In situ ATR-FTIR characterization of primary cement interfaces of the barnacle Balanus amphitrite

A method is presented for characterizing primary cement interfaces of barnacles using in situ attenuated total reflection-Fourier transform infrared spectroscopy. Primary cement of the barnacle, Balanus amphitrite (Amphibalanus amphitrite), was characterized without any disruption to the original cement interface, after settling and growing barnacles directly on double sided polished germanium wafers. High-quality IR spectra were acquired of live barnacle cement interfaces, providing a spectroscopic fingerprint of cured primary cement in vivo with the barnacle adhered to the substratum. Additional spectra were also acquired of intact cement interfaces for which the upper portion of the barnacle had been removed leaving only the base plate and cement layer attached to the substratum. This allowed further characterization of primary cement interfaces that were dried or placed in D2O. The resulting spectra were consistent with the cement being proteinaceous, and allowed analysis of the protein secondary structure and water content in the cement layer. The estimated secondary structure composition was primarily beta-sheet, with additional alpha-helix, turn and unordered components. The cement of live barnacles, freshly removed from seawater, was estimated to have a water content of 20-50% by weight. These results provide new insights into the chemical properties of the undisturbed barnacle adhesive interface.