Efeito do processo de descafeinação de café arábica em meio aquoso auxiliado por ultrassom sobre a composição química e o perfil sensorial

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Avaliação do perfil químico de fenólicos, do potencial antioxidante e fotoprotetor da torta de semente de Coffea arabica L. (Rubiaceae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

A new HPLC-DAD-MS/MS method for the simultaneous determination of major compounds in the crude extract of Lychnophora salicifolia Mart. (Brazilian arnicao) leaves: Application to chemical variability evaluation

Lychnophora salicifolia Mart., which occurs in the Brazilian Cerrado in the states of Bahia and Minas Gerais as well as in the southeast of the state of Goias, is the most widely distributed and also the most polymorphic species of the genus. This plant is popularly known to have anti-inflammatory and analgesic activities. In this work, we have studied the variation in terms of polar metabolites of ninety-three Lychnophora salicifolia Mart, specimens collected from different regions of the Brazilian Cerrado. Identification of the constituents of this mixture was carried out by analysis of the UV spectra and MS data after chromatographic separation. Twenty substances were identified, including chlorogenic acid derivatives, a flavonoid C-glucoside, and other sesquiterpenes. The analytical method was validated, and the reliability and credibility of the results was ensured for the purposes of this study. The concentration range required for analysis of content variability within the analyzed group of specimens was covered with appropriate values of limits of detection and quantitation, as well as satisfactory precision and recovery. A quantitative variability was observed among specimens collected from the same location, but on average they were similar from a chemical viewpoint. In relation to the study involving specimens from different locations, there were both qualitative and quantitative differences among plants collected from different regions of Brazil. Statistical analysis revealed that there is a correlation between geographical localization and polar metabolites profile for specimens collected from different locations. This is evidence that the pattern of metabolites concentration depends on the geographical distribution of the specimens. (C) 2012 Elsevier B.V. All rights reserved.

Medium Light and Medium Roast Paper-Filtered Coffee Increased Antioxidant Capacity in Healthy Volunteers: Results of a Randomized Trial

We compared the effects of medium light roast (MLR) and medium roast (MR) paper-filtered coffee on antioxidant capacity and lipid peroxidation in healthy volunteers. In a randomized crossover study, 20 volunteers consumed 482 +/- 61 ml/day of MLR or MR for four weeks. Plasma total antioxidant status (TAS), oxygen radical absorbance capacity (ORAC), oxidized LDL and 8-epi-prostaglandin F2 alpha, erythrocyte superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) activity were measured at baseline and after the interventions. MLR had higher chlorogenic acids-(CGA; 334 mg/150 mL) and less caffeine (231 mg/150 ml) than MR had (210 and 244 mg/150 ml, respectively). MLR also had fewer Maillard reaction products (MRP) than MR had. Compared with baseline, subjects had an increase of 21 and 26 % in TAS, 13 and 13 % in CAT, 52 and 75 % in SOD, and 62 and 49 % in GPx after MLR and MR consumption (P < 0.001), respectively. ORAC increased after MLR (P = 0.004). No significant alteration in lipid peroxidation biomarkers was observed. Both coffees had antioxidant effects. Although MLR contained more CGA, there were similar antioxidant effects between the treatments. MRP may have contributed as an antioxidant. These effects may be important in protecting biological systems and reducing the risk of diseases related to oxidative stress.

SEASONAL VARIATION OF THE MAJOR SECONDARY METABOLITES PRESENT IN THE EXTRACT OF Eremanthus mattogrossensis Less (Asteraceae: Vernonieae) LEAVES

The species Eremanthus mattogrossensis, known as "veludo do cerrado" (cerrado velvet), is native to the Brazilian Cerrado. Because the amount of metabolites present in plants may be influenced by biological and environmental factors, here we conducted an HPLC-DAD-MS/MS investigation of the metabolite concentrations found in the MeOH/H2O extract of the leaves of this species. The main compounds were identified and quantified, and the metabolites were grouped by chemical class (caffeoylquinic acids, flavonoids, and sesquiterpene lactone). Statistical analysis indicated a straight correlation between the quantity of metabolites and seasonality, suggesting that environmental properties elicit important metabolic responses.

Topical anti-inflammatory activity of yacon leaf extracts

Smallanthus sonchifolius (Poepp.) H. Rob. , Asteraceae, known as yacon, is an herb that is traditionally used for the treatment of diabetes in folk medicine. However, recent studies have demonstrated that this plant has other interesting properties such as anti-microbial and anti-inflammatory actions. Thus, the purpose of this study was to evaluate the topical anti-inflammatory property of different extracts prepared from yacon leaves and analyze the role of different chemical classes in this activity. Three yacon leaf extracts were obtained: aqueous extract, where chlorogenic acid derivatives and sesquiterpene lactones were detected; leaf rinse extract, rich in sesquiterpene lactones; and polar extract, rich in chlorogenic acid derivatives. All the extracts exhibited anti-edematogenic activity in vivo (aqueous extract: 25.9% edema inhibition at 0.50 mg/ear; polar extract: 42.7% inhibition at 0.25 mg/ear; and leaf rinse extract: 44.1% inhibition at 0.25 mg/ear). The leaf rinse extract furnished the best results regarding neutrophil migration inhibition, and NO, TNF-? and PGE2 inhibition. These data indicate that both sesquiterpene lactones and chlorogenic acid derivatives contribute to the anti-inflammatory action, although sesquiterpene lactones seem to have more pronounced effects. In conclusion, yacon leaf extracts, particularly the sesquiterpene lactone-rich extract, has potential use as topical anti-inflammatory agent.

Effetti del chitosano su composti polifenolici in colture cellulari di vite: aspetti molecolari e metabolici

Polyphenols, including flavonoids and stilbenes, are an essential part of human diet and constitute one of the most abundant and ubiquitous group of plant secondary metabolites. The level of these compounds is inducible by stress or fungal attack, so attempts are being made to identify likely biotic and abiotic elicitors and to better understand the underlying mechanism. Resveratrol (3,5,4’-trihydroxystilbene), which belongs to the stilbene family, is a naturally occurring polyphenol, found in several fruits, vegetables and beverages including red wine. It is one of the most important plant polyphenols with proved benefic activity on animal health. In the last two decades, the potential protective effects of resveratrol against cardiovascular and neurodegenerative diseases, as well as the chemopreventive properties against cancer, have been largely investigated. The most important source of polyphenols and in particular resveratrol for human diet is grape (Vitis vinifera). Since stilbenes and flavonoids play a very important role in plant defence responses and enviromental interactions, and their effects on human health seem promising, the aim of the research of this Thesis was to study at different levels the activation and the regulation of their biosynthetic pathways after chitosan treatment. Moreover, the polyphenol production in grape cells and the optimisation of cultural conditions bioreactor scale-up, were also investigated. Cell suspensions were obtained from cv. Barbera (Vitis vinifera L.) petioles and were treated with a biotic elicitor, chitosan (50 μg/mL, dissolved in acetic acid) to promote phenylpropanoid metabolism. Chitosan is a D-glucosamine polymer from fungi cell wall and therefore mimes fungal pathogen attack. Liquid cultures have been monitored for 15 days, measuring cell number, cell viability, pH and grams of fresh weight. The endogenous and released amounts of 7 stilbenes (trans and cis isomers of resveratrol, piceid and resveratroloside, and piceatannol), gallic acid, 6 hydroxycinnamic acids (trans-cinnamic, p-coumaric, caffeic, ferulic, sinapic and chlorogenic acids), 5 catechines (catechin, epicatechin, epigallocatechin-gallate (EGCG), epigallocatechin and epicatechin-gallate) and other 5 flavonoids (chalcon, naringenin, kaempferol, quercetin and rutin) in cells and cultural medium, were measured by HPLC-DAD analysis and total anthocyanins were quantified by spectrophotometric analysis. Chitosan was effective in stimulating trans-resveratrol endogenous accumulation with a sharp peak at day 4 (exceeding acetic acid and water controls by 36% and 63%, respectively), while it did not influence the production of the cis-isomer. Compared to both water and acetic acid controls, chitosan decreased the release of both trans- and cis-resveratrol respect to controls. No effect was shown on the accumulation of single resveratrol mono-glucoside isomers, but considering their total amount, normalized for the relative water control, it was possible to evidence an increase in both accumulation and release of those compounds, in chitosan-treated cells, throughout the culture period and particularly during the second week. Many of the analysed flavonoids and hydroxycinnamic acids were not present or detectable in trace amounts. Catechin, epicatechin and epigallocatechin-gallate (EGCG) were detectable both inside the cells and in the culture media, but chitosan did not affect their amounts. On the contrary, total anthocyanins have been stimulated by chitosan and their level, from day 4 to 14, was about 2-fold higher than in both controls, confirming macroscopic observations that treated suspensions showed an intense brown-red color, from day 3 onwards. These elicitation results suggest that chitosan selectively up-regulates specific biosynthetic pathways, without modifying the general accumulation pattern of other flavonoids. Proteins have been extracted from cells at day 4 of culture (corresponding to the production peak of trans-resveratrol) and separated by bidimensional electrophoresis. The 73 proteins that showed a consistently changed amount between untreated, chitosan and acetic acid (chitosan solvent) treated cells, have been identified by mass spectrometry. Chitosan induced an increase in stilbene synthase (STS, the resveratrol biosynthetic enzyme), chalcone-flavanone isomerase (CHI, that switches the pathway from chalcones to flavones and anthocyanins), pathogenesis-related proteins 10 (PRs10, a large family of defence proteins), and a decrease in many proteins belonging to primary metabolisms. A train of six distinct spots of STS encoded by the same gene and increased by chitosan, was detected on the 2-D gels, and related to the different phosphorylation degree of STS spots. Northern blot analyses have been performed on RNA extracted from cells treated with chitosan and relative controls, using probes for STS, PAL (phenylalanine ammonia lyase, the first enzyme of the biosynthetic pathway), CHS (chalcone synthase, that shares with STS the same precursors), CHI and PR-10. The up-regulation of PAL, CHS and CHI transcript expression levels correlated with the accumulation of anthocyanins. The strong increase of different molecular weight PR-10 mRNAs, correlated with the 11 PR-10 protein spots identified in proteomic analyses. The sudden decrease in trans-resveratrol endogenous accumulation after day 4 of culture, could be simply explained by the diminished resveratrol biosynthetic activity due to the lower amount of biosynthetic enzymes. This might be indirectly demonstrated by northern blot expression analyses, that showed lower levels of phenylalanine ammonia lyase (PAL) and stilbene synthase (STS) mRNAs starting from day 4. Other possible explanations could be a resveratrol oxidation process and/or the formation of other different mono-, di-glucosides and resveratrol oligomers such as viniferins. Immunolocalisation experiments performed on grape protoplasts and the subsequent analyses by confocal microscope, showed that STS, and therefore the resveratrol synthetic site, is mostly associated to intracellular membranes close to the cytosolic side of plasma membrane and in a smaller amount is localized in the cytosol. STS seemed not to be present inside vacuole and nucleus. There were no differences in the STS intracellular localisation between the different treatments. Since it was shown that stilbenes are largely released in the culture medium and that STS is a soluble protein, a possible interaction of STS with a plasma membrane transporter responsible for the extrusion of stilbenes in the culture medium, might be hypothesized. Proteomic analyses performed on subcellular fractions identified in the microsomial fraction 5 proteins taking part in channel complexes or associated with channels, that significantly changed their amount after chitosan treatment. In soluble and membrane fractions respectively 3 and 4 STS and 6 and 3 PR-10 have been identified. Proteomic results obtained from subcellular fractions substantially confirmed previous result obtained from total cell protein extracts and added more information about protein localisation and co-localisation. The interesting results obtained on Barbera cell cultures with the aim to increase polyphenol (especially stilbenes) production, have encouraged scale up tests in 1 litre bioreactors. The first trial fermentation was performed in parallel with a normal time-course in 20 mL flasks, showing that the scale-up (bigger volume and different conditions) process influenced in a very relevant way stilbenes production. In order to optimise culture parameters such as medium sucrose amount, fermentation length and inoculum cell concentration, few other fermentations were performed. Chitosan treatments were also performed. The modification of each parameter brought relevant variations in stilbenes and catechins levels, so that the production of a certain compound (or class of compounds) could be hypothetically promoted by modulating one or more culture parameters. For example the catechin yield could be improved by increasing sucrose content and the time of fermentation. The best results in stilbene yield were obtained in a 800 mL fermentation inoculated with 10.8 grams of cells and supplemented with chitosan. The culture was fed with MS medium added with 30 g/L sucrose, 25 μg/mL rifampicin and 50 μg/mL of chitosan, and was maintained at 24°C, stirred by marine impeller at 100 rpm and supplied of air at 0.16 L/min rate. Resveratroloside was the stilbene present in the larger amount, 3-5 times more than resveratrol. Because resveratrol glucosides are similarly active and more stable than free resveratrol, their production using a bioreactor could be a great advantage in an hypothetical industrial process. In my bioreactor tests, stilbenes were mainly released in the culture medium (60-80% of the total) and this fact could be another advantage for industrial applications, because it allows recovering the products directly from the culture medium without stopping the fermentation and/or killing the cells. In my best cultural conditions, it was possible to obtain 3.95 mg/L of stilbenes at day 4 (maximum resveratrol accumulation) and 5.13 mg/L at day 14 (maximum resveratroloside production). In conclusion, chitosan effect in inducing Vitis vinifera defense mechanisms can be related to its ability to increase the intracellular content of a large spectrum of antioxidants, and in particular of resveratrol, its derivates and anthocyanins. Its effect can be observed at transcriptional, proteomic (variation of soluble and membrane protein amounts) and metabolic (polyphenols production) level. The chitosan ability to elicit specific plant matabolisms can be useful to produce large quantities of antioxidant compounds from cell culture in bioreactor.

Minimização de resíduos do processamento do café solúvel através do reaproveitamento da borra para extração de óleo utilizando solvente renovável

O principal resíduo da indústria de café solúvel é a borra de café, gerada após a extração de sólidos solúveis com água, sendo usualmente queimada em caldeiras para geração de energia para a própria indústria. Entretanto, este co-produto pode conter de 15 a 20 % de óleo, componente de grande interesse na indústria alimentícia. Em paralelo, na indústria de produção de óleos vegetais o solvente frequentemente utilizado é o hexano. Contudo, por este ser um derivado de combustíveis fósseis, alternativas para sua substituição por solventes mais amigáveis ao meio ambiente, que podem ser obtidos por via biotecnológica, estão sendo estudadas. Deste modo, o objetivo principal desta dissertação de mestrado foi a viabilização técnica do emprego de solventes alcoólicos no processo de extração de óleo de borra de café proveniente da indústria de processamento de café solúvel. Foram realizadas extrações sólido-líquido em um estágio para estudar a influência das variáveis de processo temperatura (60 a 90 °C), tipo de solvente (etanol, ET ou isopropanol, IPA) e a hidratação do solvente (absoluto ou azeotrópico) nas características das fases extrato e rafinado, em termos de rendimento de extração de óleo, de ácidos clorogênicos (ACG), de carboidratos totais, teor de proteínas e índice de solubilidade de nitrogênio (ISN) da fase rafinado. Pré-tratamento enzimático ao processo de extração também foi realizado para investigar a sua atuação sobre o rendimento de extração de óleo e ACG, além do ISN das fases rafinado obtidas na temperatura de 70 °C. De forma geral, pode-se inferir que a temperatura favorece a extração de compostos lipídicos, mas a hidratação do solvente prejudica a extração destes compostos pelo aumento da polaridade do solvente. Do mesmo modo, como o ET é mais polar que o IPA, o primeiro solvente proporcionou a obtenção de menores rendimentos de extração de óleo. A temperatura de processo também influenciou a extração de ACG, a qual foi beneficiada a temperaturas mais baixas pelo aumento da polaridade dos solventes utilizados. De tal forma, que a 60 °C, nos experimentos utilizando etanol azeotrópico obteve-se os menores rendimentos de extração de óleo, porém maior rendimento de extração de ACG. O pré-tratamento enzimático apresentou diferenças significativas nas características das fases extrato e rafinado. No entanto, somente os experimentos com etanol absoluto resultaram em rendimentos de extração de óleo economicamente viáveis. De fato, será relevante um estudo mais aprofundado das variáveis de pré-tratamento enzimático para obter resultados mais expressivos em relação à extração de compostos lipídicos. Diante dos dados experimentais obtidos, conclui-se que é tecnicamente viável o emprego de solventes alcoólicos no processo de extração de óleo de borra de café. Entretanto, nota-se que as condições de processo devem ser avaliadas minuciosamente com o objetivo de se obter altos rendimentos de extração de óleo com maior teor de ACG.

Binding of polyphenols to plant cell wall analogues - Part 2: Phenolic acids

Bacterial cellulose and cellulose-pectin composites were used as well-defined model plant cell wall (PCW) systems to study the interaction between phenolic acids (PA) derived from purple carrot juice concentrate (PCJC) and PCW components. Significant PA depletion from solution occurred, with pure cellulose initially (30 s-1 h) absorbing more than cellulose-pectin composites in the first hour (ca 20% cf 10-15%), but with all composites absorbing similar levels (ca 30%) after several days. Individual PAs bound to different relative extents with caffeic acid > chlorogenic acid > ferulic acid. Extrapolation of data for these model systems to carrot puree suggests that nutritionally-significant amounts of PAs could bind to cell walls, potentially restricting bioavailability in the small intestine and, as a consequence, delivering PAs to the large intestine for fermentation and metabolism by gut bacteria. (C) 2012 Elsevier Ltd. All rights reserved.

Isolation and characterization of human colonic bacteria able to hydrolyse chlorogenic acid

Free hydroxycinnamates, including caffeic, ferulic and p-coumaric acids, exhibit antioxidant and anticarcinogenic properties both in vitro and in animal models. Given that the gut flora has a major role in human nutrition and health, some of the beneficial effects of phenolic acids may be ascribed to the microflora involved in metabolism.

Optimization of a method for determination of phenolic acids in exotic fruits by capillary electrophoresis

In this work, the separation of nine phenolic acids (benzoic, caffeic, chlorogenic, p-coumaric, ferulic, gallic, protocatechuic, syringic, and vanillic acid) was approached by a 32 factorial design in electrolytes consisting of sodium tetraborate buffer(STB) in the concentration range of 10-50 mmol L(-1) and methanol in the volume percentage of 5-20%. Derringer`s desirability functions combined globally were tested as response functions. An optimal electrolyte composed by 50 mmol L(-1) tetraborate buffer at pH 9.2, and 7.5% (v/v) methanol allowed baseline resolution of all phenolic acids under investigation in less than 15 min. In order to promote sample clean up, to preconcentrate the phenolic fraction and to release esterified phenolic acids from the fruit matrix, elaborate liquid-liquid extraction procedures followed by alkaline hydrolysis were performed. The proposed methodology was fully validated (linearity from 10.0 to 100 mu g mL(-1), R(2) > 0.999: LOD and LOQ from 1.32 to 3.80 mu g mL(-1) and from 4.01 to 11.5 mu g mL(-1), respectively; intra-day precision better than 2.8% CV for migration time and 5.4% CV for peak area; inter-day precision better than 4.8% CV for migration time and 4.8-11% CV for peak area: recoveries from 81% to 115%) and applied successfully to the evaluation of phenolic contents of abiu-roxo (Chrysophyllum caimito), wild mulberry growing in Brazil (Morus nigra L.) and tree tomato (Cyphomandra betacea). Values in the range of 1.50-47.3 mu g g(-1) were found, with smaller amounts occurring as free phenolic acids. (C) 2009 Elsevier B.V. All rights reserved.

Phenolic acids and abscisic acid in Australian Eucalyptus honeys and their potential for floral authentication

Seven phenolic acids related to the botanical origins of nine monofloral Eucalyptus honeys from Australia, along with two abscisic isomers, have been analyzed. The mean content of total phenolic acids ranges from 2.14 mg/100 g honey of black box (Eucalyptus largiflorens) honey to 10.3 mg/100 g honey of bloodwood (Eucalyptus intermedia) honey, confirming an early finding that species-specific differences of phytochemical compositions occur quantitatively among these Eucalyptus honeys. A common profile of phenolic acids, comprising gallic, chlorogenic, coumaric and caffeic acids, can be found in all the Eucalyptus honeys, which could be floral markers for Australian Eucalyptus honeys. Thus, the analysis of phenolic acids could also be used as an objective method for the authentication of botanical origin of Eucalyptus honeys. Moreover, all the honey samples analyzed in this study contain gallic acid as the main phenolic acid, except for stringybox (Eucalyptus globoidia) honey which has ellagic acid as the main phenolic acid. This result indicates that the species-specific differences can also be found in the honey profiles of phenolic acids. Further-more, the analysis of abscisic acid in honey shows that the content of abscisic acid varies from 0.55 mg/100 g honey of black box honey to 4.68 mg/ 100 g honey of bloodwood honey, corresponding to the contents of phenolic acids measured in these honeys. These results have further revealed that the HPLC analysis of honey phytochemical constituents could be used individually and/or jointly for the authentication of the botanical origins of Australian Eucalyptus honeys. (C) 2003 Elsevier Ltd. All rights reserved.

Phenolic acids in Australian Melaleuca, Guioa, Lophostemon, Banksia and Helianthus honeys and their potential for floral authentication

Eight phenolic acids and two abscisic acid isomers in Australian honeys from five botanical species (Melaleuca, Guioa, Lophostemon, Banksia and Helianthus) have been analyzed in relation to their botanical origins. Total phenolic acids present in these honeys range from 2.13 mg/100 g sunflower (Helianthus annuus) honey to 12.11 mg/100 g tea tree (Melaleuca quinquenervia) honey, with amounts of individual acids being various. Tea tree honey shows a phenolic profile of gallic, ellagic, chlorogenic and coumaric acids, which is similar to the phenolic profile of an Australian Eucalyptus honey (bloodwood or Eucalyptus intermedia honey). The main difference between tea tree and bloodwood honeys is the contribution of chlorogenic acid to their total phenolic profiles. In Australian crow ash (Guioa semiglauca) honey, a characteristic phenolic profile mainly consisting of gallic acid and abscisic acid could be used as the floral marker. In brush box (Lophostemon conferta) honey, the phenolic profile, comprising mainly gallic acid and ellagic acid, could be used to differentiate this honey not only from the other Australian non-Eucalyptus honeys but also from a Eucalyptus honey (yellow box or Eucalyptus melliodora honey). However, this Eucalyptus honey could not be differentiated from brush box honey based only on their flavonoid profiles. Similarly, the phenolic profile of heath (Banksia ericifolia) honey, comprising mainly gallic acid, an unknown phenolic acid (Phl) and coumaric acid, could also be used to differentiate this honey from tea tree and bloodwood honeys, which have similar flavonoid profiles. Coumaric acid is a principal phenolic acid in Australian sunflower honey and it could thus be used together with gallic acid for the authentication. These results show that the HPLC analysis of phenolic acids and abscisic acids in Australian floral honeys Could assist the differentiation and authentication of the honeys. © 2005 Elsevier Ltd. All rights reserved.

Proton-transfer versus nontransfer in compounds of the diazo-dye precursor 4-(phenyldiazenyl) aniline (aniline yellow) with strong organic acids: the 5-sulfosalicylate and the dichroic benzenesulfonate salts, and the 1:2 adduct with 3,5-dinitrobenzoic

The structures of two 1:1 proton-transfer red-black dye compounds formed by reaction of aniline yellow [4-(phenyldiazenyl)aniline] with 5-sulfosalicylic acid and benzenesulfonic acid, and a 1:2 nontransfer adduct compound with 3,5-dinitrobenzoic acid have been determined at either 130 or 200 K. The compounds are 2-(4-aminophenyl)-1-phenylhydrazin-1-ium 3-carboxy-4-hydroxybenzenesulfonate methanol solvate, C12H12N3+.C7H5O6S-.CH3OH (I), 2-(4-aminophenyl)-1-hydrazin-1-ium 4-(phenydiazinyl)anilinium bis(benzenesulfonate), 2C12H12N3+.2C6H5O3S-, (II) and 4-(phenyldiazenyl)aniline-3,5-dinitrobenzoic acid (1/2) C12H11N3.2C~7~H~4~N~2~O~6~, (III). In compound (I) the diaxenyl rather than the aniline group of aniline yellow is protonated and this group subsequently akes part in a primary hydrogen-bonding interaction with a sulfonate O-atom acceptor, producing overall a three-dimensional framework structure. A feature of the hydrogen bonding in (I) is a peripheral edge-on cation-anion association involving aromatic C--H...O hydrogen bonds, giving a conjoint R1/2(6)R1/2(7)R2/1(4)motif. In the dichroic crystals of (II), one of the two aniline yellow species in the asymmetric unit is diazenyl-group protonated while in the other the aniline group is protonated. Both of these groups form hydrogen bonds with sulfonate O-atom acceptors and thee, together with other associations give a one-dimensional chain structure. In compound (III), rather than proton-transfer, there is a preferential formation of a classic R2/2(8) cyclic head-to-head hydrogen-bonded carboxylic acid homodimer between the two 3,5-dinitrobenzoic acid molecules, which in association with the aniline yellow molecule that is disordered across a crystallographic inversion centre, result in an overall two-dimensional ribbon structure. This work has shown the correlation between structure and observed colour in crystalline aniline yellow compounds, illustrated graphically in the dichroic benzenesulfonate compound.