V. Payamnoor; J. Nazari; R. Jafari Hajati
Abstract
Betulin and betulinic acid are from the most important anticancer and anti-HIV metabolites, and the birch species (Betula spp.) bark is considered as the primary source of these metabolites. Due to the extinction of these tree species in Iran, it is necessary to replace the metabolites extraction from ...
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Betulin and betulinic acid are from the most important anticancer and anti-HIV metabolites, and the birch species (Betula spp.) bark is considered as the primary source of these metabolites. Due to the extinction of these tree species in Iran, it is necessary to replace the metabolites extraction from the birch bark with modern methods such as cell and tissue culture to produce the metabolites. The aim of this study was to determine the effect of explant type on the amount of betulin and betulinic acid produced in calli of two birch species B. pendula and B. litwinowii under in vitro conditions compared to the amount of metabolites extracted from the tree bark. Bark and leaf explants of two mentioned species were cultured in WPM medium containing 1 mg/L BAP and 0.1 mg/L 2,4-D for callogenesis. The amount of betulin and betulinic acid in three-month calli was measured using the HPLC technique and compared with the amount of these metabolites in one-centimeter stem bark samples taken from nature. The bark explant was more successful in callogenesis, and calli derived from this explant had more active ingredients. The amount of betulin and betulinic acid from the extract of bark sample taken from nature was respectively obtained to be 5.23 and 2.91 percent for B. pendula, and 5.65 and 2.52 percent for B. litwinowii. Moreover, calli derived from the bark explant of B. pendula and B. litwinowii contained 0.023 and 0.016 percent of betulin and 0.053 and 0.057 percent of betulinic acid, respectively. Generally, the results indicated that the bark explant was more capable of callogenesis and secondary metabolite induction than the leaf explant in both birch species under in vitro conditions.
V. Payamnoor; R. Jafari Hajati
Abstract
Birch (Betula pendula Roth.) is one of the most important species exposed to extinction due to the limited natural habitat and lack of regeneration. Plant tissue culture technique is an appropriate solution for asexual reproduction of this species and to protect it in controlled conditions. In ...
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Birch (Betula pendula Roth.) is one of the most important species exposed to extinction due to the limited natural habitat and lack of regeneration. Plant tissue culture technique is an appropriate solution for asexual reproduction of this species and to protect it in controlled conditions. In this research, callogenesis of B. pendula was investigated using five types of explants (leave, stem, nodal, petiole and bark) in WPM and NT media enriched with two hormonal combinations including A) 1mg/l BAP, 0.1 mg/l 2,4-D and B) 0.1 mg/l BAP, 0.01mg/l TDZ). The results indicated that the minimum callogenesis was observed in nodal and petiole explants in NT medium enriched with A hormonal combination. Callogenesis, fresh and dry weight of bark, stem and leave explants were significantly decreased in WPM and NT media containing B hormonal combination as compared with A hormonal combination. The maximum callogenesis was observed in bark explant in WPM (81.14) and NT (70.36) media with A hormonal combination as compared with other explants. The average fresh and dry weight of this explant in NT medium was more than that of WPM medium with the same hormonal combination.
R. Jafari Hajati; V. Payamnoor; K. Ghasemi Bezdi; N. Ahmadian Chashmi
Abstract
This study aimed to enhance the production of betulin and betulinic acid using suspension cultures of birch (Betula pendula Roth) and elicitation of the cell cultures by methyl jasmonate (MeJA) and salicylic acid (SA). To do this, at the first step, the cell growth curve was investigated in a 16-day ...
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This study aimed to enhance the production of betulin and betulinic acid using suspension cultures of birch (Betula pendula Roth) and elicitation of the cell cultures by methyl jasmonate (MeJA) and salicylic acid (SA). To do this, at the first step, the cell growth curve was investigated in a 16-day period. Then, two elicitors, namely, MeJA (at final concentration of 0, 50, 100, 150 and 200 µM) and SA (at final concentration of 0, 100, 200, 300 and 400 µM) were separately supplemented to 8-day-old cell cultures and the cells were harvested 1, 2, 3, 5 and 7 days after elicitations. Fresh weight (FW), dry weight (DW) and cell viability were measured. In addition, betulin and betulinic acid content were analyzed using HPLC. The results showed the significant effects of different concentrations of SA and MeJA on metabolites content and FW and DW. Maximum amount of betulin was observed about 4-fold (2.5 mg g-1 DW) higher than the control treatment by addition of 100 µM SA, two days after elicitation. Moreover, betulinic acid content was enhanced about 5 mg g-1 DW, 4.5-fold compared to control, one day after addition of 200 µM SA. Furthermore, the high accumulation of betulin (2.3 mg g-1 DW) was obtained in the elicited cell by 50 µM MeJA, seven days after elicitation. Also, the maximum amount of betulinic acid, about 3 mg g-1 DW, was observed in the cells elicited by 100 µM MeJA, two days after elicitation. Overall, the effect of SA on the production of betulin and betulinic acid was significantly more than the effect of MeJA.
