Effect of Color and Surface Roughness of Glass Ionomer Cements Submitted to Solutions Based on Ilex paraguariensis

ABSTRACT: The present study analyzed the color change (DE) and surface roughness (Ra) of glass ionomer cements (GICs) subjected to typical herbaceous South American beverage action - Ilex paraguariensis. 90 specimens (n = 10) were made: M1 - GIC Riva Light Cure® (SDI); M2 - GIC Riva Self Cure® (SDI); M3 - GIC Vitremer® (3M ESPE). After light curing/ polymerization of GICs, specimens were divided for being submitted to solutions for one hour a day for 21 days: S1 - Yerba Mate Pajarito Traditional (Pajarito); S2 - Erva Mate Chimarrão (Barão de Cotegipe); S3 - Water (control). Four DE and Ra readings were taken: T0 - initial; T1 - 7 days; T2 - 14 days and T3 - 21 days, with a spectrophotometer and a roughness meter respectively. The obtained data were submitted to ANOVA statistical analysis and Bonferroni. S1 and S2 at T1 and T2 there was no statistical difference between the materials, and at T3 Materials M1=M2, M1=M3, and M2>M3. For S3 at T1, T2 and T3 there was no difference between the materials. Analyzing surface roughness, there was no statistical significance. Concluded that all materials changed color with greater influence of time, but without changing surface roughness.

RESUMEN: En el presente estudio se analizó el cambio de color (DE) y la rugosidad superficial (Ra) de los cementos ionómero de vidrio (CIV) sometidos a la acción típica de las bebidas herbáceas sudamericanas - Ilex paraguariensis. Se realizaron 90 especímenes (n = 10): M1 - GIC Riva Light Cure® (SDI); M2 - GIC Riva Self Cure® (SDI); M3 - GIC Vitremer® (3M ESPE). Después de la fotopolimerización/ polimerización de los CIV, los especímenes se dividieron y fueron sometidos a soluciones durante una hora al día durante 21 días: S1 - Yerba Mate Pajarito Tradicional (Pajarito); S2 - Erva Mate Chimarrão (Barão de Cotegipe); S3 - Agua (control). Se tomaron cuatro lecturas de DE y Ra: T0 - inicial; T1 - 7 días; T2 - 14 días y T3 - 21 días, con espectrofotómetro y rugosímetro respectivamente. Los datos obtenidos fueron sometidos a análisis estadístico ANOVA y Bonferroni. S1 y S2 en T1 y T2 no hubo diferencia estadística entre los materiales y en T3 Materiales M1=M2, M1=M3 y M2>M3. Para S3 en T1, T2 y T3 no hubo diferencia entre los materiales. Al analizar la rugosidad de la superficie, no hubo significación estadística. En conclusion, todos los materiales cambiaron de color con mayor influencia del tiempo, pero sin cambiar la rugosidad de la superficie.

High polarity extracts obtained from Ilex guayusa Loes. leaves, exhibit antioxidant capacity / Extractos de alta polaridad obtenidos a partir de las hojas de Ilex guayusa Loes. exhiben capacidad antioxidante

Background: Ilex guayusa Loes. belongs to the family Aquifoliaceae. It is an evergreen tree native to the Amazon region. According to traditional uses, it is used as a diaphoretic, narcotic, purgative, among other uses. Objective: To evaluate the antioxidant capacity, quantify phenols and total flavonoids of extracts obtained from Ilex guayusa leaves. Methods: Total ethanolic extract was obtained using Soxhlet-type equipment, with subsequent liquid/liquid fractionation with solvents of different polarities (petroleum ether, dichloromethane, and ethyl acetate); were reacted with 2,2-diphenyl-1-picrylhydrazyl (DPPH•) and 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS•+), determining the inhibitory concentration 50 (IC50) and relative antioxidant activity (%AAR), with subsequent statistical analysis (ANOVA and Tukey's post-hoc). Total phenols and flavonoids were quantified using the Folin-Ciocalteu method and complex formation with AlCl3, respectively. Results: The ethanolic extract of leaves showed the highest antioxidant activity in both the DPPH• and ABTS•+ methods with IC50 values of 4.58 and 3.82 ppm, respectively. Conclusions: According to the results obtained in this study, it was possible to corroborate that the ethanolic extract (EE) obtained from the leaves of I. guayusa showed the highest antioxidant capacity, by the DPPH• and ABTS•+ methods. Additionally, it was possible to relate this capacity to the high flavonoids content present in this extract. Based on the above and accompanied by future studies, the Ilex guayusa species could be proposed as an important source of antioxidant compounds with possible application in medicine and the food industry

Antecedentes:Ilex guayusa Loes. pertenece a la familia Aquifoliaceae. Es un árbol perenne, nativo de la región amazónica. De acuerdo con los usos tradicionales es empleada como diaforético, narcótico, purgativo, entre otros usos. Objetivo: Evaluar la capacidad antioxidante, cuantificar fenoles y flavonoides totales de extractos obtenidos a partir de las hojas de Ilex guayusa. Métodos: El extracto etanólico total se obtuvo mediante un equipo tipo "Soxhlet", con posterior fraccionamiento líquido/líquido con solventes de diferentes polaridades (éter de petróleo, diclorometano y acetato de etilo); se hicieron reaccionar con 2,2-difenil-1-picrilhidracilo (DPPH•) y ácido 2,2'-azino-bis-3-etilbenzotiazolin-6-sulfónico (ABTS•+), determinando la concentración inhibitoria 50 (IC50) y la actividad antioxidante relativa (%AAR), con posterior análisis estadístico (ANOVA y post-hoc de Tukey). Los fenoles y flavonoides totales se cuantificaron con el método de Folin-Ciocalteu y el de formación de complejos con AlCl3, respectivamente. Resultados: El extracto etanólico de las hojas fue el que presentó mayor actividad antioxidante tanto en el método DPPH• como en el ABTS•+ con valores de IC50 de 4,58 y 3,82 ppm, respectivamente. Conclusiones: De acuerdo con los resultados obtenidos en este estudio, se pudo corroborar que el extracto etanólico obtenido de las hojas de I. guayusamostró la mayor capacidad antioxidante, por los métodos DPPH• y ABTS•+. Adicionalmente, se pudo relacionar esta capacidad con el alto contenido de flavonoides presentes en este extracto. Con base en lo anterior y acompañado de futuros estudios, la especie I. guayusa podría proponerse como una importante fuente de compuestos antioxidantes con posible aplicación en los campos de la medicina y la industria alimentaria

Consumption of mate Ilex paraguariensis: a folk beverage with antioxidant power against myocardial ischemic injury

The effects of the aqueous extract of Ilex paraguariensis (Ip)and the flavonoid quercetin were tested during the induction of in vivomyocardial ischemia/ reperfusion in Rattus norvegicus. The antioxidant power of the extract and quercetin were chemically determined. The experimental groups were: control, ischemia/reperfusion induction, Iporal treatment, Iporal treatment and ischemia /reperfusion, quercetin oral treatment, and quercetin oral treatment and ischemia/reperfusion. Rats were anesthetized with sodium thiopental and xylazine via intraperitoneal injection and subsequently underwent 15 minutes of ischemia followed by 15 minutes of reperfusion. Ischemia was promoted by tying the left anterior descending coronary artery. Areas of risk and infarction were stained by intravenous Evans blue and triphenyl tetrazolium chloride. Reactive oxygen species (ROS), antioxidant capacity against peroxylradicals, and lipid peroxidation of the myocardium were quantified. A significant reduction in areas of risk and infarction was detected in the ischemic myocardium treated with Ipand quercetin; ROS generation and lipid peroxidation were significantly reduced, and the antioxidant capacity was elevated. Oral administration of Ippromoted antioxidant benefits in the myocardium during ischemia and reperfusion, which reduced infarction. We suggest that Mate (a hot drink made from steeped dried leaves of Ip) consumption is a potential cardioprotective habit of indigenous people from southern South American countries, which must be better understood scientifically and ethnographically.

Content determination of eight phenols in Ilex hainanensis harvested at different time by HPLC-DAD / 中国中药杂志

This study intends to develop a high performance liquid chromatography-diode array detection(HPLC-DAD) method for simultaneous determination of chlorogenic acid, 2-hydroxymethyl-3-hydroxyl-1-butene-4-O-β-D-(6″-O-caffeoyl)-glucopyranoside, pubescenoside B, huazhongilexone-7-O-β-D-glucopyranoside, rutin, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C in Ilex hainanensis. The HPLC conditions are as follows: Waters XBridge C_(18 )column(4.6 mm×250 mm, 5 μm), mobile phase of 0.5% formic acid in water(A)-acetonitrile(B), gradient elution(0-8 min, 5%-12% B; 8-18 min, 12%-18% B; 18-30 min, 18%-25% B; 30-40 min, 25%-30% B; 40-42 min, 30%-80% B; 42-45 min, 80% B) at the flow rate of 0.8 mL·min~(-1), detection wavelengths of 282, 324, and 360 nm, column temperature of 25 ℃, and injection volume of 5 μL. The content of the 8 phenols in 8 samples was 0.30-6.29, 0.29-3.27, 0.15-10.4, 0.51-5.85, 0.49-9.02, 0.51-4.68, 1.93-13.4, and 0.87-5.95 mg·g~(-1), respectively. Moreover, the content of phenols in the samples collected in October was higher than that of samples harvested in other months. The established method is accurate and sensitive for the determination of phenols in I. hainanensis, which is useful for the quality improvement of this herbal medicine.

Triterpenoids from leaves of Ilex guayusa / 中国中药杂志

The chemical constituents from the leaves of Ilex guayusa were investigated. Sixteen triterpenoids were isolated from the 95% ethanol extract of dried leaves of I. guayusa by silica gel, Sephadex LH-20, and ODS column chromatographies and semi-prepa-rative HPLC. Those triterpenoids were identified by NMR, HR-MS, and literature analysis: 3β-hydroxy-11α,12α-epoxy-24-nor-urs-4(23)-ene-28,13β-olide(1), 3β-hydroxy-24-nor-4(23),12-oleanadien-28-methyl ester(2), oleanolic acid(3), 3β,28-dihydroxy-12-oleanene(4), 2α,3β-dihydroxy-11α,12α-epoxy-24-'nor-olean-4(23)-ene-28,13β-olide(5), ursolic acid(6), 3β,23-dihydroxy ursolic acid(7), 3β,28-dihydroxy-12-ursene(8), 3β-28-nor-urs-12-ene-3,17-diol(9), 3β-hydroxyurs-11-ene-28,13β-olide(10), 13β,28-epoxy-3β-hydroxy-11-ursene(11), 3β-hydroxy-28,28-dimethoxy-12-ursene(12), 3β-hydroxy-24-nor-urs-4(23),12-dien-28-oic acid(13), 3β-hydroxy-24-nor-urs-4(23),12-dien-28-methyl ester(14), 2α,3β-dihydroxy-11α,12α-epoxy-24-nor-urs-4(23)-ene-28,13β-olide(15) and 2α,3β-dihydroxy-11α,12α-epoxy-24-nor-urs-4(23),20(30)-dien-28,13β-olide(16). Compounds 1-2 were new compounds, and compounds 4-5, 7 and 9-16 were isolated from I. guayusa for the first time.

New triterpenoid saponins from the leaves of Ilex chinensis and their hepatoprotective activity / 中国天然药物

Seven new triterpenoid saponins, including five ursane-type saponins, ilexchinenosides R-V (1-5), and two oleanane-type saponins, ilexchinenosides W-X (6-7), with four known triterpenoid saponins (8-11) were isolated from the leaves of Ilex chinensis. Their structures were elucidated by comprehensive spectroscopic 1D and 2D NMR and HR-ESI-MS data. Their sugar moieties were determined by HPLC analysis compared with standards after hydrolysis and derivatization. Compounds 1, 2, 4, 9 and 10 exhibited potential hepatoprotective activity against N-acetyl-p-aminophenol (APAP)-induced HepG2 cell injury in vitro.

Elevated Iron Chelating Activity of Ilex paraguariensis Leaf Infusion: in vitro and in vivo Pilot Studies

Abstract This study presents an Ilex paraguariensis leaf infusion with important potential as natural iron-chelating. The impact of infusion time and the water volume to obtain an Ilex paraguariensis leaf infusion with high phenolic content and iron chelating activity, such as the stability of these proprieties in the storage time and temperature (immediately and after 24 h at 8 and 25 (C) were assessed. The acute consumption effect of this infusion to reduce iron absorption in vivo was also evaluated. A preliminary crossover trial with volunteers that ingested a meal containing non-haem iron (11.4 mg) with the treatments: Ilex paraguariensis leaf infusion with the highest phenolic content and iron chelating activity (200 mL) or control (200 mL water). Blood samples were withdrawn before and 1, 2, 3 and 4 h after the meal for serum iron measurement. The highest phenolic content (18.1 mg/mL) and iron chelating activity ((100%) were observed for 10 min infusion time using 30 g leaves/300 mL water. Storage at 8 or 25 (C for 24 h decreased total phenolics and di-caffeoylquinic acids by 23.5% and 25.5%, respectively (p< 0.05), without affecting the iron-chelating activity due to a saturating chelating effect at 3.34 mg/mL phenolic content. Inhibition of the iron absorption in vivo by infusion was 78% considering the iron recovery at peak maximum. The in vitro and preliminary in vivo results showed a functional property of the Ilex paraguariensis leaf infusion that may be useful for adjuvant management of iron overload diseases.

Morphophysiological characterization of Ceratocystis fimbriata isolates from yerba mate / Caracterização morfofisiológica de isolados de Ceratocystis fimbriata de erva-mate

ABSTRACT: This study aimed to morphologically characterize the isolates of Ceratocystis fimbriata from yerba mate and to evaluate the effect of culture medium and temperature on mycelial growth and sporulation of C. fimbriata. For the morphological characterization of the 11 monosporic isolates of the fungus, slides were prepared to determine the dimensions of the sexual and asexual structures of the fungus. Four experiments were conducted to evaluate the mycelial growth and to evaluate the sporulation of C. fimbriata in different culture mediums and temperatures. The isolates of C. fimbriata from yerba mate showed perithecia with brown to black necks, divergent ostiolar hyphae, hatshaped hyaline ascospores, single-celled, cylindrical endoconidia, and globular to ovoid aleurioconidia. PDA and V8-agar media showed the highest mycelial growth. The average optimum temperature for mycelial growth and sporulation of isolates of C. fimbriata of yerba mate were 22.5 and 22.4 ºC, respectively.

RESUMO: Este estudo teve como objetivo caracterizar morfologicamente os isolados de Ceratocystis fimbriata e avaliar o efeito do meio de cultura e da temperatura no crescimento micelial e na esporulação de C. fimbriata. Para a caracterização morfológica dos 11 isolados monospóricos do fungo foram preparadas lâminas para determinar as dimensões das estruturas sexuadas e assexuadas do fungo. Quatro experimentos foram conduzidos para avaliar o crescimento micelial e esporulação of C. fimbriata em diferentes meios de culturae temperaturas. Os isolados de C. fimbriata de erva-mate apresentaram peritécios com pescoço de marrom a preto, hifa ostiolar divergente, ascósporos hialinos em formato de chapéu, endoconídios unicelulares, cilíndricos, e aleuroconídios com formato globoso a ovoide. Os meios de cultura PDA e V8-ágar apresentaram os maiores crescimentos miceliais. A temperatura ótima média para crescimento micelial e esporulação dos isolados de C. fimbriata de erva-mate foram de 22,5 e 22,4 ºC, respectivamente.

How do clonal plantlets of mate respond to different substrate compositions and shading levels? / Como as mudas clonais de erva-mate respondem a diferentes composições de substrato e níveis de sombreamento?

Mate (Ilex paraguariensis A. St.-Hil) is an arboreal species of great economic and socio-environmental importance in South American countries. This specie presents several difficulties during seminal propagation, and studies related to plantlet production by vegetative propagation are fundamental for obtaining homogenous mate plantations with high leaf productivity. Therefore, the present study aimed to evaluate the effects of substrate and shading levels on the morphophysiological quality of mate plantlets produced by mini-cuttings. Rooted mini-cuttings of four mate clones were cultivated on commercial substrate, subsurface soil and vermiculite (2:1:1 v/v/v), commercial substrate and subsurface soil (2:1 v/v) or subsurface soil, cattle manure and carbonized rice hulls (2:1:1 v/v/v). After 120 days, the plantlets were randomly distributed on benches to evaluate the effects of 50 and 80% shading screens. Regardless of the clone, the commercial substrate and subsurface soil composition allowed plantlet production with satisfactory development of both aerial part and root system at 120 days of cultivation. Clone 06SM17 produced plantlets with high averages of stem diameter, shoot height, number of leaves, total length, surface area, total volume of roots, and number of root tips. Both shade levels resulted in similar stem diameters, shoot heights, numbers of leaves, and a, b, and total chlorophyll indices. Clonal mate plantlets with satisfactory morphophysiological quality are produced in commercial substrate and subsurface soil (2:1 v/v) under 50 and 80% shading.

A erva-mate (Ilex paraguariensis A. St.-Hil) é espécie arbórea de grande importância econômica e socioambiental para os países da América do Sul, que apresenta diversas dificuldades de propagação utilizando sementes, sendo os estudos relacionados à produção de mudas dessa espécie por propagação vegetativa fundamentais para a obtenção de ervais homogêneos e com alta produtividade foliar. Diante desse contexto, o presente estudo objetivou avaliar o efeito das composições de substratos e dos níveis de sombreamento na qualidade morfofisiológica das mudas de erva-mate produzidas por miniestaquia. Miniestacas enraizadas de quatro clones foram cultivadas em substrato comercial, solo de subsolo e vermiculita (2:1:1 v/v/v), substrato comercial e solo de subsolo (2:1 v/v) ou solo de subsolo, esterco bovino e casca de arroz carbonizada (2:1:1 v/v/v). Após 120 dias, as mudas foram aleatoriamente distribuídas em bancadas para avaliar o efeito do sombreamento de 50 e 80%. Independentemente do clone, a composição substrato comercial e solo de subsolo permitiu a produção de mudas com desenvolvimento satisfatório tanto da parte aérea, quanto do sistema radicular aos 120 dias de cultivo. O clone 06SM17 possibilitou a produção de mudas com altas médias de diâmetros de colo, altura da parte aérea, número de folhas, comprimento total, área superficial, volume total de raízes e número de extremidades das raízes. Os níveis de sombreamento não afetaram o diâmetro de colo, a altura da parte aérea, o número de folhas e os índices de clorofila a, b e total. Mudas clonais de erva-mate com satisfatória qualidade morfofisiológica podem ser produzidas em substrato comercial e solo de subsolo (2:1 v/v) em ambiente com 50% ou 80% sombreamento.

Antioxidant And Hepatoprotective Effect Of Quercus Ilex Leaves Extract In Ethanol Induced Liver Damage In Wistar Rats

Objective: This study was undertaken to investigate the hepatoprotective and antioxidant activity of Quercus ilex leaves extract (QILE) on ethanol-induced toxicity in Wistar rats. Methods: Hepatotoxicity was induced by administering ethanol (40%) at a dose of 7.9 gm/kg/day; p. o. (1:1 of ethanol in olive oil) for 28 d. Silymarin 100 mg/kg/day; p. o. was used as a standard drug. The whole study was divided into a prophylactic and curative study. In the prophylactic study, the Silymarin and QILE (test drug) 100, 200, and 400 mg/kg Body Weight(BW) given orally one hour before administration of 40% ethanol administration for 28 d. In the curative study, 7 d of treatment of Silymarin and QILE 200 and 400 mg/kg BW was given orally after 28 d of ethanol administration to different groups. Results: Hepatoprotectivity was confirmed by the highly significantly (p<0.001) restoration of elevated biochemical parameters like SGPT, SGOT, ALP, TB, and highly significantly (p<0.001) depleted Albumin and Total protein levels by 200 mg/kg BW QILE in comparison to the positive control group. QILE 200 mg/kg highly significantly (p<0.001) raised the antioxidants by draining the elevated oxidative stress markers in comparison of positive control group. At dose levels QILE 200 mg/kg, significant (p<0.05) protection from loss in body weight and in liver weight was found when the comparison was done with the positive control group. Histopathology revealed that QILE 200 mg/kg reduced the markers of cell necrosis. Conclusion: Present study revealed that Quercus ilex leaves have antioxidant and hepatoprotective activity due to its chemical constituents.

Research Process on Pharmacological Effects and Clinical Applications of Commonly Used Medicinal Plants of Genus Ilex / 中国实验方剂学杂志

Four commonly used medicinal plants of genus Ilex, including I. pubescens, I. asprella, I. rotunda, and I. latifolia, have been extensively used in clinic. The Ilex plants contain triterpenes and their glycosides, flavones, phenols and other compositions, and their have the pharmacological activities of anti-inflammatory, antibacterial, antithrombosis, cardiovascular and cerebrovascular protection, etc. Among them, I. pubescens has significant pharmacological activities in cardiovascular and cerebrovascular diseases, but there are few studies on clinical application. There are a few literature reports on the anti-inflammatory, lowering blood lipids, anti-oxidation, anti-bacterial and anti-viral activities of I. asprella, which is clinically used in the treatment of upper respiratory diseases and infectious diseases. I. rotunda has strong activities of antithrombosis and anti-inflammatory, it is clinically used for the treatment of chronic pelvic inflammatory disease. I. latifolia shows activities of hypolipidemic, antitumor, antibacterial and antioxidant, and its leaves are often used in the treatment of hypertension. This review summarized the research progress on the pharmacological effects and clinical applications of commonly used medicinal plants of this genus in recent years, in order to provide basis for their clinical applications, and provide useful reference for the further development of the research direction and the development of medicines and health products.

Inhibitory effect of BF523 from Ilex hainanensis on ox LDL-induced foam cells formation / 中国中药杂志

Cardio-cerebral vascular disease induced by atherosclerosis is a serious cause of human health. The pathogenesis of AS is very complex,and the oxidized low-density lipoprotein( ox LDL) induced foam cells formation is considered to be the most important cytological change in AS. Based on the definition of " TCM chemical biology",we clarified the chemical composition of Ilex hainanensis,the effective substances of I. hainanensis on the activity of anti-AS were screened. Then we found that saponin BF523 had the good inhibitory effect on foam cell formation. In this research,we studied the BF523 as the research object to clarify the molecular target of the active compound of I. hainanensis by foam cell formation model. The results showed that BF523 significantly inhibited the oxidation of ox LDL-induced macrophage foaming and decreased the lipid content in macrophages. BF523 had inhibited the phagocytosis of ox LDL in macrophages by reducing the mRNA and protein levels of scavenger receptor CD36,thereby inhibiting the occurrence and development of AS. These findings not only clarified the mechanism of the inhibition of foam cell formation by saponin BF523,but also provided a useful exploration for the enrichment of the theory of " TCM chemical biology".

Discovery and functional analysis of a cytochrome P450 gene involved in C-28 oxidation of the triterpenoids from Ilex pubescens / 中国药科大学学报

@#The main components of the medicinal plant Ilex pubescens Hook. et Arn. are the pentacyclic triterpenoids with various chemical structures. The modification of the triterpenoid skeleton is closely related to a class of key enzymes downstream of the biosynthetic pathway, cytochrome P450 monooxygenase(CYP450). In this study, unigenes with a length of more than 1100 bp were retrieved from the previously obtained Ilex pubescens transcriptome data, and a cytochrome P450 gene(Unigene 0036170)proposed to have the oxidative function of pentacyclic triterpene C-28 was screened through phylogenetic tree analysis. The gene was named as IpAO2. The open reading frame of IpAO2 is 1443 bp, encoding 480 amino acids. RNA was extracted from the tender leaves of Ilex pubescens, and cDNA was obtained by reverse transcription using the RNA as the template. The specific primers AO2-F and AO2-R were designed according to the DNA sequence of IpAO2, and the IpAO2 gene was amplified by PCR using high fidelity enzyme. The IpAO2 gene was ligated into the pEASY cloning vector to obtain pEASY-IpAO2 plasmid. The ligation product was transformed into E. coli Trans1-T1 competent cells and positive clones were screened. The pESC-TRP plasmid was digested with restriction endonucleases to obtain a linearized vector. Primers V-AO2-F and V-AO2-R were designed and the DNA fragment was amplified by PCR using the pEASY-IpAO2 plasmid as template. The DNA fragment was ligated to the vector pESC-TRP to obtain pESC-TRP-IpAO2 recombinant plasmid. The pESC-TRP-IpAO2 plasmid was transformed into a strain of S. cerevisiae capable of efficiently synthesizing amyrin, and the expression of IpAO2 protein was induced with galactose. After extracting the total protein of recombinant S. cerevisiae, Western blot analysis was carried out through the 6×His antigen tag linked to the C-terminus of the target protein. The results showed that the recombinant protein was synthesized in accordance with the expected size. After 7 days of induction and cultivation, the cells of recombinant yeast were collected. The metabolites of cells were extracted and detected by GC-MS after silanization. Through GC-MS analysis of recombinant yeast metabolites, it was found that the protein encoded by the IpAO2 gene can oxidize α-amyrin and β-amyrin to ursolic acid and oleanolic acid, respectively, indicating that IpAO2 is a pentacyclic triterpene C-28 oxidase gene. Meanwhile, the transmembrane domain and protein tertiary structure of the IpAO2 were predicted using the Phyre2 online tool, the results showing that IpAO2 is a transmembrane protein. This study is of great significance to elucidate the biosynthetic mechanism of triterpenoids of Ilex pubescens, and lays a foundation for the further utilization of the metabolic engineering to produce triterpenoids of Ilex pubescens in S. cerevisiae.

Quantitative and Fingerprint of Ilex pubescens Slices / 中国实验方剂学杂志

Objective:To establish HPLC-UV fingerprints of Ilex pubescens pieces,and simultaneously determine two components in 46 batches of I. pubescens in pieces of I. pubescens saponin A1 and B1,in order to provide a reference for the quality standard of I. pubescens slices. Method:Methanol was used to extract the I. pubescens saponin samples,and the extracts were measured by HPLC-UV with the absorption wavelength at 210 nm. Kromasil C18 column (4.6 mm×250 mm,5 μm) was used for determining the extracts at a flow rate of 1.0 mL·min-1. The mobile phase condition was acetonitrile-0.1% phosphoric acid aqueous solution with gradient mode. The chromatographic fingerprint similarity evaluation system of traditional Chinese medicine (2012 edition) was used to analyze I. pubescens fingerprints. SPSS 20.0 software was used to cluster the peak area of common peaks. Principal component analysis was performed to reduce the dimension of common peaks. Result:There were great differences between the root and stem parts in I. pubescens fingerprints. The fingerprints of roots and stems of I. pubescens were established respectively,cluster results assorted the roots of I. pubescens into three categories andthe branches of I. pubescens into two categories. The integrity and difference of I. pubescens decoction pieces from different parts and places of origin were compared,and the principal component analysis was performed to screen out the common components that played a decisive role in fingerprint of I. pubescens pieces. And the common peaks were determined. The content of saponin A1 and saponin B1 in Radix I. pubescens were determined. Conclusion:The established I. pubescens fingerprints and content determination methods are simple and suitable. Cluster analysis and principal component analysis are used to screen out the key components of quality control of I. pubescens. The results can provide references for quality control of I. pubescens.

Cloning and in vitro expression of sulfotransferase genes from Ilex asprella / 中草药

Objective: The medicinal plant Ilex asprella contains various biologically active sulfonated triterpenoids and triterpenoid saponins. The objective of this study was to clone sulfotransferases (ST) from I. asprella and facilitate the elucidation of the sulfonation mechanism therein. Methods: The physicochemical properties, secondary structure and tertiary structure of two ST candidates of IaST1 and IaST2 were forecasted and analyzed using related software. IaST1 and IaST2 were cloned by RT-PCR and expressed in Escherichia coli. Results: The open reading frame (ORF) of IaST1 was 1 002 bp long and encoded a protein of 333 amino acids with the calculated molecular weight of 55 500, while the ORF of IaST2 was 993 bp long and encoded a protein of 330 amino acids with the calculated molecular weight of 54 700. Both contain the five highly conserved domains of ST. Phylogenetic analysis revealed that IaST1 and IaST2 were genetically closely related and clustered together with flavonol C-3 ST of Flaveria bidentis. Conclusion: This is the first report on the cloning of STs from I. asprella, providing an important basis for further investigations into their functions and roles in the biosynthesis of sulfonated triterpenoids in I. asprella.

Comparison of Low-polarity Volatile Constituents in Supercritical CO2 Extract from the Roots and Stem of Ilex asprella and Its Effects on the Proliferation of IEC- 6 Cells / 中国药房

OBJECTIVE： To compare low-polarity volatile constituents in supercritical CO2 extract from the roots and stem of Ilex asprella and its effects on the proliferation of IEC-6 in vitro， and to provide reference for making full use of wild resources of I. asprella and expanding its medicinal parts. METHODS： The low-polarity volatile constituents were extracted from the root and stem of I. asprella with supercritical fluid CO2 extraction（SFE-CO2）. The chemical constituents were analyzed by GC-MS. IEC-6 cells were treated with different concentrations of supercritical CO2 extracts （0， 1， 5， 10， 20， 40， 60， 80， 100 μg/mL） from roots or stems of I. asprella. MTT assay was used to detect the relative viability， and cell proliferation curve was drawn and EC50 of each extract were calculated. RESULTS： Sixty-two and forty-six low-polarity volatile constituents were identified from supercritical CO2 extract in the roots and stem of I. asprella with GC-MS； there were 24 common constituents totally， mainly including pelargonic acid（14.18％ and 6.14％），octanoic acid（10.59％ and 4.35％），hexanoic acid（8.63％ and 10.86％），paeonol（7.79％ and 6.00％），2-methyl-3-phenyl-propanal（6.3％ and 0.58％），acetic acid（1.72％ and 33.77％） in root and stem， respectively. The results of cell culture in vitro showed that when the concentration of supercritical CO2 extract from the roots and stems of I. asprella was lower （≤60 μg/mL）， it could significantly promote the proliferation of IEC-6 cells and their EC50 were 16.35， 20.20 μg/mL， respectively； when the concentration of the extract was higher （≥80 μg/mL）， it showed cytotoxicity and inhibited the proliferation of IEC-6 cells. CONCLUSIONS： There are similar species of volatile constituents in roots and stems of I. asprella and similar in vitro bioactivity of the supercritical CO2 extracts to IEC-6 cells. The short-chain fatty acids may be the active ingredient to promote cell proliferation， while paeonol may be the cytotoxic active ingredient.

Anti-nociceptive and Anti-inflammatory Properties of Ilex latifolia and its Active Component, 3,5-Di-caffeoyl Quinic Acid Methyl Ester

The present study was conducted to investigate anti-nociceptive and anti-inflammatory effects of the leaves of Ilex latifolia Thunb (I. latifolia) in in vivo and in vitro. Writhing responses induced by acetic acid and formalin- and thermal stimuli (tail flick and hot plate tests)-induced pain responses for nociception were evaluated in mice. I. latifolia (50 – 200 mg/kg, p.o.) and ibuprofen (100 mg/kg, p.o.), a positive non-steroidal anti-inflammatory drug (NSAID), inhibited the acetic acid-induced writhing response and the second phase response (peripheral inflammatory response) in the formalin test, but did not protect against thermal nociception and the first phase response (central response) in the formalin test. These results show that I. latifolia has a significant anti-nociceptive effect that appears to be peripheral, but not central. Additionally, I. latifolia (50 and 100 µg/mL) and 3,5-di-caffeoyl quinic acid methyl ester (5 µM) isolated from I. latifolia as an active compound significantly inhibited LPS-induced NO production and mRNA expression of the pro-inflammatory mediators, iNOS and COX-2, and the pro-inflammatory cytokines, IL-6 and IL-1β, in RAW 264.7 macrophages. These results suggest that I. latifolia can produce antinociceptive effects peripherally, but not centrally, via anti-inflammatory activity and supports a possible use of I. latifolia to treat pain and inflammation.

The anti-obesity potential of Ilex paraguariensis: results from a meta-analysis

The dried leaves of Ilex paraguariensis, also known as yerba mate or chimarrão, are used to produce infusions consumed by over 1 million people worldwide, being a traditional drink in several South American countries, particularly, Argentina, Brazil, Uruguay, and Paraguay. The positive effects of I. paraguariensis in obesity have been described and observed in some clinical trials both in animals and in humans. The main goal of this work was to perform a systematic review, complying with the PRISMA (Preferred Reported Items for Systematic Reviews and Meta-Analysis) recommendations, followed by a meta-analysis and Trial Sequential Analysis (TSA), to summarize the results of the recent clinical trials, clarifying the anti-obesity potential of I. paraguariensis. The search for this systematic review was performed on several electronic databases (Pubmed, Web of Science, Scopus, SciELO and Cochrane Library) in July 2017. A total of 22 clinical trials were considered for the quantitative synthesis of the data, totalizing 726 patients enrolled. In conclusion, this systematic review demonstrated the positive effects of I. paraguariensis consumption on the reduction of body weight, body mass index (BMI), and waist circumference of patients, showing its anti-obesity potential.

Plant growth, nutrients and potentially toxic elements in leaves of yerba mate clones in response to phosphorus in acid soils

ABSTRACT Native to subtropical region of South America, yerba mate is responsive to P under some conditions, but the degree of influence of genetic and soil on the growth and composition of the leaf is unknown. The aim of study was to evaluate plant growth, nutrients and potentially toxic elements in leaves of yerba mate clones in response to P application in acid soils. In greenhouse condition, two yerba mate clone seedlings were grown (210 days) in pots, each clone in a completely randomized design in factorial scheme (with and without P; four acid soils). The elemental composition of leaves and the growth of plants were determined. Phosphorus promoted plant growth, but this was not accompanied by increased P in leaf tissue in all conditions tested. The P effect on the elemental composition varied: decrease/null (N, K, Mg, Mn, Cu, Ni, B, Mo, Al, Cd); increase/null (C/N, C, Ca, Fe, V); increase/decrease/null (Zn, Ba, Pb) and; null (Cr). The soils affect the elemental composition of the leaves, especially Mn, with accumulation greater than 1000 mg kg-1. The Ba, Pb, Al and Zn in the leaves varied among clones. Yerba mate response to P was affected by edaphic and plant factors.

Antipyretic, Anti-inflammatory and Analgesic Effects of Yinhaojiere Mixture and Antipyretic Effect of Ilex Asprella / 中国药师

Objective: To study the antipyretic, anti-inflammatory and analgesic effects of Yinhaojiere mixture and the antipyretic effect of Ilex asprella. Methods: The experiments of mice ear edma induced by xylene, the increase of capillary permeability by acetic acid and the sterilized cotton caused rat groin granulation tissue hyperplasia were used to study the anti-inflammatory effect of Yinhao-jiere mixture; the hot-plate tests and the acetic acid writhing test were applied to research the analgesic effect of Yinhaojiere mixture. In addition, the dry yeast induced fever in rats was employed to explore the efficacy of antipyretic agents in Yinhaojiere mixture and Ilex asprella water extract. Results: Yinhaojiere mixture could obviously inhibit the inflammation of the peritoneal cavity of mice ( P <0. 05). The inhibition of Yinhaojiere mixture against ear swelling induced by p-xylene was significant. Yinhaojiere mixture could sig-nificantly increase the pain threshold. Especially, Yinhaojiere mixture had significant antipyretic effect (P<0. 05). In the anti-inflam-matory, analgesic and antipyretic experimetns, there were no significant differences between the middle and high dose groups of Yinha-ojiere mixture and indomethacin group (P>0. 05). Conclusion: Yinhaojiere mixture with the Ilex asprella as the main ingredient has notable anti-inflammatory, analgesic and antipyretic effects, and Ilex asprella exhibits anti-inflammatory, analgesic effects, and its anti-pyretic effect is more significant.