Lab 1 purchased a further 27 beef samples, from which 79 extracts were prepared for NMR analysis. Lab 2 purchased 4 beef and 6 horse samples, from which 12 and 16 extractions were prepared, respectively. The total numbers of beef and horse extracts prepared across both labs were 91 and 16, respectively. The role of these test samples was to challenge the authenticity model created from the Training Set samples. In addition to extracts from meat samples, Lab 2 prepared a small collection

of samples from three laboratory-grade triglycerides (Sigma-Aldrich): glyceryl tristearate (C18:0), glyceryl trioleate (C18:1) and glyceryl trilinoleate (C18:3). A stock mixture was prepared containing 15% w/w C18:0 and 85% w/w C18:1. This was used to make four triglyceride mixtures containing 0, 10, 20 and 30% w/w of C18:3, respectively. These were diluted with approximately RO4929097 solubility dmso 80% by volume of chloroform before NMR analysis. Both Lab 1 and Lab 2 used similar, simple preparation and extraction procedures, Baf-A1 with the aim of establishing a protocol appropriate for a low-cost, high-throughput screening scenario. No attempt was made to determine the extraction efficiency, since the objective was to obtain representative compositional profiles suitable for speciation, rather than absolute quantitation. The extraction agent was deuterated chloroform (Lab 1) or chloroform (Lab 2), which is well-suited for the extraction

of neutral lipids such as triglycerides. The preparation for the Training Set samples at Lab 1 was as follows: A small amount of meat was cut into pieces (∼1 cm3) and homogenised in a food processor (Kenwood mini-chopper) for 30 s. Next, 1.5 ml of deuterated chloroform (Sigma-Aldrich) was added to 3-6 g homogenised meat (depending on fattiness; the lowest quantities were used

for visibly fatty samples) and the mixture vortexed for 10 min before being refrigerated for 1 h at Exoribonuclease 4 °C. The solvent extract was then recovered by pipette, filtered through paper tissue and placed in a 5 mm disposable NMR tube (Sigma-Aldrich). All samples were stored at 4 °C until NMR data were collected. Replicate extractions were obtained by homogenizing a representative cut of meat, and then preparing separate extractions from discrete subsamples. The order in which extracts were presented to the spectrometer was randomized within each batch. For the Test Set 2 samples, Lab 1’s procedure was modified slightly. In particular, the amount of sample mixed with deuterated chloroform was not weighed, and the mixture was not refrigerated after vortexing. Lab 2’s preparation for all meat samples was the same as that used by Lab 1 for the Training Set samples, with the following variations. Approximately 10 g of meat was homogenised. For each extraction, non-chloroform (analytical grade, Sigma-Aldrich) was added to a 5 ±0.05 g subsample of the homogenised meat.

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