## Materials and Methods
### Volatile Profile Determination  
#### 2.7.1 HS-SPME/GC\(\times\)GC-ToFMS  
GC\(\times\)GC-ToFMS was employed to study in-depth the \"Rocha\" pear volatile profile variation across cold storage. Sampling, reporting of chemical analysis to data preprocessing, pretreatment, processing, and interpretation were performed according to the metabolomics standards initiative (MSI), as detailed below [33]. HS-SPME/GC\(\times\)GC-ToFMS methodology was adapted from other studies [34; 35] with optimization of the following parameters: time of pre-equilibrium and HS-SPME extraction, stationary phase of the columns, temperature ramp; modulation time, transfer line temperature, and drag gas flow. The effects of these parameters were evaluated based on visual analysis of the chromatograms through the chromatographic resolution, relative abundance of the principal peaks, and chromatogram overall structure. Each analyzed pear was randomly selected and sealed in a 1 L airtight jar fitted with a rubber septum in the lid. Jars were closed and kept overnight at room temperature to reach equilibrium. Then, each jar was immersed in a water bath adjusted to 25.0 \(\pm\) 0.1 \({}^{\circ}\)C and the DVB/CAR/PDMS SPME fiber was inserted in the headspace trough the septum, for 1 h. The volatiles adsorbed and absorbed on the SPME fiber coating were determined using a LECO Pegasus 4D GC\(\times\)GC-ToFMS system (LECO, St. Joseph, MI, USA) consisting of an Agilent GC 7890A gas chromatograph (Agilent Technologies, Inc., Wilmington, DE, USA), with a dual stage jet cryogenic modulator (licensed from Zoex) and a secondary oven, and a mass spectrometer equipped with a ToF analyzer. After the extraction/concentration step, the SPME fiber was manually introduced into the port at 250 \({}^{\circ}\)C, for 3 min, for analytes desorption. The injection port was lined with a 0.75 mm I.D. glass liner. Splitless conditions (30 s) were used. An Equity-5 column (30 m \(\times\) 0.32 mm I.D., 0.25 \(\upmu\)m film thickness, Supelco, Bellefonte, PA, USA) was used as the first-dimension column (\({}^{1}\)D) and a DB-FFAP column (0.79 m \(\times\) 0.25 mm I.D., 0.25 \(\upmu\)m film thickness, J&W Scientific Inc., Folsom, CA, USA) was used as the second-dimension column (\({}^{2}\)D). The carrier gas was helium at a constant flow rate of 2.50 mL/min. The primary oven temperature was programmed from 40 \({}^{\circ}\)C (1 min) to 160 \({}^{\circ}\)C, at 3 \({}^{\circ}\)C/min, followed by a second ramp from 160 \({}^{\circ}\)C to 225 \({}^{\circ}\)C (2 min), at 15 \({}^{\circ}\)C/min. Secondary oven program was 5 \({}^{\circ}\)C offset above the primary one. The MS transfer line and MS source temperatures were both set at 250 \({}^{\circ}\)C. The modulation period was 5 s, keeping the modulator at 20 \({}^{\circ}\)C offset above primary oven, with hot and cold pulses of 0.90 and 1.60 s, respectively. The mass spectrometer ran in EI mode at 70 eV, using an \(m\)/\(z\) range of 35-300. Total ion chromatograms were processed using the automated data processing software ChromaTOF(r) (LECO, St Joseph, MI, USA) at a signal-to-noise threshold of 100. Spectral deconvolution was computationally processed, being intended to reconstruct clean mass spectra for each component; whereas the GC\(\times\)GC peak area was obtained by transforming the series of side-by-side second-dimension chromatograms into a two-dimensional chromatogram, the GC peak area being proportional to the generated signal intensity [36]. Contour plots were used to evaluate the general separation quality and for manual peak identification. For identification purposes, the mass spectrum of each detected metabolite was compared with mass spectral libraries, namely an in-house library of standards and two commercial databases (Wiley 275 and U.S. National Institute of Science and Technology (NIST) V. 2.0, Mainlib and Replib). A mass spectral match factor, similarity \(>\)700/1000, was used to decide whether a peak was correctly identified.