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A dye is a colored substance that has an affinity to the substrate to which it is being applied. The dye is generally applied in an aqueous solution, and requires a mordant to improve the fastness of the dye on the fiber. Some dyes can be precipitated with an inert salt to produce a lake pigment, and based on the salt used they could be aluminum lake, calcium lake or barium lake pigment. About 1.3 million tones of dyes, pigments and dye precursors, valued at around $23 billion were used and produced by textile industries as colorant of textiles. But the limitations of these synthetic dyes threatened to present living conditions due to their hazardous and non ecofriendly nature. Thus research has been going on to produce bioengineered natural green dyes from plant tissue and microorganisms. The majority of natural dyes are from plant sources roots, berries, bark, leaves, and wood, fungi, and lichens. Plant-based dyes such as woad, indigo, saffron, and madder were raised commercially and were important trade goods in the economies of Asia and Europe. The discovery of man-made synthetic dyes late in the 19th century ended the largescale market for natural dyes. This review involves the make use of microbes, plant and other natural substances that produce colorants. Pigments such as Anthraquinone, Annatto, Alizarin, Anthocyanins and Caretinoide mainly are obtained naturally. Such dyes can be used for dyeing natural fibers such as silk, wool etc., as well as synthetic fibers like polyester and gave good coloration. Dyeing can be performed by standard procedure for silk, wool and polyester dyeing. Good colorfastness of the dyed substrates to washing, sublimation and rubbing may be observed. If the dye displays an antimicrobial activity against Staphylococcus aureus, Corynebacterium

diphtheriae, Nocardia spps and Micrococcus luteus, its commercial value increases. Natural dyes are non hazardous and can be produced in required large quantities with cost effectiveness. Consequently even the textile manufactures would not be burdened with disposal problems of synthetic wastes and their pretreatments before disposal if they use natural dyes in place of synthetic. Thus natural occurring dyes have a presumption to be used as an alternative to synthetic dye for dyeing cotton and wool.


A dye is a colored substance that has an affinity to the substrate to which it is being applied. The dye is generally applied in an aqueous solution, and requires a mordant to improve the fastness of the dye on the fiber. Both dyes and pigments appear to be colored because they absorb some wavelengths of light more than others. In contrast with a dye, a pigment generally is insoluble, and has no affinity for the substrate. Some dyes can be precipitated with an inert salt to produce a lake pigment, and based on the salt used they could be aluminum lake, calcium lake or barium lake pigments. Textiles have been manufactured using various technologies since time immemorial. Human ingenuity and imagination, craftsmanship and resourcefulness are evident in textile products throughout the ages. We are to this day awed by beauty and sophistication of textiles sometimes found in archeological excavations. After fabricating the mansions of fashion and comfort textiles are now moving towards high-tech era of performance which has brought up diversification and expansion of technologies. This realization of technologists has coincided with rapid developments in technology and brought about a surge in research and development activities in textiles (Kumar, 2007). The textile industry produces and uses approximately 1.3 million tones of dyes, pigments and dye precursors, valued at around $23 billion, almost all of which is manufactured synthetically

(Sengupta and Singh, 2003). Biotechnology in Textiles is one of the revolutionary ways to advance the textile field (Kumar, 2007).

Dyed flax fibers have been found in the Republic of Georgia dated back in a prehistoric cave to 36,000 BP (Balter, 2009 Kvavadze, 2009). Archaeological evidence shows that, particularly in India and Phoenicia, dyeing has been widely carried out for over 5000 years. The dyes were obtained from animal, vegetable or mineral origin, with no or very little processing. By far the greatest source of dyes has been from the plant kingdom, notably roots, berries, bark, leaves and wood, but only a few have ever been used on a commercial scale. The first synthetic dye was William Perkins's mauveine in 1856, derived from coal tar. Alizarin, the red dye present in madder, was the first natural pigment to be duplicated synthetically, in 1869 (Zollinger, 2003), a development which led to the collapse of the market for naturally grown madder (Garfield, 2000). The development of new, strongly colored synthetic dyes was followed quickly, and by the 1870s commercial dyeing with natural dyestuffs was disappearing.


2600 BC Earliest written record of the use of dyestuffs in China 715 BC Wool dyeing established as craft in Rome 331 BC Alexander finds 190 year old purple robes when he conquers Susa, the Persian

capital. They were in the royal treasury and said to be worth $6 million (equivalent) 327 BC Alexander the Great mentions "beautiful printed cottons" in India
236 BC An Egyptian papyrus mentions dyers as "stinking of fish, with tired eyes and hands

working unceasingly
55 BC Romans found painted people "picti" in Gaul dyeing themselves with Woad (same

chemical content of color as indigo) 2ND and 3RD Centuries AD Roman graves found with madder and indigo dyed textiles, replacing the old Imperial Purple (purpura)
3rd Century papyrus found in a grave contains the oldest dye recipe known, for imitation

purple - called Stockholm Papyrus. It is a Greek work.

273 AD Emperor Aurelian refused to let his wife buy a purpura-dyed silk garment. It cost

its weight in gold.

Late 4TH Century Emperor Theodosium of Byzantium issued a decree forbidding the use

of certain shades of purple except by the Imperial family on pain of death

400 AD Murex (the mollusk from which purpura comes) becoming scarce due to huge

demand and over harvesting for Romans. One pound of cloth dyed with Murex worth $20,000 in terms of our money today (Emperor Augustus source) 700's a Chinese manuscript mentions dyeing with wax resist technique (batik)

A fragment of madder-dyed cotton fabric found at the Harappan sites indicates the use of natural dyes by the people of Mohenjodaro as back as 3000 B.C. The tinctorial properties of vegetable substances, recognized in the Vedic period, particularly in the Atharvavedic and the succeeding periods ranging from 1000 to 500 B.C. were kala or asikni (possibly indicating indigo), maharanjana (sunflower), manjistha (madder), lodhra (symlocis racemasa) and haridra (turmeric). The dyestuff introduced in the post-Vedic period ranging from 500B.C. to 3rd century A.D. included Kumkuma (saffron) and nila (indigo) among the plant products; krmi (kermes) and rocana (bright yellow substance prepared from cows urine) among the animal substances; gairika (red-ochre) among minerals; and khanjana (carbon black). The period from the Classical Age to Medieval Period of Indian history acknowledges the tinting capacity of a number of vegetable substances as well as of metals and minerals. The Medieval Period was marked by the discovery of the color fixation property of tuvari (alum) and the process employed for the extraction of the coloring principles from the dyestuff. The late Medieval Period (18th century) introduced the application of iron mordant for the fixation of colors like blue, green

and violet. It is during the Mugal Period that the shades of a number of colors in different tones came into the field of dyeing with natural dyes.


Throughout history, there have been several articles of clothing and dye colors that have made significant impacts on society. In the past, dyes were produced individually by harvesting natural fruits, vegetables and other items, boiling them, and submersing fabrics in the dye bath. It was a long and tedious process. This was the common practice until the mid-1800s (Joseph, 1977). Today, pre-packaged dyes are readily obtainable in almost every color, and are available to anyone who can purchase the end product. The development of different dyes and techniques has made this transition possible.

In the past, only organic matter was available for use in making dyes. Today, there are numerous options and methods for the colorization of textiles. While todays methods capitalize on efficiency, there is question as to whether the use of chemicals is harmful to the environment. A reputation for harming the earth could be detrimental to a company in a society becoming more and more focused on the environment and its preservation. Some say the discovery of dyes was probably an accident, a simple stain from a berry for fruit, and dates back shortly after the dawn of civilization. Ancient man used almost exclusively vegetable dyes. Known roots and berries and various sorts of dyestuffs were gathered, boiled, and then fabrics and textiles were submerged, resulting in the first dyed products. In this way many new dyestuffs were discovered. In fact, numerous writers have confirmed the knowledge of over one thousand sources of dyes (Leggett, 1944). As trade and commerce developed, certain dyes were regarded as better than others, and some ceased being used. Natural dyes used included vegetable, animal, and mineral dyes. In 1856, Sir William Perkin made a discovery that some even say changed the world. He discovered the color mauve. This was the first synthetic dye. His method for the dyeing of this color, using coal and tar, resulted in many scientific advances and the development of synthetic dyes that are widely used today (Garfield, 2001). No matter which dyeing technique is used, natural or synthetic, water is required to complete the process. In order for water to be usable, it must be purified. Depending on the source of the water, it may contain an excess of carbon dioxide, bicarbonates, sulphates and chlorides of calcium, magnesium and sodium. Water can be

identified in the three following broad categories: Well or spring waters, moorland waters, and surface waters. Well water is usually clear and of constant composition. It may have some sodium bicarbonate, bicarbonates of calcium, magnesium, and iron, as well as free carbon dioxide. Moorland water may be tinted and somewhat acidic. The color and acid in this water is from organic materials. The acidity and some dissolved gases may make this water corrosive. It may contain calcium chloride, magnesium chloride, and sulphates. Surface water contains sulphates, chlorides, calcium bicarbonate, and magnesium bicarbonate as well. These impurities must be removed to the limits of suitable water quality (Peters, 1967). After water is treated for turbidity and color, iron and manganese, alkalinity, and hardness of water, it can be used in textile mills (Goetz, 2008).

"The process of applying color to fiber stock, yarn or fabric is called dyeing." There may or may not be thorough penetration of the colorant into the fibers or yarns. Dyeing is the process of adding color to textile products like fibers, yarns, and fabrics. Dyeing is normally done in a special solution containing dyes and particular chemical material. After dyeing, dye molecules have uncut Chemical bond with fiber molecules. The temperature and time controlling are two key factors in dyeing. There are mainly two classes of dye, natural and manmade.



Primitive society discovered that certain roots, leaves, or bark could be manipulated, usually into a liquid form, and then used to dye textiles. They used these techniques to decorate clothing, utensils, and even the body. This was a religious, as well as functional practice. Records date all the way back to over 5,000 years ago chronicling the dyeing of fabrics. Excavated cloth fragments that date back to 3000 B.C. evidence the intricate dyeing practices and skills of India as well as hundreds of vibrant colors used to dye things in Peru (Belfer, 1972). Certain hues have historical importance and denote social standing. For example, the color royal purple was reserved for royalty and nobility not all that long ago (Belfer, 1972). The dye for this was made from the secretions of shellfish. Shellfish produce a clear fluid that oxidizes when exposed to the air; this was used to produce a red to bluish purple. This dye was difficult to create and used only on the finest garments; hence it became associated with aristocrats and royalty. Knowledge of this Tyrian purple was lost during the Middle Ages and not rediscovered until the mid- 1800s by a French scholar (Belfer, 1972). Blue dyes were particularly difficult to come across and thus were viewed as a sign of wealth (Dye). The colors one wore could even proclaim their sins, as chronicled in Nathaniel Hawthornes book The Scarlet Letter. Ancient India was particularly advanced in dyeing techniques and has been known since the sixteenth century for their vibrant colors and designs on fabrics. Resist dyeing techniques probably originated here. Also, Indians discovered the use of mordants to make dyes fast and an integral part of the fabrics, not just a pigment on the surface (Belfer, 1972) (Goetz, 2008).



Dyeing can be carried out at any of the following stages in the textile manufacturing stage:

The fibers can be dyed before they are spun. Fiber dyeing provides a deep penetration of

the dye into the fiber, giving even color and excellent color-fastness.
The yarn can be dyed after spinning but before the product is woven or otherwise

fabricated. This is called package dyeing.

Before the fabric is finished, it can be dyed in lengths (piece dyeing).This process allows

manufacturers the opportunity to produce fabrics in their natural colors, and then dye them to order.
In cross-dyeing, fabrics of two or more fibers can be dyed so that each fiber accepts a

different dyestuff and becomes a different color, through the use of appropriate dyestuffs for each fiber.

It is essential for the correct identification of the fiber or other fabric to be made before dyeing commences.


The classes of dyes defined by the application or end-use, and hence the terms most applicable to textile dyeing, are: 1. Acid - water-soluble anionic dyes 2. Basic - water-soluble cationic dyes
3. Direct - water-soluble anionic dyes which are substantive to cellulosic fibers when dyed

from aqueous solution

4. Disperse dyes - water insoluble, nonionic dyes used for dyeing hydrophobic fibers from

aqueous dispersion 5. Fluorescent brighteners - not dyes as such, applied from solution, dispersion or suspension in a mass
6. Reactive dyes - form a covalent bond with the fiber (usually cotton, wool or nylon)

7. Sulphur dyes - applied to cotton from an alkaline reducing bath

8. Vat dyes - water insoluble dyes applied mainly to cellulosic fibers as soluble leuco (i.e.,

colorless) salts after reduction in an alkaline bath 9. Pigments - printed on to surface with resin binder or dispersion in a mass
10. Solvent dyes - dissolved into the substrate (e.g., varnish, waxes, inks, oils)


The first recorded synthetic dye was picric acid which produced in the 1770s from the interaction of indigo and nitric acid. The synthetic dye industry is considered to have started when Perkins synthesized mauveine (mauve) in 1856 in the UK. The industry quickly developed, mainly in Germany, Switzerland and the UK. Since then the significant dyes discovered were: alizarin red, 1868, by Grabe and Lieberman; indigo in 1870 by von Bayaer; the azo dyes in 1880; anthraquinone vat dyes in 1901 by Bohn; disperse dyes for cellulose in 1922; fiber reactive dyes by ICI in 1956. There are very many synthetic dyes available now. Their tinctorial strength, concentration, color range and color fastness, particularly to light and detergents, make them superior to natural dyes for nearly all uses. They are relatively cheap and have other advantages, e.g., mordants may not be necessary, they can color synthetic fibers. Manufacturers and processors using dyes have to deal with potential health, effluent disposal and other environmental requirements, usually to statutory limits (Hancock, 1997).


The majority of natural dyes are from plant sources roots, berries, bark, leaves, and wood, fungi, and lichens. Throughout history, people have dyed their textiles using common, locally available materials. Scarce dyestuffs that produced brilliant and permanent colors such as the natural invertebrate dyes Tyrian purple and crimson kermes were highly prized luxury items in the ancient and medieval world. Plant-based dyes such as woad, indigo, saffron, and madder were raised commercially and were important trade goods in the economies of Asia and Europe. Across Asia and Africa, patterned fabrics were produced using resist dyeing techniques to control the absorption of color in piece-dyed cloth. Dyes from the New World such as cochineal and logwood were brought to Europe by the Spanish treasure fleets, and the dyestuffs of Europe were carried by colonists to America. Natural dyes comprises of those colorants (dyes and pigments) that are obtained from animal or vegetable matter without chemical processing. They are mainly mordant dyes although some vat, solvent, pigment, and acid types are known. Natural dyes fall into three categories on the basis of their origin; plant/vegetable origin, Insect/ animal origin and mineral origin. Some of the examples of plants used for producing natural dyes are; Al, Alkanet, Balsam, Bougainvillea, Canna, Tulsi, Terminalia Arjuna, etc. India has a very rich tradition of using natural dyes. The art and craft of producing natural dyed textile has been practiced since ages in many villages by traditional expert crafts persons in the country. Natural dyes, when used by them have many limitations of fastness and brilliancy of shade. However, when used along with metallic mordants they produce bright and fast colors. The use of metallic mordants is not always eco friendly, but the pollution problems created by metallic mordants are of very low order and can be easily overcome. Therefore, instead 14

of using unsustainable technology for producing colors one can use mild chemistry to achieve almost similar results. There is a growing demand for eco friendly/non toxic colorants, specifically for health sensitive applications such as coloration of food and dyeing of child textile/leather garments. Recently, dyes derived from natural sources for these applications have emerged as an important alternative to potentially harmful synthetic dyes and pose need for suitable effective extraction methodologies. Natural dyes also referred as mordant dyes; do not readily adhere to cotton so mordants are used. Mordants are needed to set the color when using natural dyes. It is thus a chemical agent which allows a reaction to cur between the dye and fabric. Some of the important natural dyes are blue dyes, red dyes, yellow dyes, etc. Natural dyes can produce special aesthetic qualities, which, combined with the ethical significance of a product that is environmentally friendly, gives added value to textile production as craftwork and as an industry. There are various kind of natural dyes; quinonoid dyes, cyanine dyes, azo dyes, biflvonyl dyes, omochromes, anthraquinone, coprosma gesus, Indigoid dyes, Tannins Tannins, Flavonoids, Betacyanins (betalains) etc. The use of natural dyes in cloth making can be seen as a necessary luxury to trigger off a change in habits. Dyes which stand out for their beauty and ecological attributes would never be employed on just any material but on noble fabrics such as wool, silk, linen or cotton, made to last more than one season. Market value will benefit from consumer preferences for environmentally friendly products, which will support consumption of high performance dyes and organic pigments.




Many natural dye stuff and stains were obtained mainly from plants and dominated as sources of natural dyes, producing different colors like red, yellow, blue, black, brown and a combination of these. Almost all parts of plants like root, stem, leaves, flower, wood, seeds, fruits etc. produce dyes. It is interesting to note that over 2000 pigments are synthesized by various parts of plant, of which only about 150 have been commercially exploited.


Ocher is a dye obtained from an impure earthy ore of iron or ferruginous clay, usually red (hematite) or yellow (limonite). In addition to being the principal ore of iron, hematite is a constituent of a number of abrasives and pigments.


Cochineal is a brilliant red dye produced from insects living on cactus plants. The properties of cochineal bug were discovered by pre- Columbian Indians, who dried the female insects under the sun, and then ground the dried bodies to produce a rich red powder. When mixed with water, the powder produced a deep, vibrant red color. Cochineal is still harvested today on the Canary Islands. In fact, most cherries today have a bright red appearance through the artificial color carmine, which is obtained from the cochineal insects (Siva, 1997).



Balfour-Paul has reviewed the use of textiles dyed with natural indigo by Arabic people, who perceive that wearing such textiles next to the skin has a positive health benefit, including a lower incidence of illness (Balfour, 1997). There has been much interest recently (as indicated by the number of conferences on the topic) in the pharmacological effects and possible health benefits of the use of natural dyes but, as these benefits are often very hard to quantify on scientific grounds, we should wait to see the extent to which they become substantiated by medical research. It is important not to underestimate the public perception of such benefits (Hill, 1997).

Few natural dyes are color-fast with fibers. Mordants are substances which are used to fix a dye to the fibers. They also improve the take-up quality of the fabric and help improve color and light-fastness. The term is derived from the Latin murdered, to bite. Some natural dyes, indigo for example, will fix without the aid of a mordant; these dyes are known as substantive dyes. Others dye, such as madder and weld, have a limited fastness and the color will fade with washing and exposure to light.


Traditionally, mordants were found in nature. Wood ash or stale urine may have been used as an alkali mordant, and acids could be found in acidic fruits or rhubarb leaves (which contain oxalic acid), for example. Nowadays most natural dyers use chemical mordants such as alum, copper sulphate, iron or chrome (there are concerns, however about the toxic nature of chrome and some practitioners recommend that it is not used). Mordants are prepared in solution, often with the addition of an assistant which improves the fixing of the mordant to the yarn or fiber. The most commonly used mordant is alum, which is usually used with cream of tartar as an additive or assistant. Other mordants are: Iron (ferrous sulphate) Tin (stannous chloride)
Chrome (dichromate of potash)

Copper sulphate Tannic acid Oxalic acid Using a different mordant with the same dyestuff can produce different shades, for example;
Iron is used as a saddener and is used to darken colors. Copper sulphate also darkens but can give shades which are otherwise very difficult to

obtain. 18

Tin brightens colors. Tannic acid, used traditionally with other mordants, will add brilliancy. Chrome is good for obtaining yellows. Oxalic acid is good for extracting blues from berries. Cream of Tartar is not really a mordant but is used to give a luster to wool.

Mordants are often poisonous, and in the dye-house they should be kept on a high shelf out of the reach of children. Always use protective clothing when working with mordants and avoid breathing the fumes. The mordant can be added before, during or after the dyeing stage, although most recipes call for mordanting to take place prior to dyeing. It is best to follow the instructions given in the recipe being used or experiment on a sample before carrying out the final dyeing. Later in this brief we will explain how the mordant is mixed and used as part of the dyeing process. These chemical mordants are usually obtained from specialist suppliers or from chemists. Where this is prohibitive, due to location or cost, natural mordants can be used. There are a number of plants and minerals which will yield a suitable mordant, but their availability will be dependent upon your surroundings. Some common substitutes for a selection of mordants are listed below. Some plants, such as mosses and tea, contain a small amount of aluminium. This can be used as a substitute to alum. It is difficult to know, however, how much aluminium will be present and experimentation may be necessary. 19

Iron water can be used as a substitute to ferrous sulphate. This can be made simply by adding some rusty nails and a cupful of vinegar to a bucket-full of water and allowing the mixture to sit for a couple of weeks. Oak galls or sumach leaves can be used a substitute to tannic acid.
Rhubarb leaves contain oxalic acid.

The discovery of man-made synthetic dyes late in the 19th century ended the large-scale market for natural dyes.


Use of synthetic dyes leads to gross contamination (Lokhande et al., 1999) and hence with the present national and international awareness of environment and ecology, the use of ecofriendly fibers and natural dyes has been increasing all over the globe (Verma and Venkatachalam, 2002). Synthetic dyes are known to have hazardous effects on health (Clarke and Steinle, 1995) as opposed to natural dyes that have desirable properties such as antimicrobial activity, stability to light, heat and pH (Pandey and Babitha, 2005). Thus there is an urgent need to obtain alternative colorants that are natural, cost effective and easily degradable without production of recalcitrant intermediates when they enter the ecosystem. Moreover the fact that these colors are antimicrobial in nature, the use of antimicrobial finish can be avoided completely and thus becomes cost effective (Murugkar et al., 2006). Increasing global competition in textiles has created many challenges for textile researchers and industrialists. The rapid growth in technical textiles and their end-uses has generated many opportunities for the application of innovative finishes. Novel finishes of high added value for the apparel fabrics are also greatly appreciated by a more discerning and demanding consumer market. Antimicrobial textiles with improved functionality find a variety of application such as health and hygiene products, specially the garment worn close to skin and several medical applications, such as infection control and barrier material.

Thus, even though the availability of natural dyes has been known for centuries, the reasons synthetic dyes have been so popular are: 21

They are simple to produce in large quantities, They can be manufactured at a reasonable price ($10 100/kg), They can provide the variety of colors that are demanded by todays consumers,
They provide high color-fastness (i.e., the dye is very strongly bound to the fabric and does

not detach after repeated washing cycles). However, manufacturing of synthetic dyes suffers from the following limitations:
Environmentally Unfriendly: The production of synthetic dyes requires strong acids,

alkalis, solvents, high temperatures, and heavy metal catalysts. For example, production of a dye designated as Color Index Mordant Blue 23 states, Treat 4,8-diamino-1,3,5,7tetrahydroxy-2,6- anthraquinonedisulfonic acid with boiling alkali or dilute acid and convert to the sodium salt or Treat 1,5-dinitro-anthraquinone with fuming sulfuric acid in the presence of sulfur, hydrolyze with water, and convert to the sodium salt.
Increase in Cost of Feedstock or Energy: Petroleum is the starting material for all synthetic

dyes and thus the price of dyes is sensitive to the price of petroleum. Also, since synthesis is energy intensive (uses super-heated steam, boiling acids, etc.); the process is sensitive to energy prices and also generates greenhouse gases. Hazardous Waste Generation: Since synthetic production of dyes needs very toxic and hazardous chemicals, it also generates a hazardous waste, the disposal of which is a major environmental and economic challenge. Moreover, some facilities that produced dyes in


the past are now Superfund sites due to intentional dumping or accidental spills of toxic and hazardous wastes. Increasing Transportation Costs: Since dyes are hazardous materials and are produced at central facilities, transportation of dyes from manufacturing plants to textile dyeing and printing facilities is a major cost item and a logistic challenge.
Toxic and Allergic Reactions: There are occupational safety issues involved since

production processes use the toxic and hazardous materials and conditions described above. Thus, if bioengineered natural, green dyes can be produced at a comparable price, the following benefits will be realized:
Reduce the use of toxics since starting materials are environmentally benign with

associated benefits in terms of waste disposal and occupational safety. Production can be decentralized resulting in savings in transportation costs. After extraction of the dye, the biomass can be used for energy generation (e.g., through anaerobic treatment to generate methane, which in turn, can be sued as a fuel) and the growth media can be recycled; thus, there are virtually no wastes generated. Possible beneficial aspects such as higher UV absorption by the fabric (which contains natural dye) can result in reduced incidence of melanoma. It is clear, however, that if natural dyes are to be considered as an alternative to the synthetic dyes used today, they have to manifest the same characteristics of synthetic dyes as those listed above. 23

Specifically, the major challenges in this field are: To produce natural dyes in the quantities required, To produce natural dyes at a reasonable price,
To produce natural dyes that has high color-fastness.



1. Extraction from plants

2. Extraction from arthropods and marine invertebrates (e.g., sea urchins and starfish) 3. Extraction from algae (e.g., blue-green algae) 4. Production from bacteria and fungi (Sengupta and Singh, 2003).

There are primarily 4 sources from which natural dyes are available namely specialized plant and animal sources by-product (especially lac dyes), chemical synthesis and tissue or cell culture by DNA transfer (Lee et al., 2008) (Mutnuri et al., 2009). Plants, animals and microorganisms are the sources of natural biocolorants, but few of them are available in sufficient quantities for commercial use as textile 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 (FDA/IFIC, 1993; Bridle and Timberlake, 1997). Nowadays, fermentative production of textile dye 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 (Kim et al., 1997) (Chattopadhyay et al., 2008).


Regardless of the source, it is believed that products which may be harnessed as green dyes are in essence secondary metabolites produced by the organism. These secondary metabolites are low molecular weight natural products that have a restricted taxonomic distribution, possess no obvious function in cell growth and are synthesized for a finite period by cells that are no longer undergoing balanced growth (Sengupta and Singh, 2003). In recent years, research into new sources of colorants, inclusively for textile applications, has explored different possibilities, such as:
1. Cell culture of parts of plants that could not be exploited on a big scale in a sustainable

way, such as woody plants (like al or Indian mulberry, Morinda citrifolia) in which it is the root bark of mature trees that is used and therefore, cannot be harvested without destroying the tree. In Chiangmai University, in Thailand, root cell culture of another common Morinda species, Morinda angustifolia Roxb. Var. scabridula Craib. has produced, in only 5 months, 0.6 times the amount of red dyes produced by 2-3 years old plants.
2. Bacteria and fungi can also be used for the biotechnological production of new natural

pigments. An effective biotechnology solution to manufacture of these and other dyes or dyestuff intermediates will impart the following benefits: The medium in which these plant cells or fungi or bacteria grow contain no expensive or toxic chemicals. The process is carried out at low temperature (around 30 C) compared to the fuelconsuming very high temperatures in the synthetic process 26

The process is typically run at neutral pH as opposed to very high acidic or alkaline conditions in the synthetic process The process is very environmentally friendly and sustainable However, the key factors are: High yield of the product,
High purity (Sengupta and Singh, 2003).

R&D work is under progress in genetically modified micro-organisms and dyestuffs for the textile field.


Textile auxiliaries such as dyes could be produced by fermentation or from plants in the future, before the invention of synthetic dyes in the nineteenth century many of the colors used to dye textiles came from plants e.g. indigo and madder. Biotechnological route for producing pigments for use in the food, cosmetics or textile industries is from plant cell culture. Plant cell cultures also provide effective systems for exploring plant physiology and plant biochemistry. The value of the technique of plant tissue and cell culture is that cell and tissue systems can be subjected to direct experimental control (Sengupta and Singh, 2003). One of the major success stories of plant biotechnology so far has been the commercial production since 1983 in Japan of the red pigment shikonin which has been incorporated into a new range of cosmetics. Traditionally, shikonin was extracted from the roots of five year old plants of the species Lithosperum erythrorhiz where it makes up about 1 to 2 percent of the dry weight of the roots. In 27

tissue culture, pigment yields of about 15 percent of the dry weight of the root cells have been achieved. PRODUCTION OF ALIZARIN BY CELL CULTURES: Cell cultures derived from roots of madder (Rubia tinctorum) have the ability to synthesize anthraquinones, with alizarin being extractable from the cells (Toth et al., 1993). The yield of alizarin in callus cells reached about one-third of that of field-grown roots (Lodhi et al., 1994). Further work could help understand the regulation of anthraquinone synthesis, and cell cultures may eventually also become a possible alternative to the field crop. It is clear that more research could yield an even higher level of technology in dyestuff production and dyeing with natural dyes. There is still a considerable lack of information on the biosynthetic pathways on many dye molecules. The physiological regulation of yield is poorly understood and there has been virtually no systematic plant breeding to increase yield. If we consider what research over the last 150 years has done for other crops, the yield of dyes could be greatly undervalued and underexploited. Although cell or microbial cultures could circumvent the need for agricultural production, industrial culturing has its own significant environmental impacts, and may indeed blur the distinction between synthetic and natural dyes (Hill, 1997).


PRODUCITON OF ANTHOCYANIN AND FLAVONOL FROM CRANBERRY: In cranberry fruits, the anthocyanins are cyanidin and peonidin based chromophores, whereas the flavonols are myricetin and quercetin based chromophores. A major advantage with anthocyanins and flavonols for their use as natural dyes is the fact that they have attached sugar groups, which can be removed without loss of their colors. The chemical group freed after removal of sugars could then be used to attach these dyes with cellulose in fibers. The technology is simple to implement two steps: 1. Grow the primary species in the laboratory or in a bioreactor with the right growth medium, 2. Harvest the dye or dye precursors from the plant or fungal cells. Successfully established cranberry cell culture system from cranberry (Vaccinium macrocarpon, Ericaceae) stems, leaves and leafstalks by using Gamborgs B5 medium containing 5 mM 1naphthalene acetic acid (NAA), 5 M 2,4-di-chlorophe-noxy acetic acid (2,4 D), and 2.5 mM kinetin at 25 in the dark were taken into consideration.


Production of anthocyanin from Cranberry Callus: Production of flavonoids in cranberry callus varies under different stresses such as light irradiation (red light of 660 nm and far-red light of 730 nm), temperature-changing (from 25 to 4, or 37), and wounding. Cranberry callus produces anthocyanins only on exposure to light. Production of anthocyanins in cranberry callus was induced under continuous light irradiation. However, because anthocyanin is red color, it was observed that only top layer of the callus, which received light produces anthocyanins. Production of anthocyanin and flavonol from Cranberry Cell Suspension Although callus provides more accessible uniform cells than the intact plant, the callus tissue is not uniform since only the base of the callus is exposed to the medium and the callus mass may contain cells at various stages of development. The alternative approach is to use cell suspensions. Cell suspensions show a faster growth rate, and all cells are exposed uniformly to the medium, and environment such as light. Cell suspensions are preferred for large scale and commercial production of secondary metabolic products. Initiation of cranberry cell suspension culture was done by transferring callus to liquid media of the same composition as the callus medium and gently agitating the suspension on a horizontal shaker at 150 rpm and 20. They have found out that the growth of biomass of cranberry cell suspension culture in WP and MS liquid media were greater than B5 liquid medium. Interestingly, anthocyanin content of cranberry cell suspension culture in MS liquid medium was higher than in WP liquid medium. However, the flavonol content of cranberry cell suspension culture in WP liquid medium was higher than in MS liquid medium.


Preparation of Anthocyanin and Flavonols: Cranberry anthocyanins and flavonols were extracted in a mini-blender with 20 ml of extraction solvent (85:15, ethanol: 1.5 M HCl) (Boulanger and Singh, 1998). The resulting homogenate was allowed to incubate at 4C overnight. Homogenates were filtered, and residue on the filter paper was washed with the extraction solvent to remove all pigments. The filtrate was diluted to 250 ml in a volumetric flask, and the solution spectrally analyzed. Flavonoid contents were estimated using published methods of cranberry anthocyanin and flavonol analysis (Sapers and Hargrave, 1983 Fuleki and Francis, 1968). The extract was found feasible to be used as a dyestuff at higher concentrations for Nylon-66 and Wool. Dye-affinity of the fabric increases with the increase in the concentration of the extract used. The extract gave very bright and acceptable shades for fabrics of coarser yarns, comparable to synthetic dyes, such as Nylamine Red A2B (Sengupta and Singh, 2003).


Many micro-organisms produce pigments during their growth which are substantive as indicated by the permanent staining that is often associated with mildew growth on textiles and plastics. Attempts have been made to synthesize bacterial forms of indigo as well as fungal pigments for use in the textile industry. Certain micro fungi are capable of yielding up to 30% of their biomass as pigment (Kumar, 2007).


Several of these microbial pigments have been shown to be benzoquinone, naphthoquinone, anthraquinone, perinaphthenone and benzofluoranthenequinone derivatives, resembling in some instances the important group of vat dyes.

BACTERIA: Secondary metabolites of bacterial origin include various enzymes, pigments, antibiotics etc that could be of importance to mankind in many ways. Pigments in bacteria are a source of variety of functions such as protection from photo oxidative damage, converting light energy to chemical energy, etc (Archik, 1980 Britton, 1983). These bacterial pigments are generating newer interest due to their use in food, cosmetic, textile and paper industries (Murugkar et al., 2006). Production of red color dye by fermentation: Inspire of the availability of variety of dyes from fruits and vegetables, there is an ever growing interest in the microbial dyes due to several reasons, like their natural character, safety to use and production being independent of seasons, controllable and with a predictable yield. A red colored microbe was isolated from mangrove soil. Large amount of red color was produced on Modified Nutrient agar. Such environment friendly microbial dye was used successfully to dye natural fibers such as silk, wool etc. as well as synthetic fibers such as polyester giving good pink color and polyester - wool blend in red color. Dyeing was performed by standard procedure for silk, wool and polyester dyeing. Good colorfastness of the dyed substrates to washing, sublimation and rubbing was observed but the color easily faded when dyed material was exposed to light. However since the dye displayed an antimicrobial activity against Staphylococcus aureus,


Corynebacterium diphtheriae, Nocardia spps and Micrococcus luteus, its commercial value increases (Murugkar et al., 2006). Production of indigo dye by genetic engineering and fermentation: In 1983, when scientists at Amgen Corp. (thousands Oaks, California, USA) were experimenting with enzymes engineered to eat moth balls, purely by accident, discovered that the enzymes were excreting a blue, indigo-colored dye. Hence, the company patented a technique involving the growth of genetically altered E. coli cells in a special nutrient medium which results in the excretion of indigo dye. According to them it had a significant advantage over the synthetic dyes and believed that this indigo would be regarded as Natural Indigo as it involves fermentation. Indigo synthesis has been achieved in transgenic bacteria Escherischia coli (Murdock et al., 1993). The first strains that could synthesize indigo were made in the early 1980s but they could not be used as the basis for efficient production. Murdock has prepared a strain that involves the genetic manipulation of nine genes and a modified fermentation medium, enabling more efficient synthesis directly from the glucose substrate. Molecular genetic techniques can be used to improve the stability, activity and yield of the biosynthetic pathways (Ensley and Chimia, 1994). A cluster of five genes comprising the tryptophan biosynthetic operon and an altered trpB gene causes high levels of indole synthesis from glucose. Four further genes code for enzymes that oxidise the indole to indigo, which is released into the medium. This method of synthesizing 'natural' indigo is regarded as potentially cost-competitive if successfully scaled up. An indigo producing strain of Pseudomonas putida has been biochemically characterized, which releases indigo, indirubin and isatin into the medium from indolecarboxylic acids (Eaton and Chapman, 1995). A gene coding for indole oxidase, catalysing an important step in the synthesis of indigo from indole, has been 33

cloned into Streptococcus themzophilus (Solaiman and Somkuti, 1996). This research is helping to provide an understanding of the biosynthetic pathways and regulation of indigo production. In time bacterial production may come to be a valuable alternative to agricultural crops (Hill, 1997). Among the microbial indigo producers, the majority of micro-organisms are aromatic hydrocarbon degrading bacteria such as naphthalene-degrading Pseudomonas putida NDO (Murdock et al. 1993), Ps. Putida PpG7 (O'Connor and Hartmans 1998), p-cumate and mand p-toluate-degrading Ps. putida F1 and Ps. putida mt-2, respectively (Eaton and Chapman 1995), styrene-degrading Ps. putida S12 and CA-3 (O'Connor et al. 1997) and toluene-degrading Ps. mendocina KR1 (Yen et al. 1991). The enzyme system responsible for indigo formation generally consists of one or more enzymes, typically monooxygenases, dioxygenases or hydroxylases (O'Connor et al. 1997; Doukyu et al. 1998). The genes encoding these enzymes have been cloned from Pseudomonas and Rhodococcus spp. into Escherichia coli strains (Yen et al. 1991; Hart et al. 1992; Eaton and Chapman 1995) so as to produce indigo directly from either nutrient medium (Ensley et al. 1983) or from glucose (Murdock et al. 1993). Recently, a method has been patented by O'riel and Kim (1998) for producing indigo and indirubin dyes using a recombinant Escherichia coli containing a gene encoding a phenol hydroxylase from Bacillus stearothermophilus (Bhushan et al., 2000). A gram negative rod SCV1 was isolated from oil contaminated garage soil. This bacterial strain was used for the production of indigo- a commercial dye after induction on xenobiotics like diesel, naphthalene and salicylate. The bacterial strain SCV1 was hydrophobic in nature as evident from hydrophobicity measurements. Hydrophobic nature gives the advantage to the bacterial strain in adhering to the hydrocarbons.


The results of indigo production by different substrates induced bacterial strain SCV1 suggest that the diesel induced maximum at 1.75 and 2mM concentrations. It is also suspected that uninuduced culture i.e. SCv1 enriches on nutrient broth produced other indigoid compounds other than indigo
(Mutnuri et al., 2009).

FUNGUS: Fungi cultivation certainly offers good prospects for the production of colorants. Normal fungi growing in their natural environment are being used by increasing numbers of crafts dyers (this evolution and the art of dyeing with fungi. But in addition, it is to be remembered that the colorants extracted from lichens, which are famous for their light- and wash fastness, are, in all cases, produced by the fungal partner of the symbiont. If scientific research could result in the successful cultivation of the fungi responsible for the beautiful lichen dyes, a large new class of high quality colorants would become available.

Production of anthraquinone: The anthraquinone compounds were isolated as aglycones from the ectomycorrhizal fungus Dermocybe sanguinea. The endogenous -glucosidase of the fungus was used to catalyse the hydrolysis of the O-glycosyl linkage in emodin- and dermocybin-1--D-glucopyranosides. The method, in which 10.45 kg of fresh fungi was starting material, yielded two fractions: 56.0 g of Fraction 1 (94% of the total amount of pigment,) consisting almost exclusively of the main pigments emodin and dermocybin, and 3.3 g of Fraction 2 (6%) consisting mainly of the 35

anthraquinone carboxylic acids. The anthraquinone compounds in Fractions 1 and 2 were separated by one- and two-dimensional thin-layer-chromatography (TLC) using silica plates, applying n-pentanol-pyridine-methanol (6:4:3, v/v/v) and toluene-ethyl acetate-ethanolformic acid (10:8:1:2, v/v/v/v) as eluents. Fifteen different anthraquinone derivatives were completely separated from one another. Emodin, physcion, endocrocin, dermolutein, dermorubin, 5chlorodermorubin, emodin-1--D-glucopyranoside, dermocybin-1--D-glucopyranoside and

dermocybin, and five new compounds, not earlier identified in D. sanguinea, 7-chloroemodin, 5,7dichloroemodin, 5,7- dichloroendocrocin, 4-hydroxyaustrocorticone and austrocorticone, were separated and identified on the basis of their Rf-values, UV/Vis spectra and mass spectra. One substance remained unidentified, because of its very low concentration (Raisanen, 2002)

Production of anthraquinone dyes by liquid cultures: Researchers are currently investigating the production and evaluation of microbial pigments as textile colorants. Fungi are more ecological interesting source of pigments, since some fungal species are rich in stable colorants, such as anthraquinone. Two anthraquinone compounds are described by researchers at National Research Center, Dokki, Cairo, Egypt which were produced by liquid cultures of Fusarium oxysporum, isolated from the roots of citrus trees affected with root rot disease. These anthraquinone compounds are 2-acetly-3,8-dihydroxy-6-methoxy anthraquinone or 3-acetyl-2,8-dihydroxy-6-methoxy anthraquinone. Dyeing of wool fabrics with these new anthraquinone compounds as natural dyes has been studied. The values of dyeing rate constant, half time of dyeing and standard affinity have been calculated and discussed. The effect of dye bath pH, salt concentration, dyeing time and temperature were studied. 36

Color strength values and the dye uptake were high. The results of fastness properties of the dyed fabrics were good. Thus, anthraquinone compounds which were produced by stationary cultures of F. oxysporum could be used for dyeing wool with good fastness properties and high dye uptake. They can serve as a noteworthy source of raw material in the future (Nagia FA and EL Mohamedy RSR, 2007).

Production of microbial pigments by cost effective methods: A study focused on isolation, identification of pigment producing bacteria, basidiomycetes fungi and extraction of pigment for dyeing. The growth, pigment production, optimization and characterization of 3 colors (red, yellow and pale green) from different bacteria such as Serratia marcescens, Pseudomonas fluorescens and Erwinia amylovora have been standardized on cost effective natural medium. Among the 3 pigmented bacteria Serratia marcescens was studied extensively and developed a cost effective natural solid medium containing coconut endosperm. The pigmented bacterium was harvested within 48 hrs and the methanolic extracts yielded red pigment (55% bacterial dry eight basis). The production and extraction of pigment from Serratia marcescens was evaluated for its cost benefit analysis. The pigment was produced form bacteria and extracted with an investment of Rs. 11.70 / litter of bacterial cultures. The interaction of the bacterial pigment on silk and cotton fabrics were stronger and did not fade even after exposing to sunlight drying and repeated washing whereas the paper pulp added with bacterial pigment comparatively lost its color fastness during drying and exposure to sun light.


The fungal pigment (Ganoderma lucidum wild collection) and the pigment extracted from the dry barks of Accasia melanoxylan applied on the paper pulp resulted in deep color shades. The color shades in paper sheets were not altered when exposed to sunlight drying. Twenty two mushrooms were collected, identified and extracted for pigment production, of which 14 mushrooms were collected. The fungal basidiocarp used for pigment extraction yielded different shades of yellow (Boletus edulis,), orange (Armillaria tabescens) and brown (Armillaria tabescens) color. The pigment extracts tested for dyeing in cotton yarn showed their fastness, suitability for dyeing and paves an innovative cost effective method for the use of microbial pigments for dyeing cotton and silk fabrics (Perumal et al., 2007).


What is the market for naturally dyed textiles? At present naturally dyed articles fall into small niche markets fed by craft workers and small commercial firms, and tend to be costly But what is todays small niche market can become a larger market tomorrow, as has been shown for herbal teas and natural cosmetics. There has been little attempt to define and predict the market for naturally dyed products. As always, it can be very speculative to try to describe the potential market for a new type of product. Such findings as have been published indicate that there could be a sigruficant demand for natural dyes, rising to 510% of all textiles in the next century (Das, 1994). If the world market for all dyes is about 800000 tones per year at a value of about 2.8 billion (Glover and Pierce, 1993) and if we assume about 5% could be an eventual market for natural dyes, we could predict a world market for natural dyes of about 40000 tones per year. This kind of perdition has been used to justify the funding of research by the EU. In market research, information has to be factual, usually based on data collected and used to predict future trends. Yet with natural dyes, one of the facts appears to be that most consumers are unaware of the possibility of products dyed with natural substances. The media are confused about what is meant by the term natural dyes. Many, if not most, customers would not understand how to answer simple questions such as Would you purchase naturally dyed textiles if they were available? They may not be aware of what textiles dyed with natural dyes are like, or they may erroneously believe that natural dyes are all very expensive, pale wishy-washy colors that fade. Indeed it could be argued that if textile designers specified textiles dyed with natural dyes, a demand for their production would materialize. But of course designers cannot use textiles that are not available, so the industry may need a kick start to break this catch 22 problem. 39

What are the benefits of natural dyes to the consumer? Benefits may be various and partly emotive. The following might need investigating: The textile product is made from totally natural products
Ranges of articles dyed with natural dyes tend to have a characteristic appearance; a

designer could make use of the way the colors harmonize well together for natural styles The product is manufactured without using toxic or noxious substances The product is made in an environmentally friendly way without reliance on nonrenewable resources
Articles can be represented as being unique, e.g. with slightly uneven dyeing or with

variations in shade from one batch to another, or fade characteristically in use. The principle must prevail by which a customer can be shown two identical products at the same price, one dyed with natural dyes and the other with synthetics, which he/she can most likely to choose (Hill, 1997).


Dyes currently used for dyeing textile material are classified as soluble, disperse, and pigments (Rivelin, 1992). These are all synthetic compounds, which are environmentally unfriendly compounds, as their degradation by organisms is not carried out naturally. The industry has to design expensive ways to remove these harmful compounds from the environment. Availability of natural dyes is a desired technology for dyeing fabrics with naturally produced compounds (Sengupta and Singh, 2003).


Evaluation and improvement of present and potential sources.

Feasibility of combining developments in textile technology with natural dyes. Exploiting inherent properties of natural dyes for specialty applications. To produce natural dyes in the quantities required. To produce natural dyes at a reasonable price.
To produce natural dyes that has high color-fastness.



The amount of research effort devoted to natural dyes is negligible. If there had been significant research on the use of natural dyes, it is probable that they would already be much more widely used than they currently are. As there is much catching up to do after 150 years of neglect, there is plenty of scope for rapid developments. This applies to the techniques of agricultural production and processing as well as to dyeing itself. A medium- to long-term view of research and development is needed. It is unlikely that the total cost of the R & D required to make natural dyes a profitable commercial reality need be very great when compared with many industrial products. The main are as for future work must include market research, improved crop technology and processing, dyeing methods and quality assurance (Hill, 1997).


Although, there are many cited literature, wherein efforts have been made to exploit these ecofriendly bioengineered natural dyes for textile application, but there very few studies which have carried out systematically in depth investigation. R&D work is under progress in genetically modified micro-organisms and dyestuffs for the textile field. There is a vast resource of natural antimicrobial agents, which can be used for dying as textile substrates. The major challenges in application of such dyes for textile application are that most of biomaterial used as potential source is a complex mixture of several compounds and also composition varies from source to source. The availability of such products in bulk quantities, their extraction, isolation and purification to get standardized products, durability, shelf life and anti microbial efficiency are another challenges in their application. However, because of their ecofriendly nature and non-toxic properties, they are still promising candidates for niche application such as medical and health care textiles. The application of biotechnology in a wide range of industry sectors (chemicals, plastics, food processing, natural fiber processing, mining and energy) has invariably led to both economic and environmental benefits via processes that are less costly and more environmentally friendly than the conventional processes they replace. In effect, the application of biotechnology has contributed to an uncoupling of economic growth from environmental impacts. Biotechnological applications have been increasing the eco-efficiency of industrial products and processes can provide a basis for moving a broad range of industries toward more sustainable production. To achieve this, further development of biotechnology and supporting technologies 43

will be needed, as well as policies that provide incentives for achieving more sustainable production. The main driving forces for adoption of more efficient bioprocesses and byproducts are cost savings and improved product quality/performance. Environmental considerations were an important but secondary driving force. Successful biotechnology/bioprocess development requires effective management of technology development by companies and use of tools that assess both the economic and environmental performance of technology during its development. There is a need for improved assessment tools that are easier to use and at earlier stages of the technology development process. Even large companies may not have in-house all the expertise required to develop more efficient byproducts and bioprocesses. Collaboration with university and government researchers and other companies is an important contributing factor for successful introduction of these products and processes. Long lead times are often required for introduction of paradigm shift technology into a company; but development times can be reduced considerably in subsequent development cycles. The application of biotechnology for developing industrial products and processes is still in its infancy. As awareness builds and the technology continues to be developed and diffused through different industry sectors over the next few decades, the economic and environmental benefits are predicted to grow.


This note of caution needs to be echoed across the whole spectrum of biotechnology developments. Although biological systems after many attractive possibilities and new approaches to all sorts of problems and needs, considerable advances are still being made in conventional technologies, such as, catalysis, chemical synthesis and physical fiber modification which need to be kept in perspective. There is also still great concern in society about the unbridled advance of biotechnology, especially with regard to the modification of natural species with possible unknown long-term consequences.

I am thankful to Mr. Shaik Gore Mastan, Research Scholar, Discipline of Wasteland research, CSMCRI, Bhavnagar, Gujarat, for his guidance.



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