Neurosporene lycopene from

Fig. 2.16. HPLC profile of carotenoids in an extract of vegetable soup. An expansion of the profile from 30 to 39 is shown in the inset (A). Monitored wavelengths were 436, 440, 464, and 409 nm for peaks 9,10,11,12, and 14, respectively, in the inset (A). Peak identification 1 + 1" = all-trans-lutein and cw-lutein 2 = 5,6-dihydroxy-5,6-dihydrolycopene (lycopene-5,6-diol) 3 = j3-apo-8 -carotenal (internal standard) 4 = lycopene 1,2-epoxide 5 = lycopene 5,6-epoxide 6 = 1,2-dimethoxyproly-copene (tentative identification) 7 = 5,6-dimethoxy-5,6-dihydrolycopene 8 = lycopene 9 = pheo-phytin b 10 = neurosporene 11 = (-carotene 12 = pheophytin a 13 = (-carotene 14 = pheophytin a isomer and (-carotene 15 = a-carotene 16 and 16" = all-trans-/fcarotene, cis-/J-carotene 17 and 17" = all-trans- or cA-phytofluene 18 and 18" = all-trans- or cw-phytoene. Reprinted with permisson from L. H. Tonucci et al. [40].

For example, the ion of [M-69]+, which is observed in the tandem mass spectra of lycopene, neurosporene, and y-carotene but not a-carotene, p-carotene, lutein, or zeaxanthin, indicates the presence of a terminal acyclic isoprene unit. Elimination of a hydroxyl group or a molecule of water, [M-17]+ or [MH-18]+, from carotenoids such as astaxanthin or zeaxanthin is characteristic of the presence of a hydroxyl group. Also, tandem mass spectrometry can be used to distinguish between isomeric carotenoids such as a-carotene and p-carotene, or lutein and zeaxanthin. For example, the ring of a-carotene containing the double bond that i s not conj ugated to the rest of the polyene chain shows unique retro-Diels-Alder fragmentation to form the ion of [M-56]+. In a similar manner, isomeric lutein and zeaxanthin differ by the... 

FIGURE 63.1 Starting with mevalonate, carotenoids are biosynthesized by a special branch of the terpenoid pathway. The first C-40 hydrocarbon unit formed is phytoene, a carotenoid with three conjugated double bonds, which then is enzymatically desaturated to successively yield (3-carotene, neurosporene, and lycopene. Other carotenoids such as (3-carotene and oxocarotenoids are produced from lycopene following cyclization and hydroxylation reactions. Thus, lycopene is a central molecule in the biosynthesis pathway of carotenoids. 

Two new acyclic end-groups have been found. Phytoene-1,2-oxide (15) was isolated from tomatoes. In the bacteria Rhodopseudomonas viridis it was found that although neurosporene and lycopene (16) were present, most of the carotenoids previously thought to be these two compounds, were in fact the corresponding 1,2-dihydro-derivatives [e.g. (17)]. In addition, l,2-dihydro-3,4-dehydrolycopene (18) was present. 

Usually the final product of the phytoene desaturase is lycopene, while in Rhodobacter, the final product is neurosporene. The deduced amino acid sequences of the crtl gene from Rhodobacter (final product, neurosporene), and Erb. longus and Erwinia (lycopene) show significant similarity, but the final products are different (Sandmann, 1994 Armstrong, 1995 Matsumura et al., 1997). The mechanism for recognition of the position on carotenoid by the phytoene desaturase is still unknown that is, how the intermediate of f-carotene or asymmetrical f-carotene and the final product of neurosporene or lycopene are controlled. 

A small amount of lycopene is found in Rpi. globiformis (Table 3 S. Takaichi, unpublished) and in the diphenylamine-inhibited cultures of Chr. okenii (Schmidt et al, 1963). However in diphenylamine-inhibited cultures of Rpi. globiformis, lycopene was not reported (Schmidt and Liaaen-Jensen, 1973). Although small amounts of the intermediates of phytoene desaturase are usually found in photosynthetic bacteria, if the activity of the C-1,2 hydratase is high, lycopene as the substrate may not be found. Therefore, the carotenogenesis pathways of okenone and / .g.-keto carotenoids could possibly start from lycopene rather than from neurosporene (Fig. 7). Nothing is known about how the keto group is introduced under the anaerobic conditions. 

Phytoene (150), phytofluene (151), -carotene (152) and neurosporene (153), the biosynthetic precursors of lycopene (1), also contain the y-end group. They differ from lycopene (1) in the degree of saturation of the 7,8, 7, 8, 11,12 and/or IT,12 double bond(s). Different building blocks as starting material have therefore been used for the synthesis of these compounds. 

Subsequent desaturation reactions lengthen the conjugated double-bond system to produce neurosporene or lycopene. Two completely unrelated types of phytoene desaturases exist. The enzyme found in bacteria (except cyanobacteria) and in fungi catalyzes the entire four-step desaturation process of phytoene to lycopene (Schmidt-Dannert 2000). The plant-type phytoene desaturase from cyanobacteria, algae, and plants carries out a two-step desaturation reaction with different -carotene stereoisomers as reaction products (Britton et al. 1998, Schmidt-Dannert 2000). 

Carotene desaturase catalyzes the final two desaturation steps to lycopene by introducing two double bonds at positions 7,8 of -carotene and 7, 8 of neurosporene. From cyanobacteria, two structurally unrelated genes for -carotene desaturase have been cloned. One crtQ (formerly called zds) from Anabaena is related to the bacterial crtI gene, whereas the second crtQb (also called crtQ-2) from Synechocystis is quite similar to crtP. 

In tomatoes and other higher plants, formation of lycopene (5) from Z-phytoene (11) occurs via -carotene (16). Z-phytoene (11) is converted to Z-phytofluene (14), E-phytofluene (15), -carotene (16), neurosporene (17), and, finally, lycopene (5) by extracts of tomato fruit plastids (Fig. 26.5) (Porter et al., 1984 Spurgeon and Porter, 1983). 

Studies with cell-free systems have now confirmed the proposed desaturation of phytoene to the more unsaturated carotenes. Cell-free systems obtained from tomato plastids and spinach chloroplasts will convert [ C]phy-toene (the 15-cis isomer) into lycopene and cyclic carotenes (Kushwaha et al., 1970 Subbarayan et al., 1970), as will an extract from P. blakesleeanus (Davies, 1973). Extracts ofH. cutirubrum (Kushwaha et al., 1976) incorporate radioactivity from tran.s-phytoene into the more unsaturated carotenes. With Flavobacterium extracts [ CJphytoene (the 15-cis isomer) was also converted to the more unsaturated carotenes (Brown et al., 1975). Later studies with tomato plastid extracts have demonstrated the conversion of c/i-[ C]phytofluene, rra/ij-[ C]phytofluene, and /ra/i5- -[ C]carotene to neurosporene and lycopene (Qureshiet al., 1974a,b). Cell-free extracts from H. cutirubrum (Kushwaha et al., 1976) also convert tran -phytofluene to fra/is- -carotene, -carotene to neurosporene, and neurosporene to lycopene. The conversion of neurosporene to lycopene has also been demonstrated using extracts from the Cl 15 mutant of P. blakesleeanus (Davies, 1973). 

The direct conversion of lycopene to cyclic carotenes has also been demonstrated with a number of cell-free systems. Extracts from tomato plastids and spinach chloroplasts have been shown to incorporate radioactivity from lycopene into 8-, y-, a-, and jS-carotene (Kushwaha r al., 1%9 1970 Papa-stephanou, 1973). Incorporation of radioactivity from lycopene into -caro-tene and other carotenes has also been reported with bean leaf chloroplasts (Decker and Uehleke, 1%1 Hill et al., 1971) and a cell-free system from P. blakesleeanus (Davies, 1973). The occurrence of a-zeacarotene (7, 8 -dihy-dro- ,il -carotene) and /3-zeacarotene (7, 8 -dihydro-/8,il -carotene) in some organisms suggests that neurosporene can also be cyclized. Whether or not this pathway is of significance in the biosynthesis of y-, 8-, a-, and /3-caro-tenes is not clear, however (Britton, 1976b). Possibly, the enzyme that cy-clizes lycopene also cyclizes neurosporene, but at a much slower rate. Label from neurosporene has also been incorporated into /3-carotene by cell-free... 

Phytoene an aliphatic, colorless hydrocarbon carotenoid, M, 544. P. is a polyisoprenoid containing six branch methyl groups, two terminal isopropyli-dene groups and nine double bonds, three of them conjugated. Only the A double bond has cis configuration. Biosynthetically, P. is derived from two molecules of geranylgeranyl pyrophosphate, and it serves as a C40-starter molecule in the biosynthesis of other carotenoids phytofluene, carotene, neurosporene and lycopene are formed by the stepwise dehydro-... 

Phytofluene. Porter and Lincoln s work with various tomato selections indicated the presence of phytofluene, along with other precursors (7). In an enzyme system obtained by ammonium sulfate precipitation of proteins from spinach (Spinacia oleracea) leaves, phytoene, phytofluene, and lycopene were produced from labeled isopentenyl pyrophosphate (11). Curiously, no f-carotene or neurosporene was reported. 

The simplest prototype of carotenes, called I/-carotenes, is the polyunsaturated acyclic hydrocarbon (15Z)-7,8,11,12,7, 8, llM2 -octahydro- r, Ir-carotene, known as phytoene, which is synthesised from two molecules of geranylgeranyl diphosphate. Isomerisation of phytoene yields the trans-isomer phytofluene (7,8,1 l,12,7, 8 -hexahydro- Ir, Ir-carotene). Oxidation of phytofluene gradually gives -carotene (7,8,7, 8 -tetrahydro-vIf,vIf-carotene), neurosporene (7,8-dihydro- f, Ir-carotene) and lycopene(vIf,vIr-carotene), which is the final product of the biosynthesis (9-180) and the main pigment of tomatoes (30-200 mg/kg), watermelons (33 121 mg/kg) and rose hips (101-834 mg/kg... 

Dehydrogenations (Fig. 88). The key substance phytoen is subjected to a series of dehydrogenations which lead to phytofluen, -carotene, neurosporene and, finally, lycopene. As already mentioned -carotene is the first colored carotene in this sequence. Apart from both ends of the molecule lycopene consists of a sequence of completely conjugated double bonds. Its name comes from its occurring in certain breeds of tomatoes (Lycopersicon esculeiuum). 

The desaturation of phytoene occurs in a stepwise sequence to form phytofluene (7,8,1 l,12,7, 8 -hexahydro-i/r,i/r-carotene), f-carotene (7,8,7, 8 -tetrahydro-(/, (/ -carotene), neurosporene (7,8-dihydro-i/r,i/r-carotene), and lycopene (Figure 4.3). At each stage, two hydrogen atoms are lost by trans elimination from adjacent positions to extend the polyene chromophore by two double bonds." Desaturation can be carried further to 3,4,3 4 -tetrahy-drolycopene, as demonstrated in spinach chloroplasts." ... 

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