Department of Wildlife, Fish and Conservation Biology, University of California, Davis, 1 Shields Avenue, Davis, CA, 95616, USA.
Rapid Commun Mass Spectrom. 2014 Jan 15;28(1):96-108. doi: 10.1002/rcm.6761.
Compound-specific stable isotope analysis of amino acids has proven informative to many ecological systems, but only a handful of analytical methods are routinely employed. We evaluated a simple, rapid procedure in which biological samples undergo short-duration acid hydrolysis and the resulting amino acids are derivatized with methyl chloroformate for gas chromatography/combustion/isotope-ratio mass spectrometry (GC/C/IRMS).
Amino acid derivatives were separated on a polar gas chromatography column, combusted, and δ(13)C and δ(15)N values were measured. Tests of reproducibility and accuracy were conducted for amino acid reference mixtures and biological samples. A brief case study of turtles was used to assess whether isotopic data were consistent with a priori ecological expectations.
The methyl chloroformate based reaction successfully converted 15 amino acids from acid hydrolysates of biological materials into separable derivatives. The δ(13)C and δ(15)N values had high average measurement precision (σ <1‰). Reference materials were measured accurately, with good agreement between EA/IRMS and GC/C/IRMS determinations. Analysis of turtle blood samples yielded data consistent with their trophic ecology.
This derivatization method is a rapid means of determining carbon and nitrogen isotopic ratios of amino acids present in the biological materials often sampled for ecological studies. While amino acids with charged or polar side chains do not have uniformly high recoveries, the average precision of measurements is comparable with that of other, more established methods. Batches of samples may be prepared from many raw materials in less than a day, representing a significant reduction in preparation time over prevailing methods.
对氨基酸进行特定于化合物的稳定同位素分析已被证明对许多生态系统具有重要意义，但只有少数分析方法得到了常规应用。我们评估了一种简单、快速的方法，该方法中生物样本经历短时间的酸水解，然后用氯甲酸甲酯衍生生成的氨基酸用于气相色谱/燃烧/同位素-比例质谱（GC/C/IRMS）。
氨基酸衍生物在极性气相色谱柱上分离，然后进行燃烧，测量 δ(13)C 和 δ(15)N 值。对氨基酸参考混合物和生物样本进行了重现性和准确性测试。对海龟进行了简短的案例研究，以评估同位素数据是否与先验生态预期一致。
基于氯甲酸甲酯的反应成功地将生物材料酸水解产物中的 15 种氨基酸转化为可分离的衍生物。δ(13)C 和 δ(15)N 值具有很高的平均测量精度（σ <1‰）。参考物质的测量准确，与 EA/IRMS 和 GC/C/IRMS 测定结果具有良好的一致性。对海龟血液样本的分析结果与它们的营养生态学一致。
这种衍生化方法是一种快速测定生物材料中存在的氨基酸碳和氮同位素比值的方法，这些生物材料通常是为生态研究而采样的。虽然带有电荷或极性侧链的氨基酸的回收率并不均匀，但测量的平均精度与其他更成熟的方法相当。在不到一天的时间内，就可以从许多原材料中制备出多个样本批次，与现有的方法相比，大大减少了制备时间。
