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PubMedDate not supplied

Microspectrofluorometry in biogenic amine research.

Microspectrofluorometry has played an important part in the development and understanding of fluorescence techniques for the demonstration of biogenic amines. This paper reviews briefly the techniques currently available for the characterization and localization of biogenic amines, and considers the basic design considerations applicable to the construction of a microspectrofluorometer, with special emphasis on the automated correction of spectral data. Spectral data for the commonly occurring biogenic amines dopamine, noradrenaline and 5-hydroxytryptamine have been summarized and the criteria applicable to the spectral analysis of the individual amines and their mixtures discussed. Discrepancies between spectral data derived from tissues and model systems are reviewed and emphasis is given to the importance of biochemical studies of amines prior to definitive microspectrofluorometric analysis. The conceptual basis of quantitative microspectrofluorometry is reviewed with a particular emphasis on the limitations of such studies. The analysis of terminal fields by scanning microspectrofluorometry and its application to the study of extracellular amine distribution is discussed.

Animals↗
PubMed1998

The heterocyclic amine binding receptors of Lactobacillus gasseri cells.

Lactobacillus gasseri is a common inhabitant of human intestine. The L. gasseri strains SBT10239 and SBT10241 have shown high antimutagenicity and binding properties with different heterocyclic amines. In order to identify the cell wall components involved in binding with the heterocyclic amines, the cells and cell walls of L. gasseri strains were subjected to different chemical and enzymatical treatments, prior to the binding experiments. The results indicated that the binding receptors for heterocyclic amines are the carbohydrate moieties of the cell wall. Binding of the heterocyclic amines with L. gasseri cell walls and the carbohydrate content showed high correlation coefficient, whereas it was insignificant or negative with protein content. The lectin binding studies revealed that the glucose molecules of the cell wall has a significant role in binding the heterocyclic amines. The inhibition caused by the lectin Concanavalin A was reversed when treated with methyl glucoside, a competitive inhibitor of Concanavalin A and restored the binding of heterocyclic amine with the cells.

Antimutagenic Agents↗
PubMed2000

Pathway tracing using biotinylated dextran amines.

Biotinylated dextran amines (BDA) are highly sensitive tools for anterograde and retrograde pathway tracing studies of the nervous system. BDA can be reliably delivered into the nervous system by iontophoretic or pressure injection and visualized with an avidin-biotinylated HRP (ABC) procedure, followed by a standard or metal-enhanced diaminobenzidine (DAB) reaction. High molecular weight BDA (10 k) yields sensitive and exquisitely detailed labeling of axons and terminals, while low molecular weight BDA (3 k) yields sensitive and detailed retrograde labeling of neuronal cell bodies. The detail of neuronal cell body labeling can be Golgi-like. BDA tolerates EM fixation and processing well and can, therefore, be readily used in ultrastructural studies. Additionally, BDA can be combined with other anterograde or retrograde tracers (e.g. PHA-L or cholera toxin B fragment) and visualized either by multi-color DAB multiple-labeling - if permanent labels are desired, or by using multiple simultaneous immunofluorescence - if fluorescence viewing is desired. In the same manner, BDA pathway tracing and neurotransmitter immunolabeling can be combined. Note that BDA pathway tracing can also be combined with anterograde or retrograde labeling with fluorescent dextran amines, if one wishes to exclusively use tracers with the favorable transport properties and sensitivities of dextran amines. In this case, the BDA can be visualized together with the fluorescent dextran amines using fluorescence labeling for the BDA, or the fluorescent dextran amines can be visualized together with the BDA by multicolor DAB labeling via immunolabeling of the fluorescent dextran amines using anti-fluorophore antisera. BDA is, thus, a flexible and valuable pathway tracing tool that has gained widespread popularity in recent years.

Animals↗
PubMed2001

A biogenic amine-synapse mechanism for mental retardation and developmental disabilities.

Recent studies have demonstrated that biogenic amines have a function of facilitating formation and maintenance of synapses in diverse regions of the central nervous system in developing and adult animals. The normal number of synapses maintained by biogenic amines are crucial to acquire learning and memory. The level of biogenic amines was reported to decrease in the brain by several neurodevelopmental disorders associated with mental retardation and developmental disabilities such as Rett syndrome, autism and Down syndrome. Taken into consideration this fact together with the function of biogenic amines for synapses, the density of synapses appears to decrease considerably in the brains of patients suffered from the neurodevelopmental disorders. The synaptic overproduction during the critical period of development especially 1 year after birth has been considered as a background mechanism to provide plasticity for the developing brain. Synaptic overproduction does not appear to occur in the brains of patients suffered from the neurodevelopmental disorders, which they are observed mental retardation occurring in the first 1 year after birth. Along with the neurodevelopmental disorders, environmental factors (stress, drugs and nutrition) during pre- and post-natal critical developmental periods are known to change levels of biogenic amines in the brain. In fact, maternal stress has been shown to decrease the levels of serotonin and the density of synapses in the hippocampus of the offspring, and they showed developmental disabilities in the spatial learning and memory. A cascade appears to exist from either the child neurological disorders or the environmental factors to mental retardation and developmental disabilities by decreases in the levels of biogenic amines and synaptic density.

Animals↗
PubMed2000

Tagging DNA mismatches by selective 2'-amine acylation.

BACKGROUND: Widespread characterization of genetic variation and disease at the gene-sequence level has inaugurated a new era in human biology. Techniques for the molecular analysis of these variations and their linkage with measurable phenotypes will profoundly affect diverse fields of biological chemistry and biology. RESULTS: A chemical tagging method has been developed to detect point mutations and other defects in nucleic acid sequences. The method employs oligodeoxynucleotide probes in which one 2'-ribose position (-H) is substituted with an amine (-NH(2)) group. 2'-Amine-substituted nucleotides are specifically acylated by succinimidyl esters to form a 2'-amide product. The mutation detection method exploits our observation that 2'-amine groups at the site of a mismatch are acylated more rapidly than amine substitutions at base-paired nucleotides. 2'-Amine acylation is governed primarily by local, rather than global, differences in nucleotide dynamics, such that site-specific tagging of DNA mismatches does not require discriminatory hybridization conditions to be determined. CONCLUSIONS: 2'-Amine mismatch tagging offers an approach for chemically interrogating the base-paired state of individual nucleotides in a hybridized duplex and for quantifying nucleicacid hybridization with single-base specificity.

Acylation↗
PubMed2006

Hyperbranched poly(amino ester)s with different terminal amine groups for DNA delivery.

Hyperbranched poly(amino ester)s containing tertiary amines in the core and primary, secondary, and tertiary amines in the periphery, respectively, were evaluated for DNA delivery in vitro. The same core structure facilitated the investigation on the effects of the terminal amine type on the properties of hyperbranched poly(amino ester)s for DNA delivery. The hydrolysis of the poly(amino ester)s was monitored using (1)H NMR. The results reflected that the terminal amine type had negligible effects on the hydrolysis rate but was much slower than that of linear poly(amino ester)s, probably due to the compact hyperbranched spatial structure preventing the accessibility of water. In comparison with PEI 25 K, the hyperbranched poly(amino ester)s showed much lower cytotoxicity in Cos7, HEK293, and HepG2 cells. Gel electrophoresis indicated that poly(amino ester)s could condense DNA efficiently, and the zeta potentials and sizes of the complexes formed with different weight ratios of hyperbranched poly(amino ester)s and DNA were measured. Remarkably, all the hyperbranched poly(amino ester)s showed DNA transfection efficiency comparable to PEI 25 K in Cos7, HEK293, and HepG2 cells regardless of the terminal amine type. Therefore, the terminal amine type had insignificant effects on the hydrolysis rate, cytotoxicity, DNA condensation capability, and in vitro DNA transfection efficiency of the hyperbranched poly(amino ester)s.

Animals↗
PubMed2003

Ruthenium complexes of 2-[(4-(arylamino)phenyl)azo]pyridine formed via regioselective phenyl ring amination of coordinated 2-(phenylazo)pyridine: isolation of products, X-ray structure, and redox and optical properties.

Aromatic ring amination reactions in the ruthenium complex of 2-(phenylazo)pyridine is described. The substitutionally inert cationic brown complex [Ru(pap)(3)](ClO(4))(2) (1) (pap = 2-(phenylazo)pyridine) reacts smoothly with aromatic amines neat and in the presence of air to produce cationic and intense blue complexes [Ru(HL(2))(3)](ClO(4))(2) (2) (HL(2) = 2-[(4-(arylamino)phenyl)azo]pyridine). These were purified on a preparative TLC plate. The X-ray structure of the new and representative complex 2c has been solved to characterize them. The results are compared with those of the starting complex, [Ru(pap)(3)](ClO(4))(2) (1). The transformation 1 --> 2 involves aromatic ring amination at the para carbon (with respect to the diazo function) of the pendant phenyl rings of all three coordinated pap ligands in 1. The transformation is stereoretentive, and the amination reaction is regioselective. The extended ligand HL(2) coordinates as a bidentate ligand and chelates to ruthenium(II) through the pyridine and one of the azo nitrogens. The amine nitrogen of this bears a hydrogen atom and remains uncoordinated. Similarly, the amination reaction on the mixed-ligand complex [Ru(pap)(bpy)(2)](ClO(4))(2) produces the blue complex [Ru(HL(2))(bpy)(2)](ClO(4))(2) (3) as anticipated. The reactions of [RuCl(2)(dmso)(4)] and [Ru(S)(2)(L)(2)](2+) (dmso = dimethyl sulfoxide, S = labile coordinated solvent, L = 2,2'-bipyridine (bpy) and pap) with the preformed HL(2) ligand have been explored. The structure of the representative complex [RuCl(2)(HL(2a))(2)] (5a) is reported. It has the chlorides in trans configuration while the pyridine as well as azo nitrogens are in cis geometry. Optical spectra and redox properties of the newly synthesized complexes are reported. All the ruthenium complexes of HL(2) are characterized by their intense blue solution colors. The lowest energy transitions in these complexes appear near 600 nm, which have been attributed to intraligand charge-transfer transitions. For example, the lowest energy visible range transition in [Ru(HL(2b))(3)](2+) appears at 602 nm and its intensity is 65 510 M(-1) cm(-1). All the tris chelates show multiple-step electron-transfer processes. In [Ru(HL(2))(3)](2+), six reductions waves constitute the complete electron-transfer series. The electrons are believed to be added successively to the three azo functions. In the mixed-ligand chelates [Ru(HL(2))(pap)(2)](2+) and [Ru(HL(2))(bpy)(2)](2+) the reductions due to HL(2), pap, and bpy are observed.

Journal Article↗
PubMed2003

The role of amine in the mechanism of pentathiepin (polysulfur) antitumor agents.

A computational and experimental study is presented, which provides the first evidence that amine has an opportunity to engage in bonding with pentathiepin to promote its decomposition. The study provides mechanistic insight into the process that gives rise to pentathiepin biological activity. Primary or secondary amine will allow for an intramolecular addition to the pentathiepin ring at the nearest sulfur (S1). In contrast, tertiary amine adds reversibly to S1, because nitrogen cannot lose its positive charge by deprotonation. This precludes the amine promotion step. An energetically low-lying process is characterized, corresponding to S3-loss triggered by nucleophilic activation with a primary or secondary amine. Pentathiepin desulfurization via S3-unit transfer is supported by a trapping study with norbornene. That the amine may confer an enhanced reactivity in the natural products varacin, 1, and lissoclinotoxin A, 2, adds to the understanding of the pathway for pentathiepin activation and may provide new design concepts that have potential applications for this class of biocides.

Antineoplastic Agents↗
PubMed2004

Synthetic, structural, and mechanistic aspects of an amine activation process mediated at a zwitterionic Pd(II) center.

A zwitterionic palladium complex [[Ph(2)BP(2)]Pd(THF)(2)][OTf] (1) (where [Ph(2)BP(2)] = [Ph(2)B(CH(2)PPh(2))(2)](-)) reacts with trialkylamines to activate a C-H bond adjacent to the amine N atom, thereby producing iminium adduct complexes [Ph(2)BP(2)]Pd(N,C:eta(2)-NR(2)CHR'). In all cases examined the amine activation process is selective for the secondary C-H bond position adjacent to the N atom. These palladacycles undergo facile beta-hydride elimination/olefin reinsertion processes as evident from deuterium scrambling studies and chemical trap studies. The kinetics of the amine activation process was explored, and beta-hydride elimination appears to be the rate-limiting step. A large kinetic deuterium isotope effect for the amine activation process is evident. The reaction profile in less polar solvents such as benzene and toluene is different at room temperature and leads to dimeric [[Ph(2)BP(2)]Pd](2) (4) as the dominant palladium product. Low-temperature toluene-d(8) experiments proceed more cleanly, and intermediates assigned as [Ph(2)BP(2)]Pd(NEt(3))(OTf) and the iminium hydride species [[Ph(2)BP(2)]Pd(H)(Et(2)N=CHCH(3))][OTf] are directly observed. The complex (Ph(2)SiP(2))Pd(OTf)(2) (14) was also studied for amine activation and generates dimeric [(Ph(2)SiP(2))Pd](2)[OTf](2) (16) as the dominant palladium product. These collective data are discussed with respect to the mechanism of the amine activation and, in particular, the influence that solvent polarity and charge have on the overall reaction profile.

Journal Article↗
PubMed2005

New insights into the mechanism of palladium-catalyzed allylic amination.

A comparative investigation into palladium-catalyzed allylic amination of unsubstituted aziridines and secondary amines has been carried out. The use of NH aziridines as nucleophiles favors formation of valuable branched products in the case of aliphatic allyl acetates. The regioselectivity of this reaction is opposite to that observed when other amines are used as nucleophiles. Our study provides evidence for the palladium-catalyzed isomerization of the branched (kinetic) product formed with common secondary amines into the thermodynamic (linear) product. In contrast, the branched allyl products obtained from unsubstituted aziridines do not undergo the isomerization process. Crossover experiments indicate that the isomerization of branched allylamines is bimolecular and is catalyzed by Pd(0). The reaction has significant solvent effect, giving the highest branched-to-linear ratios in THF. This finding can be explained by invoking the intermediacy of sigma-complexes, which is consistent with NMR data. The apparent stability of branched allyl aziridines towards palladium-catalyzed isomerization is attributed to a combination of factors that stem from a higher degree of s-character of the aziridine nitrogen compared to other amines. The reaction allows for regio- and enantioselective incorporation of aziridine rings into appropriately functionalized building blocks. The resulting methodology addresses an important issue of forming quaternary carbon centers next to nitrogen. The new insights into the mechanism of palladium-catalyzed allylic amination obtained in this study should facilitate synthesis of complex heterocycles, design of new ligands to control branched-to-linear ratio, as well as absolute stereochemistry of allylamines.

Journal Article↗
PubMed1979

Kojic amine--a novel gamma-aminobutyric acid analogue.

A series of compounds containing the 3-hydroxy-4H-pyran-4-one nucleus has been synthesized and tested as potential skeletal muscle relaxants. Reduction of 2-(azidomethyl)-5-hydroxy-4H-pyran-4-one (4) with HBr in HOAc--phenol yielded 2-(aminomethyl)-5-hydroxy-4H-pyran-4-one (kojic amine, 3) in 81% yield. Reaction of 2-[(tosyloxy)-methyl]-5-(benzyloxy)-4H-pyran-4-one (5) with NH3 gave a 40% yield of the O-benzyl ether of kojic amine, which was N-acylated with a series of carbobenzyloxy-protected amino acids. Complete deprotection with HBr--HOAc gave the following amino acid amides of kojic amine: glycyl (23), alpha-alanyl (24), beta-alanyl (25), gamma-aminobutyryl (26), and glycylglycyl (27). Among the analogues of kojic amine prepared was a series of one-carbon homologues: 2-[(methylamino)methyl]-5-hydroxy-4H-pyran-4-one (7a), 2-(1-aminoethyl)-5-hydroxy-4H-pyran-4-one (8), 6-(aminomethyl)-3-hydroxy-2-methyl-4H-pyran-4-one (12), and 2-(2-aminoethyl)-5-hydroxy-4H-pyran-4-one (16). Kojic amine (3) has been found to possess certain of the properties to be expected in a gamma-aminobutyric acid mimetic agent, notably skeletal muscle relaxant activity. In the chronic spinal cat preparation, ED70 values for reduction of flexor spasms of 2.2 and 4.0 mg/kg by iv and po routes of administration, respectively, were observed for kojic amine, which was the most potent of the various hydroxypyrone derivatives investigated.

Animals↗
PubMed2000

Kinetics and mechanisms of the reactions of 4-nitro- and 3-nitrophenyl 4-methylphenyl thionocarbonates with alicyclic amines and pyridines

The reactions of the title thionocarbonates (1 and 2, respectively) with a series of secondary alicyclic amines and pyridines are subjected to a kinetic investigation in 44 wt % ethanol-water, 25.0 degrees C, ionic strength 0.2 M (KCl). Under amine excess over the substrates pseudo-first-order rate coefficients (k(obsd)) are obtained for all the reactions. Those of the alicyclic amines with the two substrates show nonlinear upward plots of k(obsd) vs [amine], except the reactions of piperidine, which exhibit linear plots. For these reactions a reaction scheme is proposed with two tetrahedral intermediates, one zwitterionic (T(+/-)) and the other anionic (T(-)), with a kinetically significant proton transfer from T(+/-) to an amine to give T(-). From an equation derived from the scheme the rate microcoefficients are obtained through fitting. The rate coefficient for formation of T(+/-) (k(1)) is larger for 1 compared to 2, which can be explained by a stronger electron-withdrawal of 4-nitro in 1 than 3-nitro in 2, which leaves the thiocarbonyl carbon of 1 more positive and, therefore, more susceptible to nucleophilic attack. For the pyridinolyses of both thionocarbonates the plots of k(obsd) vs [amine] are linear, with the slope (k(N)) independent of pH. The Bronsted plots (log k(N) vs pyridine pK(a)) for these reactions are linear with slopes beta = 0.9 and 1.2 for the pyridinolysis of 1 and 2, respectively. These slopes are consistent with a mechanism through a T(+/-) intermediate on the reaction path, whereby decomposition of T(+/-) to products is the rate-determining step. The k(N) values are larger for the reactions of 1 than those of 2. This is attributed to a larger equilibrium formation of T(+/-) and a larger expulsion rate of the nucleofuge from T(+/-) in the reactions of 1 compared to those of 2.

Journal Article↗
PubMed2001

Kinetics and mechanisms of the reactions of 3-methoxyphenyl, 3-chlorophenyl, and 4-cyanophenyl 4-nitrophenyl thionocarbonates with alicyclic amines.

The reactions of 3-methoxyphenyl, 3-chlorophenyl, and 4-cyanophenyl 4-nitrophenyl thionocarbonates (1, 2, and 3, respectively) with a series of secondary alicyclic amines are studied kinetically in 44 wt % ethanol-water at 25.0 degrees C and an ionic strength of 0.2 M (KCl). Pseudo-first-order rate coefficients (k(obsd)) are obtained for all reactions (amine excess was used). The reactions of compound 1 with piperidine, piperazine, and 1-(2-hydroxyethyl)piperazine and of compounds 2 and 3 with these amines and morpholine exhibit linear k(obsd) versus amine concentration plots with slopes (k1) independent of pH. In contrast, the plots are nonlinear upward for the reactions of substrate 1 with morpholine, 1-formylpiperazine, and piperazinium ion and of substrates 2 and 3 with the two latter amines. For all these reactions, a reaction scheme is proposed with a zwitterionic tetrahedral intermediate (T+/-), which can be deprotonated by an amine to yield an anionic intermediate (T-). When the nonlinear plots are fit through an equation derived from the scheme, rate and equilibrium microcoefficients are obtained. The Brönsted-type plots for k1 are linear with slopes of beta1 = 0.22, 0.20, and 0.24 for the aminolysis of 1, 2, and 3, respectively, indicating that the formation of T+/- (k1 step) is rate-determining. The k1 values for these reactions follow the sequence 3 > 2 > 1, which can be explained by the sequence of the electron-withdrawing effects from the substituents on the nonleaving group of the substrates.

Journal Article↗
PubMed2002

NMR and ab initio studies of amination of ketenimine: direct evidence for a mechanism involving a vinylidenediamine as an intermediate.

High-level ab initio calculations were carried out in both gas phase and solvent (epsilon = 35.9) to estabilish that the amination of ketenimine proceeds via amine addition across the C=N bond rather than the C=C bond, followed by tautomerization to form amidine product. The HOMO of ketenimine is perpendicular to its molecular plane with the largest coefficient on C(beta), while the LUMO is in its molecular plane with the largest coefficient on C(alpha). Amination of ketenimine involves in-plane attack of amine nucleophile on C(alpha) (LUMO) of ketenimine. The labile vinylidenediamine intermediate trans-11 for the reaction of ketenimine 10 with n-butylamine was directly observed by means of low-temperature proton NMR spectra. The evidence confirms that the amination reaction is stepwise and proceeds via n-butylamine addition across the C=N bond of ketenimine 10 rather than the C=C bond, followed by a slower tautomerization of vinylidenediamine trans-11 to amidine 12. Even though the second step is much slower, the first step involving amine addition across the C=N bond is kinetic control. Surprisingly, in the reaction of 10 with n-BuNH(2), attack of n-BuNH(2) syn to the phenyl group on C(beta) of 10 is preferred, even though this produces a less stable product (trans-11); attack of n-BuNH(2) anti to phenyl group on C(beta) of 10 is lacking and results in serious nonbonding interactions between the two phenyls of the ketenimine, as they are pushed together in this transition state.

Journal Article↗
PubMed2003

Regioselectivity and the nature of the reaction mechanism in nucleophilic substitution reactions of 2,4-dinitrophenyl X-substituted benzenesulfonates with primary amines.

Second-order rate constants have been measured for the reaction of 2,4-dinitrophenyl X-substituted benzenesulfonates with a series of primary amines. The nucleophilic substitution reaction proceeds through competitive S-O and C-O bond fission pathways. The S-O bond fission occurs dominantly for reactions with highly basic amines or with substrates having a strong electron-withdrawing group in the sulfonyl moiety. On the other hand, the C-O bond fission occurs considerably for the reactions with low basic amines or with substrates having a strong electron-donating group in the sulfonyl moiety, emphasizing that the regioselectivity is governed by both the amine basicity and the electronic effect of the sulfonyl substituent X. The apparent second-order rate constants for the S-O bond fission have resulted in a nonlinear Brønsted-type plot for the reaction of 2,4-dinitrophenyl benzenesulfonate with 10 different primary amines, suggesting that a change in the rate-determining step occurs upon changing the amine basicity. The microscopic rate constants (k(1) and k(2)/k(-)(1) ratio) associated with the S-O bond fission pathway support the proposed mechanism. The second-order rate constants for the S-O bond fission result in good linear Yukawa-Tsuno plots for the aminolyses of 2,4-dinitrophenyl X-substituted benzenesulfonates. However, the second-order rate constants for the C-O bond fission show no correlation with the electronic nature of the sulfonyl substituent X, indicating that the C-O bond fission proceeds through an S(N)Ar mechanism in which the leaving group departure occurs rapidly after the rate-determining step.

Journal Article↗
PubMed2006

Pd(OAc)2-catalyzed carbonylation of amines.

A phosphine-free catalytic system [Pd(OAc)2-Cu(OAc)2-air] induced a substrate-specific carbonylation of amines in boiling toluene under CO gas (1 atm). Symmetrical N,N'-dialkylureas were obtained by the carbonylation of primary amines. N,N,N'-Trialkylureas were selectively formed by addition of a secondary amine to the above reaction vessel. Secondary amines did not give tetraalkylureas. However, dialkylamines with a phenyl group on their alkyl chains, such as N-monoalkylated benzylic amine or phenethylamine derivatives, underwent a direct aromatic carbonylation to afford five- or six-membered benzolactams. In the carbonylation, the chelation effect or steric repulsion between Pd(II) and the meta-substituent in the ortho-palladation and the ring sizes of cyclopalladation products that were formed prior to carbonylation were found to generate good site selectivity and increase the reaction rate. In contrast, carbonylation of omega-arylalkylamines with a hydroxyl group gave neither ureas nor benzolactams but instead produced 1,3-oxazolidinones smoothly. Hydrochlorides of amines also underwent carbonylation to afford the corresponding amides under the conditions used. This procedure made it possible to prepare ureas of amino acid esters and N-alkylcarbamates in practical yields.

Journal Article↗
PubMed2006

DNA micropatterning on polycrystalline diamond via one-step direct amination.

We report a novel method of one-step direct amination on polycrystalline diamond to produce functionalized surfaces for DNA micropatterning by photolithography. Polycrystalline diamond was exposed to UV irradiation in ammonia gas to generate amine groups directly. After patterning, optical microscopy confirmed that micropatterns covered with an Au mask were regular in size and shape. The regions outside the micropatterns were passivated with fluorine termination by C3F8 plasma, and the chemical changes on the two different surfaces--the amine groups inside the patterned regions by one-step direct amination and fluorine termination outside the patterned regions--were characterized by spatially resolved X-ray photoelectron spectroscopy (XPS). The patterned areas terminated with active amine groups were then immobilized with probe DNA via a bifunctional molecule. The sequence specificity was conducted by hybridizing fluorescently labeled target DNA to both complementary and noncomplementary probe DNA attached inside the micropatterns. The fluorescence micropatterns observed by epifluorescence microscopy corresponded to those imaged by optical microscopy. DNA hybridization and denaturation experiments on a DNA-modified diamond show that the diamond surfaces reveal superior stability. The influence of a different amination time on fluorescence intensity was compared. Different terminations as passivated layers were investigated, and as a result, fluorine termination points to the greatest signal-to-noise ratio.

Adsorption↗
PubMed1998

Effect of ring size on conformations of aromatic amine-DNA adducts: the aniline-C8 guanine adduct resides in the B-DNA major groove.

While the one-ring amine aniline (AN) has only slight genetic activity, the polycyclic aromatic amines 2-aminofluorene (AF) and 1-aminopyrene (AP) are significant mutagens and carcinogens. Moreover, the bulkier AP is more mutagenic per adduct than AF in the tetracycline-resistance gene of plasmid pBR322 [Melchior et al. (1994) Carcinogenesis 15, 889]. To elucidate possible conformational origins of the differing mutagenic effects of these three adducts, which may stem from their differing ring sizes, we have examined their conformations in two mutation-susceptible sequences from the above gene: TTGAG*GCCG (sequence I) and GAATG*GTGC (sequence II), where G* = C8-modified guanine. No experimental high-resolution NMR data are yet available for the aniline adduct in a DNA duplex. Minimized potential energy calculations were carried out, using the molecular mechanics program DUPLEX to explore the conformation space of these adducts. In the case of AN, a relatively unperturbed B-DNA helix with the amine in the major groove was strongly favored in both sequences. In the case of AF- and AP-modified DNA, however, several differing conformations were competitive in energy. They included major groove structures, as well as conformations with syn-modified guanine and the polycyclic amine in the minor groove, or the amine rings intercalated into the helix with displacement of the modified guanine, in overall harmony with high-resolution NMR solution structures. Thus, aniline distorts DNA structure to a lesser extent than larger aromatic amine ring systems, since a number of different conformations are energetically feasible and have been observed for the larger systems. This result may be relevant to their enhanced mutagenicity and their repair propensity, in contrast to aniline's low mutagenic effect.

Aniline Compounds↗