Chemical constituents from Cyclocarya paliurus (Batal.) Iljinsk

A novel triterpenoid, cyclocarioside K, together with fifteen known compounds, including seven triterpenoids (2–8), five flavonoids (9–13), three phenolic derivatives (14–16), was obtained from the leaves of Cyclocarya paliurus (Batal.) Iljinsk (Juglandaceae). Their structures were elucidated by spectroscopic methods, including two-dimensional NMR experiments (HSQC DEPT and HMBC). The chemotaxonomic relationships were also discussed. As a result, 3, 4-secodammarane triterpenoids and kaempferol glycosides may represent phytochemical fingerprints for C. paliurus, and C. paliurus has close relationships with other species of Juglandaceae.     

Cyclocarya cf. paliurus (Batal.) Iljinskaja (Juglandaceae) from the Hunchun Formation (Eocene), Jilin Province, China

The leaflet architecture of Cyclocarya cf. paliurus (Batal.) Iljinskaja from the Hunchun Formation (Middle Eocene) shows similarity to that of modern C. paliurus (Batal.) Iljinskaja and the specimen is the oldest fossil record in Europe and Asia. The distributions of C. cf. paliurus and other fossil records,such as Glyptostrobus, Metasequoia, Nyssa, and Liquidambar, in Hunchun flora show that it would have been a warmer-temperature to subtropical climate in Hunchun District during the Eocene period.     

Cyclocarya cf. paliurus （Batal.） Iljinskaja （Juglandaceae） from the Hunchun Formation （Eocene）, Jilin Province, China

The leaflet architecture of Cyclocarya cf. paliurus （Batal.） Iljinskaja from the Hunchun Formation （Middle Eocene） shows similarity to that of modem C. paliurus （Batal.） Iljinskaja and the specimen is the oldest fossil record in Europe and Asia. The distributions of C. cf. paliurus and other fossil records, such as Glyptostrobus, Metasequoia, Nyssa, and Liquidambar, in Hunchun flora show that it would have been a warmer-temperature to subtropical climate in Hunchun District during the Eocene period.     

青钱柳多糖对人胃癌MGC_803细胞生长的影响

本文研究青钱柳多糖(polysaccharides isolated from Cyclocarya paliurus(Batal.)Iljinskjk,PCP)对人胃癌MGC_803细胞生长的影响.采用噻唑蓝(MTT)法检测PCP对MGC_803细胞生长的影响;Annexin法检测PCP对MGC_803细胞诱导凋亡的作用.实验结果表明PCP在50、100、200、400 μg/mL浓度条件下,均可极显著性抑制人胃癌MGC_803细胞生长(P＜0.01),抑制率可达65.07%,细胞凋亡率可达25,44%.说明青钱柳多糖具有抑制人胃癌MGC_803细胞生长的生物活性.     

青钱柳多糖对小鼠骨髓来源树突状细胞表面分子表达的影响

为探讨青钱柳多糖（CPC）对小鼠骨髓来源树突状细胞（BMDCs）细胞形态及表面分子的影响,首先采用细胞因子诱导法,以贴壁法获得贴壁单核细胞,添加重组粒细胞-巨噬细胞集落刺激因子（rmGM—CSF）和重组白细胞介素-4（rmIL-4）进行体外诱导,倒置显微镜及透射电镜观察细胞形态的变化;流式细胞术检测培养第6dDCs的表面标志CD80,CD86和MHCⅡ类分子的表达的变化。经CPC刺激24h后,采用流式细胞术检测DCs表面MHCⅡ类分子表达的变化。结果发现：经rmGM—CSF和traiL-4诱导获得的DCs随着培养时间的延长,细胞形态发生改变,逐渐变成具有树状突起的DCs。经LPS刺激后的DCs能够发生典型的DCs的成熟,而培养第6天的DCs具有典型的DCs表型特征,可用于后续进一步实验。与阴性对照相比,CPC显著促进树突状细胞表面MHCⅡ的表达,在浓度（10～200μg／mL）范围内呈现剂量依赖性。初步表明CPC可以促进树突状细胞的成熟。     

Hypoglycemic mechanism of polysaccharide from Cyclocarya paliurus leaves in type 2 diabetic rats by gut microbiota and host metabolism alteration

Diabetes mellitus is a serious threat to human health. Cyclocarya paliurus(Batal.) Iljinskaja(C. paliurus) is one of the traditional herbal medicine and food in China for treating type 2 diabetes, and the C. paliurus polysaccharides(CP) were found to be one of its major functional constituents. This research aimed at investigating the hypoglycemic mechanism for CP. It was found that CP markedly attenuated the symptoms of diabetes, and inhibited the protein expression of Bax, improved the expression of Bcl-2 in pancreas of diabetic rats, normalized hormones secretion and controlled the inflammation which contributed to the regeneration of pancreatic β-cell and insulin resistance. CP treatment increased the beneficial bacteria genus Ruminococcaceae UCG-005 which was reported to be a key genus for protecting against diabetes, and the fecal short-chain fatty acids levels were elevated.Uric metabolites analysis showed that CP treatment helped to protect with the diabetes by seven significantly improved pathways closely with the nutrition metabolism(amino acids and purine) and energy metabolism(TCA cycle), which could help to build up the intestinal epithelial cell defense for the inflammation associated with the diabetes. Our study highlights the specific mechanism of prebiotics to attenuate diabetes through multi-path of gut microbiota and host metabolism.     

Natural population structure and genetic differentiation for heterodicogamous plant: <Emphasis Type="Italic">Cyclocarya paliurus</Emphasis> (Batal.) Iljinskaja (Juglandaceae)

Cyclocarya paliurus, a Chinese monotypic genus belonging to the Juglandaceae, exhibits a widespread distribution in southern China. To provide resource support for its future breeding and medicinal utilization, much information involving natural population succession and genetic diversity of C. paliurus should be obtained. In this study, the structures of 30 natural populations throughout its distribution were analyzed based on the investigation, showing the ratio of individuals with DBH < 40 cm accounting for 71.7%. Inter-simple sequence repeat (ISSR) and simple sequence repeat (SSR) were used for evaluation of 244 individuals from 29 natural populations. High-level polymorphism (100%) indicates high efficiency of two markers. Analysis demonstrated that genetic diversity (value of 0.18 for ISSR and 0.09 for SSR) was low among populations. While gene flow among populations reached 1.182 and 1.335 for ISSR and SSR, respectively, indicating genetic variations mainly existing within population. Twenty-six populations (size ≥5 individuals) were grouped into five clusters based on the combined ISSR and SSR loci. The value of Nei’s coefficient varying from 0.87 to 0.96 indicates small differentiation among groups. Furthermore, we speculate that C. paliurus is an expanding species in subtropical China (most of populations are increasing type (IP)) and discuss the strategy of germplasm conservation.     

Cyclocarya paliurus polysaccharides alleviate type 2 diabetic symptoms by modulating gut microbiota and short-chain fatty acids

BackgroundCyclocarya paliurus polysaccharide (CCPP), a primary active component in the leaves of Cyclocarya paliurus (Batal.) Iljinsk (C. paliurus), has the ability to treat type 2 diabetes mellitus (T2DM), but cannot be digested by our digestive system. Therefore, mechanisms of regulating the gut microbiota and intestinal metabolites might exist.PurposeTo reveal the potential mechanism of CCPP treatment, this study aimed to investigate the alterations of the gut microbiota and intestinal metabolites especially short chain fatty acids (SCFAs) in type 2 diabetic rats.Study design and methodsType 2 diabetic rat models were developed, and the therapeutic effects of CCPP were evaluated. Metagenomics analysis was utilized to analyze the alterations to the gut microbiota, and UHPLC-QTOF/MS-based untargeted metabolomics analysis of colon contents was used to identify the differential intestinal metabolites. GC/MS was used to measure the SCFAs in rat's colon contents and human fecal inoculums. Furthermore, the expression of SCFA receptors including GPR41, GPR43 and GPR109a was verified by qRT-PCR and the concentration of glucagon-like peptide-1(GLP-1) and peptide tyrosinetyrosine (PYY) was measured by Elisa.ResultsInhibition of the blood glucose levels and improvements in glucose tolerance and serum lipid parameters were observed after CCPP treatment. Eleven SCFA-producing species including Ruminococcus_bromii, Anaerotruncus_colihominis, Clostridium_methylpentosum, Roseburia_intestinalis, Roseburia_hominis, Clostridium_asparagiforme, Pseudoflavonifractor_capillosus, Intestinimonas_butyriciproducens, Intestinimonas_sp._GD2, Oscillibacter_valericigenes and Oscillibacter_ruminantium were clearly increased in the CCPP group. Furthermore, our study indicated that CCPP increases the production of SCFAs both in vivo and in vitro, and the gut microbiota are the key factor of this process. The SCFA receptors including GPR41, GPR43 and GPR109a, were significantly stimulated in the CCPP treated rats, which was accompanied by the upregulated expression of GLP-1 and PYY.ConclusionThese results demonstrated that CCPP could alleviate type 2 diabetic symptoms by increasing the SCFA-producing bacteria, promoting the production of SCFAs and upregulating SCFA-GLP1/PYY associated sensory mediators.     

Two triterpeniods from Cyclocarya paliurus (Batal) Iljinsk (Juglandaceae) promote glucose uptake in 3T3-L1 adipocytes: The relationship to AMPK activation

PurposeThe current study investigated the efficacy of Cyclocarya paliurus chloroform extract (CPEC) and its two specific triterpenoids (cyclocaric acid B and cyclocarioside H) on the regulation of glucose disposal and the underlying mechanisms in 3T3-L1 adipocytes.MethodsMice and adipocytes were stimulated by macrophages-derived conditioned medium (Mac-CM) to induce insulin resistance. CPEC was evaluated in mice for its ability by oral glucose tolerance test (OGTT) and insulin tolerance test (ITT). To investigate the hypoglycemic mechanisms of CPEC and its two triterpenoids, glucose uptake, AMP-activated protein kinase (AMPK) activation, inhibitor of NF-κB kinase β (IKKβ) phosphorylation and insulin signaling transduction were detected in 3T3-L1 adipocytes using 2-NBDG uptake assay and Western blot analysis.ResultsMac-CM, an inflammatory stimulus which induced the glucose and insulin intolerance, increased phosphorylation of IKKβ, reduced glucose uptake and impaired insulin sensitivity. CPEC and two triterpenoids improved glucose consumption and increased AMPK phosphorylation under basal and inflammatory conditions. Moreover, CPEC and its two triterpenoids not only enhanced glucose uptake in an insulin-independent manner, but also restored insulin-mediated protein kinase B (Akt) phosphorylation by reducing the activation of IKKβ and regulating insulin receptor substrate-1 (IRS-1) serine/tyrosine phosphorylation. These beneficial effects were attenuated by AMPK inhibitor compound C, implying that the effects may be associated with AMPK activation.ConclusionsCPEC and its two triterpenoids promoted glucose uptake in the absence of insulin, as well as ameliorated IRS-1/PI3K/Akt pathway by inhibiting inflammation. These effects were related to the regulation of AMPK activity.     

Hypoglycemic and antihyperglycemic activity of Syzygium alternifolium (Wt.) Walp. seed extracts in normal and diabetic rats

Aqueous, ethanolic and hexane fractions of Syzygium alternifolium seeds were prepared and given different doses of these extracts individually to different batches of rats (both normal and alloxan diabetic rats) after an overnight fast. The blood glucose levels were measured at 0, 1, 3, 5 and 7 hours after the treatment. The aqueous extract of Syzygium alternifolium at a dosage of 0.75 g/kg b.w. is showing maximum blood glucose lowering effect in both normal and alloxan diabetic rats. The ethanol and hexane fractions are also showing hypoglycemic and antihyperglycemic activity, but the effect is significantly less than that of aqueous extract. The antihyperglycemic activity of Syzygium alternifolium seed was compared with the treatment of Glibenclamide.     

Growth and triterpenic acid accumulation of Cyclocarya paliurus cell suspension cultures

Cyclocarya paliurus is a unique plant growing in central China with hypoglycaemic and hypolipaemia effects. To make better use of this functional food resource, cell suspension cultures and triterpenic acid accumulation were studied. Stable and uniform cell suspension cultures were established in liquid basal Murashige and Skoog medium supplemented with 2,4-dichlorophenoxy acetic acid (0.5 mg/L), naphthalene acetic acid (0.3 mg/L) and cytokinin (1.0 mg/L). According to the growth curve and triterpenic acid accumulation curve, the 8 ～ 10^th day postinoculation was the optimum time for subculture, and the 14^th day was the optimum time for harvest. Murashige and Skoog medium and woody plant medium were suitable for both cell growth and triterpenic acid accumulation. 3% sucrose (w/v), 60 mM total nitrogen (NO₃ ^?/NH₄ ⁺ = 2/1), 1.25 mM KH₂PO₄, 2 mM CaCl₂, and 2 mM MgSO₄ were all found to be fit for cell growth and triterpenic acid accumulation in a cell suspension culture of Cyclocarya paliurus. Total triterpenic acid, ursolic acid and oleanolic acid content in suspended cultured cells were all significantly higher than that of leaves and calluses (P ? 0.01), with levels up to 6.24, 2.28, and 0.94% (of dry weight), respectively. The betulinic acid content of suspended cultured cells also reached 0.82%, which was significantly higher than that of calluses. These results suggest that suspended cultured cells of Cyclocarya paliurus were rich in triterpenic acids and could be used for the production of total triterpenic acid, ursolic acid, oleanolic acid and betulinic acid.     

烯效唑对青钱柳试管苗生长及生理特性的影响

在含0.00(CK)、0.01、0.05、0.10和1.00 mg·L-1烯效唑的WPM培养基上继代培养120 d后,对青钱柳[Cyclocarya paliurus (Batal.)Iljinskaja]试管苗的部分生长及生理指标的变化进行了比较研究.结果显示:不同质量浓度烯效唑对青钱柳试管苗的生长及生理指标有不同的影响效应.总体上,随培养基中烯效唑质量浓度的提高,青钱柳试管苗的苗高、叶片数和可溶性蛋白质含量逐渐降低,可溶性糖与可溶性蛋白质含量的比值、SOD和POD活性逐渐提高;在培养基中添加0.01、0.05和0.10 mg·L-1烯效唑对青钱柳试管苗的成活率无显著影响,却可使试管苗的单株鲜质量增加量、叶绿素含量和可溶性糖含量均高于对照;在培养基中添加1.00 mg·L-1烯效唑能显著或极显著降低试管苗的成活率、单株鲜质量增加量、分化芽数、苗高、叶片数以及叶绿素含量、可溶性糖含量和可溶性蛋白质含量,并使苗茎出现异常增粗和矮化.而在含0.10 mg·L-1烯效唑的培养基上,虽然试管苗的苗高、分化芽数和叶片数分别较对照降低了28.03%、9.70%和12.37%,但试管苗的单株鲜质量增加量、叶绿素含量、可溶性糖含量、可溶性糖与可溶性蛋白质含量的比值、 SOD和POD活性分别较对照提高了99.39%、 14.00%、 5.00%、115.43%、129.77%和33.79%.研究结果表明,在培养基中添加0.10 mg·L-1烯效唑可有效改善青钱柳试管苗的生长和生理特性,有效控制苗高和叶片数,促进苗茎的增粗,有助于增强试管苗的抗逆能力.     

不同激素对青钱柳外植体和愈伤组织褐化的影响

以青钱柳(Cyclocarya paliurus)叶片为研究材料, 采用单因素完全随机试验法比较不同激素(6-BA、GA₃和NAA)浓度组合对叶片外植体褐化的影响; 在此基础上, 分别采用二因素、单因素完全随机试验法比较6-BA+NAA不同浓度组合、不同基本培养基对愈伤组织褐化的影响。结果表明, 改良MS+1.0 mg∙L ^-1 6-BA+1.0 mg∙L ^-1 GA₃+0.3 mg∙L ^-1 NAA是抑制青钱柳叶片外植体褐化的最佳培养基, 其褐化率为0, 愈伤组织诱导率100%; 改良MS+0.5 mg∙L ^-1 6-BA+0.2 mg∙L ^-1 NAA是抑制愈伤组织褐化的最佳培养基, 其褐化率为0, 愈伤组织增殖倍数达4.80倍, 愈伤组织呈黄绿色或黄色, 颗粒状, 颗粒小而紧密、质硬且表面干燥。该方法有效解决了青钱柳叶片外植体和愈伤组织褐化问题, 为青钱柳组织培养褐化控制提供了一条简单高效的离体培养途径, 也为青钱柳叶片离体再生体系的建立奠定基础。     

灰直纹螟-青钱柳虫茶的营养成分分析与评价

分析了灰直纹螟-青钱柳虫茶(以下简称:灰青虫茶)一般营养成分、茶特征物质、氨基酸、脂肪酸、矿物质组成,同时也对其重金属含量进行安全性评价,并以仓樟虫茶、紫白虫茶和常规茶为参照,对灰青虫茶的营养价值进行了评价。结果表明,灰青虫茶含有较高的茶多酚物质;氨基酸总含量较高,其8种必需氨基酸约占氨基酸总量的39.3%,必需氨基酸与非必需氨基酸的比值为64.76%,必需氨基酸评价显示灰青虫茶的氨基酸模式接近鸡蛋且优于大豆;矿物质含量丰富,其中以Ca含量最高,同时含有丰富的K元素,K/Na之比为208.04;按国标限量要求重金属含量无超标。灰青虫茶氨基酸和矿物质含量丰富、种类齐全,氨基酸模式较合理,是优质蛋白和安全的矿物质保健饮品,具有较高的营养价值。     

不同激素对青钱柳外植体和愈伤组织褐化的影响

以青钱柳(Cyclocarya paliurus)叶片为研究材料, 采用单因素完全随机试验法比较不同激素(6-BA、GA₃和NAA)浓度组合对叶片外植体褐化的影响; 在此基础上, 分别采用二因素、单因素完全随机试验法比较6-BA+NAA不同浓度组合、不同基本培养基对愈伤组织褐化的影响。结果表明, 改良MS+1.0 mg?L ^-1 6-BA+1.0 mg?L ^-1 GA₃+0.3 mg?L ^-1 NAA是抑制青钱柳叶片外植体褐化的最佳培养基, 其褐化率为0, 愈伤组织诱导率100%; 改良MS+0.5 mg?L ^-1 6-BA+0.2 mg?L ^-1 NAA是抑制愈伤组织褐化的最佳培养基, 其褐化率为0, 愈伤组织增殖倍数达4.80倍, 愈伤组织呈黄绿色或黄色, 颗粒状, 颗粒小而紧密、质硬且表面干燥。该方法有效解决了青钱柳叶片外植体和愈伤组织褐化问题, 为青钱柳组织培养褐化控制提供了一条简单高效的离体培养途径, 也为青钱柳叶片离体再生体系的建立奠定基础。     

青钱柳黄酮及三萜调节人体肠道菌群作用研究

以青钱柳为材料,通过热水浸提,大孔树脂柱层析分离纯化,制备得到纯度大于90%的青钱柳黄酮和三萜。继而采用体外厌氧粪样混合培养与荧光原位杂交技术,评价青钱柳黄酮和三萜对于人体肠道菌群的调节作用。结果表明青钱柳黄酮和三萜类物质能有效促进肠道中有益细菌(双歧杆菌和乳酸菌)的增殖,而对于梭状菌和拟杆菌则有抑制作用,同时未显著影响肠道总菌群的数量。研究表明,青钱柳中的黄酮和三萜类物质,对于改善人体肠道环境、维护人体健康具有重要作用。     

青钱柳组培快繁体系的初步研究

以青钱柳成熟胚为实验材料,对建立青钱柳组培快繁技术体系进行初步研究.结果表明,离体胚在添加30 g/L蔗糖、6.5 g/L琼脂的培养基中培养60 d,成苗率可达到72.67%.通过正交试验筛选出丛生芽诱导增殖的最佳培养基为WPM+0.5 mg/L 6-BA+0.01 mg/L IBA+30 g/L蔗糖,适宜的继代周期为40 d,丛生芽诱导率可达到93.33%,增殖系数最高为4.75,3种激素对丛生芽增殖系数的影响程度依次为IBA>6-BA>KT,其中IBA有显著影响.培养基中添加适当浓度的烯效唑和稀土镧对试管苗的壮苗和生长均有促进作用,其最适浓度分别为0.1 mg/L和5 mg/L.适宜浓度的IBA对试管苗生根有一定促进作用,采用WPM+0.2 mg/L IBA+20 g/L蔗糖培养基进行诱导,生根率为16.67%,经过15 d暗诱导处理,生根率可提高到23.33%.     

Hypoglycemic activity of Eriobotrya japonica seeds in type 2 diabetic rats and mice

The hypoglycemic effects of Eriobotrya japonica seeds were investigated in type 2 diabetic Otsuka Long-Evans Tokushima fatty (OLETF) rats and KK-A(y) mice. The rats and mice were fed on a diet containing 10% powdered Eriobotrya japonica seeds with the coat intact for 4 months. Although the blood glucose concentration in the OLETF rats fed on the control diet without Eriobotrya japonica seeds was increased with time, the concentration in the OLETF rats fed on the diet with Eriobotrya japonica seeds was consistently low throughout the experimental period and was comparable to the level in Long-Evans Tokushima Otsuka (LETO) rats which are normal non-diabetic rats. Serum insulin was significantly lower in the OLETF rats fed on the Eriobotrya japonica seed diet than in those fed on the control diet at the termination of the experimental period. Eriobotrya japonica seeds suppressed the increment of blood glucose for 4 months and also effectively improved the glucose tolerance in the KK-A(y) mice, these actions being mainly exerted by the ethanol extract of the seeds. These results suggest that Eriobotrya japonica seeds had a hypoglycemic property and the effect is attributable to the components extracted by ethanol.     

青钱柳法呢基焦磷酸合成酶基因的克隆及功能研究

青钱柳是集药用、材用和观赏等多种价值于一身的珍贵树种。法呢基焦磷酸合成酶(FPS)催化=牛儿基焦磷酸(GPP)与异戊烯基焦磷酸(IPP)缩合成法呢基焦磷酸(FPP),FPP是植物次生代谢产物倍半萜,三萜,甾醇等的前体。本研究通过RACE方法首次从青钱柳中扩增了法呢基焦磷酸合成酶的全长cDNA序列,序列命名为CpF-PS(Genbank登录号为GU121224),序列长度为1 420 bp,包含1 029 bp的开放阅读框,编码342个氨基酸残基,预测蛋白分子量为39.60 kDa。通过BlASTP分析,推断的青钱柳FPS蛋白序列与木本棉(Gossypium arboreum)(CAA72793.1)、橡胶树(Hevea brasiliensis)(BAF98301)等的FPS蛋白相似度较高。蛋白质保守区、特征区以及进化树分析初步证实扩增到的全长cDNA序列为青钱柳的FPS基因。将该基因连入酵母表达载体并转入麦角甾醇缺陷型酵母菌株CC25(MATa/MATalpha,deltaERG20/+),发现该基因可弥补营养缺陷使得CC25菌株在高温中正常生长,证明所得到的青钱柳CpFPS基因编码的蛋白是有功能的蛋白。