World Journal of Dairy & Food Sciences 6 (1): 71-78, 2011 ISSN 1817-308X IDOSI Publications, 2011

A Review Article on Edible Pigments Properties and Sources as Natural Biocolorants in Foodstuff and Food Industry
Ali Aberoumand Department of Fisheries, Behbahan High Educational Complex, Behbahan, Iran
Abstract: Consumers are avoiding foods containing synthetic colourants, which lead food industries to replace them by natural pigments. Food colorants may be classified into synthetic, nature-identical, inorganic and natural colorants. Natural colorants for food are made from renewable sources. Most often, the colorants are extracted from plant material, but other sources such as insects, algae, cyanobacteria and fungi are used as well. Natural colorants are usually extracted and concentrated using either water or lower alcohols for water-soluble pigments and organic solvents for lipophilic pigments. Legislation restricts which colorants are allowed, what sources may be used for that particular colorant, what solvents may be used to extract it and the purity of the pigment. Colorants are formulated to make them more suitable for a variety of foods and drinks and to increase their stability. Key words: Food colorants % Natural colourants % Pigments % Properties INTRODUCTION Color is an important characteristic of food. Based on the color of the food, first impressions are made: Is the fruit immature, ripe, or overripe? Is the toast burnt? Is the food fresh? Based on these first impressions, a judgment is made whether the food is safe to eat or not and whether it can be expected to taste good or not. Since color is closely associated with expectations, the addition of color to food is a way to fulfill these expectations. Color is added to food for one or more of the following reasons: (1) to replace color lost during processing, (2) to enhance color already present, (3) to minimize batchtobatch variations and (4) to color otherwise uncolored food. Food colors can be divided into four categories: (1) natural colors, (2) nature-identical colors, (3) synthetic colors and (4) inorganic colors. Natural colors are pigments made by living organisms. Usually, pigments made by modification of materials from living organisms, such as caramel, vegetable carbon and Cu-chlorophyllin (vide infra), are also considered natural though they are in fact (except for carbon) not found in nature. Natureidentical colors are man-made pigments which are also found in nature. Examples are carotene, canthaxanthin and riboflavin. Synthetic colors are man-made colors which are not found in nature, these are often azo-dyes. Examples of inorganic colors are titanium dioxide, gold and silver. Although structurally very diversified and from a variety of sources, natural food colorants can be grouped into a few classes, the three most important of which are: tetrapyrrols, tetraterpenoids and flavonoids. The most important member of the tetrapyrrols is chlorophyll, which is found in all higher plants. Carotenoids are tetraterpenoids that are as ubiquitous as chlorophyll, since they too are part of the photosynthetic apparatus. They also give the yelloworangered color of many fruits. Anthocyanins are a group of flavonoids that provide the redpurple shade of many fruits, in particular berries (e.g., strawberries, elderberries and black currants). Other important classes of colorants are the anthraquinones (carmine, lac, kermes and madder) and the betalains (beetroot). The terms pigment and dye are often used interchangeably. Strictly speaking, a pigment is insoluble in the given medium, whereas a dye is soluble. Thus, carotenoids are dyes in oil but pigments in water. This distinction may be difficult to maintain if nothing is assumed about the medium and in the following the term pigment will be used for colored substances in general [1-8]. The development of foods with an attractive appearance is an important goal in the food industry. Increasingly, food producers are turning to natural food

Corresponding Author: Ali Aberoumand, Department of Fisheries, Behbahan High Educational Complex, Behbahan, Iran. Postal address: No 22, second line of Ab va Barg, Zolfeghari Street, Behbahan, Khuzestan Province, Iran. Tel: +98-9166716540, Fax: +98-6712229969, E-mail: aaberoomand@yahoo.com.

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colors, since certain artificial color additives have demonstrated negative health issues following their consumption [9]. Because of the deficiencies of existing natural food colorants, the demand for natural pigments is repeatedly raised by the food industry. This demand can be fuelled by research to offer a more natural-healthy way of coloring foods and provide a clean label declaration. Therefore, part of plant pigment research is seeking new sources of pigments. This will not only directed in finding natural alternatives for synthetic dyes, but also discover new procedures for the pigment production. The diversity of tropical and subtropical vegetation offers a promising range of unknown plant compounds that might prove applicable for the needs of humans. The most common plant pigments are carotenoids, chlorophylls, anthocyanins and betalains. Most research has been focused on carotenoids and anthocyanins but betalains have recently gained interest in food science. Commonly broad arrays of anthocyanin containing extracts are used for food coloring purposes; so far, there is only one single betalainic source that has been extensively used in the food industry worldwide. Compared to anthocyanins, betalains are ideal for coloring low-acid foodstuff as they maintain their color over a wide array of pH, from 3 to 7. The most important betalainic sources for natural red coloring are selected varieties of red beet, commercial preparations of which are mainly composed of the red-purple betanin and its C15-isomer isobetanin. Betacyanins are important constituents of betalain pigments. They are the main compounds associated with the red color exhibited by flowers, fruits and other plant tissues. Red beet has been established in the market as the oldest and most abundant red food colorant, called betanin, that is known as E-162 in the European Union [10-13] and as 73.40 in the chapter 21 of the Code of Federal Regulations (CFR) section of the Food and Drug Administration (FDA) in the USA (Griffiths, 2005). Practically it has been used to color foods such as yoghurt, confectionery, ice creams, syrups, sausages and processed meats. However, the typical earthy flavor caused by geosmin and high nitrate concentrations associated with the formation of carcinogenic nitrosamines [14, 15] may affect its commercial use. Furthermore, the risk to carry-over earth-bound microorganisms from the raw material in red beet is a crucial point [16,17]. Thereby, there is a high demand for other alternative compounds that can be substitute for red beet. In recent years fruits from the Cactaceae have been introduced as a promising betalain source. The fruits of

this family plant displayed a broader color spectrum and were devoid of the mention drawbacks in red beet [18]. Selected species of cactus fruits recognized as purple-fleshed Hylocereus polyrhizus have very recently been suggested as promising betacyanin source. The primary findings on the pigment potential and the scarce knowledge about color characteristics encouraged more investigations. Preliminary data from different Hylocereus genotype were promising and further investigations are needed for process optimisation with respect to color shade and pigment yield. The main obstacle in these fruits is the pectic substances. Method of degradation of pectic material is necessary to facilitate pigment release, increasing yield and reducing processing wastes. In Malaysia fruits of Hylocereus polyrhizus (genus Hylocereus, Cactaceae) have been cultivated as fruit crop locally. Red color pigments from fruits of H. Polyrhizus could be an interesting possibility as alternative to betanin from red beet. In view of the fact that most natural food colorants are supplied as concentrated juices or extracts and to get a better understanding of how pigment profile and hue are related in making decision for commercial applicability, the present work aimed to: C C C C To extract betacyanin pigments from pulp and skin of Hylocereus polyrhizus. To identify the constituents of the betacyanin pigments from pulp of Hylocereus polyrhizus. To evaluate the effect of enzyme treatment on yield of betacyanin constituent. To compare the color of extract from Hylocereus polyrhizus to that of red beet [19-21].

Natural Distribution of Pigments: Chlorophylls and carotenoids are the most abundant pigments in nature. They are involved in fundamental processes and life on Earth depends on them. Plants, photosynthetic bacteria and protozoa (plankton) are the main sources of the organic materials that are required for the development of other living organisms such as vertebrate and invertebrate animals. Chlorophyll is not found in animals but carotenoids accumulate in some organs (e.g., eyes) and tissues (e.g., skin of fish, bird plumage). In general, animal carotenoids are obtained from the common diet. Other pigments are also found in animals; some have important functions (e.g., heme proteins, riboflavin), whereas the function of others is not yet completely clear (e.g., melanins, flavonoids). Other organisms have interesting pigments that have been used or have potential use. Lichens produce depsides, the ancient

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and most extensively used dyes which were used as textile dyeing agents. In addition, they have application as sunlight filters, as chemical indicators (litmus paper, pH indicators) and as cytological stains. Some of the pigments obtained by treatment of lichen substances are orcein and parietin: More than 1000 pigments have been identified in fungi. Consequently, the diversity of fungi pigments is the second in importance, after plant flavonoids. Fungi are not photosynthetic and do not contain chlorophyll. Carotenoid distribution in fungi is restricted to some orders (e.g., Pharagmobasidiomycetidae, Discomycetes). In addition, flavonoids are scarce in fungi whereas riboflavin imparts the yellow color in the genera Use of natural colorant in food industry appears to have multidimensional potential [22]. For example, in addition to coloring property,carotene may be used in food as an essential vitamin source or betalains as source of essential amino acid or anthocyanins as quality control marker of food stuffs. Flavonoids are colorants with high pharmacological promise. Sometimes, diets with carotenoid mixtures are recommended instead of having just one particular carotenoid, because great variability of radicals and microenvironments take place in vivo [23]. Process development and use of new and available technologies for production optimization are the important area of research on natural food grade colors. Therefore, it requires a thorough review of the biotechnological potentials of natural food grade biocolorants. History of Biocolorants: Man has always been interested in colors. The art of dyeing is of long past and many of the dyes go back into prehistory. In Europe, it was practiced during the Bronze Age. The earliest written record of the use of natural dyes was found in China dated 2600 BC. In Indian subcontinent, dyeing was known as early as in the Indus Valleyperiod (2500 BC) and has been substantiated by findings of colored garments of cloth and traces of madder dye in the ruins of Mohenjodaro and Harappa civilization (3500 BC). In Egypt, mummies have been found wrapped in colored cloth. Chemical tests of red fabrics found in the tomb of King Tutankhamen in Egypt showed the presence of alizarin, a pigment extracted from madder. The cochineal dye was used by the people of Aztec and Maya culture period of Central and North America. By the4th century AD, dyes such as woad, madder, weld, Brazilwood, indigo and a dark reddish-purple were known. Brazil was named after the woad found there [24]. Henna was used even before 2500 BC, [25] while saffron

is mentioned in the Bible [26]. Use of natural biocolarants in food is known from Japan in the shosoin text of the Nara period (8th century) contains references to colored soybean and adzuki-bean cakes, so it appears that colored processed foods had been taken at least by people of some sections. Study of color intensified, since late 19th century, to understand: C C C The phenomenon for survival of animals and plants, The relation between color and evolution theories; and The role imparting in comparative physiology.

Thus, studies on biocolorant are greatly impulsed by their multiple functions [27]. The art of coloring spread widely with the advancement of civilization [28]. Source of Biocolorants: Plants, animals and microorganisms are the sources of natural biocolorants, but few of them are available in sufficient quantities for commercial use as food colorant and mostly are plant origin. For biotechnological production of such colorants, plants and microorganisms are more suitable due to their understanding of proper cultural techniques and processing. Natural colorants obtained from plant origin are pepper, red beet, grapes, saffron [29, 30]. Nowadays, fermentative production of food grade pigments are available in the market, for example; color from Monascus sp., astaxanthin from Xanthophyllomyces dendrorhous, Arpink red color from Penicillium oxalicum, riboflavin from Ashbya gossypii and -carotene from Blakeslea trispora. Also a number of microorganisms produce biocolors in good amount that includes Serratia and Streptomyces [31]. Biocolorants in Food Industry: To consider the natural biocolors as the colorant, stability, yield and price are mostly the constrains. Most of them are sensitive to pH, heat and sunlight [32]. Inspite of such factors, the natural biocolorants are gaining importance because of health and hygiene, nutrition, pharmaceutical activities, fashion and environmental consciousness, indicates relative dependency on natural products. Colors derived from minerals (lead chromate, copper sulphate) may cause serious health problems [33]. However, in last few decades, synthetic additives are severely criticized and consumers show inhibition toward these products, consequently they prefer to use the natural colorants [35]. In the 1960s in US, the environmental activists demonstrated against the use of such food additives

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and this attitude was spread out widely. Activists campaigned for the natural colorants highlighting their nutritional characteristics as a sales tool. A strategy that has failed earlier, later converted it into a total success, resulting from changes in social attitude. Thus, a worldwide tendency to use natural colorants is generated. Currently, people interpret the content of synthetic products as contaminant and the tendency has been reinforced [36]. There are number of advantages of using natural biocolorants, over synthetic colorants, further boosted because of their pharmacological properties. Products are of good market value if they are colored with natural compounds, for example; annatto in Cheddar cheese [37]. Food Grade Natural Biocolorants: Natural biocolorants are under exempt from certificationcategory of FDA and EU for food use and are: annatto extract, dehydrated red beet, canthaxanthin, carotene, Dactylopuis coccus extract, cotton seed meal, grape skin extract, fruit and vegetable juices, tagetes extract, carrot oil, oil of corn endosperm, paprika and paprika oleoresin, riboflavin, saffron, turmeric and turmeric oleoresin and xanthophylls (flavoxanthins, rubiaxanthins, zeaxanthin). Chlorophylls, in spite of its large intake amount per day. Yellow to orange color of Annatto comes from the outer layer of seeds of the tropical tree Bixa orellana. The carotenoids, bixin and norbixin, are responsible for appearance of yellow to orange color. The pH and solubility affect the color hue; the greater the solubility in oil, the brighter is the color. Water soluble, oil soluble and oil/water dispersible forms of annatto are available. Since it precipitates at low pH, is also available as emulsion, an acid proof state [38]. Annatto has been used for over two centuries as a food color especially in cheese and in various other food products [39]. Red beet (Beta vulgaris) extract shows variety of colors, depending on their content of yellow compound and may have a good flavor. Also a bluish-red color produced by a compound known as betanin and is stable at higher pH range than red cabbage extract [40]. There is no limit on its upper usage level. It has wide application in different food commodity from beverages to candy and from dairy to cattle products [41]. Canthaxanthin, a carotenoid, is commercially produced from the algae Haematococcus lacustris. Canthaxanthin is known mainly as the natural pigment of the orangeyellow chanterelle mushroom. Canthaxanthin has various physiological functions and can be converted into vitamin A under stress. Canthaxanthin is used in poultry for the appearance of color shade of the yolk,

also in cosmeticsand foods, particularly in dairy products (cheese), confectionary (soft and hard candy), fish and meat products, fruit products, beverages, snacks, beer and wine. It is more stable in photo degradation than that of carotene [42]. However, canthaxanthin is not considered as food additive under EU regulation. Cochineal insect (Dactylopius coccus), best cultivated on Opuntia ficusindica, extract is known to have carmine or carminic acid, which appears as magenta-red upon application. Water insoluble forms of carmine exhibit a color range from pink to purple. It is resistant to light, heat and chemical oxidation, often it is more stable than synthetic food colorants but unstable at low pH. The water soluble form is used in alcoholic drinks with calcium carmine; but water insoluble form is used in a variety of products. Together with ammonium, carmine is used inmeat, sausages, processed poultry products, alcoholic drinks, beverages, bakery and dairy products, including desserts and sweets. On an average, people consume one or two drops of carminic acid annually with food [43]. Cotton seed meal is the by-product after oil extraction from cotton seed. Gossypol is a fat soluble yellow pigment that occurs in cotton seed in bound and free forms. The bound form of gossypol is combined with free amino acids which constitute a medium quality protein feed [44]. Grape skin extract (enocianina) imparts a reddish purple color to beverages [45]. Some fruits contain a single type of anthocyanin (cyanidin in apple, cherry, fig, etc), some contain two major types (cyaniding and peonidin in ascherry, canberry); or some with several anthocyanins (grape). As colorant, grape juice is available in a variety of colors: red (shades of cherry, raspberry or strawberry), purple and yellow. It gives a strawberryred shade in application. It is used to color a number ofnon-beverage foods, including gelatin desserts, fruit fillings and in certain confectionaries. Anthocyanins from banana bract and Oxalis triangularis are found as potential source of food colorants [56]. Acylated anthocyanins from different edible sources (black carrot cultivars) are also used in food industry [47]. Vegetable juice is actually fermentable liquid product, either unfermented or lactic acid fermented, obtained from the edible part of one or more vegetables for direct consumption and preserved exclusively by physical means. The juice shall be free from skins, seeds and other coarse parts of the source vegetables. Tomato juice and blends based on tomato have long been popular and account for over 90% of the non-fruit juice trade. Lycopene is the principal compound derived from tomato, enlisted under US system. Under EU legislation,

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lycopene (E 160d) is considered as food additives. It is highly stable under a wide range of temperature and pH, hence used as common food colorant. It is available in liquid form or as cold-water dispersible powder. Carrot has long been a component of tomato blends [13]. Red-cabbage juice produces a bright pink to red color to a product with a pH less than 4.0 and is soluble in water, but not in oil. Use of Marigold (Tagetes erecta) flowers as source of food colorants is known from Aztec civilization. Marigold flowers are by far the most abundant natural source for commercial lutein [48]. The antimutagenic activity of carotenoids of Aztec marigold was evaluated. The presence of some essential amino and fatty acid increases the nutritional quality of carrot seed oil. Corn endosperm oil is a reddish-brown liquid composed chiefly of glycerides, fatty acids, sitosterols and carotenoid pigments obtained by isopropyl alcohol and hexane extraction from the gluten fraction of yellow corn grain [49]. It is used in chicken feed as color additive.Paprika (Capsicum annuum) is the pioneer and widely [50] used carotenoid as food colorant. In paprika, red carotenoidsare dominated by canthaxanthin, capsorubin where as yellow xanthophylls includes cryptoxanthin, zeaxanthin, antheraxanthin and -carotene [51]. Paprika oleoresin is mainly extracted from the pods. It contains three main naturally occurring pigments: capsanthin, capsorubin and carotene. This combination produces a bright orange to red-orange in food products. The oleoresin is oil soluble, when emulsified becomes water dispersible [52]. Riboflavin (vitamin B2) has a variety of applications as a yellow food colorant. Its use is permitted in most countries. Applications include dressings, sherbet, beverages, instant desserts, ice creams, tablets and other products. Riboflavin has a special affinity for cerealbased products, but its use in these applications is somewhat limited due to its slight odour and naturally bitter taste. Other Application in Food Industry Food Preservatives: Several natural biocolorants, including anthocyanin, additionally show antagonistic activity to certain bacteria, viruses and fungi thus to protect food from microbial spoilage (Bridle and [53]. Some are also active against protozoa (Leishmania brasiliensis) and insects (Calliphora erythrrocephala). Sometimes, carotenoids can act as sun screen to maintain the qualityof food by protecting from intense light. It has beenreported that corn carotenoids inhibit the synthesis of aflatoxin by Aspergillus flavus (90%) and by most of the A. parasiticus (30%) strains [54].

Carotenoids: Carotenoids are one of the most important groups of natural pigments. It has been estimated that nature produces about 100 million tons of thesepigments per year. They give brilliant yellow and red colours to many fruits, vegetables, roots, flowers and autumn leaves; they produce the colour of the egg yolk, of many algae, yeasts and mushrooms, Crustaceae as well as of the feathers and skins of many birds. The carotenoids have attracted the curiosity of scientists since the beginning of organic chemistry, actually since 1818. Numerous outstanding chemists made valuable contributions and many tools and methods have been introduced into organic chemistry via the carotenoids'. Imention the early analyses (by Willsttter and Zechmeister), the rediscovery of column chromatography by Kuhn and Karrer and of the thin-layer chromatography by Callebaut [l2], the elucidation of the isoprene rule by Ruzicka and of the symmetrical structures of squalene, /3-carotene and lycopene by Karrer, the studies on comparative biochemistry by Goodwin3, the first rules on u.v.absorption of polyenes by Richard Kuhn, the investigation of cis-trans isomerism [54] and the first syntheses by Karrer and Inhoffen. The application of n.m.r. spectroscopy by Jackman and Weedon5 proved to be a valuable tool for the identification of carotenoids, the synthesis of which was especially advanced by the development of condensation reactions for the buildup of the conjugated chain (Wittig6, Homer7). We aimed at the development of commercial syntheses of some selected carotenoids arid the investigation of their use as food colourants. By substituting natural colouring matters for artificial dyes we fulfil a physiological desideratum that good food should not contain unnatural pigments [54]. Production of Carotenoids from Natural Sources: The classical method for producing /3-carotene and other carotenoids is solvent extraction of the plant material. The natural source, the main pigments, possible provitamin A activity of the extract and the degree of pigmentation of eggs and broilers, after feeding, are indicated. Carrots, palm oil, alfalfa, dried grass and leaves are the starting materials for practically all preparations of natural carotene with provitamin A activity. The extracts from alfalfa, grass, leaves, Tagetes and yellow maize, containing either lutein or zeaxanthin, are valuable supplements to chicken feed because allxanthophylls colour the egg yolk in contrast to the carotenes. Besides /3-carotene, annatto which is an extract of the seeds of Bixa Orellana is also used for colouring dairy products.

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For red carotenoids large quantities of paprika are extracted. Torula yeast and certain algae are other sources of red polyene pigments used as colourants for oil and cheese. The market for natural carotenes has declined since the introduction of synthetic carotene. Today, only one crystalline carotene preparation extracted from dehydrated carrots is still on the market. Recently it was announced that /3-carotene and further carotenoids will be produced in Brazil by fermentation from mated strains of Blakeslea trispora [54]. CONCLUSION Research on carotenoids has included all branches of natural sciences has led the way to pure natural food colouring matters. It is my firm belief that for food colouring it is desirable to substitute artificial dyes by pure natural pigments. Furthermore I believe that pure synthetic carotenoids will gradually replace the natural extracts in the same way as the pure synthetic vitamins have practically eliminated the impurevitamin preparations obtained from natural sources. Future Prospects: Development of new food colorants can be divided into two categories: those covered by existing legislation and those that are not. Colorants covered by existing legislation are not really new, but encompass subjects such as new sources of known pigments (e.g., a newly discovered fruit rich in carotenoids), new ways of formulating existing pigments (e.g., blue anthocyanin) and improvement of existing sources (e.g., by breeding). Breeding and selection of high-yield strains will most likely only lead to minor improvements. Major improvements could be achieved by genetic manipulation, but this is not allowed in the EU and the naturalness of the colorant would be (partly) lost. Production of colors by fermentation has a number of advantages: cheaper production, possibly easier extraction, higher yields, no lack of raw materials and no seasonal variations. Fermented colors are already used today: D. salina, B. trispora, spirulina and monascus. It is not unlikely that new, fermented colors such as lycopene from B. trispora will become allowed in the near future. A giant leap forward in color production could be achieved by combining genetic manipulation and fermentation. Microorganisms could be made to produce colorants in high yield by inserting genes coding for the colorant even colorants not naturally produced by microorganisms

(e.g., turmeric) could be made in this way. Colorants made in this way will probably face some hurdles, at least in Europe where genetically modified food is generally viewed with a large degree of skepticism by the consumers. Finally, new colorants would have to be approved by the authorities, which is very costly because of the various toxicological studies needed to confirm the safety of a new food additive. The above gives a fairly compressed overview of the most important natural food colorants. The reader is referred to a number of excellent textbooks for more detailed information on plant pigments and food colorants [12-16]. REFERENCES 1. Alexandra, P.E., G.M. Monica, R.E. Wrolstad and M.B.A. Gloria, 2001. Anthocyanins from Oxalis triangularis as potential food colorants. Food Chem., 75: 211-216. 2. Andrews, A.G., H.G. Phaff and M.P. Starr, 1976. Carotenoids of Phaffia rhodozyma, a red-pigment fermenting yeast. Phytochem., 15: 1003-1007. 3. Baranyovits, F.L.C., 1978. Cochineal carmine: an ancient dye with a modern role. Endeavor, 2: 85-92. 4. Barth, M.M., C. Zhou, K.M. Kute and G.A. Rosenthal, 1995. Determination of optimum conditions for supercritical fluid extraction of carotenoids from carrot (Daucus carota L.) tissue. J. Agric. Food Chem., 43: 2876-2878. 5. Bartley, G.E. and P.A. Scolnik, 1995. Plant carotenoids: pigments for photoprotection, visual attraction and human health. Plant Cell., 7: 1027-1038. 6. Beyer, P., S. Al-Balli, X. Ye, P. Lucca, P. Schenb, R. Welsch and I. Potmyllus, 2002. Golden Rice: Introducing the Carotene biosynthesis pathway into rice endosperm by genetic engineering to defeet Vitamin A deficiency. J. Nutr., 132: 5065-5105. 7. Bosevska, M., D. Karlovic, J. Turkulov and D. Pericin, 1993. Quality of corn germ oil obtained by aqueous enzymatic extraction. J. Am. Oil Chem. Soc., 70: 1273-1277. 8. Boyles, M.J. and R.E. Wrolstad, 1993. Anthocyanin composition of red raspberry juice: influences of cultivar, processing and environmental factors. J. Food Sci., 58: 1135-1141. 9. Bridle, P. and C.F. Timberlake, 1997. Anthocyanins as natural food colourselected aspects. Food Chem., 58: 103-109. 10. Britton, G., 1991. Carotenoids. Method. Plant Biochem., 7: 473-518.

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