DURHAM, N.C. -- Duke University chemists have identified a likely chemical pathway among the possible thousands that fruits and vegetables could use to initiate the ripening process.

The information could aid scientists seeking better ways to delay ripening, retain freshness and reduce spoilage in produce, said Duke chemistry professor Michael Pirrung. He added that his group's studies could also yield insight into seed germination and plant response to environmental stress.

Using radioactively labeled compounds, Pirrung's team traced out the pathway that a chemical called ACC follows when it is converted into ethylene -- the "ripening hormone" -- with the aid of a key enzyme.

"We have narrowed the large universe of different conceivable mechanisms ACC can use to go to ethylene down to one which looks pretty good," Pirrung said in an interview. "And we've eliminated a lot of the other possibilities."

The findings of Pirrung and two former students, Jun Cao and Jrlung Chen, were scheduled to be published Thursday on the Internet in Chemistry and Biology, a peer-reviewed electronic journal. Their work was supported by the National Science Foundation and the American Cyanamid Corp.

Scientists have known since the 1970s that ripening begins when fruits and vegetables make and release the simple hydrocarbon ethylene. Researchers have also recognized since the 1980s that this gaseous compound is naturally synthesized out of ACC -- shorthand for aminocyclopropanecarboxylic acid.

That reaction is now known to occur by the action of ethylene forming enzyme (EFE), a difficult-to-study protein that Pirrung-led teams and other research groups independently isolated in 1993.

Enzymes are natural molecular catalysts that spur biochemical reactions in organisms by temporarily binding with precursor molecules like ACC. Chemists call these precursors "substrates."

But the complex ways that enzymes and substrates interact can be very difficult to unravel. Even before EFE was discovered, scientists realized that there is an "infinite number" of possible chemical pathways ACC could follow to generate ethylene, Pirrung said.

Scientists have divided the "reasonable" pathways into two categories, Pirrung added. One group of pathways would each employ hydroxylation, a process that introduces "hydroxyl groups" -- oxygen and hydrogen atoms -- into molecules to change their structures. The other set of options requires the introduction of highly reactive groups of atoms called "free radicals."

Researchers have debated for years whether the "hydroxylation mechanism" or the "free radical mechanism" is the one EFE actually uses, Pirrung said. "The plant physiology community went for hydroxylation, and the chemical community was supporting free radical chemistry."

While both involve "oxidation" -- the removal of some of a molecule's electrons -- these two mechanisms differ fundamentally. Hydroxylation involves the simultaneous transfer of two electrons in a one step process. Free radical chemistry also transfers two electrons, but in a two step process where one electron moves first, and then the next.

Pirrung noted that a vital enzyme in the human liver, Cytochrome P450, uses the hydroxylation mechanism to perform "basically the same kind of chemistry" that EFE does in plant tissue. Cytochrome P450's task is to alter the chemistry of ingested pollutant molecules so that the liver can excrete them from the body.

Despite their chemical similarities, Pirrung said EFE and Cytochrome P450 are also showing themselves to differ in several fundamental ways.

While both enzymes contain iron, Cytochrome P450 also contains heme -- a major blood component -- while EFE doesn't. And while Cytochrome P450 acts through hydroxylation, Pirrung's research has shown that EFE instead generates a free radical that opens up a carbon ring in part of the ACC molecule.

As described in his new Chemistry and Biology report, it's that free-radical-instigated "ring expansion" process that can convert the ACC into ethylene and other byproducts. And the strongly reactive nature of free radicals probably explains why -- unlike with most enzymes -- EFE is slowly destroyed as it acts on ACC molecules, Pirrung said.

Most enzymes, Cytochrome P450 included, are highly recyclable. They repeatedly instigate chemical reactions without being destroyed by them. But EFE is far less efficient, he said. "As this enzyme works, it dies off, and it dies off pretty quickly. After it has made 50 or 100 ethylene molecules, an enzyme molecule doesn't work any more.

"That must be because it is damaging itself through oxidation. It's committing suicide."

Thus, to make enough ethylene, plant tissue must probably produce an oversupply of EFE, Pirrung said. Indeed, his team found EFE in "large amounts" when it isolated the enzyme in plant tissue.

This discovery may lower the likelihood that scientists can learn how to artificially delay the ripening process by simply developing chemicals to block the action of EFE, Pirrung added.

"That's been a kind of 'holy grail' of our program: having a simple chemical agent that could be sprayed on to keep ethylene from being produced," he said. "But we're a bit skeptical of that ourselves now. Because there is so much enzyme there, that strategy may not be effective."

But since ethylene generation only begins the ripening process, there are other potential targets for intervention, Pirrung noted. For example, a commercial genetically engineered tomato, the Flavr Savr, used a modified gene to block the production of a different enzyme that induces softening. "It's further down the pathway of ripening," he said.

Pirrung and current co-investigators are also studying another potential control point: the receptors where newly generated ethylene molecules bind to plant cells.

He has been working on ethylene and its role in ripening since 1979, when he was still a graduate student at the University of California, Berkeley.

"The ripening hormone is a really neat concept, but ethylene does a lot more than just ripen," he added. He noted that ethylene also controls seed germination in the spring. It has a role in the shedding of autumn leaves. And plants also emit it in response to environmental stress.

"Studies strongly suggest that it all goes by the same pathway," Pirrung added. "So whatever we discover in our ripening research ought to apply to any of those other things."

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