Supplemental and highly elevated tocopherol doses differentially regulate allergic inflammation: reversibility of α-tocopherol and γ-tocopherol's effects

We have reported that supplemental doses of the α- and γ-tocopherol isoforms of vitamin E decrease and increase, respectively, allergic lung inflammation. We have now assessed whether these effects of tocopherols are reversible. For these studies, mice were treated with Ag and supplemental tocopherols in a first phase of treatment followed by a 4-wk clearance phase, and then the mice received a second phase of Ag and tocopherol treatments. The proinflammatory effects of supplemental levels of γ-tocopherol in phase 1 were only partially reversed by supplemental α-tocopherol in phase 2, but were completely reversed by raising α-tocopherol levels 10-fold in phase 2. When γ-tocopherol levels were increased 10-fold (highly elevated tocopherol) so that the lung tissue γ-tocopherol levels were equal to the lung tissue levels of supplemental α-tocopherol, γ-tocopherol reduced leukocyte numbers in the lung lavage fluid. In contrast to the lung lavage fluid, highly elevated levels of γ-tocopherol increased inflammation in the lung tissue. These regulatory effects of highly elevated tocopherols on tissue inflammation and lung lavage fluid were reversible in a second phase of Ag challenge without tocopherols. In summary, the proinflammatory effects of supplemental γ-tocopherol on lung inflammation were partially reversed by supplemental levels of α-tocopherol but were completely reversed by highly elevated levels of α-tocopherol. Also, highly elevated levels of γ-tocopherol were inhibitory and reversible in lung lavage but, importantly, were proinflammatory in lung tissue sections. These results have implications for future studies with tocopherols and provide a new context in which to review vitamin E studies in the literature.     

Photosynthetic apparatus of chilling-sensitive plants IX. The involvement of α-tocopherol in the electron transport chain and the anti-oxidizing system in chloroplasts of tomato leaves

1. The role of tocopherols in tomato chloroplasts from fresh, cold and dark-stored as well as stored and illuminated leaves was studied.2. The cold and dark storage of leaves results in a loss of chloroplast α- and γ-tocopherols of about 30–40% accompanied by an increase in chloroplast δ-tocopherol of about 40%. On illumination of stored leaves, an elevation of α- and γ-tocopherol level to about 110 and 95% of the control, respectively, occurs, whilst δ-tocopherol content is not affected.3. Experiments performed with 2,2-diphenyl-1-picrylhydrazyl-treated chloroplasts show that only about 70% of total α-tocopherol is functionally active in the electron transport of Photosystem II between the diphenyl-carbazide (DPC) donation site and the inhibition site of DBMIB.4. A small amount of α-tocopherol quinone (about 10% of α-tocopherol content) is found in chloroplasts from fresh, fresh and illuminated as well as cold and dark-stored tomato leaves, whereas the illumination of the latter increases the chloroplast α-tocopherol quinone content 3-fold. Moreover, following the illumination of chloroplasts from cold and dark-stored as well as stored and illuminated leaves, the oxidation of exogenous α-tocopherol to α-tocopherol quinone is 2-fold faster then in chloroplasts from fresh leaves.5. The primary product (‘α-tocopheroxide’) formed during the α-tocopherol oxidation by illuminated chloroplasts was identified as 8a-hydroxy-α-tocopheron.6. Exogenous α-tocopherol inhibits the lipid photoperoxidation by about 40–50% in chloroplasts from all three kinds of tomato leaf.7. The results seem to suggest that chloroplast α-tocopherol is involved in both electron transport of PS II and antioxidizing system of chloroplasts.     

Vitamin E isoforms directly bind PKCα and differentially regulate activation of PKCα

Vitamin E isoforms have opposing regulatory effects on leucocyte recruitment during inflammation. Furthermore, in vitro, vitamin E isoforms have opposing effects on leucocyte migration across endothelial cells by regulating VCAM (vascular cell-adhesion molecule)-1 activation of endothelial cell PKCα (protein kinase Cα). However, it is not known whether tocopherols directly regulate cofactor-dependent or oxidative activation of PKCα. We report in the present paper that cofactor-dependent activation of recombinant PKCα was increased by γ-tocopherol and was inhibited by α-tocopherol. Oxidative activation of PKCα was inhibited by α-tocopherol at a 10-fold lower concentration than γ-tocopherol. In binding studies, NBD (7-nitrobenz-2-oxa-1,3-diazole)-tagged α-tocopherol directly bound to full-length PKCα or the PKCα-C1a domain, but not PKCζ. NBD-tagged α-tocopherol binding to PKCα or the PKCα-C1a domain was blocked by diacylglycerol, α-tocopherol, γ-tocopherol and retinol, but not by cholesterol or PS (phosphatidylserine). Tocopherols enhanced PKCα-C2 domain binding to PS-containing lipid vesicles. In contrast, the PKCα-C2 domain did not bind to lipid vesicles containing tocopherol without PS. The PKCα-C1b domain did not bind to vesicles containing tocopherol and PS. In summary, α-tocopherol and γ-tocopherol bind the diacylglycerol-binding site on PKCα-C1a and can enhance PKCα-C2 binding to PS-containing vesicles. Thus the tocopherols can function as agonists or antagonists for differential regulation of PKCα.     

α-, γ- and δ-tocopherols reduce inflammatory angiogenesis in human microvascular endothelial cells

Vitamin E, a micronutrient (comprising α-, β-, γ- and δ-tocopherols, α-, β-, γ- and δ-tocotrienols), has documented antioxidant and non-antioxidant effects, some of which inhibit inflammation and angiogenesis. We compared the abilities of α-, γ- and δ-tocopherols to regulate human blood cytotoxicity (BEC) and lymphatic endothelial cytotoxicity (LEC), proliferation, invasiveness, permeability, capillary formation and suppression of TNF-α-induced VCAM-1 as in vitro models of inflammatory angiogenesis. α-, γ- and δ-tocopherols were not toxic to either cell type up to 40 μM. In BEC, confluent cell density was decreased by all concentrations of δ- and γ-tocopherol (10–40 μM) but not by α-tocopherol. LEC showed no change in cell density in response to tocopherols. δ-Tocopherol (40 μM), but not other isomers, decreased BEC invasiveness. In LEC, all doses of γ-tocopherol, as well as the highest dose of α-tocopherol (40 μM), decreased cell invasiveness. δ-Tocopherol had no effect on LEC invasiveness at any molarity. δ-Tocopherol dose dependently increased cell permeability at 48 h in BEC and LEC; α- and γ-tocopherols showed slight effects. Capillary tube formation was decreased by high dose (40 μM) concentrations of α-, γ- and δ-tocopherol, but showed no effects with smaller doses (10–20 μM) in BEC. γ-Tocopherol (10–20 μM) and α-tocopherol (10 μM), but not δ-tocopherol, increased LEC capillary tube formation. Lastly, in BEC, α-, γ- and δ-tocopherol each dose-dependently reduced TNF-α-induced expression of VCAM-1. In LEC, there was no significant change to TNF-α-induced VCAM-1 expression with any concentration of α-, γ- or δ-tocopherol. These data demonstrate that physiological levels (0–40 μM) of α-, γ- and δ-tocopherols are nontoxic and dietary tocopherols, especially δ-tocopherol, can limit several BEC and LEC endothelial behaviors associated with angiogenesis. Tocopherols may therefore represent important nutrient-signals that limit cell behaviors related to inflammation/angiogenesis, which when deficient, may predispose individuals to risks associated with elevated angiogenesis such as inflammation and cancer; further differences seen from the tocopherols may be due to their blood or lymphatic cell origin.     

GmTMT2a from soybean elevates the α-tocopherol content in corn and Arabidopsis

Tocochromanol, or vitamin E, plays a crucial role in human and animal nutrition and is synthesized only by photosynthetic organisms. γ-Tocopherol methyltransferase (γ-TMT), one of the key enzymes in the tocopherol biosynthetic pathway in plants, converts γ, δ-tocopherols into α-, β-tocopherols. Tocopherol content was investigated in 15 soybean cultivars and GmTMT2 was isolated from five varieties based on tocopherol content. GmTMT2a was expressed in E. coli and the purified protein effectively converted γ-tocopherol into α-tocopherol in vitro. Overexpression of GmTMT2a enhanced α-tocopherol content 4–6-fold in transgenic Arabidopsis, and α-tocopherol content increased 3–4.5-fold in transgenic maize seed, which correlated with the accumulation of GmTMT2a. Transgenic corn that is α-tocopherol-rich may be beneficial for animal health and growth.     

The intracellular distribution of tocopherols in Calendula officinalis leaves

From Calendula officinalis leaves, five cellular subtractions (chloroplasts, mitochondria, Golgi membranes, microsomes and cytosol) were obtained and their purity was checked. The contents of α-,γ- and δ-tocopherols were determined in these fractions. There were no tocopherols in Golgi membranes and cytosol. γ-Tocopherol and δ-tocopherol were found in the chloroplasts, mitochondria and microsomes, whereas α-tocopherol was present only in the chloroplasts.     

Dietary α- and γ-tocopherol supplementation attenuates lipopolysaccharide-induced oxidative stress and inflammatory-related responses in an obese mouse model of nonalcoholic steatohepatitis

Oxidative stress contributes towards the development of nonalcoholic steatohepatitis (NASH). Thus, antioxidants may decrease oxidative stress and ameliorate the events contributing to NASH. We hypothesized that α- or γ-tocopherol would protect against lipopolysaccharide (LPS)-triggered NASH in an obese (ob/ob) mouse model. Five-week-old obese mice (n=18/dietary treatment) were provided 15 mg/kg each of α- and γ-tocopherol or 500 mg/kg of α- or γ-tocopherol for 5-weeks. Then, all mice were injected ip once with LPS (250 μg/kg) before being sacrificed at 0, 1.5 or 6 h. Body weight and hepatic steatosis were unaffected by tocopherols and LPS. Hepatic α- and γ-tocopherol increased (P<.05) ~9.8- and 10-fold in respective tocopherol supplemented mice and decreased in response to LPS. LPS increased serum alanine aminotransferase (ALT) by 86% at 6 h and each tocopherol decreased this response by 29–31%. By 6 h, LPS increased hepatic malondialdehyde (MDA) and tumor necrosis factor-α by 81% and 44%, respectively, which were decreased by α- or γ-tocopherol. Serum ALT was correlated (P<.05) to hepatic tumor necrosis factor-α (r=0.585) and MDA (r=0.592), suggesting that inflammation and lipid peroxidation contributed to LPS-triggered hepatic injury. α- and γ-Tocopherol similarly attenuated LPS-triggered increases in serum free fatty acid, and α-tocopherol only maintained the LPS-triggered serum triacylglycerol responses at 6 h. These findings indicate that increasing hepatic α- or γ-tocopherol protected against LPS-induced NASH by decreasing liver damage, lipid peroxidation, and inflammation without affecting body mass or hepatic steatosis. Further study is needed to define the mechanisms by which these tocopherols protected against LPS-triggered NASH.     

Engineered drought-induced biosynthesis of α-tocopherol alleviates stress-induced leaf damage in tobacco

Tocopherols are members of the vitamin E complex and essential antioxidant compounds synthesized in chloroplasts that protect photosynthetic membranes against oxidative damage triggered by most environmental stresses. Tocopherol deficiency has been shown to affect germination, retard growth and change responses to abiotic stress, suggesting that tocopherols may be involved in a number of diverse physiological processes in plants. Instead of seeking constitutive synthesis of tocopherols to improve stress tolerance, we followed an inducible approach of enhancing α-tocopherol accumulation under dehydration conditions in tobacco. Two uncharacterized stress inducible promoters isolated from Arabidopsis and the VTE2.1 gene from Solanum chilense were used in this work. VTE2.1 encodes the enzyme homogentisate phytyltransferase (HPT), which catalyzes the prenylation step in tocopherol biosynthesis. Transgenic tobacco plants expressing ScVTE2.1 under the control of stress-inducible promoters showed increased levels of α-tocopherol when exposed to drought conditions. The accumulation of α-tocopherol correlated with higher water content and increased photosynthetic performance and less oxidative stress damage as evidenced by reduced lipid peroxidation and delayed leaf senescence. Our results indicate that stress-induced expression of VTE2.1 can be used to increase the vitamin E content and to diminish detrimental effects of environmental stress in plants. The stress-inducible promoters introduced in this work may prove valuable to future biotechnological approaches in improving abiotic stress resistance in plants.     

Enhancement of α-tocopherol content through transgenic and cell suspension culture systems in tobacco

The tocopherols are amphipathic antioxidant synthesized by photosynthetic organisms, which forms the essential component in the human diet. To increase the α-tocopherol content in tobacco, two approaches have been attempted in this study: (1) transgenic approach, by constitutive overexpression of the genes encoding Arabidopsis homogentisate phytyltransferase (HPT) and tocopherol cyclase (TC) through Agrobacterium-mediated genetic transformation; (2) non-transgenic approach, by supplementation of intermediates/precursors of vitamin E biosynthesis like tyrosine, p-hydroxyphenyl pyruvic acid, homogentisic acid (HGA) and phytol in different concentrations and combinations using cell suspension culture system. Molecular analyses by PCR, RT-PCR and Southern hybridization were carried out to confirm the HPT and TC expressing transgenic tobacco lines. The α-tocopherol content in transgenic plants expressing HPT and TC increase by 5.5 and 4.1, respectively, over the wild type. These results indicate that, HPT and TC activities are important in tobacco plants for enhancing the vitamin E content. In the second approach, the supplementation of precursor in cell suspension cultures, i.e., combination of 150 μM HGA + 100 μM phytol, showed the maximum enhancement of α-tocopherol, i.e., 36-fold. These findings clearly imply that enhancement of α-tocopherol levels in tobacco system is possible, if we could modulate the vitamin E metabolic pathway. This is a very useful finding for the large-scale production of natural Vitamin E. Among the two systems tested, cell suspension culture-based system is ideal over the transgenic technology due to its efficiency and no biosafety concerns.     

Disruption of mouse cytochrome p450 4f14 (Cyp4f14 gene) causes severe perturbations in vitamin E metabolism

Vitamin E is a family of naturally occurring and structurally related lipophilic antioxidants, one of which, α-tocopherol (α-TOH), selectively accumulates in vertebrate tissues. The ω-hydroxylase cytochrome P450-4F2 (CYP4F2) is the only human enzyme shown to metabolize vitamin E. Using cDNA cloning, cell culture expression, and activity assays, we identified Cyp4f14 as a functional murine ortholog of CYP4F2. We then investigated the effect of Cyp4f14 deletion on vitamin E metabolism and status in vivo. Cyp4f14-null mice exhibited substrate-specific reductions in liver microsomal vitamin E-ω-hydroxylase activity ranging from 93% (γ-TOH) to 48% (γ-tocotrienol). In vivo data obtained from metabolic cage studies showed whole-body reductions in metabolism of γ-TOH of 90% and of 68% for δ- and α-TOH. This metabolic deficit in Cyp4f14(-/-) mice was partially offset by increased fecal excretion of nonmetabolized tocopherols and of novel ω-1- and ω-2-hydroxytocopherols. 12'-OH-γ-TOH represented 41% of whole-body production of γ-TOH metabolites in Cyp4f14(-/-) mice fed a soybean oil diet. Despite these counterbalancing mechanisms, Cyp4f14-null mice fed this diet for 6 weeks hyper-accumulated γ-TOH (2-fold increase over wild-type littermates) in all tissues and appeared normal. We conclude that CYP4F14 is the major but not the only vitamin E-ω-hydroxylase in mice. Its disruption significantly impairs whole-body vitamin E metabolism and alters the widely conserved phenotype of preferential tissue deposition of α-TOH. This model animal and its derivatives will be valuable in determining the biological actions of specific tocopherols and tocotrienols in vivo.     

Novel vitamin E forms in leaves of Kalanchoe daigremontiana and Phaseolus coccineus

In the present study, we isolated novel tocochromanols from green leaves of Kalanchoe daigremontiana and primary leaves of etiolated seedlings of Phaseolus coccineus that were identified as β-, γ-, and δ-tocomonoenols with unsaturation at the terminal isoprene unit of the side chain. The content of γ-tocomonoenol in leaves of etiolated bean increased gradually with the age of seedlings, reaching 50% of the γ-tocopherol level in 40-day-old plants. The content of this compound in leaves was increased by short illumination of etiolated plants and by addition of homogentisic acid, a biosynthetic precursor of tocopherols. These data indicated that γ-tocomonoenol is synthesized de novo from homogentisic acid and tetrahydro-geranylgeraniol diphosphate, a phytol precursor. Based on these results, a biosynthetic pathway of tocomonoenols is proposed.     

Vitamin E decreases endogenous cholesterol synthesis and apo-AI-mediated cholesterol secretion in Caco-2 cells

Intestine is the gateway for newly absorbed tocopherols. This organ also plays a crucial role in cholesterol metabolism. Because tocopherols are known to impact cholesterol metabolism in the liver, we hypothesized that tocopherols could also modulate cholesterol metabolism in the intestine. This study aimed to verify this hypothesis and to unveil the mechanisms involved, using Caco-2 cells as a model of the human intestinal cell.Both α- and γ-tocopherol significantly (P<.05) decreased endogenous cholesterol synthesis and apo-AI-mediated cholesterol secretion in Caco-2 cells. Tocopherols down-regulated (P<.05) up to half of the genes involved in the cholesterol synthesis pathway, together with CYP27A1, which is involved in oxysterol production. The activity of this enzyme, as well as the levels of intracellular oxysterols, was significantly diminished by tocopherols. Finally, tocopherols significantly reduced ABCA1 mRNA levels in Caco-2 cells.We conclude that tocopherols impair the endogenous synthesis and apo-AI-mediated secretion of cholesterol in Caco-2 cells. This effect involves a down-regulation of genes involved in the cholesterol synthesis pathway, resulting in down-regulation of CYP27A1 which, in turn, diminishes oxysterol concentrations. The outcome is a decrease of LXR activity, resulting in down-regulation of ABCA1. These data reinforce the effect of α- and γ-tocopherol on cholesterol metabolism via gene expression regulation.     

Isocratic rapid liquid chromatographic method for simultaneous determination of carotenoids,retinol, and tocopherols in human serum

An improved isocratic and rapid HPLC method was developed for the measurement of carotenoids, retinol and tocopherols in human serum. Vitamins were extracted with hexane. Mobile phase consisted of a mixture acetonitrile:methylene chloride:methanol with 20 mM ammonium acetate. This method used a small bead size (3 μm) Spherisorb ODS2 column with titane frits. Diode array and fluorescence detectors were used respectively for the detection of carotenoids and retinol/tocopherols. Chromatographic separation was complete in 13 min for β-cryptoxanthin, cis–trans-lycopene, α-carotene, β-carotene, cis-β-carotene, retinol, δ-tocopherol, γ-tocopherol and α-tocopherol. Echinenone and tocol were employed as internal standards for diode array and fluorescence detection. Accuracy was validated using standard reference material (SRM) 968C. Intra-assay and inter-assay precision were respectively 0.2–7.3% and 3.6–12.6%. Sensitivity was verified using the ICH recommendations and the limit of detection (LOD) obtained was sufficient for routine clinical application.     

Accumulation of vitamin E in potato (Solanum tuberosum) tubers

Vitamin E (tocopherol) is a powerful antioxidant essential for human health and synthesized only by photosynthetic organisms. The effects of over-expression of tocopherol biosynthetic enzymes have been studied in leaves and seeds, but not in a non-photosynthetic, below-ground plant organ. Genetic and molecular approaches were used to determine if increased levels of tocopherols can be accumulated in potato (Solanum tuberosum L.) tubers through metabolic engineering. Two transgenes were constitutively over-expressed in potato: Arabidopsis thaliana p-hydroxyphenylpyruvate dioxygenase (At-HPPD) and A. thaliana homogentisate phytyltransferase (At-HPT). α-Tocopherol levels in the transgenic plants were determined by high-performance liquid chromatography. In potato tubers, over-expression of At-HPPD resulted in a maximum 266% increase in α-tocopherol, and over-expression of At-HPT yielded a 106% increase. However, tubers from transgenic plants still accumulated approximately 10- and 100-fold less α-tocopherol than leaves or seeds, respectively. The results indicate that physiological and regulatory constraints may be the most limiting factors for tocopherol accumulation in potato tubers. Studying regulation and induction of tocopherol biosynthesis should reveal approaches to more effectively engineer crops with enhanced tocopherol content.     

The two faces of α- and γ-tocopherols: an in vitro and ex vivo investigation into VLDL, LDL and HDL oxidation

BackgroundVitamin E and its derivatives, namely, the tocopherols, are known antioxidants, and numerous clinical trials have investigated their role in preventing cardiovascular disease; however, evidence to date remains inconclusive. Much of the in vitro research has focused on tocopherol's effects during low-density lipoprotein (LDL) oxidation, with little attention being paid to very LDL (VLDL) and high-density lipoprotein (HDL). Also, it is now becoming apparent that γ-tocopherol may potentially be more beneficial in relation to cardiovascular health.ObjectivesDo α- and γ-tocopherols become incorporated into VLDL, LDL and HDL and influence their oxidation potential in an in vitro and ex vivo situation?DesignFollowing (i) an in vitro investigation, where plasma was preincubated with increasing concentrations of either α- or γ-tocopherol and (ii) an in vivo 4-week placebo-controlled intervention with α- or γ-tocopherol. Tocopherol incorporation into VLDL, LDL and HDL was measured via high-pressure liquid chromatography, followed by an assessment of their oxidation potential by monitoring conjugated diene formation.ResultsIn vitro: Both tocopherols became incorporated into VLDL, LDL and HDL, which protected VLDL and LDL against oxidation. However and surprisingly, the incorporation into HDL demonstrated pro-oxidant properties. Ex vivo: Both tocopherols were incorporated into all three lipoproteins, protecting VLDL and LDL against oxidation; however, they enhanced the oxidation of HDL.ConclusionsThese results suggest that α- and γ-tocopherols display conflicting oxidant activities dependent on the lipoprotein being oxidized. Their pro-oxidant activity toward HDL may go some way to explain why supplementation studies with vitamin E have not been able to display cardioprotective effects.     

Antioxidant Effect of Tocopherols on Autoxidation of Milk Fat

Antioxidant activity of d-α-, dl-β-, d-γ- and d-δ-tocopherol was investigated with fatty acid methylester of milk fat from which unsaponifiable matter had been removed. Autoxidation was carried out at 50°C and its degree was indicated by peroxide value, α- or β-Tocopherol was more effective at lower concentrations (0.003 and 0.01%) than at higher concentrations (0.05, 0.1 and 0.5%). The antioxidant activity of γ- and δ-tocopherol was increased with the increase of tocopherol concentration within the range of 0.001 to 0.5%. The order of antioxidant activity of these tocopherols, which was compared in terms of the time to reach 30 meq of peroxide value, varied with the concentration; γ > β > δ > α at 0.001%, α > γ > β > δ at 0.003%, γ > δ > β > α at 0.01%, and δ > γ > β > α at the concentrations more than 0.05%. α-Tocopherol at the concentration of 0.003%, which corresponded to the concentration in original milk fat, was more effective than other tocopherols at the same concentration and α-tocopherol at other concentrations. Synergism due to the combination of β-, γ-, or δ-tocopherol with 0.003% of α-tocopherol was not observed.     

Enzymatic sulfation of tocopherols and tocopherol metabolites by human cytosolic sulfotransferases

Tocopherols are essential micronutrients for mammals widely known as potent lipid-soluble antioxidants that are present in cell membranes. Recent studies have demonstrated that most of the carboxychromanol (CEHC), a tocopherol metabolite, in the plasma exists primarily in sulfate- and glucuronide-conjugated forms. To gain insight into the enzymatic sulfation of tocopherols and their metabolites, a systematic investigation was performed using all 14 known human cytosolic sulfotransferases (SULTs). The results showed that the members of the SULT1 family displayed stronger sulfating activities toward tocopherols and their metabolites. These enzymes showed a substrate preference for γ-tocopherol over α-tocopherol and for γ-CEHC over other CEHCs. Using A549 human lung epithelial cells in a metabolic labeling study, a similar trend in the sulfation of tocopherols and CEHCs was observed. Collectively, the results obtained indicate that SULT-mediated enzymatic sulfation of tocopherols and their metabolites is a significant pathway for regulation of the homeostasis and physiological functions of these important compounds.     

Response of scab-susceptible (McIntosh) and scab-resistant (Liberty) apple tissues to treatment with yeast extract and Venturia inaequalis

Yeast extract and Venturia inaequalis treated intact scab-susceptible (McIntosh) and scab-resistant (Liberty) apple plants and their organs were analyzed for phenolic metabolites. The major phenolic compounds found in both non-treated and treated leaves were phloridzin and phloretin which accumulated in mM concentrations. Untreated and treated stems and roots contained only phloridzin as the major detectable metabolite during the course of the investigation. The accumulation of phloridzin and phloretin was not developmentally regulated, since they were present in both young and old leaves, and also in the intercellular washings of both scab-susceptible and scab-resistant plants. The major metabolites of both McIntosh and Liberty fruits were cinnamyl glucose and p-coumarylquinic acid, which increased 20-fold in Liberty fruit upon yeast extract treatment. The same compounds increased only 2-fold in McIntosh fruits. Minor compounds in the fruits of both cultivars were p-coumaric acid, phloridzin and phloretin, the latter compound being present at the threshold of detection. Biphenyl and dibenzofuran compounds, the major metabolites of elicitor treated Liberty cell suspension cultures, could not be detected in the intact plants. These results indicate differential response of plant organs and cell suspension cultures to elicitor treatment or pathogen invasion.