Red Tomato Products as an Alternative to Reduce Synthetic Dyes in the Food Industry: A Review.

Most dyes used in the food industry are synthetic and can be a health hazard. Red tomato may serve as a natural alternative dye to replace synthetic colorants. This study aimed to review the literature on the addition of red tomato products (powder tomato, paste, freeze-dried, tomato peel powder, tomato pomace) to reduce the usage of synthetic dyes in the food industry. Red tomato products have been used as coloring in pasta, bologna, sausages, cookies, crackers, macaroons, hamburgers, breads, muffins, cheeses, and nuggets. The trans-cis isomerization of lycopene by oxidative processes directly affects the color of the pigment. The lycopene contained in tomato has antioxidant activity and could reduce or eliminate other oxidants and/or synthetic preservatives in food. Moreover, tomatoes in foods have high sensory scores, nutritional appeal, and marketing potential. However, its use as a food colorant has been not extensively explored. Therefore, further studies are still required, especially on the stability of carotenoids in tomatoes used in processed foods.

Effects on DNA repair in human lymphocytes exposed to the food dye tartrazine yellow.

Tartrazine is a food additive that belongs to a class of artificial dyes and contains an azo group. Studies about its genotoxic, cytotoxic and mutagenic effects are controversial and, in some cases, unsatisfactory. This work evaluated the potential in vitro cytotoxicity, genotoxicity and effects on DNA repair of human lymphocytes exposed to the dye. We assessed the cytotoxicity of tartrazine by 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide test and the response of DNA repair through comet assay (alkaline version). We used different concentrations of the dye, ranging from 0.25-64.0 mM. The results demonstrated that tartrazine has no cytotoxic effects. However, this dye had a significant genotoxic effect at all concentrations tested. Although most of the damage was amenable to repair, some damage remained higher than positive control after 24 h of repair. These data demonstrate that tartrazine may be harmful to health and its prolonged use could trigger carcinogenesis.

CYP-450 isoenzymes catalyze the generation of hazardous aromatic amines after reaction with the azo dye Sudan III.

This work describes the mutagenic response of Sudan III, an adulterant food dye, using Salmonella typhimurium assay and the generation of hazardous aromatic amines after different oxidation methods of this azo dye. For that, we used metabolic activation by S9, catalytic oxidation by ironporphyrin and electrochemistry oxidation in order to simulate endogenous oxidation conditions. The oxidation reactions promoted discoloration from 65% to 95% of Sudan III at 1 × 10(-4)molL(-1) and generation of 7.6 × 10(-7)molL(-1) to 0.31 × 10(-4)molL(-1) of aniline, o-anisidine, 2-methoxi-5-methylaniline, 4-aminobiphenyl, 4,4'-oxydianiline; 4,4'-diaminodiphenylmethane and 2,6-dimethylaniline. The results were confirmed by LC-MS-MS experiments. We also correlate the mutagenic effects of Sudan III using S. typhimurium with the strain TA1535 in the presence of exogenous metabolic activation (S9) with the metabolization products of this compound. Our findings clearly indicate that aromatic amines are formed due to oxidative reactions that can be promoted by hepatic cells, after the ingestion of Sudan III. Considering that, the use of azo compounds as food dyestuffs should be carefully controlled.

Un nuevo aporte a la toxicología de colorantes alimentarios: conjugación hepática de la indigotina con fosfolípidos / A new approach to the toxicology of food colorants: hepatic conjugation of indigotin with phospholipids

Demostramos anteriormente que la solubilidad del colorante alimentario indigotina se modifica en el hígado de mamíferos, volviéndose totalmente insoluble en solventes acuosos y altamente soluble en solventes orgánicos. En el presente trabajo se estudió la movilidad cromatográfica de la indigotina presente en un extracto lipídico de hígado de rata, 20 minutos después de su administración intravenosa. Se estudió además la presencia de grupos funcionales correspondientes a su probable conjugación con fosfolípidos, en el extracto preparado al raspar la banda del colorante separada mediante cromatografía de fosfolípidos en capa delgada. Se detectaron 0,144 +/- 0,087 µg de fósforo fosfolipídico y se demostró la presencia de ácidos grasos en dicha fracción, proveniente de un extracto preparado a partir de 2 gramos de hígado. Estos resultados confirman el aumento en la solubilidad del colorante en solventes orgánicos una vez que ha ingresado al hígado y sugieren que el mismo sufre una probable conjugación con fosfolípidos en dicho órgano.

We have demonstrated that the solubility of the food colorant indigotin was modified in mammalian liver, where it becomes totally insoluble in aqueous solvents and highly soluble in organic ones. This work presents the results from the studies on the chromatographic mobility of indigotin present in the lipidic extract obtained from rat liver 20 minutes after its intravenously administration. The presence of functional groups corresponding to its probable conjugation with phospholipids was also studied in the extract prepared from the colorant spot separated by thin layer chromatography of phospholipids. 0,144 +/- 0,087 _g of phospholipidic phosphorus were detected and fatty acid presence was demosntrated in this fraction, obtained from liver extract prepared with 2 grams of tissue. These results confirm the increment of the colorant solubility in organic solvents once it is incorporated in the liver and they suggest that it suffers a probably conjugation with phospholipids in this organ.

Absence of carcinogenic and anticarcinogenic effects of annatto in the rat liver medium-term assay.

Annatto (Bixa orellana L.) is a natural food colorant extensively used in many processed foods, especially dairy products. The lower cost of production and the low toxicity, make annatto a very attractive and convenient pigment in substitution to the many synthetic colorants. In the present study we investigate the carcinogenic and anticarcinogenic effects of dietary annatto in Wistar rat liver using the preneoplastic glutathione S-transferase (GST-P) foci and DNA damage biomarkers. Annatto, containing 5% bixin, was administered in the diet at concentrations of 20, 200, and 1000 ppm (0.07; 0.80 and 4.23 bixin/kg body wt/day, respectively), continuously during 2 weeks before, or 8 weeks after DEN treatment (200 mg/kg body wt, i.p.), to evaluate its effect on the liver-carcinogenesis medium-term bioassay. The comet assay was used to investigate the modifying potential of annatto on DEN (20 mg/kg body wt)-induced DNA damage. The results showed that annatto was neither genotoxic nor carcinogenic at the highest concentration tested (1000 ppm). No protective effects were also observed in both GST-P foci development and comet assays. In conclusion, in such experimental conditions, annatto shows no hepatocarcinogenic effect or modifying potential against DEN-induced DNA damage and preneoplastic foci in the rat liver.

Subacute toxicity assessment of annatto in rat.

Increased human use of annatto (Bixa orellana L), a red yellow food colorant, demands generation of toxicity data. The toxic effects of annatto powder (bixin 27%) have been assessed following administration of a subacute regimen (4 weeks, 20 doses) in Wistar male and female rats. A full study with three dose levels was considered unnecessary since no sign of toxicity had been noted in a preliminary experiment with 1000 mg/kg body weight/day as was recommended by the OECD guideline. In this study, annatto administered by gavage at a dose level of 2000 mg/kg/day decreased male body weight gain, but had no effect on either food intake or food conversion efficiency. Haematological and plasma biochemical examination as well necropsy performed at the end of administration (29th day) and observation (43rd day) periods revealed no alterations related with annatto administration. Kidney apoptosis occurred in 20% treated female rats in restricted areas without proliferation or tubular segments modification. The precise nature of apoptosis was not investigated in the present study. These findings suggest that annatto was no toxic to the rat.

Study on the mutagenicity and antimutagenicity of a natural food colour (annatto) in mouse bone marrow cells.

Most manufactured foods contain chemicals added as a deliberate part of the manufacturing process. The aims of the present study were to evaluate the mutagenicity and antimutagenicity of annatto, a natural pigment extracted from the Bixa orellana L. and widely used as a colorant in foods. The micronucleus test was performed in bone marrow cells from Swiss male mice treated with one of the three concentrations of annatto (1330, 5330 and 10,670 ppm), incorporated into the diet. The animals were fed with the diets for 7 days and sacrificed 24 h after the last treatment. For the evaluation of the antimutagenic potential of annatto, at day 7, the animals received an intraperitoneal injection of cyclophosphamide (50 mg/kg body weight). Under the concentrations tested annatto did not present mutagenic or antimutagenic activities on the mice bone marrow cells. However, an increased frequency of micronucleated cells was observed when the highest concentration (10,670 ppm) was administered simultaneously with cyclophosphamide. In conclusion, the data indicate that annatto colour, for the conditions used, is neither mutagenic nor an inhibitor of induced mutations, although it should be used carefully since high doses may increase the effect of a mutagen.

Evaluation of the developmental toxicity of annatto in the rat.

Annatto, a dye extracted from Bixa orellana seeds, is used as a color additive in butter, cheese and in a variety of other foods as well as in drugs and cosmetics. Toxicological data on annatto and on its main carotenoid pigment bixin are still scarce. In this study we evaluated the developmental toxicity of annatto (28% of bixin). Annatto (0, 31.2, 62.5, 125, 250 and 500 mg/kg body weight/day) was given by gavage to Wistar rats on days 6-15 of pregnancy. Ceasarean sections were performed on day 21. Implantations, living and dead fetuses and resorptions were recorded. Fetuses were weighed and examined for externally-visible anomalies. One-third of fetuses from each litter was examined for visceral anomalies by a microsectioning technique. The remaining fetuses were cleared and stained with Alizarin Red S for skeleton evaluation. No adverse effect of annatto on the mothers was noted. No increase in embryolethality and no reduction of fetal body weight were observed among annatto-exposed rats. Annatto did not induce any increase in the incidence of externally-visible, visceral or skeletal anomalies in the exposed offspring. These findings suggest that annatto was neither maternally toxic nor embryotoxic in the rat. Therefore, the no-observed-adverse-effect level (NOAEL) for annatto-induced maternal and developmental toxicity was 500 mg/kg body weight/day or greater (or > or = 140 mg bixin/kg body weight/day) by the oral route.

[Regulatory aspects in the use of natural colorants]. / Regulamentação de uso de corantes naturais.

A number of preparations of natural colours from vegetable, insect and algae sources are presently used in various foods, although many of them have not been evaluated in relation to their safety of use. In evaluating the toxicity of food additives the concept of the Acceptable Daily Intake (ADI) has been used to provide an indication of safety for use and to enable regulatory authorities to take adequate legislative measures for their control. This paper will focus on the principles for the safety assessment of food additives, with emphasis on the guidelines that have been established by the Joint Expert Committee on Food Additives and Contaminants (JECFA) for evaluating natural food colours. Recent data on the potential intake of annato extracts in Brazil and current aspects of regulation of food colours at the level of MERCOSUR will also be presented.

Subchronic toxicity study in mice fed Spirulina maxima.

The purpose of this study was to evaluate the toxicity of Spirulina maxima, a blue-green alga used as food supplement and food coloring, after 13 weeks of treatment. Groups of ten mice of each sex were given S. maxima in the diet at concentrations of 0 (control), 10, 20 or 30% (w/w) for 13 weeks. The alga ingestion had no effect on behavior, food and water intake, growth or survival. Terminal values in hematology and clinical chemistry did not reveal differences between treated and control groups. However, male and female mice showed significant changes in serum cholesterol levels at 20 and 30% algal concentrations, but a toxic effect of S. maxima was excluded. Post-mortem examination revealed no differences in gross or microscopic findings. Our results show that S. maxima up to high feeding levels did not produce adverse effects in mice after subchronic treatment.

Isolation, purification and identification of the hyperglycaemic principle of the annatto (Bixa Orellana)

The use of herbal medicines in diabetes treatment is still a reality in Third World countries. The annatto is one of the plants widely used as an oral hypoglycaemic. Preliminary findings on the annatto, by a study group at the University of the West Indies showed, however, that anaesthetized dogs fed the crude extract of this annatto exhibited signs of hyperglycaemia. This unexpected result prompted further investigation. Studies commenced in 1985 to determine which component(s) from the crude extract was responsible for the observed hyperglycaemia. The chloroform eluent of the annatto seeds was separated by a chromatography sequence and recrystallizations, yielding a single bioactive component. Analysis by GC-mass spectrophotometry showed this component to be 99.4 percent pure, having a molecular weight of 394.2 and fitting the molecular formula C25H3O4. Further analysis by infrared (IR) spectrophotometry and proton nuclear magnetic resonance (NMR) supported the suggested molecular formula. Corroborative evidence was provided by 13C-NMR, confirming the structure of the hyperglycaemic agent as being that of transbixin. When this transbixin was fed to anaesthetized dogs in the amount of 0.6 g/Kg body weight, the dogs exhibited persistent hyperglycaemia. Additionally, electron microscopy revealed that they suffered severe damage to liver, kidney and pancreatic tissues to an extent that suggested that transbixin was not just hyperglycaemic, but also diabetogenic and toxic. Annatto pigments have, however, been listed by the FAO/WHO as being safe for human consumption and is widely used as colouring for butter, cheese, soups and other foods. The findings of this study indicate that another look will have to be taken at the annatto extracts as food additives, since transbixin is a major constituent. Further, the millions of poor in developing countries who continue to use annatto as a treatment for diabetes and other systemic ailments will need to be re-educated as to the possible implications of its continued use (AU)