叶覆盆子叶绿素含量和株高差异显著,叶绿素荧光参数和生理指标也表现出优良的特性。