IL-4 inhibits osteoclast formation through a direct action on osteoclast precursors via peroxisome proliferator-activated receptor γ1

IL-4 is a pleiotropic immune cytokine secreted by activated TH2 cells that inhibits bone resorption both in vitro and in vivo. The cellular targets of IL-4 action as well as its intracellular mechanism of action remain to be determined. We show here that IL-4 inhibits receptor activator of NF-κB ligand-induced osteoclast differentiation through an action on osteoclast precursors that is independent of stromal cells. Interestingly, this inhibitory effect can be mimicked by both natural as well as synthetic peroxisome proliferator-activated receptor γ1 (PPARγ1) ligands and can be blocked by the irreversible PPARγ antagonist GW 9662. These findings suggest that the actions of IL-4 on osteoclast differentiation are mediated by PPARγ1, an interpretation strengthened by the observation that IL-4 can activate a PPARγ1-sensitive luciferase reporter gene in RAW264.7 cells. We also show that inhibitors of enzymes such as 12/15-lipoxygenase and the cyclooxygenases that produce known PPARγ1 ligands do not abrogate the IL-4 effect. These findings, together with the observation that bone marrow cells from 12/15-lipoxygenase-deficient mice retain sensitivity to IL-4, suggest that the cytokine may induce novel PPARγ1 ligands. Our results reveal that PPARγ1 plays an important role in the suppression of osteoclast formation by IL-4 and may explain the beneficial effects of the thiazolidinedione class of PPARγ1 ligands on bone loss in diabetic patients.

Inhibition of Wound-Induced Accumulation of Allene Oxide Synthase Transcripts in Flax Leaves by Aspirin and Salicylic Acid1

Allene oxide synthase (AOS) mediates the conversion of lipoxygenase-derived fatty acid hydroperoxides to unstable allene epoxides, which supply the precursors for the synthesis of the phytohormone jasmonic acid (JA). In this study the characterization of AOS gene expression in flax (Linum usitatissimum) is reported. AOS was constitutively expressed in different organs of flax plants. Additionally, AOS gene expression was enhanced after mechanical wounding in both the directly damaged leaves and in the systemic tissue located distal to the treated leaves. This wound-induced accumulation of AOS required the de novo biosynthesis of other unknown proteins involved in the signaling pathway modulating wound-induced AOS gene expression. Furthermore, the wound-induced AOS mRNA accumulation was correlated with the increase in the levels of JA. Both JA and its precursor, 12-oxo-phytodienoic acid, activated AOS gene expression in a dose-dependent manner. Thus, JA could activate its own biosynthetic pathway in flax leaves. Moreover, neither salicylic acid (SA) nor aspirin influenced AOS enzymatic activity. It is interesting that pretreatment with SA or aspirin inhibited wound-induced accumulation of AOS transcripts. These results suggest that a potent inhibition of JA biosynthetic capacity in leaves can be affected by SA or aspirin at the level of AOS gene expression.

Specific Soybean Lipoxygenases Localize to Discrete Subcellular Compartments and Their mRNAs Are Differentially Regulated by Source-Sink Status1

Members of the lipoxygenase multigene family, found widely in eukaryotes, have been proposed to function in nitrogen partitioning and storage in plants. Lipoxygenase gene responses to source-sink manipulations in mature soybean (Glycine max [L.] Merr.) leaves were examined using gene-specific riboprobes to the five vegetative lipoxygenases (vlxA–vlxE). Steady-state levels of all vlx mRNAs responded strongly to sink limitation, but specific transcripts exhibited differential patterns of response as well. During reproductive sink limitation, vlxA and vlxB messages accumulated to high levels, whereas vlxC and vlxD transcript levels were modest. Immunolocalization using peptide-specific antibodies demonstrated that under control conditions, VLXB was present in the cytosol of the paraveinal mesophyll and with pod removal accumulated additionally in the bundle-sheath and adjacent cells. With sink limitation VLXD accumulated to apparent high levels in the vacuoles of the same cells. Segregation of gene products at the cellular and subcellular levels may thus permit complex patterns of differential regulation within the same cell type. Specific lipoxygenase isoforms may have a role in short-term nitrogen storage (VLXC/D), whereas others may simultaneously function in assimilate partitioning as active enzymes (VLXA/B).

Relationship between CO2 Assimilation, Photosynthetic Electron Transport, and Active O2 Metabolism in Leaves of Maize in the Field during Periods of Low Temperature1

Measurements of the quantum efficiencies of photosynthetic electron transport through photosystem II (φPSII) and CO2 assimilation (φCO2) were made simultaneously on leaves of maize (Zea mays) crops in the United Kingdom during the early growing season, when chilling conditions were experienced. The activities of a range of enzymes involved with scavenging active O2 species and the levels of key antioxidants were also measured. When leaves were exposed to low temperatures during development, the ratio of φPSII/φCO2 was elevated, indicating the operation of an alternative sink to CO2 for photosynthetic reducing equivalents. The activities of ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, glutathione reductase, and superoxide dismutase and the levels of ascorbate and α-tocopherol were also elevated during chilling periods. This supports the hypothesis that the relative flux of photosynthetic reducing equivalents to O2 via the Mehler reaction is higher when leaves develop under chilling conditions. Lipoxygenase activity and lipid peroxidation were also increased during low temperatures, suggesting that lipoxygenase-mediated peroxidation of membrane lipids contributes to the oxidative damage occurring in chill-stressed leaves.

Arachidonate lipoxygenases as essential regulators of cell survival and apoptosis.

Arachidonic acid (AA) metabolites derived from both cyclooxygenase (COX) and lipoxygenase (LOX) pathways transduce a variety of signals related to cell growth. Here, we report that the AA LOX pathway also functions as a critical regulator of cell survival and apoptosis. Rat Walker 256 (W256) carcinosarcoma cells express 12-LOX and synthesize 12(S)- and 15(S)-hydroxyeicosatetraenoic acids as their major LOX metabolites. W256 cells transfected with 12-LOX-specific antisense oligonucleotide or antisense oligonucleotides directed to conserved regions of LOXs underwent time- and dose-dependent apoptosis. Likewise, treatment of W256 cells with various LOX but not COX inhibitors induced apoptotic cell death, which could be partially inhibited by exogenous 12(S)- or 15(S)-hydroxyeicosatetraenoic acids. The W256 cell apoptosis induced by antisense oligos and LOX inhibitors was followed by a rapid downregulation of bcl-2 protein, a dramatic decrease in the bcl-2/bax ratio, and could be suppressed by bcl-2 overexpression. In contrast, p53, which is wild type in W256 cells, did not undergo alterations during apoptosis induction. The results suggest that the LOX pathway plays an important physiological role in regulating apoptosis.

Aspirin triggers previously undescribed bioactive eicosanoids by human endothelial cell-leukocyte interactions.

Aspirin [acetylsalicylic acid (ASA)], along with its analgesic-antipyretic uses, is now also being considered for cardiovascular protection and treatments in cancer and human immunodeficiency virus infection. Although many of ASA's pharmacological actions are related to its ability to inhibit prostaglandin and thromboxane biosynthesis, some of its beneficial therapeutic effects are not completely understood. Here, ASA triggered transcellular biosynthesis of a previously unrecognized class of eicosanoids during coincubations of human umbilical vein endothelial cells (HUVEC) and neutrophils [polymorphonuclear leukocytes (PMN)]. These eicosanoids were generated with ASA but not by indomethacin, salicylate, or dexamethasone. Formation was enhanced by cytokines (interleukin 1 beta) that induced the appearance of prostaglandin G/H synthase 2 (PGHS-2) but not 15-lipoxygenase, which initiates their biosynthesis from arachidonic acid in HUVEC. Costimulation of HUVEC/PMN by either thrombin plus the chemotactic peptide fMet-Leu-Phe or phorbol 12-myristate 13-acetate or ionophore A23187 leads to the production of these eicosanoids from endogenous sources. Four of these eicosanoids were also produced when PMN were exposed to 15R-HETE [(15R)-15-hydroxy-5,8,11-cis-13-trans-eicosatetraenoic acid] and an agonist. Physical methods showed that the class consists of four tetraene-containing products from arachidonic acid that proved to be 15R-epimers of lipoxins. Two of these compounds (III and IV) were potent inhibitors of leukotriene B4-mediated PMN adhesion to HUVEC, with compound IV [(5S,6R,15R)-5,6,15-trihydroxy-7,9,13-trans-11-cis-eicosatetraenoi c acid; 15-epilipoxin A4] active in the nanomolar range. These results demonstrate that ASA evokes a unique class of eicosanoids formed by acetylated PGHS-2 and 5-lipoxygenase interactions, which may contribute to the therapeutic impact of this drug. Moreover, they provide an example of a drug's ability to pirate endogenous biosynthetic mechanisms to trigger new mediators.

Interleukin 4 suppresses c-kit ligand-induced expression of cytosolic phospholipase A2 and prostaglandin endoperoxide synthase 2 and their roles in separate pathways of eicosanoid synthesis in mouse bone marrow-derived mast cells.

Mouse bone marrow-derived mast cells (BMMCs) developed with interleukin 3 (IL-3) can be stimulated by c-kit ligand (KL) and accessory cytokines over a period of hours for direct delayed prostaglandin (PG) generation or over a period of days to prime for augmented IgE-dependent PG and leukotriene (LT) production, as previously reported. We now report that IL-4 is counterregulatory for each of these distinct KL-dependent responses. BMMCs cultured for 4 days with KL + IL-3 or with KL + IL-10 produced 5- to 7-fold more PGD2 and approximately 2-fold more LTC4 in response to IgE-dependent activation than BMMCs maintained in IL-3 alone. IL-4 inhibited the priming for increased IgE-dependent PGD2 and LTC4 production to the level obtained by activation of BMMCs maintained in IL-3 alone with an IC50 of approximately 0.2 ng/ml. IL-4 inhibited the KL-induced increase in expression of cytosolic phospholipase A2 (cPLA2) but had no effect on the incremental expression of PG endoperoxide synthase 1 (PGHS-1) and hematopoietic PGD2 synthase or on the continued baseline expression of 5-lipoxygenase, 5-lipoxygenase activating protein, and LTC4 synthase. BMMCs stimulated by KL + IL-10 for 10 h exhibited a delayed phase of PGD2 generation, which was dependent on de novo induction of PGHS-2. IL-4 inhibited the induction of PGHS-2 expression and the accompanying cytokine-initiated delayed PGD2 generation with an IC50 of approximately 6 ng/ml. IL-4 had no effect on the expression of PGHS-2 and the production of PGD2 elicited by addition of IL-1 beta to the combination of KL + IL-10. IL-4 had no effect on the immediate phase of eicosanoid synthesis elicited by KL alone or by IgE and antigen in BMMCs maintained in IL-3. Thus, the counterregulatory action of IL-4 on eicosanoid generation is highly selective for the induced incremental expression of cPLA2 and the de novo expression of PGHS-2, thereby attenuating time-dependent cytokine-regulated responses to stimulation via Fc epsilon receptor I and stimulation via c-kit, respectively.

Opioids Inhibit Lateral Amygdala Pyramidal Neurons by Enhancing A Dendritic Potassium Current

Pyramidal neurons in the lateral amygdala discharge trains of action potentials that show marked spike frequency adaptation, which is primarily mediated by activation of a slow calcium-activated potassium current. We show here that these neurons also express an alpha-dendrotoxin- and tityustoxin-Kalpha-sensitive voltage-dependent potassium current that plays a key role in the control of spike discharge frequency. This current is selectively targeted to the primary apical dendrite of these neurons. Activation of mu-opioid receptors by application of morphine or D-Ala(2)-N-Me-Phe(4)-Glycol(5)-enkephalin (DAMGO) potentiates spike frequency adaptation by enhancing the alpha-dendrotoxin-sensitive potassium current. The effects of mu-opioid agonists on spike frequency adaptation were blocked by inhibiting G-proteins with N-ethylmaleimide (NEM) and by blocking phospholipase A(2). Application of arachidonic acid mimicked the actions of DAMGO or morphine. These results show that mu-opioid receptor activation enhances spike frequency adaptation in lateral amygdala neurons by modulating a voltage-dependent potassium channel containing Kv1.2 subunits, through activation of the phospholipase A(2)-arachidonic acid-lipoxygenases cascade.

Transcriptional response of sugarcane roots to methyl jasmonate

The effect of methyl jasmonate treatment on gene expression in sugarcane roots signalling between roots and shoots was studied. A collection of 829 ESTs were obtained from sugarcane roots treated with the defence-regulator methyl jasmonate (MJ) treatment. A subset of 747 of these were combined with 4793 sugarcane ESTs obtained from stem tissues in a cDNA microarray and experiments undertaken to identify genes that were induced in roots 24-120 h following treatment with MJ. Two data analysis systems (t-statistic and tRMA) were used to analyse the microarray results and these methods identified a common set of 21 ESTs corresponding to transcripts significantly induced by MJ in roots and 23 that were reduced in expression following MJ treatment. The induction of six transcripts identified in the microarray analysis was tested and confirmed using northern blotting. Homologues of genes encoding lipoxygenase and PR-10 proteins were induced 824 It after MJ treatment while the other four selected transcripts were induced at later time points. Following treatment of roots with MJ, the lipoxygenase homologue, but not the PR-10 homologue, was induced in untreated stem and leaf tissues. The PR-10 homologue and a PR-1 homologue, but not the lipoxygenase homologue, were induced in untreated tissues after the application of SA to roots. Repeated foliar application of MJ had no apparent effects on plant growth and was demonstrated to increase lipoxygenase transcripts in roots, but did not increase transcript levels-of other genes tested. These results lay a foundation for further studies of induced pest and disease resistance in sugarcane roots. (C) 2004 Elsevier Ireland Ltd. All rights reserved.

Inhibition of COXs and 5-LOX and activation of PPARs by Australian Clematis species (Ranunculaceae)

The species of Clematis (Ranunculaceae) have been traditionally used for inflammatory conditions by indigenous Australians. We have previously reported that the ethanol extract of Clematis pickeringii inhibited COX-1. In this study, we examined the ethanol extracts and fractions of three Clematis species, Clematis pickeringii, Clematis glycinoides and Clematis microphylla, on cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX). We further examined the activating effects on the protein expression of peroxisome proliferator-activated receptor alpha (PPAR alpha) and gamma (PPAR-gamma) in HepG2 cells. The ethanol extracts of three Clematis species inhibited the activities of COX-1, COX-2 and 5-LOX in the different extents. The stem extract of Clematis pickeringii showed the highest inhibitory activities among the three species on COX-1, COX-2 and 5-LOX with the IC50 values of 73.5, 101.2 and 29.3 mu g/mL. One of its fractions also significantly elevated PPAR gamma expression by 173, 280 and 435% and PPAR gamma expression by 140, 228 and 296% at 4, 8 and 16 mu g/mL, respectively. (c) 2005 Elsevier Ireland Ltd. All rights reserved.

Effects of arachidonic acid upon the volume-sensitive chloride current in rat osteoblast-like (ROS 17/2.8) cells.

1. The effects of arachidonic acid upon the volume-sensitive Cl- current present in cultured osteoblastic cells (ROS 17/2.8) was studied using the whole-cell patch-clamp technique. 2. Arachidonate produced two distinct phases of inhibition, a rapid phase occurring within 10-15 s of application preceding a slower phase that occurred 2 min after onset of arachidonate superfusion. Accompanying the slower inhibitory phase was an acceleration of the time-dependent inactivation exhibited by the current at strongly depolarized potentials (> + 50 mV). The half-maximal inhibitory concentrations (IC50) were 177 +/- 31 and 10 +/- 4 microM for the two phases respectively. 3. Arachidonate was still effective in the presence of inhibitors of cyclo-oxygenase (indomethacin, 10 microM), lipoxygenase (nordihydroguaretic acid, 10-100 microM) and cytochrome P450 (SKF525A, 100 microM; ethoxyresorufin, 10 microM; metyrapone, 500 microM; piperonyl butoxide, 500 microM; cimetidine, 1 mM). The effects of arachidonate could not be produced by another cis unsaturated fatty acid, oleic acid. 4. Measurements of cell volume showed that arachidonate effectively inhibited the regulatory volume decrease elicited by ROS 17/2.8 cells in response to a reduction in extracellular osmolarity. 5. It is concluded that the volume-sensitive Cl- conductance in ROS 17/2.8 cells is directly modulated by arachidonate and may represent a physiological mechanism by which volume regulation can be controlled in these cells.

Downregulation of ubiquitin-dependent proteolysis by eicosapentaenoic acid in acute starvation

A number of acute wasting conditions are associated with an upregulation of the ubiquitin-proteasome system in skeletal muscle. Eicosapentaenoic acid (EPA) is effective in attenuating the increased protein catabolism in muscle in cancer cachexia, possibly due to inhibition of 15-hydroxyeicosatetraenoic acid (15-HETE) formation. To determine if a similar pathway is involved in other catabolic conditions, the effect of EPA on muscle protein degradation and activation of the ubiquitin-proteasome pathway has been determined during acute fasting in mice. When compared with a vehicle control group (olive oil) there was a significant decrease in proteolysis of the soleus muscles of mice treated with EPA after starvation for 24 h, together with an attenuation of the proteasome 'chymotryptic-like' enzyme activity and the induction of the expression of the 20S proteasome α-subunits, the 19S regulator and p42, an ATPase subunit of the 19S regulator in gastrocnemius muscle, and the ubiquitin-conjugating enzyme E214k. The effect was not shown with the related (n-3) fatty acid docosahexaenoic acid (DHA) or with linoleic acid. However, 2,3,5trimethyl-6-(3-pyridylmethyl)1,4-benzoquinone (CV-6504), an inhibitor of 5-, 12- and 15-lipoxygenases also attenuated muscle protein catabolism, proteasome 'chymotryptic-like' enzyme activity and expression of proteasome 20S α-subunits in soleus muscles from acute fasted mice. These results suggest that protein catabolism in starvation and cancer cachexia is mediated through a common pathway, which is inhibited by EPA and is likely to involve a lipoxygenase metabolite as a signal transducer. © 2001 Academic Press.

Polyunsaturated fatty acid requirements of murine colon adenocarcinomas

The polyunsaturated fatty acid (PUFA) requirements of three transplantable murine colon adenocarcinomas, the MAC13, MAC16 and MAC26, were evaluated in vitro and in vivo. When serum concentrations became growth limiting in vitro, proliferation of the MAC13 and MAC26 cell lines was stimulated by linoleic acid (LA) at 18μM and arachidonic acid (AA) at 16 or 33μM respectively. This was not demonstrated by the MAC16 cell line. MAC13 and MAC26 cells were found to be biochemically fatty acid deficient as measured by the formation of Mead acid (20:3 n-9), but the MAC16 cells were not. In vivo the growth of the MAC26 tumour was stimulated by daily oral administration of LA between 0.4-2.0g/kg. There was a threshold value of 0.4g/kg for the stimulation of MAC26 tumour growth, above which there was no further increase in tumour growth, and below which no increase in tumour growth was observed. This increased tumour growth was due to the stimulation of tumour cell proliferation in all areas of the tumour, with no effect on the cell loss factor. The growth of the MAC13, MAC16, and MAC26 cell lines in vitro were more effectively inhibited by lipoxygenase (LO) inhibitors than the cyclooxygenase inhibitor indomethacin. The specific 5-LO inhibitor Zileuton and the leukotriene D4 antagonist L-660,711 were less effective inhibitors of MAC cell growth in vitro than the less specific LO inhibitors BWA4C, BWB70C and CV6504. Studies of the hyroxyeicosatetraenoic acids (HETEs) produced from exogenous AA in these cells, suggested that a balance of eicosanoids produced from 5-LO, 12-LO and 15-LO pathways was required for cell proliferation. In vivo BWA4C, BWB70C and CV6504 demonstrated antitumour action against the MAC26 tumour between 20-50mg/kg/day. CV6504 also inhibited the growth of the MAC 13 tumour in vivo with an optimal effect between 5-10mg/kg/day. The antitumour action against the MAC16 tumour was also accompanied by a reduction in the tumour-induced host body weight loss at 10-25mg/kg/day. The antitumour action of CV6504 in all three tumour models was partially reversed by daily oral administration of 1.0g/kg LA. Studies of the AA metabolism in tumour homogenates suggested that this profound antitumour action, against what are generally chemoresistant tumours, was due to inhibition of eicosanoid production through LO pathways. As a result of these studies, CV6504 has been proposed for stage I./II. clinical trials against pancreatic cancer by the Cancer Research Campaign. This will be the first LO inhibitor entering the clinic as a therapeutic agent.

Muscle catabolism in cancer and its attenuation by eicosapentaenic acid

This work examines skeletal muscle catabolism in cancer and its attenuation by Eicosapentaenoic Acid (EPA). In vivo studies in mice bearing a cachexia inducing murine colon adenocarcinoma - MAC16, demonstrated an elevation in the gastrocnemius muscle in the activity and expression of regulatory components of the ubiquitin-proteasome proteolytic pathway. This was accompanied by an accelerated loss of muscle tissue correlating with an increase in overall weight loss, all of which were attenuated by prior daily dosing with EPA. Recently a proteolysis inducing factor (PIF) has been isolated from the MAC16 tumour, and from the serum and urine of cachectic cancer patients. Previous studies have shown that PIF induces protein degradation in vitro, and that this is possibly mediated through 15-hydroxyeicosatetraenoic acid (15-HETE), a metabolite of the n-6 polyunsaturated fatty acid- arachidonate. Employing the murine myoblast cell line C2C12, it was shown that both PIF and 15-HETE increased protein degradation and expression of proteasome subunits, processes which were again attenuated by prior incubation in EPA. Similarly, in NMRI mice which had been fasted for 24hours, EPA and the lipoxygenase inhibitor CV-6504 (but not structurally related fatty acids) inhibited skeletal muscle proteolysis and expression of various proteasome subunits, showing that firstly, EPA may be anti-cachexic partly through its ability to influence 15-HETE production; and secondly that the effect is specific for EPA as other fatty acids had no effect. Previous studies have suggested the involvement of the signal transduction family NFKB in response to PIF in the liver. It has been demonstrated here that both PIF and 15-HETE increased nuclear translocation of NFKB in the skeletal muscle of tumour bearing mice and that EPA inhibited this process by its ability to prevent the degradation of the NFKB inhibitor protein IKB. When an NFKB inhibitor was added to C2C12 myotubes, prior to the addition of PIF, proteasome activity and protein degradation was inhibited, showing that NFKB is responsible for the increased proteasome activity and muscle catabolism induced by PIF. Taken together this work suggests that 15-hydroxyeicosatetraenoic acid is the intracellular mediator for PIF induced protein degradation in skeletal muscle and that elevated muscle catabolism is accomplished through an increased functioning of the ubiquitin-proteasome pathway, a process possibly mediated through an NFKB dependent mechanism. The anticachectic (and possibly the anti-tumourigenic) effects of EPA appear to be achieved in part by its ability to inhibit the degradation of IKB and possibly by its ability to interfere with 15-HETE production.

Synthesis and biological evaluation of potential anti-cancer agents

The new technology of combinational chemistry has been introduced to pharmaceutical companies, improving and making more efficient the process of drug discovery. Automated combinatorial chemistry in the solution-phase has been used to prepare a large number of compounds of anti-cancer screening. A library of caffeic acid derivatives has been prepared by the Knoevenagel condensation of aldehyde and active methylene reagents. These products have been screened against two murine adenocarcinoma cell lines (MAC) which are generally refractive to standard cytotoxic agents. The target of anti-proliferative action was the 12- and 15-lipoxygenase enzymes upon which these tumour cell lines have been shown to be dependent for proliferation and metastasis. Compounds were compared to a standard lipoxygenase inhibitor and if found to be active anti-proliferative agents were tested for their general cytotoxicity and lipoxygenase inhibition. A solid-phase bound catalyst, piperazinomethyl polystyrene, was devised and prepared for the improved generation of Knoevenagel condensation products. This piperazinomethyl polystyrene was compared to the traditional liquid catalyst, piperidine, and was found to reduce the amount of by-products formed during reaction and had the advantage of easy removal from the reaction. 13C NMR has been used to determine the E/Z stereochemistry of Knoevenagel condensation products. Soluble polymers have been prepared containing different building blocks pendant to the polymer backbone. Aldehyde building blocks incorporated into the polymer structure have been subjected to the Knoevenagel condensation. Cleavage of the resultant pendant molecules has proved that soluble linear polymers have the potential to generate combinatorial mixtures of known composition for biological testing. Novel catechol derivatives have been prepared by traditional solution-phase chemistry with the intention of transferring their synthesis to a solid-phase support. Catechol derivatives prepared were found to be active inhibitors of lipoxygenase. Soluble linear supports for the preparation of these active compounds were designed and tested. The aim was to develop a support suitable for the automated synthesis of libraries of catechol derivatives for biological screening.