1. Introduction

Origin of physical coloration in living things has always been fascinating. It could involve diffraction, interference and scattering instead of pigments for color production [1–3]. Iridescent colors in animals and insects usually arise from selective reflectance of incident light due to unique photonic structures of their optical reflectors. Such color effect has been studied in some bird feathers, butterfly wings and beetle cuticles [2,3]. Through these studies, the nature itself has demonstrated the efficiency of physical coloration via nano-structures, which inspires other research in designing and creating photonic structures in nanotechnology [4,5]. With present nano-fabrication techniques, the replication or resembling of the natural structures has expanded from simple one-dimensional (1-D) multilayers [6] to complex systems. For example, diamond-based woodpile structure could provide wide photonic band gap [7,8] and biomimetic replicas of chiral films could give the circular polarization same as the cuticle of the beetle [9].

Coleoptera represents a large group of insects demonstrating their iridescence arising from physical nature of photonic structures. It has been known that the exocuticle of the insects, which is referred as the optical reflector, is responsible for the color effect. Peoples have been studying a number of beetles for their optical and structural properties and making progress in discovering their unique structures and mechanisms of coloration [10–14]. Neville and Caveney suggested that optically active scarabaeid beetles, particularly the outer exocuticles, behave as an optical analogue of cholesteric liquid crystals (CLCs) although these two do not bear the similarity in their chemistry and physical states [10]. The chirality of the outer exocuticles of the beetles (Coleoptera: Scarabaeidae) is due to the pitch of their helicoid structure and the material composing the structure.

The elytra of Rhomborhina Gigantea (R. Gigantea), scarabaeidae, appear bright metallic green (Fig. 1).Such iridescence, like many other beetles, originates from the optical reflector which possesses nano-structures within the exocuticle of the elytra or other parts of their bodies. The elytron of R. Gigantea was examined for its optical properties with spectral analysis. Electron microscopy was used to examine the cross-section of the elytron to verify the presence of multilayer reflector within the exocuticle. The multi-layer would act like 1-D photonic crystals which give rise to the metallic green appearance. Further experiments were performed to determine average index of refraction and half-pitch (i.e. layer thickness) of the layers. The observed layer thickness agreed with the theoretically obtained. The results show that R. Gigantea has chiral property as in CLCs just as other beetles with similar optical reflectors [15].

 

Fig. 1 Rhomborhina Gigantea (scale bar: 1 cm).

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2. Theory

Scarabaeid beetles are similar to CLCs optically, such as selective reflection of left-circularly polarized light and brilliant metallic appearance [10]. All of these optical properties resemble to that of CLCs, which have been studied comprehensively since the discovery of liquid crystalline phases [16,17]. The system of CLCs yielding a peak reflectance (${\lambda }_{\max }$) at normal incidence can be described by

3. Experiments

A flat and smooth region (3 mm x 5 mm) was dissected from the elytron of R. Gigantea (Fig. 1) for investigating its optical properties. The nano-structures of the exocuticle were observed using scanning electron microscopy (SEM).

The optical investigation of the sample included two parts: diffraction and reflection. The experiments were performed using the angle-resolved measurement setup to obtain wavelength-dependent spectra at varied angles. The light source was a Xenon lamp providing white light as an imitation of the natural light. In the diffraction experiment, unpolarized light was illuminated at normal incidence of the sample. The diffracted light was collected from the normal of the sample with angles 20° – 70° by a monochromator where the signals were amplified with a photomultiplier tube. Each set of optical data was taken for every 10° in the range of diffracted angles from 350 nm to 750 nm. In the reflection experiment, incident light with transverse electric (TE) and transverse magnetic (TM) polarizations were used to investigate the optical response of the beetle exocuticle. The angle of incidence ranged from 10° to 60°, data was recorded similar to that of diffraction experiment.

4. Results and discussions

4.1 SEM images of the cuticle

The cross-section of the elytron of R. Gigantea was imaged by SEM (Fig. 2).The inset of Fig. 2(a) is an enhanced image of the dash-lined region showing the nano-structures within the elytron, in which layered structures could be observed. From Fig. 2(a), the structure was not a simple 1-D multilayer but rather with more complex features, which made the photonic structure more complicated. In fact, the structure was basically formed by layers parallel to the outer surface and an array of perpendicular ridges supported by vertical columns (diameter ~0.55 µm).

 

Fig. 2 (a) SEM image of the elytron of R. Gigantea and an enhanced image (inset) of dash-lined region (scale bar: 2 μm); and (b) SEM image showing the layer structures of the elytron partially removed by focused ion beams (scale bar: 1 μm).

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To confirm the onset of the layered structures starts from the surface of the elytron, a small region (10 µm x 10 µm) of the surface was etched away using focused ion beams for cross-section imaging (Fig. 2(b)). The image was taken at a tilt angle of 30° from the normal plane of the sample. The lamellar spacing was approximately 0.168 µm though this image did not reveal any further fine structures.

R. Gigantea belongs to the large family Scarabaeidae. The exocuticles of several beetles belonging to this family had been studied [10,13,18,19]. The fact that the cross-section of the elytron of R. Gigantea (Fig. 2(a)) resembled to that of Dynastes Hercules [18], which is also a species of the family Scarabaeidae, implicates that R. Gigantea could also possess optical chirality analogue to CLCs.

4.2 Spectral analysis

All reflection and diffraction spectra were expressed as the ratio of the intensity of light reflected off the sample to that of the incident light source. Figure 3 shows the diffraction spectra for diffracted angles from 20° to 70°. Distinct peaks are situated within the green color of visible range. In fact, the diffraction due to the elytron of R. Gigantea was very pronounced, i.e. an intense diffracted green light could be observed without visual aids. This effect can be attributed to optimal Bragg condition such that the lamellar spacing acting as an optical reflector for R. Gigantea correlating with the wavelength of green visible region.

 

Fig. 3 Diffraction spectra of the elytron of R. Gigantea.

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Figure 4 depicts the reflection spectra of the elytron of R. Gigantea in TE and TM polarization respectively. The reflection peaks in green visible region experienced a blue-shift with increasing incident angles. Other reflection peaks can also be observed, however, they are independent of incident angle. It is worth noting that the reflection peaks were due to the nano-structures within the elytron of R. Gigantea. Since it is the parallel layers perpendicular to the incident light responsible for the angle-dependent peaks with respect to the varying incident angles [20], it is certain that the nano-layered structures as shown in Fig. 2 are responsible to the physical coloration of R. Gigantea (Fig. 1). Other reflection peaks, on the other hand, are believed to be due to the ordered ridges and the columnar supports as shown in Fig. 2(a), which were not sensitive to the incident angles in the reflection measurement setup.

 

Fig. 4 Reflection spectra in (a) TE and (b) TM polarizations of the elytron of R. Gigantea.

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The angle-dependent reflection peaks in green visible region at TM mode coincide with those at TE mode (Fig. 5).The reflection peaks in green color region denotes a typical light reflected off a simple layered nano-structure where peak wavelength varies with incident angle and that the reflection peak is associated with the physical thickness of the parallel layers. This phenomenon can also be found in Papilio butterflies [21], which possess 1-D photonic structures. However, for the elytron of R. Gigantea beetle the overall relative intensity increased with angle of incidence at TE mode while at TM mode the intensity dropped when the incident angle becomes more oblique (Fig. 4). As a reference, reflection measurements were also carried out on the wing of Papilio Ulysses, one of the Papilio butterflies. For typical multilayer as in Papilio Ulysses, the overall relative intensity of the reflections in both linear polarizations was either increase or decrease with angle of incidence. Compared the results, the opposite trends in TE and TM polarizations and the lack of single dominant reflection peak indicate that the optical reflector of R. Gigantea is not just a simple multilayer. In fact, the intensity of reflection would vary when a linearly polarized light reflects off an optically active, or chiral, material at different incident angles, and this further supports the presence of chiral structure.

 

Fig. 5 Comparing peak wavelengths in green color region of the reflection spectra in TE and TM polarizations.

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4.3 Determination of half-pitch and average refractive index

Using Eq. (3), n_av² was determined by extrapolating the linear-fit of the optical data in Fig. 5 to obtain the y-axis intercept while $-1/P_o{}^2$ was the slope of the linear-fit (Fig. 6).For TE polarization, n_av = 1.61 and P_h = 0.178 μm. For TM polarization, n_av = 1.73 and P_h = 0.162 μm. As observed from Fig. 2(b) the half-pitch was about 0.168 µm, which was comparable with the calculated values using the optical data obtained from reflection experiments. It confirms the structural and optical results and also verifies the correlation between the structure of the 1-D photonic crystals and CLCs. The refractive index of the cuticle layers for most scarabaeid beetles which had been experimentally obtained is about 1.5 – 1.6 [10], while some may have a value of 1.7 in the presence of uric acid in the cuticular reflector [19]. The experimental results agreed well with the reports.

 

Fig. 6 Linear-fit of peak wavelengths in green color region of the reflection spectra in (a) TE and (b) TM polarizations respectively.

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5. Conclusions

Spectral and structural analyses were carried out to investigate the chirality of R. Gigantea. Diffraction experiments showed distinct peak reflections in green color region when Bragg condition was fulfilled. Reflection experiments in TE and TM polarizations resulted in contrasting trends of overall relative intensity with respect to varied incident angles. When incident angle increased, the overall relative intensity increased at TE mode but dropped at TM mode. The results suggested that the structure of the optical reflector of R. Gigantea was not simply 1-D multilayer. SEM images revealed periodicity within the exocuticle implying the optical reflector of R. Gigantea. The cuticular reflector was similar to other beetles in the same family regarded as an optical analogue to CLCs. Calculation was carried out to determine n_av and P_h of the reflector medium. The half-pitch P_h obtained from the optical data agreed with the result deduced from SEM image. Also, the calculated n_av were close to the reported values of other beetles. These results indicated that the optical reflector of R. Gigantea having chiral structure is also an optical analogue to CLCs.