Downregulation of indolethylamine N-methyltransferase is an early event in the rat hepatocarcinogenesis and is associated with poor prognosis in hepatocellular carcinoma patients.

Hepatocellular carcinoma (HCC) is one of the deadliest cancers worldwide, often preceded by cirrhosis and usually diagnosed at advanced stages; therefore, identifying molecular changes at early stages is an attractive strategy for detection and timely treatment. Here, we investigated the progressive transcriptomic changes during experimental hepatocarcinogenesis to identify novel early tumor markers in an HCC model induced by chronic administration of sublethal doses of diethylnitrosamine. An analysis of differentially expressed genes showed that four processes associated with oxidation-reduction and detoxification were significantly over-represented during hepatocarcinogenesis progression, of which the Nuclear Factor, Erythroid 2 Like 2 pathway showed several dysregulated genes. Interestingly, we also identified 91 genes dysregulated at early HCC stages, but the expression of the indolethylamine N-methyltransferase gene (INMT), as well as the level of its encoding protein, were strongly downregulated. INMT was increased in perivenular hepatocytes of normal livers but decreased in livers of experimental HCC. Furthermore, a gene expression and survival analysis performed using data from the liver hepatocellular carcinoma project of The Cancer Genome Atlas Program revealed that INMT is also significantly downregulated in human HCC and is associated with poor overall survival. In conclusion, by performing a transcriptome analysis of the HCC progression, we identified that INMT is early downregulated in the rat hepatocarcinogenesis and is associated with poor prognosis in human HCC, suggesting that INMT downregulation may be a promising prognostic marker for HCC in high-risk populations.

The auxin-producing Bacillus thuringiensis RZ2MS9 promotes the growth and modifies the root architecture of tomato (Solanum lycopersicum cv. Micro-Tom).

Strains of Bacillus thuringiensis (Bt) are commonly commercialized as bioinoculants for insect pest control, but their benefits go beyond their insecticidal property: they can act as plant growth-promoters. Auxins play a major role in the plant growth promotion. However, the mechanism of auxin production by the Bacilli group, and more specifically by Bt strains, is unclear. In previous work, the plant growth-promoting rhizobacterium (PGPR) B. thuringiensis strain RZ2MS9 increased the corn roots. This drew our attention to the strain's auxin production trait, earlier detected in vitro. Here, we demonstrate that in its genome, RZ2MS9 harbours the complete set of genes required in two pathways that are used for Indole acetic acid (IAA) production. We also detected that the strain produces almost five times more IAA during the stationary phase. The bacterial application increased the shoot dry weight of the Micro-Tom (MT) tomato by 24%. The application also modified MT root architecture, with an increase of 26% in the average lateral root length and inhibition of the axial root. At the cellular level, RZ2MS9-treated MT plants presented elongated root cortical cells with intensified mitotic activity. Altogether, these are the best characterized auxin-associated phenotypes. Besides that, no growth alteration was detected in the auxin-insensitive diageotropic (dgt) plants either with or without the RZ2MS9 inoculation. Our results suggest that auxins play an important role in the ability of B. thuringiensis RZ2MS9 to promote MT growth and provide a better understanding of the auxin production mechanism by a Bt strain.

FTIR and PLS-regression in the evaluation of bioactive amines, total phenolic compounds and antioxidant potential of dark chocolates.

Cloning techniques are used to improve agronomical traits and answer to the demand for fine chocolate. The objective of this study was to predict the concentrations of bioactive amines, phenolic compounds, and the antioxidant potential of dark monoclonal chocolate from nine fine cocoa varieties by FTIR analysis and conventional techniques. Total phenolic compounds, bioactive amines and antioxidant activity varied significantly among chocolates. The antioxidant activity was also affected by the analytical method (DPPH vs. Rancimat). Chemometric models based on FTIR data provided satisfactory predictions of the concentrations of the amines: spermidine (R2 = 0.92; RMSEP = 0.39; RMSEC = 0.21), tryptamine (R2 = 0.92; RMSEP = 0.41; RMSEC = 0.20), cadaverine (R2 = 0.82; RMSEP = 1.58; RMSEC = 0.75) and tyramine (R2 = 0.87; RMSEP = 1.87; RMSEC = 0.68); as well as phenolic compounds and antioxidant activity by Rancimat® (R2 = 0.98; RMSEP = 0.32; RMSEC = 0.21) and DPPH (R2 = 0.97; RMSEP = 4.05; RMSEC = 1.66). The wavenumbers of amines vibrations are among those that most affected antioxidant prediction models, confirming the contribution of amines to the antioxidant activity of chocolates.

Insecticidal activity of indole derivatives against Plutella xylostella and selectivity to four non-target organisms.

The diamondback moth Plutella xylostella (Linnaeus, 1758) (Lepidoptera: Plutellidae) is a destructive pest of brassica crops of economic importance that have resistance to a range of insecticides. Indole derivates can exert diverse biological activities, and different effects may be obtained from small differences in their molecular structures. Indole is the parent substance of a large number of synthetic and natural compounds, such as plant and animal hormones. In the present study, we evaluate the insecticidal activity of 20 new synthesized indole derivatives against P. xylostella, and the selectivity of these derivatives against non-target hymenopteran beneficial arthropods: the pollinator Apis mellifera (Linnaeus, 1758) (Hymenoptera: Apidae), and the predators Polybia scutellaris (White, 1841), Polybia sericea (Olivier, 1791) and Polybia rejecta (Fabricius, 1798) (Hymenoptera: Vespidae). Bioassays were performed in the laboratory to determine the lethal and sublethal effects of the compounds on P. xylostella and to examine their selectivity to non-target organisms by topical application and foliar contact. The treatments consisted of two synthesized derivatives (most and least toxic), the positive control (deltamethrin) and the negative control (solvent). The synthesized compound 4e [1-(1H-indol-3-yl)hexan-1-one] showed high toxicity (via topical application and ingestion) and decreased the leaf consumption by P. xylostella, displaying a higher efficiency than the pyrethroid deltamethrin, widely used to control this pest. In addition, the synthesized indole derivatives were selective to the pollinator A. mellifera and the predators P. scutellaris, P. sericea and P. rejecta, none of which were affected by deltamethrin. Our results highlight the promising potential of the synthesized indole derivatives for the generation of new chemical compounds for P. xylostella management.

Effect of orange juice and tryptamine on the behavior and c-fos expression of Wistar rats.

Recent reports have shown that commercial orange juice is rich in biogenic amines. Consumption of foods containing large amounts of biogenic amines increase hypertensive crisis and high levels of histamine and tyramine, which have been implicated as causative agents in a number of food poisoning episodes. In addition, accumulation of tryptamine in plasma may be associated with mood disorders. The aim of this study was to determine whether chronic administration of orange juice extract and tryptamine affects the behavior and c-fos expression in the rat. For this purpose, Wistar male rats were injected with saline solution, tryptamine or orange juice extract. Sucrose preference test and elevated plus maze were evaluated to determine hedonic and anxiety behavior, respectively. Rats treated with orange juice extract showed increased anxiety behavior and sucrose consumption, similar to those treated with tryptamine. In addition, dorsal raphe nucleus, accumbens nucleus, and hippocampus showed an increase of c-fos positive cells in rats treated with orange juice extract. In conclusion, the chronic and lengthy consumption of orange juice or their derivatives in the diet could be a factor responsible to induce mood disorders and may promote excess caloric consumption.

Quality assurance of histamine analysis in fresh and canned fish.

Histamine determination is relevant for fish safety, quality and trade. Recently a study by the European Union (EU) compared the Codex and the EU mandated methods for the analysis of histamine and observed that they underestimated and overestimated the results, respectively. To solve this problem, a simple and efficient procedure for the extraction and quantification of histamine by ion-pair HPLC method with post-column derivatization and fluorimetric detection is proposed. It was optimized and validated for the analysis of histamine in fish. The method attended the performance criteria established by Commission Decision 2002/657/CE. The method was also submitted to proficiency testing; uncertainty was calculated; and the stability of solutions and standards was investigated. There was no matrix effect. The LOD, LOQ, CCα and CCß were fit for the purpose. The method was successfully used in the analyses of freshwater fish and fresh and canned tuna.

Determination of tryptamine in foods using square wave adsorptive stripping voltammetry.

Tryptamine, a biogenic amine, is an indole derivative with an electrophilic substituent at the C3 position of the pyrrole ring of the indole moiety. The electrochemical oxidation of tryptamine was investigated using glassy carbon electrode (GCE), and focusing on trace level determination in food products by square wave adsorptive stripping voltammetry (SWAdSV). The electrochemical responses of tryptamine were evaluated using differing voltammetric techniques over a wide pH range, a quasi-reversible electron-transfer to redox system represented by coupled peaks P1-P3, and an irreversible reaction for peak P2 were demonstrated. The proton and electron counts associated with the oxidation reactions were estimated. The nature of the mass transfer process was predominantly diffusion-limited for the oxidation process of P1, the most selective and sensitive analytical response (acetate buffer solution pH 5.3), being used for the development of SWAdSV method, under optimum conditions. The excellent response allowed the development of an electroanalytical method with a linear response range of from 4.7-54.5)×10(-)(8)molL(-1), low detection limit (0.8×10(-)(9)molL(-)(1)), and quantification limit (2.7×10(-9)molL(-1)), and acceptable levels of repeatability (3.6%), and reproducibility (3.8%). Tryptamine content was determined in bananas, tomatoes, cheese (mozzarella and gorgonzola), and cold meats (chicken sausage and pepperoni sausage), yielding recoveries above 90%, with excellent analytical performance using simple and low cost instrumentation.

Bioactive amines in Passiflora are affected by species and fruit development.

Bioactive amines were determined in selected passion fruit species and throughout fruit development. The same amines (spermine, spermidine, agmatine, putrescine and tryptamine) were found in four Passiflora species (2008-2010 growing seasons) at different concentrations: P. alata had higher polyamines (spermine+spermidine, 8.41mg/100g); P. setacea and P. nitida had higher putrescine (>7.0mg/100g); and P. setacea had higher agmatine contents (1.37mg/100g) compared to the others. The indolamine tryptamine was present at low concentrations in all species (~0.05mg/100g). P. nitida and P. alata had the highest soluble solids (~18°Brix); P. edulis had the lowest pH (2.97) and P. nitida the highest pH (4.19). Throughout P. setacea fruit development, the concentrations of spermidine, putrescine and agmatine decreased; spermine contents did not change; and pH decreased. Fruit shelf life and some of the health promoting properties of Passiflora and their synthesis are modulated by species.

Bioactive amines in Mozzarella cheese from milk with varying somatic cell counts.

The influence of somatic cells counts (SCC) in milk on bioactive amines in Mozzarella cheese was investigated. High SCC milk had lower lactose and higher pH compared to low and medium SCC. Low spermine levels were found in milk irrespective of SCC. The cheeses had similar characteristics, but the extension and depth of proteolysis increased with SCC. Cheese from all SCC categories contained spermine; whereas tyramine and tryptamine were only detected in cheese from high SCC milk. During 60-days refrigerated storage, significant positive effects were observed between SCC and proteolysis, storage time and pH and storage time and proteolysis. There was a significant positive effect of storage time on spermine and serotonin levels. Only cheese from high SCC milk showed significantly higher serotonin levels. Tyramine and tryptamine were found in cheese from high SCC milk.

Bioactive amines in sorghum: method optimisation and influence of line, tannin and hydric stress.

The profile and levels of bioactive amines in different sorghum lines were reported for the first time. The amines were quantified by ion-pair HPLC, post-column derivatisation with o-phthalaldehyde and fluorimetric detection. The extraction procedure was optimised: 420 µm particle size, extraction with 5% trichloroacetic acid and three extractions. The screening of 22 sorghum lines showed that four of the ten amines investigated were detected. Spermine and spermidine were the prevalent amines (100%), followed by putrescine (77%) and cadaverine (14%). Total amines ranged from 5.8 to 41.4 mg/100 g, and the polyamines represented 60-100% of the total. Sorghum without tannin had higher amines levels compared to sorghum with tannin and cadaverine was specific to samples without tannin. Hydric stress caused accumulation of spermidine in the grains and affected the levels of other amines at rates depending on the presence or not of tannin. Sorghum is a significant source of polyamines.

Synthetic indole and melatonin derivatives exhibit antimalarial activity on the cell cycle of the human malaria parasite Plasmodium falciparum.

Discovering the mechanisms by which cell signaling controls the cell cycle of the human malaria parasite Plasmodium falciparum is fundamental to designing more effective antimalarials. To better understand the impacts of melatonin structure and function on the cell cycle of P. falciparum, we have synthesized two families of structurally-related melatonin compounds (7-11 and 12-16). All synthesized melatonin analogs were assayed in P. falciparum culture and their antimalarial activities were measured by flow cytometry. We have found that the chemical modification of the carboxamide group attached at C-3 position of the indole ring of melatonin (6) was crucial for the action of the indole-related compounds on the P. falciparum cell cycle. Among the melatonin derivatives, only the compounds 12, 13 and 14 were capable of inhibiting the P. falciparum growth in low micromolar IC50. These results open good perspectives for the development of new drugs with novel mechanisms of action.

Biosynthesis of N,N-dimethyltryptamine (DMT) in a melanoma cell line and its metabolization by peroxidases.

Tryptophan (TRP) is essential for many physiological processes, and its metabolism changes in some diseases such as infection and cancer. The most studied aspects of TRP metabolism are the kynurenine and serotonin pathways. A minor metabolic route, tryptamine and N,N-dimethyltryptamine (DMT) biosynthesis, has received far less attention, probably because of the very low amounts of these compounds detected only in some tissues, which has led them to be collectively considered as trace amines. In a previous study, we showed a metabolic interrelationship for TRP in melanoma cell lines. Here, we identified DMT and N,N-dimethyl-N-formyl-kynuramine (DMFK) in the supernatant of cultured SK-Mel-147 cells. Furthermore, when we added DMT to the cell culture, we found hydroxy-DMT (OH-DMT) and indole acetic acid (IAA) in the cell supernatant at 24 h. We found that SK-Mel-147 cells expressed mRNA for myeloperoxidase (MPO) and also had peroxidase activity. We further found that DMT oxidation was catalyzed by peroxidases. DMT oxidation by horseradish peroxidase, H2O2 and MPO from PMA-activated neutrophils produced DMFK, N,N-dimethyl-kynuramine (DMK) and OH-DMT. Oxidation of DMT by peroxidases apparently uses the common peroxidase cycle involving the native enzyme, compound I and compound II. In conclusion, this study describes a possible alternative metabolic pathway for DMT involving peroxidases that has not previously been described in humans and identifies DMT and metabolites in a melanoma cell line. The extension of these findings to other cell types and the biological effects of DMT and its metabolites on cell proliferation and function are key questions for future studies.

Microorganisms as tools in modern chemistry: the biotransformation of 3-indolylacetonitrile and tryptamine by fungi

Chemical reactions performed by microorganisms have been used as a modern tool in chemistry. In this work, the ability of the fungi Beauveria bassiana and Aspergillus niger to modify the chemical structure of indole compounds was studied. Beauveria bassiana was able to transform 3-indolylacetonitrile into 3-methylindole while Aspergillus niger transformed tryptamine into 5-hydroxyindole-3-acetamide. Interestingly, these fungi were able to perform both reduction and oxidation of the indole compounds fed, the oxidation occurring with improved levels of oxygen uptake. The results prove once more the versatility of microorganisms to perform a wide range of industrially attractive chemical reactions.

O uso de microrganismos para a realização de reações químicas tem sido explorado como uma ferramenta moderna em química. Neste trabalho explorou-se a habilidade dos fungos Beauveria bassiana e Aspergillus niger de modificarem quimicamente a estrutura de compostos indólicos, uma classe de substâncias com diversas atividades biológicas relatadas. A 3-indolacetonitrila foi reduzida, com formação do 3-metilindol por Beauveria bassiana, enquanto Aspergillus niger converteu a triptamina em um derivado oxidado, a 5-hidroxiindol-3-acetamida. A oxidação ocorreu somente no experimento sob agitação e provavelmente está relacionada ao alto grau de oxigenação da reação. A ocorrência de redução e oxidação de derivados indólicos ilustra bem a versatilidade do uso de microorganismos para a condução de uma grande variedade de reações químicas de interesse industrial.