α-Glucosidase Inhibitors from Glycyrrhiza uralensis Fisch.

Aim To screen for α-glucosidase inhibitor from Glyeyrrhiza uralensis Fisch.. Methods Glycyrrhizic acid, glycyrrhetinic acid, flavonoids of glycyrrhiza, alkaloids of glycyrrhiza, and glycyrrhiza polysaccharides were isolated from the root of Glycyrrhiza uralensis Fisch. respectively. Three compounds were isolated from the flavonoids of glycyrrhiza as guided by the α-glucosidase inhibitory test in vitro. Moreover, the characteristics of inhibitory kinetics of glycyrol and glycyrrhetinic acid were investi- gated. Results The flavonoids of glycyrrhiza and glycyrrhetinic acid had the strongest α-glucosidase inhibitory activity. Glycyrol,β-sitosterol and liquifitin were isolated and identified. Glycyrol was a fast- binding, reversible, noncompetitive α-glucosidase inhibitor, showing IC50 at 0.26 μg·mL^-1 Glycyrrhetinic acid was a fast-binding, irreversible α-glucosidase inhibitor, showing IC50 at 102.4 μg·mL^-1. Conclusion Glycyrol is an effective α-glucosidase inhibitor.

Measurement and comparison of glycyrrhizic acid contents in root of licorice(Glycyrrhiza uralensis Fisch.)from different cultivating areas

The samples of licorice (Glycyrrhiza uralensis Fisch.) from 14 different cultivated areas were determined by the method of high Performance Capillary Electrophoresis (HPCE) for the contents of glycyrrihizic acid (GA) in root. The results showed that the licorice plants come from various cultivated areas of China has different contents of GA. The GA content of licorice from Zhaodong in Heilongjiang Province is the highest, followed by those from E抰uoke, Chifeng, and Hangjin Banner in Inner Mongolia. Some suggestions for establishing the production base of licorice were put forward based on the study.

Primary Speciation Analysis on 6 Kinds of Microelements in Glycyrrhiza uralensis Fisch.

[Objective]The aim was to analyze the primary speciation of 6 microelements in Glycyrrhiza uralensis Fisch. and provide theoretical basis for explaining pharmacodynamic principle of liquorice and discussing quality control of liquorice planting. [Method]The 6 elements Cu,Zn,Ca,Fe,Mg and Mn in roots of G.uralensis were extracted based on traditional decoction method and were separated into water-soluble state and suspension state by micro porous filtering film. The elements in water-soluble state were detected by flame atomic adsorption spectrophotometry （FAAS）. [Result]The results showed that extractive rates of the elements were in the range of 1.71%-60.06%,and immerse-residue ratio in 0.018 3-1.682 0; the results also indicated that the immerse-residue ratio of Zn was biggest （1.68）,Zn played an important medical role and might be considered as the best characteristic element in G.uralensis; the recoveries of the elements were ranged from 95.72% to 103.15% and relative standard deviations （RSD） were less than 2.38%. [Conclusion]Because of its high accuracy,FAAS method is feasible for analyzing primary speciation of microelements in G.uralensis.

Effect of Glycyrrhiza uralensis Fisch polysaccharide on growth performance and immunologic function in mice in Ural City,Xinjiang

Objective:To discuss the effect of Glycyrrhiza uralensis(G.uralensis) Fisch polysaccharide on growth performance and immunologic function in mice in Ural City,Xinjiang and to provide important data supporting the application of Glycyrrhiza polysaccharide.Methods:A total of100 Kunming mice aged 3 weeks old were randomly divided into 5 groups with 20 mice in each group(10 were females and 10 were males).About 0.5 mL normal saline was given to the mice of control group every day and 0.5 mL G.uralensis Fisch polysaccharide was given to the mice of other groups at the concentration of 1,20,50 and 100 mg/mL respectively.The growth performance(average body weight,average daily feed intake and feed efficiency),immune organ indexes(spleen index and thymus index) and immunologic function(serum IL-2,CD4^+/CD8^+ and the activity of NK cells) of mice in each group were detected continuously.Results:The average body weight,feed efficiency,serum IL-2,CD4^+/CD8^+ and the activity of NK cells of mice were increased with the increase of administrated time after administrating G.uralensis Fisch polysaccharide and were reached up the largest level on Day 28.At the same time,each index was proportional to the given dose and was significantly higher than those of control group and reached up the largest level at the administrated dose of 100 mg/mL.After administrating G.uralensis Fisch polysaccharide,the spleen index and thymus index of mice were increased with the increase of administrated dose and the spleen index and thymus index of mice administrated with the dose of 100 mg/mL were maximum which was more than 1.51 times and 1.43 times of that in control group respectively and the comparative differences showed statistical significance(P<0.05).The average daily feed intake of mice in each group was increased with the passage of lime and at the same time,the comparison of average daily feed intake of mice in each group was not significantly different(P>0.05).Conclusions:G.uralensis Fisch polysaccharide can significantly improve the growth performance and immunologic function of mice and laid a research basis for the clinical application of G.uralensis Fisch polysaccharide.

Molecular Cloning and Expression Analysis of an Actin Gene from Chinese Licorice(Glycyrrhiza uralensis Fisch)

A PCR-based homologous cloning strategy was used to identify an actin gene from the roots of Chinese licorice（Glycyrrhiza uralensis Fisch）. Results of sequence analysis indicate that a 1137 bp cDNA with an open reading frame encoding 377 amino acids, actin ortholog, GuActin, was successfully cloned and characterized（GenBank accession No. EU190972）. Thus far, GuActin is the first actin of Chinese licorice that has been identified at a molecular level. Analysis by Northern blot shows that GuActin was expressed strongly in the roots, particularly in radicles than in stems and leaves. These results suggest that GuActin may be a member of the vegetative subfamily of the actin family.

Effect of Ar Ion Beam Implantation on Morphological and Physiological Characteristics of Liquorice(Glycyrrhiza uralensis Fisch)Under Short-Term Artificial Drought Conditions

Ar^＋ ion beam with low energy of 30 keV was implanted into liquorice （Glycyrrhiza uralensis Fisch） seeds at the doses of 0, 600, 900 and 1200 × （2.6 × 10^13） ions/cm^2, respectively. The seeds were sowed in pots and after one month the plants were subjected to different drought conditions for two months. Then the plants＇ morphological and physiological characteristics, antioxidation enzymes and levels of endogenous hormones were investigated. The results showed that ion implantation at a proper dose can greatly enhance the liquorice seedlings＇ resistance against drought stress.

Impact of Ion Implantation on Licorice(Glycyrrhiza uralensis Fisch) Growth and Antioxidant Activity Under Drought Stress

Low energy ion beams are known to have stimulation effects on plant generation and to improve plants＇ intrinsic quality. In the present study, the growth and physiological index of licorice implanted with 0, 8, 10; 12 and 14× （2.6× 10^15） ions/cm^2 were investigated under well-watered and drought-stress conditions. The results showed that a proper dose of ion implan- tation was particularly efficient in stimulating the licorice growth and improving the plant biomass significantly in both the well-watered and drought-stress conditions. The physiological results of licorice measured by leaf water potential, lipid oxidation, soluble protein and antioxidant system showed a significant correlation between ion implantation and water regime except for leaf water potential. Therefore, the study indicated that ion implantation can enhance licorice＇s drought tolerance by increasing the activity of superoxide dismutase （SOD）, eatalase （CAT） and DPPH （1,1-diphenyl-2-picrylhydrazyl） radical scavenging ability to lower oxidative damage to lipids in plants. Ion beam implantation, therefore, provides an alternative method to enhance licorice drought tolerance.

Ethanol Extract of Glycyrrhiza uralensis Fisch:Antidiarrheal Activity in Mice and Contraction Effect in Isolated Rabbit Jejunum

Objective:To evaluate the antidiarrheal effect of ethanol extract of Glycyrrhiza uralensis Fisch root(GFR)in vivo and jejunal contraction in vitro.Methods:In vivo,50 mice were divided into negative control,positive control(verapamil),low-,medium-and high-dose GFR(250,500,1,000 mg/kg)groups by a random number table,10 mice in each group.The antidiarrheal activity was evaluated in castor oil-induced diarrhea mice model by evacuation index(El).In vitro,the effects of GFR(0.01,0.03,0.1,0.3,1,3,and 10 g/L)on the spontaneous contraction of isolated smooth muscle of rabbit jejunum and contraction of pretreated by Acetylcholine(ACh,10μmol/L)and KCl(60 mmol/L)were observed for 200 s.In addition,CaCl_(2)was accumulated to further study its mechanism after pretreating jejunal smooth muscle with GFR(1 and 3 g/L)or verapamil(0.03 and 0.1μmol/L)in a Ca^(2+)-free-high-K+solution containing ethylene diamine tetraacetic acid(EDTA).Results:GFR(500 and 1,000 mg/kg)significantly reduced El in castor oil-induced diarrhea model mice(P<0.01).Meanwhile,GFR(0.01,0.03,0.1,0.3,1,3,and 10 g/L)inhibited the spontaneous contraction of rabbit jejunum(P<0.05 or P<0.01).Contraction of jejunums samples pretreated by ACh and KCI with 50% effective concentration(EC50)values was 1.05(0.71-1.24),0.34(0.29-0.41)and 0.15(0.11-0.20)g/L,respectively.In addition,GFR moved the concentration-effect curve of CaCl_(2)down to the right,showing a similar effect to verapamil.Conclusions:GFR can effectively reduce diarrhea and inhibit intestinal contraction,and these antidiarrheal effects may be based on blocking L-type Ca^(2+) channels and muscarinic receptors.

Effects of Different Arbuscular Mycorrhizal Fungi on Growth and Protective Enzyme Activity of Glycyrrhiza uralensis

[Objective] This study aimed to investigate the effects of 5 kinds of arbuscular mycorrhizal fungi( Acaulospora mellea,Glomus mosseae,Glomus versiforme,Glomus aggregatum,Glomus etunicatum) on the growth and protective enzyme activity of Glycyrrhiza uralensis Fisch. [Method] The growth indicators and protective enzymes activity of glycyrrhiza plants inoculated and uninoculated with fungi were compared. [Result] The plant height,basal diameter,main root length,aboveground fresh weight,underground fresh weight,aboveground dry weight and underground dry weight of the inoculated glycyrrhiza plants were increased significantly compared with those in the control( non-inoculation) group. In the inoculated glycyrrhiza plants,the growth index was significantly increased compared with that in the control group( P <0. 05); the activities of superoxide dismutase( SOD) and peroxidase( POD) increased first and then decreased; and the activity of catalase( CAT) showed a continuous rising trend. The effects of different inoculants on the growth of G. uralensis were significantly different.[Conclusion]G. etunicatum,G. mosseae and G. aggregatum had a significant effect on the growth of G. uralensis,and were superior to other fungi in resisting the adverse environment.

Enhancing production of ergosterol in Pichia pastoris GS115 by over-expression of 3-hydroxy-3-methylglutaryl CoA reductase from Glycyrrhiza uralensis

The rate-limiting enzyme in the mevalonic acid(MVA)pathway which can lead to triterpenoid saponin glycyrrhizic acid(GA)is 3-hydroxy-3-methylglutaryl-CoA reductase(HMGR).In order to reveal the effect of copy number variation in the HMGR gene on the MVA pathway,the HMGR gene from Glycyrrhiza uralensis Fisch.(GuHMGR)was cloned and over-expressed in Pichia pastoris GS115.Six recombinant P.pastoris strains containing different copy numbers of the GuHMGR gene were obtained and the content of ergosterol was analyzed by HPLC.The results showed that all the recombinant P.pastoris strains contained more ergosterol than the negative control and the strains with 8 and 44 copies contained significantly more ergosterol than the other strains.However,as the copy number increased,the content of ergosterol showed an increasing–decreasing–increasing pattern.This study provides a rationale for increasing the content of GA through over-expressing the GuHMGR gene in cultivars of G.uralensis.

蒙古甘草与新疆红杆甘草再生植株体系的构建

为了建立蒙古甘草和新疆红杆甘草植株再生体系,本研究采用胚轴、叶片、子叶和茎段做外植体,探究了不同激素配比对二者愈伤组织诱导及再生植株生长情况的影响,筛选出愈伤组织诱导和植株再生的最适外植体及最佳培养基。实验结果显示:MS+2.0 mg/L NAA+2.0 mg/L 6-BA为愈伤组织诱导与植株再生诱导的最佳培养基,其中,胚轴与子叶是最佳诱导外植体,愈伤组织诱导率均高于90%。本研究建立的快繁体系可为甘草优良品种繁育及甘草生产提供理论参考与实践指导。

基于甘草全基因组序列的SSR分子标记开发

由于不同基原和产地的甘草有效成分差异较大,开发甘草特异分子标记对于种质资源鉴定意义重大。本研究以甘草全基因组序列为基础,检测到193 207个SSR位点,发生频率为73.52%。单核苷酸重复类型最多(60.73%),其次为二核苷酸(26.11%)和三核苷酸重复单元(10.95%)。分析重复序列位点发现,甘草SSR中包含284种类型重复基元,具有碱基偏好性,主要以A/T为主要重复单元,优势基元依次为A/T (58.28%)、AG/CT (10.48%)、AT/AT (10.48%)、AC/GT (5.12%)、AAT/ATT (3.57%)。优选140 294个重复位点设计出701 302对SSR引物,随机选取100对引物进行有效性检测,63对引物均扩增出目标条带,其中,12对引物能在材料间扩增出特异性条带,可用于对不同产地甘草进行区分。本研究开发的大量SSR标记不仅可用于甘草种质资源的鉴定分析,也将用于进一步的甘草分子标记辅助中。

Regulation of root secondary metabolites by partial root-associated microbiotas under the shaping of licorice ecotypic differentiation in northwest China

Interactions between plant hosts and their microbiotas are becoming increasingly evident,while the effects of plant communities on microbial communities in different geographic environments are poorly understood.Here,the differentiation of licorice plant ecotypes and the distribution of rootassociated microbiotas were investigated across five sampling sites in northwest China.The interactions between the environment,plant and microbial communities,and their effects on licorice root secondary metabolites,were elucidated.The plant community was clearly differentiated into distinct ecotypes based on genotyping-by-sequencing and was primarily driven by geographic distance and available soil nitrogen.The bulk soil and rootassociated microbiotas(rhizosphere soil and root endosphere)partially correlated with plant community,but all were significantly discriminated by plant clade.Moreover,these microbiotas were explained to different extents by distinct combinations of environment,geography,and plant community.Similarly,three structural equation models showed that licorice root secondary metabolites were complicatedly modulated by multiple abiotic and biotic factors,and were mostly explained by these factors in the rhizosphere model.Collectively,the results provide novel insights into the role of environment–plant–microbiota interactions in regulating root secondary metabolites.That should be accounted for when selecting appropriate licorice planting sites and management measures.