Study On Calcium-Encapsulated Carbon Dots To Enhance Calcium Uptake And Fruit Quality In Apple

As an essential nutrient for plant growth and development,calcium is critical to ensure crop yield and quality.Although soils are rich in calcium,there are problems such as poor solubility,easy calcification,precipitation and elemental antagonism,which greatly hinder the uptake and utilization of calcium nutrients by plants.Apple（Malus domestica）is an important fruit crop in the world,and the calcium content of the fruit is closely related to its quality,postharvest storage and shelf life.The plants absorb calcium mainly through the root system and transport it upward through the xylem to specific parts of the plant under transpiration pull,so calcium deficiency is mostly found in the tissues where transpiration is weak,such as young leaves and fruits.In agricultural production,calcium deficiency in apples often causes physiological diseases such as bitter pox,water heart disease and black spot disease,which seriously affect the quality of apple fruit.In recent years,nanomaterials have been widely studied for their small size,facile fabrication,green safety and good biocompatibility.Among nanomaterials,nanofertilizer,as one of the new nano-tools,provides a potential new method for plant nutrient delivery and shows excellent application prospects in agricultural production.However,the effects and mechanisms of application of nanofertilizers in promoting calcium uptake and improving fruit quality are still to be investigated.In this study,calcium-encapsulated carbon dots were synthesized by hydrothermal method and applied as a nano-calcium fertilizer in both laboratory and field trials to investigate their effect on calcium supplementation in apples with a view to evaluating the application potential of nano-calcium fertilizer.The main findings were as follows:（1）Calcium-encapsulated carbon dots（Ca-CDs）were prepared by hydrothermal method using calcium gluconate and urea as raw materials,and the related chemical characterizations were carried out.It was found that the as-prepared Ca-CDs showed an obvious blue fluorescence at 340 nm light excitation,with a maximum emission peak at 420 nm.Transmission electron microscopy and particle size analysis showed that the Ca-CDs exhibited a homogeneous spherical structure with an average particle size of 1.83 nm.Based on the analysis of atomic absorption spectra and thermogravimetric results,it was found that the Ca-CDs contained 1.5 w%of Ca.The crystalline phase of Ca-CDs was characterized by XRD,and it was found that the Ca-CDs had an amorphous structure.XPS results showed that the Ca-CDs material contained a little amount of Ca on the surface.The experimental results of the addition of Na₂CO₃ solution further excluded the presence of free Ca²⁺in Ca-CDs.Consequently,the desired Ca-CDs were obtained.（2）Subsequently,the transport mode of Ca-CDs in plants was investigated in this study by using the fluorescence property of Ca-CDs.The results showed that Ca-CDs were distributed in roots,stems and leaves after hydroponic cultivation of apple seedlings with blue Ca-CDs,indicating that Ca-CDs can be absorbed and transported by plants.（3）To further investigate the specific way of Ca-CDs entering into plant cells,‘Orin’apple calli were treated with Ca-CDs at 24^oC and 4^oC.As a result,Ca-CDs could be successfully observed in both calli cells.Meanwhile,‘Orin’apple calli were co-incubated with Ca-CDs using the calcium channel blocker La Cl₃ or the calcium chelator EGTA,and it was found that neither of them inhibited the entry of Ca-CDs into the cells.The results indicate that the transport of Ca-CDs is a passive transport,which is energy-independent and not affected by elemental antagonism effect.（4）After Ca-CDs treatment of apple seedlings for prolonged time period,it was found that all apple seedlings grew normally and fresh leaves grew at the top of the seedlings,which proved that the Ca-CDs were safe and non-toxic to apple seedlings.Meanwhile,apple seedlings hydroponically treated with Ca-CDs were transferred to nutrient solution culture without Ca-CDs,and by detecting the fluorescence of Ca-CDs in apple seedlings before and after the transfer,we found that Ca-CDs could be successfully degraded in the plants.（5）In order to investigate the calcium supplementation effect of Ca-CDs,calcium deficiency stress in Arabidopsis thaliana was simulated.The results showed that Ca-CDs could increase the fresh weight of shoots and roots,calcium content and leaf chlorophyll content,as well as decrease the MDA content and relative conductivity,indicating that Ca-CDs could alleviate the calcium deficiency stress in Arabidopsis.（6）Orchard applications of Ca-CDs were performed.Application of Ca-CDs significantly increased the calcium content and hardness of fruit during the growing period,both in spray and root irrigation treatments.Meanwhile,Ca-CDs increased chelated pectin,alkali-soluble pectin,cellulose content and decreased water-soluble pectin content.By measuring the ripe fruits,it was shown that Ca-CDs treatment significantly increased the content of soluble solids,soluble sugars and titratable acids compared to the blank control.In summary,we prepared a calcification-free,well-soluble and mobile calcium-encapsulated carbon dots.The experimental results showed that Ca-CDs have good calcium supplementation and ability to alleviate calcium deficiency.In addition,orchard applications by foliar spraying or root irrigation also demonstrated that Ca-CDs could significantly increase the calcium content in apple fruits and significantly improve apple quality,including fruit weight,hardness,pectin,etc.Therefore,the Ca-CDs synthesized in this study provide a new possible pathway in promoting the uptake of calcium nutrients and improving fruit quality.