Colorimetric Measurement and Functional Analysis of Selective Natural Colorants Applicable for Food and Textile Products

Abstract

Colouration of textiles as well as food products with natural colorants is an interesting subject with respect to the growing eco-concern among the consumers. Several colorants are available in nature for textile colouration and are renewable, biodegradable, and eco-friendly. Being safe for human consumption, they can serve the dual purpose of also coloring food. Several such natural dyes are available. This review chapter deals with the chemistry, extraction, application, and colorimetric analysis of colorants derived from turmeric (root), annatto (seeds), and cochineal (insect) for use on both textiles and food products.

1. Introduction

From the application in textiles, uses of natural dyes also extend to colouration of food and in other areas like medicines, cosmetics, and procession of leather products. Several sources of natural colorants used in the past have been re-identified today. Many are common and play a dual role in colouration of textiles as well as food products and drinks. Some dye-yielding plants contain compounds like curcumin, crocin, bixin, carthamin, punicalagin, nimbin that are known to have therapeutic properties and are used in various traditional medicinal therapies. Their inherent functional properties like antimicrobial, antifungal, deodorizing, UV protection, moth/insect-repellent, and others allow them to enhance the value of the dyed textiles, or the colored food products. This chapter deals with some selected natural colorants widely used in the textiles and food sectors and documents their chemistry, extraction process, application, usage and properties, separately, in relation to textiles and food. Few case studies on colourimetric measurements and analysis of functional properties of natural dyes on textiles and food are also discussed.

2. Natural colorants for textile related application

Natural dyes for textiles are dyes or colorants derived from plants, invertebrates, or minerals. From the plant source, colors are extracted from seeds, roots, stems, barks, leaves, flowers, berries, and fruits. In addition to the natural vegetable coloring matter, animal/insect coloring matter like tyrian purple, cochineal, lac and kermes, and mineral coloring matter derived from ocher, limestone, manganese, cinnabar, azurite, and malachite are also used to produce natural effects on the fabrics. With the advent of synthetic dyes, natural dyes faded into oblivion. But now with several advantages like fast and durable colors coupled with replaceable, biodegradable, and fairly non-polluting nature over the synthetic ones, natural dyes are making a comeback.

Different natural dyes yield different colors–yellow (kamala seed pods, myrobolan fruit); mustard yellow (latex from the gamboge tree); yellow to orange (pomegranate rind, turmeric, and lichens); peach to brown (chestnut hulls); orange (gold lichen, carrot and onion skin); pink (berries, rose and beets); crimson to maroon (teak leaves and cochineal); orange, pink and red (madder root); red to brown (bamboo and hibiscus flower); brown (catechu bark and coffee beans); red to purple (red sumac berries, basil leaves, hibiscus flower, logwood, lac); purple (red cabbage and murex snails), blue (indigo leaves), green (sorrel roots, spinach, and peppermint leaves); yellow, gray to black (black berries, iris root, and walnut hulls) and sepia brown (octopus/cuttlefish).

Different compounds are present in natural dye sources that impart a variety of colors on textiles; indigotin (blue and purple), anthraquinones (shades of red), carthamin from safflower (red and yellow shades), naphthoquinone (orange, red, or reddish-brown shades), flavonoid dyes (yellow to greenish-yellow and brown colors), carotenoid (orange), tannins (different colors with different mordants) and curcumin (yellow shades).

3. Natural colorants for food related application

Color is the prime sensory attribute of foods and is often used by consumers as an indicator of food quality in terms of flavor, safety, and nutritional value. Food colors are dyes, pigments, or other substances that impart color when added to a food product or a drink. Such additives make the food more attractive, appealing, and appetizing; provide color to colorless foods or enhance their natural color; offset color that is lost on exposure to air, moisture, high temperature, light, and unfavorable storage conditions; and allows the consumers to identify products on sight. Thus, one of the main applications of food colorants is the modification or preservation of its visible appearance.

Food colors can be obtained naturally as extracts from natural sources, or they can be synthesized. Natural food colors are usually extracted from seeds, fruits, vegetables, leaves, insects, algae, etc., and are used both in domestic cooking and commercial food production and are available in many forms such as liquids, powders, gels, and pastes.

Among the natural food colorants, Asian spices like turmeric and saffron are used in everyday cooking; they lend an appeasing color to the food. Saffron, as a spice finds its use in biryanis and as colorants in dairy products. Caramel is mostly used to enhance flavor in deserts. Hibiscus is a commonly used bakery product and tea-based beverage to enhance the brown tint. Marigold does not have extensive use but the petals are sometimes used to enhance colors in salads. Beet juice has several applications in many beverages, dairy products, yoghurt ice cream, sauces, jams, jellies, and candies.

Different sources of natural colorants yield different colors; dark yellow is obtained from turmeric; yellow-orange from saffron; orange from carrots, red pepper/paprika, and sweet potato; pink from strawberries and raspberries; red from carrot, beets, and tomato; deep red from beetroot and red sandalwood; green from matcha and spinach; blue from red cabbage mixed with baking soda; purple from blueberries and purple sweet potato; brown from coffee, tea, and cocoa; and black from activated charcoal and squid ink.

A variety of compounds present in natural dye sources are responsible for different colors. Anthocyanins (flavonoids) found in fruits and vegetables are responsible for blue, purple, red, and orange colors. Carotenoids in fruits and vegetables are known for imparting red, orange, and yellow colors. Betalains present in most caryophyllales plants give a pink to red color. Curcumin is responsible for the yellow color of turmeric. Safflower gives an attractive yellow color. Chlorophylls from alfalfa (Medicago sativa) are responsible for the characteristic green color. Carminic acid in carmine from cochineal is responsible for dark purplish-brown or bright red or dark red color.

4. Colorimetric measurement of natural colourants

The appearance of a textile or food material is ascertained through its surface color and is the first sensation perceived by the consumer to judge its acceptability. The color of an opaque object is described by the reflectance of light as a function of its wavelength. The human eye is versatile and can detect light and light modification by the colorant and this is interpreted by the brain as color. For any color to be perceived by a human eye, a source of light, an object, and an observer is required.

Color measurement of products can be carried out in two ways; by visual evaluation or through instrumental analysis. The chromatic attributes and different geometric factors like texture, shape, etc. of foodstuffs can be assessed qualitatively by the human eye. In this process, the observer assesses the color of the sample under standard conditions of illumination, and after comparison with defined color standards; the assessment is defined in terms of some scores generally on a 9-point scale. One of the most popular scales is the 9-point Hedonic scale in which the products can be marked from 1 to 9 depending on the appearance and acceptability rate of the food product. A lower score indicates low and least acceptable color intensity; while a high score denotes high color intensity or acceptable appearance. Such visual assessment is subjective, relative, and is dependent on the observer and environmental conditions. On the other hand, the presence of color pigments can be also be quantitatively assessed using different types of equipment. But each instrument measures only one attribute at a time and so several instruments may be needed to measure various aspects of visual perception. Basically, there are three types of instruments that measure color or its attributes, colourimeter, spectrophotometer, and spectroradiometer.

Liquid chromatography is a method for separating, identifying, and quantifying the constituents of a mixture. The interaction of the sample with the mobile and stationary phases causes this separation. Because there are so many distinct stationary/mobile phase combinations that can be used to separate a mixture, chromatography is divided into various categories based on the physical states of those phases, liquid, and gas. Liquid–solid column chromatography is the most common chromatography technique that uses a liquid phase (mobile) that filters down through the solid stationary phase, bringing the separated components with it. To separate the components that make up a sample, high-pressure liquid chromatography (HPLC) uses pumps to push a pressurized liquid solvent containing the sample mixture through a chromatography column loaded with solid absorbent materials. Each component in the sample interacts with the adsorbent material in a slightly different way, resulting in varying flow rates and separation of the components as they flow out of the column. The type of chromatography column employed determines how different chemicals are separated. Several different types of columns (size exclusion, ion exchange, normal phase, reverse phase) are used. Once the molecules make it through the column, they will be detected by a detector, which is typically a UV detector, but other detectors such as refractive index detectors, laser light scattering detectors, fluorescence detectors, and thermal conductivity detectors are also used. High-performance liquid chromatography (HPLC) is considered the ‘gold standard’ for measuring pigment concentrations in plant samples. A major drawback of this process is its high cost both in terms of time required for assessment, and the high cost of the testing equipment itself. Liquid chromatography can be combined with mass spectrometers (LC–MS) to analyze organic and inorganic compounds of biological origin. While liquid chromatography may separate mixtures with several components, mass spectrometry can identify the individual components’ structural identity with high molecular specificity and detection sensitivity.

Colorimetric or spectrophotometric analysis is another technique to evaluate color in textiles or food. Because the amount and color of light absorbed or transmitted through a solution is dependent on the concentration of pigment particles present in it, such measurements rely on detecting the concentration of material (color/pigment) in a solution. Such color evaluation measures the change in the intensity of electromagnetic radiation in the visible wavelength area of the light spectrum after it is transmitted or reflected by the object or solution through which it passes. A colorimeter or spectrophotometer thus assesses the color in various sample solutions (dyes in textiles, or colorants in food) by absorbing a particular wavelength of light and denotes the assessment in the form of some values using the Beer–Lambert law. Under Beer’s law of photometry, the amount of light absorbed is proportional to the solute concentration present in the solution. According to Lambert’s law, the amount of light absorbed is proportional to the length as well as thickness of the solution taken for analysis or in other words, when light passes through a medium, its absorption is proportional to the intermediate convergence. Beer’s law and Lambert’s law are usually taken in combination as Beer–Lambert law which indicates the relationship of absorbance with both the path length of light inside the sample and the concentration of the sample.

Thus, the principle of operation of a colorimeter is outlined as follows—in a colorimeter a beam of light of a given wavelength is directed toward a liquid sample (of the dyes in textiles, or colorants in food). While passing through a solution in the colorimeter, the beam of light travels through a series of lenses, and the photocell is able to detect the amount of light passing. The current produced by the photocell depends on the quantity of light striking on it; higher the concentration of the colorant/pigment in the solution, the higher is the absorption of light and consequently less transmission. Thus, less light passing through the solution would indicate the creation of less current by the photocell [1]. The colorimeter can qualitatively detect the presence of color pigment in a sample when the wavelength peak detected in the experimental sample matches with the peak (λ_max) of the standard pigment.

The colorimeter can also measure the amount of pigment present in the sample. In this case, calibration curves can be made using the different concentrations of the standard solution of the pigment. With the help of a calibration curve, the amount of pigment present in the sample can be estimated. In case standard solutions are not present, then various equations can be formulated using extinction coefficients, molecular weight, etc. to ascertain the amount of dye pigment in the sample.

When items are viewed under different sources of light and illuminations, their colors are frequently diverse. The discrepancy is due to differences in the spectral power distribution of the illuminations as well as changes in the lighting. An illuminant is a specific spectral power distribution incident on the object viewed by the observer, whereas a source is a physical emitter of radiant energy, such as a lamp or the sun and sky. As a result, a single source of light can provide several illuminants. Illuminants can also have a variety of spectrum power distributions. Numerical specification of color was earlier visualized by chromaticity diagram and the three chromaticity coordinates (x, y, and z) were calculated by the use of the three tristimulus values that represent the amount of standard lights (red, green, and blue) required to reproduce a color.

Over time, a slew of alternative color appearance models have arisen, as well as a numerous new color measurement related terms. To represent the color of an item, several color coordinate systems can be employed, including RGB (red, green, and blue), Hunter Lab, Commission Internationale de l’Eclairage’s (CIE) L*a*b*, CIE XYZ, CIE L*u*v*, CIE Yxy, and CIE LCH. Almost of modern color measurement is based on experimental observations in accordance with the CIE (International Commission on Illumination) color specification system. The human eye has three color receptors: red, green, and blue, according to CIE principles, and all colors are combinations of these.

Color evaluation methods such as the Hunter Lab L*,a*,b* and the modified CIE system known as CIELAB are widely used in the food and textile industries. They were created as a result of investigations that correlated tristimulus values with visual perceptions of color in order to convert the X, Y, Z system (tristimulus values) to a visually uniform color-system. Each color can be considered equivalent to a member of the greyscale lying between black and white, according to L*, which is an approximate measurement of brightness. As a result, the L value for each scale reflects the level of lightness or darkness, whereas the a and b values indicate redness or greenness, respectively. Hunter L, a, b is a color scale based on the opponent-color theory which states that color receptors in the human eye see color as pairs of opposites: light–dark, red-green, and yellow-blue. To fully define the color of an object, all three values are required. The scale consists of two color coordinates, a* and b*, as well as a psychometric index of lightness i.e. L*. The parameter a* is positive for reddish colors and negative for greenish colors, whereas the parameter b* is positive for yellowish colors and negative for bluish colors. L* is an approximate measurement of luminosity according to which each color can be considered as equivalent to a member of the greyscale lying between black and white. Thus, the L value for each scale, therefore, indicates the level of lightness or darkness; the values indicate redness or greenness, and the b values yellowness or blueness. The CIE 1976 L*a*b* color or modified CIE system called CIELAB was recommended by the CIE in 1976 to improve on the 1966 version of the Hunter L, a, b. The CIELAB color scale, like the Hunter, expresses color as three values: L* for perceived brightness, a* and b* for the four distinct hues of human vision: red, green, blue, and yellow. Under the two color scales, however, three values of L, a, and b are determined differently; the formulas for Hunter L, a, and b are square roots using CIE XYZ, whereas CIELAB uses cube roots of XYZ. The CIELAB color scales were designed to be a perceptually uniform space in which a given numerical change correlates to a corresponding perceived change in color, and so provides a better approximation to the visual judgment of color difference for very dark hues. Despite the fact that the LAB space is not genuinely perceptually uniform, it is valuable in the industry for detecting minute color changes. Because the CIE L*a*b* scale, which was released in 1976, has gained popularity, the Hunter color scale is no longer as widely used as it once was. Although CIE measured the single color space, it was not truly uniform visually throughout the color space and could not define color-difference in a singular term i.e. two colors cannot be red and green at the same time or yellow and blue at the same time. It meant that equal color difference magnitude appear of different visual magnitudes in different regions of the color space. For this reason, the CMC equation (Color Measurement Committee) or color difference (ΔE* or DE*) formula which takes the non-uniformity of the color space into account is used to assess the difference between two colors and is more preferred in textiles color assessment today. The CMC equation corrects the CIELAB color scale’s most significant flaw, which is chroma location dependency.

The total color difference, ∆E, may also be calculated. A comparison of two colors is used to determine this color difference (ΔE* or DE*). One is designated as the standard (or target), and the other as the sample. ∆E is a single value that takes into account the differences between the L, a, and b of the sample and standard. The delta values (∆L, ∆a, and ∆b) show how far a standard and sample differ in terms of L, a, and b. Different color difference formulae are used to calculate the numerical color difference between two colors.

• ΔL* (L* sample - L* standard) = difference in lightness & darkness (+ve = lighter, −ve = darker)

• Δa* (a* sample - a* standard) = difference in red & green (+ve = redder, −ve = greener)

• Δb* (b* sample - b* standard) = difference in yellow & blue (+ve = yellower, −ve = bluer)

Deltas for L* (ΔL*), a* (Δa*) and b* (Δb*) may be positive (+) or negative (−). Whether the sample is redder or greener than the standard is indicated by the sign of the delta value. For example, a sample will be redder than the standard if ∆a is positive. The total difference, Delta E (ΔE*) is always positive. For the delta values, tolerances can be established. Out-of-tolerance delta values indicate that the discrepancy between the standard and the sample is too great. If ∆E is out of tolerance, it is difficult to know the parameter that is out of tolerance. It can also be deceiving in situations when L, a, or b are out of tolerance but E is still within it.

Color values of textiles are also assessed in terms of K/S (Kubelka-Munk) values where higher values represent darker and more saturated colors. K/S values are usually calculated at the wavelength of maximum absorption of the color (λ_max); however, a calculation over the visible region may also be employed. The Kubelka-Munk equation is as follows:

$$\text{K/S}=\frac{{\left(1,-,R_{\text{λmax}}\right)}^2}{\text{2R}{\lambda }_{\text{λmax}}}$$E1

Where K: is the constant related to light absorption of the dyed fabric; S: is the constant related to light scattering of the dyed fabric; R: is the reflectance of the colored fabric that is expressed in fractional form.

The objective measurement of color is thus dependent on the quantification of the light source (E), the object’s reflectance (percent R), and the observer’s color response functions r-g-b. In food products, color quality is either measured on a spectrophotometer and expressed in terms of the chromatic attributes (L*, a*, b*) as proposed by CIE, or in terms of tint values measured using a tinctometer and interpreted as color ratio between yellow and red pigments (R and Y values). Colors on textiles can be characterized by hue (dominant shade); the amount of color present or saturation; and by the degree of lightness or darkness of the particular color. Thus in textiles color values are generally expressed in terms of the color strength (K/S values), color difference (ΔE), chromatic attributes (L*, a*, b*), as proposed by CIE and Metamerism Index (MI). Based on the respective magnitudes of ΔE, ΔC, ΔH, MI, a newer empirical index CDI (Color difference index) of assessing color for a binary mixture of dyes has also been postulated [2].

5. Some selected colourants commonly used for colouration of textiles and foodstuffs

5.1 Turmeric

Turmeric is derived from the tuberous rhizome of the Zingiberaceae family. Curcuma longa, the yellowish-brown rhizome from which the turmeric is derived develops beneath the ground and is cylindrical, tuberous, highly branched with a rough and segmented skin with a dull orange interior. The leaves are pointed and the flowers are funnel-shaped and yellow in color. C. longa is a perennial herbaceous plant that grows wild in tropical Asia. India is the largest producer, consumer, and exporter of turmeric in the world contributing 78% followed by China, Myanmar, Nigeria, and Bangladesh together contributing to 6% of the global production. Dried the turmeric rhizome gives yellow powder with a bitter, slightly acrid, but sweet flavor. C. longa is a medicinal plant that is used extensively in textile and food colouration. It is popularly used as a spice in South Asian and Middle Eastern cuisines as it lends a distinctive yellow color and flavor. C. longa also possesses antioxidant, anti-inflammatory, choleretic, antimicrobial, and carminative properties and has been used in traditional Indian ayurvedic medicine. The dye has been used to color fabrics in brilliant yellow colors. It can be used in combination with other plants like Butea monosperma flowers [3] or Nyctanthes arbor-tristis flowers [3] to give a range of yellow shades. It’s typically used as a foundation color for indigo overdyeing to achieve a fast green.

Genus: Curcuma | Species: longa | Family: Zingiberaceae.

Common name: Turmeric | Local name: Haldi.

Part of the plant used for coloring: Roots/rhizomes and leaves.

5.1.1 Coloring pigment/component

Turmeric has a volatile oil that contains turmerone, as well as other coloring compounds called curcuminoids mainly concentrated in the rhizome. Curcuminoids (1,7-bis 4-hydroxy-3-methoxyphenyl-1,6-heptadiene-3,5-dione) are natural antioxidants and curcumin is the principal curcuminoid present in turmeric. The other two curcuminoids are desmethoxycurcumin and bis-desmethoxycurcumin. Curcumin is a polyphenol and the principal coloring component of this yellow dye which has been also been classified as CI Natural Yellow 3 and considered a direct type of dye. Curcumin can be found in two different tautomeric forms: keto and enol. In the solid-state and in solution, the enol form is more energetically stable [4]. The chemical structure of curcumin is different under different pH and hence it can be used as an indicator. It remains yellow in an acidic medium, while when added to an alkaline medium above pH 8, the shift of the hydrogen atom causes the compound to change color giving a red hue. It is not soluble in water (acidic and neutral pH) at room temperature but is soluble in oil and alcohol. Curcumin also has fluorescence qualities, which extends the active life of these molecules and increases the chances of contact with oxygen in the air, making them more susceptible to photochemical oxidation. [5]. A relationship exists between the curcumin content and the L*a*b* values [6] and high curcumin content is associated with high L* (lighter) and b* (yellower) values, but with lower a* (less red) value. Where a* and b* values are high, the resultant shades are red and yellow respectively, while when both a* and b*values are similar, the resultant shade is orange (Table 1).

Curcumin content in different types of turmeric	L	a*	b*
3.5	32.6	39.1	31.5
3.8	36.6	28.4	36.1
4.3	46.3	22.1	42.0
5.1	54.7	17.5	46.1

Table 1.

Variation in color values with respect to changing curcumin content in turmeric taken from difference sources.

5.1.2 Application of turmeric in textile coloration

Very few studies have been reported on dyeing of textiles with turmeric. Cotton was dyed with purified ethalonic extract of turmeric by the exhaust technique [7]. Enhancement of dye uptake and wash fastness of cotton was achieved through modification with enzymes and chitosan [8], irradiation with gamma rays [9], and microwaves [10] before dyeing. Silk was dyed with Curcuma Longa L rhizome in brilliant shades [11] and improved dye uptake and fastness were obtained on silk pre-irradiated with methanolic extract of C. longa L rhizome [12]. Nylon dyed with turmeric gave fast colors [13].

5.1.2.1 Color produced

Turmeric yields a warm gold color on undyed natural cotton fabrics, silk, and wool. It gives a wide range of yellows without mordants. With mordants (metal salts), it gives colors like golden yellow (tin), mustard yellow (copper and chromium), and olive green (iron). Its wavelength of maximum absorption (λ_max) is 420 nm [14] or 450 nm [15] indicating that the dye can absorb color in the blue end of the spectrum. The wavelength of maximum absorption for turmeric is.

5.1.2.2 Extraction

Maximum yield (highest absorbance) of color from turmeric was obtained at pH - 6 at 100°C [16] indicating that the dye can be extracted under very mild acidic or neutral conditions. Also, maximum extraction occurs at high (boiling) temperatures [5]. The solvent extraction process gave maximum yield followed by aqueous extraction, but the purest form was obtained by spray drying [14].

5.1.2.3 Dyeing conditions

Color strength (K/S) value of the dyed fabric was maximum in pH 7 [7]. Good color strength was observed by dyeing fabric irradiated at 65°C for 40 min in dyeing bath having pH 6 [10]. Glauber’s salt tends to neutralize or reduce the negative electric charge (zeta potential) of cotton fabric, thus facilitating the approach of the dye anions to the fabric within the range of formation of hydrogen and other bonds between the dye molecules and fabric and thus the color strength of cotton dyed with turmeric extract increases with increase in salt concentrations [5].

5.1.2.4 Fastness

In general, turmeric is a fugitive dye and bleeds easily. Turmeric exhibits poor washing fastness due to the phenolic groups present in curcumin which reacts with soda ash (in washing liquor) forming curcumin salt that is soluble in water and hence can be easily washed out from the dyed fabric. The poor light fastness of turmeric is attributed to the inherent susceptibility of its chromophore to photochemical oxidation. However, both the wash and light fastness of textiles dyed with turmeric can be improved through mordanting. The improvement in light fastness can be attributed to the reduced susceptibility of the turmeric dye chromophore to photochemical oxidation in the presence of mordant. Though dyeing with turmeric exhibits good fastness to rubbing, a decrease is noted both in the dry and wet rubbing fastness in the presence of the mordant.

5.1.2.5 Functional properties of turmeric related to textile application

Turmeric also has antibacterial and anti-inflammatory effects. Natural colorants extracted from turmeric exhibited excellent antimicrobial activities and related wound healing properties [17]. Silk fabrics dyed with an extract from C. longa rhizome using copper sulphate, ferrous sulphate, and potassium aluminum sulphate as pre-mordants possessed desirable antibacterial properties and 3% (owf) copper sulphate giving complete antibacterial activity against Staphylococcus aureus (Gram-positive) and Escherichia coli (Gram-negative) [11]. The study also indicated that an increase in the dye concentration leads to a more efficient antibacterial activity and 30% (owf) of turmeric gave the optimum level of antibacterial activity. Nylon fabric dyed with various concentrations of turmeric extract using different metallic mordants displayed excellent antibacterial activity in the presence of ferric sulphate, cupric sulphate, and potassium aluminum sulphate, and exhibited good and durable fastness properties [13]. Cotton yarns colored with turmeric and coated with chitosan provide high antibacterial action against bacteria (E.coli and S.aureus). Also, the yarn coated with chitosan dyed to a darker shade compared to uncoated yarn for the amount of the dye used [18]. Colorant from turmeric can have UV protection properties and can block almost 100% UV-rays when used to dye polyester. On coating the fabric with chitosan there was no change in UV protection property though the slight change in the shade was noted [19].

5.1.2.6 Case study-1

Turmeric (C. longa L.) extract was used to dye cotton using bio-mordants (Citrus limon and Colocasia esculenta), and for comparative purposes, metallic mordants (potassium dichromate and potash alum) were also used [20]. The samples were pre-mordanted (soaked) in the bio-mordant extract for different durations before dyeing; for the metallic mordants, they were boiled with the mordant solution at 80°C for 50 min followed by cooling for 60 min in the solution itself. The effect of mordanting time on the color strength was evaluated for the bio-mordants. The surface color strength (K/S) of bio-mordants (Citrus limon and Colocasia esculenta) pre-mordanted cotton increased with an increase in mordanting time for both the bio-mordants (Table 2). Cotton pre-mordanted with lemon containing significant amounts of tannins showed the highest surface color strength (K/S) among all mordants used. The effect of moisture absorption on the hue of the dyed fabric was also studied. For this, the dyed specimens were stretched and tied over the mouth of steel tubes containing 100 ml water each. The specimens were maintained under normal conditions of 25°C and 70% relative humidity for 24 h after which the face and rear side of the specimens were visually observed for any change in hue. Under acidic conditions (below pH 4), curcumin gave a yellow appearance, in alkaline pH, it changed its hue as the dyed cotton specimen absorbed moisture changing its pH and thus showing a significant change in hue on both side of the fabric. Furthermore, the visual uniformity of the dyed samples was found to be excellent for both bio-mordants. Due to the presence of citric acid, turmeric gave uniform color in low acidic conditions (around pH 4); at higher pH (pH 4 to 5) it showed a reddish color. Color fastness to rub (dry and wet), water (EN ISO 105 E01–2013), wash (ISO 105 C06), and perspiration (EN ISO 105 E04–2013) were found to be superior for the bio-mordanted cotton and the values ranged from 3 to 4–5 in most all cases.

Treatments	Mordanting time
	1 hour	3 hours	5 hours
Unmordanted sample	4.0
Cotton pre-mordanted with Colocasia	4.1	4.5	5.1
Cotton pre-mordanted with Lemon	7.0	7.3	8.6
Cotton pre-mordanted with potassium dichromate	7.5*	—
Cotton pre-mordanted with potash alum	4.0*	—	—

Table 2.

Surface color strength of cotton dyed with turmeric pre-mordanted with different mordants for different time duration.

^*

for 1 hr. 50 min

5.1.2.7 Case study-2

Aqueous extract of turmeric was used to dye cotton fabric using aluminum sulphate as a mordant [15]. The effect of different mordanting techniques (per, post, and simultaneous) on the surface color strength of the fabric was evaluated (Table 3). Simultaneous dyeing and mordanting sequence gave maximum dye uptake probably due to the mordanting of cotton with aluminum sulphate mordant and formation of a complex between the color component of the dye curcumin and the metal mordant. Also, turmeric being a direct type of dye exhausted well in the presence of a salt-like alumnium sulphate (mordant) and hence simultaneous mordanting sequences gives better results (K/S).

Mordanting Technique	K/S at λmax (450 nm)
Pre	0.4
Post	0.3
Simultaneous	1.5

Table 3.

Surface color strength (K/S) of cotton dyed with aqueous extract of turmeric using aluminum sulphate as a mordant by the different mordanting sequences.

New and uncommon compound shades were developed through combination dyeing of the cotton combination of turmeric (yellow dye) with using madder (red dye), and turmeric (yellow dye) with red sandalwood (red dye) in different proportions by the different mordanting and dyeing process. A synergistic effect in the color interaction between the observed and calculated K/S values (calculated values were derived by adding the individual K/S value of the respective proportion of the two dye components on the fabric) was observed; the observed K/S values of the dyed cotton samples were always higher than the calculated or expected K/S values indicating the color value of the mixed dye system to be always higher. Also, an increased amount of turmeric in the mixture increased the dye uptake (K/S) values (Table 4).

Dye	Amount of dye when used singly					Proportional ratio of the dye in the mixture	Calculated value for the combined shade	Observed value for the combined shade
	100	75	50	25	0
Turmeric	—	0.7	0.3	0.2	—	100:0	—	1.5
Madder	—	0.3	0.3	0.4	—	75:25	0.7 + 0.2 = 0.9	1.0
						50:50	0.3 + 0.3 = 0.6	0.9
						25:75	0.2 + 0.4 = 0.6	0.8
						0:100	—	0.6
Turmeric	—	0.7	0.3	0.2	—	100:0	—	1.5
Red sandalwood	—	0.3	0.3	0.2	—	75:25	0.7 + 0.2 = 0.9	0.9
						50:50	0.3 + 0.3 = 0.6	0.7
						25:75	0.2 + 0.3 = 0.5	0.6
						0:100	—	0.4

Table 4.

Surface color strength (K/S) of cotton dyed with a mixture of dyes (turmeric with madder and turmeric with red sandalwood) in different proportion by the simultaneous mordanting and dyeing sequence using aluminum sulphate as a mordant.

5.1.3 Application of turmeric in food coloration

Curcumin is a polyphenol found naturally in turmeric rhizome that has antiinflammatory, antioxidant, anticancer, and immunosuppressive activities. It is used mainly in the development of dairy products as the presence of fat (triglycerides) enhances the solubility of curcumin [21]. While few studies have been carried out on colouration of food using turmeric, most of them focus on its functional aspects. Improvement in the sensory attribute and antioxidant potential of ghee has been reported by the addition of 160–350 ppm of curcumin [22]. The turmeric powder improved the oxidative stability and microbiological quality of soft cheese [23]. Turmeric extract rich in curcumin reduced the aging of fresh lamb sausages during modified atmospheric packaging by causing less generation of related volatile compounds due to its antioxidant capacity [24]. The addition of turmeric to the dough of biscuits and breads greatly improved the antioxidant potential and organoleptic properties of breads and biscuits [25].

5.1.3.1 Color produced

Turmeric when applied to food yields a bright orangish-yellow shade.

5.1.3.2 Extraction & application conditions

Curcumin is mainly dissolves in oils and alcohols. It is not stable at alkaline conditions especially at pH above 7.5 though it is quite stable in temperatures generally used for processing foods. Curcumin is complexed with aluminum ions as it is light sensitive.

5.1.3.3 Functional properties related to food application

Curcuminoids present in turmeric possesses anti analgesic, anticarcinogenic, antiinflammatory antioxidant, antiseptic properties. It also helps in the prevention, palliation, or treatment of various disorders such as diabetes, cholelithiasis, diabetes mellitus, foodborne illnesses, and circulatory disorders [26, 27, 28]. Moreover, it also acts as a potent food preservative as it slows down lipid oxidation and possesses antimicrobial activity.

5.1.3.4 Case study-3

The effect of heat treatment and conventional sun drying on the color of fresh turmeric rhizome was evaluated in terms of its hue, yellowness, and brightness (L*, a*, and b* color coordinates) [29]. Turmeric rhizomes were subjected to heat treatment at varying temperatures (50–100°C) for different time periods (10–60 minutes). The rhizomes were cooked at 100°C and then sun-dried for 15 days. The rhizomes were brightened (L*) and yellowed (b*) after being heated at 60-80°C. Heat treatment from 60 to 80°C increased the brightness (L*) and yellowness (b*) of the rhizomes; the values remained the same and did not change with further increase in temperature. The phenolic activity of oxidases in turmeric decreased with an increase in temperature and this led to a decrease in browning of the sample while inversely increasing its hue to a yellower shade and brightness. Though the heat treatment did not significantly decrease the concentration of curcuminoids, sun drying caused a significant reduction in curcuminoids (4–5%). Heat treatment thus enhanced the color of turmeric and maximum brightness was observed at 80°C for 30 minutes.

5.1.3.5 Case study-4

The impact of irradiation on the color stability of curcuminoids was examined and curcumin reagent (curcumin, DMC, and BMC; 79.4, 16.8, and 3.8% - w/w) was irradiated with fluorescent light (27 watt) for 24 hours using a household fluorescent lamp [30]. The color intensity was analyzed by measuring absorbance at 435 nm and curcuminoids before and after treatment were quantified using HPLC. Turmeric pigments (oleoresin and curcumin) were not stable under light, and their photo-degradation was lower when present in higher concentrations. An increase in concentrations of the sample (20–1000 μg/mL) resulted in a loss in color intensity of both oleoresin and curcuminoids in turmeric (Table 5).

Concentration (μg/mL) of the sample	Color intensity
	Turmeric oleoresin	Curcuminoids
20	65.4%	63.0%
200	38.9%	46.2%
1000	28.6%	27.0%

Table 5.

Loss in color intensity of different pigments (i.e. oleoresin and curcuminoids) in turmeric due to light irradiation.

5.2 Annatto

Bixa orellana is a perennial, tall shrub bearing bright white or pink flowers and red-brown fruits in the form of globular ovoid capsules or seed pods with delicate spines. The pods are grouped in clusters and each contains 30–45 cone-shaped seeds covered by a pericarp rich in the red-orange pigment, annatto. B. orellana is native and grows wild in northern South America and Central America. Later in the 16th and 17th centuries, B. orellana was distributed to the Caribbean, Hawaii, and South-Eastern North America, Southeast Asia, and Africa. It is cultivated primarily for its red seeds in India, Sri Lanka, and Java. In India, B. orellana is cultivated for its seed across Orissa, Andhra Pradesh, and Maharashtra. 70% of the world’s coloring agents derived from natural sources come from annatto [31]. Its color is used in food, textile, paint, and cosmetic industries. Also called achiote or bijol it is used as a natural orange-red condiment/spice in the food industry and is used in the bleaching of dairy food products. It is soluble in lipids and is therefore used for imparting red to orange-yellow color to processed food. Annatto is also known as lipstick tree [32] and is used in cosmetics for the production of sunscreens [33], nail gloss, hair oil, and soap. Its medicinal value is associated with its antibacterial, antifungal, antioxidant, antibiotic, and antiinflammatory properties. It has shown anticancer, enhanced gastrointestinal motility, neuropharmacological, anticonvulsant, analgesic, and antidiarrheal activities and has been used as a laxative, cardiotonic, and expectorant, and for wound healing purposes. The dye is also used in the printing and dyeing of textiles like cotton, wool, and silk.

Genus: Bixa | Species: orellana | Family: Bixaceae.

Common name: Achoite | Local name: Latkan or sinduri.

Part of the plant used for coloring: Seeds.

5.2.1 Coloring pigment/component

Of the total carotenoid pigments present in annatto, 80% consists of the red pigment, bixin, and a yellow pigment, norbixin or orelline. Bixin is a yellowish-orange-red dye that is high in carotenoid pigments and is derived from the thin seed coat of B. orellana seeds. Bixin occurs in nature as monomethyl ester of the dicarboxylic carotenoid compound [6,6′-diapo-ψ-ψ′-carotenedioic acid monomethyl ester] i.e. 16-Z (cis) form, but during extraction, it isomerizes to its 16-E (trans) form called isobixin. Norbixin is a naturally occurring demethylated derivative of bixin used for commercial purposes. Besides bixin and norbixin, other compounds such as beta-carotene, cryptoxanthin, lutein, zeaxanthin, orellin, bixein, bixol, crocetin, ishwarane, ellagic acid, salicylic acid, threonine, tomentosic acid, tryptophan, and phenylalanine are also found in the seeds of annatto. Bixin belongs to the direct/acid dye class [34]. Bixin content influences the color value of the annatto extract. With higher amounts of bixin, the L* and b* values decreased (darker and yellower) whereas a* values increased (redder) under the Hunter measurement scale [35]. For Lovibond values, for the same dyes, the R-values increased with the increase in the concentration of bixin while the Y values remained the same. The low purity dye (CFTRI method) showed a higher b*/a* values as compared to the high purity dye (new patented process by CFTRI), whereas a reverse trend was observed with respect to the Y/R values. However, a* and R-values which corresponded to red color increased with an increase in concentration in both color measuring systems irrespective of dye purity [35]. With annatto giving orange shades (combination of yellow and red) b*/a* (degree of yellowness) values were also assessed. With an increase in the bixin concentration, the b*/a* decreased indicating a more yellow color. The study also indicated that the Lovibond color was more influenced by the source of dye and its purity as compared to the Hunter values (Table 6).

Concentration of bixin in mg/L extracted by the patented method	Hunter		Lovibond
	L*	b*/a*	Y	R
10	15.2	2.3	40	6.0
20	14.1	1.9	40	8.0
30	12.9	1.5	40	9.0
40	12.4	1.3	40	10.0
50	12.2	1.2	40	11.0
100	9.7	0.8	40	17.0

Table 6.

Effect of bixin concentration on color values (hunter and Lovibond) [35].

5.2.2 Application of annatto in textile colouration

Natural fibres like cotton [34, 36], silk [37] and wool [32] and also synthetic fibers like nylon and polyester [38] have been dyed with B. orellana. Leather has been dyed in the bright red shade with excellent rub fastness using bixa extract [39].

5.2.2.1 Color produced

Yellow and orange can be produced from B. orellana. Though annatto seed extract gives an orange-red color, the hue depends on the solvent used for extraction [40]. The wavelength of maximum absorption for annatto is 458 nm [41].

5.2.2.2 Extraction

Commercial preparations consist of solutions or suspensions of the pigment in vegetable oil or as a water-soluble form in dilute alkaline solution. Content of total phenols (TP) increases with an increase in pH and higher TP contents were obtained at an extraction time of 60 h and a solvent/seed ratio of 4 ml/g of the extract [42]. The primary pigment cis-bixin is partially transformed to the trans isomer and a degradation product when heated [43]. Microwave-assisted extraction using ethyl acetate solvent also gives good pigment yield [44]. Though the total dye yield (Table 7) on the extraction of annatto seeds by the new patented process by CFTRI, Mysore, 2004 was less than the dual solvent extraction method (CFTRI method), the more purer patented process gave higher bixin and nobixin yields (g/100 g) [35].

Extraction condition	Dye yield (g/100 g)	Bixin (g/100 g)	Norbixin (g/100 g)
Bixin/norbixin dye from Indian seeds by CFTRI method	2.3	21.9	18.5
Low bixin/norbixin dye from Indian seeds by the special patented method	2.0	13.9	12.4
High bixin/norbixin dye from Indian seeds by special patented method CFTRI method	1.0	60.2	55.4

Table 7.

Total yield of dye with bixin and norbixin content in Indian seeds of annatto extracted by different processes [35].

5.2.2.3 Dyeing conditions

B. orellana gives beautiful shades on cotton in alkaline medium using inorganic salts as mordants [45]. Woolen yarns can be dyed with bixa extract in acidic, neutral, and alkaline media using ferrous sulphate, stannous chloride and alum as mordants. Regardless of the presence or absence of mordants, dyeing silk and wool fabric with an aqueous extract of annatto seeds is best successful at pH 4 [41].

5.2.2.4 Fastness

B. orellana reportedly has moderate to poor light fastness, but moderate to excellent fastness to washing, rubbing, and perspiration.

5.2.2.5 Functional properties of annatto related to textiles application

Extract of annatto has remarkable antimicrobial and antioxidant properties and a study revealed that the annatto dye had a bactericidal effect and could reduce E. coli activity [46]. Ethanolic extract form seeds showed broad spectrum antibacterial activity against Bacillus subtilis, Staphylococcus aureus, Streptococcus pyogenes, Salmonella typhi, Pseudomonas aeruginosa, Escherichia coli, and Candida albicans [47]. When compared to the traditional method, annatto dye extracted by the ultrasound aided technique has higher antibacterial and antioxidant activities [48]. Gram-positive bacteria show more sensitivity to annatto dye extracted by conventional and ultrasound-assisted extraction methods than gram-negative bacteria and. B. subtilis showed the lowest sensitivity toward annatto dye, while Escherichia coli gave the highest sensitivity. Annatto dye extracted by UAE showed a bactericidal effect against Salmonella enteritidis [48]. Annatto extracted from annatto seeds (Bixa Orellana) using ultrasound technique was used to color biodegradable films based on poly (3-hydroxybutyrate). Photo-degradation under UVA exposure of colored films showed improvement as suggested by the SEM micrographs, and the film also showed good thermal stability as confirmed by the thermo gravimetric analysis [49].

5.2.2.6 Case study-5

Cotton, wool, and silk were dyed with an aqueous extract of the B. orellana seed powder using the one-step simultaneous sequence and the two-step pre-mordanting sequence by the ultrasound technique [50]. Enzymes were used along with tannic acid to treat the fabric; cellulose and amylase for cotton and protease for silk and wool. The exhaustion of color on the fiber ranged from 55 to 63% for the ultrasound technique, while it was lower ranging between 42 and 46% for the conventional exhaust dyeing procedure for all three fibers. Because bixin and norbixin are polar and acidic, they have a stronger affinity for wool and silk (protein fibers) which have more polar groups than cotton (cellulosic fiber). Enzyme treatment increased the surface color strength (K/S) for all the fibers and lowered the L* values (indicating darker shades). It also slightly increased a* and b* values of all the dyed fibers indicating redder and yellower i.e. probably an orangish tone (Figure 1).

Figure 1.

Effect of enzyme treatment on the color related properties (L*, a* and b*) of cotton, wool, and silk were dyed with an aqueous extract of the Bixa orellana seeds.

5.2.2.7 Case study-6

Eco-friendly bamboo fiber was dyed with B. orellana using potash alum as mordant. The dyeing procedures variables of time, temperature, and pH were optimisation for the pre-mordanting process. An increase in time from 15 min to 60 min increased the color yield and a sharp liner increase in the surface color depth (K/S) was also observed with an increase in temperature from ambient temperature to 80°C. The K/S value of the mordanted bamboo fabric was significantly higher when dyeing was carried out under alkaline conditions. The optimum conditions of dyeing of the potash-alum pre-mordanted bamboo fabric with aqueous extract of seeds of annatto (B. orellana) was thus reported as dyeing time −60 min, dyeing temperature −80°C, and pH–12. Varied shades of orange (sherbet-orange to ginger-orange and apricot colors) were obtained which were dark as indicated by negative L* values. The shades were redder as indicated by positive a* values and yellower as reflected by positive b* values (Table 8). The yellower tint (b*) was more pronounced compared to the redness tint (a*) in most cases [51].

Varying Parameters		K/S at λmax	L*	a*	b*
Control (desized and potash alum pre-mordanted bamboo)		0.1	89.3	−0.2	6.6
Variation in time (in min)	15	4.4	−23.6	30.7	1.2
	30	4.5	−24.3	29.8	1.2
	60	4.7	−24.2	30.0	39.6
Variation in temperature (°C)	Ambient	3.9	−19.9	28.2	42.5
	60	4.7	−23.3	29.8	42.2
	80	5.6	−25.8	30.8	42.5
Variation in pH	2	1.4	−15.3	17.3	25.8
	7	3.8	−23.0	28.0	37.0
	10	6.5	−26.5	33.0	45.8

Table 8.

K/S (color strength) and other color related parameters of bamboo fabric pre-mordanted with potash alum and dyed with aqueous extract of annatto seeds (Bixa orellana) under variable conditions of dyeing.

L* – lightness/darkness; a* – greenness/redness difference; b* – blueness/yellowness; and CDI – color difference index

5.2.3 Application of turmeric in food coloration

Annatto (E-160B) is a natural yellow-orange dye obtained from B. orellana, with less toxicity. Compared to synthetic food colorants, it typically demonstrates superior biodegradability and compatibility with the environment [52]. Annatto colorants are used extensively in the food industry, particularly in the processing of dairy and meat products.

5.2.3.1 Color produced

Annatto gives a yellow to orange-red shade on food.

5.2.3.2 Extraction & Application conditions

Annatto is water-soluble and can be mixed with sugar powder or potassium carbonate. The pigment is not heated stable. Moreover, there is a considerable loss of pigment due to deep-fat frying at high temperatures (> 200°C). It is stable at a pH 5.0–10.

5.2.3.3 Functional properties related to food application

Extract of annatto seed possess antimicrobial properties and decrease the growth and activity of B. subtilis, Staphylococcus aureus, Streptococcus Clostridium thermophilus, perfringens, Lactobacillus casei, Lactococcuslactis, Candida albicans, Candida famata, Rodotorula species, Aspergillus species, and Neurosporacrassa. The seed extract showed strong inhibition of triglycerides oxidation in rapeseed oil [53] and norbixin was able to inhibit oxidative deterioration of olive oil [18]. Annatto seeds’ antioxidant capabilities aid in the treatment of cardiovascular disorders and have been found to decrease the oxidation of low-density lipoprotein in vitro (LDL). Extracts can protect DNA from oxidative damage thereby controlling serious consequences in some age-related cancers.

5.2.3.4 Case study-7

Color from annatto seeds is safe for human consumption compared to the synthetic colorants commonly used in sweetmeats. Jilebi and jangri are popular Indian sweetmeats colored in various shades of yellow and red. Bixin extracted from annatto seeds by a dual solvent method (washing the seeds with a non-polar solvent like hexane and then recovery of the dye in acetone) was used to color jilebi and jangri and its effectiveness was compared to the commercially available counterparts that were colored using synthetic food colorants [54]. The dye was converted to norbixin using potassium hydroxide (alkaline pH) and diluted with potassiun carbonate (K₂CO₃) to get its water-soluble form. Two water-soluble formulations of the colorant (bixin), one having potassium carbonate (with norbixin content, 10.6%) and the other with sugar (with norbixin content, 11.24%) were prepared. The first formulation with potassium carbonate was used to prepare the batter for the sweetmeat, while the other sugar formulation was used for further sweetening the sweetmeat after frying. The bixin extracts were applied in varying concentrations in both the formulations for the preparation of jilebi and jangri, and the effect in terms of red and yellow color units using Lovibond tintomer was evaluated. The effect of frying temperatures (142°C and 172°C) on the color value of both jilebi and jangri were also evaluated. With the increase in the norbixin concentrations, the R values indicated an increasing trend; while the Y values were higher and constant. Commercial jalebis’ R values matched the R and Y values of an annatto solution comprising containing 2.5 and 5 mg/kg of norbixin for sugar-based formulations; while potassium carbonate-based formulations, 5 mg/kg of norbixin gave comparable results (R values) though Y values were higher (Table 9). For jangris, 40 mg/kg of norbixin gave comparable R and Y values for both sugar and potassium carbonate-based formulations. Soluble annatto dye sugar powder formulation gave much better results with lower concentrations of dye as compared to soluble annatto dye potassium carbonate formulation. Water-soluble annatto dye sugar solutions at concentrations 5 mg/kg were found to be optimum for jalebi and 30 mg/kg for jangri as the colored developed was similar that those found in the commercial counterparts. Excellent color matching was observed and no difference in redness and yellowness in the color of the product were reported due to the effect of high temperature used in frying.

Sample	Red values	Yellow values
Commercial jalebi	1.7–4.1	9.0–20.0
2.5 mg/kg of nor-bixin (in sugar based formulation)	2.0	20.0
5 mg/kg of nor-bixin (in sugar based formulation)	3.0	20.0
5 mg/kg of nor-bixin (in potassium carbonate based formulation)	3.1	40.0
Commercial jangri	9.1–10.8	20.0–20.7
40 mg/kg of nor-bixin (in sugar based formulation)	10.0	30.0
40 mg/kg of nor-bixin (in potassium carbonate based formulation)	10.0	40.0

Table 9.

Tinctometer color values of commercial jalebis and jangris, and of the solutions containing optimized concentrations of norbixin with potassium carbonate and sugar.

5.2.3.5 Case study-8

The solubility of bixin in oil and norbixin in water determines its usage. Annatto dye formulations suitable for dairy products like cheese and butter were developed and compared to their commercially available counterparts [55]. Three formulations were prepared; water-soluble solution using K₂CO₃, oil-soluble formulation using vegetable oil, and oil/water-soluble formulation using propylene glycol solution. The formulations were applied at different concentrations in cheese and butter. Lovibond Tintometer was used to measure the color of the commercial and experimental samples. Annatto dye oil/water soluble propylene glycol formulation was found to be the most effective formulation for imparting yellow color with good brightness to various dairy products (Table 10). Butter containing 3.75 mg/kg and 5 mg/kg of oil/water propylene glycol formulation closely resembled the commercial butter samples made using synthetic dyes. In the case of cheese, creamy yellow shade imparted by oil/water propylene glycol formulation at a concentration of 3.75 mg/kg looked very similar to the color of the commercial cheese sample.

Sample	Concentration (mg/kg)	R values	Y values
Commercial butter	—	1.2 ± 0.26	4.0 ± 0.36
Butter with oil soluble annatto extract formulation	3.8	0.8 ± 0.17	2.0 ± 0.26
	5.0	1.0 ± 0.17	3.0 ± 0.26
Butter with water soluble annatto extract formulation	3.8	0.9 ± 0.10	2.5 ± 0.26
	5.0	1.0 ± 0.10	3.3 ± 0.20
Butter with oil/water annatto extract formulation	3.8	1.1 ± 0.17	4.0 ± 0.46
	5.0	1.5 ± 0.26	6.0 ± 0.36
Commercial cheese	—	1.6 ± 0.26	4.6 ± 0.26
Cheese with oil soluble annatto extract formulation	3.8	1.1 ± 0.10	4.0 ± 0.17
	5.0	1.3 ± 0.26	3.3 ± 0.20
Cheese with water soluble annatto extract formulation	3.8	1.2 ± 0.18	2.5 ± 0.26
	5.0	1.2 ± 0.20	3.0 ± 0.30
Cheese with oil/water annatto extract formulation	3.8	1.4 ± 0.12	4.6 ± 0.21
	5.0	1.8 ± 0.10	5.0 ± 0.21

Table 10.

Lovibond tintometer readings of commercial and experimental test samples of butter and cheese.

5.3 Cochineal

Cochineal is a natural dye made from the pulverized and dried corpses of a female sessile parasite found in tropical and subtropical South America and North America. Dyeing of cochineal extract is mainly practiced in Mexico and Peru. Cochineal extracts have been used over ages as colorant for food, textiles, cosmetics, pharmaceuticals, and plastic applications.

The dye has mostly been used in the dyeing of silk, wool, cotton, and natural pigments (lakes) obtained from cochineal insects were used for paintings, frescoes, and restoration processes [56]. It’s the only natural red color that’s been allowed by the FDA for use in food and cosmetics, and it’s frequently used as a substitute for the infamous Red Dye #2.

Genus: Dactylopius | Species: coccus | Family: Dactylopiidae.

Common name: Cochineals | Local name: Cochineal keet.

Part of the plant used for coloring: Cochineal insects are found on the pads of prickly pear cacti in the genus Opuntia or Nopalea and are collected by brushing them off the plants, killed (by immersing in hot water or exposure to sunlight, steam, or dry heat of an oven), dried and powdered to get the dye. One pound of cochineal nectar requires 70,000 insects.

5.3.1 Coloring pigment/component

The important color producing components in cochineal extract are carminic acid, kermesic acid and flavokermesic acid [57, 58, 59]. Cochineal’s coloring ability is due to cochinealin, or carminic acid (80–86%) with anthraquinone as the chromophore and –COOH, –OH, >C=O, and –CH₃ as auxochromes. The bodies of female insects contain up to 25% of their dry weight of this pigment. Glyceryl myristate (a lipid) and coccerin (cochineal) are also found in cochineal. Carmine is formed by precipitating carminic acid onto an alumina hydrate substrate and dried to typically 50 percent concentration. Carmine is insoluble in water but is water-soluble when treated with a strong alkali [60]. Carminic acid showed a moderately strong correlation with chromatic values (a*) from the pigment extract (Table 11). Also, there were no significant differences in the tint value of the samples containing different proportions of carminic acid [61].

Carminic acid content (percent) in different types of turmeric	L	a*	Tint (A420/A500) (ratio between yellow and red pigment)
12.8	19.5	3.9	0.44
15.8	19.4	3.8	0.44
16.0	19.1	3.7	0.45
17.9	19.4	3.6	0.46
19.7	19.4	3.3	0.44

Table 11.

Variation in color values with respect to changing carminic acid n content in turmeric taken from difference different geographical origin.

5.3.2 Application of annatto in textile coloration

Cochineal was considered as one of the great treasures of the New World in the 16th–18th centuries, and along with alkanet, madder, kermes, and lac it formed a source of natural red dye for textiles. Cochineal dyed textile fibers in intense red colors with excellent fastness and was the dyed textiles were highly prized. There are several studies on the use of cochineal for dyeing different fibers; cotton has been dyed with cochineal [62, 63] as also wool [4] and silk [64]. Cochineal extract was used to dye silk and wool by the simultaneous dyeing and mordanting process using 1 gpl and 5 gpl of the dye and 1.5 gpl potash alum and copper sulphate as mordants at pH 4 and 80°C for 90 minutes using liquor ratio 1:40 [65]. Polyamide fabric has been successfully dyed in a range of shades with cochineal using different mordants and mordanting methods [66].

5.3.2.1 Color produced

Cochineal produces scarlet, crimson, orange, and other range of fuchsias, reds, and purples on textiles. Different mordants produce different shades; blue-red/reddish-purple color with alum, maroon-red with copper, purple with iron. The addition of cream of tartar into the dye bath during the dye process will shift the color from a reddish-purple to a vivid flag red color. A combination of mordants also produces different colors like rich red when tin and alum are combined, purple-red when alum and iron are combined, and fuchsia to red shades with a combination of alum and cream of tartar. Over dyeing of cochineal with madder gives a good red, whilst cochineal over-dyed with indigo yields a range of light-fast violets and purples. Cochineal carmin has a maximum absorption wavelength (max) of 520 nm [67]. When carmin is esterified, the hydroxyl groups transform to carbonyl groups, lowering the electron cloud density and resulting in light shading effects [68].

5.3.2.2 Extraction

The bodies of the insect, Dactylopius coccus contain 19–22 percent carminic acid, which can produce crimson and scarlet colors. To preserve the dye without rotting, the insects are dried to roughly 30 percent of their original body weight. The female cochineal insects are processed by immersing them in hot water or exposing them to sunshine, steam, or the dry heat of an oven to extract carminic acid. Each process generates a different color. The dried and powdered insect corpses are cooked in ammonia or sodium carbonate solution, the insoluble debris is removed by filtration, and alum is added to the clear salt solution to precipitate the red aluminum salt to make carmine, a more pure version of cochineal. Colorant extracted from cochineal in acid solubilized medium enhance the color characteristics of bio-mordanted silk fabric [64].

5.3.2.3 Dyeing conditions

pH of dye-bath has a great influence on shades obtained with cochineal though they do not impact the fastness properties of the dyed textiles.

Since the phenolic groups in cochineal are acidic, carminic acid is pale orange in low pH, but it changes to red in slightly acidic and neutral pH, and finally turns violet in alkaline solution [69]. Alkaline medium is favorable for dyeing cotton fabrics with cochineal extract and pre-mordanting cotton with alum and tannic acid mordant mixture improves the color yield [63]. Carminic acid also forms complexes with several metals ions, which act as acceptors to electron donors to form co-ordinate bonds with water-insoluble dye molecules. This complex formation between the dye and the mordant shifts the maximum absorption in the visible range to higher wavelengths with an apparent increase in color intensity. Tin-based mordanting gives a brighter, but higher lightness (L*) value on wool dyed with cochineal than other mordants [70]. The pre-mordanting method is preferred for aluminum and chromium salts, while the post-mordanting method is preferred for copper, tin, and iron salts in order to improve the color yield of wool dyed with cochineal extracts [71]. Catonization of cotton fabric [72] or its treatment with chitosan [70, 73] increases the color value of the cochineal dyed fabric. The optimum dyeing conditions for dyeing cotton with cochineal has been identified as temperature −60°C, time −60 min, MLR–1:40 liquor ratio [74].

5.3.2.4 Fastness

Cochineal generally dyes textiles with excellent light and wash fastness. It gave moderate to good fastness properties on cotton [74] and moderate (grade 3) to very good (grade 4–5) washing fastness, and moderate (grade 5) to excellent (grade 7–8) light fastness on wool yarns [75]. Excellent fastness properties have also been reported on wool dyed with cochineal under the influence of microwave treatment and bio-mordants like heena and pomegranate [4].

5.3.2.5 Functional properties of cochineal related to textiles application

Cochineal imparted antibacterial property to wool, silk, nylon, cotton, and viscose rayon fabrics [71, 76, 77]. Nylon yarn dyed with cochineal dye showed limited antibacterial activity, which increased on mordanting with copper and tin [76]. Excellent UV protection properties (UPF > 100) were observed on wool dyed with cochineal and this was higher for copper sulpate mordant compared to alum and also improved with the increase in dye concentration [65]. UPF values for silk dyed with cochineal was less than 50 at lower concentrations of the dye, but it was very good and in the acceptable range (UPF > 50) with a higher concentration of the dye and in the presence of copper sulphate mordant [65].

5.3.2.6 Case study-9

Woolen yarns were dyed with an aqueous extract of cochineal in presence of five different mordants (aluminum sulphate, stannous chloride, ferrous sulphate, citric acid, and cream of tartar i.e. potassium hydrogen tartarate), singly and in combination, using the pre-mordanting method as well as simultaneous mordanting methods [75]. During dyeing, the carbonyl group (>C=O) and alpha hydroxyl groups (–OH) in the anthraquinone moiety of carminic acid/kermesic acid of cochineal forms a coordinate complex with the metal cation of the mordant. The carboxylic acid group of the cochineal dye can also tautomerize and easily ionize into carboxylate anion (–COO⁻) forming ionic bonding with –NH3⁺ group of the wool fiber. In this way, metal-dye-fiber coordination complexes are formed between the mordant, dye, and the fiber. The anthraquinone-metal combination formed by cochineal and the metal mordant causes a red and blue shift in the visible region, i.e. between 460 and 570 nm, resulting in scarlet-red to purple colors [78]. Due to the H-substitution of the hydroxyl group bonded to C5 of the dye molecule by each metallic ligand, carminic acid present in the cochineal dye induces a bathochromic shift of the main hue to red when it interacts with metal cations during mordanting [79]. This happens when the bonding occurs between the 2-hydroxy group of dye molecule and metal cation [80]. But if bonding between dye and metal ion occurs in 7-hydroxy group, the complex could induce a small blue shift [80]. The bluish-purple color was obtained on unmordanted wool and a range of colors from scarlet-red to black on mordanting with the various mordants. In the case when mordants were used in combination, the final color depended on the chelating property of the dominant mordant, which forms more coordination complexes with the cochineal dye than the other mordants. Thus, ferrous mordant combinations gave grayish chrome; stannous mordant combinations gave reddish chrome and aluminum mordant combinations gave purple chrome. The redness/greenness (a* values) values of dyed samples from both the pre-mordanting method and simultaneous mordanting procedures were positive, indicating that all colors obtained using cochineal dye were in the red-purplish range. All dyed samples irrespective of the mordanting procedures showed an increase in yellowness (b* values) after mordanting and consequently, the color of dyed samples shifted from bluish (higher negative b* values) to yellowish (lower negative or positive b* values). In the pre-mordanting method, the metal cation of the mordant probably diffused well inside the fiber matrix-forming ionic bonding with functional groups of wool fiber before dyeing. During this dyeing process, this metal cation fixed on the fiber probably formed coordinate bonding with the cochineal dye molecule resulting in more aggregation of the dye molecules with the metal cation and formation of dye-fiber-metal complex inside the fiber. Contrarily in the simultaneous dyeing and mordanting method, the coordinate complex between the metal cation and the cochineal dye molecule was probably formed both in the dye-bath as well as inside the fiber matrix leading to lesser aggregation of dye-metal complex inside the wool fiber. Thus darker shades were obtained by the pre-mordanting process and the lightness (L*) of dyed was found to be higher in the case of simultaneous dyeing and mordanting process.

5.3.2.7 Case study-10

Wool was dyed in purple shades with cochineal and metal mordant (aluminum sulphate) and bio-mordant (chitosan) using the pre-mordanting process [81]. Results show that K/S value of wool mordanted with chitosan was higher than when mordanted with aluminum sulphate. Dye uptake increased with an increase in the concentration of the bio-mordant but beyond 1000 mg/L concentration, the K/S decreased. The decrease in dye absorption at higher bio-mordant concentrations may be due to the aggregation of bio-mordant on the wool surface reducing the area for dye adsorption as some dye sites already occupied by the bio-mordant become inaccessible to dye molecules. Thus, by using chitosan as mordant for dyeing wool with cochineal, not only the ill effects of a metal mordant is eliminated, but appreciable depth of color is obtained with lower amounts of dye. Low dye absorption was observed for unmordanted wool at pH 7 which increases at pH 4 indicating acidic pH to be favorable for dyeing wool with this dye. Dye absorption for wool fiber is primarily controlled by ion-exchange reactions between the carboxyl group of dye and amino groups of wool. Below its isoelectric point (pH 4.2), wool, is positively charged, whereas above that point the carboxyl groups present in it render a net negative charge. As a result, at pH 6, the amino groups in wool will always be protonated (carboxylate anions). The pKa value for the carboxyl group of carminic acid in cochineal dye is 2.81, indicating that carminic acid will exist in carboxylate anion form at pH 4. As a result of its increased affinity, the weak carboxylate anion of dye substitutes that of the acid at pH 4. The anion of dye has a complicated character, and when it is bound on wool, it undergoes additional interactions with ionic forces, increasing wool’s dyeability. However, dye absorption in wool pre-treated with chitosan followed an unanticipated pattern and showed higher dye absorption at pH 7. Generally, at pH 4, bio-mordant like chitosan acts as a cationic polyelectrolyte due to protonation of its amine groups thereby significantly increasing the dye absorption capacity of treated wool and at pH 7 it has a very low positive charge. However, the reaction between cochineal and chitosan treated wool was contrary to this indicating that the contact forces them are not solely electrostatic. Hydrogen bonding formation of carminic acid with several hydroxyl and carbonyl groups reduced in the acidic media due to protonation and loss of pair electrons of amine groups of the bio-mordant, resulting in better dye absorption in neutral medium. L* (lightness/darkness) decreased on mordanting indicating darker shades on chitosn pre-mordanted wool dyed with cochineal extract. The a* values were positive indicating redder shades, which decreases on mordanting with chitosan. The b* value of wool dyed with cochineal without any mordant was negative indicating bluer tone. These values were positive and the yellowness of the shades increased (decrease in blueness) when wool was pre-mordanted with chitosan before dyeing with cochineal extract.

5.3.3 Application of cochineal in food coloration

Carmine has a color that is similar to cured pork [82]. Cochineal-derived colors are commonly found in alcoholic beverages, yoghurts, juices, ice creams, and confectionary, but they can also be found in jams and some processed meat items [83]. Typical applications of carmine dye in food are sausages and salami displaying an intense red color [84].

5.3.3.1 Color produced

Cochineal produces intense purple color and the scarlet red color is obtained on complexing with aluminum.

5.3.3.2 Extraction & application conditions

For foodstuffs, extraction conditions for cochineal/carminic acid generally involve acid and/or enzymatic hydrolysis with or without solid-phase extraction (SPE). Carminic acid from cochineal is precipitated onto an alumina hydrate substrate. The precipitated complex called carmine is dried, grounded, and used as a food colorant. Though insoluble in water, carmine can be rendered water-soluble by reaction with a strong alkali. The color of carmine is dependent on the pH; at pH–4 and below, it is orange in color; as pH increases, it becomes redder and bluer until it becomes purplish-red above pH–6.5. The color pigment shows excellent heat and light stability.

5.3.3.3 Functional properties related to food application

Although carminic acid does not produce any genotoxic or cytotoxic effects, it has been related to cause anaphylactic reactions, asthma, urticaria, and angioedema in many individuals.

5.3.3.4 Case study-11

Surimi, minced beef, and milk were colored with naturally occurring carminic acid to change their color. Color modulation of carminic acid and carminic aluminum lake colored surimi, minced meat, and milk through the addition of different food additives, proteins, and metal ions was assessed [85]. Carminic acid rendered a light purple color to surimi while carminic aluminum lake rendered a magenta color. Minced meat and milk turned red and gray-green respectively with carminic acid. Iron and copper changed the color of the samples significantly. Changes were also observed in the case of the presence of food additives. The presence of myofibrillar protein, whey protein isolates, and soy protein isolate changed the pH of the medium resulting in a red color. Sodium nitrite is used as a preservative in the meat industry and as a chromogenic agent as well. Carminic acid changed to yellow with the addition of sodium nitrite though no change was observed in the case of the carminic aluminum lake. Also, no change in color was observed for ascorbic acid. Due to the chelation of the dye in presence of calcium ions, the color of the foodstuff changed. Hence, this dye was not found suitable for food samples rich in calcium and iron.

5.3.3.5 Case study-12

Pulse polarography was used to quantify carmine food dye in strawberry-flavored milk and candies and the results were compared with the UV–visible spectrophotometric analysis [77]. A pH 2.0 Britton-Robinson (B-R) buffer solution was used to perform differential pulse polarography on a falling mercury electrode (peak at 489 mV). Strawberry flavored milk and candy samples were added into the polarographic cell containing B-R buffer (pH 2.0) and polarograms were taken. The concentrations were measured using the standard addition method. To compare the validity of this electroanalytical method, the samples were analyzed using UV–visible spectrophotometry (Figure 2). The relationship between the peak current and carminic acid concentration was linear in the range of 1 μM to 90 μM with a detection limit of 0.16 μM. The results of both methods showed similar accuracy and precision. The pulse polarographic method was advantageous as it showcased high sensitivity, low limit of determination, simple instrumentation, and easy operation (Table 12). The UV-vis curves with the peak of maximum absorbance of turmeric [7], annatto [41] and cochineal [75] along with chemical structures of the main coloring component present in turmeric [9], annatto [50] and cochineal [58] are given in Figure 2.

Figure 2.

UV–vis curves in the visible range with λ_max values of aqueous extracts, and chemical structures of the coloring pigments present in the source of different natural colors.

Sample	Concentration of carminic acid
	Differential Pulse Polarography	UV- visible spectrophotometry
Milk (μg carminic acid /mL milk)	121 ± 4	—
Candy (mg carminic acid/g candy)	28.4 ± 1.5	27.1 ± 2.5

Table 12.

Determination of carminic acid in strawberry-flavored milk and candy using differential pulse polarography and UV–visible spectrophotometry.

6. Conclusion

With the introduction of synthetic dyes like aniline, alizarin, and indigo in the mid-1800, natural dyes lost their economic and commercial significance. Synthetic dyes now dominate the market due to their wide range of colors, ease of production, and excellent fastness features. Existing limitations and technical problems in the procurement of natural dyes have further compelled the shifting of focus from natural dyes to synthetic dyes. However, within a period of 150 years, some serious drawbacks associated with synthetic dyes have come to light; synthetic dyes are suspected to release harmful chemicals that are allergic, carcinogenic, and detrimental to human health. The use of eco-friendly natural dyes that are fairly non-polluting, automatically harmonizing, more challenging, and have rare color ideas in textile and food applications is now becoming increasingly popular due to the strict environmental requirements set on the harmful chemicals used in synthetic dye production. Renewability and eco-friendliness are the two major reasons that have led to the revival of these dyes and their gradual replacement with synthetic colorants.