R. Ghodsi; B. Hosseini; A. Hedayati
Abstract
Hyoscyamus species such as H. reticulatus L. and H. pusillus L. are rich sources of tropane alkaloids, mainly hyoscyamine and scopolamine, which are used for their mydriatic, antispasmodic, anticholinergic, analgesic and sedative properties. In this study, in the first experiment, the effects of four ...
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Hyoscyamus species such as H. reticulatus L. and H. pusillus L. are rich sources of tropane alkaloids, mainly hyoscyamine and scopolamine, which are used for their mydriatic, antispasmodic, anticholinergic, analgesic and sedative properties. In this study, in the first experiment, the effects of four strains of Agrobacterium rhizogenes (ATCC 15834, A4, A7 and A13) and three explants (leaf, hypocotyls, and cotyledon) on hairy root induction were investigated in both H. reticulatus and H. pusillus species. In the second experiment, the effects of various culture media (MS, 1/2 MS, 1/4 MS, and B5) were evaluated on some morphological and phytochemical traits of hairy roots. The results showed that in H. reticulatus, maximum hairy root induction (78%) was obtained in the cotyledon explant inoculated with A7 strain and maximum hairy roots fresh weight (6.25 g) and dry weight (0.5 g) were recorded in MS medium. In this species, the highest total phenol (3.82 mg GAE g-1 FW) and total flavonoid (7 mg QUE g-1 FW) content were observed in MS medium; however, no significant changes were observed in the activity of antioxidant enzymes of hairy roots cultured in different media. In H. pusillus, the leaf explant inoculated with A13 strain showed maximum hairy root induction (90%), and the highest hairy root fresh weight (5.25 g) and dry weight (0.43 g) were observed in MS medium and the highest total phenol (2.26 mg GAE g-1 FW) and flavonoid (9.22 mg QUE g-1 FW) content were obtained in B5 medium. This study showed that basal culture medium type and plant species had a significant impact on the biomass and phytochemical characteristics of hairy roots.
F. Naseri; B. Hosseini; L. Naseri
Abstract
Rosmarinic acid (RA) is considered as one of the most important polyphenols. It is an ester of 3, 4-dihydroxyphenyl actic and caffeic acids and as an antioxidant significantly reduces the DNA damage induced by the anticancer compound doxorubicin. Hairy roots of Dracocephalum kotschyi Boiss contain rosmarinic ...
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Rosmarinic acid (RA) is considered as one of the most important polyphenols. It is an ester of 3, 4-dihydroxyphenyl actic and caffeic acids and as an antioxidant significantly reduces the DNA damage induced by the anticancer compound doxorubicin. Hairy roots of Dracocephalum kotschyi Boiss contain rosmarinic acid with biological and anticancer activities. However, due to the high production of phenolic compounds in liquid medium, the hairy roots encounter with tissue browning and death problem. In this study, the effect of antioxidant compounds on growth of D. kotschyi hairy roots and inhibition of tissue browning has been surveyed. In vitro Hairy roots were induced by inoculation of two-weeks-old cotyledons of D. kotschyi via Agrobacterium rhizogenes (15834 strain). Gene transformation confirmed by PCR using rol B gene specific primers. In order to optimize the growth and prevent the tissue browning, hairy roots were cultured in ½MS liquid medium containing different buffers and antioxidants including ascorbic acid (0.05 and 0.1 g/L), MES buffer (2.66 and 5.33 g/L), activated charcoal (1.5 and 3 g/L) and polyvinyl pyrrolidone (0.5 and 1.5 g/L) in two concentrations. In order to record the quality of hairy roots under the influence of different chemicals, the phytochemical properties of samples (total phenol and flavonoid, antioxidant activity and antioxidant enzymes activity) were analyzed by spectrophotometer. Results after 21 days showed that the maximum biomass (1.64 g) and the lowest tissue browning was obtained in ½MS media containing activated charcoal (1.5 g/L). And after AC the highest biomass was observed on both PVP concentrations (1.36 g) and the highest browning and growth loss was observed in the control samples. The maximum and minimum amount of total phenol and flavonoid were observed in control samples and 1.5 g/L of activated charcoal treatment, respectively. The highest and the lowest levels of total antioxidant activity and antioxidant enzymes were related to 1.5 g/L of activated charcoal and control samples, respectively. It is considerable that the inhibition of PAL enzyme by using antioxidants such as activated charcoal and PVP has a potential to reduce biosynthesis of phenolic compounds and prevent the browning.
S.H. Madani; B. Hosseini; Gh. Karimzadeh; A. Rahimi
Abstract
Iranian poppy (Papaver bracteatum Lindl.) is one of the medicinal plants belonging to the Papaveraceae family, which is widely used in pharmaceutical industries due to the presence of benzyl isoquinoline alkaloids. The polyploidy induction is one of the most interesting issues in the breeding and biotechnology ...
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Iranian poppy (Papaver bracteatum Lindl.) is one of the medicinal plants belonging to the Papaveraceae family, which is widely used in pharmaceutical industries due to the presence of benzyl isoquinoline alkaloids. The polyploidy induction is one of the most interesting issues in the breeding and biotechnology of medicinal plants. In this study, colchicine treatment was carried out in five concentrations (0, 0.05, 0.1, 0.2 and 0.5) and three duration times (24, 48, 72 hours) with three replications as a factorial in a completely randomized design under greenhouse conditions, which aimed at studying morphological and phytochemical changes in polyploid plants and comparing them with diploid ones. The Microscopic, morphological and chromosomal counts were used to determine ploidy level of plants. The results showed that the concentration of 0.2% of colchicine for 48 hours was the most suitable treatment for polyploidy induction. Polyploidy caused significant changes in the increasing phytochemicals amount such as phenol, flavonoids and total antioxidants (DPPH) and decreasing the stomatal density index in comparison with the diploid plants. The chromosomes number of the diploid and tetraploid plants was obtained 14 (2n=2x=14) and 28 (2n=4x=28), respectively. Therefore, it can be concluded that the increase in ploidy level increases the phytochemical and antioxidant compounds in Iranian poppy herb.
A. Norozi; B. Hosseini; M. Jafari; M. Farjaminezhad
Abstract
Henabne(Hyoscyamus reticulatus L.) is a rich source of tropane alkaloids including hyoscyamine (Hyos) and scopolamine (SCP), widely used in pharmaceutical field. The aim of the present study was to assess the effect of different concentrations (0, 50, 100 and 200 µM) of methyl jasmonate (MeJA) ...
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Henabne(Hyoscyamus reticulatus L.) is a rich source of tropane alkaloids including hyoscyamine (Hyos) and scopolamine (SCP), widely used in pharmaceutical field. The aim of the present study was to assess the effect of different concentrations (0, 50, 100 and 200 µM) of methyl jasmonate (MeJA) at two different times of exposure (24 and 48 h) on growth, antioxidant enzymes activity, tropane alkaloids content, and the expression level of hyoscyamine-6-beta-hydroxylase (h6h) gene in H. reticulatus hairy root cultures. The results showed that MeJA at high concentration (200 µM) led to the decline of fresh and dry weight of hairy roots. Catalase (CAT), guaiacol peroxidase (GPX) and ascorbate peroxidase (APX) activities were significantly increased by MeJA, especially at 200 µM in elicited hairy roots. As well, CAT and GPX activities, but not of APX, were also affected by exposure time of MeJA. According to the GC/MS analysis, the highest percentage of SCP (13.96%) and Hyos (21.9%) were respectively obtained when hairy roots were exposed to MeJA at concentrations of 100 and 200 µM for 48 and 24 h, which were, respectively, 1.6- and 1.25-fold higher than those in the control roots. According to the results of semi-quantitative RT-PCR analysis, the highest expression of h6h gene (6-fold higher than in the control) was obtained after 24 h exposure to 100 µM MeJA. These results indicate that MeJA, due to stimulation of the expression of key genes involved in the biosynthetic pathway of tropane alkaloids, could be used as an effective elicitor for increased production of Hyos and SCP.
A.R. Estaji; B. Hosseini; E. Dehghan; A. Estaji
Abstract
Salvia leriifolia Bent., belonging to Lamiaceae family, is a perennial herbaceous plant, which is endemic to Khorasan and Semnan provinces of Iran. Different pharmacological activities of this plant have been evaluated in recent years. Induction of polyploidy is an important technique for breeding of ...
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Salvia leriifolia Bent., belonging to Lamiaceae family, is a perennial herbaceous plant, which is endemic to Khorasan and Semnan provinces of Iran. Different pharmacological activities of this plant have been evaluated in recent years. Induction of polyploidy is an important technique for breeding of medicinal and aromatic plants. In the present study, we studied the effects of colchicine on morphological and physiological characteristics and chemical compositions of Salvia leriifolia. Polyploidy induction was carried out in the four-leaf stage. Apical meristem was treated by colchicine at concentrations of 0, 0.05, 0.1, 0.2, and 0.5%, and three different time levels of 24, 48 and 72 hours. Morphological, microscopic (light and electron), flow cytometric analysis and chromos counting were used for ploidy induction analysis and selection of induced tetraploids. Identification of essential oil compounds was carried out by gas chromatograph and gas chromatography mass spectrometry methods. According to the results, colchicines at a concentration of 0.05% for 48 hours at four-true-leave stage was the best treatment for induction of autotetraploidy (23.3%) in Salvia leriifolia. Tetraploid induction in Salvia leriifolia caused significant changes in morphological, cytological, physiological and physiochemical characteristics such as increased dry and fresh weight, chlorophyll content, photosynthesis, and decreased plant height. In addition, an increase in ploidy level caused significant changes in essential oil composition.
F. Moharrami; B. Hosseini; M. Farjaminezhad; A. Sharafi
Abstract
Hyoscyamus species such as Hyoscyamus reticulatus L. are rich sources of tropane alkaloids, mainly hyoscyamine and scopolamine, which are used for their mydriatic, antispasmodic, anticholinergic, analgesic and sedative properties. Due to complex chemical structures, these alkaloids are obtained from ...
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Hyoscyamus species such as Hyoscyamus reticulatus L. are rich sources of tropane alkaloids, mainly hyoscyamine and scopolamine, which are used for their mydriatic, antispasmodic, anticholinergic, analgesic and sedative properties. Due to complex chemical structures, these alkaloids are obtained from natural sources, mainly Solanaceae plants. Elicitation of secondary metabolites biosynthetic pathways by different kind of elicitors is an effective strategy to increase secondary metabolites productivity. In the present study, in order to increase production of tropane alkaloids, cotyledon-derived hairy root cultures transformed with Agrobacterium rhizogenes strain A7, were elicited with yeast extract (YE) as biotic elicitor. Effect of different concentrations (0, 125, 250, 500 and 1000mg/l) of YE elicitor at different exposure times (24, 48 and 72 h) were investigated. According to the results, YE at the concentration of 500 and 250 mg/l after 48 h treatment, significantly increased hyoscyamine (2-fold) and scopolamine (2.5-fold) production in comparison with control, respectively. The results showed that treatment with different concentrations of YE and also increasing the exposure time led to significant decrease in growth of hairy roots in comparison with control. The activity of antioxidant enzymes including catalase, guaiacol peroxidase, and ascorbate peroxidase was also elevated in treated hairy roots rather than of the control. Based on the results, it can be concluded that elicitation with YE leads to induce an oxidative stress. These results suggest that YE could be used as an effective elicitor in plant biotechnology for the production of plant secondary metabolites such as tropane alkaloids.
B. Hosseini; H. Hashemi Sohi; F. Shahriari; E. Dehghan
Abstract
Papaver somniferum L. today is considered as the commercial source of the narcotic analgesics morphine and codeine. Codeinone reductase is a key gene in metabolic engineering of isoquinoline alkaloids pathway with the ability of conversion of codeinone to codein and morphine. In this project, ...
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Papaver somniferum L. today is considered as the commercial source of the narcotic analgesics morphine and codeine. Codeinone reductase is a key gene in metabolic engineering of isoquinoline alkaloids pathway with the ability of conversion of codeinone to codein and morphine. In this project, at first optimization of the gene transfer of P. somniferum was performed via A. tumefascience containing pBI121 plasmid. This gene then was cloned in expression vectors under control of CaMV35 promoter and transferred to plants by agro transformation. After preparing the structure, hypocotyl explants of P. somniferum was inoculated by agrobacterium carrying recombinant structures. HPLC analysis indicated the variation of the amount, type and percentage of alkaloid compounds in transgenic samples compared to the control plants. The result of the evaluation showed qualitative and quantitative changes in metabolite production of transgenic and control plants.
