Analysis of Ethylphenols

The very low odor threshold limits of 4-EG and 4-EP in wine need sensitive analytical methods, including prior concentration steps. By using GC/MS analysis, detection limits of a few microgram per liter and wide linearity range can be achieved (Pollnitz et al., 2000 Martorell et al, 2002). Several methods of sample preparation were proposed liquid-liquid extraction (Chatonnet et al., 1992 Versini, 1985 Chatonnet et al., 1995 Pollnitz et al, 2000 Chatonnet and Boidron, 1988 Rocha et al., 2004) SPE (Aznar et al., 2001 Dominguez et al., 2002 Lopez et al., 2002) SPME and HS-SPME (Martorell et al., 2002 Ferreira et al., 1996 1998 Monje et al., 2002 Castro Mejias et al., 2003) and stir bar sorptive extraction (SBSE) (Dfez et al., 2004). 

A method of dispersive liquid-liquid microextraction (this technique is based on the use of a ternary component solvent system composed of an extraction and a dispenser solvent) coupled to GC/MS was recently proposed. This method provided limits of detection (LOD) 

Sampling using a 100-gm PDMS fiber by extraction of 25 mL of wine added of a suitable amount of NaCl to get a 6M solution at 25°C for 60min, allows LODs and LOQs of 1 and 5qg/L for 4-EG, and 2 and 5qg/L for 4-EP, respectively (Martorell et al., 2002). Also, the HS-SPME of NaCl saturated samples using a 85-qm polyacrylate (PA) fiber for 40min at 55 °C provided LODs in the low microgram per liter range (Monje et al., 2002). 

A method based on the use of multiple-headspace (MHS) SPME using a carbowax-divinylbenzene (CW/DVB) fiber (three consecutive extractions of the same sample to minimize the possible matrix effects), showed LODs (S/N = 3) of 0.06qg/L for 4-EG and 4-EP, 0.20qg/L for 4-VG, and 0.12 qg/L for 4-VP, below the sensory thresholds of these compounds in wines (Pizarro et al.,2007). 

The HS-SPME are usually carried out under nonequilibrium conditions. The distribution constants of analytes between the fiber and the sample, between the HS and the sample, and the volume of the three phases (sample, headspace, and coating) must be constant, like the other SPME extraction parameters (sample agitation, fiber exposure time, etc.). The total analyte area (AT) corresponding to a cumulative extraction yield after multiple extractions can be determined as the sum of the areas obtained for each individual extraction when each is exhaustive, or expressed as  

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Fig. 32.6 Influence of column length on analysis time. Analysis of phenols under isothermal conditions 1. phenol 2. o-cresol 3. 2,6-xylenol 4. p-cresol 5. n-cresol 6. o-ethylphenol 7. 2,4-xylenol 8. 2,5-xylenol 9. 2,3-xylenol 10. p-ethylphenol 11. m-ethylphenol 12. 3,5-xylenol 13. 3,4-xylenol.

Due to the labor-intensive nature of monitoring programs, alternatives have been sought. Enzyme-linked immunosorbent assay (ELISA) has been proposed (Kuniyuki et al., 1984). The advantage of ELISA is that viable yeasts need not be present. Unfortunately, the method is too sensitive for routine production applications and, at present, too costly. The volatile phenol, 4-ethylphenol, has been proposed as a marker for present/past growth of Brettanomyces and Dekkera in wine. For details regarding analysis, see Zoecklein et al. (1995). 

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