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افزایش تولید ترکیبات فلاونوئیدی و تغییر فعالیت آنزیم‌های آنتی‌اکسیدانتی در کشت کالوس گیاه گزنه تحریک شده با کیتوزان

نوع مقاله : علمی - پژوهشی

نویسندگان

فاطمه افشاری‌پور 1
محسن محمودنیا میمند 2
مریم دهجی‌پور حیدرآبادی 2
محمدرضا دهقانی 2

1 کارشناسی ارشد، گروه ژنتیک و تولید گیاهی، دانشکده کشاورزی، دانشگاه ولی‌عصر (عج) رفسنجان، ایران

2 دکتری تخصصی، گروه ژنتیک و تولید گیاهی، دانشکده کشاورزی، دانشگاه ولی‌عصر (عج) رفسنجان، ایران

https://doi.org/10.61186/JCT.15.4.317
چکیده
هدف: در این تحقیق به بررسی اثر غلظت‌ها‌‌‌‌‌‌ی مختلف الیسیتور کیتوزان و زمان در معرض الیستور بودن بر میزان تولید متابولیت‌‌‌‌‌‌های ثانویه و فعالیت آنزیم‌‌‌‌‌‌های آنتی­اکسیدانت در کشت کالوس گیاه گزنه پرداخته شد.
مواد و روش­ها: از ریزنمونه برگ گیاهچه استریل در محیط کشت MS هورمون‌‌‌‌‌‌دار جهت تولید کالوس استفاده شد. تیمار کالوس‌ها در محیط کشت مایع MS و با استفاده از کیتوزان (صفر، 50 و 100 میلی‌‌‌‌‌‌گرم بر لیتر) انجام شد و نمونه‌برداری در زمان‌های 24، 48 و 120 ساعت صورت گرفت. در نهایت میزان تولید کوئرستین، کامفرول، روتین، فنل، فلاونوئید و پروتئین کل و همچنین فعالیت آنزیم‌‌‌‌‌‌های ‌فنیل‌آلانین آمونیالیاز، پلی‌فنل اکسیداز و پراکسیداز اندازه­گیری شد. آزمایش در قالب طرح کاملا تصادفی با سه تکرار انجام شد و به‫صورت خرد شده در زمان تحلیل شد. نتایج: فعالیت آنزیم ‌فنیل‌آلانین آمونیالیاز و پلی‌فنل اکسیداز در هر سه زمان نمونه­برداری با افزایش غلظت کیتوزان نسبت به شاهد افزایش یافت. فعالیت آنزیم پراکسیداز در زمان نمونه‫برداری 120 ساعت و غلظت 50 میلی‫گرم بر لیتر کیتوزان، 57/2 برابر نمونه شاهد بود. بیشترین میزان تولید کوئرستین در تیمار 100 میلی‌‌‌‌‌‌گرم بر لیتر کیتوزان و زمان 48 ساعت و بیشترین میزان تولید کامفرول و روتین به‫ترتیب در زمان‌های 48 و 24 ساعت و غلظت 50 میلی‌‌‌‌‌‌گرم بر لیتر کیتوزان بود. نتیجه­گیری: الیسیتور کیتوزان در غلظت­ها و زمان خاصی باعث افزایش تولید متابولیت­های ثانویه و فعالیت آنزیم­های آنتی­اکسیدانتی در کشت کالوس گزنه ­شد. بنابراین در تحقیقات آتی می­توان با استفاده از این الیسیتور و بهینه­سازی شرایط تولید در مقیاس بالا به سمت تولید تجاری این مواد حرکت کرد.

کلیدواژه‌ها

الیسیتور
پلی‌فنل اکسیداز
‌فنیل‌آلانین ‌آمونیالیاز
کامفرول
گیاهان دارویی

عنوان مقاله English

Increasing the content of flavonoid compounds and changing the antioxidant enzyme activities in chitosan-elicited callus culture of nettle plant

نویسندگان English

F Afsharipour 1
M Mahmoodnia Meimand 2
M Dahajipour Heidarabadi 2
MR Dehghani 2
1 M.Sc. in Agricultural Biotechnology, Faculty of Agriculture, Vali-e-Asr University of Rafsanjan. Iran
2 Assistant Professor, Department of Genetics and Plant Production, Faculty of Agriculture, Vali-e-Asr University of Rafsanjan. Rafsanjan, Iran
چکیده English

Aim: The stinging nettle (Urtica dioica L.) is a very important medicinal plant that is used in the treatment of many diseases. Considering the important medicinal properties (flavonoid compounds and antioxidant enzymes) of nettle and the numerous advantages of using elicitors in plant tissue culture to enhance the production of medicinal compounds, this study investigated the effect of different concentrations of the chitosan elicitor on the production of some secondary metabolites and antioxidant enzyme activities in callus suspension culture of nettle.
Material and methods: Nettle seeds were cultured in 1/2MS medium after surface sterilization (with 1% sodium hypochlorite and 70% ethanol). leaf explants were isolated from sterile seedling (at the four-leaf stage). The explants were cultured in MS hormone-containing medium (NAA and BA 2.5 and 1 mg L-1, respectively) and maintained under light conditions of 16 : 8 (light : darkness) hours  at 25 °C. After several stages of subculturing, uniform callus was produced. Eight days after transfer of calli to MS liquid culture medium, calli were treated with chitosan elicitor at concentrations of 0, 50, and 100 mg L-1. Calli were sampled at 24, 48, and 120 hours after treatment. Finally, the production of secondary metabolites (quercetin, kaempferol, and rutin) was determined using HPLC and the standard line equation. Also, the activities of antioxidant enzymes (phenylalanine ammonia-lyase, polyphenol oxidase, and peroxidase) and the total content of phenol, flavonoid and protein were measured and analyzed by spectrophotometry method. The experiment was conducted in completely randomized design with three replications and was analyzed as a split-time statistical design.
Results: According to the results of the analysis of variance, the effect of chitosan elicitor concentration, sampling time after elicitor application and the interaction effect of elicitor concentration and sampling time on all measured parameters except for the total flavonoid content (the effect of elicitor concentration was not significant) were significant. The highest phenylalanine ammonia-lyase and polyphenol oxidase enzyme activities were at sampling times of 120 and 24 hours after elicitor application at 100 mg L-1 chitosan concentration, respectively. The peroxidase enzyme activity at the sampling time of 120 hours and a concentration of 50 mg L-1 of chitosan was 2.57 times that of the control sample. The total protein content decreased at concentrations of 50 and 100 mg L-1 chitosan compared with the control. The highest amount of quercetin production was in the treatment of 100 mg L-1 chitosan and sampling time of 48 hours after elicitor application, which increased 25 times compared with the control. Kaempferol and rutin had the highest production at sampling times of 48 and 24 hours and a concentration of 50 mg L-1, respectively, which increased by 72.6 and 51.2 percent compared with the control.
Conclusion: Chitosan elicitor is known as a biotic elicitor with a positive effect on medicinal and antioxidant properties in various plants. In this study, chitosan elicitor increased the production of quercetin, kaempferol, and rutin metabolites as well as the activity of antioxidant enzymes in nettle callus culture. So, chitosan elicitor can be introduced as an enhancer of medicinal properties of nettle plant under in vitro culture conditions. Obviously, for the commercial production of medicinal compounds, the conditions for large-scale callus production in bioreactors must be optimized.

کلیدواژه‌ها English

Elicitor
Kaempferol
Medicinal plants
Phenylalanine -ammonialyase
Polyphenol oxidase

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سلول و بافت
دوره 15، شماره 4
زمستان 1403
صفحه 317-335

  • تاریخ دریافت 17 بهمن 1403

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