Water permeability of plant cuticular membranes: the effects of humidity and temperature on the permeability of non-isolated cuticles of onion bulb scales

Abstract. Water permeability of cuticular membranes (CM) from the inner bulb scales of Allium cepa has been investigated. CM have a thickness ranging from 0.6 to 1.3 μm. They are composed of a thin (120–200 nm) lamellated cuticle proper and a thicker (300–900 nm) cuticular layer. Permeability coefficients for diffusion of water across these thin membranes are very low (4 × lO⁻¹⁰ms⁻¹⁰). There was no difference in permeability of CM from successive scales of the same onion. Extraction of soluble cuticular lipids (SCL) with chloroform increased permeability by a factor of 1350 to 2050. Preliminary data indicate that only 1 μg cm⁻¹⁰ of SCL are removed by this treatment, hydrocarbons being the main (75%) consistuent. Permeability coefficients of cuticular transpiration were little affected by relative humidity, showing that transport is limited by a hydrophobic barrier that lacks dipoles. However, following extraction, permeability of the membranes depended strongly on humidity due to the presence of polar functional groups in the polymer matrix. Soluble cuticular lipids undergo a phase transition around 47°C. Temperatures higher than that irreversibly increased water permeability.     

A new technique for measurement of water permeability of stomatous cuticular membranes isolated from Hedera helix leaves

Transpiration of cuticular membranes isolated from the lower stomatous surface of Hedera helix (ivy) leaves was measured using a novel approach which allowed a distinction to be made between gas phase diffusion (through stomatal pores) and solid phase diffusion (transport through the polymer matrix membrane and cuticular waxes) of water molecules. This approach is based on the principle that the diffusivity of water vapour in the gas phase can be manipulated by using different gases (helium, nitrogen, or carbon dioxide) while diffusivity of water in the solid phase is not affected. This approach allowed the flow of water across stomatal pores ('stomatal transpiration') to be calculated separately from the flow across the cuticle (cuticular transpiration) on the stomatous leaf surface. As expected, water flux across the cuticle isolated from the astomatous leaf surface was not affected by the gas composition since there are no gas-filled pores. Resistance to flux of water through the solid cuticle on the stomatous leaf surface was about 11 times lower than cuticular resistance on the astomatous leaf surface, indicating pronounced differences in barrier properties between cuticles isolated from both leaf surfaces. In order to check whether this difference in resistance was due to different barrier properties of cuticular waxes on both leaf sides, mobility of 14C-labelled 2,4-dichlorophenoxy-butyric acid 14C-2,4-DB) in reconstituted cuticular wax isolated from both leaf surfaces was measured separately. However, mobility of 14C-2,4-DB in reconstituted wax isolated from the lower leaf surface was 2.6 times lower compared with the upper leaf side. The significantly higher permeability of the ivy cuticle on the lower stomatous leaf surface compared with the astomatous surface might result from lateral heterogeneity in permeability of the cuticle covering normal epidermal cells compared with the cuticle covering the stomatal cell surface.     

Effect of temperature on cuticular transpiration of isolated cuticular membranes and leaf discs

Cuticular transpiration was measured in the temperature range between 10 degrees C and 55 degrees C using tritiated water and five species (Vinca major L., Prunus laurocerasus L., Forsythia intermedia L., Citrus aurantium L., and Hedera helix L.). Cuticular water permeabilities measured with isolated cuticular membranes were not different from cuticular water permeabilities measured with leaf discs. Depending on the species cuticular water permeabilities increased by factors between 12 (V. major) to 264 (H. helix) when temperature was increased from 10 degrees C to 55 degrees C. Arrhenius plots (lnP versus 1/T) of all investigated species were characterized by phase transitions occurring in the temperature range of 30-39 degrees C. Activation energies for water permeability across plant cuticles below and above the midpoint of phase transition were calculated from Arrhenius plots. Depending on the species they varied between 26 (F. intermedia) to 61 kJ mol(-1) (H. helix) below the phase transition and from 67 (V. major) to 122 kJ mol(-1) (F. intermedia) above the phase transition. Since the occurrence of phase transitions always lead to significantly increased rates of cuticular transpiration it is argued that temperatures higher than 35 degrees C caused structural defects to the transport-limiting barrier of the plant cuticles of all species investigated.     

Ontogenetic and seasonal development of wax composition and cuticular transpiration of ivy (Hedera helix L.) sun and shade leaves

The ontogenetic and seasonal development of wax composition and cuticular transpiration of sun and shade leaves of ivy (Hedera helix L.) was analysed by investigating leaves varying in age between 4 and 202 d. It was discovered that the total amount of solvent-extractable wax was composed of two distinct fractions, separable by column chromatography: (i) a less polar or apolar monomeric wax fraction consisting of the typical linear, long-chain aliphatics usually described as cuticular wax components and (ii) a polar, oligomeric wax fraction consisting of primary alcohols and acids mostly esterified to C₁₂-, C₁₄- and C₁₆-ω-hydroxyfatty acids. The apolar wax fraction, which could be analysed directly by gas chromatography coupled with mass spectrometry (GC-MS), exhibited pronounced seasonal changes in composition. Wax amounts in the apolar fraction reached a maximum after about 30 d and gradually decreased again during the remaining period of the season investigated. In contrast, the polar wax fraction, which was analysable by GC-MS only after transesterification, rapidly increased early in the season, reaching a plateau after 40 d, and then remained constant during the rest of the season. Thus, total amounts of solvent-extractable cuticular waxes, which can be determined gravimetrically, will only be detected by GC-MS after fractionation and transesterification, a methodological approach rarely applied in the past in cuticular wax analysis. Additionally, investigation of the cutin polymer matrix after depolymerisation through transesterification, revealed that only those primary alcohols and acids forming an essential part of the apolar and the polar wax fractions were esterified during the investigated season and incorporated in increasing amounts into the cutin polymer matrix (matrix-bound wax fraction). Thus, it can be concluded that a complete analysis of cuticular wax of ivy and its seasonal development can only be achieved if all the relevant fractions (i) the less polar or apolar, (ii) the polar and (iii) the wax fraction bound to the cutin polymer matrix are investigated. Cuticular transpiration rapidly decreased within the first 30 d and essentially remained constant during the rest of the season. Thus, changes in cuticular water permeability were closely correlated with the most prominent changes in wax amounts and composition occurring during the first 30 d of ontogenetic leaf development. However, during the remainder of the year, up to 202 d, cuticular transport properties remained constant, although significant quantitative and qualitative changes in cuticular wax composition continued to occur. Thus, our study clearly demonstrated that there will be no simple relationship between chemical composition of cuticular waxes and transport properties of isolated ivy leaf cuticles. Received: 2 March 1998 / Accepted: 26 June 1998     

Water permeability of isolated cuticular membranes: The effect of cuticular waxes on diffusion of water

Summary The water permeability of astomatous cuticular membranes isolated from Citrus aurantium L. leaves, pear (Pyrus communis L.) leaves and onion (Allium cepa L.) bulb scales was determined before and after extraction of cuticular waxes with lipid solvents. In pear, the permeability coefficients for diffusion of tritiated water across cuticular membranes (CM) prior to extraction [P _d(CM)] decreased by a factor of four during leaf expansion. In all three species investigated P _d(CM) values of cuticular membranes from fully expanded leaves varied between 1 to 2×10^-7 cm^-3 s^-1·P _d(CM) values were not affected by pH. Extraction of cuticular waxes from the membranes increased their water permeability by a factor of 300 to 500. Permeability coefficients for diffusion of THO across the cutin matrix (MX) after extraction [P _d(MX)] increased with increasing pH. P _dvalues were not inversely proportional to the thickness of cuticular membranes. By treating the cutin matrix and cuticular waxes as two resistances acting in series it was shown that the water permeability of cuticles is completely determined by the waxes. The lack of the P _d(CM) values to respond to pH appeared to be due to structural effects of waxes in the cutin matrix. Cuticular membranes from the submerse leaves of the aquatic plant Potamogeton lucens L. were three orders of magnitude more permeable to water than the cuticular membranes of the terrestrial species investigated.Abbreviations CM cuticular membrane - MX cutin matrix - WAX waxes This study was supported by a grant from the Deutsche Forschungsgemeinschaft.     

Quantitative determination of water sorption by plant cuticles

Abstract. The water sorption by cuticular membranes (CM) isolated from both leaves and fruit and representing different structural and chemical cuticular types, was measured over the whole range of relative humidities using a magnetic suspension microbalance. The sorption isotherms were generally not linear and sorption increased more rapidly at the highest values of relative humidity. The highest values of water content, at 80–99% r.h., measured for the seven species ranged from 1.1 to 7.7% of the dehydrated weight. Extraction of the soluble cuticular lipids did not result in a decrease of sorption, but only in a lower interspecific dispersion. There was a drastic reduction (63%) in water sorption when polymer matrix membranes (MX) were partially acid-hydrolysed; but methylation or charging with iron of (MX) did not have any significant effect on water sorption. The data obtained are discussed in relation to cuticular permeability. Two determinants of water permeability were determined: the partition coefficient (K) relating the equilibrium Water concentration of the cuticle to that of the surrounding atmosphere; and the diffusion coefficient (D), calculated from the half-times of the sorption process in kinetic measurements.     

DEVELOPMENT OF THE CUTICULAR LAYERS IN ANGIOSPERM LEAVES

Schieferstein , R. H., and W. E. Loomis . (Iowa State U., Ames.) Development of the cuticular layer in angiosperm leaves. Amer. Jour. Bot. 46(9): 625–635. Illus. 1959.—The cuticularized layers of leaves and other plant surfaces consist of a primary cuticle, formed by the oxidation of oils on exposed cell walls, plus various surface and subsurface wax deposits. The primary cuticle appears to form rapidly on the walls of any living cell which is exposed to air. Surface wax is present on the mature leaves of about half of the 50 or 60 species studied. In general, wax is extruded at random through the newly formed cuticle of young leaves and accumulated in various reticulate to semicrystalline patterns. No wax pores through the cuticle or primary wall can be observed in electron-micrographs of dewaxed mature leaves. Wax accumulations on older leaves are generally subcuticular and may involve the entire epidermal wall. These deposits appear to be of considerably greater ecological significance than those on the surface. Isolated cuticular membranes from Hedera helix increased slightly in permeability to water with age of the leaf, but permeability to 2,4-D decreased 50 times. Evidence based on the patterns of cellulose in primary walls, of surface wax on growing leaves, of the appearance of the cuticle at the margins of growing epidermal cells, of the forms of the cuticle plates digested from growing and older leaves, and of the marginal location of new wax deposits on growing maize leaves is presented to support the thesis that the enlargement of the outer surface of the epidermal cells of leaves occurs at the margins of the surface. Earlier formed cuticle and wax are thus undisturbed during growth. These observations, coupled with evidence for apical growth in fibers, root hairs, etc. suggest that the primary walls of angiosperm cells are formed in specific, localized growth regions, rather than by plastic extension and apposition.     

Protecting against water loss: analysis of the barrier properties of plant cuticles

The cuticle is the major barrier against uncontrolled water loss from leaves, fruits and other primary parts of higher plants. More than 100 mean values for water permeabilities determined with isolated leaf and fruit cuticles from 61 plant species are compiled and discussed in relation to plant organ, natural habitat and morphology. The maximum barrier properties of plant cuticles exceed that of synthetic polymeric films of equal thickness. Cuticular water permeability is not correlated to the thickness of the cuticle or to wax coverage. Relationships between cuticular permeability, wax composition and physical properties of the cuticle are evaluated. Cuticular permeability to water increases on the average by a factor of 2 when leaf surface temperature is raised from 15 degrees C to 35 degrees C. Organic compounds of anthropogenic and biogenic origin may enhance cuticular permeability. The pathway taken by water across the cuticular transport barrier is reviewed. The conclusion from this discussion is that the bulk of water diffuses as single molecules across a lipophilic barrier while a minor fraction travels along polar pores. Open questions concerning the mechanistic understanding of the plant cuticular transport barrier and the role the plant cuticle plays in ensuring the survival and reproductive success of an individual plant are indicated.     

Differences between water permeability of astomatous and stomatous cuticular membranes: effects of air humidity in two species of contrasting drought-resistance strategy

Cuticular water permeabilities of adaxial and abaxial leaf surfaces and their dependence on relative air humidity (RH) applied in long-term and short-term regimes have been analysed for Hedera helix, native in a temperate climate, and Zamioculcas zamiifolia, native in subtropical regions. The water permeability of cuticular membranes (CM) isolated from the adaxial (astomatous) and abaxial (stomatous) leaf sides was measured using a method which allowed the separation of water diffusion through the remnants of the original stomatal pores from water diffusion through the solid cuticle. The long-term effects of low (20-40%) or high (60-80%) RH applied during plant growth and leaf ontogeny ('growth RH') and the short-term effects of applying 2% or 100% RH while measuring permeability ('measurement RH') were investigated. With both species, water permeability of the solid stomatous CM was significantly higher than the permeability of the astomatous CM. Adaxial cuticles of plants grown in humid air were more permeable to water than those from dry air. The adaxial CM of the drought-tolerant H. helix was more permeable and more sensitive to growth RH than the adaxial CM of Z. zamiifolia, a species avoiding water stress. However, permeability of the solid abaxial CM was similar in both species and independent of growth RH. The lack of a humidity response in the abaxial CM is attributed to a higher degree of cuticular hydration resulting from stomatal transpiration. The ecophysiological significance of higher permeability of the solid stomatous CM compared to the astomatous CM is discussed.     

In situ assembly of cuticular wax

Summary Cytochemical reactions within the primary cuticle (cutinised layer) indicate that the lamellae are formed from polar lipids. The electron microscope shows that the lamellae are involved in wax formation and it is suggested that the polar lipids provide in situ precursors for the synthesis of cuticular wax.     

Polar paths of diffusion across plant cuticles: new evidence for an old hypothesis

BACKGROUND: The plant cuticle is an extracellular lipophilic biopolymer covering leaf and fruit surfaces. Its main function is the protection of land-living plants from uncontrolled water loss. In the past, the permeability of the cuticle to water and to non-ionic lipophilic molecules (pesticides, herbicides and other xenobiotics) was studied intensively, whereas cuticular penetration of polar ionic compounds was rarely investigated. RECENT PROGRESS: Recent work measuring cuticular penetration of inorganic and organic ions is presented; the effects of molecular size of ions, temperature, wax extraction, humidity and plasticizers strongly support the conclusion that ions penetrate cuticles via water-filled pores. The cuticle covering stomata and trichomes forms the preferential site of ion penetration. This indicates that cuticles possess a pronounced lateral heterogeneity: the largest fraction of the cuticle surface is covered by the lipophilic domains of cutin and wax, but to a certain extent polar domains are also present in the cuticle, which form preferential sites of penetration for polar compounds. THE FUTURE: The chemical nature of these polar domains awaits detailed characterization, which will be of major importance in agriculture and green biotechnology, since polar paths of diffusion represent the most important transport routes for foliar-applied nutrients. Furthermore, many compounds acting as inducers of gene expression in transgenic plants are ionic and need to penetrate the cuticle via polar paths in order to be active.     

Rheological Properties of Enzymatically Isolated Tomato Fruit Cuticle

Rheological properties were determined for cuticular membranes (CMs) enzymatically isolated from mature tomato (Lycopersicon esculentum Mill. cv Pik Red) fruit. The cuticle responded as a viscoelastic polymer in stress-strain studies. Both CM and dewaxed CM expanded and became more elastic and susceptible to fracture when hydrated, suggesting that water plasticized the cuticle. Dewaxing of the CM caused similar changes in elasticity and fracturing, indicating that wax may serve as a supporting filler in the cutin matrix. Exposure of the cuticle to the surfactant Triton X-100 did not significantly affect its rheological properties.     

AgCl precipitates in isolated cuticular membranes reduce rates of cuticular transpiration

Counter diffusion of chloride, applied as NaCl at the inner side of isolated cuticles, and silver, applied as AgNO₃ at the outer side, lead to the formation of insoluble AgCl precipitates in isolated cuticles. AgCl precipitates could be visualized by light and scanning electron microscopy. The presence of AgCl precipitates in isolated cuticles was verified by energy dispersive X-ray analysis. It is argued that insoluble AgCl precipitates formed in polar pores of cuticles and as a consequence, cuticular transpiration of 13 out of 15 investigated species was significantly reduced up to three-fold. Water as a small and uncharged but polar molecule penetrates cuticles via two parallel paths: a lipophilic path, formed by lipophilic cutin and wax domains, and a aqueous pathe, formed by polar pores. Thus, permeances P (m s⁻¹) of water, which is composed of the two quantities P _Lipid and P _Pore, decreased, since water transport across polar pores was affected by AgCl precipitates. Cuticles with initially high rates of cuticular transpiration were generally more sensitive towards AgCl precipitates compared to cuticles with initially low rates of transpiration. Results presented here, significantly improves the current model of the structure of the cuticular transpiration barrier, since the pronounced heterogeneity of the cuticular transport barrier, composed of lipophilic as well as polar paths of diffusion, has to be taken into account in future.     

Composition of soluble cuticular lipids and water permeability of cuticular membranes from Citrus leaves

Water permeability and composition of soluble cuticular lipids of isolated cuticular membranes from leaves of Citrus aurantium L. were investigated for 3 successive years. The average water permeability coefficient determined using 169 cuticular membranes was 1.09·10^–7 cm s^–1 with a standard deviation of 0.78·10^–7 cm s^–1. There were no significant differences in water permeability between years. Cuticular membranes are characterized by a great variability in water permeability both within and between years. Both water permeability of individual membranes and variability between membranes are shown to be determined by soluble cuticular lipids contained within the cuticular membranes. The soluble cuticular lipids of Citrus leaves are composed of fatty acids, primary alcohols, esters, and hydrocarbons. They occur in amounts of 9.84 g cm^–2, which represents approx. 3% of the total mass of isolated cuticular membranes. The specific weight of cuticular membranes (365.4 g cm^–1) and total amount of soluble cuticular lipids did not vary significantly between years. Significant differences were observed for the amounts and composition of the constituent classes of lipids. Six homologues comprise 86% of the fatty acids (C₁₆; C₁₈; C₁₉; C₂₁; C₂₄; C₂₆), 83% of the primary alcohols (C₂₄; C₂₆; C₂₈; C₃₀; C₃₂; C₃₄) and 88% of the esters (C₃₆; C₃₈; C₄₀; C₄₁; C₄₂; C₄₄). Eleven major homologues amount only to 62% of the total hydrocarbons (C₁₆; C₁₇; C₁₈; C₂₀; C₂₆; C₂₇; C₂₉; C₃₀; C₃₁; C₃₂; C₃₃). Variability in the composition of soluble cuticular lipids between years was much smaller than variability of water permeability and, therefore, no relation between composition of soluble cuticular lipids and water permeability could be found. It is suggested that this may be due to the fact that the lipid composition observed represents the averages of 20 to 30 membranes analyzed so that differences between individual membranes may have been leveled out.Abbreviations CM cuticular membranes - MX polymer matrix - P_d permeability coefficient for diffusion of water - SCL soluble cuticular lipids - MES morpholinoethane sulphonic acid     

Water permeability of plant cuticles: The effect of temperature on diffusion of water

The effect of temperature on water permeability of plant cuticles (astomatous Citrus leaf cuticles) has been investigated. The Arrhenius plot (logarithm of the permeability coefficient vs. 1/temperature) has two linear portions that intersect at 44° C. Evidence is presented to show that this intersection represents the solid/liquid phase transition of cuticular lipids. As the Arrhenius plot has only one phase transition in the temperature range of 5 to 80° C, it appears that all soluble cuticular lipids in the cuticle are present as a homogeneous mixture rather than as individual layers differing in composition. This view is supported by electron spin resonance evidence showing homogenous distribution of spin label fatty acids. The original distribution of soluble cuticular lipids is irreversibly altered by heating cuticular membranes above the transition temperature. This is accompanied by an irreversible increase in water peremeability, demonstrating the importance of the structure of cuticular lipids with regard to cuticular permeability.Abbreviations CM cuticular membranes - MX polymer matrix - SCL soluble cuticular lipids - MES morpholinoethane sulphonic acid - J flux - ESR electron spin resonance - THO tritiated water     

Cuticular permeability of the three tree species Prunus Iaurocerasus L., Ginkgo biloba L. and Juglans regia L.: comparative investigation of the transport properties of intact leaves, isolated cuticles and reconstituted cuticular waxes

Cuticular transport properties of intact leaves, isolated cuticularmembranes and reconstituted cuticular waxes of the three treespecies Prunus laurocerasus L., Ginkgo biloba L. and Juglansregia L. were measured using six different ¹⁴C-labelled compounds,benzoic acid, salicylic acid, 2,4-dichlorophenoxy acid, metribuzin,4-nitrophenol, and atrazine. For the same compound and the samespecies, the permeance of the intact leaf and the isolated cuticlewas equal. This provides strong evidence demonstrating thattransport properties of cuticles are not altered during isolation.Additionally, diffusion coefficients of the ¹⁴C-labelled compoundsin isolated and subsequently reconstituted cuticular wax ofthe three tree species were measured. Permeances of intact leavesand isolated cuticles could be predicted from diffusion coefficients,wax/water partition coefficients and the thickness of the transport-limitingwax layer with a mean deviation of about 1.7. This providesevidence that transport properties of recrystallized cuticularwaxes do indeed reflect barrier properties of isolated cuticularmembranes and intact leaves with in situ waxes. Thus, it canbe concluded that the investigation of cuticular permeabilityusing the three independent experimental systems of differentcomplexity give comparable results. Finally, it was observedthat permeances and diffusion coefficients measured with P.laurocerasus were always significantly lower than those measuredwith G. biloba and J. regia. This is interpreted as an ecologicaladaptation of the respective species. The evergreen speciesP. laurocerasus must be more adapted to environmental stresssuch as drought and frost injury compared to the two deciduousspecies G. biloba and J. regia. Key words: Cuticular permeability, diffusion coefficient, leaf surface, permeance, plant cuticle, transport     

Determination of diffusion coefficients of octadecanoic acid in isolated cuticular waxes and their relationship to cuticular water permeabilities

Transport properties of cuticular waxes from 40 different plant species were investigated by measuring desorption rates of ¹⁴C-labelled octadecanoic acid from isolated and subsequently reconstituted wax. Diffusion coefficients (D) of octadecanoic acid in reconstituted waxes, calculated from the slopes of the regression lines fitted to the linearized portions of desorption kinetics, ranged from 1.2 × 10^?19 m² s^?1 (Senecio kleinia leaf) to 2.9 × 10^?17 m² s^?1 (Malus cf. domestica fruit). Cuticular water permeabilities (cuticular transpiration) measured with intact cuticular membranes isolated from 24 different species varied from 1.7 × 10^?11 m s^?1 (Vanilla planifolia leaf) up to 2.1 × 10^?9 m s^?1 (Malus cf. domestica fruit), thus covering a range of more than 2 orders of magnitude. Cuticular water permeabilities were highly correlated with diffusion coefficients of octadecanoic acid in isolated cuticular wax of the same species. It is therefore possible to estimate cuticular barrier properties of stomatous leaf surfaces or of leaves where isolation of the cuticle is impossible by measuring D of octadecanoic acid in isolated waxes of these leaves.     

The role of ABC transporters in cuticular lipid secretion

The aerial surfaces of plants are enveloped by a waxy cuticle, which among other functions serves as a barrier to limit non-stomatal water loss and defend against pathogens. The cuticle is a complex three-dimensional structure composed of cutin (a lipid polyester matrix) and waxes (very long chain fatty acid derivatives), which are embedded within and layered on top of the cutin matrix. Biosynthesis of cuticular lipids is believed to take place solely within aerial epidermal cells. Once synthesized, both the waxes and the cutin precursors must leave the cytoplasm, pass through the hydrophilic apoplastic space, and finally assemble to form the cuticle. These processes of secretion and assembly are essentially unknown. Initial steps toward our understanding of these processes were the characterization of CER5/ABCG12/WBC12 and more recently ABCG11/WBC11, a pair of ABC transporters required for cuticular lipid secretion. ABCG12 is involved in wax secretion, as mutations in this gene result in a lower surface-load of wax and a concomitant accumulation of lipidic inclusions within the epidermal cell cytoplasm. Mutations in ABCG11 result in a similar wax phenotype as cer5 and similar cytoplasmic inclusions. In contrast to cer5, however, abcg11 mutants also show significantly reduced cutin, post-genital organ fusions, and reduced growth and fertility. Thus, for the first time, a transporter is implicated in cutin accumulation. This review will discuss the secretion of cuticular lipids, focusing on ABCG12, ABCG11 and the potential involvement of other ABC transporters in the ABCG subfamily.     

Plant response to drought stress simulated by ABA application: Changes in chemical composition of cuticular waxes

Plant cuticles form the interface between epidermal plant cells and the atmosphere. The cuticle creates an effective barrier against water loss, bacterial and fungal infection and also protects plant tissue from UV radiation. It is composed of the cutin matrix and embedded soluble lipids also called waxes. Chemical composition of cuticular waxes and physiological properties of cuticles are affected by internal regulatory mechanisms and environmental conditions (e.g. drought, light, and humidity). Here, we tested the effect of drought stress simulation by the exogenous application of abscisic acid (ABA) on cuticular wax amount and composition. ABA-treated plants and control plants differed in total aboveground biomass, leaf area, stomatal density and aperture, and carbon isotope composition. They did not differ in total wax amount per area but there were peculiar differences in the abundance of particular components. ABA-treated plants contained significantly higher proportions of aliphatic components characterized by chain length larger than C₂₆, compared to control plants. This trend was consistent both between and within different functional groups of wax components. This can lead to a higher hydrophobicity of the cuticular transpiration barrier and thus decrease cuticular water loss in ABA-treated plants. At both ABA-treated and control plants alcohols with chain length C₂₄ and C₂₆ were predominant. Such a shift towards wax compounds having a higher average chain length under drought conditions can be interpreted as an adaptive response of plants towards drought stress.