Mentha piperita as a pivotal neuro-protective agent against gamma irradiation induced DNA fragmentation and apoptosis

Ionizing radiation is classified as a potent carcinogen, and its injury to living cells, in particular to DNA, is due to oxidative stress enhancing apoptotic cell death. Our present study aimed to characterize and semi-quantify the radiation-induced apoptosis in CNS and the activity of Mentha extracts as neuron-protective agent. Our results through flow cytometry exhibited the significant disturbance and arrest in cell cycle in % of M1: SubG1 phase, M2: G0/1 phase of diploid cycle, M3: S phase and M4: G2/M phase of cell cycle in brain tissue (p < 0.05). Significant increase in % of apoptosis and P53 protein expression as apoptotic biomarkers were coincided with significant decrease in Bcl₂ as an anti-apoptotic marker. The biochemical analysis recorded a significant decrease in the levels of reduced glutathione, superoxide dismutase, deoxyribonucleic acid (DNA) and ribonucleic acid contents. Moreover, numerous histopathological alterations were detected in brain tissues of gamma irradiated mice such as signs of chromatolysis in pyramidal cells of cortex, nuclear vacuolation, numerous apoptotic cell, and neural degeneration. On the other hand, gamma irradiated mice pretreated with Mentha extract showed largely an improvement in all the above tested parameters through a homeostatic state for the content of brain apoptosis and stabilization of DNA cycle with a distinct improvement in cell cycle analysis and antioxidant defense system. Furthermore, the aforementioned effects of Mentha extracts through down-regulation of P53 expression and up-regulation of Bcl₂ domain protected brain structure from extensive damage. Therefore, Mentha extract seems to have a significant role to ameliorate the neuronal injury induced by gamma irradiation.     

Giardia lamblia: the effects of extracts and fractions from Mentha x piperita Lin. (Lamiaceae) on trophozoites

Giardia lamblia is a parasite that causes giardiasis in humans and other mammals. The common treatment includes different classes of drugs, which were described to produce unpleasant side effects. Mentha x piperita, popularly known as peppermint, is a plant that is frequently used in the popular medicine to treat gastrointestinal symptoms. We examined the effects of crude extracts and fractions from peppermint against G. lamblia (ATCC 30888) on the basis of trophozoite growth, morphology and adherence studies. The methanolic, dichloromethane and hexanic extracts presented IC(50) values of 0.8, 2.5 and 9.0microg/ml after 48h of incubation, respectively. The aqueous extract showed no effect against the trophozoites with an IC(50)>100microg/ml. The aqueous fraction presented a moderate activity with an IC(50) of 45.5microg/ml. The dichloromethane fraction showed the best antigiardial activity, with an IC(50) of 0.75microg/ml after 48h of incubation. The morphological and adhesion assays showed that this fraction caused several alterations on plasma membrane surface of the parasite and inhibited the adhesion of G. lamblia trophozoites. Cytotoxic assays showed that Mentha x piperita presented no toxic effects on the intestinal cell line IEC-6. Our results demonstrated antigiardial activity of Mentha x piperita, indicating its potential value as therapeutic agent against G. lamblia infections.     

Improved menthol production from chitosan-elicited suspension culture of Mentha piperita

The optimum concentration of chitosan to menthol production by Mentha piperita cells cultured in shake flasks was 200 mg/l, which gave 166 mg menthol/l after 12 days. Chitosan elicitation may activate the conversion of pulegone to menthol.     

椒样薄荷试管苗生根的影响因素分析

椒样薄荷(M entha piperitaL.)为多年生宿根型长日照植物,原产美国密执安、威斯康星及印第安纳等地[1],中国黑龙江、陕西、新疆、河北、江苏、浙江及安徽等地有少量引种栽培[2~4]。从其茎叶中提取的芳香挥发油具有辛辣凉爽的香气,广泛用于医药、牙膏、牙粉、糖、饮料和酒等产业     

Stimulation of menthol production in <Emphasis Type="Italic">Mentha piperita</Emphasis> cell culture

Plant cell culture provides an alternative means for producing secondary metabolites. In this study, experiments were carried out to study the impact of several parameters, independently and in combination, on the stimulation of menthol production in the cell suspension culture of Mentha piperita. Callus was obtained from leaf segments of in vitro grown plantlets on Murashige and Skoog (MS) medium supplemented with 0.2 mg l⁻¹of 2,4-dichlorophenoxy acetic acid to initiate cell suspension culture. This culture was maintained in half-strength MS medium supplemented with 0.2 mg l⁻¹of 2,4-dichlorophenoxy acetic acid at 15 d interval and used for further studies. Precursor feeding alone, i.e., menthone, at 35 μM concentration showed slightly improved productivity. γ-Cyclodextrin alone at 60 μM concentration and in combination with menthone feeding at 35 μM increased menthol yield up to 92 and 110 mg l⁻¹ in comparison to 77 mg l⁻¹ of control culture. Synergistic potentiation effect of menthone feeding at 35 μM and γ-cyclodextrin at 60 μM treatment followed by in situ adsorption with RP-8 also showed potential stimulation of menthol production in M. piperita cell culture. Fungal elicitor treatment showed enhanced production level up to 140.8 mg l⁻¹ in comparison to that of control. Further studies were carried out with the establishment of Agrobacterium tumefaciens (Ach5) gall-mediated calli, and consequently, cell suspension culture and results showed the significant enhancement of menthol yield up to 278 mg l⁻¹. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Biochemical activities of Iranian Mentha piperita L. and Myrtus communis L. essential oils

GC-MS analysis of essential oils of Iranian Mentha piperita and Myrtus communis extracted by hydrodistillation lead to identification of 26 and 32 compounds, respectively. The oils had good to excellent antimicrobial activities against Escherichia coli, Staphylococcus aureus and Candida albicans with the oil of M. piperita being more active. The findings suggest feasibility of application of M. piperita oil in treatment of the infections caused by C. albicans and E. coli. D-values on exposure to M. piperita and Myrtus communis oils were (2.14 and 2.8min), (1.4 and 12.8min) and (4.3 and 8.6min) for E. coli, S. aureus and C. albicans , respectively. The oils were screened for their possible antioxidant activities by two complementary test systems, namely DPPH free radical scavenging and beta-carotene/linoleic acid systems. M. piperirta oil exerted greater antioxidant activity than that of M. communis. Phytochemical and phytobiological characteristics of these oils may lead to extraction and production of active compounds in single or combined forms with useful applications.     

Glucosylation of esculetin by plant cell suspension cultures

Cell suspension cultures of Lithospermum erythrorhizon, Gardenia jasminoides and Nicotiana tabacum were capable of glucosylating esculetin to esculin (7-hydroxycoumarin-6-O--D-glucoside). Especially, a culture strain of Lithospermum erythrorhizon was superior in the esculetin glucosylating capability; 40 to 50% of esculetin administered to the culture medium at early stationary growth stage was converted into esculin within 24 h. The rate of glucosylation was also dependent on the growth stage and the medium composition especially growth hormones and sugar.     

Agronomical and chemical characterization of spearmint (Mentha spicata L.) originating in Turkey

The essential oil properties of spearmint (Mentha spicataL.), one of the most important spice plants, were studied and the essential oil components determined using gas chromatography. The essential oil content of wild-grown spearmint in the region was found to range from 1.00% to 2.00%, and two chemotypes were identified, one high in carvone (49.53-80.65%) and the other in pulegone (44.9-49.23%). Agronomic and essential oil properties of cultivated landraces ofM. spicata were also investigated under field conditions during the 1999 vegetation period. The examined spearmint landraces showed a great variability for each character studied, including yield and essential oil components. The crop was harvested twice during the vegetation period, and the essential oil content of the landraces varied from 0.90 to 2.70% in the first harvest and from 1.00 to 3.00% in the second one. Carvone was constantly present as the predominant essential oil in landraces, except for one sample, which was high in linalool (82.80%). Superior landraces with carvone contents were discovered; their maximum content reached 79.70% in the first cutting and 82.97% at the second cutting. The superior landraces were assayed for future improvement studies.     

Biotransformation of [ring-U-14C]4-n-nonylphenol by Agrostemma githago cell culture in a two-liquid-phase system

The biotransformation of [¹⁴C]4-n-nonylphenol (5 mg l^–1; 10 mg l^–1) by Agrostemma githago cell suspensions was studied using a batch two-liquid-phase system (medium/n-hexadecane 200:1, v/v). The highly lipophilic 4-n-nonylphenol was applied via n-hexadecane phase. After 7 d of incubation, more than 85% of applied 4-n-nonylphenol was absorbed by the cells, and 40% was transformed to 10 side-chain monohydroxylated metabolites (two with additional double bond at side-chain). The primary metabolites were analyzed by GC-EIMS. In the cells, the monohydroxylated products and residual 4-n-nonylphenol were present as glycosides. The method proved to be suitable for the production of primary metabolites of 4-n-nonylphenol on a larger scale for identification purposes and for metabolic profiling of the compound.     

Biotransformation of C13-norisoprenoids and monoterpenes by a cell suspension culture of cv. Gamay (Vitis vinifera)

A cell suspension culture of cv. Gamay was studied for its ability to metabolize two different C₁₃-norisoprenoidic volatiles, β-ionone and dehydrovomifoliol, together with monoterpenes, geraniol and linalool, biogenetically common pathways sharing compounds. β-Ionone was totally metabolized leading to fourteen norisoprenoidic volatiles oxygenated mainly at carbons 3 or 4 of the cyclohexane ring or reduced at side chain. The biotransformation of dehydrovomifoliol was at a lesser extent, giving rise to oxygenated and reduced derivatives. The norisoprenoidic metabolites were present both under free and glycosylated forms. Geraniol and linalool were also metabolized, leading to several free and glycosylated compounds. S. Mathieu, J. Wirth contributed equally to the work and should be considered joint first authors. A short part of this paper was published at the proceedings of the 10th Weurman Flavour Research Symposium, Flavour Research at the Dawn of the Twenty-first Century, J.-L.Le Quere, P.-X.Etievant, Editors; Lavoisier,2003/Intercept Ltd, 2003.     

Increased production of digoxin by digitoxin biotransformation using cyclodextrin polymer inDigitalis lanata cell cultures

Addition of β-cyclodextrin (β-CD) polymer during the biotransformation of digitoxin into digoxin using cell suspension cultures ofDigitalis lanata enhanced the conversion yield. Digitoxin showed better adsorption to CD polymer compared to digoxin, so that the optimization of addition time was found to be necessary. In the case of adding CD polymer 24 hours after the feeding of substrate digitoxin, the highest digoxin production could be achieved. At this period, digitoxin was almost consumed by cells and productivity was proportionally enhanced according as the amount of substrate was increased. Immobilization of CD polymer did not promote the biotransformation. When 3.33 g/L of CD polymer was added, 90% and 50% of digitoxin and digoxin was adsorbed respectively. Thus selective inclusion complex formation could be expected. Adsorption rate was found to be rapid and saturation was obtained within 10 hours of contact.     

椒样薄荷、薄荷和苏格兰留兰香精油与抗生素的协同抑菌功能

以开花期的椒样薄荷(Mentha × piperita)、薄荷(M. haplocalyx)和苏格兰留兰香(M. × gentilis)叶片部位提取的精油为研究对象, 通过GC-MS分析, 并采用纸片扩散法研究了3种精油单独使用及与抗生素联合使用时对金黄色葡萄球菌、蜡状芽孢杆菌、大肠杆菌、绿脓杆菌和肺炎克雷伯氏菌的抑制情况。结果表明, (1) 椒样薄荷与薄荷精油中含量最高的成分为薄荷醇、薄荷酮和异薄荷酮, 苏格兰留兰香精油的主要成分为香芹酮和柠檬烯。薄荷和苏格兰留兰香精油符合欧洲药典与ISO标准, 椒样薄荷需要继续改良以提高其精油品质与抑菌功能。(2) 精油单独使用时, Pseudomonas aeruginosa ATCC 15442对椒样薄荷精油和薄荷精油敏感; P. aeruginosa ATCC 27853对薄荷精油和苏格兰留兰香精油敏感。精油与抗生素联合使用时抑菌范围和强度均有所改变: 绿脓杆菌的2个菌株对精油与抗生素的组合最为敏感, 其中, 椒样薄荷精油与头孢他啶的组合对P. aeruginosa ATCC 15442显示出最强的增效作用, 薄荷精油与头孢他啶混合之后对P. aeruginosa ATCC 27853出现拮抗作用。Staphylococcus aureus ATCC 25923对所有精油以及精油与抗生素混合物均有抗性。(3) 椒样薄荷、薄荷和苏格兰留兰香精油的不同成分及其含量差异不仅对精油品质有影响, 而且影响精油对测试菌种的抑制作用, 可考虑将其作为薄荷属植物品质育种的参考指标。     

Improvement of phenylethanoid glycosides production by a fungal elicitor in cell suspension culture of Cistanche deserticola

When, on the 15th day of growth, an elicitor from Fusarium solani was added at 40 mg l^–1 to Cistanche deserticola cell suspension cultures, the contents of echinacoside, acteoside and total phenylethanoid glycosides (PeGs) in cultured cells all increased over the next 27 d by over 100% to 15 mg g^–1 dry wt, 9 mg g^–1 dry wt and 57 mg g^–1 dry wt, respectively. The final biomass (1.3 mg dry wt ml^–1) was not affected.     

Evaluation of acute and chronic toxicity of selected compounds in higher plants using cell culture

Summary The feasibility of using plant cell culture to measure toxicity was determined by investigating the toxicological effects of three chemical compounds, allyl alcohol, propargylglycine, and cadmium chloride, on cell cultures ofCatharanthus roseus G. Don (Madagascar periwinkle). Suspension cultures ofC. roseus were maintained in modified B5 medium and transferred every 5 d. Five-day-old cell cultures were exposed to various concentrations (10,3,1,0.3,0.1,0.03,0.01,0.003,0.001,0.0003,0.0001, 0.00003, and 0.0 mM) of the toxicants in both acute and chronic toxicity tests. In the acute test, cells were exposed to the toxicant for 24 h, washed three times with sterile medium, and plated in petri plates with an equal volume of 1.4% agar medium. Cells in the chronic test were plated with an equal volume of 1.4% agar medium containing various concentrations of the toxicant. Cells were incubated 28 d at 30°C in the dark. The colonies were counted and the results plotted as percent survival versus toxicant concentration. The results indicate, at the concentrations tested, thatC. roseus assay may be feasible in that it fulfills the criteria for a practical assay (e.g., rapid, simple, quantifiable, and reproducible). This work was submitted to the faculty of Miami University in partial fulfillment of the requirements for the degree of Master of Environmental Science, Institute of Environmental Sciences.     

Release of rosmarinic acid by Lavandula vera MM cell suspension in two-phase culture systems

Studies were conducted on the cultivation of Lavandula vera MM cell suspensions in different culture systems for the release of extracellular rosmarinic acid (RA). It was established that during cultivation with Amberlite XAD-4 as a second phase, 6.4% of the total content of RA was adsorbed. When L. vera MM cell suspension was cultivated in an aqueous two-phase system formed by adding 4% polyethylene glycol (MW 20,000) and 7.5% dextran (MW 70,000), 11.8% of the total RA content was released into the top polyethylene glycol phase.     

留兰香组织培养及快速繁殖条件的优化

以留兰香(Menthaspicata L.)茎尖为实验材料,对外植体消毒、不定芽增殖和试管苗移栽生根的最佳条件进行研究。结果表明,最佳外植体消毒方法为:用体积分数75%乙醇浸泡30s,再用1.0g·L-1HgCl2浸泡10min,培养7d后外植体生长状况良好。正交实验结果表明,在附加0.2mg·L-16-BA和0.02mg·L-1NAA的MS培养基中,留兰香不定芽的增殖倍数最高,试管苗生长状况最好。在含25mg·L-16-BA和50mg·L-1NAA的混合溶液中浸泡1h,移栽试管苗的生根率可达100%,且根较长。     

A preliminary investigation of ursolic acid in cell suspension culture of Salvia officinalis

Callus and suspension cultures of two genotypes and two morphological forms (friable and compact) were established on MS medium supplemented with 10.47 μM NAA and 4.5 μM BA. Biomass increase in 14-day-culture was calculated and ursolic acid (UA) content was determined by HPLC and MS. The growth rate and UA accumulation was found to be significant in the two genotypes. The compact biomass of both genotypes demonstrated a much slower growth rate and a lower UA accumulation than the friable biomasses. The accumulation of UA in suspension culture was constant in time when derived from the friable callus but it declined, when derived from the compact callus. This revised version was published online in June 2006 with corrections to the Cover Date.     

Morphology and monoterpene biosynthetic capabilities of secretory cell clusters isolated from glandular trichomes of peppermint (Mentha piperita L.)

Secretory cells were isolated from the monoterpene-producing glandular trichomes (peltate form) of peppermint as clusters of eight cells each. These isolated structures were shown to be non-specifically permeable to low-molecular-weight, water-soluble cofactors and substrates. Short incubation periods with the polar dye Lucifer yellow iodoacetamide (M_r=660) resulted in a uniform staining of the cytoplasm, with exclusion of the dye from the vacuole. The molecular-weight exclusion limit for this permeability was shown to be less than approx. 1800, based on exclusion of fluorescein-conjugated dextran (M_r 1800). Intact secretory cell clusters very efficiently incorporated [³H]geranyl pyrophosphate into monoterpenes. The addition of exogenous cofactors and redox substrates affected the distribution of monoterpenes synthesized from [³H]geranyl pyrophosphate, demonstrating that the cell clusters were permeable to these compounds and that the levels of endogenous cofactors and redox substrates were depleted in the isolated cells. When provided with the appropriate cofactors, such as NADPH, NAD⁺, ATP, ADP and coenzyme A, the isolated secretory cell clusters incorporated [¹⁴C]sucrose into monoterpenes, indicating that these structures are capable of the de-novo biosynthesis of monoterpenes from a primary carbon source, and that they maintain a high degree of metabolic competence in spite of their permeable nature.Abbreviations GLC gas liquid chromatography - LSCM laser scanning confocal microscopy - LY-IA Lucifer yellow iodoacetamide This investigation was supported in part by U.S. Department of Energy Grant DE-FG0688ER13869 and by Project 0268 from the Washington State University Agricultural Research Center. Light microscopy was carried out in the Plant Biology Light Microscopy and Image Analysis Facility (WSU) funded by the National Science Foundation (DIR9016138). We thank Greg Wichelns for growing the plants and Stephen Pfeiffer (BioRad Microsciences Division, Cambridge, Mass, USA), for help acquiring the confocal images.     

Production of 8-epi-tomentosin by plant cell culture ofXanthium strumarium

This study was conducted to establish a plant cell culture system for the production of medically important secondary metabolites fromXanthium strumarium. The effects of plant growth regulators including NAA, 2,4-D, kinetin, and ABA were examined in terms of callus induction, maintenance of callus and suspension cultures. It was shown that callus was induced upon treatment with NAA while embryo was induced after treatment with 2,4-D. Callus formation was further improved by treatment with ABA and NAA. The level of callusing increased by 17–29% for the seed case, cotyledon, leaf, and hypocotyl and by 96% in the case of the root. Suspension cell lines were established using calli produced from cotyledon, hypocotyl and root and cultured at 25°C under light conditions. The cells grew up to 15 g/L with NAA 2 ppm, BA 2 ppm, and ABA 1 ppm treatment. Supernatants of suspension cultures of cell lines derived from coyledon and hypocotyl produced some distinctive secondary metabolites, one of which was identified as 8-epi-tomentosin, which belongs to the xanthanolides. The amounts of 8-epi-tomentosin produced by the cotyledon-and hypocotylderived cell lines were 13.4 mg/L and 11.0 mg/L, respectively.