Neuropharmacological Characterization of Extracts from Rhodiola rosea, Oenothera paradoxa and Paullinia cupana in Comparison to Caffeine

To find possible therapeutic applications involving the Central Nervous System (CNS) for herbals is a major challenge during functional food and drug discovery and development programmes. Despite the availability of numerous in vitro and in vivo tests, there is no single agreed screening procedure for pharmacological testing of herbal extracts with anticipated CNS activity. Experience gained from more than 25 years of testing has shown that two models give reasonably reliable orientation for future CNS applications: construction of an electropharmacogram based on wireless recording of field potentials from the depth of the brain of freely moving rats (Tele-Stereo-EEG) and recording of the population spike produced by pyramidal cells from hippocampal slices in vitro. A combination of these two methods has now been used to characterize the pharmacological profile of extracts from Rhodiola rosea root, Oenothera paradoxa seeds and Paullinia cupana seeds. Spectral analysis of field potentials revealed attenuation of alpha2 and beta1 waves was common for all extracts. According to previous studies, this is interpreted as activation of the dopaminergic and glutamatergic transmission. In addition, Oenothera and Rhodiola extracts attenuated delta and theta power, probably related to interference with the cholinergic and norepinephrinergic transmission, respectively. Using discriminant analysis for comparison with reference pharmaceutical and botanical drugs, Rhodiola projected near the position of Ginkgo extract, whereas Oenothera extract was projected near the position of Tramadol, an analgesic drug. Physical motion was increased only in the presence of Paullinia extract and caffeine. Increases of long-term potentiation were observed in the presence of Rhodiola extract, Paullinia extract and caffeine. The combined information predicts stimulant and cognitive function-enhancing activities in humans for the Rhodiola extract, which could also be used as a possible caffeine-replacement, and antidepressant and analgesic activity for the Oenothera extract.

基于网络药理学探讨瓜拉纳治疗阿尔兹海默症的潜在作用机制

目的通过网络药理学方法及分子对接研究瓜拉纳治疗阿尔茨海默症(AD)的主要活性成分及其潜在作用机制。方法基于文献挖掘获取瓜拉纳的化学成分并进行化学成分的类药性分析,运用本草组鉴(HERB)、中药系统药理学数据库与分析平台(TCMSP)等数据库获取瓜拉纳活性成分的潜在作用靶点并关联靶点相关疾病。利用GeneCards人类基因数据库、DisGeNET数据库收集阿尔兹海默症的疾病相关基因,利用韦恩图构建瓜拉纳治疗阿尔兹海默症的潜在靶点,运用CytoScape软件构建相应的网络互作图。找出运用STRING在线数据库建立关键靶点之间的蛋白质-蛋白质相互作用(PPI)网络图,并对关键靶点蛋白进行基因本体(GO)及京都基因与基因组百科全书(KEGG)通路富集分析。使用AutoDock软件对重要活性化合物与关键靶点进行分子对接验证。结果瓜拉纳治疗阿尔兹海默症潜在靶点共140个。PPI蛋白互作网络分析发现白细胞介素6(IL-6)、肿瘤坏死因子(TNF)、胰岛素(INS)、丝裂原活化蛋白激酶3(MAPK3)、转录因子(JUN)、肿瘤蛋白p53(TP53)、胱天蛋白酶3(CASP3)、c-Fos蛋白质(FOS)、抗氧化酶(CAT)、连环蛋白β-1(CTNNB1)等可能是瓜拉纳治疗阿尔兹海默症的关键靶点。GO富集分析和KEGG通路分析发现瓜拉纳治疗阿尔兹海默症的潜在作用是于细胞膜外以化合物与蛋白酶结合的形式发生的。分子对接结果显示瓜拉纳中多种化合物小分子与TNF、IL-6、MAPK3、FOS等靶蛋白均具有较低的结合能。结论瓜拉纳对治疗阿尔兹海默症具有多靶点协同作用的特点,其治疗AD的关键作用靶点可能是IL-6、TNF和MAPK3,它们可能通过与β-淀粉样蛋白结合的生物学过程和癌症相关信号通路发挥治疗作用,本研究为瓜拉纳治疗阿尔兹海默症进一步的研究提供了科学依据和参考。

瓜拉纳多糖制备的响应面工艺优化及抗氧化活性研究

利用响应面法优化瓜拉纳多糖的提取工艺。采用超声辅助水提法,在单因素试验的基础上,以提取时间、超声功率、液料比、提取温度为变量,以瓜拉纳多糖得率为响应值,应用Box-Benhken设计及响应面分析法进行回归分析,得到最佳提取工艺为提取时间33.5 min,超声功率155 W,水料体积质量比35 m L︰1 g,提取温度62℃,醇沉透析后冷冻干燥得到瓜拉纳多糖,此条件下瓜拉纳多糖得率为20.80%。对瓜拉纳多糖体外抗氧化活性进行评价,并以维生素C(Vc)作对照,结果发现,瓜拉纳多糖还原能力和总的抗氧化活性均与多糖浓度成正比,但都小于Vc,其中抗氧化能力接近Vc。

瓜拉纳⁃巴西人参复合饮料处方优化

目的优化瓜拉纳⁃巴西人参复合饮料处方。方法以瓜拉纳与巴西人参比例、白砂糖用量、柠檬酸用量为影响因素,再通过模糊数学感官评价建立色泽、气味、口感、组织形态感官评分,作为评价指标。在单因素试验基础上,采用Box⁃Behnken响应面法优化处方。结果最优处方为瓜拉纳与巴西人参比例1.3∶1,主药用量40.0%,白砂糖用量13.0%,柠檬酸用量0.02%,感官评分2.603分。结论该方法稳定可靠,可用于评价瓜拉纳⁃巴西人参复合饮料的品质。

Microsatellites and the Polyploid Guarana Plant:Diversity under a Sea of Alleles

Repeat blocks, microsatellites or simple sequence repeats (SSRs) can produce good co-dominant molecular markers for genetic diversity analysis and the determination of self-pollination rates in progenies originating from open pollination of selected genotypes. The enrichment of guarana genomic libraries was underway when it was confirmed that we are working with a complex polyploid species with 210 chromosomes. The probes (CA)12, (CT)12 and (TC)14 were used to finish the enrichment of four libraries for repeat blocks and the screening of a databank of expressed sequence tags (ESTs) from guarana seeded-fruits was accomplished as well. Fifteen clonal cultivars were genotyped with three replicas at 10 out of 27 identified loci using the 59 alleles that passed the reproducibility criterion. A large number of short repeat blocks were identified and this was considered to be a consequence of the recent polyploidization event. However, blocks with eight or more repeats ideal for genotyping were scarce. Annealing of most probes to short blocks by partial complementarity could explain the scarcity of longer blocks in genomic libraries but cannot explain why they were rare in the ESTs. Due to the complexity of the genotypes, alleles were treated as dominant traits. ESTs harboring repeat blocks had the functional annotation renewed. Locus GRN07 is inserted in a homologue of the MOTHER OF FLOWERING LOCUS T AND TFL1 (MFT), in which 3’-UTR displays clear post-transcriptional regulatory features. MFT and its variants are probably involved in the determination of seed germination and embryo growth characteristics. Other accessed loci can be involved in plant architecture and defense reactions. It was concluded that the alleles described in the present work can be used to distinguish guarana cultivars and possibly to analyze segregation using the progenies of controlled pollinations between divergent genitors. Also, the fingerprints obtained can be added to the morphological and agronomic descriptors of the cultivars.

瓜拉纳化学成分的UPLC-Q-TOF-MS分析

目的:对瓜拉纳中的化学成分进行系统分析,为该植物的深入研究及开发利用提供依据。方法:参照国家标准及相关文献对瓜拉纳中所含的粗蛋白、粗脂肪、粗多糖、粗纤维等大分子成分进行含量测定,应用超高效液相色谱-四极杆-飞行时间串联质谱法(UPLC-Q-TOF-MS)对瓜拉纳的化学成分进行系统定性分析,采用ACQUITY UPLC-HSS-T3色谱柱(2.1 mm×100 mm,1.8μm),以0.1%甲酸水溶液(A)-0.1%甲酸乙腈溶液(B)为流动相进行梯度洗脱(0~5 min,2%~10%B;5~6 min,10%~20%B;6~9 min,20%~30%B;9~9.5 min,30%~35%B;9.5~10.5 min,35%~45%B;10.5~13 min,45%~55%B;13~15 min,55%~80%B;15~19 min,80%~98%B;19~20 min,98%B;20~20.3 min,98%~2%B;20.3~23 min,2%B),电喷雾离子源(ESI),正、负离子模式进行检测,扫描范围m/z 50~1 500,根据精确相对分子质量及碎片信息,结合数据库匹配和对照品比对进行结构鉴定。结果:瓜拉纳中粗蛋白、粗脂肪、粗多糖及粗纤维质量分数分别为(0.63±0.03)%,(2.73±0.09)%,(3.23±0.12)%,(8.89±0.59)%。通过UPLC-Q-TOF-MS分析从瓜拉纳中共鉴定出42个成分,包括3个甲基黄嘌呤类成分,2个核苷类成分,1个氨基酸类成分,3个有机酸,33个黄酮类成分,其中3个化合物(L-色氨酸、表没食子儿茶素没食子酸酯、大豆苷元)是从瓜拉纳中首次发现。结论:瓜拉纳中的营养成分含量丰富,作为功能食品来开发具有良好潜力。UPLC-Q-TOF-MS技术为鉴别瓜拉纳药材中化学成分提供简便、快速、准确的方法,其主要化学成分为甲基黄嘌呤类、原花青素类成分,可为瓜拉纳药材的质量评价及药效物质基础研究提供参考。

瓜拉那叶中化学成分的定性分析及咖啡因的含量测定

目的利用超高效液相色谱-线性离子阱-静电场轨道阱联用质谱(UPLC-LTQ-Orbitrap-MS)技术快速鉴别瓜拉那Paullinia cupana叶中的化学成分,建立高效液相色谱(HPLC)法测定瓜拉那叶中咖啡因的含量。方法定性分析采用Waters Acquity UPLC HSS T_(3)色谱柱(100 mm×2.1 mm,1.8μm),流动相为0.1%甲酸水-乙腈,梯度洗脱,体积流量0.3 mL·min^(−1),进样量3μL,在电喷雾正负离子模式下采集数据。定量分析采用Prevail色谱柱(250 mm×4.6 mm,5μm),柱温30℃,流动相为纯水-乙腈,梯度洗脱,体积流量为1 mL·min^(−1),进样量10μL,检测波长275 nm。结果利用UPLCLTQ-Orbitrap-MS技术,根据质谱一级精确相对分子质量和二级特征碎片,从瓜拉那叶中共鉴定出46个化学成分,包括22个酚酸类,7个黄酮类,7个生物碱类,5个长链脂肪酸类和5个其他类化合物。建立HPLC-UV测定瓜拉那叶咖啡因含量的方法中,咖啡因在0.2018~2.0180μg线性关系良好(R^(2)=0.9998),平均加样回收率(n=9)为99.52%,RSD=1.16%,瓜拉那叶中咖啡因质量分数在2.8%左右。结论首次采用UPLC-LTQ-Orbitrap-MS技术鉴定瓜拉那叶中的化学成分,提供了一种快速、高效的分析方法,所建立的HPLC-UV测定咖啡因含量的方法准确、可靠,为瓜拉那叶后续的深入研究奠定了基础。