Compressed zip folder containing two tables and nine figures.

Table A. Table with sample details for Clanculus margaritarius A & B, C. pharaonius and Calliostoma zizyphinum. Specimen number, sampling locality, analyses undertaken in this study, and corresponding photos or data. Fig B. Confocal images showing uroporphyrin distribution in areas of pink-red pigmentation for Clanculus pharaonius. Shell images on each line are of a single shell and are in approximately the same orientation and magnification. (A-C) Top of shell (specimen #4). (A) Visible pink-red pigmentation. (B) Shaded volume 3D projection of a spectrally mixed image of the same shell showing fluorescent areas congruent with visible pink-red pigment. (C) Maximum intensity projection of a spectrally unmixed image showing distribution of uroporphyrin I and III. (D-F) Area on later whorl of same specimen. (D) Visible pigmentation for approximate areas used in analyses. (E) Maximum intensity projection of spectrally mixed images showing fluorescent areas under UV light congruent with pink-red pigmentation. (F) Maximum intensity projection of spectrally unmixed images showing distribution of uroporphyrin I and III. Note that in unmixed images green corresponds to uroporphyrin I and red to uroporphyrin III and yellow to areas where both co-occur. Note also that fluorescence is quenched in black spots and nearby white areas. Visible pigmentation is marked on each image: pr–pink-red, bl–black spot, w–white area. Fig C. Results of Raman spectroscopy. Spectra excited by 532 nm light on surface of shell of Clanculus margaritarius B (specimen #7). (A) Yellow-brown area. (B) Black spot. (C) Comparison of two spectra from (A) and (B). Fig D. High performance liquid chromatography (HPLC) analyses for eumelanin in Clanculus margaritarius B. (A) Chromatogram of specimen #8 (measurable PTCA). (B) Chromatogram of specimen #12 (no obvious PTCA). Fig E. EDS maps of Clanculus margaritarius shell (specimen #11) for 14 metal ions. Black spot spectra were taken from the centre of the ESEM images; yellow-brown spectra were taken from the areas in the top left corner. Analyses were run on a FEI Quanta 650 FEG SEM equipped with a multi-channel Bruker 5060F XFlash QUAD EDS system with an acquisition time of 180 s, working distance 8.8 mm, chamber pressure 67 Pa and an accelerating voltage of 25 kV. Table F. Synchrotron XRF EDS quantification of pigmented and unpigmented regions of Clanculus margaritarius (specimen #11). 2σ errors in parentheses. Values in ppm unless where stated in weight% (%). Fig G. Microfocus map of strontium (Sr) in C. margaritarius shell (specimen #11). Sr exhibits concentration bands that are parallel to yellow-brown/white colour boundaries. Optical photo not available. Fig H. Sulfur XANES spectra obtained from the yellow-brown and black regions of a C. margaritarius shell (specimen #11). The spectrum from the yellow-brown region exhibits a higher abundance of reduced organic sulfur species than the black area, consistent with the presence of an organic film as postulated to explain the inhibited Ca fluorescence. Vertical dashed line represents the peak position for sulfate determined by a ZnSO4 standard (~2482 eV). Fig I. Attenuated Total Reflectance Infrared Spectra of C. margaritarius (specimen #11) bulk shell material compared to a dark spot and a uroporphyrin I standard. Dotted lines indicate potential peaks of similarity between the samples though this is not enough to demonstrate the presence of uroporphyrin, there are also no apparent differences between the spectra of the bulk shell material and the dark spot. Both show the presence of organic matter and protein moieties. Fig J. TofSIMS maps of Clanculus margaritarius shell (specimen #11). Secondary ion images are shown with a rainbow colour scheme from black (no counts) to red (maximum). Numbers in parentheses are the total counts given in each individual image followed by the maximum count. In positive secondary ion mode, non-polar organic molecules usually ionize as (M+H)+ which means one hydrogen atom has to be subtracted from the measured secondary ions to get the actual molecule present in the sample. Fig K. Graphs showing results of laser ablation for five selected metals. Each curve represents a different sample. There are seven samples for black pigment (black lines), three from Clanculus margaritarius and four from C. pharaonius shells; five samples for red pigment (red lines) from C. pharaonius shells; 12 samples for white areas (grey lines), seven from C. margaritarius and five from C. pharaonius shells; and nine samples for yellow-brown pigment (yellow lines) all from C. margaritarius. See Table A for list of specimens used. (ZIP)