Hydroxyapatite from solid fish waste: A review

Tạp chí Khoa học - Công nghệ Thủy sản Số 4/2014 TRƯỜNG ĐẠI HỌC NHA TRANG • 209 VAÁN ÑEÀ TRAO ÑOÅI HYDROXYAPATITE FROM SOLID FISH WASTE: A REVIEW HYDROXYAPTITE TỪ PHẾ PHẨM RẮN CỦA CÁ: VẤN ĐỀ TRAO ĐỔI Nguyễn Văn Hòa1, Trần Thị Hoàng Quyên2, Trần Quang Ngọc3 Ngày nhận bài: 18/7/2014 ; Ngày phản biện thông qua: 18/10/2014; Ngày duyệt đăng: 01/12/2014 ABSTRACT The waste from fi sh processing after the fi sh have been fi lleted can account for as much as 70% of the total catch weight. About 30% of such waste consists of scale and bone. This waste contains numerous valuable organic and inorganic components such as collagen and hydroxyapatite (HA), which have commercial value for use in manufacturing functional foods, cosmetics, and biomedical products. In particular, HA has been widely used as a biocompatible ceramic but mainly for contact with bone tissue, due to its resemblance to mineral bone. In this study, we have reviewed the characterization and applications of HA extracted from fi sh scale and bone. Moreover, several HA-based highly porous composite materials used for bone tissue engineering were also presented. Finally, a summary of HA preparation, characterization and applications will be given, together with the perspectives on this fi eld of research. Keywords: Hydroxyapatite, fi sh bone, bone engineering, biocomposites TÓM TẮT Phế phẩm từ quá trình chế biến cá có thể lên tới 70% so với tổng khối lượng cá ban đầu. Trong đó, khoảng 30% khối lượng phế phẩm này là da cá và xương cá. Chúng có chứa một số lượng đáng kể chất vô cơ và hữu cơ có giá trị như collagen và hydroxyapatite (HA). Đây là những chất có giá trị thương mại cao trong sản xuất thực phẩm chức năng, mỹ phẩm và các sản phẩm y sinh. Đặc biệt, HA đã được sử dụng rộng rãi dưới dạng gốm sinh học, chủ yếu ở phần tiếp xúc với mô xương, nhờ vào thành phần của nó tương tự như thành phần vô cơ của xương người. Trong bài viết này, chúng tôi giới thiệu tổng quan các nghiên cứu gần đây về điều chế, tính chất và khả năng ứng dụng của HA, đồng thời dự đoán triển vọng của lĩnh vực nghiên cứu này tại Việt Nam. Từ khóa: Hydroxyapatite, xương cá, kỹ thuật xương, vật liệu tổ hợp sinh học I. INTRODUCTION In the fi sh fi lleting process, only about one-third of the whole fi sh is used [4, 5, 10]. Thus, the fi lleting industry generates as much as 70% solid wastes from original fi sh materials. This waste consists of trimmings, fi ns, viscera, head, bone, and skin. Majority of fi sheries byproducts are presently employed to produce fi sh oil, fi shmeal, fertilizer, pet food and fi sh silage [4, 5]. However, most of these recycled products possess low economic value. Recent studies have identifi ed a number of bioactive compounds from remaining fi sh muscle proteins, collagen and gelatin, fi sh oil, fi sh bone, internal organs and shellfi sh and crustacean shells [14, 15, 16]. Generally, a far better profi tability is obtained by producing human consumables and the highest profi tability is currently expected from bioactive compounds. These bioactive compounds can be extracted and purifi ed with technologies varying from simple to complex and such compounds may include preparation and isolation of bioactive peptides, oligosaccharides, fatty acids, enzymes, water-soluble minerals and biopolymers for biotechnological and pharmaceutical applications. Furthermore, some of these bioactive compounds have been identifi ed to possess nutraceutical potentials that are benefi cial to human health promotion [8]. Therefore, the development of new technologies in search of novel bioactive compounds from marine processing by-products will bring more value out of what is today considered a waste and it represents unique challenges and opportunities for the seafood industry. 1 TS. Nguyễn Văn Hòa, 2TS. Trần Thị Hoàng Quyên, 3TS. Trần Quang Ngọc: Khoa Công nghệ thực phẩm - Trường Đại học Nha Trang Tạp chí Khoa học - Công nghệ Thủy sản Số 4/2014 210 • TRƯỜNG ĐẠI HỌC NHA TRANG Hydroxyapatite (HA, Ca10(PO4)6(OH)2) was demonstrated to be an attractive material for biomedical applications mainly in orthopedics and dentistry since it presents a chemical composition close to that of the bone (65 - 70 wt%) and teeth (~ 99 wt%). HA has good biocompatibility, bioactivity, high osteoconductive and non-infl ammatory behavior, and non-immunogenicity properties [21]. It can also easily be processed to matrices with interconnecting pores bone ingrowth. Therefore, many methods have been used to prepare HA such as ultrasonic irradiation, emulsion liquid membrane, microwave-mediated metathesis, expeditious microwave irradiation, RF thermal plasma, chemical precipitation, microemulsion and hydrolysis [20]. However, these synthesis processes might be either complicated or biologically unsafe, therefore, recently natural HA bioceramics has been extracted by normal calcination of some biowastes such as fi sh bones, bovine bones, teeth and bones of pig, etc. Moreover, extraction of HA from bio-waste is both economically and environmentally preferable. In the study, we would like to present an overview of HA preparation, characterization and the perspectives of HA applications. II. PREPARATION METHODS AND CHARACTERIZATION OF HA FROM FISH SCALE AND BONE So far, a variety of methods for synthesizing HA have been developed. The methods involve various types of known chemical synthesis routes, including hydrothermal [30], liquid membrane [13], precipitation [26], radio frequency thermal plasma [29], ultrasonic precipitation [2], reverse micro emulsion [11], sol–gel [9] and polymer-assisted methods [27]. On the one hand, most of the procedures for synthesizing HA are biologically hazardous and they have a complicated process. These synthetic procedures have also led to the formation of non-stoichiometric products [17]. On the other hand, synthetic HA with a Ca/P ratio near 1.67 is stable when sintered in dry or wet air below 12000C, but HA often loses its OH groups gradually at higher temperature. It can be transformed to oxyapatite such as Ca10O(PO4)6 or Ca(PO4)6O, and after that these oxyapatites are normally dissociates into the products α-Ca3(PO4)2, Ca2P2O7 and Ca4P2O9 at 1450 0C [31]. Moreover, it is diffi cult to prepare HA crystals from aqueous solutions due to the high chemical affi nity of the materials to some ions, the complex nature of the calcium phosphate system, and the roles of kinetic parameters [17]. Recently, HA has been prepared from various bio-waste, including corals [7, 19], cuttlefi sh shells [23], fi sh scales and bones, porcine teeth and bones, and bovine bones [1, 25]. In particular, extraction of HA from these sources is an interesting process. It is not only because of some superior characteristics of the extracted HA, but also due to the environmental benefi ts of waste recovery. Moreover, compared with HA produced by synthetic methods as described above, HA extracted from bio-waste is a biologically safe (i.e., no chemicals are often required) and potentially lucrative process, especially given the growing global demand for HA bioceramics. Nowadays, a variety of techniques for producing HA from bio-wastes with appreciable quantities have been developed, especially for fi sh bone and fi sh scale. Generally, preparation of HA using these biowastes usually involves a few hours annealing during which the organic materials in the bone get removed, leaving pure HA as the residue (fi gure 1). There are now fi ve primary methods including calcination, enzymatic hydrolysis, plasma processing, subcritical water processing, and hydrothermal hydrolysis, the most widely applied technique of which is the calcination of by-products at high temperature due to its ease of large scale production and relatively low cost. Figure 1. Preparation of HA via extraction of minerals from some biowastes. Copyright Elsevier and reproduced with permission [24] Tạp chí Khoa học - Công nghệ Thủy sản Số 4/2014 TRƯỜNG ĐẠI HỌC NHA TRANG • 211 As indicated in Table 1, a few studies have recently reported on the utilization of fi sh scales and bones to produce HA. For example, Ozawa and Suzuki [22] prepared HA with microstructural particles through treatment. They reported that fi sh bone heated at temperatures <1300°C maintains a porous structure, with a sintered wall and a major crystalline phase of hydroxyapatite (fi gure 2). Recently, Huang et al. [12] fabricated nano-sized HA particles with Ca/P ratio of 1.76 from fi sh scale using enzymatic hydrolysis method (fi gure 3). A tilapia fi sh-scale was hydrolyzed using protease N for 2.5 h, and fl avourzyme for 0.5 h at an optimal pH and temperature, followed by stirring in a boiling water bath for 10 mins and subsequent sintering at 800◦C. Figure 2. SEM micrograph of fi sh-bone ceramic heated at (a) 800° and (b) 1000°C for 1 h in air Copyright John Wiley and Sons with permission [22] Table 1. Recent progress in the synthesis of HAp from fi sh scale and bone Authors/ Year Brief description Characteristics of powder Ref. Huang et al. 2011 Hydrolysis of a tilapia fi sh-scale using protease N for 2.5 h, and fl avourzyme for 0.5 h at an optimal pH and temperature, followed by stirring in a boiling water bath for 10 min and subsequent sintering at 8000C Agglomerated HA nanoparticles [12] Mondal et al. 2010 Treatment of Labeo rohita fi sh-scale using HCl and then NaOH, followed by stirring in a boiling water bath for 20 min and subsequent calcination at various temperatures (800 - 14000C) HA powder of submicrometric size [18] Coelho et al. 2006 Thermal treatment of bones originated form Brazilian river fi sh at 9000C for 4 - 12 h, followed by milling at 300 rpm for 2 - 16 h using a high-energy milling Irregular stoichiometric HA nanoparticles of various sizes [6] Ozawa and Suzuki 2002 Thermal treatment of Japanese sea bream fi sh bone at 600 - 13000C HA of macroporous structure [22] Kim et al. 1997 Thermal treatment of Tuna bone at 850 ◦C for 2 h HA of microporous structure [3, 28] Figure 3. SEM images of the HA powder (a), sintered HA (b) and EDS analysis of HA powder. Copyright Elsevier with permission [12] More recently, Kim et al. [28] isolated successfully pure natural nano-HA from tuna bone by employing the alkaline hydrolysis and thermal calcination methods. The thermal calcination method produces good crystallinity with dimensions 0.3 - 1.0 μm at 2500C for 5h, whereas the alkaline hydrolysis method produces nanostructured HA crystals with 17 - 71 nm length and 5 -10 nm width at 9000C for 5h. Tạp chí Khoa học - Công nghệ Thủy sản Số 4/2014 212 • TRƯỜNG ĐẠI HỌC NHA TRANG Figure 4. SEM of the nanostructured HA powder obtained from the bones of Brazilian river fi sh after milling for (a) 2h, (b) 4h, (c) 8h, and (d) 16h. Copyright AIP Publishing LLC with permission [6] The elemental composition or purity of obtained HA is an important key for further applications. Raw fi sh bone contains large amount of organics, and even after heat treatment up to 6000C, it has partially shown grayish or black color with carbon residue (0.5%) [22]. However, powder can become completely white by heating at 9000C and reduced to powder using a high-energy ball mill, in order to obtain a natural nanostructured HA powder (Figure 4) [6]. The Ca:P ratio was not be changed by high temperature, while the main natural impurities such as sodium, magnesium, and potassium were slightly lost by heating at 13000C [22]. The chemical analyses in the previous studies suggest good quality of fi sh-bone-extracted HA as a mineral resource for further application [3, 6, 18, 22, 28]. III. APPLICATIONS OF HA PREPARED FROM FISH SCALE AND BONE Today the properties of HA extracted from fi sh scale and bones have been taken into investigation with the purpose of applying HA in many fi elds, especially in orthopedics and dentistry. For example, Huang et al. [12] evaluated the biocompatibility of HA crystals from fi sh scale by cytotoxicity and cell proliferation with human osteoblast-like cell MG-63 [12]. Under the osteogenic-inductive cultural condition, HA promoted osteogenic differentiation and mineralization of MG63 cells. As indicated by the in vitro mineralization, MG-63 cells can differentiate into bone forming cells (Figure 5). However, Kim et al. [28] checked the cytotoxicity and cell proliferation of MG-63 human osteosarcoma cell on micro and nanosized HA prepared from Thunnus obesus bone at different concentrations and days. They found that MG-63 cells grew rapidly on the control plate, whereas limited and lesser cell growth was observed on treated HA particles. The cell proliferation rate on HA was slower than that under the control. Moreover, the experiments demonstrated that micro and nanoparticles had similar cytotoxicity on the MG-63 cell line. Figure 5. Von Kossa stains of MG63 cells cocultured with (A) 20mg/ml HA extracted from fi sh scale particles and (B) 20mg/ml HA from Sigma particles. Mineralized matrix formation is the most reliable indicator of the osteoinductive capacity of materials. Copyright Elsevier with permission [12] Tạp chí Khoa học - Công nghệ Thủy sản Số 4/2014 TRƯỜNG ĐẠI HỌC NHA TRANG • 213 IV. CONCLUSIONS AND PERSPECTIVES Besides the synthetic HA, a variety of HA extracted from fi sh scales and bones was also developed for potential applications. The HA from fi sh scales and bones has the size from micro to nano. Their composition is suitable for medical and biotechnological applications. However, when considered for use in daily life, the health risk associated with HA-based materials needs to be evaluated through the investigation of the toxicity and biocompatibility of these materials. 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