## Materials and Methods
### Solid Phase Microextraction (SPME) Optimization  
Three SPME fibers (Supelco, Bellefonte, PA, USA) were assessed for their sampling efficiency of the volatile components of cider headspace. These were: polyacrylate 85 \(\upmu\)m (PA), divinylbenzene/carboxoen/polydimethylsiloxane 50/30 \(\upmu\)m (DVB/CAR/PDMS), and carboxen/polydimethylsiloxane 75 \(\upmu\)m (CAR/PDMS). _S. cerevisiae_ var. _diasticus_ yeast (French Saison, strain 3711) and a mixture of local, Montezuma County, CO, USA heirloom variety apples were utilized for the cider sample fermentation for all method validations reported herein. As shown in Figure 2, the DVB/CAR/PDMS fiber was selected as the optimal sampling fiber for VOCs originating from cider samples due to the greater extraction efficiency as well as the wider variety of cider constituents (primarily esters, higher alcohols, and phenols) compared to the other fibers. Sampling efficiency was determined through the peak integration of the chromatograms (Figure 2) for each SPME fiber. In contrast to previous work [12], we observed that the extraction efficiency of the DVB/CAR/PDMS fiber (100%) was significantly greater than that of the CAR/PDMS (51%) and the PA fibers (2%), due to the affinity of cider headspace molecules for this polymer blend. Based on these data, every result reported herein utilized DVB/CAR/PDMS SPME fibers.  
All cider aroma measurements were performed through the introduction of a SPME fiber into the headspace of a 10 mL cider aliquot sealed in a 20 mL 22.5 \(\times\) 75 mm glass headspace vial (Sigma Aldrich, St. Louis, MO, USA). Each vial was capped airtight with a 20 mm aluminum release seal PTFE/silicone liner cap (Sigma Aldrich) using a handheld manual vial crimper. For temperature consistency across all samples, SPME fibers were introduced into the headspace after samples were submerged in a 40 \({}^{\circ}\)C water bath for 10 min. Prior to headspace sampling, all SPME fibers were conditioned for 30 min at the temperature specified by the manufacturer (Supelco). The SPME fiber exposure times were then compared, and 30 min was determined to be optimal for the time-efficient and abundant sampling of esters and higher alcohols, as illustrated in Figure 3. Longer SPME fiber exposures (90 min) did not result in additional analyte detections despite the increased time, because a stable analyte deposition  
Figure 2: Overlaid chromatograms of a single cider sample (EsoTerra Cider) analyzed with 3 different solid phase microextraction (SPME) fibers are shown. Polyacrylate (PA: Red), carboxen/polydimethylsiloxane, (CAR/PDMS: Green), and divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/PDMS: Blue) were compared. A wider variety of esters (**1**: Ethyl acetate, **2**: Methyl formate, **6**: Hexanoic acid ethyl ester, **9**: Benzoic acid ethyl ester, **10**: Octanoic acid ethyl ester, **12**: Nonanoic acid ethyl ester, **13**: Ethyl 9-decanoate, **14**: Decanoic acid ethyl ester, **15**: Dodecanoic acid ethyl ester), higher alcohols (**3**: 3-Methyl-1-Butanol, **4**: 1-Hexanol, **7**: 1-Octanol, **8**: 2-Phenylethyl alcohol), phenols (**11**: 4-ethyl-2-methoxyphenyl), and an amine (**5**: methoxy-phenyl-oxime) were detected consistently at higher abundance in the DVB/CAR/PDMS fiber than the other fibers analyzed.  
equilibrium had already been established [26]. Shorter SPME fiber exposures (5 min) resulted in several minor species dropping below the detection limit.