br Fourier transform infrared FTIR attenuated total reflectance ATR The
2.5.3. Fourier transform infrared (FTIR) attenuated total reflectance ATR The Fourier transform infrared (FTIR) spectroscopy was applied to
infer some structure characteristics from papaya Haloperidol and to deter-mine the degree of O-methyl esterification  using an Alpha FTIR spectrometer (Bruker Optic, Ettlingen, Germany) equipped with a deu-terated triglycine sulfate detector and a single-bounce attenuated total reflectance (ATR) accessory (diamond crystal). FTIR–ATR spectra of samples were obtained with a resolution of 4 cm−1 and 50 scans. GRAMS/AI 9.1 software (Thermo Scientific) was used for spectra analy-sis. Methyl esterified and free uronic acids corresponded to bands at 1749 cm−1 and 1630 cm−1, respectively. Commercially available pec-tins with known degrees of O-methyl esterification (28%, 64%, 91%) and their mixtures (14%, 46%, 78%) were used to produce a standard curve of O-methyl esterification.
Samples diluted in water (2.5 μL/mL) were sonicated, dropped onto freshly cleaved mica, and dried in a vacuum at 30 °C for 20 min. Samples were maintained in a desiccator until analysis. To-pography images were obtained in an NX-10 Atomic Force Micro-scope (Park Systems, Suwon, South Korea) in an acrylic glove box with controlled temperature (~22 °C) and humidity (~3%). AFM im-aging was acquired on tapping mode using an NCHR probe (NanoWorld) with a spring constant of 42 N/m and 320 kHz reso-nance frequency. The scan speed and scanning resolution were
0.5 Hz and 512 × 512 points, respectively. For each sample, at least ten images were collected. Image measurements and automatic pro-cessing (plane subtraction and row alignment) were performed using Gwyddion 2.47 software (http://gwyddion.net/).
1D and 2D 1H spectra of 3CSF were recorded using a 500 MHz NMR spectrometer (Bruker Biospin, Rheinstetten, Germany) with a triple res-onance probe. Approximately 5 mg of each sample were dissolved in 0.5 mL of 99.9% deuterium oxide (Cambridge Isotope Laboratory, Cam-bridge, MA). All spectra recorded at 35 °C with deuterated water exhib-ited a peak due to exchange with residual H2O (HOD), suppressed by presaturation. For 1D 1H NMR (zgpr from Bruker Library) spectra, 64 scans were recorded using an interscan delay equaling 1 s. 2D 1H-1H COSY (cosyphpr from Bruker Library) and 1H\\1H TOCSY (mlevphpr from Bruker Library) spectra were recorded using states time propor-tion phase incrementation for quadrature detection in the indirect di-mension. TOCSY spectra were run with 4096 × 512 points with a spin lock field of 10 kHz and a mixing time of 60 ms. The 1H/13C Multiplicity-edited HSQC (HSQC) spectra were recorded with 1024
× 512 points and 256 scans using an Echo-Antiecho acquisition mode
with globally optimized alternating phase rectangular pulses for decoupling. The 1H/13C HMBC spectra were recorded with 2048 × 200 points and 512 scans, with a 50 ms delay for evolution of long-range couplings and set with no decoupling during the acquisition time, a
low-pass filter, and a QF magnitude acquisition mode. Chemical shifts were displayed relative to external trimethylsilylpropionic acid at 0 ppm for 1H. The data were processed using TopSpin3.1 (Bruker Biospin) .
2.6. Statistical analysis
The results were expressed as the mean ± standard deviation (SD), and the significance was set at P b 0.05. Data were analyzed using GraphPad Prism 6.0 software (GraphPad, San Diego, CA). One-way ANOVA with Tukey's (to assess differences among all groups) or Dunnett's (to assess differences among the control and two or more groups) were used as post hoc tests.
3.1. Definition of ripening-time points of papayas
Papayas were analyzed for pulp softness as well as ethylene and CO2 production (Fig. 1) to define papaya ripening-time points: papayas with 1 and 2DAH had unripe characteristics, as observed by no pulp softening (firmness higher than 2 N·cm−2 (10−2)) and the low CO2 and ethylene production (CO2 lower than 0.25 mL kg−1 h−1 and ethylene lower than 0.25 μL kg−1 h−1); papayas with 3DAH had an intermediate ripening-time point (almost ripe), which is marked by the onset of ethylene pro-duction and start of pulp softening; papayas with 4 and 5DAH were fully ripe, with high ethylene production (higher than 0.5 μL kg−1 h−1) and a soft pulp (lower than 0.2 N·cm−2 (10 −2)). The WSF and CSF had the yield amounts measured. As expected, the WSF yields increased as the fruit ripened, and the CSF was a little bit higher on very unripe 1DAH fruit. Ash, starch, proteins and phenolic compounds contents were in-significant in the CSF fractions.