Synthesis and Characterization of Calcium Carbonate Obtained from Green Mussel and Crab Shells as a Biomaterials Candidate

Green mussel and crab shells are natural sources of CaCO3, which is widely used as a bioceramic for biomedical applications, although they are commonly disposed of in landfills. The improper disposal of green mussel and crab shells can cause environmental pollution, reducing the quality of life in the community. Many studies have reported the preparation of CaCO3 from green mussels and crab shells. However, there are limited studies comparing the characteristics, including the crystal phase obtained, weight percentage (%) of crystal, crystal size, crystal system, and elemental composition of CaCO3 from green mussel shells, crab shells, and commercial CaCO3. The objective of this research was to compare the calcium carbonate properties formed from green mussel (PMS) and crab (PCS) shells to commercial CaCO3. Green mussel and crab shells were crushed to powder and were calcined at 900 °C for 5 h. Precipitated Calcium Carbonate (PCC) was synthesized from calcined green mussel and crab shells using a solution of 2M HNO3, NH4OH, and CO2 gas. The effect of setting parameters on the synthesized product was analyzed using XRD and SEM-EDX methods. This study shows that the chemical composition of PMS is nearly identical to that of commercial CaCO3, where no contaminants were identified. In contrast, PCS has N components other than Ca, C, and O. Furthermore, the predominance of the vaterite crystal phases in PMS and PCS, with respective weight percentages of 91.2% and 98.9%, provides a benefit for biomaterial applications. The crystallite sizes of vaterite in PMS, PCS, and calcite in commercial CaCO3 are 34 nm, 21 nm, and 15 nm, respectively.


Introduction
Shellfish are animals that live in water (salt water or fresh water) and have a shell or shell-resembling exterior. Commonly, shellfish can be divided into crustaceans and mollusks [1]. Crustaceans are aquatic animals that have jointed legs, a hard shell, and no backbone, such as crabs, crayfish, lobsters, prawns, and shrimp. Most mollusks have a hinged two-part shell, including clams, green mussels, oysters, and scallops, and various octopuses, snails, and squids. Shellfish has been a popular and favorite food in ancient and modern civilizations.
This has occurred because fresh shellfish is an excellent source of protein and a good source of minerals for humans. Additionally, most shellfishes are low in fat, cholesterol, and sodium [1].

Materials and Methods
The green mussel and crab shells in this study were obtained from the province of Central Java, Indonesia. After cleaning, the green mussel and crab shells were oven-dried at 100 • C for 2 h. The dried green mussel and crab shells were reduced using a crusher machine and sieved with 100 mesh screens. This process produced the green mussel and crab shell powders. The calcination process was carried out on the green mussel and crab shell powders using a Thermolyne Furnace Chamber F6010 at a temperature of 900 • C for 5 h. The green mussel and crab shell powders before and after calcination were characterized using SEM-EDX and XRD methods.
In this study, Precipitated Calcium Carbonate (PCC) was synthesized using calcined green mussel and crab shell powders. Seventeen grams of calcined crab shell powder were mixed with 300 mL of 2M HNO 3 .
To produce a homogeneous mixture, stirring was carried out using a magnetic stirrer at 60 • C for 30 min with a rotation speed of 30 rpm. After 30 min, NH 4 OH was added until the pH of the solution reached 12, which was then filtered using Whatman 42 filter paper. The filtrate was precipitated by slowly flowing CO 2 gas. The resulting milky white precipitate was then washed and filtered with distilled water to a pH of 7 and then dried at Materials 2022, 15, 5712 3 of 15 110 • C for 2 h. The exact process was also carried out to produce PCC made from calcined green mussel shell powder [19].
SEM-EDX (SEOL JSM-6510LA) method was used to determine PCC's morphology and chemical elements made of green mussel and crab shells. The Shimadzu XRD-7000 was also used to analyze PCC's resulting phase and crystallite size from the green mussel and crab shells. Crystalline phases presented in the samples were identified with the help of the Joint Committee on Powder Diffraction Standards (JCPDS). The Rietveld analysis was conducted using the High Score Plus software version 3.0e from PANalytical X'Pert, Cambridge, UK. The description of the diffraction line profiles at Rietveld refinement was achieved using the pseudo-Voigt function. For comparison, SEM-EDX dan XRD tests were held to commercial-grade calcium carbonate gained from Merck. Figure 1 shows a graphic comparison of the XRD test results on green mussel and crab shell powder. Calcite and aragonite crystalline phases were found in powdered green mussel and crab shells. Following JCPDS card number 05-0586, the phase of calcite crystals in mussel and crab shell powder is generally denoted as 2θ: 29 until the pH of the solution reached 12, which was then filtered using Whatman 42 f paper. The filtrate was precipitated by slowly flowing CO2 gas. The resulting milky w precipitate was then washed and filtered with distilled water to a pH of 7 and then d at 110 °C for 2 h. The exact process was also carried out to produce PCC made from cined green mussel shell powder [19].

Characterization of Green Mussel and Crab Shell Powders
SEM-EDX (SEOL JSM-6510LA) method was used to determine PCC's morphol and chemical elements made of green mussel and crab shells. The Shimadzu XRD-7 was also used to analyze PCC's resulting phase and crystallite size from the green mu and crab shells. Crystalline phases presented in the samples were identified with the h of the Joint Committee on Powder Diffraction Standards (JCPDS). The Rietveld anal was conducted using the High Score Plus software version 3.0e from PANalytical X'P Cambridge, UK. The description of the diffraction line profiles at Rietveld refinem was achieved using the pseudo-Voigt function. For comparison, SEM-EDX dan XRD t were held to commercial-grade calcium carbonate gained from Merck. Figure 1 shows a graphic comparison of the XRD test results on green mussel crab shell powder. Calcite and aragonite crystalline phases were found in powde green mussel and crab shells. Following JCPDS card number 05-0586, the phase of ca crystals in mussel and crab shell powder is generally denoted as 2θ: 29.404, 39.399, 43.143. Meanwhile, the aragonite crystal phase is shown at 2θ: 26.312, 31.176, and 33 (JCPDS Card No. 05-0453). In this study, the powder of green mussel shells was do nated by an aragonite crystal phase with a little calcite crystal phase. This contrasts w the crystalline phase found in powdered crab shells dominated by the calcite crystal phase with a little aragonite crystalline phase. The comparison of the weight percentage (%) phase of calcite and aragonite crys on green mussel and crab shell powders is shown in Figure 2. Weight percentage (% calcite and aragonite crystals were generated from a Rietveld analysis carried out us High Score Plus software version 3.0e, as shown in Figure 3a,b. In the green mussel sh powder, the aragonite crystal had a weight percentage (%) and crystallite size of 98 and 59 nm, respectively, with an orthorhombic crystal system. Meanwhile, the cal The comparison of the weight percentage (%) phase of calcite and aragonite crystals on green mussel and crab shell powders is shown in Figure 2. Weight percentage (%) of calcite and aragonite crystals were generated from a Rietveld analysis carried out using High Score Plus software version 3.0e, as shown in Figure 3a,b. In the green mussel shells powder, the aragonite crystal had a weight percentage (%) and crystallite size of 98.6% and 59 nm, respectively, with an orthorhombic crystal system. Meanwhile, the calcite phase in the crab shells powder showed a trigonal crystal system with a weight percentage (%) and crystallite size of 91.8% and 19 nm.   The XRD test results on green mussel and crab shell powders showed the crystal phase of aragonite and calcite dominance, respectively. This is supported by the SEM test phase in the crab shells powder showed a trigonal crystal system with a weight percentage (%) and crystallite size of 91.8% and 19 nm.  The XRD test results on green mussel and crab shell powders showed the crystal phase of aragonite and calcite dominance, respectively. This is supported by the SEM test  The XRD test results on green mussel and crab shell powders showed the crystal phase of aragonite and calcite dominance, respectively. This is supported by the SEM test results shown in Figure 4a,b. Scanning Electron Microscopy (SEM) was used to study the morphology of the synthesis product [20][21][22]. The morphology of the green mussel shell powder is irregular in shape and resembles a small branching rod, which represents the characteristics of aragonite crystals [23]. The common morphologies of aragonite are rod-like, multilayered, pseudohexagonal, needle-like, and dendrite-like [24]. results shown in Figure 4a,b. Scanning Electron Microscopy (SEM) was used to study the morphology of the synthesis product [20][21][22]. The morphology of the green mussel shell powder is irregular in shape and resembles a small branching rod, which represents the characteristics of aragonite crystals [23]. The common morphologies of aragonite are rod-like, multilayered, pseudohexagonal, needle-like, and dendrite-like [24]. Meanwhile, the morphology of the crab shell powder shows a predominance of a nail head and cubic-like structure, which is the characteristic of calcite crystals [25,26].  Energy Dispersive X-ray Analysis (EDX), referred to as EDS or EDAX, is used to determine the mineral content of green mussel and crab shell powder. Table 1 shows a summary of the EDX results of green mussel and crab shell powder, with the major components found to be Ca, C, and O. The EDX test results show the presence of Na in green mussel shells, although in small amounts. Meanwhile, in crab shells, Na, Mg, P, and Zr are found in addition to Ca, C, and O elements.

Characterization of Green Mussel and Crab Shell Powders after Calcination
In this study, the calcination process was carried out on the powder of the green mussel and crab shells using a Thermolyne Furnace Chamber F6010 at a temperature of 900 °C for 5 h. Here, calcination produces Calcined Green Mussel Shells (CMS) and Calcined Crab Shells (CCS). Figure 5 shows a graphic comparison of the results of XRD testing on CMS and CCS with the dominance of the portlandite crystal phase. Portlandite was observed at 2θ: 18.0123, 34.1015, and 50.7966 (JCPDS Card No. . Calcination is generally used to remove organic compounds and impurities in green mussel powder and crab shells. In addition, calcination in green mussel powder and crab shells with a temperature of 900 °C for 5 h to convert the CaCO3 compound into CaO is given in the reaction equation [27]: Meanwhile, the morphology of the crab shell powder shows a predominance of a nail head and cubic-like structure, which is the characteristic of calcite crystals [25,26]. Energy Dispersive X-ray Analysis (EDX), referred to as EDS or EDAX, is used to determine the mineral content of green mussel and crab shell powder. Table 1 shows a summary of the EDX results of green mussel and crab shell powder, with the major components found to be Ca, C, and O. The EDX test results show the presence of Na in green mussel shells, although in small amounts. Meanwhile, in crab shells, Na, Mg, P, and Zr are found in addition to Ca, C, and O elements.

Characterization of Green Mussel and Crab Shell Powders after Calcination
In this study, the calcination process was carried out on the powder of the green mussel and crab shells using a Thermolyne Furnace Chamber F6010 at a temperature of 900 • C for 5 h. Here, calcination produces Calcined Green Mussel Shells (CMS) and Calcined Crab Shells (CCS). Figure 5 shows a graphic comparison of the results of XRD testing on CMS and CCS with the dominance of the portlandite crystal phase. Portlandite was observed at 2θ: 18.0123, 34.1015, and 50.7966 (JCPDS Card No. . Calcination is generally used to remove organic compounds and impurities in green mussel powder and crab shells. In addition, calcination in green mussel powder and crab shells with a temperature of 900 • C for 5 h to convert the CaCO 3 compound into CaO is given in the reaction equation [27]: (1) temperature, the furnace was opened to collect the green mussel and crab shell powde During the cooling process, there was contact between the CaO compound in green mussel and crab shell powders with air containing water vapor so that hydrat could occur, and portlandite or Ca(OH)2 was formed through the reaction equation [2 This is what causes the calcite and aragonite crystals in the powder of green mus and crab shells to transform into calcium hydroxide or portlandite (Ca(OH)2) after ca nation [27,28]. In this study, only a portlandite crystal phase was found in CMS, while in CCS, th was still a small amount of aragonite crystal phase. The weight percentage (%) of portlandite crystal phase was generated from a Rietveld analysis using High Score P software version 3.0e, as shown in Figure 6a,b. In CMS, the portlandite crystal ha weight percentage (%) and crystallite size of 100% and 14 nm, respectively, with trigonal crystal system. Meanwhile, the portlandite phase on CCS shows a trigonal cr tal system with a weight percentage (%) and crystallite size of 99.7% and 12 nm, resp tively. Apart from the portlandite phase, aragonite crystals in CSS were also found wit weight percentage (%) of 0.3%. The XRD test results on the CMS and CCS are suppor by the SEM test results, as shown in Figure 7a,b. As a result of the study, portland crystallized in the shape of imperfect cubic and irregular shapes. The portlandite cryst morphology in this study is consistent with the literature [29,30]. Table 2 shows a summary of the EDX results on CMS and CCS with the ma components found being Ca, C, and O. The EDX test results show that the Na elemen green mussel shells powder before calcination was not found in CMS. Meanwhile CCS, there were still elements of Na, Mg, and P, and Ca, C, and O. After calcination, Ca content in CMS and CCS was more than the Ca content in the green mussel and c shell powders. Meanwhile, the C content in CMS and CCS was less than the C conten the green mussel and crab shell powders. This happened because of the decomposit process, where C bound to O to form CO2 gas and Ca bound to O to form CaO. After 5 h, cooling down to room temperature was held by turning off the furnace without opening it (cooling in the furnace). It was done to avoid the possibility of damage to the furnace walls due to the sudden change of temperature. After reaching room temperature, the furnace was opened to collect the green mussel and crab shell powders.
During the cooling process, there was contact between the CaO compound in the green mussel and crab shell powders with air containing water vapor so that hydration could occur, and portlandite or Ca(OH) 2 was formed through the reaction equation [27]: This is what causes the calcite and aragonite crystals in the powder of green mussel and crab shells to transform into calcium hydroxide or portlandite (Ca(OH) 2 ) after calcination [27,28].
In this study, only a portlandite crystal phase was found in CMS, while in CCS, there was still a small amount of aragonite crystal phase. The weight percentage (%) of the portlandite crystal phase was generated from a Rietveld analysis using High Score Plus software version 3.0e, as shown in Figure 6a,b. In CMS, the portlandite crystal has a weight percentage (%) and crystallite size of 100% and 14 nm, respectively, with the trigonal crystal system. Meanwhile, the portlandite phase on CCS shows a trigonal crystal system with a weight percentage (%) and crystallite size of 99.7% and 12 nm, respectively. Apart from the portlandite phase, aragonite crystals in CSS were also found with a weight percentage (%) of 0.3%. The XRD test results on the CMS and CCS are supported by the SEM test results, as shown in Figure 7a,b. As a result of the study, portlandite crystallized in the shape of imperfect cubic and irregular shapes. The portlandite crystals' morphology in this study is consistent with the literature [29,30].      Table 2 shows a summary of the EDX results on CMS and CCS with the major components found being Ca, C, and O. The EDX test results show that the Na element in green mussel shells powder before calcination was not found in CMS. Meanwhile, in CCS, there were still elements of Na, Mg, and P, and Ca, C, and O. After calcination, the Ca content in CMS and CCS was more than the Ca content in the green mussel and crab shell powders. Meanwhile, the C content in CMS and CCS was less than the C content in the green mussel and crab shell powders. This happened because of the decomposition process, where C bound to O to form CO 2 gas and Ca bound to O to form CaO.

Characterization of Green Mussel and Crab Shell Powders after Precipitation Process
In this study, the Precipitated Calcium Carbonate (PCC) produced from green mussel shells powder was labeled as PMS, whilst the Precipitated Calcium Carbonate (PCC) produced from crab shells powder was labeled as PCS. Figure 8 shows a comparison graph of the XRD test results on PMS and PCS. The crystalline phases of vaterite, calcite, and aragonite were found in PMS and PCS. Following JCPDS card number 13-0192, the phase of the vaterite crystal in PMS and PCS is denoted by 2θ: 24.9011, 27.0705, and 32.7760. In addition to that, there were calcite and aragonite crystal phases that correspond to the JCPDS card numbers 05-0586 and 05-0453. In this study, PMS and PCS were dominated by the vaterite crystalline phase, with a few calcite and aragonite crystalline phases.

Characterization of Green Mussel and Crab Shell Powders after Precipitation Proces
In this study, the Precipitated Calcium Carbonate (PCC) produced from mussel shells powder was labeled as PMS, whilst the Precipitated Calcium Ca (PCC) produced from crab shells powder was labeled as PCS. Figure 8 shows a comparison graph of the XRD test results on PMS and P crystalline phases of vaterite, calcite, and aragonite were found in PMS and P lowing JCPDS card number 13-0192, the phase of the vaterite crystal in PMS and denoted by 2θ: 24.9011, 27.0705, and 32.7760. In addition to that, there were cal aragonite crystal phases that correspond to the JCPDS card numbers 05-0586 and In this study, PMS and PCS were dominated by the vaterite crystalline phase, wi calcite and aragonite crystalline phases. In this study, CMS and CCS with portlandite or Ca(OH)2 as the dominant ph converted to PMS and PCS, which had CaCO3 (vaterite) as the predominant cryst precipitation process using CO2 gas. CaCO3 may be obtained when portlandite (C was exposed to atmospheric carbon dioxide (CO2). The transformation of portla In this study, CMS and CCS with portlandite or Ca(OH) 2 as the dominant phase was converted to PMS and PCS, which had CaCO 3 (vaterite) as the predominant crystal phase precipitation process using CO 2 gas. CaCO 3 may be obtained when portlandite (Ca(OH) 2 ) was exposed to atmospheric carbon dioxide (CO 2 ). The transformation of portlandite to CaCO 3 is under the equation [30]: The formation of the vaterite dominant phase in PMS and PCS can occur due to several things, such as the reaction temperature, Ph, and CO 2 flow rate at the time of carbonation [31,32]. In this study, the carbonation process was carried out at a temperature of 30 • C, pH 12 and a low flow rate of CO 2 . The dominant vaterite phase was caused by an increase in the concentration of CO 2 gas. That happens because the amount of CO 2 gas added will increase the solubility of CO 2 gas in the solution. The increasing solubility of CO 2 gas in the solution causes an increase in the supersaturation of the solution so that vaterite is formed as the dominant crystalline phase [33].
A comparison of the weight percentage (%) phase of vaterite, calcite, and aragonite crystals on PMS and PCS is shown in Figure 9.  The weight percentage (%) was generated from Rietveld analysis conducted using High Score Plus software version 3.0e, as shown in Figure 10a,b. In PMS, the vaterite crystal had a weight percentage (%) and crystallite size of 91.2% and 34 nm, respectively, with the monoclinic crystal system. Meanwhile, the weight percentage (%) on calcite and aragonite was 4.9% and 3.9%, respectively. The vaterite phase of PCS shows the monoclinic crystal system with a weight percentage (%) and crystallite size of 98.9% and 21 nm, respectively. Apart from the vaterite phase, PCS also found aragonite and calcite crystals with a weight percentage (%) of 0.8% and 0.3%, respectively.
The test results on PMS and PCS showed the dominance of the vaterite crystal phase. The results of the SEM tests support these results carried out as shown in Figure 11a,b. As a result of the study, vaterite was crystallized in a spherical shape. The vaterite crystals' morphology in this study is consistent with the literature [34,35]. Table 3 summarizes the EDX results on PMS and PCS, and the major components found are Ca, C, and O. After precipitation, PMS and PCS experience a decrease in Ca content due to the addition of CO 2 gas into the solution. This is proved by the increase in the C and O content. EDX testing showed that only Ca, C, and O were found in PMS. Meanwhile, PCS found N elements other than Ca, C, and O. Element N found in PCS was caused by mixing less homogeneous Ca(OH) 2 with 2M HNO 3 , leading to a less perfect reaction of NO 3 − ions.  The test results on PMS and PCS showed the dominance of the vaterite crystal phase. The results of the SEM tests support these results carried out as shown in Figure 11a,b. As a result of the study, vaterite was crystallized in a spherical shape. The vaterite crystals' morphology in this study is consistent with the literature [34,35]. Table 3 summarizes the EDX results on PMS and PCS, and the major components found are Ca, C, and O. After precipitation, PMS and PCS experience a decrease in Ca content due to the addition of CO2 gas into the solution. This is proved by the increase in the C and O content. EDX testing showed that only Ca, C, and O were found in PMS. Meanwhile, PCS found N elements other than Ca, C, and O. Element N found in PCS was caused by mixing less homogeneous Ca(OH)2 with 2M HNO3, leading to a less perfect reaction of NO3 − ions.    The XRD test results' analysis on commercial CaCO 3 were carried out with Rietveld analysis using High Score Plus software version 3.0e to determine the crystal phase formed, weight percentage (%), and crystallite size, as shown in Figure 12. In commercial CaCO 3 , there was only a calcite phase as a single phase with a trigonal crystal system. The weight percentage (%) and crystallite size of calcite on commercial CaCO 3 were 100% and 15 nm, respectively. The crystallite size for commercial CaCO 3 was 15 nm, while the crystallite size for PMS and PCS were 34 nm and 21 nm, respectively. A comparison of the weight percentage (%) for each crystal phase found in PMS, PCS, and commercial CaCO 3 is shown in Figure 13. In commercial CaCO 3 , there is only calcite as a single phase. Whereas in PMS and PCS, vaterite is the dominant crystalline phase. presence of vaterite (55.20 wt %) and calcite (44.40 wt %) minerals following the carbonation of the calcined powder product [36]. According to , no polymorphic difference was noticed between the PCC product and the CO2 stream in contact with the reaction solution. Importantly, PCC-800 and PCC-900 products include a high concentration of vaterite, a promising biomaterial for use in drug delivery systems [19].  The crystalline phase of calcium carbonate is generally in the form of calcite, aragonite, and vaterite. The different morphological forms are due to different synthesis conditions [37,38]. Calcite has stable structures, mechanical properties, and thermodynamic properties, so it is widely used in biomedical applications [39]. Aragonite is formed in the orthorhombic system and is biocompatible. Aragonite can be broken down, combined, presence of vaterite (55.20 wt %) and calcite (44.40 wt %) minerals following the carbonation of the calcined powder product [36]. According to , no polymorphic difference was noticed between the PCC product and the CO2 stream in contact with the reaction solution. Importantly, PCC-800 and PCC-900 products include a high concentration of vaterite, a promising biomaterial for use in drug delivery systems [19].  The crystalline phase of calcium carbonate is generally in the form of calcite, aragonite, and vaterite. The different morphological forms are due to different synthesis conditions [37,38]. Calcite has stable structures, mechanical properties, and thermodynamic properties, so it is widely used in biomedical applications [39]. Aragonite is formed in the orthorhombic system and is biocompatible. Aragonite can be broken down, combined, In this study, after the carbonation process of the calcined powder was produced from green mussel and crab shells, vaterite was the dominant crystalline phase in the calcium carbonate polymorph. Similar findings were reported by Prihanto et al. (2022). A quantitative XRD Rietveld examination of PCC products generated from green mussel shells revealed the presence of vaterite (55.20 wt %) and calcite (44.40 wt %) minerals following the carbonation of the calcined powder product [36]. According to , no polymorphic difference was noticed between the PCC product and the CO 2 stream in contact with the reaction solution. Importantly, PCC-800 and PCC-900 products include a high concentration of vaterite, a promising biomaterial for use in drug delivery systems [19].
The crystalline phase of calcium carbonate is generally in the form of calcite, aragonite, and vaterite. The different morphological forms are due to different synthesis conditions [37,38]. Calcite has stable structures, mechanical properties, and thermodynamic properties, so it is widely used in biomedical applications [39]. Aragonite is formed in the orthorhombic system and is biocompatible. Aragonite can be broken down, combined, and it can also replace bone. Aragonite is denser than calcite and has also been used for biomedical applications [40,41]. Vaterite has low stability and belongs to the hexagonal crystal system. In contact with water, vaterite can slowly dissolve and recrystallize to a stable form [42]. Due to its nontoxicity, good biocompatibility and affinity, low cost, and ease of large-scale production, vaterite can be used as an ideal nominee for biomedical applications [35,42].The XRD results on commercial CaCO 3 are supported by the SEM test results shown in Figure 14. Commercial CaCO 3 has a morphology with a cube-like shape, which is characteristic of calcite [37]. Meanwhile, PMS and PCS have spherical morphology, which is characteristic of vaterite crystals. The results of this study are the same as the research conducted by Hamester et al. (2012) [43]. The EDX test results show that PMS and commercial CaCO3 only contain Ca, C, and O. Meanwhile, PCS found the presence of N elements besides Ca, C, and O. The results of the research conducted by [44] stated that the chemical composition of commercial Ca-CO3 (calcite) consists of only Ca, C, and O. The common elements such as Ca, C, and O were present in the CaCO3 derived from green mussel shells (PMS), indicating the success of the precipitation method [45]. The comparison of the chemical composition of PMS, PCS, and commercial CaCO3 is shown in Figure 15. In this study, the Ca content in commercial CaCO3 was higher than the Ca content in PMS and PCS. While the C and O content in commercial CaCO3 was lower than the C and O content in PMS and PCS. In this study, the chemical composition of PMS was better than the chemical composition of PCS. Moreover, the chemical composition of PMS was almost the same as the chemical composition of commercial CaCO3. In addition, in PMS, there were no impurities. The EDX test results show that PMS and commercial CaCO 3 only contain Ca, C, and O. Meanwhile, PCS found the presence of N elements besides Ca, C, and O. The results of the research conducted by [44] stated that the chemical composition of commercial CaCO 3 (calcite) consists of only Ca, C, and O. The common elements such as Ca, C, and O were present in the CaCO 3 derived from green mussel shells (PMS), indicating the success of the precipitation method [45]. The comparison of the chemical composition of PMS, PCS, and commercial CaCO 3 is shown in Figure 15. In this study, the Ca content in commercial CaCO 3 was higher than the Ca content in PMS and PCS. While the C and O content in commercial CaCO 3 was lower than the C and O content in PMS and PCS. In this study, the chemical composition of PMS was better than the chemical composition of PCS. Moreover, the chemical composition of PMS was almost the same as the chemical composition of commercial CaCO 3 . In addition, in PMS, there were no impurities.
were present in the CaCO3 derived from green mussel shells (PMS), indicating the success of the precipitation method [45]. The comparison of the chemical composition of PMS, PCS, and commercial CaCO3 is shown in Figure 15. In this study, the Ca content in commercial CaCO3 was higher than the Ca content in PMS and PCS. While the C and O content in commercial CaCO3 was lower than the C and O content in PMS and PCS. In this study, the chemical composition of PMS was better than the chemical composition of PCS. Moreover, the chemical composition of PMS was almost the same as the chemical composition of commercial CaCO3. In addition, in PMS, there were no impurities. Based on the EDX analysis, secondary reactions involving the formation of solid solutions correlating to the existence of Cu and Zn may also exist, especially along the adsorption-precipitation boundary [36]. A similar result was stated by . The EDX analysis revealed that PCC samples derived from green mussel shells (PCC-800 and PCC-900) included primarily Ca, C, and O. Introducing a stream of CO 2 into the solution caused a decrease in the Ca concentration of PCC samples. Conversely, the carbonization process raises the C and O concentration [19]. Prihanto et al. (2022) discovered differences showing that in addition to Ca, C, and O, PCC derived from green mussel shells contained Zn and Cu. Based on the EDX analysis, secondary reactions involving the formation of solid solutions correlating to the existence of Cu and Zn may also exist, especially along the adsorption-precipitation boundary [36].

Conclusions
The use of green mussel and crab shells after industrial and consumption activities has many benefits. It has the potential to be applied in various fields, including in the field of biomaterials. This research shows that the calcination and precipitation processes significantly affect the chemical composition, crystal phase, crystal size, and crystal system of the CaCO 3 obtained. The crystalline phases of aragonite and calcite in green mussel and crab shell powders were converted to Ca(OH) 2 or portlandite by calcination. In addition, calcination can remove organic compounds and reduce the impurity content in green mussel powder and crab shells. The chemical composition of PMS and commercial CaCO 3 shows that they existed only as Ca, C, and O. In contrast, in PCS, it was found that the element N is an impurity in addition to Ca, C, and O. The results of this study show that PMS can be used as a candidate for biomaterials because the chemical composition in PMS is almost the same as the chemical composition in commercial CaCO 3 , where no impurities were found. In addition, the dominance of the vaterite crystal phase in PMS is a distinct advantage for biomaterial applications because it has nontoxicity, good biocompatibility and affinity, a low cost, and ease of large-scale production.
The crystallite size produced in PMS is more prominent than commercial CaCO 3 . The crystallite size of vaterite in PMS is 34 nm, while the crystallite size of calcite in commercial CaCO 3 is 15 nm.