Solubilization of skin collagen improves the accuracy and reliability of stable isotope measurements.

Abstract

Stable isotope analysis of skin collagen is useful for detecting short-term or seasonal diet. Preparation of skin for stable isotope analysis varies across laboratories, and this may impact the comparability of data. It is important to understand the effects of different preparatory protocols on the stable isotopic and elemental compositions of skin samples. Using a Eurovector 3,300 elemental analyzer coupled to a Nu Horizon isotope ratio mass spectrometer, we tested the impact of three treatment variants (refluxing at three temperatures to remove non-collagenous proteins, treatment with sodium hydroxide (NaOH), and chemical lipid extraction using 2:1 chloroform:methanol) on the stable isotope (δ ¹³carbon (C) and δ ¹⁵nitrogen (N)) and elemental (wt% C, and wt% N) composition of pig (Sus scrofa domesticus) skin. The refluxing step produced pig skin with higher δ ¹³C values, lower C:N_Atomic ratios, less variable C:N_Atomic ratios, wt% C, and wt% N. The chemical lipid extraction also produced higher, more reliable δ ¹³C values and lower, less variable C:N_Atomic ratios. The isotopic data in the lipid-extracted and refluxed samples were more consistent in the refluxed samples than the non-refluxed and non-lipid-extracted samples, as determined by the elemental compositions.