背景:骨骼的稳定同位素分析提供了对动物觅食的见解,并允许随着时间的推移进行生态重建,然而,需要预处理来分离胶原蛋白。预处理通常包括去矿质以去除无机成分和/或脂质提取以去除脂肪。但是这些方案可以不同地影响稳定的碳(δ13C)和氮(δ15N)同位素值,这取决于化学物质,组织,和/或涉及的物种。特定物种的方法为跨研究的可比性创造了标准,并增强了对现代鲸目动物骨骼中胶原蛋白分离的理解。
方法:使用与同位素比质谱仪耦合的元素分析仪来测量完整(对照)或经受以下三种实验条件之一的粉状虎鲸(Orcinusorca)骨的δ13C和δ15N值:去矿质,脂质提取,去矿质和脂质提取。此外,评估C:N比率作为胶原纯度的代表。最后,检查了对照C:N比与之间的相关性历史年龄和对照C:N比与样本特征。
结果:对于任何实验方案,没有观察到δ15N值的显著差异。然而,通过所有三个实验方案,δ13C值均显着增加:去矿质,脂质提取,两种治疗相结合。最有影响力的方案是去矿质和脂质提取。C:N比的测量也显着降低了去矿质和两种处理相结合,表明处理后材料更接近纯胶原蛋白。通过C:N比显示的胶原蛋白纯度与历史年龄或样品特征无关。
结论:如果只有虎鲸骨骼的δ15N值值得分析,似乎没有必要进行预处理。如果对δ13C值感兴趣,样品应同时脱矿质和脂质提取。由于历史年龄和标本特征与样品污染无关,所有样品都可以平等对待。
BACKGROUND: Stable isotope analysis of bone provides insight into animal foraging and allows for ecological reconstructions over time, however pre-treatment is required to isolate collagen. Pre-treatments typically consist of demineralization to remove inorganic components and/or lipid extraction to remove fats, but these protocols can differentially affect stable carbon (δ13C) and nitrogen (δ15N) isotope values depending on the chemicals, tissues, and/or species involved. Species-specific methodologies create a standard for comparability across studies and enhance understanding of collagen isolation from modern cetacean bone.
METHODS: Elemental analyzers coupled to isotope ratio mass spectrometers were used to measure the δ13C and δ15N values of powdered killer whale (Orcinus orca) bone that was intact (control) or subjected to one of three experimental conditions: demineralized, lipid-extracted, and both demineralized and lipid-extracted. Additionally, C:N ratios were evaluated as a proxy for collagen purity. Lastly, correlations were examined between control C:N ratios vs. historical age and control C:N ratios vs. sample characteristics.
RESULTS: No significant differences in the δ15N values were observed for any of the experimental protocols. However, the δ13C values were significantly increased by all three experimental protocols: demineralization, lipid extraction, and both treatments combined. The most influential protocol was both demineralization and lipid extraction. Measures of the C:N ratios were also significantly lowered by demineralization and both treatments combined, indicating the material was closer to pure collagen after the treatments. Collagen purity as indicated via C:N ratio was not correlated with historical age nor sample characteristics.
CONCLUSIONS: If only the δ15N values from killer whale bone are of interest for analysis, no pre-treatment seems necessary. If the δ13C values are of interest, samples should be both demineralized and lipid-extracted. As historical age and specimen characteristics are not correlated with sample contamination, all samples can be treated equally.