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Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites
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Original Article
Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites Wendell Bruno Almeida Bezerra ∗ , Sergio Neves Monteiro, Michelle Souza Oliveira, Fernanda Santos da Luz, Fabio da Costa Garcia Filho, Luana Cristyne da Cruz Demosthenes, Ulisses Oliveira Costa Military Institute of Engineering — IME, Department of Materials Science, Prac¸a General Tibúrcio, 80, Praia Vermelha, CEP 22290-270, Rio de Janeiro, RJ, Brazil
a r t i c l e
i n f o
a b s t r a c t
Article history:
The arapaima is a large Amazonian freshwater fish and an example of a natural protective
Received 27 April 2019
system against predators such as the piranha fish. In this work, both the plain scales and a 30
Accepted 13 January 2020
vol% of arapaima scales reinforced epoxy composite were characterized for their structure,
Available online xxx
composition and morphology. The characterization was performed by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS) and Fourier transform infrared spec-
Keywords:
troscopy (FTIR). The SEM images showed that the surface morphology of the scales was not
Arapaima scales
altered by the flattening process applied before composite manufacturing. The EDS results
Epoxy composite
confirm that the percentage of calcium is higher in the scale outer layer, which also shows
SEM
the presence of phosphorous. The evidence of collagens in the plain scales as well as the
EDS
presence of hydroxyl groups and absorption bands related to the epoxy group in the com-
FTIR
posites were revealed by FTIR. Mechanical bend tests disclosed the toughening contribution
Characterization
of arapaima scales to the composite epoxy matrix. Nanoindentation testing confirms the higher hardness of the scale outer layer associated with calcium participation. These experimental results provide, for the first time, an initial view of the arapaima scales potential for use as reinforcement in novel polymer composites. © 2020 Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
1.
Introduction
The bio-inspiration ideas for novel solutions to engineering applications usually come from many aspects of nature, specially from animals. Indeed, animals have developed and improved their protective systems over millions of years
through a process of convergent evolution. The dermal scales of fish constitute a protective system that exemplifies this evolution process [1–4]. The fish scales possess many features suitable to protective systems: they are thin, flexible, light weight, and can resist puncture from attacks by predators, or from collision with obstacles [5–7]. Many recent studies have focused on the structure and mechanical properties of indi-
∗
Corresponding author. E-mails: [email protected] (W.B. Bezerra), [email protected] (S.N. Monteiro), [email protected] (M.S. Oliveira), [email protected] (F.S. Luz), [email protected] (F.C. Garcia Filho), [email protected] (L.C. Demosthenes), [email protected] (U.O. Costa). https://doi.org/10.1016/j.jmrt.2020.01.051 2238-7854/© 2020 Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/ licenses/by-nc-nd/4.0/). Please cite this article in press as: Bezerra WB, et al. Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites. J Mater Res Technol. 2020. https://doi.org/10.1016/j.jmrt.2020.01.051
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vidual fish scales. In particular, components such as type I collagen provide the basis of the tensile strength and toughness of such fish scales [8]. Tensile tests have revealed the scale’s capability to absorb large amounts of deformation and energy, associated with progressive failure mechanisms [9–11]. Some specific articles have been devoted to the study of the structure and properties of arapaima scales [12–18]. The arapaima (Arapaima gigas), also known as pirarucu in Brazil, is a fresh water fish weighing up to 200 kg with large hard scales. It is considered a referential example of a natural protective system against predators in the Amazonian rivers and lakes, particularly the famous piranha fish [14]. In recent years, Torres et al. [12] studied the structure and composition of the scales through X-ray diffraction and FTIR analysis, confirming the presence of hydroxyapatite and collagen, which is by far the most common and important structural component of arapaima scales [8]. Sherman et al. [18] carried out comparative studies of the structures found in the arapaima scales with those observed in other species of fish. Torres et al. [15] analyzed the influence of the presence of different water contents on the thermal transition of the scales through differential calorimetry analysis. Arola et al. [7] studied the structure, composition and mechanical behavior of the limiting layer of three different fish, namely Arapaima gigas, Megalops atlanticus and Cyprinus carpio.Despite these various articles on the structure and composition of fish scales, there is only a limited number of studies on the application of the fish protective systems in composite materials [19,20]. Satapathy et al. [19] studied the utilization of Labeo rohita scales as reinforcement in epoxy matrix composites, obtaining better wear performance when compared to neat epoxy resin. Gopi et al. [20] analyzed the application of fish scales in vynil ester matrix composites, observing improved mechanical properties with 10 wt.% scales reinforced composites. However, the arapaima scales have not yet been considered for composite reinforcement. Thus, the present investigation aims to explore, for the first time, the use of arapaima scales in polymer composites. The main objective of the present investigation is to study the structure and morphology of the arapaima scales, before and after a flattening process. The chemical structure of both the plain scales and their epoxy composites was investigated by means of FTIR, while morphological and qualitative analysis by SEM/EDS. Hardness changes along the scale thickness was determined by nanoindentation. Mechanical properties of arapaima scales composites were evaluated by bend tests.
2.
Materials and methods
2.1.
Materials
Arapaima scales were provided in their natural state, without any type of treatment, by the company Amazon Forest, Brazil. Fig. 1 shows one of the curved scales in the as received state. The polymer used as composite matrix was a commercial epoxy resin diglycidyl of bisphenol-A (DGEBA) cured with triethylene tetramine (TETA), which were supplied by Epoxyfiber, Brazil.
2.2.
Fabrication of composites
Before manufacturing the composites, the curved scales were flattened according to the process illustrated in Fig. 2. First, they were placed in a water bath for 24 h, followed by drying under pressure in a stove, for about three hours at 80 ◦ C, which result in perfectly flat scales. After the flattening process, the scales were layered together with the resin and hardener mixture, using the stoichiometric phr of 13 parts of hardener to 100 parts of resin. The layers were set in a metal mold with dimensions of 150 × 120 × 11.9 mm and put under pressure for 24 h for proper curing at room temperature (RT ∼ 25◦ ). The flattened scales, the composite, and the metal mold are presented in Fig. 3.
2.3.
Characterization
The surface morphology of the samples was studied by means of scanning electron microscopy (SEM), adapted with energy dispersive X-ray spectroscopy (EDS), in a model Quanta FEG FEI equipment with a TM3000 Hitachi detector. In order to enhance the conductivity of the samples, a thin film of nickel was vacuum-evaporated, using a model LEICA EM ACE600 equipment. Fourier transform infrared (FTIR) analysis was conducted in a model IR Prestige 21-FTIR Shimadzu equipment. Samples of both plain arapaima scale and flattened scale reinforced epoxy composites were first milled and sieved for 20 mesh, before mixed with KBr. The mixture was then pressed to produce a film suitable for the analysis.Mechanical properties of the arapaima scale incorporated epoxy composites were evaluated by 3-points bend tests in a model DL10000 EMIC machine, Brazil, with 10 tonnes of capacity, operating as per ASTM D790 [21] with 2 mm/min of loading velocity at RT. Nanoindentation was performed across the thickness of arapaima scale using procedures described by Brito et al. [22]. Ten indentations were performed at the distinct outer, inner and intermediate thickness layers of the scale.
3.
Results and discussion
In Fig. 1 some morphological characteristics of the arapaima scales can be seen with the naked eye. The scales have two distinct parts. First, a thicker and darker exposed region (≈2 mm). Second, a thinner and lighter embedded region (≈1 mm) [12–14]. While swimming, the dark region is exposed to the water while the light embedded region is covered by other scales in the fish skin. The external region of the darker region is referred in literature as the limiting layer [5,7,14]. The arrow in Fig. 1a indicates the direction of growth of the scales, which is perpendicular to the scale long axis, forming a ridge structure. This structure is oriented along the semicircle formed around the vertice point shown in this figure [12,14]. The arapaima scales, as other elasmoid scales, have a structure composed of three layers across the thickness: the limiting layer, the external (outer) layer and the internal (inner) layer. The outer and inner layers differ basically
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Fig. 1 – Arapaima gigas scale in the as-received state showing the (a) front view; (b) side view; (c) rear and (d) front views of the curved arapaima scales.
Fig. 2 – Hierarchical representation of the flattening process to which the curved scales where submitted before composite manufacture.
Please cite this article in press as: Bezerra WB, et al. Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites. J Mater Res Technol. 2020. https://doi.org/10.1016/j.jmrt.2020.01.051
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Fig. 3 – (a) A layer of arranged arapaima scales after submitted to flattening process; (b) Epoxy matrix composite reinforced with 30 vol% of arapaima scales and (c) Metallic mold used in the manufacturing of composite plates.
by their mineral content. The outer layer presents a higher degree of mineralization [1,5,12–16], while the inner is basically composed of collagen [8]. An intermediate region may be considered in between the outer and inner layers. Fig. 4a and b are, respectively, SEM images of the exposed and embedded regions of the as-received scales. On the other hand, Fig. 4c and d are, respectively, images of the exposed and embedded scale regions after the flattening process. From Fig. 4a and c one can observe the transition from the ridge structure in the embedded part to the corrugated zone in the exposed part, which presents ridges with larger spacings in between. In these figures, one can also see the presence of cracks located between the ridges in the exposed part and across the ridges in the embedded region. This cracking formation may have occurred during the drying step of the flattening process.
From Fig. 4b and d, in the ridge structure of the embedded region, it is possible to see the presence of cracks mostly in the veins for the as-received scales. The flattened scales present the same type of aforementioned cracks with addition of some cracks across the ridges. The presence of cracks on the scales surface was also observed in previous studies and was associated with the natural drying process of the scales, which produce shrinking stresses [13,14]. Fig. 5a and b show SEM images of the fracture surface of the 30 vol% arapaima scale reinforced epoxy matrix composite. In the left side of Fig. 5a, one should notice the lamellar structure of the inner part of the scales. The lamellae present different contrast due to their different orientation forming a Bouligand-type structure, in agreement with previous works [13,14,23]. In Fig. 5a, it is also noted the presence of a crack running through the inner layer of the scale, which might have been nucleated in one of the ridges of the scales. In Fig. 5b
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Fig. 4 – (a) Transition region from the embedded part to the exposed part of as received scale; (b) Embedded part of an as received scale showing the ridge structure; (c) Transition region from the embedded part to the exposed part of a scale submitted to the flattening process and (d) Embedded part of a flattened scale showing the ridge structure with some cracks.
Fig. 5 – SEM images of the fracture surface of the 30 vol% arapaima scales reinforced composites evidencing: (a) the epoxy matrix and the scale and (b) the different layers of the scales internal structure.
it is possible to notice the interface region showing a good adhesion between the scale and the matrix phase. EDS scans where performed to assess the qualitative chemical composition of the scales. Fig. 6 shows the EDS results. In this figure, it is possible to see from the mapping of calcium (Ca) the presence of a higher degree of mineralization
in the limiting layer, where there is a higher concentration of Ca, associated with hydroxyapatite. Phosphorous (P) from the hydroxyapatite is also reported to exist in arapaima scales [13,14]. Fig. 7 shows the wavelength transmittance graphs obtained by the FTIR analysis for the epoxy composite reinforced with
Please cite this article in press as: Bezerra WB, et al. Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites. J Mater Res Technol. 2020. https://doi.org/10.1016/j.jmrt.2020.01.051
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Fig. 6 – Energy dispersive X-ray spectroscopy (EDS) results for the arapaima scales: (a) SEM image of the analyzed region; (b) elemental mapping obtained and (c) graphs for the elemental mapping.
Fig. 7 – FTIR spectrum of the 30 vol% arapaima scales reinforced composite.
Fig. 8 – Comparison between the FTIR spectra of both the plain arapaima scales and the 30 vol% arapaima scales reinforced composite.
arapaima scales. In this figure, several bands associated with the scales as well as bands associated with resin may be noticed. When compared with the results obtained by Torres et al. [12] and Lin et al. [13], a small variation of transmittance intensity with respect to the plain scale and the composite could be detected. This is basically due to the interference caused by the resin, which is seen in Fig. 8. However, all transmittance bands related to the arapaima scales could be found and are presented in Table 1.
In this table, bands 2, 3 and 5 are associated with organic components of the scale that are related to type I collagen [8], while bands 4, 6 and 7 are related to the inorganic part of the scale, which is composed basically of hydroxyapatite [12,13]. In addition, the identification of the isolated bands from studies related to the arapaima scales indicates that there was no chemical interaction between the reinforcement and the matrix phases [9,12,13]. Table 2 shows a comparison between the bands obtained in previous articles and the ones in the
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Table 1 – FTIR absortion peaks observed and their related components. Peak
Wavelength (cm− 1 )
Related peak
1 2 3 4 5 6 7
3425.6 1631.1 1558.5 1458.2 1265.3 1033.9 864.1
Amide I Amide II Phosphate groups Amide III Phosphate groups Carbonate groups
Table 2 – Comparison of FTIR wavelengths of amide I, II and III.
Ikoma et al. [9] Torres et al. [12] Lin et al. [13] Present work
Amide I (cm− 1 )
Amide II (cm− 1 )
Amide III (cm− 1 )
1657 1662 1637 1631
1520 1560 1546 1558
1447 1242 1239 1265
Table 3 – Flexural properties by 3-points bend tests of 30 vol% arapaima scale/epoxy composite and plain DGEBA/TETA epoxy specimens. 3-points bend test specimens
Flexural Strength (MPa)
Flexural Modulus (GPa)
Maximum Deflection (mm)
Plain epoxy 30 vol% arapaima scales/epoxy
71.6 ± 25.5 43.6 ± 10.9
2.82 ± 0.49 4.50 ± 0.70
10.8 ± 4.0 5.6 ± 1.5
Table 4 – Nanoindentation hardness across the thickness of an arapaima scale. Hardness (GPa) across the thickness
Arapaima scale
Outer layer
Intermediate layer
Inner layer
0.323 ± 0.082
0.268 ± 0.011
0.170 ± 0.108
present work. These results indicate similar chemical interaction in both scales and epoxy reinforced with scales. Table 3 presents the flexural properties of the 30 vol% arapaima flattened scales incorporated into epoxy matrix composite, obtained in 3-points bend tests. Corresponding flexural strength and modulus as well as maximum deflection before rupture for plain DGEBA/TETA epoxy are also shown in Table 3. The results in this table reveal that, within the standard deviation (SD), strength and deflection are comparable for both plain epoxy and 30 vol% arapaima scale composite. On the other hand, the flexural modulus is sensibly increased with incorporation of arapaima scales into epoxy matrix. This characterizes a toughening effect, which can be attributed to the stiffer nature of the arapaima scales [13,17], as well as their collagen content [8]. Table 4 depicts the average and SD values for nanoindentation at the outer, intermediate and inner layers, Fig. 5a, of an arapaima scale. In this table one should notice the comparatively greater hardness at the outer layer, which is an expected result in view of the higher degree of mineraliza-
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tion due to calcium content in Fig. 6, as found and discussed in most works on characterization of arapaima scales [12–18]. The combination of a hard outer layer and a collagen-based [8] stronger inner layer contributes to reinforce epoxy composites as presented in Table 3.
4.
Conclusions
• The surface of the arapaima scales presented cracks located in the veins between the ridges in both as received scales and flattened scales, which were associated with the shrinkage stresses induced during the natural drying process. • Arapaima scales after the flattening process showed surface cracks going through the ridge structure, that could be related to the drying step of the flattening process. • The fracture surface of the arapaima scale epoxy composite revealed the lamellar structure of the scales. These also confirmed the physical interaction in the interface of the composite. • EDS results showed a high content of calcium in the external layer, which correlated to the higher degree of mineralization present in this outer layer and leads to greater hardness. Phosphorous peaks were also detected and related to the composition of the hydroxyapatite present in the scales. • The FTIR results were obtained for both the plain scales and the arapaima scales reinforced epoxy composites. It was possible to observe in corresponding graphs the characteristic bands associated with type I collagen as well as those constituents of the hydroxyapatite. Despite the variation in the transmittance intensity of the material, there was no chemical interaction between the composite phases. • Flexural properties disclosed a toughening effect, in terms of sensibly increase in the elastic modulus, caused by the incorporation of 30 vol% of arapaima scales into epoxy matrix. This novel cost effective composite material might be included among those based on discarded natural waste (scales in fish market) with potential engineering applications. • The improved composite toughening is supported by the combination of arapaima scale calcium-based hard outer layer with collagen-based stronger inner layer.
Conflicts of interest The authors declare no conflicts of interest.
Acknowledgements The authors thank the support to this investigation by the Brazilian agencies: CNPq, CAPES and FAPERJ.
Appendix A. Supplementary data Supplementary material related to this article can be found, in the online version, at doi:https://doi.org/10.1016/j. jmrt.2020.01.051.
Please cite this article in press as: Bezerra WB, et al. Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites. J Mater Res Technol. 2020. https://doi.org/10.1016/j.jmrt.2020.01.051
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references [13] [1] Bruet BJ, Song J, Boyce MC, Ortiz C. Materials design principles of ancient fish armour. Nat Mater 2008;7(9):748–56. [2] Yang W, Chen IH, Gludovatz B, Zimmermann EA, Ritchie RO, Meyers MA. Natural flexible dermal armor. Adv Mater 2013;25:31–48. [3] Ebenstein D, Calderon C, Troncoso OP, Torres FG. Characterization of dermal plates from armored catfish Pterygoplichthys pardalis reveals sandwich-like nanocomposite structure. J Mech Behav Biomed Mater 2015;45:175–82. [4] Murcia S, Lavoie E, Linley T, Devaraj A, Ossa EA, Arola D. The natural armors of fish: a comparison of the lamination pattern and structure of scales. J Mech Behav Biomed Mater 2017;73:17–27. [5] Sire J-Y, Akimenko M-A. Scale development in fish: a review, with description of sonic hedgehog (shh) expression in the zebrafish (Danio rerio). Int J Dev Biol 2004;48:233–47. [6] Yang W, Gludovatz B, Zimmermann EA, Bale HA, Ritchie RO, Meyers MA. Structure and fracture resistance of alligator gar (Atractosteus spatula) armored fish scale. Acta Biomater 2013;9(4):5876–89. [7] Arola D, Murcia S, Stossel M, Pahuja R, Linley T, Devaraj A, et al. The limiting layer of fish scales: structure and properties. Acta Biomater 2018;67:319–30. [8] Yang W, Meyers MA, Ritchie RO. Structure architecture with toughening mechanisms in nature: a review of the materials science of type-I collagenous materials. Prog Mater Sci 2019;103:425–83. [9] Ikoma T, Kobayashi H, Tanaka J, Walsh D, Mann S. Microstructure, mechanical, and biomimetic properties of fish scales from Pagrus major. J Struct Biol 2003;142:327–33. [10] Zhu D, Ortega CF, Motamedi R, Szewciw L, Vernerey F, Barthelat F. Structure and mechanical performance of a “modern” fish scale. Adv Eng Mater 2012;14:B185–B194. [11] Garrano AMC, La Rosa G, Zhang D, Niu LN, Tay FR, Majd H, et al. On the mechanical behavior of scales from Cyprinus carpio. J Mech Behav Biomed Mater 2012;7:17–29. ´ [12] Torres FG, Troncoso OP, Nakamatsu J, Grande CJ, Gomez CM. Characterization of the nanocomposite laminate structure
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Please cite this article in press as: Bezerra WB, et al. Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites. J Mater Res Technol. 2020. https://doi.org/10.1016/j.jmrt.2020.01.051
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Original Article
Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites Wendell Bruno Almeida Bezerra ∗ , Sergio Neves Monteiro, Michelle Souza Oliveira, Fernanda Santos da Luz, Fabio da Costa Garcia Filho, Luana Cristyne da Cruz Demosthenes, Ulisses Oliveira Costa Military Institute of Engineering — IME, Department of Materials Science, Prac¸a General Tibúrcio, 80, Praia Vermelha, CEP 22290-270, Rio de Janeiro, RJ, Brazil
a r t i c l e
i n f o
a b s t r a c t
Article history:
The arapaima is a large Amazonian freshwater fish and an example of a natural protective
Received 27 April 2019
system against predators such as the piranha fish. In this work, both the plain scales and a 30
Accepted 13 January 2020
vol% of arapaima scales reinforced epoxy composite were characterized for their structure,
Available online xxx
composition and morphology. The characterization was performed by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS) and Fourier transform infrared spec-
Keywords:
troscopy (FTIR). The SEM images showed that the surface morphology of the scales was not
Arapaima scales
altered by the flattening process applied before composite manufacturing. The EDS results
Epoxy composite
confirm that the percentage of calcium is higher in the scale outer layer, which also shows
SEM
the presence of phosphorous. The evidence of collagens in the plain scales as well as the
EDS
presence of hydroxyl groups and absorption bands related to the epoxy group in the com-
FTIR
posites were revealed by FTIR. Mechanical bend tests disclosed the toughening contribution
Characterization
of arapaima scales to the composite epoxy matrix. Nanoindentation testing confirms the higher hardness of the scale outer layer associated with calcium participation. These experimental results provide, for the first time, an initial view of the arapaima scales potential for use as reinforcement in novel polymer composites. © 2020 Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
1.
Introduction
The bio-inspiration ideas for novel solutions to engineering applications usually come from many aspects of nature, specially from animals. Indeed, animals have developed and improved their protective systems over millions of years
through a process of convergent evolution. The dermal scales of fish constitute a protective system that exemplifies this evolution process [1–4]. The fish scales possess many features suitable to protective systems: they are thin, flexible, light weight, and can resist puncture from attacks by predators, or from collision with obstacles [5–7]. Many recent studies have focused on the structure and mechanical properties of indi-
∗
Corresponding author. E-mails: [email protected] (W.B. Bezerra), [email protected] (S.N. Monteiro), [email protected] (M.S. Oliveira), [email protected] (F.S. Luz), [email protected] (F.C. Garcia Filho), [email protected] (L.C. Demosthenes), [email protected] (U.O. Costa). https://doi.org/10.1016/j.jmrt.2020.01.051 2238-7854/© 2020 Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/ licenses/by-nc-nd/4.0/). Please cite this article in press as: Bezerra WB, et al. Processing and characterization of Arapaima gigas scales and their reinforced epoxy composites. J Mater Res Technol. 2020. https://doi.org/10.1016/j.jmrt.2020.01.051
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vidual fish scales. In particular, components such as type I collagen provide the basis of the tensile strength and toughness of such fish scales [8]. Tensile tests have revealed the scale’s capability to absorb large amounts of deformation and energy, associated with progressive failure mechanisms [9–11]. Some specific articles have been devoted to the study of the structure and properties of arapaima scales [12–18]. The arapaima (Arapaima gigas), also known as pirarucu in Brazil, is a fresh water fish weighing up to 200 kg with large hard scales. It is considered a referential example of a natural protective system against predators in the Amazonian rivers and lakes, particularly the famous piranha fish [14]. In recent years, Torres et al. [12] studied the structure and composition of the scales through X-ray diffraction and FTIR analysis, confirming the presence of hydroxyapatite and collagen, which is by far the most common and important structural component of arapaima scales [8]. Sherman et al. [18] carried out comparative studies of the structures found in the arapaima scales with those observed in other species of fish. Torres et al. [15] analyzed the influence of the presence of different water contents on the thermal transition of the scales through differential calorimetry analysis. Arola et al. [7] studied the structure, composition and mechanical behavior of the limiting layer of three different fish, namely Arapaima gigas, Megalops atlanticus and Cyprinus carpio.Despite these various articles on the structure and composition of fish scales, there is only a limited number of studies on the application of the fish protective systems in composite materials [19,20]. Satapathy et al. [19] studied the utilization of Labeo rohita scales as reinforcement in epoxy matrix composites, obtaining better wear performance when compared to neat epoxy resin. Gopi et al. [20] analyzed the application of fish scales in vynil ester matrix composites, observing improved mechanical properties with 10 wt.% scales reinforced composites. However, the arapaima scales have not yet been considered for composite reinforcement. Thus, the present investigation aims to explore, for the first time, the use of arapaima scales in polymer composites. The main objective of the present investigation is to study the structure and morphology of the arapaima scales, before and after a flattening process. The chemical structure of both the plain scales and their epoxy composites was investigated by means of FTIR, while morphological and qualitative analysis by SEM/EDS. Hardness changes along the scale thickness was determined by nanoindentation. Mechanical properties of arapaima scales composites were evaluated by bend tests.
2.
Materials and methods
2.1.
Materials
Arapaima scales were provided in their natural state, without any type of treatment, by the company Amazon Forest, Brazil. Fig. 1 shows one of the curved scales in the as received state. The polymer used as composite matrix was a commercial epoxy resin diglycidyl of bisphenol-A (DGEBA) cured with triethylene tetramine (TETA), which were supplied by Epoxyfiber, Brazil.
2.2.
Fabrication of composites
Before manufacturing the composites, the curved scales were flattened according to the process illustrated in Fig. 2. First, they were placed in a water bath for 24 h, followed by drying under pressure in a stove, for about three hours at 80 ◦ C, which result in perfectly flat scales. After the flattening process, the scales were layered together with the resin and hardener mixture, using the stoichiometric phr of 13 parts of hardener to 100 parts of resin. The layers were set in a metal mold with dimensions of 150 × 120 × 11.9 mm and put under pressure for 24 h for proper curing at room temperature (RT ∼ 25◦ ). The flattened scales, the composite, and the metal mold are presented in Fig. 3.
2.3.
Characterization
The surface morphology of the samples was studied by means of scanning electron microscopy (SEM), adapted with energy dispersive X-ray spectroscopy (EDS), in a model Quanta FEG FEI equipment with a TM3000 Hitachi detector. In order to enhance the conductivity of the samples, a thin film of nickel was vacuum-evaporated, using a model LEICA EM ACE600 equipment. Fourier transform infrared (FTIR) analysis was conducted in a model IR Prestige 21-FTIR Shimadzu equipment. Samples of both plain arapaima scale and flattened scale reinforced epoxy composites were first milled and sieved for 20 mesh, before mixed with KBr. The mixture was then pressed to produce a film suitable for the analysis.Mechanical properties of the arapaima scale incorporated epoxy composites were evaluated by 3-points bend tests in a model DL10000 EMIC machine, Brazil, with 10 tonnes of capacity, operating as per ASTM D790 [21] with 2 mm/min of loading velocity at RT. Nanoindentation was performed across the thickness of arapaima scale using procedures described by Brito et al. [22]. Ten indentations were performed at the distinct outer, inner and intermediate thickness layers of the scale.
3.
Results and discussion
In Fig. 1 some morphological characteristics of the arapaima scales can be seen with the naked eye. The scales have two distinct parts. First, a thicker and darker exposed region (≈2 mm). Second, a thinner and lighter embedded region (≈1 mm) [12–14]. While swimming, the dark region is exposed to the water while the light embedded region is covered by other scales in the fish skin. The external region of the darker region is referred in literature as the limiting layer [5,7,14]. The arrow in Fig. 1a indicates the direction of growth of the scales, which is perpendicular to the scale long axis, forming a ridge structure. This structure is oriented along the semicircle formed around the vertice point shown in this figure [12,14]. The arapaima scales, as other elasmoid scales, have a structure composed of three layers across the thickness: the limiting layer, the external (outer) layer and the internal (inner) layer. The outer and inner layers differ basically
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Fig. 1 – Arapaima gigas scale in the as-received state showing the (a) front view; (b) side view; (c) rear and (d) front views of the curved arapaima scales.
Fig. 2 – Hierarchical representation of the flattening process to which the curved scales where submitted before composite manufacture.
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Fig. 3 – (a) A layer of arranged arapaima scales after submitted to flattening process; (b) Epoxy matrix composite reinforced with 30 vol% of arapaima scales and (c) Metallic mold used in the manufacturing of composite plates.
by their mineral content. The outer layer presents a higher degree of mineralization [1,5,12–16], while the inner is basically composed of collagen [8]. An intermediate region may be considered in between the outer and inner layers. Fig. 4a and b are, respectively, SEM images of the exposed and embedded regions of the as-received scales. On the other hand, Fig. 4c and d are, respectively, images of the exposed and embedded scale regions after the flattening process. From Fig. 4a and c one can observe the transition from the ridge structure in the embedded part to the corrugated zone in the exposed part, which presents ridges with larger spacings in between. In these figures, one can also see the presence of cracks located between the ridges in the exposed part and across the ridges in the embedded region. This cracking formation may have occurred during the drying step of the flattening process.
From Fig. 4b and d, in the ridge structure of the embedded region, it is possible to see the presence of cracks mostly in the veins for the as-received scales. The flattened scales present the same type of aforementioned cracks with addition of some cracks across the ridges. The presence of cracks on the scales surface was also observed in previous studies and was associated with the natural drying process of the scales, which produce shrinking stresses [13,14]. Fig. 5a and b show SEM images of the fracture surface of the 30 vol% arapaima scale reinforced epoxy matrix composite. In the left side of Fig. 5a, one should notice the lamellar structure of the inner part of the scales. The lamellae present different contrast due to their different orientation forming a Bouligand-type structure, in agreement with previous works [13,14,23]. In Fig. 5a, it is also noted the presence of a crack running through the inner layer of the scale, which might have been nucleated in one of the ridges of the scales. In Fig. 5b
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Fig. 4 – (a) Transition region from the embedded part to the exposed part of as received scale; (b) Embedded part of an as received scale showing the ridge structure; (c) Transition region from the embedded part to the exposed part of a scale submitted to the flattening process and (d) Embedded part of a flattened scale showing the ridge structure with some cracks.
Fig. 5 – SEM images of the fracture surface of the 30 vol% arapaima scales reinforced composites evidencing: (a) the epoxy matrix and the scale and (b) the different layers of the scales internal structure.
it is possible to notice the interface region showing a good adhesion between the scale and the matrix phase. EDS scans where performed to assess the qualitative chemical composition of the scales. Fig. 6 shows the EDS results. In this figure, it is possible to see from the mapping of calcium (Ca) the presence of a higher degree of mineralization
in the limiting layer, where there is a higher concentration of Ca, associated with hydroxyapatite. Phosphorous (P) from the hydroxyapatite is also reported to exist in arapaima scales [13,14]. Fig. 7 shows the wavelength transmittance graphs obtained by the FTIR analysis for the epoxy composite reinforced with
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Fig. 6 – Energy dispersive X-ray spectroscopy (EDS) results for the arapaima scales: (a) SEM image of the analyzed region; (b) elemental mapping obtained and (c) graphs for the elemental mapping.
Fig. 7 – FTIR spectrum of the 30 vol% arapaima scales reinforced composite.
Fig. 8 – Comparison between the FTIR spectra of both the plain arapaima scales and the 30 vol% arapaima scales reinforced composite.
arapaima scales. In this figure, several bands associated with the scales as well as bands associated with resin may be noticed. When compared with the results obtained by Torres et al. [12] and Lin et al. [13], a small variation of transmittance intensity with respect to the plain scale and the composite could be detected. This is basically due to the interference caused by the resin, which is seen in Fig. 8. However, all transmittance bands related to the arapaima scales could be found and are presented in Table 1.
In this table, bands 2, 3 and 5 are associated with organic components of the scale that are related to type I collagen [8], while bands 4, 6 and 7 are related to the inorganic part of the scale, which is composed basically of hydroxyapatite [12,13]. In addition, the identification of the isolated bands from studies related to the arapaima scales indicates that there was no chemical interaction between the reinforcement and the matrix phases [9,12,13]. Table 2 shows a comparison between the bands obtained in previous articles and the ones in the
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Table 1 – FTIR absortion peaks observed and their related components. Peak
Wavelength (cm− 1 )
Related peak
1 2 3 4 5 6 7
3425.6 1631.1 1558.5 1458.2 1265.3 1033.9 864.1
Amide I Amide II Phosphate groups Amide III Phosphate groups Carbonate groups
Table 2 – Comparison of FTIR wavelengths of amide I, II and III.
Ikoma et al. [9] Torres et al. [12] Lin et al. [13] Present work
Amide I (cm− 1 )
Amide II (cm− 1 )
Amide III (cm− 1 )
1657 1662 1637 1631
1520 1560 1546 1558
1447 1242 1239 1265
Table 3 – Flexural properties by 3-points bend tests of 30 vol% arapaima scale/epoxy composite and plain DGEBA/TETA epoxy specimens. 3-points bend test specimens
Flexural Strength (MPa)
Flexural Modulus (GPa)
Maximum Deflection (mm)
Plain epoxy 30 vol% arapaima scales/epoxy
71.6 ± 25.5 43.6 ± 10.9
2.82 ± 0.49 4.50 ± 0.70
10.8 ± 4.0 5.6 ± 1.5
Table 4 – Nanoindentation hardness across the thickness of an arapaima scale. Hardness (GPa) across the thickness
Arapaima scale
Outer layer
Intermediate layer
Inner layer
0.323 ± 0.082
0.268 ± 0.011
0.170 ± 0.108
present work. These results indicate similar chemical interaction in both scales and epoxy reinforced with scales. Table 3 presents the flexural properties of the 30 vol% arapaima flattened scales incorporated into epoxy matrix composite, obtained in 3-points bend tests. Corresponding flexural strength and modulus as well as maximum deflection before rupture for plain DGEBA/TETA epoxy are also shown in Table 3. The results in this table reveal that, within the standard deviation (SD), strength and deflection are comparable for both plain epoxy and 30 vol% arapaima scale composite. On the other hand, the flexural modulus is sensibly increased with incorporation of arapaima scales into epoxy matrix. This characterizes a toughening effect, which can be attributed to the stiffer nature of the arapaima scales [13,17], as well as their collagen content [8]. Table 4 depicts the average and SD values for nanoindentation at the outer, intermediate and inner layers, Fig. 5a, of an arapaima scale. In this table one should notice the comparatively greater hardness at the outer layer, which is an expected result in view of the higher degree of mineraliza-
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tion due to calcium content in Fig. 6, as found and discussed in most works on characterization of arapaima scales [12–18]. The combination of a hard outer layer and a collagen-based [8] stronger inner layer contributes to reinforce epoxy composites as presented in Table 3.
4.
Conclusions
• The surface of the arapaima scales presented cracks located in the veins between the ridges in both as received scales and flattened scales, which were associated with the shrinkage stresses induced during the natural drying process. • Arapaima scales after the flattening process showed surface cracks going through the ridge structure, that could be related to the drying step of the flattening process. • The fracture surface of the arapaima scale epoxy composite revealed the lamellar structure of the scales. These also confirmed the physical interaction in the interface of the composite. • EDS results showed a high content of calcium in the external layer, which correlated to the higher degree of mineralization present in this outer layer and leads to greater hardness. Phosphorous peaks were also detected and related to the composition of the hydroxyapatite present in the scales. • The FTIR results were obtained for both the plain scales and the arapaima scales reinforced epoxy composites. It was possible to observe in corresponding graphs the characteristic bands associated with type I collagen as well as those constituents of the hydroxyapatite. Despite the variation in the transmittance intensity of the material, there was no chemical interaction between the composite phases. • Flexural properties disclosed a toughening effect, in terms of sensibly increase in the elastic modulus, caused by the incorporation of 30 vol% of arapaima scales into epoxy matrix. This novel cost effective composite material might be included among those based on discarded natural waste (scales in fish market) with potential engineering applications. • The improved composite toughening is supported by the combination of arapaima scale calcium-based hard outer layer with collagen-based stronger inner layer.
Conflicts of interest The authors declare no conflicts of interest.
Acknowledgements The authors thank the support to this investigation by the Brazilian agencies: CNPq, CAPES and FAPERJ.
Appendix A. Supplementary data Supplementary material related to this article can be found, in the online version, at doi:https://doi.org/10.1016/j. jmrt.2020.01.051.
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