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Commercial coffee is available as a mixture of two varieties of coffee beans, namely Arabica, which is more expensive, and Canephora (Robusta), less expensive. To assess the correspondence between the composition indicated on the label and the real composition of commercially available coffee, it would be desirable to be able to differentiate between the two varieties. This would also help to avoid any possible commercial frauds. This work identifies parameters based on the fatty acid composition to differentiate between Arabica and Canephora coffee in a mixture. Total monounsaturated fatty acids (ΣMUFA), linolenic acid (cis18:3n–3) concentration, the 18:0/cis18:1n–9 ratio, and the ΣMUFA/ΣSFA ratio could be used to determine the relative amounts of Arabica and Canephora in a coffee blend.