Improvement in physio-biochemical characteristics of shallot plants with nano silica at several levels of drought stress

Plants are stressed if the environment in which they grow changes, which can reduce the rate of vegetative and generative growth and finally the production of plant. One of the factors that causes stress in plants is the availability of water which greatly influences the productivity of shallot plants as shown by changes in physiological and biochemical characters. Analysis of physiobiochemical characteristics of shallots for the application of nano silica at several levels of water stress is the aim of this research. Parameters analyzed at the Central Pharmacy Research Laboratory Faculty of Pharmacy, Tissue Culture Laboratory and Disease Laboratory, Faculty of Agriculture, Universitas Sumatera Utara, November 2022 to March 2023. This study used RBCD with 2 treatment factors. The first factor namely water stress condition (80, 60 and 40% field capacity). The second factor namely the application of nano silica which consisted of 4 levels of concentration (0, 6, 12 and 18 g/l). The results showed that the addition of water based on field capacity at 80% optimal conditions significantly increasing chlorophyll a, b, and total, leaf relative water content, H2O2 and SOD enzymes. Application of nanosilica at concentration of 18 g/l significantly increased chlorophyll a, b and total.


Introduction
Changing climate and environmental conditions pose a threat to crop production throughout the world, new solutions are needed to increase the adaptability of plants to environmental changes.The abiotic stress that has the most negative impact on the growth and production of cultivated plants is drought stress.Shallots are plants that have shallow roots which limit the ability of the roots to get water from deeper zones.This morphological condition makes shallots very sensitive to water deficit stress.Plant responses to water deficit conditions will disrupt various biochemical and physiological processes in plants which are reflected in phenotypic variations [1].
Apart from inhibiting physiological functions, drought stress will reduce hormonal activity, synthesis of chlorophyll and increase temperature of canopy which results in a decrease in photosynthetic activity and plant metabolism.Water deficit conditions also reduce membrane permeability and increase the production of reactive oxygen species [2].
Currently, the use of nanoparticles is an effort to increase crop production in conditions of climate change and increasing drought intensity.The application of nanoparticles can significantly increase plant growth and production under drought stress through the mechanism of increasing photosynthetic efficiency, protecting photosynthetic organelles and oxidative damage, increasing accumulation of osmolytes and hormones and antioxidant activity [3].
Conditions of water shortage can cause an increase in ROS and accumulation of malondialhehid which causes plants to experience oxidative damage.The use of silicon nanoparticles can increase enzymatic antioxidant activity and reduce malondialhehid content in plants experiencing water deficit [4].
Analyze the physiobiochemical characteristics of shallots for the giving of nano silica at several levels of water stress as objective of this research.

Materials and methods
This research starts from November 2022 to March 2023 in Central Pharmacy Research Laboratory Faculty of Pharmacy, Tissue Culture Laboratory and Disease Laboratory Faculty of Agriculture.Using a randomized block design with two factors, the first factor namely water stress with 3 levels (80%, 60% and 40% field capacity) and the second factor namely application of nano silica with 4 levels (0, 6, 12 and 18 g/l).
The study began with land preparation, determination of soil water content and field capacity, nursery, transplanting at 40 DAP (Days After Planting), watering based on field capacity, application of nano silica, plant maintenance, leaf sampling an observing parameters at 63 DAP.
The observed variables were the chlorophyll a, b, and total, leaf relative water content, Hydrogen Peroxide (H₂O₂), Superoxide Dismutase (SOD).Data on observed variables were analyzed using analysis of variance.Testing of the influence of significant observed variable data was tested using the Duncan Multiple Range Test (DMRT) at alpha level = 5%.

Results and discussion
Climate change causes the environment for plant growth to change which reduces plant productivity.One of the most frequent environmental changes is water shortage.Plants need sufficient water available to grow and produce optimally.In plants, water plays a very vital role, namely functioning as a solvent for various nutrient compounds in the soil to meet plant needs, a transport medium for dissolved organic and inorganic compounds, maintaining cell turgidity, a raw material in the photosynthesis process, the hydrolysis process, regulating plant temperature and various other chemical reactions [2].[5] stated that chlorophyll synthesis, photosystem II efficiency and electron flow behavior decrease due to drought stress, causing photosynthetic efficiency in plants to decrease.Note : Based on DMRT at the alpha level 5%, numbers followed by different letters show significant differences.
The organic matter production index and plant growth are determined by the chlorophyll content.The photosynthetic response to drought stress is very important to understand through biochemical characteristics.The decrease in water content in the planting media caused a decrease in the chlorophyll a, b and total content showed a significant decrease as shown in Table 1.The results showed a decrease in chlorophyll a content of 24.16%, chlorophyll b 19.49% and total chlorophyll 7.10% at 40% field capacity conditions when compared to optimal conditions (80% field capacity).One indicator of plants experiencing drought stress can be seen from the chlorophyll content in the leaves.This is because the rate of photosynthesis will increase in line with the increase in chlorophyll content in plants.[6] stated Water stress will affect the formation of chlorophyll and can cause changes in the structure of chlorophyll.The decrease in chlorophyll content under drought stress conditions was due to accelerated chlorophyll degradation caused by high expression of chlorophyllase genes and pheophytinase activity.On the other hand, the genes encoding chlorophyl synthesis characteristics and enzymatic activity did not show significant changes due to stress.
The application of nano silica increased the chlorophyll a, b and total content significantly (Table 1).Research data shows that shallot plants treated with foliar application of nano silica at a concentration of 18 g/l produced the highest chlorophyll content.The percentage increase in chlorophyll a content was 31.49%,chlorophyll 22.90% and total chlorophyll 24.17% when compared to the chlorophyll content in shallot leaves that were not treated with nano silica.The research results of [8] also showed an increase in chlorophyll content in several varieties treated with foliar application of SiO2 nanoparticles at concentrations of 60 and 90 ppm when compared to the control.The smaller particle size of silica nanoparticles makes the absorption process easier and increases the chlorophyll content more quickly.Plants given silica have leaves that are more resistant to stress conditions with a rougher texture and a more horizontal leaf position, have more chlorophyll content, and can reduce the activity of the carboxylase enzyme and delay the senescence process which ultimately can increase the efficiency of photosynthesis [7].Note : Based on DMRT at the alpha level 5% , numbers followed by different letters show significant differences.
One of the important parameters that indicates the water status of plants is the relative water content (RWC) of leaves which shows the balance between the plant's transpiration rate and the water supply to the leaf tissue.The data in Table 2 shows that the RWC of leaves experiencing drought stress decreased significantly when compared to optimal conditions, namely at 80% field capacity (Table 2).The research results of [8] also showed that the RWC of bean leaves decreased significantly due to water stress.[9] stated that the RWC of leaves decreased due to less frequent watering.the less water can be transported from the roots to the leaves, causing the relative water content of the leaves to be low.Reducing the relative water content of leaves is a form of avoidance mechanism to regulate water use for plants during drought stress so that limited water can be utilized properly.Meanwhile, the application of nano silica to shallots had no significant effect on the relative water content of the leaves (Table 2).Drought stress causes reactive oxygen species (ROS) to be produced in large quantities in cellular systems.One type of ROS is hydrogen peroxide (H2O2).The research results showed a significant increase in H2O2 production in onion plants experiencing drought stress (Table 3).Shallots planted at optimal soil water content (80% field capacity) produce lower peroxide than those under drought conditions (40% field capacity).The research results of [10] also showed that watering shallot plants every 5 days caused the H2O2 formed to increase significantly compared to watering water carried out every day.[11] stated that ROS production increased several times under water stress conditions.Drought stress causes disruption of plasma membrane integrity, increased lipid peroxidation, early senescence which ultimately results in plant production decreasing significantly.
The hydrogen peroxide (H2O2) content produced by shallot plants under drought stress showed no significant effect with the use of nanosilica (Table 3).The lowest H2O2 content was produced by shallot plants treated with nanosilica with a concentration of 6 g/l, namely 0.126 μmol/g.According to [12], abundance of H2O2 which is one of the Reactive Oxygen Species will be dangerous because it can cause molecular damage, proteins and even DNA.In plant cells, there are molecules that reactively bind to free radicals called ROS.One of the ROS that can signal changes in physiological processes in plants is hydrogen peroxide (H2O2).Based on the research results of [13] stated that silica fertilizer is able to suppress the presence of H2O2 as ROS so that it can increase physiological resistance such as chlorophyll content and relative water content in leaves.Note: Based on DMRT at the alpha level 5%, numbers followed by different letters show significant differences.
Plants attempt to survive damage caused by abundant ROS production by increasing antioxidants, both through enzymatic antioxidant mechanisms and non-enzymatic anti-oxidants which play a role in protecting plants from damage due to oxidative stress [14].The data that can be seen in Table 4 shows that the activity of the superoxide dismutase enzyme in drought conditions increased significantly compared to other treatments, namely 18.935 units/mg protein.Water treatment based on 80% field capacity tends to have a lower SOD enzyme, namely 16.931 units/mg protein.When plants experience high drought stress, the activity of the superoxidase dismutase enzyme will increase.[14] stated that in plants experiencing water stress, excessive production of ROS and SOD which is an antioxidant enzyme produced by plants aims to stimulate plant resistance and scavenging excess ROS.ROS production induced by drought stress causes changes in metabolic activity, damage to cell membranes and increased cell death in plant cells.
The application of nano silica to shallot plants that were stressed by drought had no significant effect in increasing the activity of the SOD enzyme (Table 4).Although it can be seen that giving nano silica with a concentration of 12 g/l tends to increase the SOD enzyme by 5.06% when compared to treatment without giving nano silica.The SOD enzyme is an antioxidant enzyme that functions to increase plant resistance to oxidative stress.The SOD enzyme can convert superoxide radicals into H2O2.Even though H2O2 is not as reactive as superoxide, it is still in the form of free radicals which are unstable and dangerous for plants in plant metabolism.In organisms, H2O2 and other superoxides will be converted into oxygen and water by SOD [10].

Conclusions and suggestions
The results showed that the addition of water based on field capacity at 80% optimal conditions had a significant effect on increasing chlorophyll a, b, and total, leaf relative water content, H2O2 and SOD enzymes.Application of nanosilica at concentration of 18 g/l significantly increased chlorophyll a, b and total.The interaction of water stress level and nano silica application had no significant effect on all observed physiobiochemical parameters.

Table 1 .
Mean of chlorophyll a, b, and total of shallot plants with nano silica at several levels of drought stress.

Table 2 .
Mean of leaf relative water content of shallot plants with nano silica at several levels of drought stress.

Table 3 .
Mean of hydrogen peroxide (H₂O₂) of shallot plants with nano silica at several levels of drought stress.
Note : Based on DMRT at the alpha level 5% , numbers followed by different letters show significant differences.

Table 4 .
Mean of superoxide dismutase (SOD) at of shallot plants with nano silica at several levels of drought stress.