A simple photometric device analysing cuticular transport physiology: surfactant effect on permeability of isolated cuticular membranes of Prunus laurocerasus L.

4-Nitrophenol permeabilities of astomatous cuticular membranesisolated from the upper surface of Prunus laurocerasus L. leaveswere measured applying a newly developed photometric device.Isolated cuticles were mounted between donor and receiver compartmentsof a stainless steel transport chamber. 4-Nitrophenol was appliedas non-dissociated species in citric buffer at pH 3.0 in thedonor compartment and sampled as dissociated species in thereceiver compartment in borate buffer at pH 9.0. Permeances,calculated from steady-state rates of 4-nitrophenol permeation,ranged from 1.73 10^–10 m s^–1 up to 38.410^–10ms^–1. They were in the same order of magnitude comparedto published permeances obtained with a different method usingradiolabelled 4-nitrophenol and isolated cuticles of Citrusaurantium L. In the presence of the surfactant Brij 30, whichis a polydisperse alcohol ethoxylate, cuticular permeabilitiesincreased on average by a factor of 37. Cuticles, initiallyhaving the lowest permeabilities, exhibited the highest increaseof their permeabilities due to the surfactant and vice versa.This increase ofcuticular permeabilities in the presence ofa surfactant is interpreted as a plasticizing effect of thesurfactant molecules on the cuticular wax forming the cuticulartransport barrier. Furthermore, surfactant-induced increasesof cuticular permeabilities were reversible to a large extent.Permeabilities decreased again after the removal of Brij 30reaching final values about 6-times higher compared to the initialpermeabilities. This demonstrates that the surfactant and thepermeating molecule must be present simultaneously in the cuticlein order to enhance cuticular permeation. Possible applicationsof this simple photometric device analysing further aspectsof cuticular transport physiology are finally suggested. Key words: Cuticular transport, leaf surface, permeability, plant cuticle, surfactant     

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.     

Water permeability of plant cuticles: permeance,diffusion and partition coefficients

Summary Using isolated cuticular membranes from ten woody and herbaceous plant species, permeance and diffusion coefficients for water were measured, and partition coefficients were calculated. The cuticular membranes of fruit had much higher permeance and diffusion coefficients than leaf cuticular membranes from either trees or herbs. Both diffusion and partition coefficients increased with increasing membrane thickness. Thin cuticles, therefore, tend to be better and more efficient water barriers than thick cuticles. We compared the diffusion coefficients and the water content of cuticles as calculated from transport measurements with those obtained from water vapor sorption. There is good to fair agreement for cuticular membranes with a low water content, but large discrepancies appear for polymer matrix membranes with high permeance. This is probably due to the fact that diffusion coefficients obtained from transport measurements on membranes with high permeance and water content are underestimated. Water permeabilities of polyethylene and polypropylene membranes are similar to those of leaf cuticular membranes. However, leaf cuticles have much lower diffusion coefficients and a much greater water content than these synthetic polymers. This suggests that cuticles are primarily mobility barriers as far as water transport is concerned.     

A radioactive assay allowing the quantitative measurement of cuticular permeability of intact <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> leaves

Cuticular penetration of five different ¹⁴C-labeled chemicals (benzoic acid, bitertanole, carbaryl, epoxiconazole and 4-nitrophenol) into Arabidopsis thaliana leaves was measured and permeances P (ms⁻¹) were calculated. Thus, cuticular barrier properties of A. thaliana leaves have been characterized quantitatively. Epoxiconazole permeance of A. thaliana was 2.79 × 10⁻⁸ ms⁻¹. When compared with cuticular permeances measured with intact stomatous and astomatous leaf sides of Prunus laurocerasus, frequently used in the past as a model species studying cuticular permeability, A. thaliana has a 48- to 66-fold higher permeance. When compared with epoxiconazole permeability of isolated cuticles of different species (Citrus aurantium, Hedera helix and P. laurocerasus) A. thaliana permeability is between 17- to 199-fold higher. Co-permeability experiments, simultaneously measuring ¹⁴C-epoxiconazole and ³H₂O permeability of isolated cuticles of three species (C. aurantium, H. helix and P. laurocerasus) showed that ³H₂O permeability was highly correlated with epoxiconazole permeability. The regression equation of this correlation can be used predicting cuticular transpiration of intact stomatous leaves of A. thaliana, where a direct measurement of cuticular permeation using ³H₂O is impossible. Water permeance estimated for A. thaliana was 4.55 × 10⁻⁸ ms⁻¹, which is between 12- and 91-fold higher than water permeances measured with isolated cuticles of C. aurantium, H. helix and P. laurocerasus. This indicates that cuticular water permeability of the intact stomatous leaves of the annual species A. thaliana is fairly high and in the upper range compared with most P values of perennial species published in the past.     

Structure and dynamics of reconstituted cuticular waxes of grape berry cuticle (Vitis vinifera L.)

Cuticular waxes from grape berry cuticle of Vitis vinifera L. have been investigated by X-ray diffraction, differential scanning calorimetry and gas chromatography. The waxes were mainly composed of n-alcohols and n-fatty acids and a considerable amount (about 30%) of the cyclic terpenoid oleanolic acid. The physical techniques used showed that the waxes have a high degree of molecular order in spite of the presence of the cyclic component. Molecular dynamics of the reconstituted waxes were measured to characterize the transport properties of the cuticular waxes that form the transport-limiting barrier of plant cuticles. For this purpose, the diffusion coefficient of labelled cholesterol, imitating the terpenoic acid, was measured. The value obtained was around 10^-21 m² s^-1 indicating a low mobility of the cyclic part of the reconstituted waxes. Temperature dependence of the diffusion coefficient was studied in the range of 5-45C. Arrhenius plot analysis yielded a high activation energy, 196.4 kJ mol^-1, of the diffusion process. This indicates dense molecular packing of reconstituted cuticular waxes.     

Isolation of cuticular membranes from various conifer needles

Summary A method of isolating intact needle cuticles is presented. Cuticles were separated enzymatically from needles of Abies alba Mill., Picea abies (L.) Karst., Picea pungens Engelm., Pinus mugo Turra, and Taxus baccata L. Cuticle separation depended on the enzyme concentration, the developmental stage of the needles and the duration of incubation in the hydrolytic pectinase/cellulase solution. Cuticles could not be removed from needles older than 2 years. Scanning electron micrographs of enzymatically isolated cuticles are presented. The permeance coefficients for water and oxygen transport across the isolated cuticular membranes indicate their functional intactness. But permeance coefficients also show that isolation of cuticular membranes with chromic acid is an unacceptable method, since they are lo longer structurally or functionally intact following isolation by this method.     

Transport properties of cuticular waxes of Fagus sylvatica L. and Picea abies (L.) Karst.: Estimation of size selectivity and tortuosity from diffusion coefficients of aliphatic molecules

Diffusion coefficients for thirteen lipophilic organic compounds in reconstituted waxes of Fagus sylvatica L. and Picea abies (L.) Karst. were measured to characterise the transport properties of the cuticular waxes that form the transport-limiting barrier of plant cuticles. Desorption kinetics (relative amounts desorbed versus time) were asymptotic, but could be linearized up to 50% desorption by plotting relative amounts desorbed versus the square root of time. Diffusion coefficients calculated from the slopes of the linear regressions ranged from 10^–22 to 10^–17 m²·s^–1 and decreased with increasing molecular size. This size dependency of diffusion coefficients was analysed, assuming an exponential dependence of the diffusion coefficients on molar volumes, which allowed cuticular transport properties to be related to the physical structure of the wax. Furthermore, the fact that the barrier properties of Fagus wax are less pronounced than those of Picea is interpreted as an ecological adaptation of the respective tree species to their habitats at the level of their cuticular transport barriers.Abbreviation D diffusion coefficient The authors gratefully acknowledge financial support by the Schwerpunkt Programm Baumphysiologie of the Deutsche Forschungsgemeinschaft.     

Co-permeability of 3H-labelled water and 14C-labelled organic acids across isolated Prunus laurocerasus cuticles: effect of temperature on cuticular paths of diffusion

Co‐permeability of ³H‐labelled water and ¹⁴C‐labelled benzoic acid or 2,4‐dichlorophenoxyacetic acid across isolated cuticular membranes of Prunus laurocerasus L. was measured at temperatures ranging from 15 to 50 °C. The water and benzoic acid permeances were highly correlated over the whole temperature range investigated, whereas water and 2,4‐dichlorophenoxyacetic acid permeances were only correlated between 15 and 30 °C. The activation energies of cuticular permeability calculated from Arrhenius plots were 40 kJ mol^?1 for water and benzoic acid and 115 kJ mol^?1 for 2,4‐dichlorophenoxyacetic acid. The slopes of the Arrhenius plots of 2,4‐dichlorophenoxyacetic acid were linear between 15 and 50 °C, whereas pronounced phase transitions around 30 °C were observed for water and benzoic acid permeability. However, with isolated polymer matrix membranes, where cuticular waxes forming the transport‐limiting barrier of cuticles have been extracted, phase transitions were not observed for water and benzoic acid. It is concluded that temperatures above 30 °C caused structural changes in the transport‐limiting barrier of the cuticles leading to additional paths of diffusion for water and benzoic acid but not for 2,4‐dichlorophenoxyacetic acid.     

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.     

The Morphology and Permeability of Isolated Cuticular Membranes of Hoya carnosa R. Br. (Asclepiadaceae)

This investigation is in part an extension of previous leafcuticle observations made on 52 other taxa among 34 families.Dewaxed, chemically isolated, adaxial and abaxial cuticularmembranes and transverse leaf sections of the wax-flower plant(Hoya carnosa R. Br.) were examined using ordinary stainingtechniques and light-microscopy methods. Evidence is presentedfor the existence of ubiquitous, discrete, naturally occurringcuticular pores, concomitant with anticlinally oriented trans-cuticularcanals, distributed randomly throughout the cuticular matrix.The surface of the adaxial cuticular membrane contains approx.6540 unclustered pores per mm², the abaxial approx. 4680 poresper mm². Pore and canal diameters range between 0.5 and 0.75µm. The canals are often arcuate and their lengths aredirectly related to cuticle thickness. No correlations werefound between cuticle thickness and either pore numbers or poreand canal diameters. Based upon experiments with various pHindicators, solutions, and stains, the dewaxed, dry cuticularmembrane of H. carnosa appears to be both distinctly hydrophilicand selectively permeable through a myriad of microscopicallyvisible pores and canals permeating its matrix. A de novo interpretationof gross cuticle morphology based solely upon light microscopyobservations is presented by semi-diagrammatic illustrations. Hoya carnosa R. Br., wax-flower (wax-plant), cuticular membranes, cuticular pores, transcuticular canals, permeability     

Apple Fruit Cuticles and the Occurrence of Pores and Transcuticular Canals

Developmental studies were made on the dewaxed thin-sectionedapple fruit cuticles of 10 Malus sylvestris Mill, cultivarsfrom 4 weeks before anthesis through fruit maturation and harvest.Cuticular development appears to correlate well with the generalgrowth of the fruit. However, no correlation exists betweenfruit size and cuticle thickness. Cuticular pores were evidentas early as 1 week before anthesis and transcuticular canalsbecame evident by 1 week following anthesis. Dewaxed thin-sectioned,as well as isolated, mature fruit cuticles of 16 cultivars andfour crab-apples consistently revealed the distinct presenceof ubiquitous pores and canals. Evidence is provided by lightmicroscopy and scanning electron microscopy (SEM). Measurementswere taken of cuticle thicknesses during development and ofcuticular pore dimensions, and calculations were made of poreand canal numbers. Fruit size alone is not directly indicativeof total pore numbers per surface area. Canal lengths are directlyrelated to the developmental thickness of the cuticle. No correlationwas found between the thickness of the mature cuticle and eitherthe number of pores present or the pore diameters. Malus sylvestris Mill, apple fruit, morphology, cuticle, cuticular flanges, cuticular pores, transcuticular canals, ultrastructure     

Cuticular Pores and Transcuticular Canals in Diverse Fruit Varieties

Dewaxed thin-sectioned and dewaxed isolated mature fruit cuticlesrevealed the unequivocal presence in situ of visibly discrete,ubiquitous, cuticular pores or orifices concomitant with anticlinally-orientedtranscuticular canals in 51 varieties of fruit among 20 plantfamilies. More than 66 per cent of the fruit cuticles have poresand/or canals. No correlation exists between either fruit sizeor pore size and cuticle thickness. Dewaxed cuticles rangedfrom 1.25–22.5 µm in thickness. Canal lengths aredirectly related to cuticle thickness. Cuticular occlusionsof the epidermal cells were found in 76 per cent of the fruitsexamined. Evidence is provided by light microscopy photomicrographs. Fruit cuticles, cuticle morphology, cuticular pores, transcuticular canals     

Phase behaviour and crystallinity of plant cuticular waxes studied by Fourier transform infrared spectroscopy

The phase behaviour of cuticular waxes from leaves of Hedera helix L. and Juglans regia L. was studied by Fourier transform infrared spectroscopy. For this purpose reconstituted waxes, isolated cuticular membranes, dewaxed polymer matrix membranes and whole leaves were studied in the horizontal attenuated total reflection and transmission modes. Melting curves of cuticular waxes were derived from temperature-dependent changes in the absorption maximum of the symmetric stretching mode of CH₂ groups (ν_s, at approx. 2856–2848 cm⁻¹). With increasing temperature absorption band doublets due to CH₂ scissoring (δ_sciss) and rocking (δ_rock) movements (at approx. 1473–1471 and 730–720 cm⁻¹, respectively) indicative of an orthorhombic arrangement of alkyl chains merged into a single peak. The area ratio of the peaks at approx. 720 and 730 cm⁻¹ was used as a measure for aliphatic crystallinity of plant cuticular waxes at a given temperature. The investigations of reconstituted cuticular waxes and those still embedded in isolated cuticles or in situ on the leaf produced comparable results. The findings are discussed in terms of the properties of the cuticular transport barrier. Received: 21 March 1997 / Accepted: 25 April 1997     

The positional sterile (ps) mutation affects cuticular transpiration and wax biosynthesis of tomato fruits

Cuticular waxes are known to play a pivotal role in limiting transpirational water loss across primary plant surfaces. The astomatous tomato fruit is an ideal model system that permits the functional characterization of intact cuticular membranes and therefore allows direct correlation of their permeance for water with their qualitative and quantitative composition. The recessive positional sterile (ps) mutation, which occurred spontaneously in tomato (Solanum lycopersicum L.), is characterized by floral organ fusion and positional sterility. Because of a striking phenotypical similarity with the lecer6 wax mutant of tomato, which is defective in very-long-chain fatty acid elongation, ps mutant fruits were analyzed for their cuticular wax and cutin composition. We also examined their cuticular permeance for water following the developmental course of fruit ripening. Wild type and ps mutant fruits showed considerable differences in their cuticular permeance for water, while exhibiting similar quantitative wax accumulation. The ps mutant fruits showed a five- to eightfold increase in water loss per unit time and surface area when compared to the corresponding wild type fruits. The cuticular waxes of ps mutant fruits were characterized by an almost complete absence of n-alkanes and aldehydes, with a concomitant increase in triterpenoids and sterol derivatives. We also noted the occurrence of alkyl esters not present in the wild type. Quantitative and qualitative cutin monomer composition remained largely unaffected. The significant differences in the cuticular wax composition of ps mutant fruits induced a distinct increase of cuticular water permeance. The fruit wax compositional phenotype indicates the ps mutation is responsible for effectively blocking the decarbonylation pathway of wax biosynthesis in epidermal cells of tomato fruits.     

Characterization of hydrophilic and lipophilic pathways of Hedera helix L. cuticular membranes: permeation of water and uncharged organic compounds

The permeability of astomatous leaf cuticular membranes of Hedera helix L. was measured for uncharged hydrophilic (octanol/water partition coefficient log K(O/W) < or =0) and lipophilic compounds (log K(O/W) >0). The set of compounds included lipophilic plant protection agents, hydrophilic carbohydrates, and the volatile compounds water and ethanol. Plotting the mobility of the model compounds versus the molar volume resulted in a clear differentiation between a lipophilic and a hydrophilic pathway. The size selectivity of the lipophilic pathway was described by the free volume theory. The pronounced tortuosity of the diffusional path was caused by cuticular waxes, leading to an increase in permeance for the lipophilic compounds after wax extraction. The size selectivity of the hydrophilic pathway was described by hindered diffusion in narrow pores of molecular dimensions. A distinct increase in size selectivity was observed for hydrophilic compounds with a molar volume higher than 110 cm3 mol(-1). Correspondingly, the size distribution of passable hydrophilic pathways was interpreted as a normal distribution with a mean pore radius of 0.3 nm and a standard deviation of 0.02 nm. The increased permeance of the hydrophilic compounds by the removal of cuticular waxes was attributed to an increase in the porosity, a decrease in the tortuosity, and a widening of the pore size distribution. Cuticular transpiration resulted from the permeation of water across the hydrophilic pathway. The far-reaching implications of two parallel pathways for the establishment of correlations between cuticular structure, chemistry, and function are discussed.     

Co-Permeability of 3H-Labeled Water and 14C-Labeled Organic Acids across Isolated Plant Cuticles : Investigating Cuticular Paths of Diffusion and Predicting Cuticular Transpiration

Penetration of ³H-labeled water (³H₂O) and the ¹⁴C-labeled organic acids benzoic acid ([¹⁴C]BA), salicylic acid ([¹⁴C]SA), and 2,4-dichlorophenoxyacetic acid ([¹⁴C]2,4-D) were measured simultaneously in isolated cuticular membranes of Prunus laurocerasus L., Ginkgo biloba L., and Juglans regia L. For each of the three pairs of compounds (³H₂O/[¹⁴C]BA, ³H₂O/[¹⁴C]SA, and ³H₂O/[¹⁴C]2,4-D) rates of cuticular water penetration were highly correlated with the rates of penetration of the organic acids. Therefore, water and organic acids penetrated the cuticles by the same routes. With the combination ³H₂O/[¹⁴C]BA, co-permeability was measured with isolated cuticles of nine other plant species. Permeances of ³H₂O of all 12 investigated species were highly correlated with the permeances of [¹⁴C]BA (r² = 0.95). Thus, cuticular transpiration can be predicted from BA permeance. The application of this experimental method, together with the established prediction equation, offers the opportunity to answer several important questions about cuticular transport physiology in future investigations.     

Studies on Foliar Penetration: VIII. EFFECTS OF CHLORINATION ON THE MOVEMENT OF PHENOXYACETIC AND BENZOIC ACIDS THROUGH CUTICLES ISOLATED FROM THE FRUITS OF LYCOPERSICON ESCULENTUM L.

The effects of chlorine substitution on the movement of phenoxyaceticand benzoic acids through enzymatically-isolated cuticles ofLycopersicon fruits were determined by following the transferof each acid containing ¹⁴C from a donor to a receiver solution.This cuticle is characterized by an isotropic cutin matrix,within which patches of birefringent cuticular waxes are foundnear the outer surface. The outer, morphological surface isrelatively smooth while at the junction with the outer wallsof the epidermal cells there is extensive cuticular developmentextending down between the anticlinal walls. The epicuticularwax appears as a soft sheet-like covering of which the surfaceis relatively featureless. Chlorination of phenoxyacetic acid results in an enhanced transferacross the isolated cuticle. The order was 2,4,5- and 2,4,6-trichlorophenoxyacetic> 2,4- and 3,5-dichlorophenoxyacetic > 2-chlorophenoxyacetic> phenoxyacetic acid. Removal of the epicuticular wax resultedin greater permeability for all compounds; transfer of the morepolar acids was favoured. In contrast, chlorination of benzoicacid decreases passage through the cuticle; the rate is highestfor benzoic acid followed in descending order by 2-chlorobenzoic,2,4- and 2,5-dichlorobenzoic and 2,3,6-trichlorobenzoic acid.Chlorination also depresses the passage of both phenoxyaceticand benzoic acid through a dialysis membrane. The effects ofchlorination on the lipid solubility of both series of compoundsare discussed in relation to differences in transfer acrossthe cuticle.     

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.     

Cuticular water permeability and its physiological significance

Cuticles act as solution-diffusion membranes for water transport.Diffusion in pores does not contribute to cuticular transpiration.An extensive literature survey of cuticular permeances (P) andminimum leaf conductances (g_min) to water is presented. Thetwo variables cannot be distinguished with most experimentaltechniques. Results from different experiments are in good agreementwith each other for some species, for example, Fagus sytvaticaL., but not for others, such as Picea abies (L.) Karst. In adata set of 313 values of P or g_min from 200 species, distributionsof results obtained with different techniques were found todiffer significantly. Likely reasons include water loss fromincompletely closed or incompletely sealed stomata, and thedependence of P on moisture content of the cuticle and on storagetime of isolated cuticles. Contrasting evidence for an interactionbetween cuticular transpiration and stomatal sensitivity toair humidity is presented. The occurrence of unusually highg_min in trees growing at the alpine treeline and its physiologicalsignificance are discussed. It is shown that g_min is of littlevalue as a predictor for drought resistance of crops, with thepossible exception of Sorghum bicolor L. Moench. Possible wateruptake from fog or dew across cuticles is considered briefly. Key words: Epidermal conductance, VPD-response, water absorption, waxes, winter desiccation     

Leaf cuticular waxes are arranged in chemically and mechanically distinct layers: evidence from Prunus laurocerasus L.

The composition and spatial arrangement of cuticular waxes on the leaves of Prunus laurocerasus were investigated. In the wax mixture, the triterpenoids ursolic acid and oleanolic acid as well as alkanes, fatty acids, aldehydes, primary alcohols and alcohol acetates were identified. The surface extraction of upper and lower leaf surfaces yielded 280 mg m ^{? 2} and 830 mg m ^{? 2}, respectively. Protocols for the mechanical removal of waxes from the outermost layers of the cuticle were devised and evaluated. With the most selective of these methods, 130 mg m ^{? 2} of cuticular waxes could be removed from the adaxial surface before a sharp, physically resistant boundary was reached. Compounds thus obtained are interpreted as ‘epicuticular waxes’ with respect to their localization in a distinct layer on the surface of the cutin matrix. The epicuticular wax film can be transferred onto glass and visualized by scanning electron microscopy. Prunus laurocerasus epicuticular waxes consisted entirely of aliphatic compounds, whereas the remaining intracuticular waxes comprised 63% of triterpenoids. The ecological relevance of this layered structure for recognition by phytotrophic fungi and herbivorous insects that probe the surface composition for sign stimuli is discussed.