Sites of auxin action: regulation of geotropism, growth, and ethylene production by inhibitors of auxin transport

The inhibitors of auxin transport-NPA (N-1-naphthylphthalamic acid), DPX1840 (3,3a-dihydro-2-(p-methoxyphenyl)-8H-pyrazolo[5,1-a] isoindol-8-one), and TIBA (2,3,5-triiodobenzoic acid)-inhibited geotropism in roots of intact Pisum sativum L. seedlings. NPA and DPX1840 also caused cellular swelling in the roots. The swelling was due to a greater inhibition of elongation than increase in weight and looked identical to the one caused by ethylene. However, ethylene did not act as an intermediate in the action of auxin transport inhibitors because all three failed to stimulate ethylene production and some of their growth-inhibiting effect was retained in the presence of saturating levels of ethylene. In the presence of 10 mum indoleacetic acid the growth-inhibiting effect of auxin transport inhibitors was lost after 18 hours. On the other hand, auxin transport inhibitors did not interfere with the ability of auxin to promote ethylene production. Growth inhibition caused by auxin transport inhibitors was reversible. Pea root sections resumed normal growth following flushing of treated sections with inhibitor-free solutions. Experiments with (14)C-2, 4-dichlorophenoxyacetic acid revealed that the herbicide and auxin transport inhibitors may have the same binding site. It was concluded that a class of structurally dissimilar compounds may share a similar physiological role since they all appear to compete with endogenous auxin for certain binding sites and they all have similar growth-regulating activities.     

The Control of Lateral Root Development in Cultured Pea Seedlings. II. Root Fasciation Induced by Auxin Inhibitors

Lateral root development in cultured seedlings of Pisum sativum (cv. Alaska) was modified by the application of auxin transport inhibitors or antagonists. When applied either to replace the root tip or beneath the cotyledonary node, two auxin transport inhibitors, 2,3,5-triiodobenzoic acid (TIBA) and 3,3a-dihydro-2-(p-methoxyphenyl)-8H-pyrazolo[5,1-α]isoindol-8-one (DPX-1840), increased cell division activity opposite the protoxylem poles. This resulted in the formation of masses of cells, which we are calling root primordial masses (RPMs), 2 to 3 days after treatment. RPMs differed from lateral root primordia in that they lacked apical organization. Some roots however developed both RPMs and lateral roots indicating that both structures were similar in terms of the timing and location of cell division in the pericycle and endodermis leading to their initiation. Removal of the auxin transport inhibitors allowed many of the RPMs to organize later into lateral root primordia and to emerge in clusters. When the auxin, indoleacetic acid (IAA) was added to the growth medium along with DPX-1840, 3 ranks of RPMs now in the form of fasciated lateral roots emerged from the primary root. The auxin antagonist, p-chlorophenoxy-isobutyric acid (PCIB), also induced RPM formation. In contrast to DPX-1840 treatment, the addition of IAA during PCIB treatment caused normal lateral root development.     

Auxin transport: a new synthetic inhibitor

The new synthetic plant growth regulator DPX1840 (3,3a-dihydro-2-(p-methoxyphenyl)-8H-pyrazolo [5,1-a] isoindol-8-one) was examined for its effects on auxin transport. At a concentration of 0.5 mm in the receiver agar cylinders DPX1840 significantly inhibited the basipetal transport of naphthaleneacetic acid-1-¹⁴C in stem sections of Vigna sinensis Endl., Pisum sativum L., Phaseolus vulgaris L., Glycine max L., Helianthus annuus L., Gossypium hirsutum L., and Zea mays L. without significantly reducing total auxin uptake or recovery. The time sequence of the effect varied with the plant species. A similar inhibition of the basipetal movement of indoleacetic acid-1-¹⁴C was observed in intact seedlings of Phaseolus vulgaris L. In contrast to basipetal auxin transport DPX1840 had no significant effect on the acropetal movement of indoleacetic acid-1-¹⁴C in stem sections of Gossypium hirsutum L. Qualitatively the effect of DPX1840 on basipetal auxin transport was similar to that of other known auxin transport inhibitors. Quantitative differences, however, suggested the following order of activity: Naptalam>morphactin[unk]DPX1840>2,3,5-triiodobenzoic acid.     

Failure of Ethylene to Change the Distribution of Indoleacetic Acid in the Petiole of Coleus blumei X frederici during Epinasty

The effect of ethylene on the distribution of applied indoleacetic acid in the petiole of Coleus blumei Benth. X C. frederici G. Taylor has been investigated during the development of epinastic curvature. Using intact plants, ¹⁴C-IAA was applied to the distal region of the leaf lamina and the accumulation of label in the abaxial and adaxial halves of 5 mm petiole sections was determined after 1.5, 3, and 6 hours. Over this period the label was transported out of the lamina into the petiole at a rate of at least 66 mm hr⁻¹. Of the total amount of label in the petiole sections, 24 to 30% was located in the adaxial half and this distribution was not altered significantly by exposing plants to an atmosphere containing 50 μl/l ethylene. Thus when epinastic curvature is induced by ethylene there is no associated increase in the IAA content of the expanding adaxial half. The role of endogenous IAA in petiole epinasty was studied by restricting its movement with DPX 1840 (3,3a-dihydro-2-[p-methoxyphenyl]-8H-pyrozolo{5,1-a}isoindol-8-one). The leaf petioles still showed an initial epinastic response to ethylene. It is concluded that ethylene-induced epinasty is not dependent upon either any change in the transport of IAA or its redistribution within the petiole.     

DPX 1840-induced Organogenesis in Xanthi-nc Tobacco Plants

The synthetic growth regulant DPX 1840 (3,3a-dihydro-2-(p-methoxyphenyl)-8H-pyrazolo[5, 1-a]isoindol-8-one) induced callus growth and subsequent tissue differentiation on cut surfaces of decapitated Xanthi-nc tobacco plants (Nicotiana tabacum). Callus formation and organogenesis induced by DPX 1840 depended on the presence of leaves. The adventitious meristems developed into either vegetative or flowering shoots. Pedicels that bore single flower buds developed two abscission zones that caused the buds to abscise before anthesis. The various morphological and physiological processes affected by DPX 1840 suggests that this growth regulator affects the endogenous hormonal distribution and/or activity.     

Structure-activity relationships of compounds targeting mycobacterium tuberculosis 1-deoxy-D-xylulose 5-phosphate synthase

We report on a target-based approach to identify possible Mycobacterium tuberculosis DXS inhibitors from the structure of a known transketolase inhibitor. A small focused library of analogs was assembled in order to begin elucidating some meaningful structure-activity relationships of 3-(4-chloro-phenyl)-5-benzyl-4H-pyrazolo[1,5-a]pyrimidin-7-one. Ultimately we found that 2-methyl-3-(4-fluorophenyl)-5-(4-methoxy-phenyl)-4H-pyrazolo[1,5-a]pyrimidin-7-one, although still weak, was able to inhibit M. tuberculosis DXS with an IC(50) of 10.6 microM.     

Synthesis and selective cyclooxygenase-2 (COX-2) inhibitory activity of a series of novel bicyclic pyrazoles

Novel series of pyrazolo[5,1-b]1,3-oxazolidines, pyrazolo[5,1-b]1,3-oxazines and imidazolidino[1,2-d]pyrazoles were synthesized. These compounds were evaluated in vitro for their ability to inhibit cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) in human whole blood (HWB). Several of the compounds were found to be novel and selective COX-2 inhibitors, the most potent and selective being 1-(5-cyclohexyl (2H,3H-pyrazolo[5,1-b]-1,3-oxazolidin-6-yl)-4-(methylsulfonyl)benzene, 7a (IC(5o) for COX-1>100 microM; for COX-2=1.3 microM).     

Design, synthesis and preliminary pharmacological evaluation of new analogues of DM232 (unifiram) and DM235 (sunifiram) as cognition modulators

A series of amides, structurally related to DM232 (unifiram) and DM235 (sunifiram), characterized by a 1,2,3,4-tetrahydropyrazino[2,1-a]isoindol-6(2H)-one, 1,4-diamino-cyclohexane or 1,4-diaminobenzene ring, have been synthesized and tested for cognition-enhancing activity in the mouse passive-avoidance test. Some of the compounds display good antiamnesic and procognitive activity, with higher potency than piracetam, while some cyclohexane derivatives are endowed with amnesia inducing properties.     

Microwave-assisted synthesis of antimicrobial dihydropyridines and tetrahydropyrimidin-2-ones: novel compounds against aspergillosis

Ten 4-aryl-1,4-dihydropyridine and three 4-aryl-1,2,3,4-tetrahydropyrimidin-2-one derivatives have been synthesized and examined for their activity against pathogenic strains of Aspergillus fumigatus and Candida albicans. Although none of the three compounds belonging to pyrimidin-2-one series showed any activity against two pathogens, two of the compounds of the dihydropyridine series, that is, diethyl 4-(4-methoxyphenyl)-2,6-dimethyl-1,4-dihydropyridin-3,5-dicarboxylate and dimethyl 4-(4-methoxyphenyl)-2,6-dimethyl-1,4-dihydropyridin-3,5-dicarboxylate, exhibited significant activity against A. fumigatus in disc diffusion, microbroth dilution and percent spore germination inhibition assays. The most active diethyl dihydropyridine derivative exhibited a MIC value of 2.92 microg/disc in disc diffusion and 15.62 microg/ml in microbroth dilution assays. The MIC(90) value of the most active compound by percent germination inhibition assay was found to be 15.62 microg/ml. The diethyl dicarboxylate derivative of dihydropyridine also exhibited appreciable activity against C. albicans. The in vitro toxicity of the most active diethyl dihydropyridine derivative was evaluated using haemolytic assay, in which the compound was found to be non-toxic to human erythrocytes even at a concentration of 625 microg/ml. The standard drug amphotericin B exhibited 100% lysis of erythrocytes at a concentration almost 16 times less than the safer concentration of the most active dihydropyridine derivative.     

Diarylheptanoids from the rhizomes of Zingiber officinale

Seven new diarylheptanoids, i.e., (3S,5S)-3,5-diacetoxy-1,7-bis(4-hydroxy-3-methoxyphenyl)heptane, (3R,5S)-3-acetoxy-5-hydroxy-1,7-bis(4-hydroxy-3-methoxyphenyl)heptane, (3R,5S)-3,5-dihydroxy-1-(4-hydroxy-3,5-dimethoxyphenyl)-7-(4-hydroxy-3-methoxyphenyl)heptane, (5S)-5-acetoxy-1,7-bis(4-hydroxy-3-methoxyphenyl)heptan-3-one, 5-hydroxy-1-(3,4-dihydroxy-5-methoxyphenyl)-7-(4-hydroxy-3-methoxyphenyl)heptan-3-one, 5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-7-(3,4-dihydroxy-5-methoxy-phenyl)heptan-3-one and 1,5-epoxy-3-hydroxy-1-(4-hydroxy-3,5-dimethoxyphenyl)-7-(4-hydroxy-3-methoxyphenyl)heptane were isolated from the rhizomes of Chinese ginger (Zingiber officinale Roscoe), along with 25 known compounds, i.e., 8 diarylheptanoids, 14 gingerol analogs, a diterpene and 2 steroids. Their structures were elucidated by spectroscopic and chemical methods.     

Synthesis and anti-inflammatory activities of N4,N5-disubstituted-3-methyl- H-pyrazolo[3,4-c]pyridazines

The synthesis and anti-inflammatory activity of 4,5-dihydroxy-3-methyl-1H-pyrazolo[3,4-c]pyridazine (4), 4,5-dichloro-3-methyl-1H-pyrazolo[3,4-c]pyridazine (5), 4,-benzoyloxy-3-methyl-1-benzoyl-1H-pyrazolo[3,4-c]pyridazin-5yl benzoate (6), 3-methyl-N4,N5-bis(4-methylphenyl)-1H-pyrazolo[3,4-c]pyridazine-4,5-diamine (7), 4[[5-(4-carboxyanilino)-3-methyl-1H-pyrazolo[3,4-c]pyridazin-4yl]amino]benzoic acid (8), N-[5-(benzoylamino)-3-methyl-1H-pyrazolo[3,4-c]pyridazin-4-yl]benzamide (9) and 3-methyl-N4,N5-bis[4-(1H-benzimidazol-2yl)phenyl]-1H-pyrazolo[3,4-c]pyridazine-4,5-diamine (10) are being reported.     

Auxin activity of substituted benzoic acids and their effect on polar auxin transport

Six dichloro-, 3 trichloro-, 2 triiodo-, and 3 heterosubstituted benzoic acids (amiben, dinoben, dicamba), and N-1-naphthylphthalamic acid have been tested for effects on growth and on polar auxin transport. Growth activity with and without kinetin was measured by effects on fresh and dry weights of 30-day cultures of fresh tobacco pith. Transport inhibition was measured by following uptake and output of IAA-2-¹⁴C through 10 mm bean epicotyl sections. The distribution of callus growth on vascularized tobacco stem segments was also observed. Avena first internode extension assays established the relative activities: dicamba > amiben > dinoben suggested by pith growth results. Growth effects of active compounds were similar with and without kinetin, except that amiben was less active with kinetin, while 2,3,6-trichlorobenzoic acid was more active with kinetin than alone. The weak auxin activity of NPA was confirmed. Transport experiments showed that NPA was the most inhibitory compound tested, followed by TIBA. Other compounds tested were at least 300 times less inhibitory to IAA transport. The best growth promoters were the least inhibitory to transport, and the most effective transport inhibitors were at best poor auxins. It is suggested that the weak auxin and auxin synergistic activity of TIBA (and perhaps 2,3-dichlorobenzoic acid) in extension growth tests arises from its inhibition of transport of endogenous or added auxin out of the sections, rather than from its intrinsic auxin activity. Chemically induced apolar callus growth on vascularized tobacco stem explants can arise from inhibition of native auxin transport, apolar growth stimulation by auxinic action of the test compound, or both.     

生姜中6个姜辣素类成分的抗菌活性研究

为了探究生姜化学成分的抗菌活性及初步构效关系,采用色谱法从生姜中分离得到6个姜辣素类化合物,采用波谱法对这6个成分进行鉴定,分别为5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-4-decen-3-one(1)、5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-4-dodecen-3-one(2)、5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-4-tetradecane-3-one(3)、[6]-姜酚(4)、[8]-姜酚(5)和[10]-姜酚(6)。采用抗菌纸片扩散法测定6个化合物对15株病原菌株的抗菌活性。结果表明化合物1和4抗菌活性最好,而6对所有菌株均无活性。初步构效关系分析表明:烯醇型化合物对革兰氏阳性菌的抗菌活性优于姜酚型化合物;而姜酚型化合物对革兰氏阴性菌的抗菌活性优于烯醇型化合物。此外,姜辣素类成分脂肪链的长度增加,可能导致抗菌活性降低。     

Development of imidazopyrazinone red-chemiluminescent probes for detecting superoxide anions via a chemiluminescence resonance energy transfer method.

During the development of useful probes for detecting superoxide anions via chemiluminescence with longer wavelengths than that of green, chemiluminescent probes that emit red light (lambda(max) 610 nm) when induced by superoxide anions were synthesized and characterized. These red-chemiluminescent probes consist of a 6-(4-methoxyphenyl)imidazo[1,2-a] pyrazin-3(7H)-one moiety, which reacts with superoxide anions to generate energy, and a sulphorhodamine 101 moiety, which accepts the energy and emits red light. Using a hypoxanthine-xanthine oxidase system for the generation of superoxide anions, it was shown that the superoxide anion-induced chemiluminescences of red-chemiluminescent probes (3 and 4) were more intense than those of the blue- and green-chemiluminescent probes 2-methyl-6-(4-methoxyphenyl)imidazo[1,2-a] pyrazin-3(7H)-one (MCLA) and 6-[4-[2-[N'-(5-fluoresceinyl)thioureido]-ethoxy]phenyl]-2-methylimidazo[1,2-a]pyrazin-3(7H)-one (FCLA), respectively, which are generally considered to be the most sensitive chemiluminescent probes. The ratio between the superoxide-dependent and background chemiluminescence intensities for 3 was comparable to those of MCLA and FCLA, but higher than that of 4. Due to its highly intense superoxide anion-induced chemiluminescence at low probe concentrations, red-chemiluminescent probe 3 is superior to MCLA and FCLA for measurement of superoxide anions.     

Ligand Specificity of Bean Leaf Soluble Auxin-binding Protein

The soluble bean leaf auxin-binding protein (ABP) has a high affinity for a range of auxins including indole-3-acetic acid (IAA), α-napthaleneacetic acid, phenylacetic acid, 2,4,5-trichlorophenoxyacetic acid, and structurally related auxins. A large number of nonauxin compounds that are nevertheless structurally related to auxins do not displace IAA from bean ABP. Bean ABP has a high affinity for auxin transport inhibitors and antiauxins. The specificity of pea ABP for representative auxins is similar to that found for bean ABP. The bean ABP auxin binding site is similar to the corn endoplasmic reticulum auxin-binding sites in specificity for auxins and sensitivity to thiol reagents and azide. Qualitative similarities between the ligand specificity of bean ABP and the specificity of auxin-induced bean leaf hyponasty provide further evidence, albeit circumstantial, that ABP (ribulose 1,5-bisphosphate carboxylase) can bind auxins in vivo. The high incidence of ABP in bean leaves and the high affinity of this protein for auxins and auxin transport inhibitors suggest possible functions for ABP in auxin transport and/or auxin sequestration.     

Syntheses of (±)-Hinokiresinol Dimethyl Ether and Related Compounds

Dimethyl ethers of (±)-hinokiresinol, dihydro- and tetrahydro-hinokiresinol have been synthesized. The key compounds, 1,3-bis-(p-methoxyphenyl)-pentan-1-one (4) and 1,3-bis-(p-methoxyphenyl)-4-penten-1-one (8) were obtained by treatment of 4,4′-dimethoxybenzal-acetophenone (3) with ethyl magnesium iodide and vinyl magnesium chloride, respectively. On Clemmensen reduction of the compound (4), tetrahydrohinokiresinol dimethyl ether was obtained. On reduction followed by dehydration of the compounds (4) and (8), dimethyl ethers of dihydrohinokiresinol and hinokiresinol were obtained, respectively.     

Phytochemical investigation of Millettia dorwardi Coll. et Hemsl

The first phytochemical investigation on the vine stems of Millettia dorwardi Coll. et Hemsl led to the isolation of ten flavonoids (isoafrormosin 1, formononetin 2, afrormosin 3, padmakastein 4, liquiritigenin 5, 4H-1-Benzopyran-4-one,7-hydroxy-5,8-dimethoxy-3-(4-methoxyphenyl)-isoflavone 6, 4H-1-Benzopyran-4-one,7-hydroxy-3-(3-hydroxy-5-methoxyphenyl)-6-methoxy 8, 4H-1-Benzopyran-4-one,6-methoxy-3-(4-methoxyphenyl)-6,4′-dimethoxyisoflavone 9, irisolidone 10, prunetin 11), one heterocycle (5-5′-dibuthoxy-2-2′-bifuran 7) and one new isoflavone glycoside (4H-1-Benzopyran-4-one,5-hydroxymethyl-3-(4-methoxyphenyl)-6-β-d-glucopyranoside-isoflavone 12). Their structures were determined by extensive analysis of their spectroscopic data. Among them, compounds 4, 6–10, 12 were for the first time isolated from this genus. The chemotaxonomic importance of these compounds was also summarized.     

Identification of dehydrocostus lactone and 4-hydroxy-β-thujone as auxin polar transport inhibitors

The survey of naturally occurring of auxin polar transport regulators in Asteraceae was investigated using the radish (Raphanus sativus L.) hypocotyl bioassay established in this study. Significant auxin polar transport was observed when radiolabeled indole-3-acetic acid (IAA) was applied at the apical side of radish hypocotyl segments, but not when it was applied at the basal side of the segments. Almost no auxin polar transport was observed in radish hypocotyl segments treated with synthetic auxin polar transport inhibitors of N-(1-naphthyl)phthalamic acid (NPA) and 9-hydroxyfluorene-9-carboxylic acid (HFCA) at 0.5 μg/plant. 2,3,5-Triiodobenzoic acid (TIBA) at 0.5 μg/plant was less effective than NPA and HFCA, and p-chlorophenoxyisobutyric acid (PCIB) at 0.5 μg/plant had almost no effect on auxin polar transport in the radish hypocotyl bioassay. These results strongly suggest that the radish hypocotyl bioassay is suitable for the detection of bioassay-derived auxin polar transport regulators. Using the radish hypocotyl bioassay and physicochemical analyses, dehydrocostus lactone (decahydro-3,6,9-tris-methylene-azulenol(4,5-b)furan-2(3H)-one) and 4-hydroxy-β-thujone (4-hydroxy-4-methyl-1-(1-methylethyl)-bicyclo[3.1.0]hexan-3-one) were successfully identified as auxin polar transport inhibitors from Saussurea costus and Arctium lappa, and Artemisia absinthium, respectively. About 50 and 40 % inhibitions of auxin polar transport in radish hypocotyl segments were observed at 2.5 μg/plant pre-treatment (see “Materials and methods”) of dehydrocostus lactone and 4-hydroxy-β-thujone, respectively. Although the mode of action of these compounds in inhibiting auxin polar transport has not been clear yet, their possible mechanisms are discussed.     

2-aryl-8-chloro-1,2,4-triazolo[1,5-a]quinoxalin-4-amines as highly potent A1 and A3 adenosine receptor antagonists

Some 2-aryl-8-chloro-1,2,4-triazolo[1,5-a]quinoxaline derivatives 2-18, obtained by introducing different substituents on either the 4-amino moiety (acyl or carbamoyl groups) or the 2-phenyl ring (4-OCH3) of previously reported 8-chloro-2-phenyl-1,2,4-triazolo[1,5-a]quinoxalin-4-amine (1), have been synthesized and tested in radioligand binding assays at bovine A1 and A(2A) and at cloned human A1 and A3 adenosine receptors. The rationally designed 8-chloro-2-(4-methoxy-phenyl)-1,2,4-triazolo[1,5-a]quinoxalin-4-acetylamine (14) can be considered one of the most potent and hA3 versus hA1 selective AR antagonists reported till now. The structure-activity relationships of compounds 2-18 are in agreement with those of previously reported 2-aryl-1,2,4-triazolo[4,3-a]quinoxalines (series A) and 2-arylpyrazolo[3,4-c]quinolines (series B), thus suggesting a similar AR binding mode. In fact, the importance for the A3 receptor-ligand interaction of both a strong acidic NH proton donor and a C=O proton acceptor at position-4, able to engage hydrogen-bonding interactions with specific sites on the A3 AR, has been confirmed. Using our recently published hA3 receptor model, to better elucidate our experimental results, we decided to theoretically depict the putative TM binding motif of the herein reported 1,2,4-triazolo[1,5-a]quinoxaline derivatives on human A3 receptor. Structure-activity relationships have been explained analyzing the three-dimensional structure of the antagonist-receptor models obtained by molecular docking simulation.