Nutrient controlled release behaviors and plant growth of NPK encapsulated hydroxyapatite/alginate biocomposite toward agricultural and environmental sustainability

Controlled release fertilizers (CRFs) promote sustainable agriculture by gradually releasing nutrients into the soil while also mitigating environmental pollution. Nitrogen-phosphorus-potassium embedded hydroxyapatite/alginate (NPK-HA/Alg) biocomposite beads were developed using a simple, cost-effective, and environmentally friendly dropping and external gelation method. Addition of eggshell biowaste-derived HA to the alginate matrix improved the structural, thermal, and structural stability of the alginate beads, and enabled the inclusion of significantly high plant nutrients. The biocomposite beads exhibited a prolonged and controlled nutrient release in deionized water over 35 days. Biocomposite bead addition was assessed for the growth of flowering Chinese cabbage in a controlled greenhouse environment. Results confirmed vegetative growth with high values of plant height, number of leaves, and fresh and dry weights. The non-toxic and cost-effective NPK-HA/Alg biocomposite beads demonstrated controlled nutrient release as promising CRF materials to promote sustainable agricultural production.


Introduction
Sustainable agriculture has now emerged as an innovative alternative approach to reduce negative impacts on the environment.Controlled release fertilizers (CRFs) play an essential role in promoting sustainable agricultural systems, especially for crop production.CRFs offer an environmentally sustainable approach to increase crop yields by providing proper nutrient release patterns that suit both the physiological and biochemical aspects of plant growth.CRFs have been increasingly deployed in crop production to reduce issues of nutrient volatilization and leaching; however, their application in larger-scale agricultural practices remains sparse.CRFs are expensive, have low nutrient release efficiency, and CRF materials are typically composed of fossil fuel-based synthetic polymers including polystyrene, polyurethane, and polyethersulfone [1,2].Naturallyderived non-toxic and biodegradable polymers such as cellulose and its derivatives, starch, chitosan and alginate are interesting materials for CRF preparation [3,4].Alginate hydrogels have recently been developed as CRFs, with benefits including good availability, low cost, and high water absorption and retention [5][6][7].To minimize manufacturing costs, alginate was chosen as the CRF material, allowing easy gel formation under mild conditions.Alginate-Ca 2+ hydrogel beads incorporating potassium dihydrogen phosphate as a source of phosphorus fertilizer were previously produced as CRFs.They showed reduced initial burst phosphate release in water compared with conventional fertilizers [8].Bacteria and fungi use alginate as a carbon source, thereby facilitating the biodegradation of alginate in soil over several months [9].Alginate is an eco-friendly and nontoxic material suitable for soil addition and plant growth.
However, alginate has limited mechanical strength and stability because it decomposes in the presence of monovalent cations.Several strategies have been proposed to improve the mechanical properties and stability of alginate including blending it with other polymers [10,11] and incorporating inorganic particles to form a composite [12][13][14].Polyvinyl alcohol (PVA), chitosan and gelatin were blended with alginate-based ureacontrolled release fertilizers to improve the structural and thermal stabilities [7].Inorganic particles/polymer composites have advantages over polymer blending systems because they act as polymer reinforcements.Recently, composites based on montmorillonite/chitosan [15], bentonite/carnauba wax [16], and poly(acrylic acid-co-acrylamide)-grafted silica/deproteinized natural rubber [17] were reported as CRF materials.The drive toward more sustainable agriculture has motivated research efforts to minimize and utilize biowastes.Inorganic particles derived from biowaste such as hydroxyapatite (HA) synthesized from eggshells, were used to reduce the cost of the alginate matrix by acting as a particulate filler.The HA enhanced the mechanical properties of the alginate matrix by acting as a reinforcement.HA [Ca 10 (PO 4 ) 6 (OH) 2 ] inorganic mineral found in human bone and teeth has been widely used in bone regeneration studies and dental repair [18].During the last decade, HA has also attracted interest as an absorbent material for wastewater treatment [19][20][21].Currently, HA is recognized as a promising material for boosting soil quality and has been used as a phosphorus biofertilizer in agriculture [22].Nanohydroxyapatite/cellulose-grafted polyacrylamide composite hydrogels have been studied for their suitability as slow-release fertilizer materials, with HA composite hydrogels releasing mineral nitrogen content over 16 weeks compared to 4 weeks shown by conventional fertilizers [22].In an earlier study, we prepared and investigated the suitability of electrospun NPK-polyvinyl alcohol/polylactic acid core/shell fiber mats as CRF material [23].The fibers exhibited NPK-controlled release behavior with significantly reduced initial burst release compared to conventional fertilizers.However, fiber encapsulation efficiency was limited, as indicated by the low amount of loaded total nitrogen content.The development of nutrient carriers with enhanced ability to entrap fertilizers was thus the motivation for this study.
Here, NPK fertilizer-entrapped HA/alginate biocomposite beads were developed as controlled release fertilizers.Adhering to the zero waste philosophy, eggshells were used as the calcium precursor to synthesize HA particles as a value-added product using a simple, cost-effective, and environmentally friendly precipitation method.HA particles were incorporated in alginate beads to form biocomposites with enhanced structural and thermal stability of alginate.NPK-loaded HA/alginate biocomposite beads were expected to provide sustained nutrient release behavior and sufficient nutrient content after application at the start of the plant growth cycle.

Preparation of biocomposite beads
An alginate solution (alginic acid sodium salt from brown algae, molecular weight 100,000-200,000 g/mol, Sigma-Aldrich, USA) at a concentration of 1.5%w/v was prepared in deionized (DI) water.Urea (CO(NH 2 ) 2 99%, KemAus, Australia), diammonium phosphate ((NH 4 ) 2 (HPO 4 ) 99%, KemAus, Australia), and potassium dihydrogen phosphate (KH 2 PO 4 99%, KemAus, Australia) at a ratio of 2:1:1 were added to the alginate solution at a 50%w/w concentration of alginate.The mixture was stirred at room temperature until becoming a homogeneous solution.Eggshell powder was used as a calcium precursor to prepare hydroxyapatite (HA) using the chemical precipitation method, as reported in our previous study [18].The successful formation of HA particles was confirmed by the XRD result and HA morphology was determined by SEM (Supplementary figure S1).As-synthesized HA particles (d 50 ∼ 8.3 ± 0.3 μm) were added to the NPK-alginate solution at 10%w/w of alginate and continuously stirred for 2 h.Biocomposite beads were formed by the dropping method, followed by external gelation.The mixture was loaded into a 10 ml plastic syringe and then dropped into 100 ml of 0.1 M calcium chloride dihydrate (CaCl 2 •2H 2 O, Sigma-Aldrich, USA) crosslinking solution under continuous stirring, leading to bead gelation.The nitrogen-phosphorus-potassium embedded hydroxyapatite/alginate (NPK-HA/ Alg) biocomposite beads were filtered and washed with distilled water several times to remove the excess CaCl 2 solution and then dried at 50 °C in a hot air oven (UN110-230V, Memmert, Germany) until constant weight.The Alg and NPK-Alg beads were prepared following the same procedure.

Bead morphological analysis
The morphology of the Alg, NPK-Alg, and NPK-HA/Alg beads was analyzed using a scanning electron microscope (SEM, JSM-IT-500HR, JEOL, Japan).Before SEM analysis, the beads were coated with gold using a sputtering device (JFC-1200, JEOL, Japan).The beads were photographed, and the sizes of 200 randomly selected beads were determined from the photographs using ImageJ software.Histograms showing bead size distribution were constructed following Sturges' rule and the curves were then fitted using a log-normal function [24].The means of random bead sizes, along with standard deviations were reported, with variance and coefficient of dispersion statistically analyzed to describe the distribution of data and assess the level of variability.Energy dispersive x-ray spectroscopy (EDS, Oxford X-Max 50, Oxford Instruments, UK) was performed for elemental mapping analysis.In each SEM image of Alg, NPK-Alg and NPK-HA/Alg beads, the area attributed to calcium, phosphorus and nitrogen rich region was evaluated.

Chemical composition analysis
The chemical compositions of Alg, NPK-Alg, and NPK-HA/Alg beads were characterized using a Fourier Transform Infrared Spectrometer (ATR-FTIR; Nicolet170-SX, Thermo Nicolet Ltd., USA) over wavenumber range 4000 cm −1 to 400 cm −1 with a resolution of 4 cm −1 for 64 scans at room temperature.Energy Dispersive x-ray Spectroscopy analysis was performed to support the results of the FTIR analysis and qualitatively confirm the bead elemental compositions.

Determination of entrapment efficiency
The Kjeldahl method (ASTM International) was used to determine the total nitrogen (N) content loaded in the Alg, NPK-Alg, and NPK-HA/Alg beads, following our previous study [23].The Kjeldahl method for nitrogen determination of the beads was performed by digestion in sulfuric acid, distillation, and acid-base titration [25].Encapsulation efficiency (EE) was calculated using equation (1).
where N L is the total nitrogen content presented in the beads and N I is the total nitrogen content initially added in the beads (mg/g sample).The investigation was performed in five replicates.

Swelling ratio study
The swelling ratio of Alg, NPK-Alg, and NPK-HA/Alg beads was studied to estimate the water absorption capacity of the beads by measuring their geometry change after immersion in DI water.The diameters of dry beads were measured and used to calculate bead volume (V d ).Then, 0.5 g of each bead type was added to 50 ml of DI water in a polystyrene bottle and statically incubated at 25°C.After 24 h, the beads were removed and blotted with tissue paper to remove excess water from the surface.The diameters of the swollen beads were measured and used to calculate the bead volume at equilibrium (V w ).The swelling degree (SR) was determined through the equation (2).
Five sample sets were determined at each time point, and the results were reported as mean percentage ± standard deviation (SD).

Thermal analysis
The thermal behaviors of Alg, NPK-Alg, and NPK-HA/Alg beads were tested using a Diffraction Scanning Calorimeter (DSC, NETZSCH 204F1 Phoenix, NETZSCH, USA) at temperatures ranging from 25 °C to 200 °C and a heating rate of 10 °C min −1 .The thermal stability of the beads was analyzed using a Simultaneous Thermal Analyzer (STA, NETZSCH5 STA 449F3, NETZSCH, USA) under nitrogen atmosphere from 50 °C to 900 °C at a heating rate of 10 °C min −1 .

Nutrient release behaviors and kinetics study
The release behaviors of NPK-Alg beads and NPK-HA/Alg biocomposite beads were investigated by soaking in DI water for 35 days at room temperature and compared with conventional NPK granules.One gram of each bead type was placed in a polystyrene bottle containing 50 ml of DI water, and 1 ml of the aqueous medium was withdrawn at each time point and used to determine the total nitrogen content using the Kjeldahl method.Five replicates were investigated for each bead type.The release amount of the beads was calculated by using the equation (3).

( ) ( )
where N t is the nitrogen content released at time t and N i is the initial nitrogen content.
A well-known kinetic model like the Korsmeyer-Peppas mathematic model was used to fit the release results.The dissolution mechanism is represented in the equation (4).
where M t /M ∞ is a fraction of nitrogen content released at time t, k is the Korsmeyer-Peppas rate constant, and n is the diffusional exponent.The slope of the logarithm plot of M t /M ∞ and time (t) determined the n value.The n value indicates the release mechanism, including Fickian diffusion when n < 0.43, anomalous transport when 0.43 < n < 0.85, and the case II transport mechanism (zero-order kinetic) when n > 0.85 [26].

Plant growth assessment
Pot experiments to investigate the effects of the controlled release fertilizers on plant growth were conducted following the OECD 208 Terrestrial Plant Growth Test [25] with a slight modification.Plant growth with NPKfree Alg, NPK-Alg, and NPK-HA/Alg beads was assessed under good horticultural practices in a controlled greenhouse environment (Internet of Things monitoring system) using flowering Chinese cabbage (Brassica rapa subsp.chinensis var.parachinensis) and compared with conventional fertilizer (NPK 21-21-21, Twenforty ® , Thailand) and with no additional fertilizer as the controls.The soil was sieved (0.5 cm) to remove coarse fragments and 450 g was mixed with 0.5 g of each bead type.The mixtures were filled in five-inch cultivation pots and the experiment was carried out with five replicates.The plant seeds were first cultivated in seedling trays containing peat moss for 5 days, then transferred to the cultivation pots prepared as above (one seedling per pot) and maintained in the greenhouse for a further 35 days with adequate daily watering.The environmental conditions were controlled by a venting and cooling system at 32 °C ± 2 °C under constant humidity of 70% ± 25% and a photoperiod of 12-hour light (natural light from July to August 2023).Plant growth performance was monitored at least weekly by taking digital photographs during the observation period (7 to 35 days).At the end of the test, the height, number of leaves, and fresh weight of the harvested plants were determined, with the dry weight measured after drying in a hot air oven at 60 °C for 48 h.

Statistical analysis
The results obtained from replicate studies were reported as an average ± SD.A one-way ANOVA was used to analyze statistical differences.A p-value of < 0.05 presents significance among the comparative results of the samples.

Results and discussion
3.1.Physical appearance, size distribution and surface morphology Alginate (Alg) beads were fabricated using a simple, cost-effective, and environmentally friendly external gelation method with the presence of calcium cations leading to ionic crosslinking.Irregular-shaped and compact beads were produced (figure 1(a), (a´)) with diameters of 2.6 ± 0.3 mm.The Alg beads were used as plant nutrient carriers, leading to the formation of NPK-Alg beads.The addition of NPK nutrients did not improve the structural shape of the Alg beads (figure 1(b)).NPK-Alg bead formation and uniformity were more difficult to achieve compared to the production of NPK-free Alg beads, possibly caused by the additional NPK composition interfering with bead formation during the gelation method.Average bead sizes increased to 2.7 ± 0.4 mm with entrapment of NPK nutrients (figure 1(b´)).Bead size range also increased from 1.64 mm to 2.17 mm, the variance increased from 0.08 to 0.12, and the coefficient of dispersion increased from 0.03 to 0.05 with addition of NPK nutrients.Hydroxyapatite (HA) in the Alg beads formed NPK-HA/Alg biocomposite beads.HA is a calcium phosphate compound and acts as an additional plant nutrient.Addition of HA improved bead formation as spherical and elliptical shapes (figure 1(c)).The size of the NPK-HA/Alg beads was 3.6 ± 0.4 mm, and significantly higher than the NPK-free Alg and NPK-Alg beads.The increased viscosity of dropping alginate solution when adding HA particles led to an increase in bead size.This finding agreed with a study of bentonite/alginate beads [27].Compact beads were obtained with a more uniform and smaller size distribution than NPK-Alg beads with size range, variance, and coefficeint of dispersion 2.06 mm, 0.15, and 0.04, respectively.Inclusion of HA in the beads decreased the variability in bead formation and gave better control over bead geometry and bead size distribution.HA acted as a polymer filler and enhanced the structural integrity and stability of the beads.HA addition facilitated uniform distribution within the alginate matrix due to its inherent hydrophilic properties and excellent compatibility with alginate aqueous solutions.
The surface morphology of Alg, NPK-Alg, and NPK-HA/Alg beads is shown in figures 2(a), (b), (c).All bead formulations had very rough and irregular surfaces.The morphological structure of Alg beads (figure 2(a)) was similar to chitosan/alginate or gelatin/alginate beads, as previously reported [11].The bead surface was similar to orange peel as a typical characteristic of calcium-crosslinked alginate beads [28].Figure 2(b) shows plate-and rod-like crystals distributed on the bead surface and embedded in the alginate matrix.These plate-and rod-shaped crystals were assumed to be particulate NPK nutrients.Figure 2(c) shows aggregated HA particles well distributed in an alginate matrix, combined with plate-and rod-shaped crystals of NPK nutrients.The addition of HA particles in the beads gave a fine and dense morphology, indicating harder and more compact biocomposite beads compared to Alg beads without HA.
EDS mapping analysis was performed on the Alg, NPK-Alg and NPK-HA/Alg beads in order to investigate the distribution of hydroxyapatite (HA) particles in the alginate matrix (figure 3).Observation of calcium (Ca)and phosphorus (P)-rich regions revealed the distribution of HA particles, alongside Ca-crosslinked alginate.The Ca and P elements were found to be concentrated on the surface of NPK-HA/Alg biocomposite beads, as evidenced by the presence of rich regions displaying red and green spots, respectively.and demonstrated a uniform distribution of HA particles within the alginate matrix.Additionally, the distribution of NPK fertilizer was confirmed through the EDS mapping of the nitrogen (N)-rich region (depicted by blue spots).The nitrogen element's blue spots were well-dispersed on the maps of both NPK-Alg beads and NPK-HA/Alg beads, consistent with expectations.A higher nitrogen content was detected in the NPK-HA/Alg beads, likely attributed to the enhanced interaction between the hydroxyapatite (HA) particles and the NPK fertilizer.

Chemical composition
The FTIR spectra of calcium crosslinked Alg beads and neat sodium alginate (neat Alg) were analyzed.The characteristic peaks of alginate at 1589 cm −1 corresponded to the asymmetric vibration of carboxylate salt ion, while the peak at 1404 cm −1 was attributed to its symmetric vibration (figure 4(a)).Stretching vibration of the O-H bond exhibited as a broad band ranging from 3500 cm −1 to 3000 cm −1 [29][30][31].After crosslinking with calcium cations, the peak of carboxylate (-COO − ) groups in the alginate structure showed reduced intensity due to ionic interaction between carboxylate anions and calcium cations.Alginate beads were formed through the egg-box mechanism [29], confirming the successful crosslinking of alginate with calcium ions.When NPK nutrients were loaded in the beads, the spectrum of NPK-Alg beads showed additional characteristic peaks of urea (NH 2 CONH 2 ) [13,32].A doublet peak with broad bands at 3351 cm −1 and 3292 cm −1 was attributed to N-H stretching vibration [11], while the absorption peak at 1658 cm −1 was due to the carbonyl group.The peak at 1575 cm −1 was assigned to N-H deformation with vibration of C-N stretching appearing at 1456 cm −1 [11].All the characteristic peaks of urea were found in the spectrum of NPK-HA/Alg beads, indicating the existence of fertilizers in the Alg and HA/Alg beads.The addition of HA to the beads led to the appearance of small new peaks at 1415 cm −1 , 1012 cm −1 , 600 cm −1 , and 557 cm −1 in the spectrum of NPK-HA/Alg beads.The peaks at 1012 cm −1 and 557 cm −1 were reported as PO 4 3− of hydroxyapatite, the characteristic peak of HPO 4 2− was observed at 600 cm −1 , and the peak corresponding to asymmetric stretching of the carbonate (CO 3 2− ) group was detected at 1415 cm −1 [18].This observation confirmed the presence of HA in the Alg beads.The entrapment of NPK fertilizers in the HA/Alg beads was confirmed by EDX analysis, with the EDX spectrum of NPK-HA/Alg beads (figure 4(b)) demonstrating the compositional peak of N, P, and K, supporting the FTIR spectra.Detection of the calcium peak indicated sources as both the calcium crosslinking agent and HA.

Entrapment capacity
The result of the EDS analysis (figure 4(b)) was used to qualitatively confirm the entrapment of NPK fertilizers in Alg beads and HA/Alg beads.The Kjeldahl method was also performed to quantitatively determine the ability of Alg beads and HA/Alg beads to entrap plant nutrients, and presented as total nitrogen content (mg/g sample), as shown in figure 5. Total N content was determined at 102 ± 2 mg g −1 of NPK-Alg beads and 114 ± 6 mg g −1 of NPK-HA/Alg beads, indicating that both types of beads could carry a considerable amount of nutrients.The presence of HA particles improved the entrapment capacity of Alg beads ( * p < 0.05) in the alginate matrix, while increased ionic strength enabled the bead materials to better interact with the fertilizer [33].Compared to our previous study, the NPK carrier in the form of core-shell polymeric fibers achieved a lower amount of encapsulated total nitrogen at 10.6 ± 0.5 mg/g sample [23].Fertilizer consumed during a plant growth cycle required more polymeric materials, leading to increased production costs.HA/Alg biocomposite beads can contain 10 times more plant nutrients than fiber bead types.This simple production method to produce biocomposite beads offers a suitable alternative with low cost of materials and high loading ability.The bead formation method in this study facilitated large-scale production of the released fertilizer product.

Swelling ratio
Besides acting as a plant nutrient carrier, alginate beads can also uptake and retain large amounts of water to support plant growth and moisturize the soil.The networking structure of crosslinked alginate hydrogel promotes high water absorption by the beads and, subsequently a high swelling ratio.The diameters of swollen Alg, NPK-Alg, and NPK-HA/Alg beads were 4.3 ± 0.3, 5.0 ± 0.3, and 5.7 ± 0.7 mm, respectively.Bead sizes of all formulations in the wet state doubled compared to dry beads.The swelling ratio (SR) was reported as a dimensional change of Alg, NPK-Alg, and NPK-HA/Alg beads at 320%, 505%, and 300%, respectively (figure 6) indicating that they contained high amounts of water.The ability to absorb water was influenced by bead size, while the incorporation of HA reduced the swelling ratio (%SR) of Alg beads.The compact nature of the HA/Alg beads inhibited water penetration through the matrix network.This result concurred with the swelling ability of the bentonite-starch-alginate bead network [27].The SR value at 300% of NPK-HA/Alg biocomposite beads was sufficiently high to control the release of fertilizer and induce the diffusion mechanism.Plant water absorption ability resulted in lower water requirements during growth and was beneficial for agricultural production, especially during the dry season.

Thermal behavior and stability
Thermal transitions of NPK-HA/Alg biocomposite beads were analyzed by the DSC technique and compared to NPK-Alg beads, Alg beads, and neat Alg.The thermogram of neat sodium alginate as a raw material used in bead production (figure 7(a)) showed a broad peak of endothermic transition ranging from 50 °C to 200 °C that was related to water evaporation [34].An additional small peak observed at 170.6 °C was assigned to impurities contaminating the sodium alginate powder.The DSC curve of Alg beads prepared by the ionotropic gellification method showed similar thermal transition behavior to neat alginate.The endothermic peak of water evaporation shifted to a lower range of 50 °C to 125 °C.The DSC curves of NPK-Alg and NPK-HA/Alg beads differed from the curves of Alg beads in the presence of NPK nutrients and HA particles.Addition of NPK and HA flattened the dehydration endothermic peaks, with peaks of NPK-Alg and NPK-HA/Alg beads appearing at 110.3 °C and 94.5 °C, respectively and shifting at lower temperatures due to strong interaction among all the components in both formulations [35].This result agreed with the compact structure of the NPK-HA/Alg biocomposite beads.The DSC curve of Alg beads exhibited a sharp endothermic band at 180.9 °C, related to the cleavage phenomenon of an ordered molecular arrangement within the beads [36].After HA was incorporated into the beads, the cleavage peak shifted to a higher temperature close to 200 °C because addition of HA enhanced the thermal stability of the beads.
TGA analysis was used to investigate the compositions of biocomposite beads and evaluate whether NPK fertilizers were lost during bead production.dehydration at 70 °C due to the evaporation of moisture trapped on the bead surface, and 150 °C due to evaporation of the water molecules bound inside the beads.This finding concurred with the DSC results.The third weight loss, stage III ranged from 150 °C to 400 °C and was associated with thermal decomposition of the alginate matrix [28].Stage III weight loss was 36% to 38%.The last weight loss (stage IV) was observed at 450 °C to 550 °C in the curve of Alg beads but was not found in the curves of NPK-Alg and NPK-HA/Alg composite  beads.This transition was due to the pyrolysis of the polymer matrix and the elimination of volatile products [37].The residual weights of Alg, NPK-Alg, and NPK-HA/Alg beads were determined at 30.34%, 32.41%, and 35.02%, respectively as the inorganic components in the beads.These corresponded to the combination of crosslinked calcium cations, embedded NPK fertilizer, and filled HA particles and confirmed that the ingredients were not lost during the fabrication method.The bead formulation decomposition temperature (T d ) increased in the presence of NPK and HA, indicating improved thermal stability of the Alg beads caused by hydrogen bonding interaction between the alginate matrix, plant nutrients, and HA particles through the ionic crosslinking of calcium cations.

Fertilizer release behavior
NPK-Alg beads with and without adding HA were investigated in DI water and compared to the conventional NPK granules without coating.The nutrient release profiles are shown in figure 8(a).The conventional NPK granules dissolved in water in a shorter time, as indicated by burst release, and reached 100% at the initial stage (30 min) of investigation.The NPK-Alg beads and NPK-HA/Alg biocomposite beads exhibited similar release behavior, with an initial burst release of up to 50% within 100 h of immersion followed by a gradual release until the end of the investigation.After 7 days of immersion, NPK fertilizer loaded in Alg beads was released faster than fertilizer loaded in HA/Alg biocomposite beads.The dense and compact structure of NPK-HA/Alg beads retarded water penetration through the alginate network, thereby slowing down the diffusion of entrapped nutrients throughout the medium.This finding concurred with a previous study that investigated adding clay to an alginate matrix.Results showed that the release of active compounds was controlled by the addition of clay to the polymer matrix [27].The release results of NPK-Alg and NPK-HA/Alg beads differed from CRF granules produced using polymer coating, as previously reported [1].The CRF-based capsules, comprising the nutrientencapsulated core region and the polymeric barrier, showed slower release than the polymeric spheres based on the nutrients embedded in the polymeric matrix [38].The capsules demonstrated three stages of a sigmoidal release pattern including the lag phase, constant release phase, and decay phase [1].The initial burst release occurred in the release patterns of both NPK-Alg and NPK-HA/Alg beads instead of the lag phase.The first release stage of nutrients embedded in the alginate matrix probably occurred together with the nutrients adsorbed on the surface of the beads.The nutrients on the surface of the beads rapidly dissolved at first contact with the aqueous medium, followed by the second constant release phase caused by water penetration through the alginate matrix and nutrient diffusion.To confirm this assumption, the release kinetics and mechanisms of NPK fertilizer released from the biocomposite beads were identified using the Korsmeyer-Peppas mathematical model.As shown in figure 8(b), the release parameters and the average release exponent (n) values for both NPK-Alg beads and NPK-HA/Alg beads were 0.4807 with a standard deviation of 0.04 and 0.6049 with a standard deviation of 0.0007, respectively.The R 2 coefficient values of both bead types were higher than 0.99, indicating that the NPK release behaviors fitted well with the Korsmeyer-Peppas model.The n values in the range 0.43 < n < 0.85 indicated anomalous (non-Fickian) transport as mixed diffusion and chain relaxation mechanisms.Results confirmed that the release mechanism of the biocomposite beads in this study was controlled by diffusion control and swelling control.Water penetrated the interior matrix and swelling occurred.The alginate chains then relaxed, allowing the nutrients to diffuse, and gradual release occurred after the initial burst release.To investigate the influence of hydroxyapatite (HA) particles within the alginate beads on release behaviors, the Korsmeyer-Peppas release rate constant (k) for both NPK-Alg and NPK-HA/Alg beads was determined, with values of 0.0404 and 0.0137, respectively.The inclusion of HA particles in the alginate beads hindered the release rate of nutrients.

Plant growth performance
The plant growth performance of flowering Chinese cabbage was assessed over 35 days with treatments of NPK-Alg beads and NPK-HA/Alg biocomposite beads and compared with the application of conventional fertilizers, fertilizer-free Alg beads, and the absence of both fertilizer and beads as the control.As shown in figure 9, the tested plants survived under all the treatments, thereby confirming the non-toxicity of the materials.The treatments (c) conventional NPK, (d) NPK-Alg beads, and (e) NPK-HA/Alg beads showed faster growth than plants treated with Alg beads and the control.Yellow and small leaves, indicating nitrogen deficiency, were found in treatments (a) the control and (b) Alg beads.After 28 days of growth, the positive effects of NPK-Alg beads and NPK-HA/Alg beads were more pronounced.Plants applied with (d) NPK-Alg beads and (e) NPK-HA/Alg beads grew healthily with larger leaf size and higher leaf numbers compared to plants treated with (c) conventional NPK granules.After 35 days, treatments of NPK-Alg beads and NPK-HA/Alg biocomposite beads showed increased plant height with several large green leaves, similar to plants treated with conventional fertilizer.Treatments c, d, and e significantly promoted leaf numbers, plant height, and fresh and dry weights compared to treatments a and b, as shown in figures 10(a)-(c), and (d), respectively.Results among the c, d, and e treatments showed no significant differences, indicating that the utilization of bead-based CRFs, as only one application at the beginning of the plant growth cycle, exhibited an effective performance by providing sufficient nutrients for vegetative growth.Fertilizer granules without the polymer carrier rapidly dissolved and were applied every two weeks.The nutrients were partially leached out of the fertilizer granules by the daily watering, leading to ineffective uptake of nutrients by the plants.Conventional fertilizer granule treatment was applied a few times during the entire growth stage.By contrast, a one-time application of the biocomposite-based CRFs was sufficient for the entire growth stage and avoided nutrient loss by providing nutrients in a controlled manner, subsequently reducing excess fertilizer used for planting, and minimizing adverse impacts on the environment.

Conclusions
This study successfully enhanced the encapsulation efficiency of plant nutrients using NPK-embedded biocomposite beads based on alginate and hydroxyapatite (HA) particles.Eggshell biowaste was used as the calcium source to synthesize the bio-based HA using a simple and inexpensive precipitation method.NPK-HA/ Alg biocomposite beads were fabricated using a simple, cost-effective, and environmentally friendly dropping and external gelation methods.Beads added with inorganic HA were dense and compact, leading to enhanced structural and thermal stability.The biocomposite beads entrapped large amounts of nutrients at 114 ± 6 mg nitrogen content per gram of sample.NPK-HA/Alg biocomposite bead formulation showed potential as a carrier of fertilizer using cost-effective materials with low energy production requirements in a short time.The biocomposite beads exhibited a controlled release of plant nutrients over a longer period, compared to the release behavior of commercial fertilizers without polymeric carriers.The release mechanisms of the NPK-Alg and NPK-HA/Alg beads were driven by diffusion and chain relaxation mechanisms, as confirmed by the Korsmeyer-Peppas model.The results of plant growth assessment confirmed that NPK-Alg beads and NPK-HA/Alg biocomposite beads promoted high efficiency of vegetative growth, as qualitatively evidenced by visual observation after 35 days and quantitatively confirmed by the highest dry weight, numbers of leaves, and height of harvested plants among all treatments.The NPK-HA/Alg biocomposite beads exhibited higher efficiency in terms of controlled release rate and plant growth performance compared to the NPK-Alg beads.Plants, subjected to bead application at the beginning of the growth cycle grew effectively, thereby promoting agricultural production following the philosophy of sustainable development.The NPK-HA/Alg biocomposite beads actively functioned as a controlled release fertilizer to enhance and promote agricultural and environmental sustainability.

Figure 1 .
Figure 1.Photographs showing shape and histograms illustrating bead size distribution of (a), (a´) Alg beads, (b), (b´) NPK-Alg beads and (c), (c´) NPK-HA/Alg biocomposite beads.The log-normal function is depicted by the solid line.Mean bead diameter, standard deviation (SD), and size dispersion parameters including variance and range are reported.

Figure 3 .
Figure 3. EDS mapping of calcium, phosphorus and nitrogen in Alg beads, NPK-Alg beads and NPK-HA/Alg beads.

Figure 7 (
b) shows the TGA curves of NPK-Alg and NPK-HA/Alg beads compared with the curve of Alg beads.All formulations exhibited two transition stages (I-II) of

Figure 4 .
Figure 4. (a) FTIR spectra of Alg beads, NPK-Alg beads, and NPK-HA/Alg beads in comparison with the spectra of neat Alg and eggshell-derived HA and (b) EDS analysis showing the chemical compositions of NPK-HA/Alg beads.

Figure 5 .
Figure 5. Entrapment capacity of NPK fertilizer with HA/Alg biocomposite beads compared to that of Alg beads as presented by total nitrogen content (mg/g sample).* Symbol shows the significant difference (p < 0.05) of NPK-HA/Alg beads in comparison with NPK-Alg beads.

Figure 8 .
Figure 8.(a) Release behaviors and (b) release kinetics of Korsmeyer-Peppas mathematical model determined by release study of Alg beads, NPK-Alg beads, and NPK-HA/Alg biocomposite beads by immersion in DI water for 35 days compared to those of DI water and conventional fertilizer.

Figure 9 .
Figure 9. Representative digital photographs of flowering Chinese cabbage growth treating with (a) an absence of fertilizers as control, (b) NPK-free Alg beads, (c) conventional chemical fertilizers, (d) NPK-Alg beads and (e) NPK-HA/Alg beads, after 14, 21, 28 and 35 days of growth.

Figure 10 .
Figure 10.Effects of controlled release fertilizers treatments on the vegetative growth parameters of flowering Chinese cabbage after 35 days of planting including (a) number of leaves, (b) height, (c) fresh weight, and (d) dry weight of harvested plants.* p < 0.05 indicates statistically significant differences among the treatments.