Comparative analysis of chemical components between barks and leaves of Eucommia ulmoides Oliver

The chemical components of the essential oils in the barks and leaves of Eucommia ulmoides Oliver were analyzed and compared by chromatograms and mass spectra technique, heuristic evolving latent projections (HELP), alternative moving window factor analysis (AMWFA) algorithms and normalization method based on the peak areas; the flavones in the barks and leaves of Eucommia ulmoides Oliver were separated on an ODS column by gradient elution carried out with the flow phase consisting of water, methanol and phosphoric acid (0.1%), and their contents were quantitatively determined by standard curve method and diode array detection (DAD) at 362 nm. The results show that 68 and 73 compounds respectively from essential oils of the barks and leaves of Eucommia ulmoides Oliver are identified, and there are 33 mutual compounds among 108 compounds determined. The total contents of these volatile components of the two samples possess 92.9% and 97.75% of the gross of the relevant essential oils, respectively; the contents of the rutin, quercetin and kaempferol in the barks and leaves of Eucommia ulmoides Oliver are 0.016 9, 0.003 6, 0.002 1 and 0.064 4, 0.030 2, 0.010 0 mg/g, respectively, and the determination recoveries are 95.2%-106.2%. The comparative analysis shows that for the barks and leaves of Eucommia ulmoides Oliver, there are significant differences in their components of the relevant essential oils and flavones.

Chemical constituents,biological functions and pharmacological effects for comprehensive utilization of Eucommia ulmoides Oliver

Eucommia ulmoides Oliver is a native plant and valuable tonic Chinese medicine in China with a long history,great economic value and comprehensive development potential.Traditionally,the comprehensive utilization rate of E.ulmoides Oliv.is still very low,only bark has been used as medicine and other parts of Eucommia ulmoides Oliv.cannot be fully utilized,even the leaves have been well utilized in food products in Japan in the past decades.In order to improve the comprehensive utilization efficiency of E.ulmoides Oliv.,in this review,we summarized the varieties and contents of main active compounds,biological functions and pharmacological effects in different parts of E.ulmoides Oliv.The findings suggest that other parts of E.ulmoides Oliv.could replace the bark of E.ulmoides Oliv.to some extent besides of their respective applications.The unique and extensive physiological functions between different parts of E.ulmoides Oliv.indicate that the comprehensive utilization of E.ulmoides Oliv.has a wide space to develop,which is also an effective way to protect E.ulmoides Oliv.resources and improve its the utilization rate.

Isolation of a novel antifungal peptide from the bark of Eucommia ulmoides Oliv

A novel Eucommia antifungal peptide, named EAFP3, was isolated from the bark of Eucommia ul- moides by NaC1 extract, gel filtration and reverse phase high performance liquid chromatography. The molecular mass of EAFP3 is 4157.3 Da, and its partial amino acid sequence is -. LYQQLIAGITLNK.-. EAFP3 exerts an inhibitory activity against Candida albicans in vitro and the drug concentration required for 50% growth inhibition （IC50） is 31.25μg/mL.

Cloning and Characterization of EuGID1 in Eucommia ulmoides Oliver

Gibberellic acid controlled the key developmental processes of the life cycle of landing plants,and regulated the growth and development of plants.In this study,a novel gibberellin receptor gene EuGID1 was obtained from Eucommia ulmoides Oliver.The cDNA of EuGID1 was 1556 bp,and the open reading frame was 1029 bp,which encoded 343 amino acids.EuGID1 had the homology sequence with the hormone-sensitive lipase family.Amino acid sequence alignment confirmed EuGID1 protein had the highest homology with the GID1 protein of Manihot esculenta.EuGID1 was located in the nucleus and cell membrane and had expression in four plant organs.Overexpression of EuGID1 in transgenic Arabidopsis plants promoted plant elongation and increased siliques yield.

Eucommia ulmoides Oliv.antagonizes H_2O_2-induced rat osteoblastic MC3T3-E1 apoptosis by inhibiting expressions of caspases 3,6,7,and 9

Eucommia ulmoides Oliv.(EuO),also known as Duzhong,native to China,has been reported to have antioxidative function,but its cellular mechanism is not fully examined yet.We investigated inhibitory effects of EuO leaf ethanol extracts on H2O2-induced apoptosis in rat osteoblastic MC3T3-E1 cells and underlying mechanisms.Locally-grown Duzhong leaves were extracted with ethanol.MC3T3-E1 cells were treated with EuO (6.25,12.5,25,50,and 100 μg/ml) for 24 h,and then H2O2 (800 μmol/L) for an additional 24 h.Cell survival rate,percentage of apoptosis,and expressions of caspases 3,6,7,and 9 were examined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay,microscopic analysis,Western blotting,and reverse transcription polymerase chain reaction (RT-PCR).The final EuO leaf ethanol extract powder was detected to contain caffeotannic acid at 58 mg/g and geniposide at 3.45 mg/g by high performance liquid chromatography (HPLC).EuO remarkably restrained cell oxidative damage and increased cell survival rate in a dose-dependent manner: 0 μg/ml,0.21;6.25 μg/ml,0.28;12.5 μg/ml,0.31;25 μg/ml,0.48;50 μg/ml,0.54;and 100 μg/ml,0.66 (P<0.05),with the half-effective concentration being around 25 μg/ml.MTT results were confirmed by microscopic analysis.Western blotting and RT-PCR analyses showed that the expressions of caspases 3,6,7,and 9 were significantly decreased in the EuO-treated cells compared with the control (EuOand H2O2-free) (P<0.05),with the half-effective concentration of EuO ranging from 12.5 to 25 μg/ml.We conclude that the ethanol-extracted EuO leaf extracts promoted the growth of MC3T3-E1 cells,and suppressed the H2O2-induced apoptosis in a rat MC3T3-E1 osteogenic cell model,likely due to the inhibition of caspases’ activities.The results indicate that EuO is a potent antioxidant,which may contribute to its many cellular protective functions,including the promotion of bone growth.

Plant regeneration from dry mature endosperm cultures of Eucommia ulmoides Oliv.

EUCOMMIA ulmoides is not only a famous medicinal plant,but also a most promising rubber-source plant in the temperate zone.All the individuals present in this species are diploid(2n=2x=34),and the objective of this work is to establish a novel triploid type of it so as to im-prove its overall biomass output or rubber content in the leaf cells by the characteristic cell en-

杜仲提取物对鲫抗氧化性能的影响

该试验探讨在鲫(Carassius auratus)的饲料中添加不同水平的杜仲提取物对其机体抗氧化性能的影响,确定杜仲(Eucommia ulmoides Oliv.)提取物在鲫养殖中的最适添加水平。试验选择体重约为50g的健康的鲫600尾,按照单因素试验设计,将其均分4个处理,每个处理5个重复,每个重复30尾鲫。对照组鲫投喂基础饲料,试验组鲫投喂添加0.1%、0.2%和0.4%的杜仲提取物的饲料。试验鲫驯养1周,正式试验期6周。试验结束,检测各组鲫机体的抗氧化相关指标。结果显示,投喂0.2%和0.4%的杜仲提取物可以显著降低血清中丙二醛水平(P<0.05),显著提高了过氧化氢酶活力、谷胱甘肽过氧化物酶活力、谷胱甘肽-硫转移酶活力(P<0.05);投喂0.1%、0.2%和0.4%的杜仲提取物显著提高了超氧化物歧化酶活力(P<0.05)。研究结果表明,杜仲提取物可以有效改善鲫机体的抗氧化功能,且杜仲提取物的最适添加量为0.2%。

Extraction Process of Total Flavonoids from Eucommiae Cortex and Its in vitro Antioxidant Activity

[Objectives]To optimize the extraction process of total flavonoids from Eucommiae Cortex,establish the extraction and content determination methods of its medicinal materials,and study its in vitro antioxidant activity.[Methods]The total flavonoids from Eucommiae Cortex were extracted by reflux extraction method,and the effects of extraction method,extraction solvent concentration,extraction volume,and extraction time on the total flavonoids content of the medicinal materials were explored through the single factor experiment.Orthogonal experiment was carried out to optimize the extraction process conditions,and the optimal extraction process for total flavonoids from Eucommiae Cortex was screened out.The total antioxidant capacity index was used to determine the free radical scavenging ability of the total flavonoids from Eucommiae Cortex.[Results]The optimal extraction process of total flavonoids from Eucommiae Cortex was 50%ethanol,1∶40 solid-to-liquid-ratio,and 1.5 h reflux extraction time.Through the antioxidant experiment in vitro,it is found that the total flavonoids from Eucommiae Cortex have a higher scavenging ability for the total antioxidant capacity,and there is a certain positive correlation with the mass concentration of total flavonoids.[Conclusions]This method can effectively determine the total flavonoids content of Eucommiae Cortex medicinal material,and provide a certain scientific basis for the quality standard research of the medicinal material.The total flavonoids from Eucommiae Cortex have good in vitro antioxidant activity.And the method for extracting total flavonoids from Eucommiae Cortex medicinal material in this experiment has good repeatability,high stability and feasibility.

中药杜仲皮总黄酮提取工艺及抗氧化活性研究

优化杜仲(Eucommia ulmoides Oliv.)皮总黄酮提取工艺,建立了总黄酮含量测定方法,并对其体外抗氧化活性进行研究。采用回流提取法提取杜仲皮总黄酮,通过单因素试验,考察了提取方法、溶剂浓度、溶剂体积、提取时间对总黄酮含量的影响,采用正交试验优化提取工艺,筛选出杜仲皮总黄酮的最佳提取工艺;测定总抗氧化能力、羟基自由基清除自由基能力判断杜仲皮总黄酮的体外抗氧化活性。结果表明,杜仲皮总黄酮的最佳提取工艺为50%乙醇、料液比1∶40 g/mL、回流提取时间1.5 h;总黄酮具有较强的还原能力,浓度为3.0 mg/mL时其总抗氧化能力为132.2 U/mL,总抗氧化能力随着总黄酮浓度的增加而增强,相同溶液浓度下较维生素C强;总黄酮对羟基自由基有较强的清除能力,溶液浓度为3.0 mg/mL时,羟基自由基清除率为90%,清除率随总黄酮浓度的增加而增大,最后趋于平稳。该方法能有效测定杜仲皮总黄酮的含量,提取的总黄酮具有良好的体外抗氧化活性,精密度、准确性、稳定性、重复性较好,为该药材的质量标准研究提供参考。

Pharmacokinetic study about compatibility of Eucommia ulmoides and Psoralea corylifolia

Objective:The compatibility of Eucommia ulmoides(Eu)and Psoralea corylifolia(Pc)on the pharmacokinetic(PK)properties in the rat was explored in this study.Methods:Eu extract,Pc extract and the combined extracts(crude drug ratio was 2:1)was administered by gavage,respectively.Two PK experiments were conducted.In first one,the blood samples were collected via the occuli chorioideae vein to get the PK properties of the components.In second one,the blood samples were simultaneously collected via the internal jugular vein or portal vein at different time points and the concentrations of target ingredients were detected by LC/MS/MS to clear the location where the interaction of Eu and Pc took place in vivo.Results:Eight of 11 ingredients in Eu and Pc extract were determined in rat plasma.The exposure levels of geniposidic acid(GPA),aucubin(AU),geniposide(GP),pinoresinol diglucoside(PDG),psoralen glycosides(PLG)and isopsoralen glycosides(IPLG)were decreased 1/5–2/3 after administration of combined extracts.Comparing to the combined administration,the exposure of GPA and AU in plasma of single Eu administration collected via the portal vein were decreased 1/3–2/3,and the values of AUC0-24h and AUC0-∞ of GP collected from the portal vein or internal jugular vein were double increased.The other components’parameters were not significantly changed.Conclusion:In summary,the Pc and Eu combined administration could affect the exposure of the main components of Eu extract in rats due to the changed intestinal absorption.The research on the compatibility of Pc and Eu was helpful to guide the clinical administration of Eu and Pc simultaneously.

Ectopic Overexpression of EuCHIT30.7 Improves Nicotiana tabacum Resistance to Powdery Mildew

Various strains of powdery mildew(PM),a notorious plant fungal disease,are prevalent and pose a significant threat to plant health.To control PM,transgenic technology can be used to cultivate more resistant plant varieties.In the present study,we utilized the rapid amplification of cDNA ends(RACE)technique to clone the full-length cDNA sequence of the EuCHIT30.7 gene to explore plant genes with disease resistance functions.Bioinformatics analysis revealed that this gene belongs to the GH18 family and is classified as a class III chitinase.The EuCHIT30.7 gene is expressed throughout the Eucommia ulmoides plant,with the most abundant expression in male flowers.Subcellular localization analysis indicated that the protein encoded by this gene was detected within both the cell membrane and cytoplasm.Upon PM inoculation,overexpression of EuCHIT30.7 in tobacco plants led to a significantly reduced relative lesion area and a decreased spore count compared to both wild-type and empty vector control plants.Activities of the protective enzymes,namely,peroxidase(POD),superoxide dismutase(SOD),catalase(CAT),and phenylalaninammo-nialyase(PAL),in tobacco plants overexpressing EuCHIT30.7 were significantly greater than those in wild-type and empty vector tobacco plants.Furthermore,the rate of increase in malondialdehyde(MDA)content was significantly lower in tobacco plants expressing EuCHIT30.7 compared to control tobacco plants.In EuCHIT30.7 transgenic tobacco,the expression of pathogen-related protein genes,namely,PR2,PR5,PR1a,PDF1.2,and MLP423,along with the tobacco PM negative regulatory gene,MLO2,were significantly higher compared to control tobacco plants.These findings suggested that EuCHIT30.7 significantly enhances the resistance of tobacco to PM.

杜仲活性成分四氢呋喃骈四氢呋喃型木脂素的快速分析

杜仲(eucommia ulmoides Oliv)具有补肝肾、强筋骨、安胎和降压等功效.其中四氢呋喃骈四氢呋喃型木脂素松脂素二葡萄糖苷是其主要的降压有效成分;丁香脂素二葡萄糖苷在治疗淋巴细胞白血病中有较好的活性[1].……

双水相体系分离纯化杜仲总黄酮和绿原酸的研究

采用双水相萃取法辅助醇提法来提取纯化杜仲(Eucommia ulmoides Oliver)中的总黄酮和绿原酸。通过单因素试验和Box-Behnken响应面法确定最佳提取工艺,利用乙醇/无机盐双水相体系将杜仲提取物中的总黄酮和绿原酸进行有效分离。结果表明,在最佳提取条件下,杜仲总黄酮的最佳双水相萃取体系为乙醇的质量分数50%、K2HPO4的质量分数20%,此时,相比R为2.93,分配比D为10.35,杜仲总黄酮的萃取率达77.94%。绿原酸的最佳双水相萃取体系为乙醇的质量分数60%、K2HPO4的质量分数20%,此时,相比R为3.43,分配比D为12.32,杜仲绿原酸的萃取率达78.22%。开发条件温和、成相稳定、用时较短的双水相萃取技术,对杜仲的开发利用有重要意义。

Two-step emulsification for the fabrication of homogeneous PCL microspheres encapsulating geniposidic acid with antioxidant properties

The mortality rate of neurological disorders is increasing globally,and natural antioxidant geniposidic acid(GPA)holds great potential in the treatment of neuronal oxidative damage.Nevertheless,its inherent instability constrains its pragmatic utilization.Herein,we introduced a drug delivery system capable of protecting unstable natural active compounds from degradation.Among the various methods for preparing drug-loaded microspheres,the emulsification-solvent evaporation technique is one of the most commonly employed due to its efficiency and simplicity.Nevertheless,this method results in microspheres with heterogeneous particle sizes.To address this limitation,we developed a two-step emulsification method involving stirring and homogenization.Using the biocompatible,synthetic,biodegradable polymer polycaprolactone(PCL)as the drug delivery carrier,we prepared GPA-loaded PCL microspheres via the two-step emulsification method.The results demonstrated that the microspheres possessed uniform particle size(polydispersity index=0.12),excellent drug loading capacity(∼4.86%),sustained drug release profiles(∼68.55%in 264 h),and biocompatibility(cell viability>85%).The in vitro tests showed that the microspheres exerted antioxidant effects by scavenging reactive oxygen species(ROS)induced by oxidative stress,thereby protecting neuronal cells from oxidative damage.This work presents a promising new approach for the treatment of neuronal oxidative damage.

Quality assessment of processed Eucommiae Cortex based on the color and tensile force

Eucommiae Cortex(EC),the dried stem bark of Eucommia ulmoides Oliv,has been traditionally used to strengthen the muscle and bone tissues and improve liver and kidney functions in East Asian countries,including China,Japan,and Korea.Salty-fried EC(SFEC)is made by using mixing EC and saline together based on the protocol of Chinese Materia Medica Processing(CMMP)for clinical use.However,the clinical effectiveness of SFEC is directly impacted by the frying temperature and time.But precise techniques for evaluating the caliber of SFEC have yet to be developed.Thus,this study aimed to establish a fast and accurate quality-check method for SFEC decoction pieces.According to the frying temperature and time,four different categories of SFEC had been got according to the Chinese Pharmacopoeia method as Raw(R),Under(U),Moderately(M),and Overly(O).The red(R),green(G),blue(B),and light(L)color values of the decoction pieces were quantitated using the Photoshop software to determine the standard value range of the L color(raw[104.44±15.06],under[67.28±8.20],moderately[39.94±6.40],and over[15.02±5.03]).Additionally,the tensile strengths and pinoresinol diglucoside(PDG)levels of EC gum-silks were measured using the mechanical tensile test and HPLC,respectively.We also conducted Pearson correlation analysis on the L color value,EC gum-silk tension(FbcN),and PDG level,and established the following muliple-linear regression equation:the decline in PDG level(w)=0.829-0.001×FbeN-0.009×I.In conclusion,the L color value and FbeN could be used for cluster analysis of four categories of SFEC.Additinally,based on the correlation among the L color value,gum-silk tensile test,and PDG level,a rapid and accurate quality-control method for SFEC decoction pieces with different frying temperatures and durations was estab-lished.This method facilitates the manufacturing of efficacious SFEC.

外源激素对杜仲苗形态及光合作用的影响

为给杜仲(Eucommia ulmoides Oliv.)栽培生理研究提供一定的理论基础,对杜仲苗喷施不同外源激素后,对杜仲苗的株高、节间距、基茎、叶片数、叶长、叶宽及光合作用等指标进行了测定。结果表明:30 mg/L GA_3和20 mg/L ABA对杜仲苗株高有极显著促进生长的作用(P<0.01),且杜仲苗叶片数显著增多(P<0.05),其中以GA_3效果最为显著;而净光合速率(Pn)、气孔导度(Gs)、蒸腾速率(Tr)与对照差异不显著。20 mg/L 2,4-D对杜仲苗各形态指标无显著影响,但喷施后第5天和第20天的净光合速率、气孔导度、蒸腾速率均显著高于对照,第10天的净光合速率、气孔导度、蒸腾速率在8:00—14:00显著高于对照,与对照差异达到极显著水平(P<0.01),表明20 mg/L 2,4-D能有效增强杜仲苗的光合速率。