Free volatile carboxylic acids (FVCA) such as those unbranched from C1 to C6, and isoforms of butyric, valeric and caproic acids, are formed during cheese ripening as desired and also as undesired metabolites. Although in the literature these acids are often also referred to as free volatile fatty acids, strictly speaking only applies to butyric acid and n-caproic acid which compose milk fat. All these compounds are formed by hydrolysis of milk fat and by lactate and protein, such as during heterofermentative lactic acid fermentation or amino acid deamination. In most cases, all mechanisms can be observed during cheese ripening to a greater or lesser extent depending on the type of cheese. In addition, the presence of undesirable microorganisms can lead to unintended fermentations in which FVCAs are formed in sometimes high concentrations (Fox, Guinee, Cogan, & McSweeney, 2017). On the other hand, these compounds are contributors to the typical and desirable flavors of many cheeses. For example, the conversion of lactate into propionic acid by propionibacteria is a known metabolic pathway for the formation of the typical propionic acid flavor in Emmental cheese (Fröhlich-Wyder, Bisig, Guggisberg, Jakob, Turgay, & Wechsler, 2017). The quantitative composition of FVCA is an important quality characteristic of a cheese. Variations in the composition indicate disturbances during ripening and are very helpful in the search for manufacturing defects. For example, Obligat heterofermentative lactobacilli form acetate and Faktulativ heterofermentative lactobacilli form formate and acetate (Fröhlich-Wyder, Guggisberg, Badertscher, Wechsler, Wittwer, & Irmler, 2013). In addition to propionate, propionic acid bacteria can also form acetate (Fröhlich-Wyder et al., 2017) while Coliform bacteria form lactate, formate and acetate (Eugster, Jakob, & Wechsler, 2012). An early butyric acid fermentation is observed with Clostridium butyricum or as late bloating with Clostridium tyrobutyricum. The Putrifikus (white rot) is caused by Clostridium sporogenes spores, which produces a whole spectrum of carboxylic acids such as acetate, propionate, butyrate, isovalerate, isobutyrate and isocaproate through their proteolytic activities (Gómez-Torres, Garde, Peirotén, & Ávila, 2015). Similarly, an atypical butyric fermentation with little gas (H2 and CO2) and butyric acid is induced by Clostridium beijerinckii (Klijn, Nieuwenhof, Hoolwerf, van der Waals, & Weerkamp, 1995). Certain yeasts can produce acetate at an early stage of ripening and cause undesirable eye formation. The new method is based on an unpublished in-house development that has been continuously improved. In the previous method, 20 g of cheese was first distilled in an acidic medium with steam and the distillate titrated with NaOH to determine the total acidity. Subsequently, 1 mL of the over-titrated solution was esterified and the relative concentrations of each FVCA were determined by headspace injection on a GC-FID. Together with the total acidity, the individual absolute contents could then be calculated. Over the years, various optimizations have been made to improve recoveries and reduce workload. Initially, propanol was used as the esterification reagent. Later, due to blank value problems, a switch was made to ethanol. The optimal esterification and headspace conditions (time, temperature) were systematically determined. For the switch from steam distillation to basic extraction, the amount of ethanol used for esterification as well as different sodium hydroxide concentrations and extraction times were optimized. Special attention was paid to avoid possible saponification of the milk fat.
