Aminonaphthoquinone induces oxidative stress in Staphylococcus aureus.

The biological activity of 5-amino-8-hydroxy-1,4-naphthoquinone (ANQ) on Staphylococcus aureus was investigated in comparison with the unsubstituted 1,4-naphthoquinone (NQ). Complete inhibition of microbial growth was observed with ANQ and NQ at 50 and 10 microg/mL, respectively. The antibacterial effect of naphthoquinones decreased in the presence of sodium ascorbate, but the superoxide scavenger 4,5-dihydroxy-1,3-benzene-disulfonic acid (Tiron) was able to protect S. aureus only from the harmful effect of ANQ. Naphthoquinones blocked oxygen uptake and induced cyanide-insensitive oxygen consumption. When combining rotenone or salicylhydroxamic acid with ANQ or NQ, a slight decrease in respiratory activity was observed. Assays in the presence of naphthoquinones induced an increase of lipid peroxidation in S. aureus, as determined by thiobarbituric acid reactive substances. These results showed that 1,4-naphthoquinones effectively act as electron acceptors and induce an increase in reactive oxygen species that are toxic to S. aureus cells.     

Synthesis and cytotoxicity of new heterocyclic terpenylnaphthoquinones

Several 2-arylamino-, 2-aryloxy- and 2-arylsulfanyl-6(7)-alkyl-1,4-naphthoquinones (NQ) have been prepared and further transformed into the corresponding heterocyclic-fused naphthoquinones by palladium (II)-catalyzed oxidative cyclization. The compounds synthesized have been evaluated against neoplastic cell lines. The extension of the polycyclic system clearly decreased the cytotoxic potency of the 2-substituted terpenylnaphthoquinones.     

Photochemical transformation of 1-naphthol in aerated aqueous solution.

The phototransformation of 1-naphthol in aerated aqueous solution was investigated by means of product studies and laser flash photolysis. The quantum yield as measured at 313 nm was found to be equal to 3.2 x 10(-2) in oxygen-saturated medium while being 10-fold lower in deoxygenated solution. The main photoproducts in aerated medium were 1,4-naphthoquinone, 2-hydroxy-1,4-naphthoquinone and 6-hydroxy-1,4-naphthoquinone. Traces of 1,2-naphthoquinone and 5-hydroxy-1,4-naphthoquinones were detected too. Solvated electrons were detected by laser flash photolysis. The quantum yield of monophotonic ionisation was found to be lower than that of 1-naphthol photolysis indicating that other reaction pathways took place. The mechanisms of naphthoquinones and hydroxynaphthoquinones formation are discussed.     

Multivariate analysis of Photosystem II fluorescence quenching by substituted benzoquinones and naphthoquinones

The present investigation examines the action of substituted benzoquinones and naphthoquinones as inhibitors of chlorophyll fluorescence in barley chloroplasts. Stern-Volmer analyses have been used to determine the fractions of chlorophyll fluorescence accessible to quinone as a measure of quinone-membrane solubility and the Stern-Volmer quenching constant as a measure of quinone intrinsic activity. Correlations between quinone structural properties and experimentally measured chlorophyll fluorescence parameters have been made to determine the contribution of lipophilic and electronic factors to quinone solubility and intrinsic activity. In particular, 1,4-benzoquinones must have hydrophobic substituents or hydrogen atoms at positions 2, 3, 5 and 6 for fluorescence quenching activity. The magnitude of the Stern-Volmer quenching constant is affected by the electronic character of hydrophobic substituents, with a maximum value observed in the presence of both electron-releasing and electron-withdrawing moieties. Similarly, the lipophilic character of 2,3-substituents of 1,4-naphthoquinones determines the fraction of chlorophyll fluorescence accessible to quinone, with a maximum accessibility observed for hydrophobic substituents. Naphthoquinone intrinsic activity is also correlated with the electronic character of the substituents and is observed to be at a maximum value with only electron-withdrawing substituents.     

DT-diaphorase-catalysed reduction of 1,4-naphthoquinone derivatives and glutathionyl-quinone conjugates. Effect of substituents on autoxidation rates.

DT-diaphorase catalysed the reduction of 1,4-naphthoquinones with hydroxy, methyl, methoxy and glutathionyl substituents at the expense of reducing equivalents from NADPH. The initial rates of quinone reduction did not correlate with either the half-wave reduction potential (E1/2) value (determined by h.p.l.c. with electrochemical detection against an Ag/AgCl reference electrode) or the partition coefficient of the quinones. After their reduction by DT-diaphorase the 1,4-naphthoquinone derivatives autoxidized at distinct rates, the extent of which was influenced by the nature of the substituents. Thus for the 1,4-naphthoquinone series the following order of rate of autoxidation was found: 5-hydroxy-1,4-naphthoquinone greater than 3-glutathionyl-1,4-naphthoquinone greater than 5-hydroxy-3-glutathionyl-1,4-naphthoquinone greater than 1,4-naphthoquinone greater than 2-hydroxy-1,4-naphthoquinone. For the 2-methyl-1,4-naphthoquinone (menadione) series the following order was observed: 5-hydroxy-2-methyl-1,4-naphthoquinone greater than 3-glutathionyl-5-hydroxy-2-methyl-1,4-naphthoquinone greater than 3-glutathionyl-2-methyl-1,4-naphthoquinone greater than 2-methyl-1,4-naphthoquinone greater than 3-hydroxy-2-methyl-1,4-naphthoquinone. The autoxidized naphthohydroquinone derivatives were re-reduced by DT-diaphorase, thus closing a cycle of enzymic reduction in equilibrium autoxidation. This was expressed as an excess of NADPH oxidized over the initial concentration of quinone present as well as H2O2 formation. These findings demonstrate that glutathionyl conjugates of 1,4-naphthoquinone and 2-methyl-1,4-naphthoquinone and those of their respective 5-hydroxy derivatives are able to act as substrates for DT-diaphorase and that they also autoxidize at rates higher than those for the unsubstituted parent compounds. These results are discussed in terms of the cellular role of DT-diaphorase in the reduction of hydroxy- or glutathionyl-substituted naphthoquinones as well as the further conjugation of these hydroquinones with glucuronide or sulphate within the cellular milieu, thereby facilitating their disposal from the cells.     

2-chloro-3-substituted-1,4-naphthoquinone inactivators of human cytomegalovirus protease.

A random screening approach has identified 2-chloro-3-substituted-1,4-naphthoquinones as potent inactivators of HCMV protease. Enzyme inactivation is due to modification of Cys202. Two of the most potent compounds maintain activity against HCMV in a plaque reduction assay.     

Elucidation of structure-activity relationships for 2- or 6-substituted-5,8-dimethoxy-1,4-naphthoquinones

1,4-Naphthoquinones have already been recognized to possess a wide range of biological activities. We have developed quantitative structure activity relationships (QSAR) for different series of 2- or 6-substituted-5,8-dimethoxy-1,4-naphthoquinones to understand the chemical-biological interaction governing antiproliferative/cytotoxic activities against L1210 cells. QSAR results have shown that these activities of 2- or 6-substituted-5,8-dimethoxy-1,4-naphthoquinones depend largely on their hydrophobicity.     

Evaluation of selected benzoquinones, naphthoquinones, and anthraquinones as replacements for phylloquinone in the A1 acceptor site of the photosystem I reaction center

Selected substituted 1,4-benzoquinones, 1,4-naphthoquinones, and 9,10-anthraquinones were investigated as possible replacement quinones in spinach photosystem I (PSI) preparations that had been depleted of endogenous phylloquinone by extraction with hexane/methanol. As a criterion for successful biochemical reconstitution, the restoration of electron transfer was determined by measuring P-430 turnover at room temperature from flash-induced absorbance transients. Restoration of complete electron transfer between A0- and P-430 (terminal iron-sulfur centers, FAFB) was demonstrated by using phylloquinone, 2-methyl-3-decyl-1,4-naphthoquinone, 2-methyl-3-(isoprenyl)2-1,4-naphthoquinone, and 2-methyl-3-(isoprenyl)4-1,4-naphthoquinone. All other quinones tested did not restore P-430 turnover but acted as electron acceptors and oxidized A0-. It is concluded that the specificity of the replacement quinone for interaction with the primary acceptor, A0-, is low but additional structural constraints are required for the quinone occupying the A1 site to donate to the iron-sulfur center, Fx. It is suggested that the 3-phytyl side chain of phylloquinone and the 3-alkyl tails of the three naphthoquinones that restored P-430 turnover may be required for interaction with a hydrophobic domain of the A1 site in the PSI core to promote electron transfer to Fx and then to FAFB.     

Synthesis and biological evaluation of novel (L)-alpha-amino acid methyl ester, heteroalkyl, and aryl substituted 1,4-naphthoquinone derivatives as antifungal and antibacterial agents

A series of (S)-N-(1,4-naphthoquinon-2-yl)-alpha-amino acid methyl esters 3-9, 2-N,N-dialkylamino-1,4-naphthoquinones 10-11 and 2-hydroxy-3-(2'-mercaptoimidazolyl)-1,4-naphthoquinones and their cyclic analogs 12-15 were synthesized and evaluated for antifungal and antibacterial activities. The structure-activity relationships of these compounds were studied and the results show that the compounds 9b and 13c exhibited in vitro antifungal activity against Candida albicans, Cryptococcus neoformans, and Sporothrix schenckii, whereas compound 6a showed in vitro antibacterial activity against Streptococcus faecalis, K. pneumoniae, Escherichia coli, and Staphylococcus aureus.     

Synthesis and evaluation of novel 1,4-naphthoquinone derivatives as antiviral, antifungal and anticancer agents

The synthesis and evaluation of some 2-substituted-1,4-naphthoquinones 2, S-(1,4-naphthoquinon-2-yl)-mercaptoalkanoic acid amides 4, related benzoquinone and naphthoquinone derivatives 6-9 and 2,3-disubstituted 1,4-naphthoquinones 10-11 were carried out. The antifungal, antibacterial, antiviral and anticancer activities were determined by using the standard assay. The results show that compounds 2b and 10a showed in vitro antiviral activity against Influenza-A Virus and Herpes Simplex Virus and possess pronounced antifungal profile whereas 4a showed anticancer activities against Lymphoid Leukaemia P 388.     

Comparative toxicity of alkyl-1,4-naphthoquinones in rats: Relationship to free radical production in vitro

2-Methyl-1,4-naphthoquinone causes haemolysis in vivo. This toxic effect is believed to result from oxidative damage to erythrocytes by “active oxygen” species formed via one-electron reduction of the naphthoquinone by oxyhaemoglobin. In the present investigation, seven 2-alkyl-1,4-naphthoquinones have been studied with regard to their haemolytic activity in rats, their ability to cause oxidative damage in erythrocytes in vitro, and their reactivity toward oxyhaemoglobin. A close correlation was observed between the in vivo and in vitro parameters, suggesting that the proposed mechanism of toxicity of 2-methyl-1,4-naphthoquinone is correct and is also applicable to other alkylnaphthoquinones.     

Antiproliferative activities and SAR studies of substituted anthraquinones and 1,4-naphthoquinones

STAT3 is constitutively active in a large variety of cancers. The search for STAT3 inhibitors led to the discoveries of LLLs 3 and 12, which are substituted anthraquinones. LLL12 is an extremely potent compound that exhibits high levels of antiproliferative activity. Herein the synthesis and evaluation of compounds containing either an anthraquinone or 1,4-naphthoquinone moiety are reported. Analogs were evaluated in several cancer cell lines. Interestingly, it was found that the anthraquinones did not follow the same trends as the 1,4-naphthoquinones in regards to potency. LLL12, which contains a sulfonamide at position 1, was found to be the most potent of the anthraquinones. In contrast, the methyl ketone and methyl ester derivatives (LLLs 3.1 and 5.1) were found to be the most potent of the 1,4-naphthoquinones. Selected 1,4-naphthoquinones were also evaluated in the STAT3 fluorescence polarization assay in order to evaluate their abilities to bind to the STAT3 SH2 domain. They were found to have similar affinities, and their activities suggest that STAT3 is one of their molecular targets.     

Targeting of Helicobacter pylori thymidylate synthase ThyX by non-mitotoxic hydroxy-naphthoquinones

ThyX is an essential thymidylate synthase that is mechanistically and structurally unrelated to the functionally analogous human enzyme, thus providing means for selective inhibition of bacterial growth. To identify novel compounds with anti-bacterial activity against the human pathogenic bacterium Helicobacter pylori, based on our earlier biochemical and structural analyses, we designed a series of eighteen 2-hydroxy-1,4-naphthoquinones (2-OH-1,4-NQs) that target HpThyX. Our lead-like molecules markedly inhibited the NADPH oxidation and 2′-deoxythymidine-5′-monophosphate-forming activities of HpThyX enzyme in vitro, with inhibitory constants in the low nanomolar range. The identification of non-cytotoxic and non-mitotoxic 2-OH-1,4-NQ inhibitors permitted testing their in vivo efficacy in a mouse model for H. pylori infections. Despite the widely assumed toxicity of naphthoquinones (NQs), we identified tight-binding ThyX inhibitors that were tolerated in mice and can be associated with a modest effect in reducing the number of colonizing bacteria. Our results thus provide proof-of-concept that targeting ThyX enzymes is a highly feasible strategy for the development of therapies against H. pylori and a high number of other ThyX-dependent pathogenic bacteria. We also demonstrate that chemical reactivity of NQs does not prevent their exploitation as anti-microbial compounds, particularly when mitotoxicity screening is used to prioritize these compounds for further experimentation.     

2,3-Dimethoxy-5-methyl-1,4-benzoquinones and 2-methyl-1,4-naphthoquinones: glycation inhibitors with lipid peroxidation activity

Anti-glycation activity of our anti-oxidant quinone library was measured and several 2,3-dimethoxy-5-methyl-1,4-benzoquinones and 2-methyl-1,4-naphthoquinones were identified as novel inhibitors of glycation, of which 2,3-dimethoxy-5-methyl-1,4-benzoquinones 13b is the most potent glycation inhibitor with around 50 microM of the IC(50) value.     

Growth inhibitory activity for cancer cell lines of lapachol and its natural and semi-synthetic derivatives

A series of 17 selected natural and semisynthetic 1,4-naphthoquinones were synthesized, and their growth inhibitory activity was evaluated in vitro. The compounds were tested on six human cancer cell lines using the MTT colorimetric assay. The data revealed that of the chemicals under study only lapachol, its acetate and 3-geranyllawsone displayed the highest activity, recording mean IC₅₀ values ranging from 15 to 22 μM.     

An unusual reduction on the quinonoid ring of 5-amino-8-hydroxy-1,4-naphthoquinone by Staphylococcus aureus

AIMS: The objective of this study was to investigate the interactions between 5-amino-8-hydroxy-1,4-naphthoquinone (ANQ) and Staphylococcus aureus. METHODS AND RESULTS: The compound ANQ display antimicrobial activity against S. aureus. During incubation with 50 microg ml(-1) of ANQ, an unusual reduction reaction takes place and leads to the isolation of 2,3-dihydro-5-amino-8-hydroxy-1,4-naphthoquinone (ANQ-H(2)), fully characterized by means of (13)C-NMR and (1)H-NMR, plus infrared, UV-visible and mass spectroscopy. Oxygen uptake by S. aureus cells was inhibited by ANQ, but in a significantly minor extent by ANQ-H(2). CONCLUSIONS: The ability of S. aureus to reduce the double bond at C2-C3 of the ANQ is a unusual behaviour for biological transformation of naphthoquinones. SIGNIFICANCE AND IMPACT OF THE STUDY: This uncommon reaction may provide valuable understanding of the S. aureus regarding to the antimicrobial effect and the acquisition of resistance to naphthoquinones.     

Characterization of the quinone reductase activity of the ferric reductase B protein from Paracoccus denitrificans

The ferric reductase B (FerB) protein of Paracoccus denitrificans exhibits activity of an NAD(P)H: Fe(III) chelate, chromate and quinone oxidoreductase. Sequence analysis places FerB in a family of soluble flavin-containing quinone reductases. The enzyme reduces a range of quinone substrates, including derivatives of 1,4-benzoquinone and 1,2- and 1,4-naphthoquinone, via a ping-pong kinetic mechanism. Dicoumarol and Cibacron Blue 3GA are competitive inhibitors of NADH oxidation. In the case of benzoquinones, FerB apparently acts through a two-electron transfer process, whereas in the case of naphthoquinones, one-electron reduction takes place resulting in the formation of semiquinone radicals. A ferB mutant strain exhibited an increased resistance to 1,4-naphthoquinone, attributable to the absence of the FerB-mediated redox cycling. The ferB promoter displayed a high basal activity throughout the growth of P. denitrificans, which could not be further enhanced by addition of different types of naphthoquinones. This indicates that the ferB gene is expressed constitutively.     

Singlet oxygen production by pyrano and furano 1,4-naphthoquinones in non-aqueous medium

The influence of ring size on the photobehaviour of condensed 1,4-naphthoquinone systems, such as pyrano- and furano-derivatives (1 and 2, respectively) has been investigated. The absorption spectra for both families of naphthoquinones reveal clear differences; in the case of 2 they extend to longer wavelengths. A solvatochromic red shift in polar solvents is consistent with the π,π* character of the S(0)→ S(1) electronic transition in all cases. Theoretical (B3LYP) analysis of the HOMO and LUMO Kohn-Sham molecular orbitals of the S(0) state indicates that they are π and π* in nature, consistent with the experimental observation. A systematic study on the efficiency of singlet oxygen generation by these 1,4-naphthoquinones is presented, and values larger than 0.7 were found in every case. In accordance with these results, laser flash photolysis of deoxygenated acetonitrile solutions led to the formation of detectable triplet transient species with absorptions at 390 and 450 nm (1) and at 370 nm (2), with φ(ISC) close to 1. Additionally, the calculated energies for the T(1) states relative to the S(0) states at UB3LYP/6-311++G** are ca. 47 kcal mol(-1) for 1 and 43 kcal mol(-1) for 2. A comparison of the geometrical parameters for the S(0) and T(1) states reveals a marked difference with respect to the arrangement of the exocyclic phenyl ring whilst a comparison of electronic parameters revealed the change from a quinone structure to a di-dehydroquinone diradical structure.     

Role of menaquinone in the reduction of fumarate, nitrate, iron(III) and manganese(IV) by Shewanella putrefaciens MR-1

Abstract Mutants of Shewanella putrefaciens MR-1 deficient in menaquinone and methylmenaquinone, but which have wild-type levels of ubiquinone, retain the ability to use trimethylamine N -oxide as an electron acceptor, but they lose the ability to use nitrate, iron(III), and fumarate as electron acceptors. These mutants also show a reduced rate of manganese(IV) reduction. One of these mutants could be restored to essentially wild-type phenotype by supplementing the medium with 1,4-dihydroxy-2-naphthoic acid. A requirement for naphthoquinones in iron(III) reduction and a preference for naphthoquinones in manganese(IV) reduction provide further support that the metal reducing systems in MR-1 are linked to anaerobic respiration.     

Investigation of chemical reactivity of 2-alkoxy-1,4-naphthoquinones and their anticancer activity

To establish the structure-activity relationship of 5-hydroxy-1,4-naphthoquinones toward anticancer activity, a series of its derivatives were prepared and tested for the activity (IC₅₀ in µM) against three cell lines; colo205 (colon adenocarcinoma), T47D (breast ductal carcinoma) and K562 (chronic myelogenous leukemia). Among them 2 (IC₅₀: 2.3; 2.0; 1.4?µM), 6 (IC₅₀: 1.9; 2.2; 1.3?µM), 9 (IC₅₀: 0.7; 1.7; 0.9?µM) and 10 (IC₅₀:1.7; 1.0; 1.2?µM) showed moderate to excellent activity. Our perception toward the DNA substitution of alkoxy groups at the C2 position of these naphthoquinones for the anticancer activity led us to investigate their reactivity of substitution toward dimethylamine as a nucleophile. The ease of the substitution of alkoxy groups at the C2 position with dimethylamine is strongly accelerated by hydroxyl group at C5 position and is well correlated with the found anticancer activity results.