This dataset contains stable isotope values from eye lens laminae, vertebral bands, and muscle tissue of wild Leopard Sharks (Triakis semifasciata) collected from multiple estuaries in Central California between 2020 and 2023. Each record includes individual identifiers, tissue type, collection date, locality, sex, total length, lamina/layer designation, tissue diameter, δ¹³C, δ¹⁵N, C:N ratio, and predicted FTIR absorbance for lens laminae. Eye lens laminae provide a sequential biochemical archive, enabling reconstruction of lifetime diet and habitat use at fine temporal resolution. Vertebral and muscle tissues offer complementary isotopic baselines for comparison and tissue discrimination factor estimation. The dataset structure allows tracking of isotopic variation across ontogeny, among individuals, and between estuarine populations. Potential reuse includes ecological studies of long-lived chondrichthyans, life history reconstructions, trophic ecology analyses, and comparative studies of isotopic incorporation across tissue types. All data were collected under appropriate permits and comply with ethical standards for wildlife research. No sensitive or personally identifiable information is included.

Materials and Methods

1.1 Sampling

To verify the use of eye lens laminae in chondrichthyans, we compared the stable isotope values of muscle, eye lenses, and vertebrae within individual Leopard Sharks. We then used eye lenses to study their ecology through ontogeny in Central California. Here, we collected sub-adult and adult Leopard Sharks across four sites (San Francisco Estuary [SFE; n = 9], Drakes Estero State Marine Conservation Area [hereby after referred to Drakes Estero; n = 10], Tomales Bay [n = 10], and Bodega Bay [n = 9]) via hook and line, set gill nets, and salvage donation from fishers (Fig 2). Individuals were caught and euthanized with methods approved by the Institutional Animal Care and Use Committee of the University of California, Merced (protocol # AUP20-0013), California Department of Fish & Wildlife (permit S-201820001-20182-001 and S-201840003-20196-001), and the National Park Service (#PORE-2020-SCI-0017). Leopard Sharks were transported on ice to the University of California, Merced for dissection and further analyses. We opportunistically collected Leopard Shark fetuses (n = 15) from a single, near-term pregnant individual from Tomales Bay post necropsy; these fetuses were used for the eye lens diameter aging methodology.

We dissected and chronologically sampled eye lens laminae to reconstruct the isotopic life histories of sub-adult and adult Leopard Sharks (n = 38). Eye lenses were dissected from frozen whole eyes and once thawed, serially delaminated from the outermost pre-apoptotic lamina to the innermost post-apoptotic laminar core. During delamination, lenses were photographed at 10x magnification using an imaging camera mounted on a stereoscope, and laminar diameter (mm) was measured along four evenly spaced axes using ImageJ following [1]. Laminar midpoints were calculated following [2]. In addition, the total eye lens diameter (capsule diameter) was measured for each individual (n = 53) to compare with total length and facilitate back-calculation of age. Eye lens laminae were desiccated in a drying oven at 55 °C for 12 hours before weighing and preparation for SIA. Urea was previously confirmed in the outer pre-apoptotic eye lens laminae, but urea inclusion resulted in minimal isotopic effect (mean δ¹⁵N offset ≈ 0.2‰, within analytical error) [1]; therefore we did not remove urea prior to analysis in this study. However, to conservatively account for potential urea contamination isotopic effects, samples exhibiting C:N ratios ≤ 2.47—outside the range of C:N values evaluated by the study—were excluded from further analyses [1].

To establish eye lenses as life history recorders, vertebrae and muscle from Leopard Sharks were also analyzed. The 15th thoracic vertebra was dissected from six individuals across three sites (SFE [n = 3], Drakes Estero [n = 1], and Tomales Bay [n = 2]). Each vertebral centrum was mounted in epoxy resin, and two parallel sagittal sections were taken via diamond saw. The first 1-mm section was polished to identify annual bands for age determination under dissecting microscope, while the second 2-mm section was sampled for SIA. The small size of Leopard Shark vertebrae required combining multiple annual bands in some sections to achieve the minimum sample size for SIA (~0.2 mg). Early, wider annual bands could be segmented and analyzed individually, whereas later, smaller bands were segmented and later combined. Similarly to eye lens laminar measurements described in [3], images were taken on a dissecting microscope of the vertebral section after removal of each segmented sample, and a vertebral band midpoint was measured (mm) to compare with eye lens laminae. Each vertebral section was demineralized using ethylenediaminetetraacetic acid (EDTA) following [4]. Demineralized vertebral section samples were frozen 8+ hours and then lyophilized overnight prior to weighing and preparation for SIA. We also collected muscle tissue from the dorsal region above the pectoral fin for comparative analysis with outer eye lenses. Muscle tissue was lipid and urea extracted following [4] prior to lyophilization and preparation for SIA.

1.2 Stable Isotope Analysis

1.2.1 Analytical Procedure

Eye lens laminae and vertebral banding sections were analyzed for δ¹³C and δ¹⁵N at the Stable Isotope Ecosystem Laboratory, University of California, Merced. Samples were weighed into tin capsules and combusted using a Costech Elemental Analyzer coupled via a Conflo IV interface to a Thermo Delta V Plus continuous-flow isotope ratio mass spectrometer, allowing simultaneous measurement of carbon and nitrogen isotope ratios.

1.2.2 Traceability

Isotope ratios are reported in standard delta (δ) notation in per mil (‰), with δ¹³C values normalized to Vienna Pee Dee Belemnite (VPDB) and δ¹⁵N values normalized to atmospheric nitrogen (V-AIR). Instrument calibration, normalization, and linearity were established using internationally recognized reference materials measured throughout each analytical sequence, including USGS40 (δ¹³C = −26.4 ± 0.1‰; δ¹⁵N = −4.5 ± 0.2‰; n = 129), USGS41a (δ¹³C = +36.5 ± 0.1‰; δ¹⁵N = +47.5 ± 0.1‰; n = 76), acetanilide 1 (δ¹³C = −28.3 ± 0.1‰; δ¹⁵N = −0.6 ± 0.1‰; n = 19), and acetanilide 2 (δ¹³C = −26.5 ± 0.1‰; δ¹⁵N = −4.9 ± 0.2‰; n = 31). At the start of each analytical sequence, USGS40 was measured in quintuplicate to establish calibration and was reanalyzed throughout the run to account for linearity. USGS41a and either of the in house acetanilides were also measured repeatedly within each analytical sequence to account for drift.

1.2.3 Data Processing

Raw isotope-delta values were corrected for instrumental drift and linearity using calibrated reference material means and corresponding standard deviations. Drift corrections were applied based on quality control (QC) materials analyzed at regular intervals, with two QC measurements interspersed after every ten samples. Final isotope-delta values were normalized using multi-point normalization.

1.2.4 Analytical Uncertainty and Quality Control

Analytical uncertainty was estimated from the standard deviation of repeated measurements of reference materials and internal standards and is reported alongside isotope-delta values where appropriate. Analytical accuracy and consistency were independently assessed using an internal laboratory reference material (Mb Squid; δ¹³C = −18.8 ± 0.1‰; δ¹⁵N = +11.9 ± 0.3‰; n = 46), which was measured repeatedly throughout each analytical run and exhibits a similar long-term instrument average and standard deviation (δ¹³C = −18.7 ± 0.1‰; δ¹⁵N = +11.9 ± 0.2‰).

1.3 ATR-FTIR Spectroscopy

ATR-FTIR spectra of Leopard Shark eye lens laminae (n = 2 individuals, 29 laminae) were previously collected using a Bruker Vertex 70 FTIR spectrometer (Nuclear Magnetic Resonance Facility, UC Merced), with 32 scans averaged per lamina across 400–4000 cm^-1 at 4 cm^-1 resolution. We reanalyzed previously these published spectra to quantify protein compaction across eye lens laminae [1]. Spectra were baseline-corrected using flattened background regions and smooth spline fits [5,6] and normalized to the maximum absorbance of the amide II band per lamina to facilitate comparison of peak shape and relative intensity independent of absolute signal magnitude. To account for variation in absolute absorbance due to sampling geometry, tissue thickness, and ATR crystal contact, peak-to-peak ratios were calculated [7]. Protein compaction was quantified as the ratio of the two amide II sub-peaks (peak 1: ~1517 cm^-1; peak 2: ~1535 cm^-1), yielding an FTIR-based index of structural maturity (FTIR_i). Higher FTIR_i values correspond to less compact, hydrated β-sheet structures, whereas lower values indicate increased crystallin fiber elongation, twisting, and compaction [8]. FTIR_i was subsequently modeled as a function of normalized laminar position and predicted across all laminae.

References

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