Science
Nitrogenous air pollutants: bigger hazard Findings about nitrous oxide and peroxynitrates indicate that they may be more serious health hazards than had been previously suspected
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ANAHEIM
Ongoing research into the chemistry of nitrogenous air pollutants by environ mental chemists has brought no single spectacular finding. But progress in many laboratories filled a four-day symposium held by the Division of Environmental Chemistry. Together this work begins to develop a more detailed picture of both the chemical reactions these compounds may be undergoing in the atmosphere and how the pollutants affect animals. Among the findings: • The peroxynitrates, a rarely moni tored class of nitrogenous air pollutants, are ^particularly stable at lower air tem peratures and may be more important than ozone in air pollution in northern cities such as Chicago and New York. • Nitrous oxide can react with benzo[a]pyrene or other polycyclic aromatic hydrocarbons in laboratory simulations of polluted air to give products that are mutagenic in the Ames test without acti vation. On the other hand, reactions that form mutagens from previously nonmutagenic air contaminants also seem to take place during air sampling processes, so that measurements of mutagens found in air samples may be artificially high. • Large doses of vitamin Ε seem to protect mice, as well as rats, from the ad verse effects of nitrous oxide. At the levels found in the normal human diet, however, the vitamin has no measurable effect in the animal model. Scientists have assumed that the health hazards associated with photochemical smog are less serious in northern cities than in southern ones because ozone does not form in significant amounts in these colder climates, explains Dr. Dale G. Hendry of SRI International, Menlo Park, Calif. However, he and Dr. Richard A. Kenley find in laboratory experiments simulating polluted urban air that two classes of nitrogenous air pollutants, the peroxyacyl nitrates and the peroxyalkyl 22
C&EN March 27, 1978
Pitts: more investigation needed nitrates, are stable for several hours at 72° F and become increasingly more stable at lower temperatures. The peroxyacyl ni trates (PAN's) have been recognized as important components of smog since the 1950's, Hendry says, and they are known to cause damage to plants at concentra tions as low as 10 ppb. The SRI work in dicates that peroxyalkyl nitrates also may be important, and, since both classes of compounds are stable at lower air tem peratures, they could be serious contam inants of polluted air in northern cities. Hendry and Kenley have not done any actual air monitoring in northern cities, but they say that much of the monitoring presently being done, which uses ozone levels as an indication of air quality, could be misleading in colder areas. At the Chicago meeting of the Ameri can Chemical Society last fall, Dr. James N. Pitts and his colleagues at the Uni versity of California, Riverside, pointed out that air samples taken from the Los Angeles basin contain more mutagens, as determined by the Ames test on bacteria, than can be accounted for by the known carcinogens detected in the air samples. Furthermore, one of the important known carcinogens in the air samples, benzo[a]pyrene, is not directly active in the Ames test. It must be activated by reaction with a liver microsomal extract, whereas the Los Angeles air samples are directly active in the test. At that time Pitts proposed that reactions were taking place in the atmosphere to convert benzo[a]pyrene and other molecules into active muta gens. Now Pitts says that nitrous oxide will react with benzo[a]pyrene in laboratory simulations of polluted air to give several products that are direct mutagens in the
Ames test. Two of the active compounds have been tentatively identified as 6-nitrobenzo[a]pyrene and a mixture of 1nitro and 3-nitrobenzo[a]pyrene. In addition, Pitts finds that nitrous oxide will react with perylene, a wellstudied polycyclic aromatic hydrocarbon that is not mutagenic or carcinogenic and is also present in polluted urban air, to form mutagenic compounds. The muta gen formed in this case is identified as 3-nitroperylene. Relatively high levels of polycyclic aromatic hydrocarbons and oxides of nitrogen exist together in auto mobile exhaust and plumes from coalfired power plants, Pitts points out. Pitts also treated a glass fiber filter with benzo[a]pyrene and placed it in series with an untreated glass filter. The un treated filter removed particulates from the air, but the treated filter still con tained substances that, when extracted, were directly mutagenic in the Ames test. Pitts says this finding suggests that some of the mutagens found in air samples collected by filtering may be "artifacts" formed because of the unusual chemistry that can occur on the surface of the fil ter. New data on the effects of nitrous oxide on animal models come from the labora tory of Dr. Daniel B. Menzel at Duke University's medical center. Menzel and his colleagues find that large doses of vi tamin Ε make mice less susceptible to the lung damage caused by nitrous oxide, ozone, and other free-radical-producing air pollutants. This finding is in accord with his earlier work, which found the same protective effect in rats fed high doses of the vitamin. As in rats, mice with either vitamin Ε-deficient diets or normal diets develop diseased lungs which begin to fill with fluid making them heavier when the ani mals are exposed to 13 ppm of nitrous oxide for long periods. Menzel finds that mice that receive 100 international units (IU) of vitamin Ε per day, or 10 times normal dietary levels, show no increase in lung weight after the same exposure for eight hours per day. Menzel and a number of other bio chemists think that nitrous oxide, ozone, and other free-radical-forming air pollu tants affect animals by interfering with their synthesis of prostaglandin E 2 , a hormone manufactured in the lungs to regulate blood circulation there. Vitamin E, a free-radical scavenger, removes the extra free radicals caused by the polluted air before they can interfere with prosta glandin synthesis, if the vitamin is given in large enough doses, Menzel says. The Duke chemists examined the level
of prostaglandin E2 in lungs following exposure to nitrous oxide in rats that did not receive supplemental vitamin E. Prostaglandin levels in these animals were not affected for three hours following ex posure. Then levels began to fall, reaching a minimum of 70% inhibition of the hor mone 18 hours after exposure. Recovery to half the original level took 200 hours. These long delays in physiological ef fect have not been observed before, Menzel says, and explain why other re searchers have not seen a correlation be tween nitrous oxide exposure and pros taglandin levels. Atmospheric nitrous oxide concentrations fluctuate daily, passing through two maxima in a 24-hour period. Thus, if the findings can be ap plied to man, not only has the body not fully recovered from one high exposure when the next occurs, but it has not even fully reacted to the first exposure, Menzel says. Under these circumstances, air pol lution control measures based on average nitrous oxide levels, rather than peak levels, are probably not adequate. D
Scientists close to synthesis of morphine ANAHEIM V
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No field in chemistry seems livelier than the study of the opiates, the drugs known for their addictive properties and mindlulling qualities. Dr. Arthur G. Schultz of Cornell University in Ithaca, N.Y., told the Organic Chemistry Division of his and graduate student Ying K. Yee's progress toward synthesizing morphine chemically from simple starting materials. Mean while, Dr. Kenner C. Rice of the National Institutes of Health, Bethesda, Md., outlined for the Division of Medicinal Chemistry his group's efforts to make the mirror-image isomer of naloxone, an opiate antagonist. "All of the morphine produced in the U.S. originates from the opium poppy/' Schultz notes. "Our starting material is 1,3-cyclohexanedione, a commercially cheap chemical.,, But beyond the question of cost, synthesis of morphine in a con trolled, industrial setting "might be a way of curbing illicit drug traffic," he says. "However, we're really in the very early stages of developing a synthesis method for morphine," Schultz says. His strategy relies on photoarylation, a method using light to make a carbon-carbon bond to an aromatic ring. The key step is "a photocyclization of an aryl vinylether to give an intermediate carbonyl-ylide that undergoes rearrangement to a dihydrofuran," he says. "The synthesis is at least one step away from making morphine." And it's considerably farther away from being commercially practical. The photoreaction is useful because of
its high chemical yields, close to 90%, and its high quantum efficiency, Schultz ex plains. Beyond that, however, the lightdependent reaction "gives an entirely different way to look at the structure of the molecule and to consider different functional groups—a way that wasn't possible with earlier approaches." Although morphine is yet to be syn thesized, a molecule having many similar structural features to morphine has been made. Called lycoramine, it's a minor al kaloid from the Amaryllis plant family. Overall yield in its synthesis, using pho toarylation, is about 8%. Schultz feels it's too early to be concerned about yields in the morphine scheme. "Worrying over yields isn't appropriate until the last step in the synthesis works," he says. But if it does prove successful, Schultz expects the path to be highly stereoselective. Stereoselectivity is the key word for describing the work that Rice and his colleagues are doing at NIH. They are making the unnatural stereoisomers of opiates, such as morphine and its antag onist, naloxone. Availability of (+)-nal oxone "permits testing for the first time of the stereospecificity of the biochemical and pharmacological actions of its mo lecular mirror image [the natural iso mer]," according to Rice. Natural nalox one is used clinically to reverse narcotic overdoses and is used in research on opi ate receptors in nerve cells. "The regulation of physiological re sponses such as pain, pleasure, and mood may be controlled by such receptors" and the materials that bind to them, Rice notes. With the two stereoisomers of naloxone now available, distinguishing
New synthesis of morphine being readied...
. .."wrong" stereoisomer of naloxone now available
between real and spurious binding effects on these receptors will be possible. So far, the unnatural isomer has been tested in three assay systems generally used for naloxone. The unnatural isomer has less than a thousandth the activity of natural naloxone, according to Rice. Synthesis of the unnatural isomer be gins with extraction of the alkaloid sinomenine from roots of a vine grown in Japan and Korea. The 13-step procedure gives an approximate yield of 10%. Rice predicts unnatural naloxone will be "a valuable tool for unraveling certain com plex functions of the brain." D
Protein mask allows tumor cell growth
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Certain tumor cells wear a glycoprotein "mask" that enables them to evade host defense mechanisms and thus grow faster, Dr. John F. Codington explained to the Division of Carbohydrate Chemistry. He and colleagues at Massachusetts General Hospital in Boston are studying the properties of tumor cell-surface glyco proteins. One, which they call epiglycanin, ap pears on tumor cells that can grow very rapidly in certain mice. Ordinarily, such tumors grow only in mice that carry ap propriate markers (antigens) on the sur faces of normal cells throughout their bodies. But tumor cells carrying epigly canin behave as if disguised, fooling the defense mechanisms of mice that nor mally would destroy the tumor cells. Codington says that epiglycanin masks antigens on the tumor cell surface that usually signal the host defense mecha nisms to come to action. Thus, for exam ple, antibodies produced by the host mouse can't get past the epiglycanin at the tumor cell surface to the proper rec ognition points. When so prevented, the antibodies fail to halt the growth of the tumor cells. Electron micrographs (made in col laboration with Dr. Elizabeth D. Hay and Dr. Scott C. Miller of Harvard medical school) of ordinary tumor cells and of ones containing epiglycanin show dramatically their structural differences. For example, epiglycanin-containing cells have a net work of fine filaments extending more than 100 nm from the cell surface, Codington notes. Sometimes, these fila ments aggregate into thicker, rodlike shapes. By contrast, cells that are sus ceptible to normal host defenses possess no visible extracellular coat. Codington backs up the contention that the "mask" is glycoprotein with specific staining studies. The filament network stains with a polycationic ferritin label, a substance that fixes to sialic-acid-containing glycoproteins such as epiglycanin. March 27, 1978 C&EN
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