Cutin, components

Hydroxy acids are major aliphatic components of cutin and suberin and these are readily identified by GLC-mass spectrometry. Major ions generated from the usual cutin components are listed in Kolattukudy (1977). The position of the hydroxyl group in the chain is easily seen because cleavage occurs on either side of the substituent (Fig. 6.13). The rather simple phenolic compounds yielded by reductive depolymerization of cutin and suberin are also very... 

Plants were probably the first to have polyester outerwear, as the aerial parts of higher plants are covered with a cuticle whose structural component is a polyester called cutin. Even plants that live under water in the oceans, such as Zoestra marina, are covered with cutin. This lipid-derived polyester covering is unique to plants, as animals use carbohydrate or protein polymers as their outer covering. Cutin, the insoluble cuticular polymer of plants, is composed of inter-esterified hydroxy and hydroxy epoxy fatty acids derived from the common cellular fatty acids and is attached to the outer epidermal layer of cells by a pectinaceous layer (Fig. 1). The insoluble polymer is embedded in a complex mixture of soluble lipids collectively called waxes [1], Electron microscopic examination of the cuticle usually shows an amorphous appearance but in some plants the cuticle has a lamellar appearance (Fig. 2). 

Fig. 3. (Top left) Chemical methods used to depolymerize the polyesters. (Top right) Thin-layer and gas-liquid chromatograms (as trimethylsilyl derivatives) of the monomer mixture obtained from the cutin of peach fruits by LiAlD4 treatment. In the thin-layer chromatogram the five major spots are, from the bottom, C18 tetraol, C16 triol, and C18 triol (unresolved), diols, and primary alcohol. Nx = C16 alcohol N2= C18 alcohol Mj = C16 diol M2 = C18 diol D = C16 triol D2 and D3 = unsaturated and saturated C18 triol, respectively, T4 and T2, unsaturated and saturated C18 tetraol, respectively. (Bottom) Mass spectrum of component D3 in the gas chromatogram. BSA = bis-N,O-trimethylsilyl acetamide...

The most common major components of cutin are derivatives of saturated C16 (palmitic) acid and unsaturated C18 acids (Fig. 4). The major component of the C16 family of acids is 9- or 10,16-dihydroxyhexadecanoic acid (and some mid-chain positional isomers), with less 16-hydroxyhexadecanoic acid and much smaller amounts of hexadecanoic acid. In some cases other derivatives are significant constituents. For example, in citrus cutin 16-hydroxy-10-oxo-C16 acid, and in young Vicia faba leaves 16-oxo-9 or 10-hydroxy C16 acid are significant... 

Table 1. Fatty acids with one or more additional functional groups that have been reported as components of cutin or suberin3. Adapted from [16]...

The unique suberin components that are not found as significant components of cutin are the very long chain molecules and the dicarboxylic acids. Therefore, chain elongation and conversion of co-hydroxy acids to the corresponding dicarboxylic acids constitute two unique biochemical processes involved in the synthesis of suberin. Incorporation of labeled acetate into the very long chain components of suberin was demonstrated and this ability developed during suberization in potato tuber disks [73]. The enzymes involved... 

The major function of cutin is to serve as the structural component of the outer barrier of plants. As the major component of the cuticle it plays a major role in the interaction of the plant with its environment. Development of the cuticle is thought to be responsible for the ability of plants to move onto land where the cuticle limits diffusion of moisture and thus prevents desiccation [141]. The plant cuticle controls the exchange of matter between leaf and atmosphere. The transport properties of the cuticle strongly influences the loss of water and solutes from the leaf interior as well as uptake of nonvolatile chemicals from the atmosphere to the leaf surface. In the absence of stomata the cuticle controls gas exchange. The cuticle as a transport-limiting barrier is important in its physiological and ecological functions. The diffusion across plant cuticle follows basic laws of passive diffusion across lipophylic membranes [142]. Isolated cuticular membranes have been used to study this permeability and the results obtained appear to be valid... 

Kolattukudy PE, Espelie KE (1985) Biosynthesis of cutin, suberin, and associated waxes. In Higuchi T (ed) Biosynthesis and biodegradation of wood components. Academic Press, New York p 161... 

The aliphatic components of SOM, derived from various sources, tend to persist in soil (Almendros et al. 1998 Lichtfouse et al. 1998a Lichtfouse et al. 1998b Mosle et al. 1999 Poirier et al. 2000). The principal source of aliphatic materials in soil is plant cuticular materials, especially cutin, an insoluble polyester of cross-linked hydroxy-fatty acids and hydroxy epoxy-fatty acids (Kolattukudy 2001). Some plant cuticles also contain an acid and base hydrolysis-resistant biopolymer, comprised of aliphatic chains attached to aromatic cores known as cutan (Tegelaar et al. 1989 McKinney et al. 1996 Chefetz 2003 Sachleben et al. 2004). 

Kolattukudy, P. E. Espelie, K. E. Biosynthesis of Cutin, Suberin and Associated Waxes In Biosynthesis and Biodegradation of Wood Components Higuchi, T., Ed Academic Press New York, 1985 pp. 161-207. 

High-resolution 13C NMR studies have been conducted on intact cuticles from limes, suberized cell walls from potatoes, and insoluble residues that remain after chemical depolymerization treatments of these materials. Identification and quantitation of the major functional moieties in cutin and suberin have been accomplished with cross-polarization magic-angle spinning as well as direct polarization methods. Evidence for polyester crosslinks and details of the interactions among polyester, wax, and cell-wall components have come from a variety of spin-relaxation measurements. Structural models for these protective plant biopolymers have been evaluated in light of the NMR results. 

Cutin-Wax Interactions. In order to obtain a more complete structural picture of plant cuticle, 13C CPMAS data were also obtained for the polymeric assembly prior to removal of waxes (Figure 5). A second (CH2)n peak appeared in the spectrum, and additional signal intensity in the carboxyl region produced a single broadened peak. Bulk methylene carbons from cutin and wax components exhibited identical values of Tip(H), indicating that they were mixed intimately and shared a common 1H spin reservoir... 

Suberized Cell Walls. An analogous set of CPMAS experiments is presented for suberin in Figure 6. Because this polymer is an integral part of the plant cell wall, the 13C NMR spectrum had contributions from both polysaccharide and polyester components. Chemical-shift assignments, summarized in Table IV, demonstrated the feasibility of identifying major polyester and sugar moieties despite serious spectral overlap. Semiquantitative estimates for the various carbon types indicated that, as compared with cutin, the suberin polyester had dramatically fewer aliphatic and more aromatic residues. A similar observation was made previously for the soluble depolymerization products of these plant polymers (1,8,11). 

For the study of complex cuticular mixtures, measurements of cross-polarization dynamics proved to be especially informative. The equality of Ti/>(H) values in cutin-wax assemblies demonstrated that these cuticular materials were mixed intimately. By contrast, Ti >(H) measurements showed that the polymeric components of suberized cell walls were present in distinct domains, suggesting that suberin was attached at a few structural sites rather than being embedded in the polysaccharide wall. 

Surface lipids of plants. The thick cuticle (Fig. 1-6) that covers the outer surfaces of green plants consists largely of waxes and other lipids but also contains a complex polymeric matrix of cutin (stems and leaves) or suberin (roots and wound surfaces).135/135a Plant waxes commonly have C10 - C30 chains in both acid and alcohol components. Methyl branches are frequently present. A major function of the waxes is to inhibit evaporation of water and to protect the outer cell layer. In addition, the methyl branched components may inhibit enzymatic breakdown by microbes. Free fatty acids, free alcohols, aldehydes, ketones, 13-dike tones, and alkanes are also present in plant surface waxes. Chain lengths are usually C20 - C35.136 Hydrocarbon formation can occur in other parts of a plant as well as in the cuticle. Thus, normal heptane constitutes up to 98% of the volatile portion of the turpentine of Pin us jeffreyi.81... 

Nile Blue is used as a 0.01 to 0.1 %W/V aqueous solution and is simply added to or mixed with the substrate. The active component of the dye is actually a minor contaminant of the solution, not the blue-colored material [31]. The preparations are viewed with 450-490 nm excitation (an FTTC filter set. Figure 6). Emulsion stability is sometimes an issue in the presence of the cationic blue component of Nile Blue. In this case we use Nile Red, the pure form of this colorant. Nile Red solution is made fresh from a stock solution (0.1%W/V in acetone). This stock is added dropwise to water until a moderate blue color is seen and the solution is used immediately (it deteriorates quickly). For either colorant, the active molecule is fluorescent only when it is in a suitably hydrophobic environment. This usually means neutral lipid droplets [31] but other sites (aggregates of surfactants, the center of casein micelles, cutin plates in some seeds) are possibilities. 

The two major polymeric lipid components found in plant cuticles are cutin and cutan. Whereas cutin is the polyester biopolymer that is solubilized upon saponification treatment, cutan is a nonsaponifiable and nonextractable polymeric substance... 

Rontani, J.F., de Rabourdin, A., Pinot, F., Kandel, S. and Aubert, C. (2005) Visible light-induced oxidation of unsaturated components of cutins a significant process during the senescence of higher plants. Phytochemistry 66(3), 31 3-321. 

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