6. Representative Results
Chemical shift analysis of the CPMAS 13C NMR spectra (Fig. 3) identified the major functional groups present in the exhaustively dewaxed tomato cuticle (cutin). The key carbon moieties in the cutin biopolyester were found to be long-chain aliphatics (0-45 ppm), oxygenated aliphatics (45-110 ppm), multiply-bonded and aromatics (110-160 ppm), and carbonyls (160-220 ppm). The oxygenated alkyl moieties (CHO + CH2O) play a crucial role in establishing covalent connections between the monomeric units of the cutin biopolymer, thereby forming the molecular architecture of the cutin matrix. Differences in relative peak areas observed in the spectral region between 45 and 100 ppm suggest that the mutant cutin has a relatively large proportion of cross-link forming CHO structural moieties compared to the wild-type cutin; careful quantitative measurements using direct polarization (DPMAS) NMR5 methods support this inference (data not shown).
CPMAS 13C NMR spectra also showed a progressively diminishing wax peak at 31 ppm (Fig. 4), indicating the sequential removal of epi- and intracuticular waxes from the cutin-wax composite while retaining the principal chemical architecture of the cutin biopolymer. Parallel AFM image analysis (Fig. 5) revealed surface irregularities due to the stepwise extraction of epi- and intracuticular waxes from the fruit cuticle, signifying alterations in the organization of the cuticular assembly.