Butyric acid (butyrate) is a candidate marker of milk fat in complex fat blends, since it is exclusive of milk triacylglycerols (TAGs) from different ruminant species. In this work, we determined the amount of milk fat used for the preparation of fat blends by ¹³C-Nuclear Magnetic Resonance (¹³C-NMR) spectroscopy-based quantification of butyrate. When tested on fat samples spiked with known amounts of reference bovine milk fat (BCR-519 certified material), the relative composition of the mixtures was reliably assessed through the integration of the diagnostic ¹³C-NMR carbonyl (C1) or α-carbonyl methylene (C2) resonances of butyrate. NMR data exhibited strict correlation with high resolution-gas chromatography (GC) of fatty acid methyl esters (R² = 0.99), which was used as an independent and well-established method for the determination of butyrate. Thus, ¹³C-NMR can be used for the direct assessment of milk fat content in fat mixtures, at a limit of detection lower than 5%, with clear advantages over the traditional GC methods in terms of speed, robustness and minimal sample handling. The natural variability of butyrate in milk has been taken into account to estimate the uncertainty associated with the milk fat content in unknown fat blends.

丁酸（丁酸酯）是复杂脂肪混合物中乳脂的候选标记，因为它不包含来自不同反刍动物物种的乳三酰甘油（TAG）。在这项工作中，我们通过基于¹³ C-核磁共振（¹³ C-NMR）光谱的丁酸酯定量确定了用于制备脂肪混合物的乳脂量。当在掺有已知量参考牛乳脂肪（BCR-519认证材料）的脂肪样品上进行测试时，通过诊断¹³ C-NMR羰基（C1）或α-羰基亚甲基的整合，可以可靠地评估混合物的相对组成（C2）丁酸酯的共振。NMR数据与脂肪酸甲酯（R ²= 0.99），这是用于测定丁酸盐的独立且完善的方法。因此，¹³ C-NMR可用于直接评估脂肪混合物中的牛奶脂肪含量，检出限低于5％，与传统的GC方法相比，在速度，稳定性和最小的样品处理方面具有明显的优势。已考虑到牛奶中丁酸酯的自然变异性，以估计与未知脂肪混合物中的牛奶脂肪含量相关的不确定性。