Chapter 8 Pigments
8.1 Introduction
To understand colorants in foods some terms need to be defined. Color refers to human perception of colored materials---red, green, blue, etc. A colorant is any chemical, either natural or synthetic, that imparts color. Foods have color because of their ability to reflect or emit different quantities of energy at wavelengths able to stimulate the retina in the eye. The energy range to which the eye is sensitive is referred to as visible light. Visible light, depending on an individual’s sensitivity, encompasses wavelengths of approximately 380-370 nm. This range makes up a very small portion of the electromagnetic spectrum. In addition to obvious colors (hues), black, white, and intermediate grays are also regarded as colors.
Pigments are natural substances in cells and tissues of plants and animals that impart color. Dyes are any substances that lend color to materials. The term dye is commonly used in the textile industrial. In the U .S. food industry, a dye is a fetid-grade water-soluble colorant certified by the U.S. Food and Drug Administration (FDA). These specific dyes are referred to as “certified colors,” and each one is assigned an FD&C number. The FD&C designation means that the dye may be used in foods, drugs, and cosmetics. Added to the approved list of certified colors are the FD&C lakes. Lakes are dyes extended on a substratum and they are oil dispersible. The dye/substratum combination is achieved by adsorption, coprecipitation, or chemical reaction. The complex involves a salt of a water-soluble primary dye and an approved insoluble base stratum. Alumina is the only approved substratum for preparing FD&C lakes. In addition, there are other dyes or lakes approved for use in other countries, where specifications are established by the European Economic Community (EEC) or the World Health Organization (WHO). Colorants exempt from certification may also be used. These are natural pigments or sunbatance synthesized, but identical to the natural pigment.
Color and appearance are major, if not the most important, quality, attributes of foods. It is because of our ability to easily perceive these factors that they are the first to be evaluate by the consumer when purchasing foods. One can provide consumers the most nutrition, safest, and most economical foods, but if they are not attractive, purchase will not occur. The consumer also relates specific colors of foods to quality. Specific colors of fruits are often associated maturity---while redness of raw meat is associated with freshness, a green apple may be judge immature (although some are green when ripe), and brownish-red meat as not fresh.
Color also influences flavor perception. The consumer expects red drinks to be strawberry, raspberry, or cherry flavored, yellow to be lemon, and green to be lime flavored. The impact of color on sweetness perception has also been demonstrated. It should also be noted that some substances such as β-carotene or riboflavin are not only colorants but nutrients as well. It is clear therefore that color of foods has multiple effects on consumers, and it is wrong to regard color as being purely' cosmetic.
Many food pigments are, unfortunately, unstable during processing and storage. Prevention of undesirable changes is usually difficult or impossible. Depending on the pigment, stability is
impacted by factors such as the presence or absence of light, oxygen, heavy metals, and oxidizing or reducing agents, temperature and water activity, and pH. Because of the instability of pigments, colorants are sometimes added to foods.
The purpose of this chapter is to provide an understanding of colorant chemistry---an essential prerequisite for controlling the color and color stability of foods.
8.2 Chlorophyll
8.2.1 Structure of Chlorophylls
Chlorophylls are the major fight harvesting pigments in green plants, algae, and photosynthetic bacteria. Several chlorophylls are found in nature. Their structures differ in the substituents around the pborbin nucleus. Chlorophyll a and b are found in green plants in an approximate ratio of 3:1. They differ in the carbon C-3 substituent. Chlorophyll a contains a methyl group while chlorophyll b contains a formyl groups (Fig. 1). Both chlorophylls have a vinyl and an ethyl group at the C-2 and C 4 position, respectively; a carbomethoxy group at the C- l0 position of the isocylic rings, and a phytol group esterfied to pmpionate at the C-7 position. Phytol is a 20-carbon monounsaturated isoprenoid alcohol. Chlorophyll c is found in association with chlorophyll a in marine algae, dinoflagellates, and diatoms. Chlorophyll d is a minor constituent accompanying chlorophyll a in red algae. Bacteriochlorophylls and chlorobium chlorophylls are principal chlorophylls found in purple photosynthetic bacteria and green sulfur bacteria, respectively.
Figure 1 Structure of chlorophyll.
8.2.2 Physical Characteristics
Chlorophyll are located in the lamellae of intercellular organelles of green plants known as chloroplasts. They are associated with carotenoids, lipids, and lipoproteins. Weak linkages (noncovalent bonds) exist between these molecules. The bonds are easily broken; hence chlorophylls can be extracted by macerating plant tissue in organic solvents. Various solvents have been used. Polar solvents such as acetone, methanol ethanol, ethyl acetate, pyridine, and dimethylformamide are most effective for complete extraction of chlorophylls. Nonpolar solvents such as hexane or petrolleum ether are less effective. High-performance liquid chromatography (HPLC) today is the method of choice for separating individual chlorophylls and their derivatives. In the case of chlorophyll a and b, for example, the increase in polarity contributed by the C-3 formyl substituent of chlorophyll b causes it to be more strongly adsorbed on a normal-phase column and more weakly absorbed on a reverse-phase column than chlorophyll a.
8.2.3 Color Loss during Thermal Processing
Loss of green color in thermally processed vegetables results from formation of pheophytin and pyropheophytin. Blanching and commercial heat sterilization can reduce chlorophyll content by as much as 80-100%. The greater amount of pheophytin detected in frozen spinach as compared to spinach blanched for canning is most likely attributable to the greater severity of the blanch treatment that is generally applied to vegetables intended for freezing. One of the major reason for blanching of spinach prior to canning is to wilt the tissue and facilitate packaging, whereas blanching prior to freezing must be sufficient not only to wilt the tissue but also to inactivate enzymes. The pigment composition shown for the canned sample indicated that total conversion of chlorophylls to pheophytins and pyropheophytins has occurred.
Degradation of chlorophyll within plant tissues postharvest is initiated by heat-reduced decompartmentalization of cellular acids as well as the synthesis of new acids. In vegetables several acids have been identified, including oxalic, malic, citric, acetic, succinic, and pyrrolidone carboxylic acid (PCA). Thermal degradation of glutamine to form PCA is believed to be the major cause of the increase in acidity of vegetables during heating. Other contributors to increased acidity may be fatty acids formed by lipid hydrolysis, hydrogen sulfide liberated from proteins or amino acids, and carbon dioxide from browning reactions.
8.2.4 Technology of Color Preservation
Efforts to preserve green color in canned vegetables have concentrated on retaining chlorophyll, forming or retaining green derivatives of chlorophyll, that is, chlorophyllides, or creating a more acceptable green color through the formation of metallo complexes.
Acid Neutralization to Retain Chlorophyll
The addition of alkalizing agents to canned green vegetables can result in improved retention of chlorophylls during processing. Techniques have involved the addition of calcium oxide and
sodium dihydrogen phosphate in blanch water to maintain product pH or to raise the pH to 7.0. Magnesium carbonate or sodium carbonate in combination with sodium phosphate has been tested for this purpose. However, all of these treatments result in softening of the tissue and an alkaline flavor.
Blair in 1940 recognized the toughening effect of calcium and magnesium when added to vegetables. This observation led to the use of calcium or magnesium hydroxide for the purpose of raising pH and maintaining texture, This combination of treatments became known as the “Blair process”. Commercial application of these processes has not been successful because of the inability of the alkalizing agents to effectively neutralize interior tissue acids over a long period of time, resulting in substantial color loss after less than 2 months of storage.
Another technique involved coating the can interior with ethycellulose and 5% magnesium hydroxide. It was claimed that slow leaching of magnesium oxide from the lining would maintain the pH at or near 8.0 for a longer time and would therefore help stabilize the green color. These efforts were only partially successful, because increasing the pH of canned vegetables can also cause hydrolysis of amides such as glutamine or asparagine with formation of undesirable ammonia-like odors. In addition, fatty acids formed by lipid hydrolysis during high pH blanching may oxidize to form rancid flavors. In peas an elevated pH (8.0 or above) can cause formation of struvite, a glass-like crystals consisting of a magnesium and ammonium phosphate complex. Struvite is believed to result from the reaction of magnesium with ammonium generated from the protein in peas during heating.
High-Temperature Short-Time processing
Commercially sterilized foods processed at a higher than normal temperature for a relatively short time (HTST) often exhibit better retention of vitamins, flavor and color than do conventionally processed foods. The greater retention of these constituents in HTST foods results because their destruction is more temperature dependent than that for inactivation of Clostridium botulinum spores. Temperature dependence can be expressed in terms of z value or activation energy. The z-value is the change in oC required to effect a tenfold change in the destruction rate. The large values for both as compared to that for Clostridium botulinum spores (z =10oC), result in greater color retention when HTST processing is used. However, this advantage of HTST processing is lost after about 2 months of storage, apparently because of a decrease in product pH during storage.
Other studies of vegetable tissue have combined HTST processing with pH adjustment. Samples treated in this manner were initially greener and contained more chlorophyll than control samples (typical processing and pH). However, the improvement in color was generally lost during storage.
Enzymatic Conversion of Chlorophyll to Chlorophyllides to Retain Green Color
Blanching at lower temperatures than conventionally used to inactivate enzymes has been suggested as a means of achieving better retention of color in green vegetables, in tile belief that the chlorophyllides produced have greater thermal stability than their parent compounds. Early studies showed that when spinach was blanched for canning at 71 oC (168oF) for a total of 20 min
better color retention resulted. This occurred as long as the blanch temperature was maintained between 54oC (130oF) and 76oC (168oF). It was concluded that the better color of processed spinach blanched under low temperature conditions (65oC for up to 45 min) was caused by heat-induced conversion of chlorophyll to chlorophyllides by the enzyme chlorophyllase. However, the improvement in color retention achieved by this approach was insufficient to warrant commercialization of the process.
Commercial Application of Metallo Complex
Current efforts to improve the color of green processed vegetables and to prepare chlorophylls that might be used as food colorants have involved the use of either zinc or copper complexes of chlorophyll derivatives. Copper complexes of pheophytin and pheophorbide are available commercially under the names copper chlorophyll and copper chlorophyllin, respectively. The chlorophyll derivatives cannot be used in foods in the United States. Their use in canned foods, soups, candy, and dairy products is permitted in most European countries under regulatory control of the European Economic Community. The Food and Agriculture Organization (FAO) of the United Nations has certified their use as safe in foods, provided no more than 200 ppm of free ionizable copper is present.
Commercial production of the Cu pigments was described by Humphry. Chlorophyll is exacted from dried grass or alfalfa with acetone or chlorinated hydrocarbons. Sufficient water is added, depending on the moisture content of the plant material, to aid penetration of the solvent while avoiding activation of chlorophyllase. Some pheophytin romps spontaneously during extraction. Copper acetate is added to form oil-soluble copper chlorophyll. Alternatively, pheophytin can be acid hydrolyzed before copper ion is added, resulting in formation of water-soluble copper chlorophyllin. The copper complexes have greater stability than comparable Mg complexes; for example, after 25 hr at 25oC, 97% of the chlorophyll degrades while only 44% of the copper chlorophyll degrades.
Regreening of Thermal Processed Vegetables
It has been observed that when vegetables purees ale commercially sterilized, small bright-green areas occasionally appear. It was determined that pigments in the bright green areas contained zinc and copper. This formation of bright-green areas in vegetable purees was termed “regreening.” Regreening of commercially processed vegetables has been observed when zinc and/or copper ions are present in process solutions. Okra when processed in brine solution containing zinc chloride retains its bright green color, and this is attributed to the formation of zinc complexes of chlorophyll derivatives.
8.3 Myoglobin/hemoglobin
Structure of Heme Compounds
Myoglobin is a globular protein consisting of a single polypeptide chain. Its molecular mass is 16.8 kD and it is comprised of 153 amino acids. This protein portion of the molecule is known as globin. The chromophore component responsible for light absorption and color is a porphyrin known as heme. Within the porphyrin ring, a centrally located iron atom is complexed with four
chapter 7 Pigments and colors(
