Eggshells of the Western Painted Turtle Sequester CO₂: Implications for Gas Exchange, pH Homeostasis, and Overwintering in Hatchling Turtles

Newly hatched painted turtles remain in the nest and generally emerge 7 to 8 months after hatching. During that time, atmospheric conditions inside the nest potentially become hypoxic and hypercapnic. However, recent nest gas sample analysis has shown lower than expected CO 2 levels (unpublished data). One possible explanation for the reduced CO 2 levels may be due to the presence of eggshells. The calcareous portion of avian and most reptilian eggshells is composed of CaCO 3 primarily in the crystalline form of calcite, but the crystalline portion of turtle eggshells is composed of aragonite. Compared to rigid archosaur eggshells, painted turtle eggs are thin and lightly mineralized. Does the extent of eggshell mineralization relate to its ability to sequester CO 2 ? To test the hypothesis that turtle eggshells sequester CO 2 , we measured CO 2 uptake gravimetrically in alligator, chicken, and painted turtle eggshells. Whole eggshells (n=3) were placed inside separate 30 ml glass syringes and a rubber stopper connected to a one-way stopcock valve was used to seal each chamber. The one-way stopcock valves on both ends of the syringes were then opened and the syringes were flushed with nitrogen gas overnight to remove any lingering CO 2 . Afterwards, they were flushed with increasing concentrations (25%, 50%, 100%) of CO 2 and weighed until constant mass before switching to the next concentration. To determine the reversibility of the process, nitrogen gas was flushed through the chambers that were saturated with 100% CO 2 . Powdered x-ray diffraction was also used to assess compositional differences of dried eggshell powder, and eggshell that was ashed at 900°C. The results showed that turtle eggshell was significantly better at sequestering CO 2 at all concentrations measured. At 25% CO 2 , alligator, chicken, and turtle mean sequestered 0.029 ± 0.002, 0.017 ± 0.002, and 0.306 ± 0.081 mmol CO 2 /g eggshell, respectively. At 50% CO2, alligator, chicken, and turtle mean sequestered 0.051 ± 0.005, 0.044 ± 0.003, and 0.622 ± 0.174 mmol CO 2 /g eggshell, respectively. At 100% CO 2 , alligator, chicken, and turtle mean sequestered 0.112 ± 0.009, 0.106 ± 0.008, and 1.366 ± 0.374 mmol CO 2 /g eggshell, respectively. On average 87.06 ± 0.62% of the sequestered CO 2 could be washed from the dried turtle eggshells following nitrogen flush. Powdered x-ray diffraction analysis confirmed calcite as the crystalline structure in raw chicken and alligator eggshell and aragonite in painted turtle eggshell, but no consistent differences in composition in ashed eggshells. These results suggest that the turtle eggshell can remove CO 2 from the nest cavity and that a portion of this process is reversible. We posit that painted turtle eggshells may play an important role in preventing environmental hypercapnia in overwintering hatchlings. NSF CAREER grant to DEW. Award number 1253939 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.