1. Introduction

A qualitative correlation between visual appearance by reflection and structural morphology of wings’ scales of a blue Morpho Rhetenor butterfly and iridescent Anoplognathus viridis beetles’ elytron was carried out by the fourth Lord Rayleigh (Robert John Strutt) during the Twentieth Annual Exhibition of the Physical and Optical Societies [1]. His dissertation was based on analogies between optical trends observed when illuminating the biological systems (change of color with angle of incidence, for example) and already known optical properties of layered materials due to interference of light rays. In this way, the presence of layered structures in these biological systems was inferred although in fact without a detailed description of them. The basic idea was previously suggested by Hooke [2] and Newton [3] when considering the changeable colors of some birds’ feathers, and followed by the third Lord Rayleigh (John William Strutt) [4]. What began with qualitative correlations has been carried out towards quantitative levels involving photometric optical measurements, color analysis, radiative transfer models, and electron microscope images to characterize the structural morphology of the biological nanostructures. Nowadays we call this research field structural color.

In recent years the number of publications focusing on reflection properties of natural photonic systems has increased significantly due to the desire to understand in each case the correlation between main features of measured reflection spectra and internal morphology of the samples considered [5]. Samples have been taken from the exoskeleton of scarabs, bird feathers, scales of butterfly wings or spiders, and from scales and corneas of fishes [6–10]. The plant world also shows some examples of coloration arising from structural arrangements of organic materials [11, 12]. A review of the different biologically structured systems considered during the last years is included in the book of Kinoshita [13], and in the paper of Yu et al. [14]. Both multilayer structures and more complex tridimensional photonic crystal arrangements have been considered in the search to explain the observed colors and measured reflection spectra [15, 16]. Successful correlations between microstructure and visible light reflection open the possibility of synthetizing similar structures for specific applications [17]. Samples have been studied under non- or circularly polarized incident radiation, with chitin as their major constituent when they were taken from animals and cellulose in the case of samples from plants. In this work we consider measured reflection spectra of non-polarized, diffuse or linearly polarized light by the dorsal surface of Chrysina aurigans scarabs and their correlation with the morphological structure found by means of Scanning Electron Microscope (SEM) analysis, throughout the procuticle of these scarabs. The contribution to the reflected light from diffuse radiation reflected by a thick layer of pigmented micron-sized structures, located beneath the chitin-protein procuticle, is established from optical transmission microscope, and from additional optical measurements carried out with an integrating sphere. The reflection measurements are carried out in a spectral range which covers near ultraviolet, visible, and near infrared wavelengths. The left handed circular polarization of the light reflected by the C. aurigans’ cuticle has also been determined.

2. Sample characterization and measurements

This section is focused on describing the morphological properties of the biological samples considered in this study. The different optical measurements performed are described in section 3, where the appearance of C. aurigans scarabs under different illumination conditions is qualitatively analyzed in order to acquire knowledge about the physical mechanisms contributing to the reflected light and the corresponding visual appearance of the scarabs’ cuticle.

2.1 General aspects about Chrysina aurigans scarabs

The so called “metallic” scarabs of the genus Chrysina are beetles about 20 – 40 mm long [18] which can be found in locations from Southern United States to Northwestern South America [19]. Chrysina aurigans [20] is a species mainly reported from Costa Rica [21], where it has been found on the slopes of the Caribbean side of the Guanacaste, Tilarán, Central and part of Talamanca mountain ranges, at elevations between 600 and 1250 m (see Fig. 1(a)), according to the database of the National Biodiversity Institute (Instituto Nacional de Biodiversidad: InBio) [22]. These scarabs display a striking golden-like appearance (see Fig. 1(b)). They live in the canopy in places that still retain its natural forest, where they feed on the leaves of the trees. As adults they are nocturnal, resting during the day in the foliage of the treetops. After mating and copulation, the females descend to the forest floor to locate trunks of dead trees where they lay their eggs. The larvae develop inside the trunk feeding on decaying wood. Figure 1(b) shows a picture of a C. aurigans specimen indicating the different sections of its elytron.

 

Fig. 1 (a) Distribution through the territory of Costa Rica of those regions, indicated by the contour defined by the blue line inside the map, where specimens of C. aurigans scarabs have been collected. (b) Chrysina aurigans scarab with labeled sections of its elytron: (1) the scuttelum, (2/3) left/right dorsal-lateral areas, and (4/5) left/right posterior-lateral sections of the elytron. The RGB values just above the point indicated by the symbol “x” are R = 210, G = 173, and B = 25. The Adobe Photoshop^® tools were used to determine these R, G, and B values, by assuming a D65 illuminant.

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2.2 Microscopy of the surface and cross section of the cuticle

Figure 2(a) displays a section of a locally flat surface in the elytron of a C. aurigans scarab. The presence of micron-sized close packed scales on the surface of the elytron, with sizes close to 10 μm, is observed. These scales are part of the epicuticle which consists of a thick waxy layer. For comparison purposes we have included in Fig. 2(b) a picture of the surface which corresponds to another beetle of the same genus Chrysina, the green C. aurora scarab, which shows a random distribution of rather circular cells over its elytron’s surface. In an initial interpretation of the SEM images, the flat surface of C. aurigans is correlated with a morphological structure consisting of a sequence of flat layers located beneath the epicuticle while the corresponding structure for the C. aurora consists of concave layers. A flat or concave layered structure has great influence on the appearance of the cuticle and on their corresponding optical properties [23]. Now we are reporting on reflection spectra of flat structures and concave multilayered arrangements will be considered in the near future.

 

Fig. 2 Close-up view of the surface of (a) C. aurigans´ and (b) C. aurora’s elytra covering a surface of 100x100 μm² in both cases. The reflective layers of C. aurigans shows a locally rather flat surface structured from close packed scales, while the elytron’s surface of the C. aurora shows micron sized cells well differentiated from each other.

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To obtain the SEM images, sections of the elytra from specimens of this golden-like scarab were cooled with liquid nitrogen and freeze-fractured using a razor blade. The fractured samples were coated with Pt-Pd. The edges of the fractured specimens showing vertical sections through the cuticles were examined using Hitachi Models S-570 and S3700N Scanning Electron Microscopes.

Figure 3 displays cross sections of the cuticle showing what resembles sequences of layers whose thicknesses change with depth. A small section of the elytron’s surface is displayed on the top of Fig. 3(b), showing a very smooth outer surface. The epicuticle is a homogenous layer whose thickness is about 0.8 μm. A sequence of around 94 thick layers is seen through the procuticle with thicknesses changing between 150 and 275 nm up to 16 μm in depth. On initial observations, what looks like thin layers are displayed between thick layers, with thicknesses around 20 nm and showing a small thickness variation with depth through the procuticle. In a previous work we assumed the presence of a chirped structure in the cuticle of C. aurigans scarabs [6]. The current SEM analysis shows a more complex structure rather than a simple chirped layered system which is characterized by decreasing thickness with depth.

 

Fig. 3 SEM images of the C. aurigans’ cuticle cross section showing a sequence of about 94 thick layers through the procuticle. The white bars at left and right figures correspond to 10 and 5 μm respectively. The outer surface of the cuticle is: (a) above on the right, and (b) at the top.

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3. Reflection of collimated light normally incident on the cuticle

Direct reflection spectra were taken from C. aurigans’ cuticles normally illuminated with non-polarized light (300 to 1100 nm in wavelength) on, at least, 4 different places in the dorsal elytron (see Fig. 1(b)). To decrease as much as possible the effect of the elytron’s curvature, and to approach perpendicular incidence and repeatability, a device was constructed to allow motion in the axial and azimuth angles as well as proper positioning and distance adjustment of the fiber optic probe at any point on the elytron (see Fig. 2 in [6] and linked multimedia video). The spectra at the various sites show very similar features but differ in their intensities, probably due to a remaining sensitivity of the reflection measurements to the local curvature of the elytron. Measurements were made using a fiber optics spectrometer (AvaSpec 3648) and a Halogen Deuterium lamp (AvaLight DHc). A specular reflectance standard of aluminum (Ocean Optics STAN-SSH) was used for normalization. Figure 4 depicts typical reflection spectra when the elytron’s surface of a specimen was normally illuminated with non-polarized radiation. As seen, a well-defined broad reflection band is displayed with a width of about 500 nm and an average reflection around 30%, a pronounced reflection edge close to 525 nm, and low reflection values (R_o = 3%) through short wavelengths. We attribute this reflection background R_o to the epicuticle, which is transparent to visible light at longer wavelengths. R_o = 2.5% in Fig. 4(a) and it is 4% in Fig. 4(b). From the measured values of R_o and the relation: R_o = ((n_w-1)/(n_w + 1))² where n_w is the refractive index of the waxy epicuticle layer, the average value of the refractive index in the visible range is determined between ${\overline{n}}_{\text{w}}\simeq 1.38$ and ${\overline{n}}_{\text{w}}\simeq 1.50$. The lack of structure in the R_o spectrum in the near ultraviolet and through short visible wavelengths means that this waxy layer could act as an absorbing and scattering layer. The epicuticle is made of waxes hardened through an oxidative crosslinking process which incorporates oxygen atoms in the form of organic functional groups that can be responsible for ultraviolet light absorption with tails at short visible wavelengths. A similar protective layer is found in plants’ leaves [24] where the light absorbing character has been attributed to the presence of cuticle waxes [25]. The low reflection values at short wavelengths means that under illumination with white light, most of it is transmitted through the procuticle of the C. aurigans, including bluish and reddish colors.

 

Fig. 4 Measured reflection spectra under normal incidence of non-polarized radiation taken when illuminating the elytron’s surface of a C. aurigans beetle at (a) its head, and (b) in section 2. The background reflection R_o from the epicuticle is shown. The thin solid lines are smoothed curves obtained from the experimental values. The colors of the squares showing the values of x, y, Y, R, G, and B are determined by the same values of these parameters.

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In a previous work this broad reflection band was partially characterized in the wavelength range from 300 to 750 nm [6]. Now we report new measurements taken over a broader spectral range. As seen in Fig. 4, the reflection band shows a ripple structure: a sequence of maxima and minima for wavelengths larger than around 600 nm. About 20 reflection peaks are displayed with a wavelength separation between successive peaks of $14\pm 1$ nm. In the near infrared, beyond 900 nm, the reflection values decrease gradually with wavelength. The peaks superimposed to the reflection band show some modulation which could be attributed to a diffuse background reflection from a structured layer containing pigments, located beneath the procuticle (see thin solid lines in Fig. 4). Other mechanisms contributing to modulate the fine structure of the reflection band will be mentioned later. About 57% of the reflected light corresponds to infrared wavelengths, and it could be used by the scarabs as camouflage between green leaves against nocturnal predators capable of detecting infrared light [26, 27].

We have included in Fig. 4 the chromaticity coordinates (x,y), the relative luminance value (Y) and the RGB parameters corresponding to the displayed measured reflection spectra. They determine the color of the background squares were their values are displayed. The evaluation of the first three parameters requires the spectral irradiance of the illuminant which we have assumed as the AM1.5 solar spectrum ($I_S$) [28], and of the color matching functions ($\overline{x},\overline{y},\overline{z}$) specified by the Commission Internationale de l’Eclairage: CIE [29]. In the CIELab color space, the stimulus Y is related to the luminance $L$ through the expression $L=116{(Y/Y_n)}^{1/3}-16$ with Y_n as a normalized reference value. L gives an approximation of the color brightness, i.e. L and Y are correlated with the attribute which depends on the amount of light reflected by the surface [30]. The explicit expression to evaluate the parameter Y from a reflection spectra R(λ) is given by

3.1 Diffuse reflection by a thick layer of pigmented structures

An integrating sphere device was attached to the spectrophotometer to measure total and diffuse reflection when a small section of the C. aurigan’s cuticle was normally illuminated with collimated non-polarized light, for wavelengths between 450 and 800 nm. A white barium sulfate standard was used as reference. Spectral reflection measurements were also carried out after removal of the multilayer structure by sanding and illuminating the sanded surface with normally incident collimated radiation. In this way we obtain the collimated-diffuse reflection spectrum (R_cd) arising from what seems to be a thick layer of structures containing micron sized pigments located beneath the procuticle. This layer was displayed during analysis based on optical transmission microscope which shows the presence of a reddish micron-sized structure consisting of cross-linked thick fibers. These structures probably are the result of a sclerotization process and the presence of pigments through them is expected [32]. The collimated-collimated contribution (R_cc) can also be obtained when subtracting the diffuse component from the total reflection (R_cc + R_cd). Figure 5(a) shows the results: some amount of diffuse reflection is displayed. For wavelengths larger than 650 nm, the measured total reflection as well as the collimated-diffuse component increase with wavelength, which is consistent with the presence of micron-sized pigments. Another fact consistent with the presence of pigments in the bottom of the procuticle is shown in Fig. 5(b): when the cuticle is illuminated with white light, reddish light is transmitted through some sutures in the ventral side of the scarab. The bluish colors of the white light transmitted through the procuticle are extinguished probably due to absorption and scattering by the micron sized pigments. This method of sanding the cuticle in a progressive way, until its characteristic color disappears to be replaced by red, brown or black colors, was applied by Neville and Caveney to infer the presence of a pigmented layer at the bottom of the structure producing the light interference effect giving the optical appearance of the cuticle [33].

 

Fig. 5 (a) Integrating sphere measurements of total (R_cc + R_cd) and diffuse (R_cd) reflection when illuminating a section of the C. aurigans cuticle with normally incident non-polarized light. The R_cd spectrum corresponds to measurements taken after removal of the procuticle by grinding, and R_cc values are obtained from the difference between R_cc + R_cd and R_cd. (b) Incidence of white light on the cuticle of a C. aurigans specimen, and transmitted reddish light emerging through sutures in the ventral side of the scarab.

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3.2 Reflection of diffuse incident light and circular polarization

Other measurements carried out by illuminating the cuticle of C. aurigans specimens with non-polarized light involved the use of polarizers to detect the circular polarization of the reflected light. A diffuser filter (a parchment paper) was adapted to the light source so that the illuminating beam was as similar as possible to ambient atmospheric sun light, i.e. the sample is illuminated with diffuse light similar to that of the cloud forests where these scarabs inhabit (see Fig. 4(a) in [34]). Resulting measurements proved to be highly repeatable. Measurements were made using a fiber optics spectrometer (AvaSpec 3648) and a quartz-halogen vibration-less light source at a color temperature of 3100 K (Dolan Jenner Fiber-Lite, Model 190). A quarter wave plate followed by a linear polarizer were used as a polarization analyzer in order to determine the circular polarization of the reflected light. The quarter wave plate was rotated at either 45° for right handed polarized light or 315° for left handed polarized light, instead of 355° as erroneously indicated in a previous publication [34]. In this way we assembled the polarizers for left handed and right handed circularly polarized light, PLHCPL and PRHCPL respectively.

The measurements of selective reflection of circularly polarized light by the cuticle of many species of scarabs is a well-known phenomenon [35,36], and this has been confirmed in our measurements, as shown in Fig. 6 where a dominant reflection band of left handed circularly polarized (LHCP) light is displayed. In a previous work we reported measurements based on illuminating the scarab’s cuticle with left handed or right handed circularly polarized light, in order to determine the polarization of the reflected light. The conclusion is consistent with a left handed circular polarization of the reflected light (see Section 4 in [6]). This selective reflection effect was first reported by Michelson in 1911 when he considered the brassy appearance of C. resplendens scarabs [37]. A correlation between this optical property and the structure of the cuticle was first developed by Neville and Caveney who interpreted Transmission Electron Microscope (TEM) images displaying white and dark layers in terms of a twisted helical structure similar to those characterizing cholesteric liquid crystals [33]. With better TEM magnification, the helical structure was revealed much more clearly (see Fig. 2 in [38] and also Fig. 3(b) in [39]). These structural arrangements, so common in natural systems, are now called Bouligand-type structures in recognition of the work done by Ives Bouligand and his very significant contributions in the field of morphogenesis of living organisms [40]. In this case they consist of protein-chitin nano-fibrils whose azimuth orientations change with depth through the cuticle until completing several cycles (see Fig. 6(a) in [34]).

 

Fig. 6 Reflection of left handed circularly polarized (LHCP) and non-LHCP light when illuminating the cuticle of a C. aurigans with diffuse light. Both spectra have been normalized to a total reflection value close to 3% at short wavelengths, which is the average value corresponding to the two spectra displayed in Fig. 4 in this spectral range. The lines display smoothed curves only for visual aid, and dots are measured values.

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The presence of a dominant left handed circularly polarized reflected light band requires of interpreting the structure displayed by the SEM images of Fig. 3 in terms of a left handed chiral Bouligand-type structure: the dark layers correspond to planes where the protein-chitin nano-fibrils are parallel to the cut of the cuticle. There is a difference of 180° in the azimuth angle specifying the orientation of the fibrils, for two successive dark layers. This is the same assumption introduced by Neville and Caveney as above mentioned, and followed by other researchers [41–43]. The diffuse character of the incident light suppresses the spectral ripple structure observed when illuminating the cuticle with collimated non polarized radiation. The presence of this ripple structure in the optical measurements reported in Fig. 4 indicates that the Bouligand structure in the cuticle of a C. aurigans scarab is characterized by a depth-dependent pitch distribution more complex than those previously reported in the literature for other biological systems (see Fig. 6 in [34]) also characterized by selective reflection of circularly polarized light. This idea is consistent with the observed variation of the layers thickness through the cuticle, as shown in the SEM images.

According to the Bragg conditions, for a twisted structure with a single spatial period or pitch P_o, a reflection peak is displayed in its spectrum, centered at wavelength λ = nP_o where n is the average refractive index of the anisotropic structure, and by assuming normal illumination. The presence of a pitch step in the structure leads to two dominant peaks in its reflection spectrum [41]. The successive peaks displayed in the reflection spectra of the C. aurigans’ cuticle are consistent with the presence of a depth-dependent pitch distribution. As mentioned, the thin dark layers displayed in Fig. 3 correspond to those planes were chitin nano-fibrils are parallel to the cross section of the cuticle once cut. A half pitch is completed every two thin layers, from the middle of the first to the middle of the next one, with a thick layer between them [41–43]. Consequently the thickness variation of the thick layer with depth is directly related to a depth dependence of the pitch characterizing the left handed chiral structure. This modulation in the pitch of the structure is responsible of the spectral ripple structure superimposed to the broad reflection band. The depth dependence of the pitch displays increasing values beyond the exocuticle, through the endocuticle, which is responsible of the background reflection band, as seen in the reflection spectra of cholesteric liquid crystal circular polarizers [44]. The presence of the ripple structure in the reflection spectra, when illuminating the cuticle with non-polarized light, has not been reported for other living organisms.

At short wavelengths the intensity of the light reflected through the PLHCPL and PRHCPL shows similar structure-less spectral behaviors. This probably corresponds to the previously mentioned reflection background from the waxy epicuticle. Through the red part of the visible spectrum the intensity of the LHCP light is about twice of that measured with the PRHCPL. This selective character has been correlated with the presence of a left handed helical or twisted plywood structure through the cuticle of the beetles [45], similar to that found in cholesteric liquid crystals (CLCs) [46]. The presence of some reflected non-LHCP radiation, which contributes with a brownish orange coloration of the cuticle, could be an indication of some contribution from non-polarized, non-coherent, or diffuse light. The role of the circular polarizers consists of suppressing one of the two circular polarization components. If the reflected light contains diffuse, non-polarized or non coherent light, about 50% of this contribution is transmitted through the polarizers with a further decrease due to absorption by the polarization system. Some jewell scarabs like C. aurigans have a remarkably golden shine that is believed to actually provide cryptic coloration as a defense against diurnal predators such as birds when they stand still and reflect the surrounding foliage. On the other hand, reflections from Chrysina scarabs’ cuticle mainly consist of left-handed circularly-polarized light, and there is currently much interest in knowing if they are able to detect it as a means of signaling to members of their own species [47–49]. The reflection thus might do double-duty making them invisible to predators but highly visible to members of their own species [50, 51]. We have shown that C. aurigans scarabs have a broad band of reflection that extends into the near infrared. As these scarabs are nocturnal, it would be very interesting to see if their very light-sensitive eyes are also able to detect red and near infrared light. In most rhodopsin-based visual systems heat interferes with detection of low light levels in this region [52]. However, a chlorophyll-based visual system that allows long-wave detection by some fish has recently been discovered [53, 54], and the possibility that some nocturnal insects can use this spectral region for navigation or intra-species communication should be investigated further.

3.3 Scarabs’ images based on contribution of different reflection mechanisms

The predominant left handed circular polarization of the light reflected by C. aurigans’ cuticle can be easily shown by taking photographs using circular polarizers located between the scarab and the camera. These polarizers absorb about 70% of any non-polarized or diffuse light contained in the radiation reflected by the cuticle. Figure 7 shows pictures taken with circular polarizers that can be compared with that obtained with no polarizer. The luminance values indicated in the figures were obtained by using Adobe PhotoShop^® tools, with the gray color of the surface supporting the beetle as reference, whose normalized brightness is Y_n = 64.6. From each L-value, the corresponding Y one is obtained. This parameter, Y approaches the reflectance factor of a surface for wavelengths in the middle of the visible wavelength range, where the matching function $\overline{y}$ reaches their maximum values [30]. The two spectra displayed in Fig. 4 are examples of this relationship between the brightness parameter and the reflectance spectrum. A first comparison between Figs. 7(a) and 7(b) indicates that the polarization of the reflected radiation is mainly left handed, and the color of the scarab is golden or yellowish. The intensity of the LHCP reflected light shows the largest values for wavelengths close to 600 nm, as shown in Fig. 6. When the PLHCPL is used, as in Fig. 7(b), the scarabs appear more brilliant with the contour of the beetle appearing more greenish. The appearance of the contour is determined by light rays at grazing angles of incidence. It is then expected that the reflection band for LHCP reflected light moves to shorter wavelengths giving this greenish yellow color to the contour of the scarab. When the PRHCPL is used, and according to measurements reported in Fig. 6, the non-LHCP component has its largest values for wavelengths close to 675 nm (the red side of the visible spectrum), which is consistent with the appearance of the scarabs displayed in Fig. 7(c). If the non-LHCP component of the reflected light is negligible, the general appearance of the beetle should be blackish (see Fig. 2 in [55] for example) instead of brownish orange as shown in Fig. 7(c). The reflection band for non-LHCP shown in Fig. 6 will also move to shorter wavelengths at grazing angles of incidence. The contour of the scarab in Fig. 7(c) should look yellowish orange in the presence of some amount of non-LHCP component in the reflected light, as is the case. Now we discuss the nature of the light giving the brownish orange color of the beetle, as displayed in Fig. 7(c). By taking as base background the gray color of the surface supporting the scarab, a semi-quantitative comparison of the luminance values (brightness) reported in the caption of this figure (Y_c < Y_d < Y_a < Y_b) suggests that the reflected radiation spectrum, labeled as non-LHCP in Fig. 6, corresponds to light non-polarized, non-correlated or diffuse light. The inequality Y_b > Y_a would be less significant with no additional absorption from the polarizer. If the non-LHCP radiation is correlated to right handed circularly polarized light we must have that Y_a > Y_b. In Fig. 7(d) the contribution of non-circularly polarized components is canceled by subtracting image (c) from (b). The fact that Y_a-Y_d = 3.6, and remembering that due to absorption only about 30% of the non-circularly polarized components reach the camera’s detector, this means that the contribution of the non-LHCP reflected (non-polarized, non-correlated, or diffuse) light is close to 12%.

 

Fig. 7 (a) Picture of C. aurigans’ scarabs taken with no circular polarizer (CP). (b and c) Photographs taken with the use of a CP for left handed and right handed circularly polarized light located between the camera and the beetle, respectively. (d) Digital subtraction of images [(b)-(c)]. For the position indicated by the symbol “x” in Fig. 1(b), the corresponding luminance (luminous reflectance) values are: L_a = 63 (Y_a = 20.4), L_b = 79 (Y_b = 35.5), L_c = 24 (Y_c = 2.6), and L_d = 58 (Y_d = 16.8).

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A final experiment corroborates our interpretation that the brownish orange color of the scarab displayed through a PRHCP light is due to non-polarized, non-correlated or diffuse light reflected by the beetle’s cuticle, with the yellowish color due to LHCP reflected light. It consists of illuminating the scarab with linearly polarized light and on taking a picture of the beetle with a linear polarizer between the camera and the reflecting surface. The results are displayed in Fig. 8. From the same source, two orthogonal components of incident light are selected to illuminate the surface of the scarab. These two components define corresponding states of linearly polarized incident light. Each one of these states can be considered as a superposition of in-phase left and right handed circularly polarized waves. A left handed twisted plywood structure is transparent to the RHCP components. The LHCP components corresponding to the vertical and horizontal polarized waves are also mutually orthogonal, i.e. they are out of phase by 90 degrees. The left handed twisted structure acts as a filter for these two LHCP components. It is transparent for one of the LHCP component and the other is reflected. For the linear polarization whose LHCP and RHCP components are transferred through the twisted structure (photographs (a) and (b) in Fig. 8), the color of the scarab will be determined by additional reflection of non-polarized, non-correlated or diffuse light. This could be due to the presence of a deep optically isotropic and non-correlated section in the cuticle’s structure and/or a pigmented thick layer beneath the procuticle, respectively. For the linear polarization whose LHCP component is reflected by the left handed helical structure and the RHCP component is transferred (photographs (c) and (d) in Fig. 8), the color of the scarab will be mainly determined by the LHCP reflected light. The approximate differences in the luminance values are due to the diffuse background emerging from a pigmented structured layer or from others optical mechanisms contributing to the reflection spectrum.

 

Fig. 8 Photographs of a C. aurigans scarab taken with incident linearly polarized light and with a linear polarizer between the illuminated cuticle and the camera, and according to the following setups: (a) vertical polarization (NS) and horizontal polarizer (EW), (b) vertical polarization and vertical polarizer, (c) horizontal polarization and horizontal polarizer, and (d) horizontal polarization and vertical polarizer.

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4. Summary and conclusions

The procuticle of C. aurigans scarabs is a natural broad band reflector whose reflection spectra are very similar to those characterizing broad band cholesteric liquid crystal polarizers. The outstanding features of the measured reflection spectra consist of a broad reflection band of left handed circularly polarized light whose width approaches 500 nm, with average values close to 30%, and displaying a ripple structure which consists of a sequence of narrow peaks whose spacing in wavelengths is almost constant. This property of the reflected light implies the presence of a twisted or helical structure through the procuticle of the C. aurigans characterized by a left handed chirality, with a spatial period or pitch dependent on the depth through the optically anisotropic structure. The visual appearance of C. aurigans scarabs is determined by two well differentiated contributions: a main structured broad reflection band of left handed circularly polarized radiation displayed for wavelengths between 525 and 950 nm, and an additional radiation which arises from three optical mechanisms: around 3% of the reflected light comes from the epicuticle, a small fraction corresponds to diffuse radiation due to light scattering and absorption by a micron-sized pigmented layer located beneath the procuticle, and up to around 8% of the reflected light could be due to the presence of defects through the twisted left handed procuticle. These three mechanisms contribute with an amount of reflected light which agrees with the estimation carried out in Section 3.3 from brightness values and by subtracting the contribution of the left handed circularly polarized light, i.e. they contribute with about 12% of the reflected light, and around 17% corresponds to left-handed circularly polarized radiation.