Cell and Tissue Journal

Abstract Aim: Oregano (Mentha longifolia) is an important medicinal plant from the mint family, which is widely used in food and pharmaceutical industries due to its active compounds. As an efficient approach, the use of biological elicitors (yeast, bacterial, and fungal agents) as well as non-biological elicitors (inactive enzymes, ultraviolet rays, heavy metal salts, polysaccharides, salicylic acid, jasmonic acid, and melatonin) can provide a way to increase the production of secondary metabolites in plant cell cultures. Given the importance of Oregano in the pharmaceutical and therapeutic industries, this study was carried out to assess melatonin's effect on the percentage of cell viability and the level of important bioactive compounds of oregano.  
Materials and methods: This study was performed in a factorial experiment with a completely randomized design and three replications in the lab. Melatonin (0, 100, and 200 μM based on the previous studies) was added to the MS culture medium containing 2,4-D (1 mg.L^-1). Then, 24, 48, and 72 h after treatment, the percentage of cell survival and the level of important oregano compounds were measured by tetrazolium and HPLC tests, respectively. In SAS software version 9.4, analysis of variance (ANOVA) was used to analyze the data and the LSD method to compare the means.
Results: Based on the results of the tetrazolium test, there was no significant difference in the percentage of cell viability in various concentrations of melatonin. However, the survival percentage of cells decreased with the passage of time, so that the lowest survival percentage was observed 72 h after melatonin treatment. Adding melatonin to the culture medium increased the production of secondary metabolites. At the concentration of 100 µM, the amount of menthol, menthone, pulegone, and 1,8-cineole increased by 77, 108, 46, and 80%, respectively, after 48 hours in contrast to the control. The increase in the level of menthol, menthone, pulegone, and 1,8-cineole compounds in the concentration of 200 µM of melatonin was respectively 125, 130, 140, and 120%, after 48 hours when compared to the control. Therefore, increasing melatonin concentration from 100 to 200 µM in cell culture significantly increased the level of compounds compared to the control. The levels of menthol, menthone, pulegone, and 1,8-cineole increased up to 48 hours after treatment, but after that time, they showed a significant decrease. Since the ultimate goal is to increase the production of biologically active compounds and not the highest cell viability, therefore, 48 hours after applying the treatment of 200 μM melatonin provides the best conditions for obtaining a high level of valuable oregano metabolites. The above treatment increased menthol, menthone, pulegone, and 1,8-cineole by 125, 130, 140, and 120%, respectively.
Conclusion: In this study, for the first time, the effect of melatonin elicitor on important secondary metabolites of the Oregano plant was investigated to determine the potential of using this metabolic stimulant in research and commercial areas. Overall, The use of melatonin at a concentration of 200 μM along with the extraction of metabolites 48 hours after the treatment can enhance menthol, menthone, pulegone, and 1,8-cineol, while maintaining the viability of oregano cells

Abstract Aim: Salinity stress is one of the most important environmental stress in the world and one of the important factors in reducing growth in many plants, especially in arid regions of the world. Salinity stress and increased sodium ion lead to the induction of oxidative stress and consequent cell death. Melatonin is a multiple function molecule spread in different plant and triggers several physiologic responses to different environmental stress. Exogenous application of melatonin to several plants can improve crop growth and development in response to many abiotic and biotic stresses with adjusting the antioxidant system of plants. Current studies reported that, the exogenous melatonin can increase plants' stress resistance by regulating both the enzymatic and non-enzymatic antioxidant defense process. In this study, the effect of melatonin on alfalfa roots under tissue culture condition was investigated. The aim of the present study was to investigate how melatonin regulated the antioxidant system and non-enzymatic antioxidants such as reduced glutathione and ascorbate under salt stress
Materials and methods: In this study, alfalfa seeds (Medicago sativa) of the Isfahani variety were used and we studied the effect of melatonin and salinity stress on the alfalfa root. In order to sterilize the seeds, they were placed in a 70% ethanol solution for one minute and then in a 20%  sodium hypochlorite solution for 20 minutes. After disinfecting the seeds, seeds were placed in each culture dish containing MS (Murashige and Skoog) culture medium. After germination, alfalfa seeds were transferred to MS culture medium containing concentrations of 0, 0.1, 10, and 15 micromolar melatonin and concentrations of 0, 150 and 200 mM salt. After 10 days of growth, total antioxidant capacity,
the activity level of catalase, ascorbate peroxidase, superoxide dismutase, guaiacol peroxidase, and glutathione reductase, ascorbate and glutathione levels in alfalfa roots were measured.
 
Results: Also, in the salinity stress, melatonin treatment significantly increased the total antioxidant power, while no significant difference was observed between different concentrations of melatonin, 0.1µM melatonin 82 درصد  and 62 درصد  raised antioxidant activity under 150 and 200 mM NaCl, so melatonin can reduce the levels of reactive oxygen species by scavenging of them through antioxidant enzyme or non- antioxidant system. Based on the results Melatonin treatment caused a significant increase in antioxidant power, the activity of CAT, APX, POD, SOD, GR enzymes, and antioxidant compounds in the glutathione-ascorbate cycle including DHA, ASC/DHA, GSH, and GSH/GSSG. along with increasing salinity concentration. On the other hand, salt stress increased oxidized compounds including DHA and GSSG in alfalfa roots. The data were carried out by two-way analysis of variance (ANOVA), followed by Duncan’s multiple range tests.




Conclusion: In addition to its direct role in clearing free radicals, melatonin activates the antioxidant defense system of the root, i.e. both antioxidant enzymes and antioxidant compounds in the ascorbate-glutathione cycle, which increases the resistance to damage oxidative effects caused by salinity stress in alfalfa root. These findings proposed that exogenous melatonin utilization dramatically activated ROS scavenging systems including enzymatic and non-enzymatic antioxidants to keep a relatively low amount of ROS and increased the tolerance of alfalfa root against salinity stress.

Abstract Aim: The aim of this study was to determine the effect of stress oxidative (H2O2) and different concentrations of melatonin on nuclear maturation of immature oocytes in sheep.
Material and Methods: Ovary collection and oocyte recovery was carried out by standard method. Oocytes culture was in the following conditions: A: TCM199+10% FBS, 5µg/ml FSH, 0.01IU/ml LH, 100 IU/ml penicillin and 100 IU/ml streptomycin, B: A + H2O2 (300 µM) at 38/5 C^˚, C: B + 1 µM melatonin and D: B + 10µM melatonin.
Results: This study showed that H2O2 significantly (p < 0.05) decreases nuclear maturation in compare to control (14.7 vs. 84. 9). 0, 1 and 10 µM melatonin could improve oocytes to reach to metaphase-II stage (respectively 14.7 vs. 43.29, 54.12). But increasing melatonin dose from 1 to 10 µM, did not have any significant effect on oocytes maturation.
Conclusion: The results of this study showed that melatonin improves sheep oocytes maturation during oxidative stress.