Dioxetanone

OC-Peroxylactones (1,2-Dioxetanones). Alkyl-substituted 1,2-dioxetanones (21) are prepared using low temperature techniques (54,55). The... 

Dioxetanones decompose near or below room temperature to aldehydes or ketones (56). The decomposition reactions are weakly chemiluminescent Qc ca 10 ein/mol) because the products are poorly fluorescent. However, addition of 10 M mbrene provides 2iQc ca 10 ein/mol, and 2iQc on the order of was calculated at mbrene concentrations above 10 M after correcting for yield loss factors (57). The decomposition rates are first order ia... 

Long before 1,2-dioxetanones were isolated, they were proposed as key intermediates in bioluminescence (58—60). This idea led to the discovery of a number of new chemiluminescent reactions. For example, (23) reacts with to give (25). The hydroperoxide (24) has been isolated and is... 

X = Cl) was based independently on the dioxetanone (61) and concerted peroxide decomposition (6,8,62) theories. Possible examples of dioxetanones in bioluminescence are discussed later. 

Subsequent studies (63,64) suggested that the nature of the chemical activation process was a one-electron oxidation of the fluorescer by (27) followed by decomposition of the dioxetanedione radical anion to a carbon dioxide radical anion. Back electron transfer to the radical cation of the fluorescer produced the excited state which emitted the luminescence characteristic of the fluorescent state of the emitter. The chemical activation mechanism was patterned after the CIEEL mechanism proposed for dioxetanones and dioxetanes discussed earher (65). Additional support for the CIEEL mechanism, was furnished by demonstration (66) that a linear correlation existed between the singlet excitation energy of the fluorescer and the chemiluminescence intensity which had been shown earher with dimethyl dioxetanone (67). 

The carbonyl compound (43) has also been synthesi2ed, and its fluorescence spectmm has been shown to match the bioluminescence spectmm under equivalent conditions (214). The details of the excitation step are unclear and a dioxetanone mechanism (59,215) may apply to the reaction. 

Fig. 1.12 Mechanism of the bioluminescence reaction of firefly luciferin catalyzed by firefly luciferase. Luciferin is probably in the dianion form when bound to luciferase. Luciferase-bound luciferin is converted into an adenylate in the presence of ATP and Mg2+, splitting off pyrophosphate (PP). The adenylate is oxygenated in the presence of oxygen (air) forming a peroxide intermediate A, which forms a dioxetanone intermediate B by splitting off AMP. The decomposition of intermediate B produces the excited state of oxyluciferin monoanion (Cl) or dianion (C2). When the energy levels of the excited states fall to the ground states, Cl and C2 emit red light (Amax 615 nm) and yellow-green light (Amax 560 nm), respectively.

In the luminescence reaction of firefly luciferin (Fig. 1.12), one oxygen atom of the product CO2 is derived from the molecular oxygen while the other originates from the carboxyl group of luciferin. In the chemiluminescence reaction of an analogue of firefly luciferin in DMSO in the presence of a base, the analysis of the product CO2 has supported the dioxetanone pathway (White et al., 1975). 

Contrary to the dioxetanone pathway, DeLuca and Dempsey (1970) proposed a mechanism of the bioluminescence reaction that involves a multiple linear bond cleavage of luciferin peroxide... 

However, the linear bond cleavage hypothesis of the firefly bioluminescence was made invalid in 1977. It was clearly shown by Shimomura et al. (1977) that one O atom of the CO2 produced is derived from molecular oxygen, not from the solvent water, using the same 180-labeling technique as used by DeLuca and Dempsey. The result was verified by Wannlund et al. (1978). Thus it was confirmed that the firefly bioluminescence reaction involves the dioxetanone pathway. Incidentally, there is currently no known bioluminescence system that involves a splitting of CO2 by the linear bond cleavage mechanism. 

Fig. 3.3.4 Reaction mechanism of the coelenterazine bioluminescence showing two possible routes of peroxide decomposition, the dioxetanone pathway (upper route) and linear decomposition pathway (lower route). The Oplopborus bioluminescence takes place via the dioxetanone pathway. The light emitter is considered to be the amide-anion of coelenteramide (see Section 5.4).

One is the concerted decomposition of a dioxetanone structure that is proposed for the chemiluminescence and bioluminescence of both firefly luciferin (Hopkins et al., 1967 McCapra et al., 1968 Shimomura et al., 1977) and Cypridina luciferin (McCapra and Chang, 1967 Shimomura and Johnson, 1971). The other is the linear decomposition mechanism that has been proposed for the bioluminescence reaction of fireflies by DeLuca and Dempsey (1970), but not substantiated. In the case of the Oplopborus bioluminescence, investigation of the reaction pathway by 180-labeling experiments has shown that one O atom of the product CO2 derives from molecular oxygen, indicating that the dioxetanone pathway takes place in this bioluminescence system as well (Shimomura et al., 1978). It appears that the involvement of a dioxetane intermediate is quite widespread in bioluminescence. 

Based on the available knowledge on the chemiluminescence and bioluminescence reactions of various luciferins (firefly, Cypridina, Oplophorus and Renilla), the luminescence reaction of coelenterazine is considered to proceed as shown in Fig. 5.4 (p. 171). The reaction is initiated by the binding of O2 at the 2-position of the coelenterazine molecule, giving a peroxide. The peroxide then forms a four-membered ring dioxetanone, as in the case of the luminescence... 

The decomposition of dioxetanone may involve the chemically initiated electron-exchange luminescence (CIEEL) mechanism (McCapra, 1977 Koo et al., 1978). In the CIEEL mechanism, the singlet excited state amide anion is formed upon charge annihilation of the two radical species that are produced by the decomposition of dioxetanone. According to McCapra (1997), however, the mechanism has various shortfalls if it is applied to bioluminescence reactions. It should also be pointed out that the amide anion of coelenteramide can take various resonance structures involving the N-C-N-C-O linkage, even if it is not specifically mentioned. 

Usami, K., and Isobe, M. (1995). Two luminescent intermediates of coelen-terazine analog, peroxide and dioxetanone, prepared by direct photooxygenation at low temperature. Tetrahedron Lett. 36 8613-8616. 

FMNH2 requirement in bacterial luminescence Crystallization of Cypridina luciferin Crystallization of firefly luciferin Cypridina luciferin in fishes the first cross reaction discovered Structure of firefly luciferin Discovery of aequorin and GFP (green fluorescent protein) Structure of Cypridina luciferin Concept of photoprotein Structure of Latia luciferin Dioxetanone mechanism proposed in firefly and Cypridina luminescence... 

It was pointed out that these values are only approximate but they nevertheless demonstrate the high exergonicity of dioxetane decomposition. The sft2 carbons in the dioxetanones and -diones appear to stabilize the 4-membered ring peroxide. 

Another method of preparing a dioxetanone is ring closure in a a-hydroperoxy-carboxylic acid, as was demonstrated very recently 89> ... 

The reaction is first-order in 88 and in persulfate. It proceeds very probably via a monopersulfate ester 90 and a dioxetanone derivative 91 ... 

The brilliant emissions resulting from the oxidation of certain oxalic acid derivatives, especially in the presence of a variety of fluorophores, are the bases of the most active area of current interest in CL. This group of chemiluminescent reactions has been classified as peroxyoxalate chemistry because it derives from the excited states formed by the decomposition of cyclic peroxides of oxalic acid derivatives called dioxetanes, dioxetanones, and dioxetanediones. 

Several N-methyl-9-acridinecarboxylic acid derivatives (e.g., 10-methyl-9-acridinecarboxylic chloride and esters derived therefrom [39]) are chemiluminescent in alkaline aqueous solutions (but not in aprotic solvents). The emission is similar to that seen in the CL of lucigenin and the ultimate product of the reaction is N-methylacridone, leading to the conclusion that the lowest excited singlet state of N-methylacridone is the emitting species [40], In the case of the N-methyl-9-acridinecarboxylates the critical intermediate is believed to be either a linear peroxide [41, 42] or a dioxetanone [43, 44], Reduced acridines (acridanes) such as N-methyl-9-bis (alkoxy) methylacridan [45] also emit N-methylacridone-like CL when oxidized in alkaline, aqueous solutions. Presumably an early step in the oxidation process aromatizes the acridan ring. 

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