نوع مقاله : مقاله پژوهشی
نویسندگان
1 گروه اصلاح نباتات و بیوتکنولوژی. دانشکده کشاورزی. دانشگاه زابل. زابل. ایران
2 گروه اصلاح نباتات و بیوتکنولوژی، دانشکده کشاورزی، دانشگاه زابل، زابل، ایران
3 پژوهشکده زیست فناوری کشاورزی، دانشگاه زابل، زابل، ایران
چکیده
کلیدواژه‌ها
موضوعات
عنوان مقاله [English]
نویسندگان [English]
Considering the low amount of important bioactive compounds such as glycyrrhizin and glycyrrhetinic acid in licorice and their complex biosynthesis, in vitro production of hairy roots is an efficient method for licoric secondary metabolites production. Therefore, this experiment aimed to investigate the efficiency of licorice hairy roots in the presence of Agrobacterium rhizogenes and the effect of methyl jasmonate (MeJA) at 100 µm, as an elicitor, on inducing glycyrrhizin and glycyrrhetinic acid production.. After inoculation, two kinds of the cefotaxime were used to eliminate the bacterium. The hairy roots were then harvested after one, two, three, and five days of elicitation. Overall, The results showed that the amount of glycyrrhizin, glycyrrhetinic acid and total flavonoids decreased in all treatments compared to the control, and the magnitude of this reduction in hairy roots treated with one inoculation of cefotaxime sodium salt and no MeJA was the highest in terms of glycyrrhizin (0.959 times) and glycyrrhetinic acid (0.925 times) production. In addition, the amount of total phenol (31.22 GAE/g D.W) and antioxidant activity (90.85%) was highest in in the hairy roots subjected to MeJA for two days and the control. MeJA had negative or no effect on production of glycyrrhizin and glycyrrhetinic acid, which can be related to type of species, ecotype, and antibiotics as well as the number of subcultures after inoculation, the concentration of the elicitor, and a period of time at which hairy roots exposed to a given elicitor. Based on our findings, cefotaxime is recommended to omit the bacteria from culture media. Overall, our findings could be used as an economically method to increase bioactive compounds, especially glycyrrhizin and glycerethnic acid, at a short time and low cost.
کلیدواژه‌ها [English]
Extended Abstract
Introduction
Licorice, belonging to Fabaceae, is well known for its medicinal properties. Glycyrrhizin and glycyrrhetinic acid are of the most important bioactive compounds mainly accumulated in Licorice roots and stolons. Glycyrrhizin possesses a wide array of pharmacological activities including anti-inflammatory, liver protective, gastric ulcer, antiallergic, and antiviral activities against various RNA or DNA viruses, such as human acquired immunodeficiency virus (HIV) and acute respiratory syndrome. Likewise, glycerhethnic acid has numerous medicinal properties, such as anti-inflammatory, antitumor, and so on. Plants generally contain low amount of above-mentioned compounds; therefore, using hairy roots and eliciting them by an appropriate substance has attracted the attentions in order to produce such compounds (as secondary metabolites) at in vitro condition. Many studies have concentrated on inducting hairy roots production in licorice. Hairy roots generally are able to produce glycyrrhizin. The greatest advantage of hairy roots, as compared to other parts of a plant, is contributed to their biosynthetic capacity for producing secondary metabolites. Recently, the beneficiary effects of methyl jasmonate (MeJA) on inducing secondary metabolites have been documented in different reports, because of its role in the biosynthesis of flavonolignans. This study aimed to investigate the effect of MeJA, as an elicitor, on in vitro production of glycyrrhizin and glycyrrhetinic acid in hairy roots of licorice.
Materials and Methods
Licorice seeds were initially collected from the Semiram region of Isfahan Province and soked in MS media for germination. Hypocotyls and cotyledons were used as explants for producing bioactive compounds. In this context, Agrobacterium rhizogenes A4 strain was used to induce hairy roots. The two types of antibiotics were also employed to remove bacteria including cefotaxime and cefotaxime sodium salt. After emerging, the 21-day-old roots were immersed in a liquid ½ MS medium for 35 days. In our research, MeJA (100 µM) was used as an elicitor. Hair roots were harvested at one, two, three, and five days after elicitation. In this experiment, numerous traits were measured including glycyrrhizin, glycyrrhetinic acid, total phenol, total flavonoid, and antioxidant activity. In order to analyze the data and draw graphs, SAS (ver. 9.2) and Excel softwares were used, respectively.
Results
The results of this research showed that the content of glycyrrhizin and glycyrrhetinic acid was higher in the control (bacteria eliminated using general cefotaxime and no MeJA). The lower amount of glycyrrhizin and glycyrrhetinic acid was produced using MeJA. The total flavonoid content was lower in all treatments than in the control, but the extent of this decrease was less than the decrease in glycyrrhizin and glycyrrhetinic acid. The roots that were exposed to MeJA for two days contained the highest amount of total phenol, followed by the roots of the control group, as indicated by the results of the mean comparisons. Hairy root in other treatments had the lowest amount of phenolic compounds. Furthermore, the activity of antioxidants declined in all the treatments, except for hairy roots exposed to MeJA for two days as well as control. These roots showed the highest percentage of free radical inhibition as 90.85 and 88.75, respectively, whereas the lowest was recorded in hairy roots treated with MeJA for five days. The percentage of free radical inhibition in hairy roots were derived in media containing sodium salt cefotaxime for bacteria elimination, and without elicitation, as well as those were subjected to Me JA for five day showed 78.9 and 77.34, respectively.
Conclusion
The results of this research showed that the lack or negative effects of MeJA on bioactive compounds, especially glycyrrhizin and glycyrrhetinic acid, are different based on type of species, ecotype, and antibiotic used to remove bacteria as well as the number of sub culture after inoculation, the concentration of given elicitor, and the duration of time subjecting to the elicitor(s). In a vast majority of previous studies, the rate of secondary metabolites produced by elicited hairy roots was lower or approximately close to the amount recorded in our work. Overall, our findings could be used as an economically method to increase bioactive compounds, especially glycyrrhizin and glycerethnic acid, at a short time and low cost.
Maryam Allahdou contributed to the conception of the study, doing laboratory work, data collection and analysis, interpretation of data, and drafting the manuscript. Leila Mehravaran assisted in the analysis and interpretation of results, and Hamideh Khajeh did laboratory work, data collection, and analysis.
All data supporting the findings of this study are available within the paper and the datasets during the current study are available from the corresponding author on reasonable request.
This article is taken from research project number PR-UOZ99-9. The authors of the article thank University of Zabol for the financial support of this research.
This article does not contain any studies involving human and animal subjects.
Conflict of interest
On behalf of all authors, the corresponding author states that there is no conflict of interest.
)