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ارزیابی تأثیر نانو کیتوزان و NPK بر محتوای کاتچین‌های برگ سبز چای (رقم کاشف) از طریق آنالیز برخی پارامترهای بیوشیمیایی، فیزیولوژیکی و مولکولی | ||
| تحقیقات گیاهان دارویی و معطر ایران | ||
| مقاله 6، دوره 40، شماره 1، فروردین 1403، صفحه 79-103 اصل مقاله (1.02 M) | ||
| نوع مقاله: مقاله پژوهشی | ||
| شناسه دیجیتال (DOI): 10.22092/ijmapr.2023.363015.3348 | ||
| نویسندگان | ||
| حسین عسکری* 1؛ امینه زینالی2؛ میترا پارسا3؛ منا کاشانچی4؛ رضا آزادی گنبد5؛ اقدس بنایی6؛ صنم صفائی‌چائی‌کار7؛ احسان کهنه8؛ علی سراجی9 | ||
| 1دانشیار گروه بیولوزی سلولی و مولکولی، دانشکده علوم زیستی و زیست‌فناوری، دانشگاه شهید بهشتی، تهران ، ایران | ||
| 2دانشجوی دکتری ومربی پژوهش گروه پژوهشی بیوتکنولوژی صنعت و محیط، پژوهشکده علوم پایه کاربردی جهاد دانشگاهی، دانشگاه شهید بهشتی، | ||
| 3مربی پژوهش گروه پژوهشی بیوتکنولوژی محیط و صنعت، پژوهشکده علوم پایه کاربردی جهاد دانشگاهی، دانشگاه شهید بهشتی، تهران | ||
| 4کارشناس آزمایشگاه، گروه پژوهشی بیوتکنولوژی صنعت و محیط، پژوهشکده علوم پایه کاربردی جهاد دانشگاهی، دانشگاه شهید بهشتی، تهران. | ||
| 5رضا ازادی گنبد. پژوهشکده چای,موسسه تحقیقات علوم باغبانی ، سازمان تحقیقات ، آموزش و ترویج کشاورزی ، لاهیجان ، ایران | ||
| 6استادیار گروه پژوهشی نانو و بیوفیزیک، پژوهشکده علوم پایه کاربردی جهاد دانشگاهی، دانشگاه شهید بهشتی، تهران، | ||
| 7عضو هیات علمی پژوهشکده چای، موسسه تحقیقات علوم باغبانی، سازمان تحقیقات، آموزش و ترویج کشاورزی، لاهیجان، ایران آدرس: گیلان، لاهیجان، | ||
| 8عضو هیات علمی پژوهشکده چای، موسسه تحقیقات علوم باغبانی، سازمان تحقیقات، آموزش و ترویج کشاورزی، لاهیجان، ایران | ||
| 9استادیار گروه فناوری و مدیریت تولید پژوهشکده چای؛ موسسه تحقیقات علوم باغبانی؛ سازمان تحقیقات، آموزش و ترویج کشاورزی؛ لاهیجان، | ||
| چکیده | ||
| سابقه و هدف: امروزه استفاده از محرک‌ها به‌منظور افزایش رشد گیاهان و تولید متابولیت‌های ثانویه مورد توجه هستند. رشد کند و کیفیت پایین برگ سبز چای (Camellia sinensis (L.) O. Kuntze) در مناطق کشت چای در ایران، از مهمترین چالش‌های تولیدکنندگان چای است. محلول‌پاشی برگی کیتوزان و نانو کیتوزان (NC) در بسیاری از گیاهان باعث القای رشد و افزایش متابولیت‌های ثانویه شده و بر شاخص‌های فیزیولوژیکی و بیوشیمیایی گیاهان اثر مثبت دارد. هدف از این پژوهش، بررسی تأثیر نانو کیتوزان بر برخی پارامترهای بیوشیمیایی و فیزیولوژیکی با و بدون تغذیه معدنی NPK و ارزیابی کیفیت برگ سبز چای از طریق مقایسه سطح بیان نسبی 3 آنزیم درگیر در مسیر فلاونوئیدها در گیاه چای رقم کاشف در شمال ایران بود. مواد و روش‌ها: بدین منظور از دو طرح استفاده شد: 4 غلظت متفاوت (0، 25، 50 و 100 میلی‌گرم بر لیتر) از محلول نانو کیتوزان به همراه تغذیه معدنی (NPK) و 4 غلظت متفاوت (0، 25، 50 و 100 میلی‌گرم بر لیتر) از محلول نانو کیتوزان بدون تغذیه معدنی تهیه شد. این دو آزمایش به‌صورت محلول‌پاشی پس از اولین چین بهاره دو بار با فاصله دو هفته‌ای در سال 1400 در مرکز تحقیقات چای لاهیجان (ایران) اجرا شد. 20 روز پس از محلول‌پاشی اولیه، نمونه‌برداری گیاهی انجام شد. نمونه‌های برگی از شاخساره جوان شامل جوانه و برگ اول و دوم برای آزمایش‌های متابولیکی و مولکولی و برگ سوم و چهارم برای آزمایش‌های فیزیولوژیکی و بیوشیمیایی برداشت شد. بررسی ویژگی‌های فیزیولوژیکی شامل محتوای کلروفیل و محتوای نسبی آب (RWC)، ویژگی‌های بیوشیمیایی شامل محتوای پروتئین و آنزیم‌های آنتی‌اکسیدانی کاتالاز (CAT)، سوپراکسید دیسموتاز (SOD) و پلی‌فنل اکسیداز (PPO) اندازه‌گیری شدند. همچنین، اندازه‌گیری ویژگی‌های متابولیکی شامل میزان ترکیب‌های فنلی تام از طریق روش فولین سیو کالتئو و تعیین میزان فلاونوئیدهای کاتچین، اپی‌گالوکاتچین (EGC) و گالوکاتچین (GC) با روش HPLC انجام شد. آنالیز مولکولی از طریق بررسی بیان نسبی 3 آنزیم کلیدی مسیر بیوسنتز فلاونوئیدها فلاونون 3 هیدروکسیلاز (F3H)، دی‌هیدروفلاونول 4 ردوکتاز (DFR) و لوکوآنتوسیانیدین ردوکتاز (LAR) انجام شد. نتایج: نتایج نشان داد مقدار پلی‌فنل کل در تیمارهای مورد بررسی افزایش معنی‌داری نسبت به شاهد (بدون NC و NPK) داشت. میزان کاتچین در تیمارهای مختلف، کاهش 4 تا 6 برابری داشت. محتوای اپی‌گالوکاتچین با افزایش غلظت نانو کیتوزان افزایشی بود. محتوای گالوکاتچین نیز تنها در غلظت mgL-1 100 نانو کیتوزان کمی روند افزایشی نشان داد. محتوای کلروفیل در تیمارهای با غلظت پایین نانو کیتوزان تفاوت معنی‌دار با روند کاهشی داشت؛ در حالی‌که در غلظت mgL-1 100 نانو کیتوزان تفاوت معنی‌دار با روند افزایشی مشاهده شد. البته، محتوای پروتئین در تیمارهای مختلف نانو کیتوزان تفاوت معنی‌داری نسبت به شاهد نشان نداد. فعالیت آنزیم SOD در تیمارهای مختلف نانو کیتوزان به استثنای تیمار mgL-1 50 NC+NPK تفاوت معنی‌دار آشکاری با روند افزایشی داشت. فعالیت آنزیم CAT نیز در تیمارهای مختلف با افزایش غلظت نانو کیتوزان افزایش یافت. فعالیت آنزیم PPO در تیمارهای با غلظت پایین نانو کیتوزان کاهش معنی‌داری نشان داد. محتوای نسبی آب برگ در تیمارهای مختلف، افزایش یافت. بیان نسبی آنزیم F3H، تنها در غلظت mgL-1 100 NC+NPK افزایش حدود 3 برابری نسبت به شاهد داشت و در سایر تیمارها بیان نسبی چندان معنی‌دار نبود. بیان نسبی DFR در تیمارهای مختلف NC+NPK افزایشی و بالاترین میزان بیان (4 برابر) در غلظت mgL-1 100 نانو کیتوزان مشاهده شد. بیان نسبی LAR در غلظت‌های mgL-1 0، 50 و 100 NC به همراه NPK نسبت به شاهد روند افزایشی داشت. بالاترین میزان بیان LAR در تیمار فاقد NC و همراه NPK با بیان حدود 5/2 برابری نسبت به شاهد مشاهده شد. نتیجه‌گیری: استفاده از نانوذرات کیتوزان در غلظت‌های مختلف به همراه NPK منجر به افزایش تولید ترکیب‌های کاتچین شد که علاوه‌بر مقابله با تنش اکسیداتیو در کیفیت برگ سبز چای نقش مؤثر دارد. نانو کیتوزان می‌تواند جایگزین مناسبی برای مواد شیمیایی برای القای رشد و افزایش کیفیت برگ سبز در گیاه چای باشد و با توجه به زیست تخریب‌پذیر بودن آن میزان آلودگی محیط‌زیستی را کاهش دهد. | ||
| کلیدواژه‌ها | ||
| گیاه چای؛ Camellia sinensis؛ تغذیه معدنی؛ نانو کیتوزان؛ فلاونوئید؛ دی‌هیدروفلاونول 4 ردوکتاز؛ فلاونون 3 هیدروکسیلاز؛ لوکوآنتوسیانیدین ردوکتاز | ||
| عنوان مقاله [English] | ||
| Evaluation of the effect of foliar application of nano-chitosan and mineral nutrition (NPK) on the catechins content in green tea (Kashef var.) leaves through analysis of some biochemical, physiological and molecular parameters | ||
| نویسندگان [English] | ||
| Hossein Askari1؛ Amineh Zeinali2؛ Mitra Parsa3؛ mona kashanchi4؛ Reza Azadi Gonbad5؛ Aghdas Banaei6؛ Sanam Safaei-Chaeikar7؛ Ehsan Kahneh8؛ Ali Seraji9 | ||
| 1Associate Professor, Department of Cellular and Molecular Biology, Faculty of Life Sciences and Biotechnology, Shahid Beheshti University, Tehran, Iran. | ||
| 2Industrial and Environmental Biotechnology Department, Research Institute of Applied Science, ACECR, Shahid Beheshti University, Tehran, Iran, | ||
| 3Industrial and Environmental Biotechnology Department, Research Institute of Applied Science, ACECR, Shahid Beheshti University, Tehran, Iran, Po box: 198396411 | ||
| 43. Industrial and Environmental Biotechnology Department, Research Institute of Applied Science, ACECR, Shahid Beheshti University, Tehran, Iran, Po box: 198396411 | ||
| 5Assist. Professor, Tea Research Center, Horticultural Sciences Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Lahijan, Iran | ||
| 6Department of Nano and Biophysics, Research Institute of Applied Sciences , Academic Center of Education, Culture and Research (ACECR), Tehran, Iran | ||
| 7Tea Research Center, Horticultural Science Research Institute, AREEO, Lahijan, Iran. | ||
| 8Tea Research Center, Horticultural Science Research Institute, AREEO, Lahijan, Iran. | ||
| 9Assistant Professor of Tea Research Center, Horticultural Science Research Institute, AREEO, Lahijan, Guilan, Iran. | ||
| چکیده [English] | ||
| Background and objectives: Elicitors are used more frequently to promote plant growth and secondary metabolites. One of the main challenges for tea producers is the slow growth and poor quality of green tea leaves (Camellia sinensis) in tea-growing regions of Iran. In many plants, foliar application of chitosan or nano-chitosan (NC) enhances secondary metabolite production. It has a positive impact on plants' physiological and biochemical indicators. This study investigated the impact of nano-chitosan on some biochemical and physiological parameters with and without mineral nutrition (NPK), as well as assessing the quality of green leaves by comparing the relative expression levels of three enzymes involved in the flavonoids pathway in the Kashef cv. tea plant in northern Iran. Methodology: For this, two scenarios were used: four different concentrations of nano chitosan solution (0, 25, 50, and 100 mgL-1) were prepared and combined with NPK (1%-1%-0.5%), and without NPK were prepared. At the Lahijan Tea Research Center in Iran, two experiments were conducted as foliar treatments applied twice at two-week intervals, after the first season's harvest in 2021. Twenty days following the initial foliar treatment, plant sampling was conducted to examine physiological, biochemical, metabolic, and molecular characteristics. The first and second leaves, as well as the buds, were gathered for metabolic and molecular testing. The third and fourth leaves were collected for physiological and biochemical analyses. Chlorophyll content and relative water content were measured in physiological experiments. Protein content and the antioxidant enzymes CAT, SOD, and PPO were investigated biochemically. Additionally, metabolic properties were determined using the folin-sio-catheo method as well as HPLC to determine catechins, epigallocatechins, and gallocatechins. Molecular analysis was also performed by examining the relative expression of three critical enzymes in the flavonoid biosynthesis pathway, F3H, DFR, and LAR. Results: The results showed that utilizing NC along with NPK significantly increased the content of total polyphenols in tea compared to the control (without NC and NPK). All treatments reduced catechin content 4- to 6-fold. With an increase in NC concentration, epigallocatechin content increased. Gallocatechin content also revealed a slight increase in 100 mg.L-1 NC concentration. Chlorophyll content indicated a significant difference with a falling trend in treatments with low concentrations of NC; however, a significant difference with a growing trend was seen in treatments with 100 mgL-1 of nano chitosan. In comparison to the control, various NC treatments had similar protein content. Except for the 50 mg.L-1 NC+NPK treatment, there was an apparent significant difference in the SOD enzyme activity in each NC treatment, with a positive trend. With increasing NC concentrations, CAT enzyme activity also rose in various treatments. In treatments with insignificant NC concentrations, PPO enzyme activity significantly decreased. In different treatments, leaf water content rose. Only at a dose of 100 mg.L-1 NC+NPK did the relative expression of the F3H enzyme rise nearly three times compared to the control; in contrast, other treatments had no meaningful effect on relative expression. Different NC+NPK treatments raised DFR relative expression, and 100 mg.L-1 NC demonstrated the highest expression (4 times). Compared to the control, LAR relative expression increased at 0, 50, and 100 mg.L-1 NC and NPK. The treatment without NC and with NPK displayed the highest level of LAR expression, with an expression almost 2.5 times higher than the control. Conclusion: In Kashef cultivar tea plants, chitosan nanoparticles in various concentrations coupled with NPK increased the production of catechin compounds. This effectively reduced oxidative stress and enhanced green tea leaf quality. In addition to addressing oxidative stress, NC may play a practical role in green tea quality. Due to its biodegradable properties, nano chitosan can be used instead of chemicals to improve tea plants' green leaves quality and lower environmental pollution. | ||
| کلیدواژه‌ها [English] | ||
| Tea plant, Camellia sinensis (L.) O. Kuntze, NPK, nano chitosan, flavonoid, Dihydro flavonol 4 reductase, Flavonone 3 hyroxilasr, Leuoco anthocyanin reductase | ||
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