Objective: Antinociceptive and anti-inflammatory activities of hydroalcoholic extract of Teucrium Oliverianum were investigated by formalin test model. This study was conducted in on the male Wistar rats, weighting 150-180 g. The animals were divided into seven groups (n = 7) and received 200, 400, 600 and 800 mg kg(-1) of hydroalcoholic extract of teucrium oliverianum intraperitoneally, respectively. Negative control group received normal saline (5 mL kg(-1)) and the positive control groups received 2.5 mg kg(-1) morphine and 300 mg kg(-1) aspirin, intraperitoneally respectively. The results showed that all doses of extract have significant analgesic effect (p < 0.05) in all studies times in comparison with negative control. The best result achieved with 600 mg kg(-1) of extract. The result revealed that the analgesic effect of the extract (600 mg kg(-1))\was less than aspirin (300 mg kg(-1)) on the second phase of pain and less than morphine (2.5 mg kg(-1)) in both phases of the pain, more than aspirin in first phase of pain. One group of animals was treated with naloxone (1 mg kg(-1), i.p.) and suitable dose of extract (600 mg kg(-1), i.p.). Also, Naloxone inhibited analgesic effect of alcoholic extract of Teucrium Oliverianum. It can be concluded that the alcoholic extract of Teucrium oliverianum may exert its effect through opioid receptors, stimulating GABAergic system or promotes the release of endogenous opipeptides or decreasing free radicals.

The chemical composition of the essential oil of the aerial parts of Teucrium polium ssp. capitatum collected during the flowering period from rocky places and dry pastures,(Serbia, Nis, Kamenica), and dunes along the sea-side,(Bulgaria, Burgas, Primorsko) has been studied by GC and GC/MS. The identified compounds, 45 for the oil from Serbia and 44 for that from Bulgaria, amounted to 97.3% and 96.4% of the oils, respectively. The dominant constituents of the Serbian oil were sesquiterpenes {59.6%, the most abundant components being germacrene D (31.8%), trans-caryophyllene (8.8%) and bicyclogermacrene (6.2%)}, while monoterpenes accounted for 37.5%{the most abundant components being linalool (14.0%) and beta-pinene (10.7%)}. The essential oil from Bulgaria was characterized by a high percentage of monoterpenes, amounting to 62.9%{the most abundant components being beta-pinene (26.8%), alpha-pinene (9.3%) and limonene (6.4%)}, while sesquiterpenes accounted for 33.5%(the most abundant component was germacrene D 17.7%)}.

The volatile constituents of Teucrium scordium L. ssp. scordioides, T. polium, and T. montanum, obtained by hydrodistillation, were investigated by GC-FID and GC/MS analyses. A total of 296 constituents were identified, representing 89.8-98.4% of the oil compositions. The oils of T. polium and T. montanum consisted mainly of sesquiterpenes (64.3 and 72.7%, resp.), with germacrene D (4; 31.0%) and δ-cadinene (10; 8.1%) as the main constituents, respectively. In contrast, the monoterpene menthofuran (1; 11.9%) predominated in the oil of T. scordium ssp. scordioides, and this clearly distinguished this species from the other Teucrium taxa investigated up to date. The chemistry of the volatiles of eight Teucrium taxa from Serbia and Montenegro were compared using multivariate statistical analysis, and this provided chemotaxonomically important conclusions.

Chemical and genetic diversity of Teucrium scorodonia L. subsp. scorodonia from Corsica and T. scorodonia L. subsp. baeticum from western Algeria were investigated. Diversity within and among the two populations of subspecies was assessed according to the chemical composition of their essential oils, and genetic diversity was evaluated using three polymorphic genetic markers. Chemical analysis was performed using a combination of capillary GC-RI and GC/MS after fractionation using column chromatography. Genetic structures were mapped using two chloroplast markers (RPL32-TRNL and TRNL-F) and ribosomal nuclear markers (ITS region). The statistical analysis showed that the two subspecies were clearly distinguished by these chemical and genetic markers. The chemical composition of oil differed qualitatively and quantitatively between the subspecies. Corsican oil samples contained germacrene B (4.2-8.8%) and γ-elemene (2.6-5.7%), which were not detected in Algerian oil samples. The oils of the scorodonia and baeticum subspecies were dominated by sesquiterpene hydrocarbon compounds (75.6-82.9% and 69.6-79.4%, respectively), but they differed in oxygenated sesquiterpene content (3.1-8.9% and 8.4-20.3%, respectively). Neighbor-joining trees constructed from chloroplast DNA and ITS region sequences showed the existence of two groups associated with taxonomic and chemical characteristics. One group consisted of T. scorodonia subsp. scorodonia and the other of T. scorodonia subsp. baeticum, indicating that variation in the essential oil composition of T. scorodonia subspecies depends more on genetic background than environmental characteristics.

BACKGROUND Saponins isolated from plant sources have a number of traditional and industrial applications. Saponins have pharmacological effects like anti-inflammatory, molluscicidal, antimicrobial, antispasmodic, antidiabetic, anticancer, anticonvulsant, anthelmintic, antitussive and cytotoxic activities. The current work describes the anthelmintic and cytotoxic activities of crude saponins of Achillea Wilhelmsii and Teucrium Stocksianum as these plants are rich with saponins. METHODS Brine shrimp cytotoxic activity of crude saponins was determined by Meyer et al.(1982) at test concentrations of 1000 μg/ml, 100 μg/ml, 10 μg/ml, 7.5 μg/ml, 5.0 μg/ml, 2.5 μg/ml and 1.25 μg/ml. Percentage mortality of test concentrations was determined. Similarly, in vitro anthelmintic activity was determined against roundworms, tapeworms and earthworms. Albendazole and piperazine citrate at concentration 10 mg/ml were used as standard anthelmintic drugs. RESULTS Crude saponins of Achillea wilhelmsii (CSA) and Teucrium stocksianum (CST) had, respectively, cytotoxic activity with LC50 values 2.3±0.16 and 5.23±0. 34 μg/ml. For in vitro anthelmintic activity, time for paralysis and death of parasites (parasiticidal activity) was noted. At concentration 40 mg/ml, crude saponins of Achillea wilhelmsii are 1.96 and 2.12 times more potent than albendazole against Pheretima posthuma and Raillietina spiralis, respectively. Similarly, at concentration 40 mg/ml, crude saponins of Teucrium stocksianum (CST) has 1.89, 1.96 and 1.37 times more parasiticidal activity than albendazole against Pheretima posthuma, Raillietina spiralis and Ascardia galli, respectively. CONCLUSION Crude saponins of Achillea wilhelmsii and Teucrium stocksianum have cytotoxic and anthelmintic activity. The crude saponins may be excellent sources of cytotoxic and anthelmintic constituents that warrant its isolation and purification for new drug development.

Amoebic keratitis is difficult to treat without total efficacy in some patients because of cysts, which are less susceptible than trophozoites to the usual treatments. The aim of this study is to evaluate the in vitro amoebicidal activity of the methanolic extracts of Teucrium polium and Teucrium chamaedrys. In the presence of methanolic extracts (ranging from 1.0 to 32.0 mg/ml), numbers of the viable Acanthamoeba castellani trophozoites and cysts were decreased during the experimental process. Both extracts showed time- and dose-dependent amoebicidal action on the trophozoites and cysts. Among the extracts tested, T. chamaedrys showed the strongest amoebicidal effect on the trophozoites. In the presence of 16 mg/ml or above extract concentrations, no viable trophozoites were observed within 48 h. In the case of T. polium, no viable trophozoites were observed within 48 h at 32 mg/ml concentration. As expected, cysts were found more resistant to the extracts than the trophozoites.

Fractionation of the chloroformic extracts from Teucrium ramosissimum leaves resulted in the isolation of three flavonoids: genkwanin (1), cirsimaritin (2) and 4',7-dimethoxy apigenin (4) and one sesquiterpene: β-eudesmol (3). The structures were determined using data obtained from (1)H and (13)C NMR spectra, as well as by various correlation experiments (COSY, HMQC and HMBC). The antioxidant activities of the isolated flavonoids from T. ramosissimum leaves were evaluated by measuring their ability to scavenge the radical ABTS(+) and through chemical assays: cupric reducing antioxidant capacity (CUPRAC), reducing power (RP) and ferric reducing antioxidant power (FRAP). Furthermore, the effects of T. ramosissimum isolated molecules, on inhibition of cell proliferation and induction of apoptosis in human leukemia cells, were also examined. Cirsimaritin showed the best activity in the ABTS assay with TEAC value 2.04μM, whereas apigenin and 4',7-dimethoxy apigenin exhibited the highest antioxidant activity using the CUPRAC, RP and FRAP assays with TEAC values 10.5, 1.39 and 0.71μM respectively. The cytotoxic activity revealed that the β-eudesmol inhibited significantly the proliferation of K562 cells (IC(50)=20μg/ml).

The essential oil of Teucrium arduini L. was characterized by a high concentration of sesquiterpene hydrocarbons (43.8%) of which beta-caryophyllene (19.9%) being the major compound, followed by oxygenated sesquiterpenes (19.6%) of which caryophyllene-oxide (14.6%) was dominant. When applied to plants of Chenopodium amaranticolor and Ch. quinoa for two successive days prior inoculation, the oil was effective in reducing lesion numbers on plants infected with Tobacco mosaic virus (25.7%) and Cucumber mosaic virus (21.9%). The main components of oil, beta-caryophyllene and caryophyllene oxide showed potent antiviral activity against CMV, but weak activity against TMV infection.

Seven neo-clerodanes (teupolins VI-XII) and eleven known compounds were isolated and characterized from leaf extracts of Teucrium polium L., a medicinal plant used in traditional and herbal medicine for its hypolipidemic, hypoglycemic, antioxidant and antiproliferative properties. The structures of these compounds were elucidated by 1D (1H, 13C and DEPT) and 2D (COSY, TOCSY, HSQC, HMBC) NMR experiments and by mass spectrometry analysis. The complete stereostructure of each compound was defined with a NOESY experiment. Because the overexploitation of herbal remedies containing T. polium extracts has resulted in several cases of hepatitis, the hepatotoxic activity of pure metabolites against the human hepatoblastoma cancer cell line HepG2 was assessed by the MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) test. All of the compounds showed low toxicity values at the highest concentration tested (200 μM).

Teucrium chamaedrys L. is a herbaceous plant common in European woods that has been used for many purposes for centuries. Recently, T. chamaedrys L. has become popular as a slimming decoction without any scientific proof of efficacy notwithstanding its well-known hepatotoxicity. Hydroalcoholic extracts are currently used as flavourings in the preparation of wines, bitters and liqueurs. Teucrin A and teuchamaedryn A are the major toxic components of the diterpenoid fraction of T. chamaedrys L.

ETHNOPHARMACOLOGICAL RELEVANCE Teucrium chamaedrys L. and Nepeta cataria L.(Lamiaceae) are species with traditional uses that relate to the treatment of inflammation. Extracts of both species were found to inhibit calcineurin; an important regulator of T-cell mediated inflammation that has received little attention in ethnopharmacological research. MATERIALS AND METHODS Extracts and isolated compounds were tested against calcineurin in its calmodulin-activated and basal un-activated state. Active compounds were isolated using Sephadex LH-20 gel filtration and HPLC then identified using NMR spectroscopy. RESULTS AND CONCLUSIONS Activity-guided fractionation of Teucrium chamaedrys and Nepeta cataria led to the isolation of the caffeoyl phenylethanoid glycosides teucrioside, verbascoside and lamiuside A (teupolioside). The three compounds inhibited calcineurin both in the presence and absence of calmodulin, suggesting a direct interaction with calcineurin. Calcineurin inhibition should be considered as a potential mode of action when investigating the immunomodulatory activity of caffeoyl phenylethanoid glycoside containing plants.

Scutellaria lateriflora, commonly known as skullcap, is used as an ingredient in numerous herbal products. However, it has been occasionally adulterated/contaminated with Teucrium canadense and/or Teucrium chamaedrys, commonly known as germander, due to the morphological similarities between the two genera. The latter contains hepatotoxic diterpenes. Despite the potential hepatotoxicity introduced by germander contamination, analytical methodologies for the authentication and quality assessment of S. lateriflora-based dietary supplements have not been reported. In this study, a flow-injection/mass spectrometry fingerprinting method in combination with principal component analysis was used to survey S. lateriflora-based dietary supplements sold in the USA.

Teucrium ramosissimum (Lamiaceae), a native and endemic plant from South Tunisia, has traditionally been used as a treatment for inflammation and for ulcers. Though the plant and its products are widely used, very few studies have analyzed the pharmacological/toxicological properties of this plant. Thus, the aim of these studies was to evaluate the anti-inflammatory/anti-ulcerogenic activities of various extracts (i.e., methanolic, aqueous, and total oligomer flavonoid [TOF]-enriched) from leaves of T. ramosissimum. In vitro, the effects from each extract on lysosomal enzyme activity and proliferation of, respectively, freshly isolated peritoneal macrophages and splenic lymphocytes were assessed. The extracts alone clearly affected macrophage function, as evidenced by a significant modulation of cell lysosomal enzyme activity and ability to form and/or release nitric oxide. These extracts were also found to be able to significantly modify the proliferation of splenocytes, even when lipopolysaccharide or lectin mitogens were absent. With respect to the anti-ulcerogenic activity of the extracts, these studies found that the leaf extracts were able to exert significant protective effects against ethanol-induced ulcers in a rat model; at some doses, the extract effects were even greater than that obtained using a cytoprotective histamine H2-antagonist, cimetidine. Based on these studies, we conclude that the extracts from T. ramosissimum appear to be potentially potent modulators of innate immunity and that their efficacy against ulcer formation may be due, in part, to a cytoprotective effect. Further, these results fortify the ethnopharmacological importance of the use of T. ramosissimum products as anti-inflammatory and anti-ulcer agents. Nevertheless, ongoing/further studies are needed to clarify more precisely mechanisms underlying effects against ulcers and on lymphocyte and macrophage functionality, as well as the causative agents.

CONTEXT The main objective of the study was to investigate the biochemical mechanism of the antidiabetic activities of the dry extracts of Teucrium polium L. ssp. capitatum (L.) Arcangeli (Lamiaceae), from Republic of Macedonia, traditionally used to treat diabetes. MATERIALS AND METHODS Aerial parts of the plant were extracted in alcohol and freeze- or spray-dried, analyzed by high performance liquid chromatography (HPLC) and examined for insulinotropic effect in INS-1E cells in vitro. Their effect on blood glucose, lipids and carbohydrate-related enzymes was tested in normo- and streptozotocin hyperglycemic rats. RESULTS AND DISCUSSION HPLC analyses revealed several flavonoids: luteolin, apigenin, cirsiliol, diosmetin, cirsimaritin and cirsilineol as both free aglycons and glycosides. The extract and mixture of commercial flavonoids showed a distinct insulinotropic effect on INS-1E cells at 500 µg/ml. Intragastric (i.g.) administration of identical doses of the extract (125 mg/kg) in both normo- and hyperglycemic rats was more efficient in lowering the blood glucose than intraperitoneal injection (35% vs. 24% reduction) with highest effect (50% reduction) 8 h after administration. After 10 days of treatment, the magnitude of the effect was comparable to i.g. administration of 2.5 mg/kg of glibenclamide (38% reduction). No effect was seen on blood lipid profiles. In OGTT (oral glucose tolerance test), the extract lowered blood glucose levels by ~35%. The treatment reduced hepatic glycogen and tended to normalize the activity of gluconeogenic enzymes. CONCLUSION The results demonstrate that examined plant extracts contain flavonoids with insulinotropic and antihyperglycemic effects.

Corsica Island exhibited the particularity to display Teucrium flavum subsp. glaucum and subsp. flavum on the same territory with the same bioclimatic conditions. For the first time, volatile components extracted from aerial parts and genetic diversity of both Corsican T. flavum L. subspecies have been investigated through (i) the characterization of the chemical composition of essential oils and (ii) the study of three polymorphic genetic markers. Chemical analysis were performed using combination of capillary GC/RI, GC-MS after fractionation on column chromatography and the definition of the genetic structure were carried out using two chlororoplast markers (RPL32-TRNL and TRNL-F) and ribosomal nuclear markers (ITS region). According to statistical analysis, both subspecies were clearly distinguished by the chemical and genetic studies. Chemical compositions of oils from both subspecies were qualitatively similar but they differed by the normalized% abundances of their major components; oils from subsp. flavum were dominated by large amounts of hydrocarbon monoterpenes while oils obtained from subsp. glaucum were characterized by higher amounts of oxygenated compounds. The genetic analysis divided T. flavum L. populations in two groups, the first displayed subsp. glaucum populations and the latter group exhibited subsp. flavum populations. The presence of two groups is weakly consistent with chemical differentiation. These data suggest that the differences in the volatile composition of the two T. flavum subspecies depends more on the genetic background and less on environmental factors.

The essential oils of four Teucrium species were studied and 131 components, in all, were identified. All oils were rich in sesquiterpenes (50.0-61.9%). Caryophyllene and caryophyllene oxide were the main components of Teucrium arduini; germacrene D, delta-cadinene and gamma-cadinene predominated in Teucrium maghrebinum. Carvacrol and caryophyllene predominated in Teucrium polium ssp. capitatum, while carvacrol, caryophyllene oxide and caryophyllene were the most abundant components in Teucrium montbretii ssp. heliotropiifolium. The germination of radish and garden cress was less sensitive to the four essential oils. The radicle elongation, above all, of radish was significantly inhibited by all oils, in particular by the essential oil of T. arduini, at the highest doses tested. Among the main components of the oils, monoterpenes resulted the more active compounds.

ETHNOPHARMACOLOGICAL RELEVANCE Achillea santolina L., Pistacia atlantica Desf, Rheum ribes L., Sarcopoterium spinosum (L.) Spach and Teucrium polium L. have traditionally been used as herbal antidiabetic medicines. However their alleged benefits and mechanisms remain elusive. AIM OF THE STUDY This study aimed to evaluate the effect of water extracts of these plants in in vitro and in vivo experiments. MATERIALS AND METHODS In vitro enzymatic starch digestion with aqueous extracts from plants at concentrations of 1, 5, 10, 12.5, 25, 50 and 100 mg/ml was assayed using α-amylase and α-amyloglucosidase. Acarbose was used as control and glucose liberation was determined by glucose oxidase method. Oral starch tolerance test (OSTT) and oral glucose tolerance test (OGTT) were determined for the plant extracts at concentrations 125, 250 and 500 mg/kg b.wt. on Sprague-Dawley rats. Blood glucose levels in rats treated with plant extracts and drugs (acarbose or metformin and glipizide) were measured at -30, 0, 45, 90 and 135 min. RESULTS Compared to acarbose (IC(50)=1.2 μg/ml), water extracts of Pistacia atlantica, Rheum ribes and Sarcopoterium spinosum exerted significant dose dependent dual inhibition of α-amylase and α-glucosidase in in vitro enzymatic starch digestion bioassay, with IC(50)s; 46.98, 58.9 and 49.9 mg/ml, respectively. Comparable in vivo results were obtained for starch-fed rats, exhibiting significant acute postprandial antihyperglycemic efficacies. While Achillea santolina and Teucrium polium extracts lacked any favourable in vitro anti-α-amylase and anti-α-glucosidase effect, other modes of action can possibly explain their substantial acute antihyperglycemic activities in starch-treated rats. Except for Pistacia atlantica extracts, none of the investigated extracts qualified for improving the glucose intolerance in fasted rats on glucose loading. CONCLUSIONS Pistacia atlantica, Rheum ribes and Sarcopoterium spinosum can be considered as potential candidates for amelioration/management of type 2 diabetes.

The mutagenic and antimutagenic effects of the essential oil extracted from the aerial parts of Teucrium ramosissimum were evaluated by the bacterial reverse mutation assay in Salmonella typhimurium TA98, TA100, and TA1535, with and without exogenous metabolic activation (S9 fraction). The T. ramosissimum essential oil showed no mutagenic effect. In contrast, our results established that it possessed antimutagenic effects against sodium azide (SA), aflatoxin B1 (AFB1), benzo[a]pyrene (B[a]P), and 4-nitro-o-phenylenediamine (NOPD). The antioxidant capacity of the tested essential oil was evaluated using enzymatic, i.e., the xanthine/xanthine oxidase (X/XOD) assay, and nonenzymatic systems, i.e., the nitro-blue tetrazolium (NBT)/riboflavin and the DPPH assays. A moderate free radical-scavenging activity was observed towards DPPH(.) and O2(.-). In contrast, T. ramosissimum essential oil showed no effect for all the tested concentrations in the X/XOD assay.

Nonalcoholic fatty liver disease, the most common chronic liver disorder, is frequently associated with the clinical features of metabolic syndrome such as insulin resistance. We aimed to determine the effect of the crude and the ethyl acetate extracts of Teucrium polium on insulin resistance in rats with nonalcoholic steatohepatitis. Rats were divided into four groups. Group A was fed a normal diet for 11 weeks. Nonalcoholic steatohepatitis was induced in the remaining groups using a methionine/choline deficient (MCD) diet for 8 weeks. After nonalcoholic steatohepatitis development, group B continued with receiving the MCD diet alone; group C rats were given the MCD diet along with crude extract of T. polium (equivalent to 1 g leaves powder/kg body weight/day); group D rats were given the ethyl acetate fraction of T. polium by intragastric administration for 3 weeks. MCD diet led to grade 1 liver steatosis. In group C and D, these factors abated to grade 0 in 80% of the rats. In the groups receiving the extract, lipoprotein profiles were significantly improved relative to those not receiving the extract. Also, a dramatic reduction was observed in sera alkaline phosphatase, aspartate aminotransferase and alanine aminotransferase activities. In addition, in groups C and D, an increase in the activities of liver superoxide dismutase, glutathione peroxidase and glutathione reductase enzymes were also associated with a decrease in the malondialdehyde level relative to group B. Moreover, both extracts significantly decreased plasma glucose and insulin levels along with insulin resistance. In conclusion, both extracts of T. polium could reverse the adverse effects of an MCD diet.

Teucrium divaricatum Sieb. ssp. villosum (Celak.) Rech. fil. is a medicinal plant used in Lebanon. The hydrodistillation of the aerial parts of T. divaricatum yielded 0.27%(wt/wt) of essential oil; gas chromatography and gas chromatography/mass spectrometry analysis enabled the identification of 60 compounds representing 94.5% of the total oil. On the whole, the oil was constituted mainly by sesquiterpenes (64.6%), particularly sesquiterpene hydrocarbons (50.0%). The most abundant compounds were (E)-caryophyllene (30.1%) and caryophyllene oxide (6.1%). The in vitro activity of the essential oil against some microorganisms in comparison with gentamicin by the broth dilution method was determined. The oil exhibited a good activity as an inhibitor of growth of Gram-positive bacteria.