Lilian Nassi

ENERGY TRANSFER FROM ENZYMICALLY‐GENERATED TRIPLET SPECIES TO ACCEPTORS IN MICELLES

Abstract— Electronically excited triplet species generated during the peroxidase catalyzed aerobic oxidation of appropriate substrates efficiently elicit fluorescence from acceptors in micelles, as shown with 9, 10‐dibromoanthracene and chlorophyll solubilized by various surfactants. In the case of 9, 10‐dibromoanthracene excited by triplet acetone, phosphorescence also can be detected near O2 depletion.

Chlorophyll: an efficient detector of electronically excited species in biochemical systems

Micelle-solubilized chlorophyll efficiently detects electronically excited species generated in enzymatic systems. In most, if not all, systems the chemiexcited species is formed in the triplet state; chlorophyll fluorescence is observed as result of energy transfer. Red emission can also be elicited from chlorophyll in chloroplasts or bound to microsomes.

RED EMISSION FROM CHLOROPLASTS ELICITED BY ENZYME‐GENERATED TRIPLET ACETONE AND TRIPLET INDOLE‐3‐ALDEHYDE

—Enzyme‐generated triplet acetone and triplet indole‐3‐aldehyde transfer energy very efficiently to chloroplasts, as indicated by the intensity of the sensitized red emission that is observed. The intermediacy of excited species of oxygen (1O2, O2−, HO) has been excluded. Our results open the way for investigating energy transfer in architecturally organized systems in the absence of light.

Triplet energy transfer to chloroplasts from peroxidase-generated excited aliphatic aldehydes

Abstract Linear aldehydes trigger red emission from chloroplasts. If horseradish peroxidase is present, the aldehyde is oxidized to the next lower homologue in the triplet state, which in turn sensitizes chlorophyll fluorescence. Only certain chlorophylls are activated.

EXCITATION OF CHLOROPLASTS INDUCED BY PHENYLACETALDEHYDE

Abstract Chloroplasts promote a slow oxygen consumption by phenylacetaldehyde. The latter elicits a sustained red emission and induces reduction of tetrazolium blue. Addition of horseradish peroxidase greatly increases both O2 uptake and the initial light emission, but has little or no effect upon the reduction of tetrazolium blue. These results indicate that chlorophylls in chloroplasts can be efficiently excited in the absence of light.

ENERGY TRANSFER FROM ENZYME-GENERATED TRIPLET CARBONYLS TO THYLAKOID MEMBRANE FRACTIONS ENRICHED IN PHOTOSYSTEMS I and II

Abstract— Enzyme‐generated triplet species transfer energy very efficiently to thylakoid membrane fractions enriched in either photosystem‐I or photosystem‐II. Independent of the nature of the triplet donor, the emission is always more intense with photosystem‐I. Since the fluorescence quantum yield of chlorophyll in PS‐I is lower and the rate of energy transfer usually smaller than to chlorophyll in PS‐II, the yield of 1S chlorophylls in PS‐I is substantially higher. This is tentatively attributed to more favorable reverse intersystem crossing from an upper triplet state in PS‐I.

Induction of the SOS function sfiA in E. Coli by systems which generate triplet ketones

Generation of triplet ketones, either chemically through thermal decomposition of 3-hydroxymethyl-3,4,4-trimethyl-1,2-dioxetane and 3-[N-(pyridino)carbamoyl]methyl-3,4,4-trimethyl-1,2-dioxetane++ + or enzymatically via the aerobic oxidation of isobutyraldehyde trimethylsilyl enol ether catalyzed by horse-radish peroxidase, triggers the SOS function sfiA in E. coli. Although the observed effects are relatively weak and the triplet ketone scavenger tryptophan was ineffective in this system, our results provide evidence for the involvement of triplet ketones in this type of DNA damage. Possible mechanisms are discussed.

Excitation of micelle-solubilized chlorophyll during the peroxidasecatalyzed aerobic oxidation of isonicotinic acid hydrazide

Addition of micelle (hexadecyl-trimethylammoniumbromide)-solubilized chlorophyll alpha to the isoniazid/peroxidase/Mn2+/O2 system promotes light emission, identified as chlorophyll fluorescence. Based on O2 consumption, the quantum yield of chlorophyll excitation to the S1 state exceeds 6 X 10(-6). At least part of the excitation has its origin in the conversion of an intermediate--presumably a diazene--to pyridine-4-carboxaldehyde. On the basis of the present and earlier results [K. Zinner, C. C. C. Vidigal, N. Durán, and G. Cilento (1977) Arch. Biochem. Biophys. 180, 452-458], it is inferred that isoniazid, an important chemotherapeutic and also a carcinogenic agent, can lead to a substantial generation of electronically excited states.

EXCITATION OF CHLOROPHYLL DURING THE PEROXIDASE‐CATALYZED OXIDATION OF ETHYL α‐FORMYLPHENYLACETATE

Abstract When micelle‐solubilyzed chlorophyll is present during the horseradish peroxidase catalyzed aerobic oxidation of ethyl α‐formylphenylacetate its fluorescence is observed. The excitation of chlorophyll may occur via energy transfer from the enzyme‐generated triplet ethyl benzoylformate. These results imply that excited states may be generated in the roots of Datura innoxia.

Enzymatic generation of triplet acetone: A window to photobiochemistry without light

In most bioluminescent processes, a fluorescent molecule is generated in the excited singlet state as a result of an enzymatic reaction. This species then relaxes to the ground state with emission of light. Work carried out in this and associated laboratories during the last decade has shown that triplet (carbonyl) species can also be generated enzymatically. Since triplet species have much longer intrinsic lifetimes than singlet species, they are potentially of importance in biological systems. The aim of this paper is to provide pertinent, detailed information regarding one of the most striking and widely investigated enzymatic systems which gives rise to an elec- tronically excited product in its triplet state. Students should find this unique system intriguing and interested investigators may find it worthwhile to consider using this system to replace light in order to accomplish certain photochemical and photobiological transformations. The enzymatic generation and transfer of triplet energy is an entirely new field, which opens the way for understanding the biological occurrence of photochemical-like phenomena in the absence of light. 1