Conjugates of several lupane,oleanane, and ursane triterpenoids with the antituberculosis drug isoniazid and pyridinecarboxaldehydes

Conjugates of betulinic, oleanolic, ursolic, and glycyrrhetic acids and several of their derivatives with the antituberculosis drug isoniazid and 3- and 4-pyridinecarboxaldehydes were synthesized.     

Analysis of four pentacyclic triterpenoid acids in several bioactive botanicals with gas and liquid chromatography and mass spectrometry detection

Several pentacyclic triterpenoid acids including betulinic, oleanolic, and ursolic acids were reported to have health beneficial properties such as antiviral and anti‐inflammatory properties, as well as the capability to inhibit “in vitro” the development of various cancer cell types. For this reason betulinic, oleanolic, and ursolic acids are used as neutraceuticals. For the analysis of the pentacyclic triterpenoid acids in complex plant materials, an improved scheme was developed, involving a qualitative screening using silylation and gas chromatography with mass spectrometry analysis, followed by quantitation using a novel liquid chromatography with tandem mass spectrometry procedure. The use of the two methods provides more reliable information regarding the plant materials with unknown composition. Besides betulinic, oleanolic, and ursolic acids that were analyzed, by this procedure a fourth pentacyclic triterpenoid acid was identified and quantitated that was not previously reported to be present in plants. This acid has been identified as 3β‐3‐hydroxy‐lupa‐18,20(29)‐dien‐28‐oic acid. The newly identified acid has a structure as a derivative of lupane, although lupane with a double bond in the 18‐position was not previously reported as present in plants. The new liquid chromatography with tandem mass spectrometry procedure developed for this study offers a very low limit of quantitation, excellent precision, and robustness. Rosemary was found to contain the largest levels of pentacyclic triterpenoid acids among all the analyzed botanicals.     

Mass spectrometric identification of oleanolic and ursolic acids

p-Toluidides of 3-acetoxy ursolic and 3-acetoxy oleanolic acids were prepared. On the basis of different intensities of the m/e 452 and 453 peaks the identification of oleanolic and ursolic acids is possible.     

Simultaneous determination of five triterpene acids in rat plasma by liquid chromatography–mass spectrometry and its application in pharmacokinetic study after oral administration of Folium Eriobotryae effective fraction

Folium Eriobotryae effective fraction (FEA), the extract of Folium Eriobotryae, had been used as anti‐hyperglycemia and anti‐hyperlipemia medicine in China. A previous study indicated that euscaphic acid, maslinic acid, corosolic acid, oleanolic acid and ursolic acid, the five structurally similar triterpene acids (containing two groups of structural isomers), are the major components of FEA. In the present study, we developed a specific and reliable LC‐MS method for simultaneous determination of the five triterpene acids in rat plasma, and further investigated their pharmacokinetic properties after oral administration of FEA. Following a simple sample preparation, chromatographic separation was achieved on a C₁₈ column with a mobile phase composed of methanol–0.1% ammonium acetate (80:20, v/v). Quantification was achieved by monitoring the selected ions at m/z 487.6 for euscaphic acid, m/z 471.5 for maslinic acid and corosolic acid, m/z 455.5 for oleanolic acid and ursolic acid and m/z 469.5 for internal standard. The method was validated to be specific, accurate and precise over the concentration ranges of 10–3000 ng/mL with limits of detections of 5 ng/mL for the five triterpene acids. Finally, the method was successfully applied to the pharmacokinetic study of the five structurally similar triterpene acids in rats after oral administration of FEA. Copyright © 2015 John Wiley & Sons, Ltd.     

Determination of oleanolic acid and ursolic acid in cornel by cyclodextrin-modified micellar electrokinetic chromatography

A cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) method was established for the determination of oleanolic acid and ursolic acid in cornel. The two components were separated in the running buffer of 40 mmol/L sodium borate containing 5% methanol, 25 mmol/L SDS and 15 mmol/L hydroxypropyl-beta-cyclodextrin (HP-beta-CD). The applied voltage was 24 kV. The wavelength of detection was 200 nm. The temperature was kept at 25 C. Cinnamic acid was used as the internal standard. The analytical performance of the method was tested with respect to linearity, precision and recovery. The calibration curves were linear in the range of 10.15-243.6 microg/mL, r=0.9993 (oleanolic acid) and 10.07-241.7 microg/mL, r=0.9994 (ursolic acid); the intra-day precision (RSD) was less than 3.7% (oleanolic acid) and 4.1% (ursolic acid); the inter-day precision (RSD) was less than 4.2% (oleanolic acid) and 4.9% (ursolic acid). The limits of detection were 1.6 microg/mL for both components. The method proved to be sensitive, rapid, accurate and suitable for the determination of oleanolic acid and ursolic acid in cornel.     

齐墩果酸与熊果酸结构修饰物的药理活性和构效关系研究进展

齐墩果酸(OA)和熊果酸(UA)均属于五环三萜类化合物,广泛存在于自然界中,具有多种显著的生物活性。本文综述了近年来齐墩果酸及熊果酸结构修饰物的药理活性和构效关系的研究进展。     

Novel biotransformation of pentacyclic triterpenoid acids by Nocardia sp. NRRL 5646

Six pentacyclic triterpene acids, ursolic acid, oleanolic acid, betulinic acid, 23-hydroxybetulinic acid, glycyrrhetinic acid, and senegenin, were metabolized by the microbe Nocardia sp. NRRL 5646 to selectively furnish their corresponding 28-methyl esters. Notably, ursolic acid (1) was converted to oleanolic acid methyl ester (4) via two intermediates, oleanolic acid (2), and ursolic acid methyl ester (3), which are formed by participation of ‘retro-biosynthetic’ methyl migration from C-19 to C-20. Senegenin (11) was selectively converted to a nortriterpene methyl ester, senegenic acid methyl ester (12), via an unprecedented C-C bond cleavage. The stereochemical assignments of compounds 11 and 12 were made unambiguously for the first time using 2D NMR spectroscopy.     

Production of triterpene acids by cell suspension cultures of Olea europaea

Olive (Olea europaea) contains large quantity of triterpene acids including oleanolic acid (6) as a major one. Varieties of biological activities exhibited by triterpene acids attracted our attentions, especially from pharmaceutical viewpoints. Cell culture of olive plant was induced and its triterpene constituents were studied. From the cell suspension cultures, six ursane type triterpene acids; ursolic acid (9), pomolic acid (10), rotundic acid (11), tormentic acid (12), 2alpha-hydroxyursolic acid (13) and 19alpha-hydroxyasiatic acid (14), and two oleanane type acids; oleanolic acid and maslinic acid (7), have been isolated. Quantity of ursane type triterpene acids produced by cell cultures was larger than that of oleanane type. Further, a multifunctional oxidosqualene cyclase (OSC) named OEA was cloned by homology based PCRs from the same cultured cells. Major product of OEA is alpha-amyrin (ursane skeleton), showing good accordance to higher content of ursane-type triterpene acids in the cultured cells, and strongly suggesting OEA to be a major contributor OSC for their production.     

Determination of Oleanolic, Betulinic and Ursolic Acid in Lamiaceae and Mass Spectral Fragmentation of Their Trimethylsilylated Derivatives

Phytochemical investigations of different species of Lamiaceae family (rosemary-Rosmarinus officinalis L., sage-Salvia officinalis L., winter savory-Satureja montana L., clary sage-Salvia sclarea L. and sticky sage-Salvia glutinosa L.), using gas chromatography and mass spectrometry (GC-MS) were performed. The studies were focused on oleanolic, betulinic and ursolic acid. Since oleanolic and ursolic acid are position isomers with very similar structures, the difficulties in their separation and identification have been reported by several authors. However, both compounds can be well distinguished by order of elution during gas chromatography and by intensities of the fragment ion signals in their mass spectra, where the retro-Diels-Alder reaction was primarily observed. A combination of ultrasonic extraction, solid phase extraction, size exclusion chromatography, trimethylsilylation, and GC-MS resulted in simultaneous separation, identification and quantification of the mentioned compounds. The compounds have been identified by retention time and comparison of mass spectra with those of pure standards. The mass spectral fragmentation behavior of all three derivatised acids was investigated. The obtained characteristic fragment patterns are discussed in the presented work. Good linearity over the concentration range 1–50 mg L⁻¹ for all three compounds was confirmed. The correlation coefficients (r ² ) were in the range of 0.9980–0.9983. Quantitative analyses of different Lamiaceae extracts showed that the oleanolic acid content ranged from 0.09 to 0.9% dry weight, content of betulinic acid ranged from traces to 0.6%, and that of ursolic acid varied from 0.09 to 1.6% dry weight.     

Rapid and novel discrimination and quantification of oleanolic and ursolic acids in complex plant extracts using two-dimensional nuclear magnetic resonance spectroscopy—Comparison with HPLC methods

A novel strategy for NMR analysis of mixtures of oleanolic and ursolic acids that occur in natural products is described. These important phytochemicals have similar structure and their discrimination and quantification is rather difficult. We report herein the combined use of proton-carbon heteronuclear single-quantum coherence (¹H-¹³C HSQC) and proton-carbon heteronuclear multiple-bond correlation (¹H-¹³C HMBC) NMR spectroscopy, in the identification and quantitation of oleanolic acid (OA) and ursolic acid (UA)in plant extracts of the Lamiaceae and Oleaceae family. The combination of ¹H-¹³C HSQC and ¹H-¹³C HMBC techniques allows the connection of the proton and carbon-13 spins across the molecular backbone resulting in the identification and, thus, discrimination of oleanolic and ursolic acid without resorting to physicochemical separation of the components. The quantitative results provided by 2D ¹H-¹³C HSQC NMR data were obtained within a short period of time (∼14 min) and are in excellent agreement with those obtained by HPLC, which support the efficiency of the suggested methodology.     

Rapid analysis of triterpenic acids by liquid chromatography using porous graphitic carbon and evaporative light scattering detection

An original system which uses Porous Graphitic Carbon as support and a mixture of organic solvents as mobile phase is proposed for the analysis of triterpenic acids by liquid chromatography. The separation of betulinic acid, ursolic acid, oleanolic acid, and 18alpha- and 18beta-glycyrrhetinic acids was carried out within a short time and monitored by evaporative light scattering detection as universal detection method. Molecular modelling studies show that the main contribution to the selectivity comes from the electrostatic interaction characterised by the dipole moment of the products.     

Simultaneous determination of oleanolic acid,ursolic acid,quercetin and apigenin in Swertia mussotii Franch by capillary zone electrophoresis with running buffer modifier

The method of capillary zone electrophoresis (CZE) with direct UV detection was developed for the determination of oleanolic acid, ursolic acid, quercetin and apigenin. and then for the first time successfully applied to the analysis of four analytes in Swertia mussotii Franch and its preparations. Various factors affecting the CZE procedure were investigated and optimized, and the optimal conditions were: 50 × 10^?3 mol/L borate‐phosphate buffer (pH 9.5) with 5.0 × 10^?3 mol/L β‐cyclodextrin, 15 kV separation voltage, 20 °C column temperature, 250 nm detection wavelength and 5 s electrokinetic injection time (voltage 20 psi). Under the conditions, oleanolic acid, ursolic acid, quercetin and apigenin could be determined within the test ranges with a good correlation coefficient (r² > 0.9991). The limits of detection for conditions, oleanolic acid, ursolic acid, quercetin and apigenin were 0.3415, 0.2003, 0.0062 and 0.2538 µg/mL, respectively, and the intra‐ and inter‐day relative standard deviations were no more than 4.72%. This procedure provided a convenient, sensitive and accurate method for simultaneous determination of oleanolic acid, ursolic acid, quercetin and apigenin in S. mussotii Franch. Copyright © 2014 John Wiley & Sons, Ltd.     

熊果酸与齐墩果酸高效液相色谱分离条件优化

描述了熊果酸、齐墩果酸色谱保留行为与流动相pH和有机溶剂体积分数之间的函数关系,并求解了各自模型中的9个参数,得到在流动相中熊果酸的电离常数pKa=4.8145,齐墩果酸电离常数pKa=4.9317。文章还对模型外推过程中存在的误差进行了校正,并运用校正后的模型预测熊果酸的容量因子,实验值接近预测值,误差小于5%。最后采用熊果酸和齐墩果酸的容量因子与流动相pH和有机溶剂组成之间的函数关系,进行了两者在同一流动相中分离条件的优化,获得了最佳的分离条件:流动相中甲醇体积分数为85.2%,pH为6.50。     

Preparation and characterization of monodisperse molecularly imprinted polymers for the recognition and enrichment of oleanolic acid

Monodisperse molecularly imprinted polymers for oleanolic acid were successfully prepared by a precipitation polymerization method using oleanolic acid as a template, methacrylic acid as a functional monomer, and divinylbenzene/ethylene glycol dimethacrylate as a crosslinker in a mixture of acetonitrile and ethanol (3:1, v/v). The imprinted polymers and nonimprinted polymers were characterized by using scanning electron microscopy, Fourier transform infrared spectroscopy, and thermogravimetric analysis. The resulting imprinted polymers had average diameters of 3.15 μm and monodispersity values of 1.024. The results clearly demonstrate that use of ethanol as a cosolvent is indeed exceedingly effective in promoting the dissolution of oleanolic acid and in obtaining uniform microspheres. Molecular recognition properties and binding capability to oleanolic acid were evaluated by adsorption testing, which indicated that the imprinted polymers displayed optimal binding performance with a maximum adsorption capacity of 17.3 mg/g and a binding saturation time of 80 min. Meanwhile, the produced imprinted polymers exhibited higher selectivity to oleanolic acid than that for ursolic acid and rhein. Herein, the studies can provide theoretical and experimental references for the oleanolic acid molecular imprinted system.     

Rhodamine 101 Conjugates of Triterpenoic Amides Are of Comparable Cytotoxicity as Their Rhodamine B Analogs

Pentacyclic triterpenoic acids (betulinic, oleanolic, ursolic, and platanic acid) were selected and subjected to acetylation followed by the formation of amides derived from either piperazine or homopiperazine. These amides were coupled with either rhodamine B or rhodamine 101. All of these compounds were screened for their cytotoxic activity in SRB assays. As a result, the cytotoxicity of the parent acids was low but increased slightly upon their acetylation while a significant increase in cytotoxicity was observed for piperazinyl and homopiperazinyl amides. A tremendous improvement in cytotoxicity was observed; however, for the rhodamine B and rhodamine 101 conjugates, and compound 27, an ursolic acid derived homopiperazinyl amide holding a rhodamine 101 residue showed an EC₅₀ = 0.05 µM for A2780 ovarian cancer cells while being less cytotoxic for non-malignant fibroblasts. To date, the rhodamine 101 derivatives presented here are the first examples of triterpene derivatives holding a rhodamine residue different from rhodamine B.     

Anti-Diabetic Potential of Plant-Based Pentacyclic Triterpene Derivatives: Progress Made to Improve Efficacy and Bioavailability

Diabetes mellitus (DM) results from the inability of the pancreas to produce sufficient insulin or weakened cellular response to the insulin produced, which leads to hyperglycemia. Current treatments of DM focus on the use of oral hypoglycemic drugs such as acarbose, alpha-glucose inhibitors, sulphonylureas, thiazolidinediones, and biguanides to control blood glucose levels. However, these medications are known to have various side effects in addition to their bioavailability, efficacy, and safety concerns. These drawbacks have increased interest in the anti-diabetic potential of plant-derived bioactive compounds such as oleanolic and maslinic acids. Although their efficacy in ameliorating blood glucose levels has been reported in several studies, their bioavailability and efficacy remain of concern. The current review examines the anti-diabetic effects of oleanolic, maslinic, asiatic, ursolic, and corosolic acids and their derivatives, as well as the progress made thus far to enhance their bioavailability and efficacy. The literature for the current review was gathered from leading academic databases—including Google Scholar and PubMed—the key words listed below were used. The literature was searched as widely and comprehensively as possible without a defined range of dates.     

Solid-phase extraction of ursolic acid from herb using beta-cyclodextrin-based molecularly imprinted microspheres

A new method was employed to solid-phase extract ursolic acid from Ilex kudingcha C. J. Tseng using molecularly imprinted microspheres (MIMs) as the sorbent. Using a surface molecular imprinting technique, MIMs for ursolic acid were prepared with bonded beta-CD and acrylamide in combination based on functionalized poly(glycidyl methacrylate) microspheres (F-PGMA). Compared with non-MIMs (NIMs), MIMs showed high adsorption capacity, significant selectivity, and good site accessibility for ursolic acid. The maximum static adsorption capacities of the MIMs and NIMs for ursolic acid were 42.5 and 4.9 micromol/g, respectively. Chromatographic analysis shows that ursolic acid and oleanolic acid could be separated well when MIMs were used as the stationary phase of HPLC. The conditions of molecularly imprinted SPE (MISPE) of ursolic acid from the herb extract were optimized using different concentrations of ethanol solutions as loading, washing, and eluting solutions. The successful extraction of ursolic acid by MIMs provided a possible innovative approach to separate ursolic acid from herb.     

Synthesis and characterization of folic acid‐modified carboxymethyl chitosan‐ursolic acid targeted nano‐drug carrier for the delivery of ursolic acid and 10‐hydroxycamptothecin

Carboxymethyl chitosan (CMCS), as a water‐soluble, biocompatible, and biodegradable polymer, is an excellent carrier for a sustained drug delivery system. In this study, a amphiphilic carboxymethyl chitosan‐ursolic acid nano‐drug carrier modified by folic acid (FPCU) were prepared, and then the nano‐drug carrier wrapped another anticancer drug 10‐hydroxycamptothecin were self‐assembled into nanoparticles (FPCU/HCPT NPs). The FPCU/HCPT NPs had a suitable size, high drug loading efficiency of ursolic acid (6.4%) and 10‐hydroxycamptothecin (14.1%). The drug release study in vitro indicated that the nanoparticles have obviously sustained effect and pH sensitive behaviors, the drug release amount was higher at pH 5.5 than at pH 7.4. in vitro and in vivo study showed that the nanoparticles displayed a high antitumor efficiency to tumor cells compared with free drug. The nano delivery system as a carrier for ursolic acid (UA) and 10‐hydroxycamptothecin (HCPT) has good application prospects in cancer treatment.     

Quantification of eugenol, luteolin, ursolic acid, and oleanolic acid in black (Krishna Tulasi) and green (Sri Tulasi) varieties of Ocimum sanctum Linn. using high-performance thin-layer chromatography

Ocimum sanctum (family Lamiaceae) is a reputed drug of Ayurveda, commonly known as Tulasi. In the present work, we quantified 4 marker compounds, viz., eugenol, luteolin, ursolic acid, and oleanolic acid, from the leaf of green and black varieties of O. sanctum using high-performance thin-layer chromatography (HPTLC) with densitometry. The methods were found to be precise, with relative standard deviation (RSD) values for intraday analyses in the range of 0.52 to 0.91%, 0.77 to 1.29%, 0.11 to 0.16%, and 0.34 to 0.42% and for interday analyses in the range of 0.73 to 0.96%, 1.02 to 2.08%, 0.11 to 0.12%, and 0.39 to 0.64% for different concentrations of eugenol, luteolin, ursolic acid, and oleanolic acid, respectively. Instrumental RSD values were 0.24, 0.39, 0.21, and 0.18% for eugenol, luteolin, ursolic acid, and oleanolic acid, respectively. Accuracy of the methods was checked by conducting a recovery study at 3 different levels for the 4 compounds, and the average recoveries were found to be 99.73, 99.3, 100.58, and 100.57%, respectively. Eugenol content ranged from 0.175 to 0.362% (w/w) and luteolin from 0.019 to 0.046% (w/w) in the samples analyzed. Green variety was found to contain higher amounts of ursolic acid [0.478 and 0.348% (w/w), from Sources 1 and 2, respectively] than the black variety [0.252 and 0.264% (w/w) from Sources 1 and 2, respectively]. Black variety had 0.174 and 0.218% (w/w) of oleanolic acid from Sources 1 and 2, respectively, while it was not detected in the green variety. Ursolic acid and oleanolic acid ran at the same Rf value and could not be resolved in several solvent systems tried. However, we observed that only ursolic acid gave yellow fluorescence under 366 nm ultraviolet light after derivatization with anisaldehyde-sulfuric acid reagent. The HPTLC-densitometry methods for the quantification of the 4 markers in O. sanctum leaf will have the applicability in quality control.     

Rapid access to ketones related to oleanolic and ursolic acids

We describe in this article a process with which to obtain different ketones related to oleanolic and ursolic acids from a natural source, Salvia canariensis L., with minimal use of chromatography columns. Amongst the isolated compounds was 12α-bromo-3-oxo-olean-13, 28-olide (3), which was fully characterised, including a characterisation of its molecular configuration by X-ray crystallographic analysis by 1 and 2-dimensional NMR techniques.