structural design and mechanical behavior of alligator (alligator mississippiensis) osteoderms

Acta Biomaterialia xxx (2013) xxx–xxx

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Acta Biomaterialia

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Structural design and mechanical behavior of alligator (Alligatormississippiensis) osteoderms

1742-7061/$ - see front matter � 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.http://dx.doi.org/10.1016/j.actbio.2013.07.016

⇑ Corresponding author. Tel.: +886 3 571 5131x33889.E-mail address: [email protected] (P.-Y. Chen).

Please cite this article in press as: Sun C-Y, Chen P-Y. Structural design and mechanical behavior of alligator (Alligator mississippiensis) osteodermBiomater (2013), http://dx.doi.org/10.1016/j.actbio.2013.07.016

Chang-Yu Sun, Po-Yu Chen ⇑Department of Materials Science and Engineering, National Tsing Hua University, 101, Sec. 2, Kuang-Fu Rd, Hsinchu 30013, Taiwan

a r t i c l e i n f o a b s t r a c t

Article history:Received 9 April 2013Received in revised form 12 July 2013Accepted 16 July 2013Available online xxxx

Keywords:OsteodermArmorCompositesMechanical propertyToughening mechanisms

Alligator is a well-adapted living fossil covered with dorsal armor. This dermal shield consists of bonyplates, called osteoderms, interconnected by sutures and non-mineralized collagen fibers, providing adual function of protection and flexibility. Osteoderm features a sandwich structure, combining an innerporous core and an outer dense cortex, to offer enhancements for stiffness and energy absorbance. In thisstudy, we investigated the multi-scale structure and mechanical behaviors of the American alligator (Alli-gator mississippiensis) osteoderm. Microcomputed tomography was applied to reveal the complex neuro-vascular network. Through the observation under optical and scanning electron microscopes, theosteoderm was found to consist of woven bone in the dorsal region and lamellar-zonal bone in the ven-tral region. Nanoindentation and compressive tests were performed to evaluate the mechanical proper-ties of osteoderms. The varying mineral contents and porosity result in a graded mechanical property: ahard and stiff dorsal cortex gradually transform to a more compliant ventral base. Three protective mech-anisms optimized for alligator osteoderms were proposed and elucidated.

� 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

1. Introduction

Many structural biological materials have been extensivelyinvestigated in recent years due to their superior mechanical prop-erties, considering the weak building blocks of which they arecomposed [1–3]. Currently, flexible and lightweight dermal armorshave aroused increasing interest due to their intriguing designs forprotection [4,5], including fish scales [6–12], turtle shells [13–16]and armadillo carapaces [17,18]. Fish scales, such as P. senegalus[7], A. gigas [8] and A. spatula [10,12], have been widely studied la-tely. Despite the differences in material compositions, mineral con-tent and thicknesses, they all applied a similar strategy ofcombining a stiffer and harder external region with a softer inter-nal base. The scales of these marine species exhibit flexibilitythrough interlocking and overlapping [4,5].

On the other hand, armadillo carapace [17,18] and turtle shell[13–16] utilize rather distinct strategies from fish scales. These ar-mors share many similar structural features: (1) the main constit-uents of these mineralized tissues are bone, consisting of collagenfibers and hydroxyapatite minerals; (2) the bony plates are con-nected by soft tissues or joints; (3) they are covered by keratinouslayers on the outer surface; (4) they are both sandwich compositeswith a dense cortex and a porous core. Chen et al. [17] found thatthe non-mineralized collagen fibers are responsible for the

macroscopic mechanical responses of the armadillo carapace. Thestretching of these connective fibers between hexagonal plates isthe major contribution to tensile and shear strengths [17]. In turtleshells, the bony segments are juxtaposed with zigzag joints inter-locking in between, called sutures. The sutures are three-dimen-sional (3-D) and complicated structures with organic tissues,giving rise to effortless deformation under small loads and trans-ferring to stiffer responses after locking under higher degrees ofmovement [15]. Rhee et al. [13] reported that the porous core ofthe turtle shell is made of closed-cell foam, causing the sandwichstructure to undergo a nonlinear deformation, which leads to ahigher specific energy absorption compared with the dense cortexalone. Recent investigations conducted by Achrai and Wagner [14]revealed that the dorsal and ventral cortices of the sandwich struc-ture own various mechanical properties as a result of different fi-ber arrangements. The randomly oriented fibrillar network in thedorsal cortex can sustain sharp impact isotropically, while the ply-wood arrangement of fibers in the ventral cortex possesses aniso-tropic mechanical properties and is beneficial for structuralsupport [14]. The turtle carapace also appears to be a functionallygraded material (FGM) in terms of composition, porosity andmechanical properties.

Crocodilian osteoderm is another interesting topic in naturalflexible dermal armors. These ancient reptiles have long been con-sidered as fierce carnivorous tetrapods with heavily armored skins.Although they seldom encounter predators, territorial fightsamong the same species can often be deadly because of their

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extremely high bite force, reaching �10 kN, the highest value thathas been reported for living animals to date [19]. Thus, well-devel-oped armor designs for excellent mechanical performances are de-manded, along with some flexibility for speedy and agilemovements in order to capture preys. As a matter of fact, the dorsalsheaths of crocodilians have been used as armor suits for ancientwarriors since they are found to repel knives and arrows, and areeven bulletproof under certain conditions, as discovered recently[20]. However, the microstructure, mechanical properties anddeformation mechanisms have not been thoroughly investigated.

In this study, we investigated the osteoderm of American alliga-tor (Alligator mississippiensis) by multi-scale structural character-izations using materials science approaches. Mechanicalbehaviors were evaluated and related to the structure features atvarying length scales, and the deformation as well as tougheningmechanisms of this biological armor when subjected to externalforces were proposed. We hope this study can provide furtherunderstanding of biological defensive designs, and offer inspirationfor novel synthetic armors and advanced composites.

2. Background

Reptiles are cold-blooded animals featuring scales that covertheir whole body. Among them, crocodilians, including crocodiles,alligators and caimans, are amazing living fossils which appeared150 million years ago and have evolved into one of the most adap-tive modern animals on the planet. These large tetrapods possessnot only keratinous scales on their external surfaces, but also un-ique bony plates underneath the keratinous scales for reinforcedprotection, called osteoderms. Crocodilian osteoderms are foundmainly on the dorsal dermis (also on the abdomen for some spe-cies, such as most caimans), sheltering areas from the nuchal tothe caudal region. These natural armors are composed of mineral-ized bony plates which are connected by fibrous tissues, similar toarmadillo and turtle carapaces. The hierarchical structure of thedermal armor of Alligator mississippiensis is schematically pre-sented in Fig. 1. The whole armor includes about 70 pieces of bonyplates. Each plate has a longitudinal keel in the middle. Through atransverse cross-section, various structural features are demon-strated. The external surface of the bony plates is covered by a thinlayer of keratinous scutes. These scutes, or scales, cover the dorsalarmor of alligator as well as all other parts of its body, and mayvary in shape, composition and formation mechanism. They result

Fig. 1. Hierarchical structure of alligator osteoderm from m

Please cite this article in press as: Sun C-Y, Chen P-Y. Structural design and mBiomater (2013), http://dx.doi.org/10.1016/j.actbio.2013.07.016

from morphological transitions through differentiation andkeratinization of the crocodilian’s epidermis [21]. Harder andtougher b-keratin outer layer coats the osteoderms to provide wearresistance, while the a-keratin forms mainly the matrix and hingeregions, acting as a barrier to water and electrolyte exchange[21,22]. In addition, connective fibers are found at the junction oflaterally neighboring bony plates.

Osteoderm is not an original element in evolutionary for croc-odilians. This type of integumentary skeleton is a plesiomorphictrait for tetrapods [23,24], and has been well demonstrated inmany dinosaurs, such as the renowned stegosaurs [25], ankylo-saurs [20], and other extinct relatives such as squamates [26]and archosaurs [27]. The osteoderms in various species differ insize, shape, ornamentation and functions. In addition to protectionfrom claws and teeth of predators, other functions of osteoderms,including heat transfer [25,28], mineral storage [29] and locomo-tion aid [30], have been suggested. Since these integumentary skel-etons of reptiles are not subjected to external forces and do notlikely undergo bone remodeling, the ages of these animals can beestimated by counting the growth marks in the osteoderms corre-sponding to the seasonal changes in growth rate [31], which is anequally valid yet much more convenient method to obtain agesthan counting or observing the growth marks in the interior bones(e.g. long bones) from living or preserved species [32].

3. Materials and methods

3.1. Sample preparation

A complete dorsal armor of an American alligator was obtainedfrom Jernigan’s Taxidermy (Waco, Texas, USA). The alligator armorwas prepared using a relatively harmless and natural methodwithout using strong chemicals, which may alter the natural stateof the samples. The longitudinal length of the armor is �0.85 m,indicating the animal may have had a body length of �1.6–1.8 msince the armored part, excluding the head and about half the tail,accounts for �50% of the length of the entire animal. It it is likely tobe a mature alligator, 8–10 years old, depending on the gender. Ithas been reported that male alligators in Texas can be 8 ft longat the age of 10 and female alligators 6–7 ft long at the same age[33].

Keratin coverings on the dorsal surface and dried dermis on theventral surface were removed in order to observe the structural

acro-, meso- and micro-, to nanometer-length scales.

echanical behavior of alligator (Alligator mississippiensis) osteoderms. Acta


C.-Y. Sun, P.-Y. Chen / Acta Biomaterialia xxx (2013) xxx–xxx 3

features of individual osteoderms. All samples prepared for micro-structural characterization and mechanical testing were takenfrom the central and caudal parts of the whole armor to maintainconsistency, since the cervical and transverse terminal osteodermspossess rather irregular shapes and non-uniform keel heights. Itshould be noted that the experimental samples are taken from asingle alligator, and may not be representative of the entirespecies.

3.2. Elemental analysis

3.2.1. Ash content measurement17 samples sectioned from five mid-dorsal osteoderms (3–4

samples from each) with regions varied from the keel to the edgewere used to determine the average mineral content of the osteo-derm by the ash-weight method [17]. Since the samples for themeasurement were all from the mid-dorsal osteoderms, the resultsmay stand for the major portion of the alligator armor. However, itshould be noted that the mineral content may change for osteo-derms at different locations on the body. Samples were dried ona hot plate at 105 �C for 12 h and the dry weights were measuredusing an electronic balance. Samples were then ashed at 600 �Cfor 24 h and ash weights were measured. The water content andash content (in wt.%) were calculated.

3.2.2. X-ray diffractionX-ray diffraction (XRD) was carried out on powders of ground

alligator osteoderms utilizing a powder X-ray diffractometer(XRD-6000, Shimadzu Co., Kyoto, Japan). A continuous scan usingCu Ka1 (k = 0.154 nm) as the radiation source was performed in ah–2h mode scanning from 2h = 20� to 60�, with a step size of0.02� at a rate of 2� min�1.

3.2.3. Electron probe microanalysisThe localized elemental compositions were analyzed by field-

emission electron probe microanalysis (FE-EPMA) with a JEOLJXA-8500F EPMA (JEOL Ltd., Tokyo, Japan). Three cross-sectionalsamples (�10 � 5 � 5 mm3) with both dorsal and ventral regionsof the osteoderm were sectioned and embedded in epoxy followedby grinding and polishing. The samples were coated with a thinlayer of carbon instead of other common conductive coatings suchas gold or platinum because these heavy metal coatings can se-verely suppress the emission of X-rays induced by the incidentelectron, serving as a barrier layer for the signals to come outand be detected. Five quantitative measurements were taken fromthe dorsal and ventral region, respectively, for each specimen, andthe results were then averaged to compare the compositional dif-ferences between the two regions.

3.2.4. Energy-dispersive spectroscopyElemental mapping at interfacial regions was achieved by an

energy-dispersive spectroscope (EDS) within a field-emission scan-ning electron microscope (FE-SEM) (JSM-7600F, JEOL Ltd.). Sam-ples were sectioned and ground from the edge of an osteodermand coated with a thin layer of carbon (�10 nm) to prevent elec-tron charging. X-rays were produced from electron bombardmentsunder an accelerating voltage of 10 kV and a working distance of15 mm. A silicon-drift detector (SDD) (X-Max SDD, Oxford Instru-ments, Abingdon, Oxfordshire, UK) was used to collect the charac-teristic X-rays from the sample, while the software AZtec (OxfordInstruments) was applied to analyze and map the elementaldistributions.


3.3. Structural characterization

3.3.1. Macroscopic observationExternal shape and morphology of osteoderms were taken from

central and edge regions of the whole armor. For cross-sectionalobservation, a sample was cut by a hand saw and through the keelregion followed by grinding and polishing. Photographs of top, bot-tom and cross-sectional views were taken by a digital camera.

3.3.2. Microcomputed tomography (l-CT)l-CT was accomplished by unmonochromatized synchrotron

hard X-rays with energy ranging from 5 keV to 35 keV at the Na-tional Synchrotron Radiation Research Center (NSRRC) in Hsinchu,Taiwan [34]. l-CT images were obtained from a CCD camera (mod-el 211, Diagnostic instruments, 1600 � 1200 pixels) after convert-ing the X-rays into visible lights by a scintillator. The resolution ofthe corresponding images (in pixel size) of a 2� lens was�2.8 lm � 2.8 lm. In order to reconstruct 3-D tomographic mod-els, images were collected with a regular step size of 0.3� over a to-tal 180� rotation of the specimen stage. Xradia software (XradiaInc., Pleasanton, CA, USA) was applied to reconstruct the raw data,which was then visualized by Amira software (Visualization Sci-ence Group, a FEI company, Burlington, MA, USA).

3.3.3. Optical microscopy and stereoscopyCross-sectional samples through the keel region were ground

and polished before observing under an optical microscope(BX51M, Olympus Co., Tokyo, Japan) equipped with a 0.8 mega-pixel digital camera (DP12, Olympus Co., Tokyo, Japan). An inte-grated view of the keel cross-section was achieved by combining17 micrographs of consecutive regions.

Stereoscopic images were taken from an Olympus SZX7 ZoomStereomicroscope (Olympus Co., Tokyo, Japan) with a 2.0 mega-pixel CCD camera (Infinity 1, Lumenera Co., Ontario, Canada). Themagnification of the stereoscope ranged from 8� to 56�.

3.3.4. Scanning electron microscopyMicrostructural characterization of the fracture surfaces were

observed by a FE-SEM (JSM-7600F, JEOL Ltd.). Fracture surfaceswere created by exerting a bending force through a clamp and awrench. The specimens were coated with a thin layer of platinumto enhance electron conductivity on the surface. Secondary elec-tron images (SEIs) were taken with an accelerating voltage of10 kV and a working distance of 10 mm.

3.4. Mechanical testing

Schematic representations of specimens prepared for mechani-cal testing are shown in Fig. 2. The system of coordinates weadopted throughout the text is illustrated in Fig. 2a. Longitudinalis defined as the direction along the keel long axis, transverse is re-ferred to the direction perpendicular to the keel and vertical is thedirection through the thickness of the osteoderm. Longitudinal andtransverse are both included when the term ‘‘horizontal’’ isreferred.

3.4.1. NanoindentationA sample for nanoindentation was taken from the keel region of

a caudal osteoderm. The sample was mounted with the longitudi-nal cross-sectional area (Fig. 2a) revealed, followed by grinding andfine polishing with Al2O3 suspensions from particle size of 1 lm,0.3 lm and finally 0.05 lm. The average surface roughness of thefinal sample was �20 nm, measured with atomic force microscopy(Dimension Icon, Bruker Corp., Billerica, MA, USA). Nanoindenta-tion tests were conducted by using a Hysitron TI900 TriboIndenter(Hysitron Inc., Eden Prairie, MN, USA) with a Hysitron TI-0039



Fig. 2. Schematic illustrations of sample preparations for mechanical tests. The shades of the color in (b) and (c) denote different regions, i.e. dorsal and ventral region. (a) Thehead–tail direction along which the keel is oriented is defined as the longitudinal (or parasagittal) direction. The direction along the lateral row of osteoderm is defined as thetransverse direction. Both directions are included when ‘‘horizontal’’ direction is referred. The horizontal direction is perpendicular to the vertical direction. (b) An illustrationof the direction where nanoindentation tests were performed. (c) An illustration of the locations where compression samples of different orientations were taken. Due to thegeometrical limitations, vertical samples are taken near the keel and possess more dorsal region, while the horizontal samples are taken mainly within ventral regions.


Berkovich diamond tip under a load-controlled mode. The radius ofthe tip was �100 nm. The load function can be separated into threestages: linear loading, holding and linear unloading, and the dura-tion for each stage was 5 s, with the peak load set to be 1000 lN.The area function of the tip was calibrated before conducting thetests with a fused quartz bulk specimen as a standard materialdue to its low elastic-modulus-to-hardness ratio [35]. A series ofindentations was performed vertically across the cross-section(Fig. 2b) with an interval of 300 lm and a total number of 53groups. Each group contained eight indent points forming a 2 � 4rectangular region with a lateral space of 15 lm between neigh-boring indents, which is a significantly large distance comparedwith the indent size to avoid effects of adjacent indents. The hard-ness and reduced modulus values of each group were then aver-aged. The area function was calculated again after the test andshowed no significant variations (�0.3%) on the tip geometry, sug-gesting that the results were reliable even after a large amount ofindentations were performed.

3.4.2. Compressive testingSamples for compressive testing were cut into 2.3 �

2.3 � 4.5 mm3 rectangular pieces by a rotating diamond blade.The dimensions were chosen to prevent buckling by the Euler’s cri-teria. Each facet of the samples was then ground carefully using aclamp to ensure that the two surfaces in contact with the upperand lower load cells are parallel to each other, while keeping theside surfaces perpendicular to the ends as precise as possible toeliminate eccentric loading. 80 vertical and 80 horizontal sampleswere prepared, as shown in Fig. 2c. The horizontal samples weretaken from flat (non-keel) regions, since the large deviation onthe vertical direction of the keel cross-section may have significantinfluences on experimental results. However, for vertical samples,


the height 4.5 mm cannot be satisfied at locations far away fromthe keel due to the decrease in thickness from the keel towardthe edge. Therefore, vertical samples were taken from non-keel re-gions near the keels. The difference in regional distribution be-tween the horizontal and vertical samples led to differentportions of dorsal and ventral regions, which is clearly demon-strated in Fig. 2c: vertical samples contain more dorsal region thanthe horizontal samples. 40 samples from each group (longitudinaland vertical directions) were immersed in Hank’s balanced salinesolution (HBSS) (H2387, Sigma–Aldrich Co., St Louis, MO) for24 h before mechanical testing. The rehydrated samples weretested immediately after taking out of HBSS in order to prevent fur-ther drying. The other set of samples (40 each direction) weretested in ambient dry condition. Compressive tests were conductedby using a universal testing machine (Instron 3343 Single ColumnTesting System, Norwood, MA, USA) with a 1 kN load cell at a strainrate of �1 � 10�3 s�1.

3.4.3. Flexibility demonstrationTwo adjacent osteoderms were taken from the mid-dorsal re-

gion of the alligator armor to demonstrate the flexibility of thejoints. The bony plates were sectioned transversely to better dis-play the angles bent by bare hands.

3.4.4. Whole osteoderm compressionA transversely cross-sectioned large osteoderm sample with

keel height of �15 mm was used to demonstrate the deformationof the sandwich structure under compression. The sample wasground at the bottom to create a flat contact surface, followed byimmersion in HBSS for 24 h before the test. The large-scalecompressive tests were carried out by a universal testing machine



Fig. 3. Photographs showing the top views of mid-dorsal (a) and transverse terminal (b) osteoderms show different shapes and features. The ventral surface (c) of a mid-dorsal osteoderm shows that arterial grooves were used to hold vessel branches. The arrows indicate pits where the artery bifurcates into nutrient foramina entering theosteoderm. The edge of the osteoderm between laterally neighboring plates is shown in (d), which was covered by non-mineralized connective fibers. By removing the non-mineralized connective fibers, the sutures can be observed in (e). A SEM image in (f) shows the 3-D feature of the sutures, which contain pits that are connected to theneurovascular system within the plates.


(Instron 4468, Double Column Testing System, Norwood, MA, USA)with a 50 kN load cell at a strain rate of � 1 � 10�3 s�1.

4. Results and discussion

4.1. Macroscopic observation

An osteoderm from the mid-dorsal region appears in a quadrateshape �5 cm in length and width with a parasagittally aligned keelof �1.5 cm in height (Fig. 3a). Osteoderms taken from different


locations show distinct appearances, in both shape and keel height.An osteoderm from the transverse terminal is shown in Fig. 3b forcomparison. The large concave regions on the external surface con-tain small cavities that connect to the vascular channels, whichwere proposed to be the evidence for the thermoregulation func-tion of alligator osteoderms [30]. The neurovascular foramina enterthe bony plates from the ventral surface, as shown in Fig. 3c, wherethe grooves are traces of bifurcated dorsal median arteries circulat-ing across the surface [30]. The transverse edge of an alligatorosteoderm contains connective collagenous fibers (Fig. 3d) be-tween two adjacent plates, similar to the armadillo carapace



Fig. 4. Minerals in alligator osteoderm. (a) The amount of mineral content withinthe osteoderm was measured by ash-weight method, where the results showeddecrease from the keel to the edge (keel: 67.13 ± 0.58 wt.%, transition:65.54 ± 0.56 wt.%, edge: 62.89 ± 1.60 wt.%). (b) X-ray diffraction (XRD) pattern ofthe alligator osteoderm confirmed that hydroxyapatite is the main mineralcomponent.


[17]. These fibers account for connectivity and flexibility enhance-ments. By removing the connective soft fibers, serrated sutures(Fig. 3e) are revealed, which share the same functional design forinterlocking as the turtle shell [15]. From SEM, we can clearly ob-serve the 3-D characteristic of the sutures (Fig. 3f). Numerous tinypits are also shown in Fig. 3f, which are connected to the neurovas-cular foramina, presumably for vascularization, sensing and nutri-ent transportation.

4.2. Mineral content measurement and elemental analysis

From the ash-weight measurement, the water content is10.70 ± 0.58 wt.% and the mineral content of the dried specimenis 65.77 ± 2.26 wt.%, giving 34.23 ± 2.26 wt.% of the dried alligatorosteoderm to be organic components. The osteoderm of Americanalligator possesses a similar mineral content to that of armadillocarapace (�65 wt.%) [17] and bovine femur (�67 wt.%) [36], whichis higher than other natural armors such as tortoise shell(�53 wt.%) [37] and fish scale (�46 wt.%) [6], as well as that ofsome mammalian compact bones, for instance, elk antler(�57 wt.%) [38]. It is also found that the mineral content decreasedgradually from the keel (67.13 ± 0.58 wt.%) through the transitionregion (65.54 ± 0.56 wt.%) to the edge (62.89 ± 1.60 wt.%) as shown


in Fig. 4a, which is functionally graded since the edge with suturesand connective fibers serve for flexibility enhancements, and maybe a result of evolutionary convergence with the turtle shell [14].

To confirm the mineral constituents, the crystalline phase wasdetected by XRD. The resulting pattern in Fig. 4b can be indexedto JCPDS 09-0432, revealing hydroxyapatite as the main compo-nent, which is the same as the minerals in bone and other bony tis-sues [6,17,38]. Furthermore, localized elemental analysis measuredby FE-EPMA shows that the dorsal region contains more calcium(29.58 ± 1.15 wt.%) and phosphorus (13.13 ± 0.68 wt.%) than theventral region (Ca: 24.53 ± 1.62 wt.%, P: 10.38 ± 0.75 wt.%). The re-sults indicate that the dorsal region contained more minerals,which are primarily non-stoichiometric hydroxyapatite, while theventral region appeared to be less-mineralized and had more or-ganic constituents.

Elemental mapping by EDS under FE-SEM was conducted toanalyze the compositional difference between the bony plateand the connective fibers (Fig. 5). As the secondary electron im-age (SEI) in Fig. 5a shows, the area of interest was the interfacebetween sutures, which is still a part of the bony plate, and theconnective fibers at the edge of an osteoderm. Carbon mappingin Fig. 5b indicated that both regions contain organic contents,which is collagen [24], and that the amount of organic contentsis obviously much richer in the connective fibers. On the otherhand, distribution of Ca (Fig. 5c) and P (Fig. 5d) clearly illus-trated that hydroxyapatite (Ca10(PO4)6(OH)2), as the major min-eral content, is confined in the suture. Therefore, it wasconfirmed that the fibers connecting bony plates together arenot mineralized.

4.3. l-CT imaging

A cross-sectional view in the longitudinal direction of an osteo-derm keel reveals a sandwich structure (Fig. 6a), where the porousinterior is surrounded by compact cortex. A sandwich structurealso appears in various light-weight designs, such as leaves [39],bird beaks [40] and feathers [41], as well as many defensive de-vices against impact and bending, including human skull [39], tur-tle shell [14], armadillo carapace [17], fish armor [11] andhorseshoe crab exoskeleton [1]. The main advantage of sandwichstructures is to provide high bending stiffness with minimumweight. Defensive designs applying this principle also serve thefunction of energy absorbance under impact loads by deformationsof the cellular core through elastic bending, brittle fracture or plas-tic buckling of the walls or trabeculae before undergoing densifica-tion [39]. The reduced burden of these lightweight armors can thusenhance locomotion along with improvements in mechanicalproperties.

Since cross-sections can only show porosities and incompletechannels, a computed tomographic technique was applied to gain3-D information of the complex neurovascular network in thebony plates. By collecting the transmitted X-ray signals and thecorresponding intensities, we can identify the porosity since theabsorbance along the X-ray path is different for materials withporosity and without porosity. Through rotation during scanning,the information of the whole specimen can be obtained and 3-Dmodels can be further reconstructed. A sectioned image of thereconstructed 3-D model from l-CT scans is shown in Fig. 6b.The neurovascular channels form an intricate 3-D network, wherethe major cavity in the center branches out toward the dorsal andventral regions with much smaller pipes. The bifurcation withinthe dorsal region tends to be more complex compared with theventral region, whereas both regions contain evidences of sea-sonal and/or annual growth, indicated by the white arrows inFig. 6b.



Fig. 5. Elemental mapping by energy-dispersive spectroscopy (EDS) at the edge of the osteoderm. (a) An SEI image of the area being analyzed, which shows no significantdistinguishing structural features of the two regions. (b) Carbon mapping indicates that the carbon content is much richer in the connective fibers. (c, d) Calcium andphosphate mapping clearly demonstrates the lack of minerals, which is mainly hydroxyapatite, within the connective fibers.


4.4. Microstructural characterization

4.4.1. Optical microscopyFour different regions with different microstructural morpholo-

gies of bone can be distinguished from the optical micrograph, asshown in Fig. 7a. At the outer sheath, randomly oriented wovenbone can be observed along with extensive vascularization. Thevascular channels connect to small pits on the external surface,mainly located within the large concave regions, as previouslyshown in Fig. 3a and b. These pits and vascular systems are sug-gested to be the major evidence of the role of osteoderm in ther-moregulation of the body [30]. The second region beneath theouter sheath is composed of dense woven bone and scatteredlamellar rings deposited around the neurovascular channels. AnSEM image taken from this region is shown in Fig. 7b. The patternson the surface indicate collagen fiber bundles being ruptured andpulled out, where no preferred orientation can be observed, illus-trating bundles entangled in a randomly woven manner. Scatteredlamellar rings can be recognized in this region. These concentriclamellar structures are not likely to be secondary osteons sinceno canaliculi and well-developed vascular systems (Haversianand Volkmann’s channels) are observed. Moreover, osteodermsare not constantly subjected to external loading and bone remod-eling may be restricted and limited. The third region contains por-ous woven bone with large neurovascular channels. The wovenbone in this region is similar to that of the outer sheath. The majorneurovascular foramen and branches mainly locate in this regionand the large cavities can reach up to hundreds of micrometersor several millimeters in diameter. Lamellar bone is also foundaround the channels, forming circular rings. At the bottom, the ba-sal region consists of lamellar-zonal bone [42]. This type of bone iscommonly seen in reptiles, and is related to the poor vasculariza-tion, which derives from low metabolic rate [42]. The lamellar-zo-nal bones correspond to seasonal or annual growth, where bone


growth stops or slows down in winters, leaving lines of arrestedgrowth (LAGs). The lamellae in this region are not the same asthe lamellar bone, but rather a parallel-fibered bone, which is con-structed by woven collagen fibers with a preferred orientation(Fig. 7c). Furthermore, Sharpey-fibered bone is also found mostlyin the ventral region and sometimes on the edge of dorsal region,extending in oblique directions from the margin to the core withwavy or zigzag structures [20,23,24]. This type of bone is derivedfrom non-mineralized Sharpey’s fibers functioning for connection,which fuse into the osteoderm and anchor the bony plates to theepidermis. The Sharpey-fibered bone in the ventral region indi-cated locations where the osteoderm contact the epidermis, whilethose in the dorsal region implied that the genesis of osteodermtook place within the skin. It is thus discovered that the alligatorosteoderm consists of various types of bone, resulting in a hetero-geneous composition. The differences in fiber orientation at differ-ent locations are schematically presented in Fig. 1. It should benoted that the structural variations between adjacent regions un-dergo a gradual change.

Thin sections of the osteoderm were observed under the stereo-scope (Fig. 8). LAGs are clearly observed in the ventral region, whileonly vague annuli can be recognized at the dorsal region becauseno complete halt occurred during growth. The growth rate is alsodifferent in the two regions. Although this is not a precise quanti-tative evaluation due to the limited resolution, we can still distin-guish that the interval between annual growth marks in the ventralregion (�0.5 mm) is much narrower compared to those in the dor-sal region (�1.1 mm). Hence, we proposed a growth model, asshown in the schematic illustration (Fig. 8). The growth rate inthe keel region is higher than the basal region, leading to a uniqueshape of a ridged keel. In addition, according to Vickaryous andHall [24], the mineralization process of the osteoderm initiatesfrom the keel, and then extends radially across the whole plate.Therefore, combining the non-simultaneity in calcification and



Fig. 6. (a) The sandwich structure of an osteoderm keel cross-section. The porosityis caused by the foramina branching system. (b) 3-D image reconstructed from l-CTscans by synchrotron X-rays. The white arrows indicate annual growth of thebranches.


the anisotropy in growth rate, it is implied that the degree of min-eralization may not be uniform within the entire osteoderm, espe-cially along the vertical direction.

4.4.2. Scanning electron microscopyMicrostructural characterization of a fractured osteoderm keel

is shown in Fig. 9, where comparisons between the dorsal, ventralregions and the non-mineralized collagen fibers are made. At alower magnification, the dorsal region (Fig. 9a) shows a denselypacked and relatively flat fracture surface, indicating a direct fail-ure and rather brittle behavior upon fracture. The mineralized col-lagen fiber bundles are fused together and the patterns areobscured. Under a more detailed view (Fig. 9b), granular morphol-ogies of mineral aggregates on the surface are observed. Individual


fibers can hardly be recognized in this region, implying that the fi-bers are highly mineralized and form bundles which cannot be eas-ily separated. In comparison, the fracture surface taken from theventral region (Fig. 9c) exhibits a fibrous feature. These collagen fi-bers were stretched and twisted upon breakage, revealing a moreductile behavior. Individual fibrils can be easily distinguished athigher magnification (Fig. 9d), where the surfaces of these fibrilsappear to be smoother compared with those in Fig. 9b. It is there-fore suggested that these collagenous fibers should possess a lowerdegree of mineralization, in contrast to those in the dorsal region,which corresponds to the growth rate and mineralization processas previously proposed in Fig. 8. The SEM observation is also ingood agreement with the previous EPMA results, which suggestedmineral content difference in dorsal and ventral regions.

Fig. 9e shows the microstructural features of the non-mineral-ized connective fibers between neighboring bony plates to provideflexibility for the dorsal shield. These organic bundles are ran-domly oriented, with smooth surface morphologies as shown inthe higher magnification (Fig. 9f). Characteristic patterns of67 nm periodicity in collagen fibrils, which derives from the stag-gered molecular arrangement, can also be observed in Fig. 9f.

In summary, the alligator osteoderm is a complex, heteroge-neous, hierarchically structured bio-composite with varying de-grees of mineralization and porosity at different locations.Through a thorough compositional and structural characterizationat multiple levels with XRD, EPMA, EDS, l-CT, OM (optical micros-copy) and SEM, the whole dermal armor is revealed to be a hybridsystem of mineralized bony plates and non-mineralized connectivecollagen fibers, whereas each bony plate is also a combination of aheavily-mineralized interwoven dorsal cortex and a less-mineral-ized parallel-fibered ventral base, with complex 3-D neurovascularchannels branching from the core of the osteoderm. These featuresare integrated and organized in the schematic illustrations of Fig. 1.

4.5. Mechanical behavior

4.5.1. Nano-mechanical evaluationNanoindentation tests along the cross-section of an alligator

osteoderm reveal the difference in mechanical properties for dorsaland ventral regions. Hardness and reduced modulus of each indentare calculated from the load–depth curves according to the Oliver–Pharr method [43]. Fig. 10a shows the hardness variation acrossthe keel while the photo above the plot corresponds to the inden-tation position. The average hardness values are 367 ± 94 MPa forthe ventral region and 690 ± 170 MPa for the dorsal region.Fig. 10b shows the change in reduced modulus through the keelcross-section, where the trend is the same as the hardness values.The ventral region was found to possess an average reduced mod-ulus value of 13.9 ± 2.1 GPa, gradually increasing to 20.3 ± 3.4 GPain the dorsal region. Drops at the exterior region for both hardnessand reduced modulus are observed. The relatively high standarddeviation for both hardness and reduced modulus across the wholesample can be related to the non-uniform porosity within theosteoderm. The projected area of contact of nanoindentation onthe specimen is �1–5 lm2, and if we consider the deformed sur-face profiles around the indents, the area influenced by the inden-tation can reach up to �10 lm2. On the other hand, the size of theporosity in the osteoderm spreads over a wide range from sub-micrometer to millimeter, where those smaller than tens ofmicrometers may cause serious effects on the indentation results.We have avoided the larger pores at the scales from millimetersdown to several tens of micrometers under the optical microscopewhile choosing the indent positions. However, smaller voids be-yond the limit of light microscopy can still exist, which we wereunable to identify and avoid. Therefore, it is possible that we haveset our indents on or nearby the porosity and affected the results,



(a)

Woven & lamellar bone

(b)

Lamellar-zonal bone

(c)

Fig. 7. (a) A combined optical micrograph of an osteoderm keel cross-section. Four regions can be distinguished as marked in the Fig.: outer sheath, woven bone and lamellarbone, woven bone with neurovascular channels and lamellar-zonal bone. (b) An SEM image of the dorsal region showing tangled woven collagen fiber bundles. (c) An SEMimage from the ventral region showing parallel-fibered bone (woven bone with a preferred orientation).

Fig. 8. A schematic drawing demonstrating the proposed growth mechanism of an alligator osteoderm. The insets are stereoscopic images of thin-sectioned samples from thedorsal (left) and ventral regions (right). The black arrows indicate growth marks (LAGs or annuli).


giving rise to the high standard deviation. Furthermore, since theporosity is three-dimensional, it is very likely that there are voidsunderneath the indentation surface at any depth, which we cannot


observe but may have significant effects on the measured values. Inaddition, the osteoderm is composed mainly of woven bone, whichis a loosely structured mineralized tissue. Thus, it is also possible



Dorsal Region

(a) (b)

Ventral Region

(c) (d)

Non-mineralized

Collagen Fibers

(e) (f)

Fig. 9. SEM fractographs at low and high magnifications of the dorsal (a, b) and ventral (c, d) region of an alligator osteoderm keel and the connective fibers (e, f). Differentmorphologies indicate the varied amount of minerals on the collagen fibers: the dorsal region is highly mineralized, the ventral region is less mineralized and the connectivefibers are non-mineralized.


that we have indented onto the spacing between the collagen fiberbundles, resulting in large deviations.

The distribution of localized mechanical properties is in goodagreement with the microstructural features in different regionspreviously described in Fig. 7a. The second region (II) from theexternal surface contains highly mineralized, relatively dense wo-ven bone and has the highest hardness and reduced modulus;whereas less-mineralized lamellar-zonal bone in the basal region(region IV) possesses the lowest values in both properties. Theexterior region (I) and the porous core (III) both seem to have anintermediate mechanical property between the dense dorsal re-gion and the ventral base, corresponding to their similar microcon-stituents (woven bone with high porosity). Fig. 10c shows the


typical load–depth curves of the dorsal and ventral regions, illus-trating the distinct difference in mechanical properties. Under aconstant maximum load, it is apparent that the dorsal regionshowed a higher stiffness and hardness with smaller indentationdepth and larger slope of the unloading path, whereas the ventralregion showed an advantage of higher energy dissipation esti-mated by the area under the curves. The different mechanicalproperties between the dorsal and ventral region may relate tothe compositional difference in mineral content and variation inporosity, as demonstrated in elemental analysis and microstruc-tural characterization.

The combination of hard external region and soft internal re-gion is analogous to many biological composites, such as arthropod



Fig. 10. Nanoindentation tests along the cross-section of an osteoderm show difference in (a) hardness and (b) reduced modulus in various regions. Note the gradienttransition between the two regions. (c) Representative load–depth curves for dorsal and ventral regions show distinct difference in mechanical properties at the maximumload of 1000 lN.


exoskeleton [44] and fish scales [7,8]. This is beneficial for defen-sive armors since the stiff and hard exterior can resist penetrationsby sharp claws or teeth, whereas the compliant interior can arrestcrack propagation and provide toughness, acting as a cushion byabsorbing impact energy. However, most of them reported a dis-continuity (or extremely steep gradations) in mechanical re-sponses between the stiff and compliant layers. In this study,hardness and elastic modulus change gradually rather than therebeing discrete interfaces, as found in the scales of alligator gar(Atractosteus spatula), which are composed of two distinct materi-als (ganoin and bone) [10,12,45]. The alligator osteoderm containsonly ‘‘bone’’ but undergoes gradual transitions in microstructuresat different locations due to their growth, leading to mechanicalfunctions. The advantages of comprising a homogeneous materialwith mechanical property gradient (i.e. FGM) instead of abruptchanges include better stress redistribution and enhanced resis-tance to interfacial failure [7].

4.5.2. Compressive mechanical behaviorFig. 11a and b shows the representative stress–strain curves of

the compressive tests for vertical and horizontal specimens, respec-tively, under dry and rehydrated conditions. In the dry condition,vertical samples can sustain higher stresses (142.1 ± 21.4 MPa)but eventually underwent a direct fracture in a relatively brittlemanner. On the contrary, the horizontal samples showed lowerstrengths (117.5 ± 15.9 MPa) with several stages of deformation be-fore ultimate failure, fractured in a more ductile mode. It should be


noticed that for both orientations, the ultimate strengths occurredat about the same amount of strain (�30%), meaning that bothdirections are able to suffer certain amounts of plastic deforma-tions. However, the horizontal samples yielded a much higher com-pressive strain (>40%) before totally breaking down due to theimpedance to direct failure, providing a greater toughness. Further-more, the osteoderm samples of both orientations were also testedunder rehydrated conditions in HBSS to simulate the actual envi-ronments of the biological system, where both vertical(124.2 ± 21.8 MPa) and horizontal samples (88.6 ± 12.5 MPa)showed a decrease in compressive strength. It was discovered thatin both dry and rehydrated conditions, the vertical samples exhib-ited higher strengths than the horizontal samples (dry vertical:142.1 MPa vs. dry horizontal: 117.5 MPa, rehydrated vertical:124.2 MPa vs. rehydrated horizontal: 88.6 MPa); moreover, the ver-tical samples in both conditions failed similarly in a direct andrather brittle way, whereas the horizontal samples in both condi-tions underwent multiple toughening mechanisms upon breakage(Fig. 11a and b). This implies that the anisotropic mechanical re-sponse is actually employed by the armor within biological sys-tems. The compressive mechanical properties are summarized inTable 1.

Typical fracture samples after compressive tests in both direc-tions are examined under the stereoscope, as demonstrated inFig. 11c and d. Direct crack propagation for vertical samples canbe found in Fig. 11c, which is in accordance with the relativelybrittle failure shown in the s–s curve. Horizontal samples, on



Fig. 11. Representative compressive stress–strain curves for (a) vertical and (b) horizontal samples under dry and rehydrated conditions. Stereoscopic images showing typicalfailure mechanisms in (c) vertical and (d) horizontal samples after compressive deformation.

Table 1Mechanical properties of dry and rehydrated alligator osteoderm samples inhorizontal and vertical directions (40 samples for each condition).

Direction Hydrationstate

Ultimate compressivestrength ravg (MPa)

Compressive elasticmodulus E (GPa)

Vertical Dry 142.1 ± 21.4 1.04 ± 0.11Vertical Rehydrated 124.2 ± 21.8 0.94 ± 0.15Horizontal Dry 117.5 ± 15.9 1.19 ± 0.22Horizontal Rehydrated 88.6 ± 12.5 1.07 ± 0.17


the other hand, exhibit wavy and complex fracture paths and re-veal evidence of lamellae buckling and fiber bridging, as shown inFig. 11d. These toughening mechanisms can prevent catastrophiccollapse of the whole structure and correspond to the resistingsteps before ultimate fracture observed in the s–s curve(Fig. 11b). The observation further illustrates the anisotropy inmechanical properties for alligator osteoderm.

Table 2Elastic modulus measured from nanoindentation on dorsal and ventral regions of alligatorthe elastic modulus calculated by the Bonfield and Clark equation.

Nanoindentation modulus (GPa) 10% porosity

Alligator Osteoderm (Dorsal) 20.0 16.4Alligator Osteoderm (Ventral) 13.6 11.2Human Femur 20.1 [47] –Bovine Femur 23.1 [48] –


To establish a relationship connecting the macroscopic com-pressive behaviors and the localized nanoindentation results, theelastic modulus from the two tests were compared. To begin with,the elastic modulus was first calculated from the reduced modulusreported from the nanoindentation test. The relationship betweenreduced modulus and elastic modulus can be presented as

1Er¼ 1� m2

s

Es� 1� m2

i

Ei

where Er is the reduced modulus, Es is the elastic modulus of thespecimen and Ei is the elastic modulus of the indenter tip. ms repre-sents the Poisson’s ratio of specimen and mi means the Poisson’s ra-tio of the indenter tip. For a Berkovich diamond tip, Ei is 1140 GPaand mi is 0.07 [35]. Since we are unable to determine the Poisson’sratio of the osteoderm, the Poisson’s ratio of a typical compact bone,which is 0.18 for bovine femur [46], is used to calculate the elasticmodulus. From the above equation, the elastic moduli of alligatorosteoderm from nanoindentation tests are �13.6 GPa in the ventralregion and 20.0 GPa in the dorsal region (using the average reduced

osteoderm, with comparison to bovine and human femurs, and the porosity effect on

(GPa) 20% porosity (GPa) 30% porosity (GPa) 40% porosity (GPa)

10.7 5.5 2.17.3 3.7 1.4– – –– – –



Fig. 12. Experimental demonstrations of two proposed deformation mechanisms: (a) Flexibility provided by sutures and connective fibers allows limited bending. Theneighboring plates could be bent upward to �10� and downward to �20�. (b) The major porosity in the center core of the osteoderm is able to absorb some energy atdeformations lower than �10% before cracks start to propagate in the cortex. Large channels and small voids (circled) were found to be squeezed and distorted from 0% and4% to 7% deformation.


Please cite this article in press as: Sun C-Y, Chen P-Y. Structural design and mechanical behavior of alligator (Alligator mississippiensis) osteoderms. ActaBiomater (2013), http://dx.doi.org/10.1016/j.actbio.2013.07.016


Fig. 13. Schematic illustrations of three deformation mechanisms of alligator dermal armor: (a) Sutures and non-mineralized collagen fibers provide flexibility; (b) sandwichstructure absorbs energy; (c) graded mechanical property from dorsal to ventral regions offers optimization in load redistribution and energy dissipation.


moduli), which showed comparable values with the elastic moduliof human [47] and bovine [48] femur. However, it is significantlyhigher and cannot relate to the compressive modulus. We sug-gested that it should be a result of porosity effect for macroscopicmechanical tests. By applying the Bonfield and Clark [49] equation,which is a modified version of the Mackenzie [50] equation, we canestimate the contribution of porosity to the elastic modulus:

E ¼ E0ð1� 1:9pþ 0:9p2Þ

where E is the elastic modulus with porosity, E0 is the elastic mod-ulus of the solid phase only and p represents porosity. Table 2gives the elastic modulus of bulk material at different porositieswhere the elastic modulus from the nanoindentation test is usedas E0. As the result showed, the elastic modulus of compressive


tests clearly cannot reach the calculated value, even when an over-estimated 40% porosity (the average porosity was �13% as esti-mated from the l-CT scans) is taken into account, implying thatthe alligator osteoderm is a complex composite and may incorpo-rate factors other than porosity upon mechanical responses at themacroscale.

4.6. Deformation mechanisms

Apart from the localized and global mechanical responses of themineralized bony tissues within the osteoderm, two additionaldeformation mechanisms were proposed: flexibility and sandwichstructure. Thus, simple tests were conducted to demonstrate thesemechanisms, as shown in Fig. 12. The sutures and non-mineralizedconnective fibers at the lateral edge of osteoderms provide a




limited flexibility that allows bending to some extent. As illus-trated in Fig. 12a, the connection device of alligator armor is ableto be bent upward to �10�, while the bending downward can goup to �20�. This implied that the osteoderm is designed to suffermore flexure downward, corresponding to the main function ofload dissipation for protection. On the other hand, a whole pieceof osteoderm with a cross-sectional surface revealed was subjectedto a large-scale compressive test to demonstrate the function of thesandwich structure. We focused on the deformation of the majorchannel in the center core and the surrounding small voids beforecracks started to propagate in the dense cortex, which is �10%deformation. The three successive images at the bottom half ofFig. 12b showed the cross-section at 0%, 4% and 7% strain, wherethe major channel was found to be squeezed and distorted, andwere thus able to absorb additional energy. Also, the void at thelower right became smaller as the bony plate is being compressed,possibly resulting from plastic buckling and wall collapsing of thecellular foam. These simple demonstrations illustrated that flexi-bility and sandwich structure are both incorporated in the protec-tion mechanisms of the dorsal shield of alligator.

Based on the experimental results, we summarized three defor-mation mechanisms of the alligator armor under external loads(Fig. 13). When an external compressive force acts on the osteoderm,sutures and non-mineralized collagenous fibers connecting adja-cent bony plates can dissipate loads by providing a limited amountof flexibility (Fig. 13a) [15]. The movements between neighboringosteoderms can avoid further deformation under small loads. Asthe load increases, the sandwich structure of a bony plate functionsas a preliminary route for energy absorbance (Fig. 13b). The porousinterior acts as a cellular foam, undergoing deformations such aselastic bending, plastic buckling or wall breakage to absorb a certainamount of energy. This mechanism may prevent cracking and thusmaintain structural integrity of the cortex at small strains. The thirdmechanism incorporates the structural and mechanical propertiesof the material itself by combining the dorsal and ventral cortex(Fig. 13c). The hard and highly mineralized dorsal region can sustainhigher vertical stresses as the mechanical testing indicates, and be-cause of the unique shape of the osteoderm, the loads then transferdown to the less-mineralized, more compliant ventral region. Theventral region thus suffers from stresses oriented preferentially inthe horizontal direction. The basal region consists mainly of paralleloriented collagen fibrils (lamellar-zonal bone) and possesses bettercapability to absorb energy horizontally through various toughen-ing mechanisms. In addition, the dorsal and ventral region is joinedthrough a mechanical property gradient, where interfacial tough-ness is enhanced and stress is better redistributed. The integrationof these three deformation mechanisms may lead to a synergistic ef-fect and therefore an optimized dermal armor for alligator.

5. Conclusions

Dermal armors developed in reptiles as well as some mammalsand fish are considered to be optimized for both protection andflexibility through millions of years of evolution. In the presentstudy, we investigated the structure and mechanical behaviors ofthe American alligator (Alligator mississippiensis) osteoderms atmultiple length scales. The dermal armor of alligator is a hierarchi-cally structured composite consisting of mineralized rigid bonyplates connected by non-mineralized collagen fibers. Through theexperiments, we established the structural–mechanical propertyrelationships and deformation mechanisms of the osteoderm, andeventually proposed how the dorsal shield of alligator protectsagainst external threats from the whole armor system to individualplates. The major discoveries of this study are concluded asfollows:


(1) Complex 3-D sutures and the non-mineralized connectivefibers between neighboring osteoderms provide flexibilityfor the whole armor. Strategies such as bridging and stretch-ing of the collagen fibers and interlocking of the sutures areutilized, which are similar to those observed in armadillocarapace and turtle shell.

(2) The sandwich structure of the osteoderm shows a compactcortex surrounding the porous core, enhancing bending stiff-ness and energy absorption ability with reduced weight. Theintricate 3-D network of the neurovascular system is respon-sible for the spongy interior.

(3) The osteoderm is composed of four different bone morphol-ogies, vertically from the outmost surface to the ventralregion: outer sheath (woven bone with porosity), wovenand lamellar bone, woven bone with major neurovascularcavities and lamellar-zonal bone. The varying mineral con-tents and porosity result in different localized hardnessand reduced modulus values across the osteoderm: from ahard and stiff dorsal cortex gradually transform to a morecompliant ventral base. Similar design strategies have beenapplied in various natural armors as well, implying evolu-tionary convergence for defensive functionality.

(4) Cross-sectional fracture surfaces of osteoderm through keelregion indicate various degrees of mineralization and thusdifferent microstructures in the dorsal and the ventralregions: the dorsal region is highly mineralized, showinggranular morphology and flat fracture surface while the ven-tral region is less mineralized, showing flexible and twistedfibrils. Incorporating the two regions with mild mechanicalgradient leads to the anisotropy in compressive behaviors:the vertical orientation is able to bear higher loads, whilethe horizontal orientation can absorb more energy throughmultiple toughening mechanisms including lamellae buck-ling and fiber bridging.

(5) Three deformation mechanisms are proposed for the dermalarmor of alligator: (1) the flexibility provided by sutures andnon-mineralized collagen fibers can dissipate energy undersmall loads; (2) deformations of the cellular foam interiorabsorb impact energy without cortex cracking; (3) a combi-nation of the hard dorsal region and the compliant ventralregion with graded mechanical properties offers optimiza-tion in load re-distribution and energy absorbance.

Acknowledgements

The authors gratefully thank Yu-Chen Chan, Hsien-Wei Chen, Su-Yueh Tsai and Prof. Jenq-Gong Duh (MSE Department, NationalTsing Hua University) for their support and assistant with the FE-SEM, FE-EPMA and ash-content measurements. We acknowledgeChia-Chi Chien, Bai-Hong Ke, Tsung-Tse Lee and Prof. Yeu-KuangHwu (Institute of Physics, Academia Sinica) for help with the techni-cal work and advices on l-CT scans and 3-D image reconstruction atthe National Synchrotron Radiation Research Center (NSRRC). Wewould also thank Hsi-Ming Yang, Li-Chi Hsu and Prof. Jyh-Wei Lee(MSE Department, Ming Chi University of Technology) for their helpwith nanoindentation measurements. This research is supported byNational Science Council, Taiwan (NSC100-2218-E-007-016-MY3and NSC101-2628-E-007-017-MY3).

Appendix A. Figures with essential colour discrimination

Certain figures in this article, particularly Figs. 1–13, are diffi-cult to interpret in black and white. The full colour images can




be found in the on-line version, at doi: http://dx.doi.org/10.1016/j.actbio.2013.07.016.

References

[1] Meyers MA, Chen P-Y, Lin AY-M, Seki Y. Biological materials: structure andmechanical properties. Prog Mater Sci 2008;53:1–206.

[2] Chen P-Y, McKittrick J, Meyers MA. Biological materials: functional adaptationsand bioinspired designs. Prog Mater Sci 2012;57:1492–704.

[3] Dunlop JWC, Fratzl P. Biological composites. Ann Rev Mater Res 2010;40:1–24.[4] Yang W, Chen IH, Gludovatz B, Zimmermann EA, Ritchie RO, Meyers MA.

Natural flexible dermal armor. Adv Mater 2013;25:31–48.[5] Currey JD. Mechanical properties and adaptations of some less familiar bony

tissues. J Mech Behav Biomed Mater 2010;3:357–72.[6] Ikoma T, Kobayashi H, Tanaka J, Walsh D, Mann S. Microstructure, mechanical,

and biomimetic properties of fish scales from Pagrus major. J Struct Biol2003;142:327–33.

[7] Bruet BJ, Song J, Boyce MC, Ortiz C. Materials design principles of ancient fisharmour. Nat Mater 2008;7:748–56.

[8] Lin YS, Wei CT, Olevsky EA, Meyers MA. Mechanical properties and thelaminate structure of Arapaima gigas scales. J Mech Behav Biomed Mater2011;4:1145–56.

[9] Marino Cugno Garrano A, La Rosa G, Zhang D, Niu LN, Tay FR, Majd H, et al. Onthe mechanical behavior of scales from Cyprinus carpio. J Mech Behav BiomedMater 2012;7:17–29.

[10] Yang W, Gludovatz B, Zimmermann EA, Bale HA, Ritchie RO, Meyers MA.Structure and fracture resistance of alligator gar (Atractosteus spatula) armoredfish scales. Acta Biomater 2013;9:5876–89.

[11] Song J, Reichert S, Kallai I, Gazit D, Wund M, Boyce MC, et al. Quantitativemicrostructural studies of the armor of the marine threespine stickleback(Gasterosteus aculeatus). J Struct Biol 2010;171:318–31.

[12] Allison PG, Chandler MQ, Rodriguez RI, Williams BA, Moser RD, Weiss Jr CA,et al. Mechanical properties and structure of the biological multilayeredmaterial system, Atractosteus spatula scales. Acta Biomater 2013;9:5289–96.

[13] Rhee H, Horstemeyer MF, Hwang Y, Lim H, El Kadiri H, Trim W. A study on thestructure and mechanical behavior of the Terrapene carolina carapace: apathway to design bio-inspired synthetic composites. Mater Sci Eng C2009;29:2333–9.

[14] Achrai B, Wagner HD. Micro-structure and mechanical properties of the turtlecarapace as a biological composite shield. Acta Biomater 2013;9:5890–902.

[15] Krauss S, Monsonego-Ornan E, Zelzer E, Fratzl P, Shahar R. Mechanical functionof a complex three-dimensional suture joining the bony elements in the shellof the red-eared slider turtle. Adv Mater 2009;21:407–12.

[16] Balani K, Patel RR, Keshri AK, Lahiri D, Agarwal A. Multi-scale hierarchy ofChelydra serpentina: microstructure and mechanical properties of turtle shell. JMech Behav Biomed Mater 2011;4:1440–51.

[17] Chen IH, Kiang JH, Correa V, Lopez MI, Chen PY, McKittrick J, et al. Armadilloarmor: mechanical testing and micro-structural evaluation. J Mech BehavBiomed Mater 2011;4:713–22.

[18] Rhee H, Horstemeyer MF, Ramsay A. A study on the structure and mechanicalbehavior of the Dasypus novemcinctus shell. Mater Sci Eng C 2011;31:363–9.

[19] Erickson GM, Lappin AK, Vliet KA. The ontogeny of bite-force performance inAmerican alligator (Alligator mississippiensis). J Zool 2003;260:317–27.

[20] Scheyer TM, Sander PM. Histology of ankylosaur osteoderms: implications forsystematics and function. J Vertebr Paleontol 2004;24:874–93.

[21] Alibardi L, Thompson MB. Fine structure of the developing epidermis in theembryo of the American alligator (Alligator mississippiensis, Crocodilia, Reptilia).J Anat 2001;198:265–82.

[22] Grigg G, Gans C. Morphology and physiology of the Crocodylia. In: Glasby CG,Ross GJB, Beesley PL, editors. Fauna of Australia, vol. 2A. Amphibia andReptilia. Canberra: Australian Government Publishing Service; 1993.

[23] Vickaryous MK, Sire JY. The integumentary skeleton of tetrapods: origin,evolution, and development. J Anat 2009;214:441–64.


[24] Vickaryous MK, Hall BK. Development of the dermal skeleton in Alligatormississippiensis (Archosauria, Crocodylia) with comments on the homology ofosteoderms. J Morphol 2008;269:398–422.

[25] Farlow JO, Hayashi S, Tattersall GJ. Internal vascularity of the dermal plates ofStegosaurus (Ornithischia, Thyreophora). Swiss J Geosci 2010;103:173–85.

[26] Buffrenil V, Sire JY, Rage JC. The histological structure of glyptosaurineosteoderms (Squamata: Anguidae), and the problem of osteodermdevelopment in squamates. J Morphol 2010;271:729–37.

[27] Cerda IA, Desojo JB. Dermal armour histology of aetosaurs (Archosauria:Pseudosuchia), from the Upper Triassic of Argentina and Brazil. Lethaia2011;44:417–28.

[28] Farlow JO, Thompson CV, Rosner DE. Plates of the dinosaur stegosaurus: forcedconvection heat loss fins? Science 1976;192:1123–5.

[29] Curry Rogers K, D’Emic M, Rogers R, Vickaryous M, Cagan A. Sauropod dinosaurosteoderms from the Late Cretaceous of Madagascar. Nat Commun2011;2:564.

[30] Seidel MR. The osteoderms of the American alligator and their functionalsignificance. Herpetologica 1979;35:375–80.

[31] Hutton JM. Age determination of living nile crocodiles from the corticalstratification of bone. Copeia 1986:332–41.

[32] Tucker AD. Validation of skeletochronology to determine age of freshwatercrocodiles (Crocodylus johnstoni). Mar Freshw Res 1997;48:343–51.

[33] Saalfeld DT, Webb KK, Conway WC, Calkins GE, Duguay JP. Growth andcondition of American alligators (Alligator mississippiensis) in an InlandWetland of East Texas. Southeast Nat 2008;7:541–50.

[34] Song YF, Chang CH, Liu CY, Chang SH, Jeng US, Lai YH, et al. X-ray beamlines forstructural studies at the NSRRC superconducting wavelength shifter. JSynchrotron Radiat 2007;14:320–5.

[35] Rho JY, Zioupos P, Currey JD, Pharr GM. Variations in the individual thicklamellar properties within osteons by nanoindentation. Bone 1999;25:295–300.

[36] Currey JD. Mechanical properties of bone tissues with greatly differingfunctions. J Biomech 1979;12:313–9.

[37] Kienzle E, Kopsch G, Koelle P, Clauss M. Chemical composition of turtles andtortoises. J Nutr 2006;136:2053S–4S.

[38] Chen PY, Stokes AG, McKittrick J. Comparison of the structure and mechanicalproperties of bovine femur bone and antler of the North American elk (Cervuselaphus canadensis). Acta Biomater 2009;5:693–706.

[39] Gibson LJ. Biomechanics of cellular solids. J Biomech 2005;38:377–99.[40] Seki Y, Kad B, Benson D, Meyers MA. The toucan beak: structure and

mechanical response. Mater Sci Eng C 2006;26:1412–20.[41] Bodde SG, Meyers MA, McKittrick J. Correlation of the mechanical and

structural properties of cortical rachis keratin of rectrices of the Toco Toucan(Ramphastos toco). J Mech Behav Biomed Mater 2011;4:723–32.

[42] Currey JD. The structure of bone tissue. Bones: structure andmechanics. Princeton, NJ: Princeton University Press; 2002. pp. 3-26.

[43] Oliver WC, Pharr GM. An improved technique for determining hardness andelastic modulus using load and displacement sensing indentationexperiments. J Mater Res 1992;7:1564–83.

[44] Chen PY, Lin AY, McKittrick J, Meyers MA. Structure and mechanical propertiesof crab exoskeletons. Acta Biomater 2008;4:587–96.

[45] Chen P-Y, Schirer J, Simpson A, Nay R, Lin Y-S, Yang W, et al. Predation versusprotection: fish teeth and scales evaluated by nanoindentation. J Mater Res2011;27:100–12.

[46] Pithioux M, Lasaygues P, Chabrand P. An alternative ultrasonic method formeasuring the elastic properties of cortical bone. J Biomech 2002;35:961–8.

[47] Zysset PK, Edward Guo X, Edward Hoffler C, Moore KE, Goldstein SA. Elasticmodulus and hardness of cortical and trabecular bone lamellae measured bynanoindentation in the human femur. J Biomech 1999;32:1005–12.

[48] Rho JY, Pharr GM. Effects of drying on the mechanical properties of bovinefemur measured by nanoindentation. J Mater Sci Mater Med 1999;10:485–8.

[49] Bonfield W, Clark EA. Elastic deformation of compact bone. J Mater Sci 1973;8:1590–4.

[50] Mackenzie JK. The elastic constants of a solid containing spherical holes. ProcPhys Soc B 1950;63:2–11.




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