Isoangoroside C, a phenylpropanoid glycoside from Scrophularia scorodonia roots

A new phenylpropanoid glycoside isoangoroside C was isolated from the roots of Scrophularia scorodonia. Its structure was determined on the basis of spectral data as: 3-hydroxy-4-methoxy-beta-phenylethoxy-O-alpha-L-arabinopyranosy l-(1-->6)alpha L-rhamnopyranosyl-(1-->3)-4-O-Z-feruloyl-beta-D-glucopyranoside. Additionally, one known phenylpropanoid, angoroside C, and five known iridoid glycosides, harpagoside, bartsioside, 8-O-acetyl-harpagide, aucuboside and harpagide were isolated and identified.     

Harpagoside attenuates MPTP/MPP⁺ induced dopaminergic neurodegeneration and movement disorder via elevating glial cell line-derived neurotrophic factor

Parkinson's disease is a chronic neurodegenerative movement disorder characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta. New therapeutic approaches aiming at delaying or reversing the neurodegenerative process are under active investigations. In this work, we found that harpagoside, an iridoid purified from the Chinese medicinal herb Scrophularia ningpoensis, could not only prevent but also rescue the dopaminergic neurodegeneration in MPTP/MPP(+) intoxication with promising efficacy. Firstly, in cultured mesencephalic neurons, harpagoside significantly attenuated the loss of TH-positive neuron numbers and the shortening of axonal length. Secondly, in a chronic MPTP mouse model, harpagoside dose-dependently improved the loco-motor ability (rotarod test), increased the TH-positive neuron numbers in the substantia nigra pars compacta (unbiased stereological counting) and increased the striatal DAT density ((125) I-FP-CIT autoradiography). Thirdly, harpagoside markedly elevated the GDNF mRNA and GDNF protein levels in MPTP/MPP(+) lesioned models. However, the protecting effect of harpagoside on the dopaminergic degeneration disappeared when the intrinsic GDNF action was blocked by either the Ret inhibitor PP1 or the neutralizing anti-GDNF antibody. Taken together, we conclude that harpagoside attenuates the dopaminergic neurodegeneration and movement disorder mainly through elevating glial cell line-derived neurotrophic factor.     

Harpagoside: from Kalahari Desert to pharmacy shelf

Harpagoside is an iridoid glycoside that was first isolated from Harpagophytum procumbens (devil’s claw, Pedaliaceae), a medicinal plant in which it is the major constituent of the iridoid pool. Both the pure compound and devil’s claw extracts have potent anti-rheumatic, anti-inflammatory and analgesic effects. According to the European Pharmacopoeia commercial devil’s claw products should contain at least 1.2% harpagoside. However, the compound has also been isolated from several other plant species and in vitro plant culture systems. Recent advances in knowledge of harpagoside distribution, biosynthesis/accumulation and pharmacology are summarized in this review. We also discuss the possible synergism and/or antagonism between major constituents in harpagoside-containing phytopharmaceutical products. Finally, future perspectives for its potential application are highlighted.     

Iridoids from Scrophularia genus

We report here an updated summary about iridoid composition of a series from the genus Scrophularia which have been investigated until now from a phytochemistry point of view. In addition a list is included about iridoids isolated in our laboratory from different plant parts of Scrophularia scorodonia L., which are compared with iridoids from some species of the Scrophularia genus. The present study may serve as a current information to researchers working on phytochemistry and pharmacological aspects from the Scrophularia genus and possibly to serve as a new starting point for future investigations.     

Effects of Devil's Claw (Harpagophytum procumbens) on the multidrug transporter ABCB1/P-glycoprotein

Devil's Claw (Harpagophytum procumbens) a plant native to Southern Africa, has historically been used in traditional medicine to treat a wide range of diseases and currently is widely employed as anti-inflammatory and pain-relieving natural remedy in Europe and other parts of the world.Aim of the studyLittle is known about possible herb-drug interactions arising from effects of Devil's Claw on the major drug metabolizing enzymes or transporters. This study evaluated in vitro the effects of Devil's Claw on the multidrug transporter ABCB1/P-glycoprotein.Materials and methodsThe effects of three commercially available Devil's Claw preparations and that of pure harpagoside were studied in the human kidney (HK-2) proximal tubule cell line, constitutively expressing ABCB1/P-glycoprotein (P-gp). Pgp activity and expression were tested by the calcein-AM test and by Western blotting, respectively.ResultsCommercial preparations inhibited P-gp activity, even if to a different extent, while pure harpagoside was almost ineffective. In cells cultured for three days in the presence of Devil's Claw preparations or pure harpagoside, a dose-dependent P-gp upregulation was found.ConclusionsOur results demonstrate for the first time that Devil's Claw may interact with the multidrug transporter ABCB1/P-gp, the effect not appearing strictly related to the harpagoside relative content. Modulation of both P-gp activity and P-gp expression by Devil's Claw raise the possibility of herb-drug interactions, to be further explored in depth.     

Physicochemical properties of harpagoside and its in vitro release from Harpagophytum procumbens extract tablets.

The objective of this investigation was to characterize the active-component harpagoside of Harpagophytum extract from a physico-chemical perspective and to determine its in-vitro release from tablets according to DAB 1996. It was found that both pure harpagoside and harpagoside in Harpagophytum extract have an octanol-water distribution coefficient of approximately 4 which is neither dependent on temperature nor on pH. The mean harpagoside content in Harpagophytum tablets of Batch 9102 was 16.4 mg (S.D. 0.2; S.E. 0.03). Related to a tablet weight of 365 mg (100%), this corresponds to a haragoside content of 4.5% (S.D. 0.049; S.E. 0.006). On average the tablets disintegrate after 18 +/- 3 minutes (mean +/- SD). The tablets taken from Batch 9102 released the active component harpagoside well, with a t50 of 13.5 min, a t90 of 23 min and a t95 of 25 min in relation to 16.5 mg of harpagoside per dose. Harpagoside content decreased by about 10% in artificial gastric fluid within a period of 3 hours and remained stable in artificial intestinal fluid for a period of 6 hours.     

Changes of phenylethanoid and iridoid glycoside distribution in various tissues of shoot cultures and regenerated plants of Harpagophytum procumbens (Burch.) DC. ex Meisn

Harpagophytum procumbens is a medicinal plant containing several compounds with pharmaceutical activity. Previously, we established shoot culture and in vitro regenerated plants of H. procumbens. In this study, HPLC and LC-ESI-MS were used to identify harpagoside, harpagide, verbascoside and isoverbascoside in various tissues (stems, leaves and callus) of shoots multiplied on Schenk and Hildebrandt (SH) solid medium supplemented with 0.57 μM indole-3-acetic acid (IAA) and 8 μM 6-benzylaminopurine (BAP), as well as in stems, leaves and root tubers of in vitro propagated plants grown in the greenhouse for 3, 6 and 12 months. The content of the compounds was also determined by HPLC. For comparison, control H. procumbens plants initiated from seeds were analyzed. H. procumbens shoots grown under in vitro conditions accumulated lower amounts of iridoids and phenylethanoids than the plants derived from them. The levels of analyzed compounds were higher in the organs of 3- or 6-month-old plants than in those of 12-month-old plants. Differences in the distribution of secondary metabolites were also observed between organs. The aerial parts (stems, leaves) of 3-month-old in vitro regenerated plants were characterized by the highest amounts of phenylethanoids, which significantly exceeded those detected in control plants. Total iridoid content, calculated as the sum of harpagoside and harpagide, was highest in the root tubers of 6-month-old plants. In these organs the level of harpagoside was comparable to that in root tubers of 6-month-old seed-propagated plants, but the level of harpagide was much lower.     

Teucrium francoi M. Seq., Capelo, J.C. Costa & R. Jardim, a new species of Teucrium gr. scorodonia (Lamiceae) from Madeira

The species formerly recognized as Teucrium scorodonia in Madeira is here described as new: Teucrium francoi M. Seq., Capelo, J.C. Costa & R. Jardim. Morphologically close to species of Teucrium gr. scorodonia [ T. scorodonia L., T. pseudoscorodonia Desf., T. siculum (Raf.) Guss. and T. kotschyanum Poech], it exhibits, nonetheless, some distinct diagnostic characters. The indumentum density and type of hairs of T. francoi are clearly distinct from those of related species, as are the shape and dimensions of the leaves and bracts, calyx, and corolla, which are all taken as taxonomically significant diagnostic features. A diagnosis and a distribution map are presented for this new species. Morphology, ecology, biogeography, and conservation issues are discussed. Teucrium francoi , which is an endemic from Madeira (Portugal), is to be found mostly in the scope of the association Teucrio francoi–Origanetum virentis J.C. Costa, Capelo, Jardim, Sequeira, Lousã & Rivas-Martínez, but also occurs in somewhat humid habitats, such as open stands of Rosa mandonii Déségl. associated with small streams.  © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society , 2008, 156 , 639–647.     

Preparative isolation and purification of harpagoside from Scrophularia ningpoensis hemsley by high-speed counter-current chromatography

The bioactive component harpagoside was successfully separated from the crude extract of Scrophularia ningpoensis Hemsley by one-step purification using high-speed counter-current chromatography (HSCCC). A two-phase solvent system containing n-butanol:ethyl acetate:water (1:9:10) was selected following consideration of the partition coefficient of the target compound. A 276 mg quantity of the crude extract was loaded onto a 250 mL HSCCC column and yielded 11 mg harpagoside at over 97% purity. The chemical structure of harpagoside was determined by HPLC-ESI/MS and 1H-NMR.     

Occurrence of iridoid glycosides in in vitro cultures and intact plants of <Emphasis Type="Italic">Scrophularia nodosa</Emphasis> L.

Shoot, root, and callus cultures of Scrophularia nodosa L. (Scrophulariaceae) were established and cultivated in vitro. Iridoid glycosides, such as harpagoside, aucubin, and catalpol were identified by LC-ESI-MS and their contents determined by HPLC. For comparison intact plants of S. nodosa were analysed. In shoot cultures slightly lower amounts of detectable iridoid glycosides (4.36% dry weight) were determined than in the field grown plants (4.88%). Concentration of harpagoside was highest in leaves of field plants (1.05%) and in flowers of in vitro plantlets (1.10%). For aucubin the highest amount was found in the leaves of in vitro plantlets (1.67%) whereas the levels of aucubin in the leaves of field plants were remarkably lower. Catalpol was produced as a trace compound in intact plants and shoot cultures. Callus and root cultures were apparently not able to synthesise iridoid glycosides.     

High anti-inflammatory activity of harpagoside-enriched extracts obtained from solvent-modified super- and subcritical carbon dioxide extractions of the roots of Harpagophytum procumbens

Solvent-modified carbon dioxide extractions of the roots of Harpagophytum procumbens have been investigated with respect to extraction efficiency and content of harpagoside, and compared with a conventional extract. The effects of pressure, temperature, type and concentration of the modifier have been examined. Two extraction steps were necessary in order to achievehigh anti-inflammatory harpagoside-enriched extracts. The first extraction step was carried out in the supercritical state using carbon dioxide modified with n-propanol to remove undesired lipophilic substances. The main extraction was performed either in the supercritical or in the subcritical state with carbon dioxide modified with ethanol. The supercritical fluid extraction resulted in extracts containing up to 30% harpagoside. The subcritical extracts showed a harpagoside content of ca. 20%, but the extraction yield was nearly three times greater compared with supercritical conditions. The total harpagoside recovery resulting from the sum of the extract and the crude drug residue was greater than 99% in all experiments. The conventional extract and two carbon dioxide extracts were tested for in-vitro inhibition of 5-lipoxygenase or cyclooxygenase-2 biosynthesis. Both carbon dioxide extracts showed total inhibition on 5-lipoxygenase biosynthesis at a concentration of 51.8 mg/L. In contrast, the conventional extract failed to show any inhibition of 5-lipoxygenase biosynthesis.     

Phenylpropanoid glycosides from Scrophularia scorodonia: in vitro anti-inflammatory activity

Five phenylpropanoid glycosides isolated from Scrophularia scorodonia L. (Scrophulariaceae), namely angoroside A (1), angoroside C (2), angoroside D (3), acteoside (4) and isoacteoside (5), had been evaluated as potential inhibitors of some macrophage functions involved in the inflammatory process. These compounds have been tested in two experimental systems: ionophore-stimulated mouse peritoneal macrophages and human platelets serve as source of COX-1 and 5-LOX, and mouse peritoneal macrophages stimulated with E. coli LPS are the means of testing for COX-2, NO and TNF-alpha activity. None of compounds assayed had a significant effect on LTC(4)-release from calcium ionophore-stimulated mouse peritoneal macrophages. However, the release of PGE(2) by mouse peritoneal macrophages stimulated with calcium ionophore was inhibited by most of these compounds. In the TXB(2)-release assay, acteoside (4), angoroside A (1) and angoroside C (2) showed a significant effect. These five compounds, except angoroside C (2) significantly inhibited LPS-induced PGE(2), NO and TNF-alpha in a concentration-dependent manner. In LPS-stimulated macrophages, the phenylpropanoid glycoside angoroside C (2) only had activity on NO. These results indicate that the pharmacology of these compounds may participate in the anti-inflammatory effect of Scrophularia scorodonia.     

Comparative phytochemical analysis of wild and in vitro-derived greenhouse-grown tubers, in vitro shoots and callus-like basal tissues of Harpagophytum procumbens

Comparative phytochemical analysis of wild and in vitro-derived greenhouse-grown tubers, in vitro shoots and callus-like basal tissues of Harpagophytum procumbens was done. Dried samples were ground to fine powders and their total iridoid (colorimetric method), phenolic [Folin-Ciocalteu (Folin C) method] and gallotannin (Rhodanine assay) contents as well as anti-inflammatory activity [cyclooxygenase assays (COX-1 and COX-2)] were determined. The tissue culture-derived tubers had the highest total iridoid content which was significantly higher than that of the tubers collected from the wild and other tissue cultured materials evaluated. This suggests that cultivated plants can be a viable alternative source of the active principle(s). The total phenolic and gallotannin contents of the wild tubers were significantly higher than the tissue culture-derived tubers and other in vitro-derived plant materials. The presence of phenolic compounds including gallotannins in the tissue cultured materials is of interest from a pharmacological point of view given the pharmacological role of phenolics. In general, extracts from wild tubers demonstrated better inhibitory activities in both COX-1 and COX-2 assays when compared to the tissue culture-derived tubers. All the petroleum ether (PE) and dichloromethane (DCM) extracts showed moderate (50-70%) to good (> 70%) inhibitory activities whereas the ethanol (EtOH) extracts showed poor or no inhibition in both assays. Based on previous reports indicating weak inhibition of COX-2 enzyme by harpagoside, the inhibitory activities of both COX enzymes exhibited by PE and DCM extracts in the current study could be due to the presence of other constituents in the extracts. This points towards the need to identify other active constituents and evaluate their role and mode of action in relation to harpagoside — the main active principle.     

金沙江干热河谷种子植物区系特征的初探

研究涉及金沙江云南境内的干热河谷和干暖河谷的种子植物区系。建立名录共920种,有野生植物748种。这748种,分属于111科,427属。427属的分布区类型的格局为：l－44,2—127,3—10,4—39,5—13,6—37,7—29,8—47,9—12,10—25,11—3,12—7,13—l,14—20,15—13．在748种中,三江（金沙江、元江、怒江）干热河谷共有的标志植物73种,金沙江的标志植物156种,它们是研究区系特征的基础。经分析,本区植物区系有以下特征：(1)属、种均以热带成分为主,热带属占66.58%,种的热带成分更多,均交错少数温带成分;(2)三江干热河谷共同标志植物中,多数是优势种或常见种,例如Heteropogon contortus, Bombax ceiba,Phyllanthus emblica,Euphorbia royleana,Tribulus terrestris等,地区特有成分则少,如Terminalia franchetii,Andropogon yunnanensis;(3)滇北、川西南河谷特有成分则较多,如Cycas panzhihuaensis,Nouelia insignis,Aristolochia delavayi等;(4)西南特有成分也较多,且在河谷常见,如Delavaya yunnanensis,Croton yunnanensis,Munronia delavayi等;(5)有一些非特有的特殊种类,如Heteropogon melanocarpus,Dinebra retroflexa等.鉴于特征分析,本区植物区系有以下几个更深层的特征:(1)热带起源与近代交错现象;(2)有古地中海区系的后裔;(3)与近代非洲与南亚区系有联系;(4)与华北区系有联系;(5)特有现象明显而成因复杂,等。     

An in vivo microdialysis measurement of harpagoside in rat blood and bile for predicting hepatobiliary excretion and its interaction with cyclosporin A and verapamil

Harpagoside, a major bioactive iridoid glucoside in genus Scrophularia, has been widely used in clinical practice for the treatment of pain in the joints and lower back for its neuroprotective and anti-inflammation activities. To investigate the pharmacokinetics and hepatobiliary excretion, an in vivo microdialysis method coupled with high performance liquid chromatography was developed to monitor the concentration of harpagoside in blood and bile. The harpagoside bile-to-blood distribution ratio (AUC_bile/AUC_blood) up to 986.28 ± 78.46 significantly decreased to 6.41 ± 0.56 or 221.20 ± 18.92 after co-administration of cyclosporin A or verapamil. The results indicated that harpagoside went through concentrative elimination from the bile which was probably regulated by P-glucoprotein, providing possible clinical trials of co-administration of transporter inhibitors to decrease drug efflux, thus to enhance the curative effects.     

Screening of plant extracts for antimicrobial activity against bacteria and yeasts with dermatological relevance

There is cumulative resistance against antibiotics of many bacteria. Therefore, the development of new antiseptics and antimicrobial agents for the treatment of skin infections is of increasing interest. We have screened six plant extracts and isolated compounds for antimicrobial effects on bacteria and yeasts with dermatological relevance. The following plant extracts have been tested: Gentiana lutea, Harpagophytum procumbens, Boswellia serrata (dry extracts), Usnea barbata, Rosmarinus officinalis and Salvia officinalis (supercritical carbon dioxide [CO2] extracts). Additionally, the following characteristic plant substances were tested: usnic acid, carnosol, carnosic acid, ursolic acid, oleanolic acid, harpagoside, boswellic acid and gentiopicroside. The extracts and compounds were tested against 29 aerobic and anaerobic bacteria and yeasts in the agar dilution test. U. barbata-extract and usnic acid were the most active compounds, especially in anaerobic bacteria. Usnea CO2-extract effectively inhibited the growth of several Gram-positive bacteria like Staphylococcus aureus (including methicillin-resistant strains - MRSA), Propionibacterium acnes and Corynebacterium species. Growth of the dimorphic yeast Malassezia furfur was also inhibited by Usnea-extract. Besides the Usnea-extract, Rosmarinus-, Salvia-, Boswellia- and Harpagophytum-extracts proved to be effective against a panel of bacteria. It is concluded that due to their antimicrobial effects some of the plant extracts may be used for the topical treatment of skin disorders like acne vulgaris and seborrhoic eczema.     

P2X7-Receptor Pathway Involvement in the Anti-Inflammatory Activity of Medicinal Plants

Medicinal plants used in European folk medicine attached to Lamiales, Gentianales or Asterales orders are used to treat inflammatory disorders. Many targets have been identified but to date, implication of purinergic receptor P2X7 activation has not yet been investigated. We managed to evaluate the protective effect on P2X7 activation by plant extracts used as anti-inflammatory in European folk medicine by the YO-PRO-1 uptake dye in vitro bioassay. Results revealed that among our selected plants, species from Scrophularia and Plantago genus were able to decrease significantly P2X7 activation (>50 % at 0.1 and 1 μg/mL). UPLC/MS, dereplication and metabolomic analysis of Scrophularia extracts, allowed us to identify the cinnamoyl-iridoid harpagoside as putative inhibitor of P2X7 activation. These results open a new research field regarding the anti-inflammatory mechanism of cinnamoyl-iridoids bearing plants, which may involve the P2X7 receptor.     

CHEMICAL CONSTITUENTS FROM SCROPHULARIA NINGPOENSIS

A new acyi sugar along with four know iridoidal and phenyl propanoid glycosides, harpagoside, harpagide, 0-methyl catalpol and angoroside C, were isolated from the roots of Scrophularia ningpoensis Hemsl., a famous traditional Chinese medicine known as "Xuan-Shem". By means of chemical and spectral evidence its structure was elucidated as 4-0-(p-methoxycinnamoyl)-α-L-rhamnopyranose.     

Production of Iridoids and Phenolics by Transformed Harpagophytum procumbens Root Cultures

Harpagophytum procumbens (Devil's claw) is an important medicinal plant, which tubers are used for the treatment of different inflammatory diseases. In this study, the time courses of growth of Devil's claw hairy roots and accumulation of intracellular total iridoids and phenolics were investigated during cultivation under submerged conditions. After 21 days of growth in liquid hormone‐free MS medium, a growth index of 81.3 was achieved and the accumulated biomass reached 0.58 g/flask. It was found that the time courses of total iridoids and phenolics production showed almost the same patterns with a maximum on day 21 from the beginning of cultivation (15.93 mg harpagoside equivalents/L and 261 mg gallic acid equivalents/L, respectively). The methanolic extracts possessed relatively high DPPH‐radical scavenging properties (IC₅₀ 4.08 mg/mL), while for the culture media no antiradical activity was detected. According to our best knowledge, the present report is the first one dealing with the investigation of transformed root cultures from Harpagophytum procumbens plants.