Determination of anthraquinones in Rhamnus purshiana using high-performance liquid chromatography coupled to diode array detector and simple ultraviolet spectroscopic analysis.

A new method based on Ultraviolet spectrophotometry was developed and compared with that based on high-performance liquid chromatography for the determination and quantification of anthraquinones in the extracts of Rhamnus purshiana bark. A validated quantitative analysis of cascaroside A, cascaroside B, emodin, and aloe-emodin in these herbal products has been previously performed using high-performance liquid chromatography coupled with a diode array detector. In the high-performance liquid chromatography analysis, all the anthraquinones showed satisfactory regression (r2 > 0.98) within the test ranges, and the recovery was in the range of 94-117%. The limits of detection and quantification were 0.008-0.010 and 0.029-0.035 µg/mL, respectively. Hierarchical cluster analysis and principal component analysis showed differences in the anthraquinones determined from herbal samples. Subsequently, a simple and low-cost ultraviolet spectrophotometric methodology for the quantitative analysis of the same compounds in the extracts was applied, and all the contents were determined. A paired t-test confirmed that there were no significant differences between the two methods. Our results revealed that the developed method is simple and provides the ability to discriminate and control the quality of anthraquinones in herbal products.

Assessment of Rhamnus alaternus Leaves Extract: Phytochemical Characterization and Antimelanoma Activity.

Rhamnus alaternus (Rhamnaceae) has been used as a laxative, purgative, diuretic, antihypertensive, and depurative. However, few scientific research studies on its antimelanoma activity have been reported. This study aimed to investigate the in vitro antimelanoma effect of an enriched total oligomer flavonoid (TOF) extract, from R. alaternus, and to identify its phytochemical compounds. The chemical composition of TOF extract was assessed by HPLC-electrospray ionization tandem mass spectrometry (HPLC/ESI-MS2) analysis. Antimelanoma activity was determined on cultured tumor cell B16F10 by the crystal violet assay, the alkaline comet assay, acridine orange/ethidium bromide (AO/EB), annexin V-fluorescein isothiocyanate/ propidium iodide (V-FITC/PI) staining, the cell cycle distribution, and the wound healing assay. Regarding chemical composition, a mixture of quercetin diglucoside, quercetin-3-O-neohesperidoside, kaempferol-3-O-(2G-α-L-rhamnosyl)-rutinoside, rhamnetin hexoside, kaempferol-3-O-rutinoside, rhamnocitrin hexoside, pilosin hexoside, apigenin glucoside, and kaempferol-3-O-glucoside was identified as major phytochemical compounds of the extracts. TOF extract inhibits melanoma B16F10 cell proliferation in dose-dependent manner. The induction of apoptosis was confirmed by comet assay, AO/EB, and annexin V-FITC/PI test. TOF extract could also induce S phase cell cycle, inhibit, and delay the cell migration of B16F10 cells. The findings showed that TOF extract from R. alaternus could be a potentially good candidate for future use in alternative antimelanoma treatments.

Isolation, Characterization, Complete Structural Assignment, and Anticancer Activities of the Methoxylated Flavonoids from Rhamnus disperma Roots.

Different chromatographic methods including reversed-phase HPLC led to the isolation and purification of three O-methylated flavonoids; 5,4'-dihydroxy-3,6,7-tri-O-methyl flavone (penduletin) (1), 5,3'-dihydroxy-3,6,7,4',5'-penta-O-methyl flavone (2), and 5-hydroxy-3,6,7,3',4',5'-hexa-O-methyl flavone (3) from Rhamnus disperma roots. Additionlly, four flavonoid glycosides; kampferol 7-O-α-L-rhamnopyranoside (4), isorhamnetin-3-O-ß-D-glucopyranoside (5), quercetin 7-O-α-L-rhamnopyranoside (6), and kampferol 3, 7-di-O-α-L-rhamnopyranoside (7) along with benzyl-O-ß-D-glucopyranoside (8) were successfully isolated. Complete structure characterization of these compounds was assigned based on NMR spectroscopic data, MS analyses, and comparison with the literature. The O-methyl protons and carbons of the three O-methylated flavonoids (1-3) were unambiguously assigned based on 2D NMR data. The occurrence of compounds 1, 4, 5, and 8 in Rhamnus disperma is was reported here for the first time. Compound 3 was acetylated at 5-OH position to give 5-O-acetyl-3,6,7,3',4',5'-hexa-O-methyl flavone (9). Compound 1 exhibited the highest cytotoxic activity against MCF 7, A2780, and HT29 cancer cell lines with IC50 values at 2.17 µM, 0.53 µM, and 2.16 µM, respectively, and was 2-9 folds more selective against tested cancer cell lines compared to the normal human fetal lung fibroblasts (MRC5). It also doubled MCF 7 apoptotic populations and caused G1 cell cycle arrest. The acetylated compound 9 exhibited cytotoxic activity against MCF 7 and HT29 cancer cell lines with IC50 values at 2.19 µM and 3.18 µM, respectively, and was 6-8 folds more cytotoxic to tested cancer cell lines compared to the MRC5 cells.

Comparison between the Antioxidant and Antidiabetic Activity of Fenugreek and Buckthorn in Streptozotocin-Induced Diabetic Male Rats.

This study is aimed at comparing the antidiabetic and antioxidant potential of fenugreek and buckthorn which are commonly used in modulating diabetes in the Middle East. In this study, the antioxidant and antidiabetic activity of the aqueous extracts of the leaf and seed of fenugreek and buckthorn was tested in streptozotocin-induced diabetic male rats fed with a fat-rich diet for 8 weeks. Thirty-six male albino rats were divided into 6 groups (n = 6); the 1st group was the negative control. Diabetes was induced in the other 30 rats using streptozotocin, which were then divided into 5 groups; the 2nd was the untreated positive diabetic group, the 3rd was treated with fenugreek leaf aqueous extract, the 4th was treated with the fenugreek seed aqueous extract, the 5th was treated with buckthorn leaf aqueous extract, and the 6th was treated with buckthorn seed aqueous extract. The positive control group showed an increase in blood sugar, glycated hemoglobin, liver function enzymes, lactate dehydrogenase, kidney indices, total cholesterol, triglycerides, low- and very-low-density lipoprotein, immunoglobulins, and lipid peroxidation and a decrease in high-density lipoprotein, albumin, and antioxidant activity. The histology of the liver and testes showed severe histopathological alterations. Rats of groups 4-6 that were treated with the aqueous extract of the leaf and seed extract of fenugreek and buckthorn showed improvement of all biochemical and histopathological parameters. The seed extract of fenugreek and buckthorn showed more antioxidant activity than their leaves.

Bioguided Isolation of Active Compounds from Rhamnus alaternus against Methicillin-Resistant Staphylococcus aureus (MRSA) and Panton-Valentine Leucocidin Positive Strains (MSSA-PVL).

Despite intensified efforts to develop an effective antibiotic, S. aureus is still a major cause of mortality and morbidity worldwide. The multidrug resistance of bacteria has considerably increased the difficulties of scientific research and the concomitant emergence of resistance is to be expected. In this study we have investigated the in vitro activity of 15 ethanol extracts prepared from Moroccan medicinal plants traditionally used for treatment of skin infections. Among the tested species I. viscosa, C. oxyacantha, R. tinctorum, A. herba alba, and B. hispanica showed moderate anti-staphylococcal activity. However, R. alaternus showed promising growth-inhibitory effects against specific pathogenic bacteria especially methicillin-susceptible Staphylococcus aureus Panton-Valentine leucocidin positive (MSSA-PVL) and methicillin-resistant S. aureus (MRSA). The bioguided fractionation of this plant using successive chromatographic separations followed by nuclear magnetic resonance (NMR) and mass spectrometry (MS) including EIMS and HREIMS analysis yielded the emodin (1) and kaempferol (2). Emodin being the most active with MICs ranging between 15.62 and 1.95 µg/mL and showing higher activity against the tested strains in comparison with the crude extract, its mechanism of action and the structure-activity relationship were interestingly discussed. The active compound has not displayed toxicity toward murine macrophage cells. The results obtained in the current study support the traditional uses of R. alaternus and suggest that this species could be a good source for the development of new anti-staphylococcal agents.

Phytofabrication of cobalt oxide nanoparticles from Rhamnus virgata leaves extract and investigation of different bioactivities.

The tunable cobalt oxide nanoparticles (CoONPs) are produced due to the phytochemicals present in Rhamnus virgata (RhV) leaf extract which functions as reducing and stabilization agents. The synthesis of CoONPs was confirmed using different analytical techniques: UV-Vis spectroscopy, X-ray diffraction (XRD), scanning electron microscopy (SEM), dynamics light scatterings (DLS), Fourier-transform infrared spectroscopy (FTIR), energy dispersive X-ray, and Raman spectroscopy analyses. Furthermore, multiple biological activities were performed. Significant antifungal and antibacterial potentials have been reported. The in vitro cytotoxic assays of CoONPs revealed strong anticancer activity against human hepatoma HUH-7 (IC50 : 33.25 µg/ml) and hepatocellular carcinoma HepG2 (IC50 : 11.62 µg/ml) cancer cells. Dose-dependent cytotoxicity potency was confirmed against Leishmania tropica (KMH23 ); amastigotes (IC50 : 58.63 µg/ml) and promastigotes (IC50 : 32.64 µg/ml). The biocompatibility assay using red blood cells (RBCs; IC50 : 4,636 µg/ml) has confirmed the bio-safe nature of CoONPs. On the whole, results revealed nontoxic nature of RhV-CoONPs with promising biological potentials.

Isolation of six anthraquinone diglucosides from cascara sagrada bark by high-performance countercurrent chromatography.

In this study, high-performance countercurrent chromatography was employed to isolate six anthraquinone diglucosides, namely, cascarosides A-F, from cascara sagrada (Rhamnus purshiana DC [Rhamnaceae]) bark. The n-butanol-soluble extract of cascara sagrada was separated by off-line two-dimensional high-performance countercurrent chromatography. The first-dimensional high-performance countercurrent chromatography resolved the n-butanol-soluble extract (510 mg) of cascara sagrada using the flow-rate gradient method with a chloroform-methanol-isopropanol-water (6:6:1:4, v/v/v/v, normal-phase mode) system to afford four anthraquinone diglucoside fractions (groups I [cascarosides C-D, 71 mg], II [cascarosides E-F, 56 mg], III [cascaroside A, 53 mg], and IV [cascaroside B, 31 mg]). Groups I and II were separated by the second-dimensional high-performance countercurrent chromatography using an ethyl acetate-n-butanol-water (7:3:10, v/v/v, normal-phase mode) system to yield cascarosides C (34 mg), D (26 mg), E (19 mg), and F (15 mg). Additionally, one-step preparative-scale high-performance countercurrent chromatography method was developed to isolate large amounts of cascarosides A (389 mg) and B (187 mg) from the water-soluble extract (2.1 g) of cascara sagrada using an ethyl acetate-n-butanol-water (2:8:10, v/v/v, normal-phase mode) system. The current study demonstrated that high-performance countercurrent chromatography is a powerful technique for the isolation of marker compounds from herbal materials.

Rhamnus prinoides (gesho) stem extract prevents co-culture biofilm formation by Streptococcus mutans and Candida albicans.

Streptococcus mutans and Candida albicans exhibit a symbiotic relationship to form polymicrobial biofilms that exacerbate oral infections including early-childhood caries, periodontitis and candidiasis. Rhamnus prinoides (gesho) has traditionally been used for the treatment of a variety of illnesses and was recently found to inhibit Gram-positive bacterial biofilm formation. We hypothesized that Rhamnus prinoides extracts have anti-biofilm activity against S. mutans and C. albicans mono- and dual-species biofilms. Ethanol extracts were prepared from gesho stems and leaves; then anti-biofilm activity was assessed using crystal violet, resazurin and XTT staining. Ethanol extracts significantly inhibited Streptococcus mutans and Candida albicans mono-species biofilm formation up to 97 and 75%, respectively. The stem ethanol extract disrupted S. mutans and C. albicans co-culture synergism, with 98% less polymicrobial biofilm formation than the untreated control. Additionally, this extract inhibited planktonic S. mutans cell growth and decreased biofilm polysaccharide production up to 99%. The reduction in polysaccharide production is likely a contributing factor in the anti-biofilm activity of GSE. These findings indicate that gesho or gesho-derived compounds may have potential as additives to oral hygiene products. SIGNIFICANCE AND IMPACT OF THE STUDY: Oral Streptococcus mutans and Candida albicans biofilms are associated with a variety of illnesses. When occurring together, the resulting infections are especially challenging to treat due to enhanced biofilm formation and antibiotic resistance. More therapeutics that can effectively prevent polymicrobial biofilm formation and disrupt interspecies synergism are needed. Rhamnus prinoides ethanol extracts significantly inhibited dual-species biofilm formation and disrupted interspecies synergism.

Physicochemical Aspects and Sensory Profiles as Various Potential Factors for Comprehensive Quality Assessment of Nü-Er-Cha Produced from Rhamnus heterophylla Oliv.

Nü-Er-Cha, produced from the leaves of Rhamnus heterophylla Oliv., is known as an herbal tea and used in the treatment of bleeding, irregular menstruation and dysentery. A method was developed for the quality assessment of herbal tea, Nü-Er-Cha, adopting physical parameters, chemical constituents and sensory profiles as various potential factors. Their inner relationship was mined by multivariate statistical analysis tools, and the three factors were integrated by a technique for order preference by a similarity to ideal solution (TOPSIS) approach to comprehensively analyze the characters of Nü-Er-Cha. Viscosity was also introduced to the physical parameter determination besides conductivity, pH and color. Seven common peaks of eight batches of Nü-Er-Cha were marked by a high performance liquid chromatography (HPLC) fingerprint. They were further identified by HPLC mass spectrometry/mass spectrometry (HPLC-MS/MS) as hydroxybenzoic acids and flavanol glycosides. Fifty trained members participated in the sensory evaluation. Significant correlations between total sensory scores and conductivity, viscosity as well as pH were observed, a relatively innovative result for the quality assessment of herbal teas. The common peaks, belonging to hydroxybenzoic acids and flavanol glycosides, were mainly related to the color of infusions and leaves. The result of the TOPSIS analysis showed that S3 and S4 ranked as the top two in the comprehensive quality assessment. This may be related to rhamnetin triglycoside with a galactose/glucose and two rhamnoses, which had a higher peak response in S3 and S4 than that in the other samples. The present study may contribute to a better understanding of the relationship regarding physical properties, chemical composition and sensory profiles, and it may supply ideas for the comprehensive quality assessment of the herbal tea Nü-Er-Cha.

Two-dimensional qNMR of anthraquinones in Frangula alnus (Rhamnus frangula) using surrogate standards and delay time adaption.

In the absence of adequate reference material, a problem often encountered in natural product chemistry, we investigated the use of surrogate standards in two-dimensional qNMR for the quantification of anthraquinones in the bark of alder buckthorn (Frangula alnus). Using the integrals of cross signals in the HSQC spectrum obtained from commercial standards rutin and duroquinone and adapting the delays for the 1JCH coupling, we quantified the total amount of anthraquinones and anthraquinone glucosides, as well as the content of the value-determining glucofrangulins and frangulins. Thereby, duroquinone was used as an external standard to establish the calibration curve for the methylated anthraquinone scaffold, whereas calibration curves for the glycosides were obtained using the anomeric proton signals of the rutinose disaccharide. The method was validated for accuracy, precision, specificity, linearity and limit of quantitation and shows clear advantages over the method of the European Pharmacopeia, especially in terms of specificity and meaningfulness of the results. Apart from being a useful alternative in the quality control of alder buckthorn, the presented approach demonstrates, moreover, the versatility of sophisticated 2D measurements in quantitative NMR.

Phytochemical profiles, antioxidant and anti-acetylcholinesterasic activities of the leaf extracts of Rhamnus lycioides subsp. oleoides (L.) Jahand. & Maire in different solvents.

In this work, the extracts obtained with different solvents from the leaves of Rhamnus lycioides subsp. oleoides (L.) Jahand. & Maire were studied for their phytochemical profile and then for their antioxidant and acetylcholinesterase inhibitory activities. The phytochemical profiles of the extracts in n-hexane, dichloromethane, ethyl acetate, methanol, anthraquinone rich and water, showed the presence of different compounds belonging to several classes of natural products such as flavonoids, anthraquinones, saccharides and fatty acids. For what concerns the biological tests, the ethyl acetate, methanol and anthraquinone rich extracts showed the highest activities in both assays due to the high amount of compounds possessing those properties such as flavonoids and anthraquinones. By consequence, these specific extracts of the species may be considered to be potential sources of natural antioxidant and anti-acetylcholinesterasic compounds.

Isolation and identification of new anthraquinones from Rhamnus alaternus L and evaluation of their free radical scavenging activity.

From the butanolic and the ethyl acetate extracts of Rhamnus alaternus L root bark and leaves, three new anthraquinone glycosides, alaternosides A-C (1,4,6,8 tetrahydroxy-3 methyl anthraquinone 1-O-ß-D-glucopyranosyl-4,6-di-O-α-L-rhamnopyranoside (1); 1,2,6,8 tetrahydroxy-3 methyl anthraquinone 8-O-ß-D-glucopyranoside (2) and 1, 6 dihydroxy-3 methyl 6 [2'-Me (heptoxy)] anthraquinone (3)) were isolated and elucidated together with the two known anthraquinone glycosides, Physcion-8-O-rutinoside (4) and emodin-6-O-α-L-rhamnoside (5) as well as with the known kaempferol-7-methylether (6), ß-sitosterol (7) and ß-sitosterol-3-O-glycoside (8). Their chemical structures were elucidated using spectroscopic methods (1D-, 2D-NMR and FAB-MS). Free radical scavenging activity of the isolated compounds was evaluated by their ability to scavenge DPPH. free radicals. Compounds (3), (4) and (6) showed the highest activity with IC50 values of 9.46, 27.68 and 2.35 µg/mL, respectively.

Inhibitory effects of various solvent extracts from Rhamnus frangula leaves on some inflammatory and metabolic enzymes.

Many enzymes are involved in numerous pathologies which are related to metabolic reactions and inflammatory diseases such as pancreatic lipase, α-amylase, α-glucosidase and xanthine oxidase and secreted phospholipases A2 (Group IIA, V and X), respectively. Therefore, inhibiting these enzymes offer the potential to block production of more inflammatory substances and decrease the risk factors for cardiovascular diseases. The purpose of this study was to investigate some potent, bioavailable and selective inhibitors of some catalytic proteins implicated to metabolic syndrome and their antioxidant effects from various solvent extracts of R. frangula leaves. The anti-inflammatory, obesity, diabete and XO potentials were evaluated through analyses of inhibition activities of corresponding metabolites.The water extract exhibited an important inhibitory effect on human, dromedary and stingray sPLA2-G IIA achieved an IC50 of 0.16±0.06, 0.19±0.05 and 0.07±0.01 mg/mL, respectively. Likewise, the same fraction demonstrated the highest pancreatic lipase inhibitory activity using two different substrates. Indeed, 50% of dromedary pancreatic lipase inhibition was demonstrated for 5 min and 15 min using olive oil and TC4 substrates, respectively. Besides, it was established that methanolic extract had more effective inhibitory lipase activity than ORLISTAT used as a specific inhibitor of gastric, pancreatic and carboxyl ester lipase for treating obesity, with an IC50 of 5.51±0.27 and 91.46±2.3 µg/mL, respectively. In the case of α-amylase, α-glucosidase and xanthine oxidase, the crude methanolic extract showed a potential inhibitory effect with an IC50 of 45±3.45, 3±0.15 and 27±1.71 µg/mL, respectively. Conclusively, R. frangula leaves extracts showed a potential value of some sPLA2, some metabolic enzymes and XO inhibitors as anti-inflammatory and metabolic syndrome drugs.

Emodin, Physcion, and Crude Extract of Rhamnus sphaerosperma var. pubescens Induce Mixed Cell Death, Increase in Oxidative Stress, DNA Damage, and Inhibition of AKT in Cervical and Oral Squamous Carcinoma Cell Lines.

There have been few studies on the pharmacological properties of Rhamnus sphaerosperma var. pubescens, a native Brazilian species popularly known as "fruto-de-pombo." The aim of this study was to investigate the scavenging capacity of emodin, physcion, and the ethanolic crude extract of Rhamnus sphaerosperma var. pubescens against reactive oxygen and nitrogen species, as well as their role and plausible mechanisms in prompting cell death and changes in AKT phosphorylation after cervical (SiHa and C33A) and oral (HSC-3) squamous cell carcinoma treatments. Emodin was shown to be the best scavenger of NO• and O2•-, while all samples were equally effective in HOCl/OCl- capture. Emodin, physcion, and the ethanolic extract all exhibited cytotoxic effects on SiHa, C33A, HSC-3, and HaCaT (immortalized human keratinocytes, nontumorigenic cell line), involving mixed cell death (apoptosis and necrosis) independent of the caspase activation pathway. Emodin, physcion, and the ethanolic extract increased intracellular oxidative stress and DNA damage. Emodin decreased the activation of AKT in all tumor cells, physcion in HSC-3 and HaCaT cells, and the ethanolic extract in C33A and HaCaT cells, respectively. The induction of cancer cell death by emodin, physcion, and the ethanolic crude extract of Rhamnus sphaerosperma var. pubescens was related to an increase in intracellular oxidative stress and DNA damage and a decrease in AKT activation. These molecules are therefore emerging as interesting candidates for further study as novel options to treat cervical and oral carcinomas.

Antihyperlipidemic effect of a Rhamnus alaternus leaf extract in Triton-induced hyperlipidemic rats and human HepG2 cells.

The Mediterranean buckthorn, Rhamnus alaternus L., is a plant used in traditional medicine in Mediterranean countries. We aimed at characterizing its phenolic compounds and explore potential antihyperlipidemic activity of this plant. The profile of phenolic compounds in R. alaternus leaf crude methanolic extract (CME) and its liquid-liquid extraction-derived fractions were analyzed by high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS2). Effects of CME on: circulating lipids in rats with Triton WR-1339-induced hyperlipidemia, intracellular lipid accumulation and expression of genes of fatty acid metabolism in human hepatoma HepG2 cells, and adipogenesis in the 3T3-L1 murine adipocyte cell model were assessed. The HPLC/ESI-MS2 analytical profile revealed a total of fifteen compounds, of which eleven were identified. Oral CME administration decreased blood levels of cholesterol and triacylglycerols in hyperlipidemic rats (by 60% and 70%, respectively, at 200 mg CME/kg). In HepG2 cells, CME exposure dose-dependently decreased intracellular lipids and up-regulated gene expression of carnitine palmitoyltransferase 1 involved in fatty acid oxidation. In the 3T3-L1 model, CME favored preadipocyte proliferation and adipogenesis, pointing to positive effects on adipose tissue expandability. These results suggest novel uses of R. alaternus by showing that its leaves are rich in flavonoids and flavonoid derivatives with an antihyperlipidemic effect in vivo and in hepatic cells.

Screening for anti-proliferative and anti-inflammatory components from Rhamnus davurica Pall. using bio-affinity ultrafiltration with multiple drug targets.

Rhamnus davurica Pall. (R. davurica) has been used as a traditional medicine for many years in China and abroad and shown a wide spectrum of biological activities. Previously, we reported the phytochemical fingerprinting profile of R. davurica, its distinct anti-proliferative activities against HT-29 and SGC-7901 cell lines, and the topoisomerase I (Top I) ligands based on bio-affinity ultrafiltration and HPLC-MS (UF-HPLC-MS). Nevertheless, among the 32 peaks detected in the fingerprinting profile, the common bioactive constituents responsible for the anti-inflammatory and anti-proliferative activities in the extracts remain elusive. To further explore the specific responsible components for their diversified activities and their potential action targets/mechanisms, the method based on bio-affinity UF-HPLC-MS using therapeutic targets like Top I and cyclooxygenase 2 (COX-2) was established to rapidly screen and identify the ligands binding to these known target enzymes. As a result, 12 components were revealed as potential Top I ligands along with 11 components as potential COX-2 ligands, where several components were revealed to possess both activities. Further validations of these bioactive components have also been conducted and confirmed their highlighted activities. This integrated method of UF-HPLC-MS exhibits high efficiency in rapidly screening for multi-target bioactive components responsible for multiple pharmacological effects from the complex natural products and could be very useful to explain the complex action mechanisms of herb medicines in a complex multi-component and multi-target mode at the molecular level. Graphical abstract Schematic diagram of UF-HPLC-MS assay to screen for Top I and COX-2 ligands. The principle of the assay usually involves the following steps: incubation, ultrafiltration, and identification.

An optimized protocol for anthraquinones isolation from Rhamnus frangula L.

Different from works described in the literature, which use expansive analytical methods to separation of anthraquinones derivatives (AQs), this communication reported a simple and inexpensive methodology to get them. In this way, the expensive commercial AQs: Chrysophanol, physcione and emodine were extracted from plant material (Rhamnus frangula L.) and isolated by classical column chromatography technique under optimised binary mobile phase gradients (CHCl3 : AcOEt(a), a = 1 to 5%) in excellent yields.

In-Silico UHPLC Method Optimization for Aglycones in the Herbal Laxatives Aloe barbadensis Mill., Cassia angustifolia Vahl Pods, Rhamnus frangula L. Bark, Rhamnus purshianus DC. Bark, and Rheum palmatum L. Roots.

For the European Pharmacopoeia (Ph. Eur.) herbal monograph draft of Cassia angustifolia Vahl. and Cassia senna L. leaves and pods, a safety limitation of aloe-emodin and rhein was proposed, due to toxicological concerns. A quantitative, analytical method of the anthraquinone aglycones in all Ph. Eur. monographed herbal laxatives is of interest. A rational method development for the aglycones aloe-emodin, rhein, emodin, chrysophanol, and physcion in five herbal drugs was realized by using 3D chromatographic modelling (temperature, solvent, and gradient time) and design of experiment (DOE) software (DryLab® 4). A methodical approach suitable for the challenging peak tracking in the chromatograms of the herbal drugs in dependence on the changes in the chromatographic conditions is described by using a combination of mass spectroscopy (MS) data (UHPLC-QDa), UV/Vis-spectra, and peak areas. The model results indicate a low robust range and showed that with the selected chromatographic system, small interferences could not be averted. The separation achieved shows a pure UV/Vis spectrum for all aglycones except for chrysophanol in Aloe barbadensis and emodin in Cassia angustifolia fruit. A gradient with the best resolution of the aglycones in all five drugs is proposed, and its suitability demonstrated for the quantification of aglycones in these herbal drugs.

Phytochemicals from the Bark of Rhamnus caroliniana.

Rhamnus caroliniana (Rhamnaceae), or Carolina buckthorn, is a plant commonly found in the southeastern United States that was used in Cherokee traditional medicine. To date, there have been no reports on the phytochemical constituents of R. caroliniana. The bark of R. caroliniana was collected and extracted with chloroform. The crude chloroform extract was subjected to preparative column chromatography on silica gel leading to the isolation of two anthraquinones (chrysophanol and physcion), the bianthrone ararobinol, the dihydroxynaphthalene torachrysone, and the fatty alcohol 1-docosanol. The bark essential oil of R. caroliniana was obtained by hydrodistillation and analyzed by gas chromatography - mass spectrometry. The major bark volatiles were the anthrone chrysarobin-(24.2%), the piperidine alkaloid piperine (15.4%), and the dibenzoxepin pacharin (7.5%).

Rhamnella gilgitica Attenuates Inflammatory Responses in LPS-Induced Murine Macrophages and Complete Freund's Adjuvant-Induced Arthritis Rats.

Rhamnella gilgitica Mansf. et Melch, which belongs to the rhamnus family (Rhamnaceae), is traditionally used to treat rheumatism, swelling and pain in China. However, little is known about the pharmacological activities of this plant. The anti-inflammatory activities of the 70% ethanol extract of R. gilgitica (RG) in RAW264.7 macrophages and complete Freund's adjuvant (CFA)-induced arthritic rats are investigated in this study for the first time. The effects of RG on cell viability were determined by a MTT assay, and the effects of RG on pro-inflammatory mediators were analyzed by ELISA and Western blot. The effects of RG on paw thickness, thymus and spleen index were also examined in CFA-induced arthritic rats. RG suppressed the induction of proinflammatory mediators, including iNOS (inducible nitric oxide synthase), NO (nitric oxide), COX-2 (cyclooxygenase-2) and PG (prostaglandin) E2 in LPS stimulated RAW264.7 macrophages. RG also inhibited the phosphorylation and degradation of I[Formula: see text]B-[Formula: see text], as well as the nuclear translocation of nuclear factor kappa B (NF-[Formula: see text]B) p65. In addition, RG treatment significantly reduced the paw thickness in CFA-induced arthritic rats. Oral administration of RG led to a significant decrease of both the thymus and spleen index at a concentration of 100[Formula: see text]mg/mL. Taken together, these findings suggest that RG might be an agent for further development in the treatment of a variety of inflammatory diseases.