CarotenoidsCarotenoids

By: Jonathan Lord R. Aquino

IntroductionIntroduction

• Nature’s widespread pigments

• biosynthesis by ocean algae

• masked by dominant chlorophyll

• photosynthesis and photoprotection

• ability inactivate reactive oxygen

• precursors of vitamin A

• Provitamin A presents 30 -100 % of the vitamin A requirement

• retinoid structure vitamin A activity

IntroductionIntroduction

Structures of carotenoids IStructures of carotenoids I

• Hydrocarbons carotenes • Oxygenated xanthophylls

Two groups of structure

Structures of carotenoids IIStructures of carotenoids II

• groups on xantophylls:

• hydroxyl, epoxy, aldehyde and ketone

• isoprene unit = basic structure

CH2

C CH

CH3

CH2

Some types ofSome types of carotenoidscarotenoids

Occurrence and distributionOccurrence and distribution

Fruit and vegetables:

• Tomatoes (lycopene)

• Carrots (-, -carotenes)

• Red peppers (capsanthin)

• Pumpkins (-carotene)

• Sweet potatoes (-carotene)

Occurrence and distributionOccurrence and distribution

• All green vegetables contain carotenoids but their color is masked by the green chlorophylls

Physical propertiesPhysical properties

• All carotenoids are lipophilic compounds and thus are soluble in oils and organic solvents

• they can be isomerized by heat, acid, light

• many carotenoids exhibit spectral shifts with various reagents and these spectral changes are used for identification

Chemical propertiesChemical properties

• Carotenoids are easily oxidized because of large number of conjugated double bonds

• Such reactions cause color loss of carotenoids in foods

• Destability of a particular pigment to oxidation is highly dependent on its environment

INTRODUCTIONINTRODUCTION• Carotenoids are familiar to all of us through the orange-

red colors of popular foods like oranges, tomatoes and carrots, and the yellow colors of many flowers.

• They are also added as colorants to many manufactured foods, drinks and animal feeds, either in the form of natural extracts (e.g annatto) or as pure compounds manufactured by chemical synthesis. The production of carotenoids by biotechnology is of increasing interest. Carotenoids are essential to plants for photosynthesis, acting in light-harvesting and, especially, in protection against destructive photooxidation. Without carotenoids photosynthesis in an oxygenic atmosphere would be impossible.

• But carotenoids are not simply pigments of terrestrial plants. They occur widely in bacteria, fungi and algae, where they can be useful taxonomic markers. The production of carotenoids in seaweed runs to hundreds of million tons per year.

• Some animals use carotenoids for coloration, especially birds (yellow and red feathers), fish (g.g. goldfish and salmon) and a wide variety of invertebrate animals, where complexion with protein may modify their color to blue, green or purple.

Carotenoids are important factors in human health. The essential role of beta-carotene and others as the main dietary source of vitamin A has been known for many years. More recently, protective effects of carotenoids against serious disorders such as cancer, heart disease and degenerative eye disease have been recognized, and have stimulated intensive research into the role of carotenoids as antioxidants and as regulators of the immune response system.

• Currents carotenoid research encompasses a wide veriety of fields and interests including plant physiology, food science, environmental science, taxonomy, industrial chemical synthesis, biotechnology and medical research. All the work must be based on a solid foundaation of carotenoid chemistry and reliable methods for handling and analysing these rather unsatble substances.

CAROTENOIDS?CAROTENOIDS?

Of the various classes of pigments in Nature the carotenoids are among the most widespread and important ones, especially due to their most varied functions.In 1831 Wackenroder isolated carotene from carrots and in 1837 Berzelius named the yellow pigments from autumn leaves xanthophylls. This marks the beginning ofcarotenoid research and since then continuous developments have taken place.Because of their ubiquitous occurrence, different functions (see below), and interesting properties carotenoids are the subject of interdisciplinary research in chemistry, bio-chemistry, biology, medicine, physics, and many other branches of science.

OCCURRENCEOCCURRENCE

• As already mentioned, the carotenoids are a class of natural pigments that is very widespread and it was demonstrated that they occur in all the three domains of life, i.e. in the eubacteria, the archea and in the eucarya. A rich source for carotenoids are the algae and more than 100 carotenoids have been isolated and characterized from these organisms. For humans the most important source for carotenoids are plants, where often the brilliant colors of the carotenoids are masked by chlorophyll, e.g. in green leaves.

The carotenoids are responsible for the beautiful colors of many fruits (pine-apple, citrus fruits, tomatoes, paprika, rose hips) and flowers (Eschscholtzia, Narcissus), as well as the colors of many birds (flamingo, cock of rock, ibis, canary), insects (lady bird), and marine animals (crustaceans, salmon).

EXAMPLESEXAMPLES

• Carotenoids found in yellow maize are located in the corn predominantly. Main carotenoids are Zeaxanthin and Cryptoxanthin.

MAIZE

• Zeaxanthin contains a hydroxy function at each end group. Therefore, increased polarity results. Zeaxanthin-glucosides and Zeaxanthin-diglucosides have been found in Archaea (e.g. Sulfolobus shibatae).

• Due to their amphiphilic properties, it was postulated, that these carotenoids act as reinforcers in membrane architecture.

PEPPER PEPPER

Capsanthin is the main carotenoid in red pepper (Capsicum annuum).

Natural carotenoid extracts of pepper are available as food colours. Oleoresin of paprika is the oil extract of paprika, Capsicum annuum. The colour of a food product containing paprika may range from a deep red to a pale pinkish-yellow, depending on the concentrations used. The material can be used in salad dressings, sauces and particularly in meat products, including sausages, where it is allowed as an additive. It is often used in combination with annatto to dye processed cheese.

• This oleoresin contains about 37 to 54 pigments depending on the mode of preparation (extract of unbleached or bleached paprika), of which only few could be completely or even tentatively identified. The main pigments are in general esters of capsanthin and capsorubin. Most assay methods are therefore based on the determination of these two carotenoids.

• According to a tradition, a lot of research is done in Hungary, where pepper is cultivated extensively. The picture below shows the separation of a natural paprika extract by means of open-top column chromatography.

GREEN PEPPERGREEN PEPPER

CARROTCARROT

In natural products the most common carotenoid is the yellow-orange pigment of the carrot (Daucus carota), the ß,ß-carotene. It was isolated in crystalline form as early as in 1831 by Wackenroder. Later, it gave the name to the entire class of these compounds.

• Total synthesis of b,b-Carotene was developped in 1950 by Karrer, Eugster, Inhoffen and Milas. Just four years later b,b-carotene was produced commercially and used as pigment in food and feed. In 1995 capacity for the synthesis of more than 500 tons of b,b-Carotene per year was planned.

• Because of its pro-vitamin A activity b,b-carotene is one of the most important carotenoids. The first step in biosynthesis is the cleavage of the central double bond of b,b-carotene which results in two molecules of retinal. This reaction is catalyzed by the enzyme b-carotene-15,15'-deoxygenase. Afterwards, retinal is reduced to vitamin A (retinol).

the b,b-carotene content can vary depending on the variety, the season and the degree of ripening; • the bioavailability of b,b-carotene from fruits and vegetables depends on the method of preparation before ingestion b,b-Carotene is one of six individual carotenoids produced industrially by Hoffmann-La Roche AG and BASF AG companies. These are namely canthaxanthin, astaxanthin and the apo-carotenoids b-apo-8'-carotenal, b-apo-8'-carotenoic acid ethyl ester and citranaxanthin.

The chart below shows the content of b,b-carotene in variousThe chart below shows the content of b,b-carotene in various fresh fruits and vegetablesfresh fruits and vegetables. The values indicated can only . The values indicated can only serve as an approximation since:serve as an approximation since:

TOMATOTOMATO

The main carotenoids in tomato fruit (Lycopersicon esculentum) are:

Phytoene, phytofluene, z-carotene, neurosporene, lycopene, b-zeacarotene, b,y-carotene and b,b -carotene (The carotenoid content and composition are highly dependent on the tomato variety and on the ripeness of the fruit.)

• All carotenoids may be derived formally from the acyclic C40H56 structure, having a long central chain of conjugated double bonds, by hydrogenation, dehydrogenation, cyclization, or oxidation or any combination of these processes.

Lycopene (y,y-Carotene)

Lycopene (y,y-Carotene) Lycopene (y,y-Carotene)

Tomato paste, tomato sauce, and tomato-based soups are rich in carotenoid compounds and are frequently consumed. Foods such as these, which are high in carotenoid content, were individually identified and quantified by reversed-phase HPLC . The carotenoids detected and quantified included lycopene, lutein, alpha-, beta-, gamma-, and zeta-carotenes, neurosporene, phytoene, and phytofluene. As expected, lycopene was the most abundant carotenoid, ranging in concentration from 0.3 mg/100 g in vegetable beef soup to 55 mg/100 g in tomato paste. The concentration of beta-carotene ranged from 0.23 mg/100 g in tomato soup to 1.51 mg/100 g in vegetable beef soup.

SOFT DRINKS (e.g. FANTA)SOFT DRINKS (e.g. FANTA)

A common composition (%w) of soft drinks is as follows:

•   80 % Water   • 15 % Sugar   • 4 % Fruit Juice     • Carbon dioxide     • Citric Acid     • Vitamin C     • Natural and Synthetical Flavours     Stabilizer    

Colourants

NOMENCLATURE AND STRUCTURENOMENCLATURE AND STRUCTURE

• Carotenoids are a class of hydrocarbons (carotenes) and their oxygenated derivatives (xanthophylls). They consist of eight isoprenoid units joined in such a manner that the arrangement of isoprenoid units is reversed at the center of the molecule so that the two central methyl groups are in a 1,6-position relationship and the remaining nonterminal methyl groups are in a 1,5-position relationship.

Figure 1

All carotenoids may be formally derived from the acyclic C40H56 structure (I) (Fig. 1), having a long central chain of conjugated double bonds, by (I) hydrogenation, (2) dehydrogenation, (3) cyclization, or (4) oxidation, or any combination of these processes. The class also includes compounds that arise from certain rearrangements or degradations of the carbon skeleton (I), provided that the two central methyl groups are retained.

• About 600 carotenoids have been isolated from natural sources; they are listed with their trivial and semisystematic names which also includes literature references for their spectroscopic and other properties. It must be pointed out, however, that for many of the carotenoids listed the structure (this term includes the stereochemistry) is still uncertain and in all these cases a reisolation, followed by structural elucidation with all the modern spectroscopic methods (especially high resolution nuclear magnetic resonance (NMR) spectroscopy) is absolutely necessary. About 370 of the naturally occurring carotenoids are chiral, bearing from one to five asymmetric carbon atoms, and in most cases one carotenoid occurs only in one configuration in Nature.

Figure 2Rules for the nomenclature of carotenoids (semisystematic names) have been published by the International Union of Pure and Applied Chemistry (IUPAC) and IUPAC-International Union of Biochemists (IUB) Commissions on Nomenclature (1975). For the most common carotenoids trivial names are normally used. If these trivial names are used in a paper, the semisystematic name should always be given, in parentheses or in a footnote, at the first mention. All specific names are based on the stem name carotene, which corresponds to the structure and numbering in II (Fig. 2).

The name of a specific compound is constructed by adding two The name of a specific compound is constructed by adding two Greek letters as prefixes (Figure 3) to the stem name carotene; Greek letters as prefixes (Figure 3) to the stem name carotene; the Greek letter prefixes are cited in alphabetical order.the Greek letter prefixes are cited in alphabetical order.

The oxygenated carotenoids (xanthophylls) are The oxygenated carotenoids (xanthophylls) are named according to the usual rules of organic named according to the usual rules of organic chemical nomenclature. The functions most chemical nomenclature. The functions most frequently observed are hydroxy, methoxy, carboxy, frequently observed are hydroxy, methoxy, carboxy, oxo, and epoxy. oxo, and epoxy.

• Important and characteristic carotenoids (Fig. 4) are lycopene (gamma,gamma-carotene) (I),

• beta-carotene (beta,beta-carotene) (III), • alpha-carotene ((6’R)-beta,epsilon-carotene) (IV), • beta-cryptoxanthin ((3R)-beta,beta-caroten-3-ol) (V), • zeaxanthin ((3R,3'R)-beta,beta carotene-3,3'-diol) (VI), • lutein ("xanthophyll", (3R,3'R,6'R)-beta,epsilon -

carotene-3,3'-diol) (VII), • neoxanthin ((3S,5R,6R,3'S,5'R,6'S)-5',6'-epoxy-6,7-

didehydro-5,6,S',6'-tetrahydro-beta,beta-carotene-3,5,3'-triol) (VIII),

• violaxanthin ((3S,5R,6R,3’S,5'R,6'S)-5,6,5',6'-diepoxy-5,6,5',6'-tetrahydro-beta,beta-carotene-3,3'-diol) (IX),

• fucoxanthin ((3S,5R,6S,3'S,5'R,6'R)-5,6-epoxy-3,3',5'-trihydroxy-6',7'-didehydro-5,6,7,8,5',6'-hexahydro-beta,beta-caroten-8-one 3'-acetate) (X),

• canthaxanthin (beta,beta-carotene-4,4'-dione) (XI), • astaxanthin ((3S,3'S)-3,3'-dihydroxy-beta,beta-carotene-

4,4'-dione) (XII).• Derivatives in which the carbon skeleton has been

shortened by the formal removal of fragments from one or both ends of a carotenoid are named apo- and diapocarotenoids, respectively, e.g. beta-apo-8'-carotenal (8'-apo-beta-caroten-8'-al) (XIII). Other structural variations are encountered in the norcarotenoids, in which one or more carbon atoms have been eliminated from within the typical C40-skeleton. A prominent example is the C37-skeleton of peridinin ((3S,5R,6R,3’S,5’R,6’R)-epoxy-3,5,3’-trihydroxy-6,7-didehydro-5,6,5’,6’-tetrahydro-10,11,20-trinor-beta,beta-caroten-19’,11’-olide 3-acetate) (XIV) characteristic of diatoms.

• This cis-trans or (E/Z)-isomerism of the carbon-carbon double bonds is another interesting feature of the stereochemistry of the carotenoids, because it was demonstrated that the (E/Z)-isomers may have different biological properties. The literature in this field is extensive: the first comprehensive review of the cis-trans isomerism of carotenoids and vitamin A was published in 1962 (Zechmeister, 1962). According to the number of double bonds a great number of (E/Z)-isomers exist for each carotenoid, e.g. 1056 for lycopene (I) and 272 for b-carotene (III).

• In view of the (E/Z)-isomerism the double bonds of the polyene chain can be divided into two groups: (I) double bonds with no steric hindrance of the (Z)-isomer (central 15,15'-double bond and the double bonds bearing a methyl group, such as the 9-, 9'-, 13-, and 13'-double bonds) and (2) double bonds with steric hindrance (7-, 7'-, 11-, and 11’-double bonds). Although isomers with sterically hindered (Z)-double bonds are known ((11Z)-retinal) the number of possible (Z)-isomers is in practice reduced considerably, e.g., for lycopene (I) to 72.

• Normally carotenoids occur in Nature as the (all-E)-isomer; however, exceptions are known, such as the (15Z)-phytoene isolated from carrots, tomatoes, and other organisms. On the other hand, some carotenoids undergo isomerization very easily during workup; therefore many (Z)-isomers that are described in the literature as natural products are artifacts.

• For experimental work it must be kept in mind that (E/Z)-isomerization may occur when a carotenoid is kept in solution. Normally the percentage of the (Z)-isomers is rather low, but it is enhanced at higher temperature. Furthermore, the formation of (Z)-isomers is increased by exposure to light.

BIOSYNTHESISBIOSYNTHESIS

Carotenoids are synthesized in Nature by plants and many microorganisms. Animals can metabolize carotenoids in a characteristic manner, but they are not able to synthesize carotenoids.

Carotenoids, being terpenoids, are synthesized from the basic C5-terpenoid precursor, isopentenyl diphosphate (IPP) (XVII) (Fig.1). This compound is converted to geranylgeranyl diphosphate (C20) (XVIII). The dimerization of XVIII leads to phytoene (7,8,11,12,7’,8’,11’,12’-octahydro-gamma,gamma-carotene) (XIX) and the stepwise dehydrogenation via phytofluene (15Z,7,8,11,12,7’,8’-hexahydro-gamma,gamma-carotene (XX), zeta-carotene (7,8,7’,8’-tetrahydro-gamma,gamma-carotene) (XXI), and neurosporene (7,8-dihydro-gamma,gamma-carotene) (XXII) gives lycopene (I). Subsequent cyclizations, dehydrogenations, oxidations, etc., lead to the individual naturally occurring carotenoids, but little is known about the biochemistry of the many interesting final structural modifications that give rise to the hundreds of diverse natural carotenoids.

There are now exciting prospects for rapid progress through the application of molecular genetics techniques in combination with other biochemical and chemical approaches. The benefits of this are not purely academic. The industrial production of natural carotenoids through microbial biotechnology is already established and expanding, mainly through the exploitation of some microalgae (particularly Dunaliella) which can synthesize large amount of carotenoid.

Carotenoids in Human RetinaCarotenoids in Human Retina• Macula: retinal area of highest visual

acuity

• High concentrations of xanthophyll carotenoids lutein and zeaxanthin (yellow coloration)

• Role of carotenoids: optical filtering; antioxidants (protection of macula from light-induced damage)

• Individuals with high dietary intakes and blood levels of lutein and zeaxanthin have a lower rate of visual loss from age-related macular degeneration (AMD), the leading cause of blindness in the elderly.

Macula

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