Authors: Rachel Carson
The research that led to our present understanding of cellular oxidation is one of the most impressive accomplishments in all biology and biochemistry. The roster of contributors to this work includes many Nobel Prize winners. Step by step it has been going on for a quarter of a century, drawing on even earlier work for some of its foundation stones. Even yet it is not complete in all details. And only within the past decade have all the varied pieces of research come to form a whole so that biological oxidation could become part of the common knowledge of biologists. Even more important is the fact that medical men who received their basic training before 1950 have had little opportunity to realize the critical importance of the process and the hazards of disrupting it.
The ultimate work of energy production is accomplished not in any specialized organ but in every cell of the body. A living cell, like a flame, burns fuel to produce the energy on which life depends. The analogy is more poetic than precise, for the cell accomplishes its "burning" with only the moderate heat of the body's normal temperature. Yet all these billions of gently burning little fires spark the energy of life. Should they cease to burn, "no heart could beat, no plant could grow upward defying gravity, no amoeba could swim, no sensation could speed along a nerve, no thought could flash in the human brain," said the chemist Eugene Rabinowitch.
The transformation of matter into energy in the cell is an ever-flowing process, one of nature's cycles of renewal, like a wheel endlessly turning. Grain by grain, molecule by molecule, carbohydrate fuel in the form of glucose is fed into this wheel; in its cyclic passage the fuel molecule undergoes fragmentation and a series of minute chemical changes. The changes are made in orderly fashion, step by step, each step directed and controlled by an enzyme of so specialized a function that it does this one thing and nothing else. At each step energy is produced, waste products (carbon dioxide and water) are given off, and the altered molecule of fuel is passed on to the next stage. When the turning wheel comes full cycle the fuel molecule has been stripped down to a form in which it is ready to combine with a new molecule coming in and to start the cycle anew.
This process by which the cell functions as a chemical factory is one of the wonders of the living world. The fact that all the functioning parts are of infinitesimal size adds to the miracle. With few exceptions cells themselves are minute, seen only with the aid of a microscope. Yet the greater part of the work of oxidation is performed in a theater far smaller, in tiny granules within the cell called mitochondria. Although known for more than 60 years, these were formerly dismissed as cellular elements of unknown and probably unimportant function. Only in the 1950's did their study become an exciting and fruitful field of research; suddenly they began to engage so much attention that 1000 papers on this subject alone appeared within a five-year period.
Again one stands in awe at the marvelous ingenuity and patience by which the mystery of the mitochondria has been solved. Imagine a particle so small that you can barely see it even though a microscope has enlarged it for you 300 times. Then imagine the skill required to isolate this particle, to take it apart and analyze its components and determine their highly complex functioning. Yet this has been done with the aid of the electron microscope and the techniques of the biochemist.
It is now known that the mitochondria are tiny packets of enzymes, a varied assortment including all the enzymes necessary for the oxidative cycle, arranged in precise and orderly array on walls and partitions. The mitochondria are the "powerhouses" in which most of the energy-producing reactions occur. After the first, preliminary steps of oxidation have been performed in the cytoplasm the fuel molecule is taken into the mitochondria. It is here that oxidation is completed; it is here that enormous amounts of energy are released.
The endlessly turning wheels of oxidation within the mitochondria would turn to little purpose if it were not for this all-important result. The energy produced at each stage of the oxidative cycle is in a form familiarly spoken of by the biochemists as ATP (adenosine triphosphate), a molecule containing three phosphate groups. The role of ATP in furnishing energy comes from the fact that it can transfer one of its phosphate groups to other substances, along with the energy of its bonds of electrons shuttling back and forth at high speed. Thus, in a muscle cell, energy to contract is gained when a terminal phosphate group is transferred to the contracting muscle. So another cycle takes place—a cycle within a cycle: a molecule of ATP gives up one of its phosphate groups and retains only two, becoming a diphosphate molecule, ADP. But as the wheel turns further another phosphate group is coupled on and the potent ATP is restored. The analogy of the storage battery has been used: ATP represents the charged, ADP the discharged battery.
ATP is the universal currency of energy—found in all organisms from microbes to man. It furnishes mechanical energy to muscle cells; electrical energy to nerve cells. The sperm cell, the fertilized egg ready for the enormous burst of activity that will transform it into a frog or a bird or a human infant, the cell that must create a hormone, all are supplied with ATP. Some of the energy of ATP is used in the mitochondrion but most of it is immediately dispatched into the cell to provide power for other activities. The location of the mitochondria within certain cells is eloquent of their function, since they are placed so that energy can be delivered precisely where it is needed. In muscle cells they cluster around contracting fibers; in nerve cells they are found at the junction with another cell, supplying energy for the transfer of impulses; in sperm cells they are concentrated at the point where the propellant tail is joined to the head.
The charging of the battery, in which ADP and a free phosphate group are combined to restore ATP, is coupled to the oxidative process; the close linking is known as coupled phosphorylation. If the combination becomes uncoupled, the means is lost for providing usable energy. Respiration continues but no energy is produced. The cell has become like a racing engine, generating heat but yielding no power. Then the muscle cannot contract, nor can the impulse race along the nerve pathways. Then the sperm cannot move to its destination; the fertilized egg cannot carry to completion its complex divisions and elaborations. The consequences of uncoupling could indeed be disastrous for any organism from embryo to adult: in time it could lead to the death of the tissue or even of the organism.
How can uncoupling be brought about? Radiation is an uncoupler, and the death of cells exposed to radiation is thought by some to be brought about in this way. Unfortunately, a good many chemicals also have the power to separate oxidation from energy production, and the insecticides and weed killers are well represented on the list. The phenols, as we have seen, have a strong effect on metabolism, causing a potentially fatal rise in temperature; this is brought about by the "racing engine" effect of uncoupling. The dinitrophenols and pentachlorophenols are examples of this group that have widespread use as herbicides. Another uncoupler among the herbicides is 2,4-D. Of the chlorinated hydrocarbons, DDT is a proven uncoupler and further study will probably reveal others among this group.
But uncoupling is not the only way to extinguish the little fires in some or all of the body's billions of cells. We have seen that each step in oxidation is directed and expedited by a specific enzyme. When any of these enzymes—even a single one of them—is destroyed or weakened, the cycle of oxidation within the cell comes to a halt. It makes no difference which enzyme is affected. Oxidation progresses in a cycle like a turning wheel. If we thrust a crowbar between the spokes of a wheel it makes no difference where we do it, the wheel stops turning. In the same way, if we destroy an enzyme that functions at any point in the cycle, oxidation ceases. There is then no further energy production, so the end effect is very similar to uncoupling.
The crowbar to wreck the wheels of oxidation can be supplied by any of a number of chemicals commonly used as pesticides. DDT, methoxychlor, malathion, phenothiazine, and various dinitro compounds are among the numerous pesticides that have been found to inhibit one or more of the enzymes concerned in the cycle of oxidation. They thus appear as agents potentially capable of blocking the whole process of energy production and depriving the cells of utilizable oxygen. This is an injury with most disastrous consequences, only a few of which can be mentioned here.
Merely by systematically withholding oxygen, experimenters have caused normal cells to turn into cancer cells, as we shall see in the following chapter. Some hint of other drastic consequences of depriving a cell of oxygen can be seen in animal experiments on developing embryos. With insufficient oxygen the orderly processes by which the tissues unfold and the organs develop are disrupted; malformations and other abnormalities then occur. Presumably the human embryo deprived of oxygen may also develop congenital deformities.
There are signs that an increase in such disasters is being noticed, even though few look far enough to find all of the causes. In one of the more unpleasant portents of the times, the Office of Vital Statistics in 1961 initiated a national tabulation of malformations at birth, with the explanatory comment that the resulting statistics would provide needed facts on the incidence of congenital malformations and the circumstances under which they occur. Such studies will no doubt be directed largely toward measuring the effects of radiation, but it must not be overlooked that many chemicals are the partners of radiation, producing precisely the same effects. Some of the defects and malformations in tomorrow's children, grimly anticipated by the Office of Vital Statistics, will almost certainly be caused by these chemicals that permeate our outer and inner worlds.
It may well be that some of the findings about diminished reproduction are also linked with interference with biological oxidation, and consequent depletion of the all-important storage batteries of ATP. The egg, even before fertilization, needs to be generously supplied with ATP, ready and waiting for the enormous effort, the vast expenditure of energy that will be required once the sperm has entered and fertilization has occurred. Whether the sperm cell will reach and penetrate the egg depends upon its own supply of ATP, generated in the mitochondria thickly clustered in the neck of the cell. Once fertilization is accomplished and cell division has begun, the supply of energy in the form of ATP will largely determine whether the development of the embryo will proceed to completion. Embryologists studying some of their most convenient subjects, the eggs of frogs and of sea urchins, have found that if the ATP content is reduced below a certain critical level the egg simply stops dividing and soon dies.
It is not an impossible step from the embryology laboratory to the apple tree where a robin's nest holds its complement of blue-green eggs; but the eggs lie cold, the fires of life that flickered for a few days now extinguished. Or to the top of a tall Florida pine where a vast pile of twigs and sticks in ordered disorder holds three large white eggs, cold and lifeless. Why did the robins and the eaglets not hatch? Did the eggs of the birds, like those of the laboratory frogs, stop developing simply because they lacked enough of the common currency of energy—the ATP molecules—to complete their development? And was the lack of ATP brought about because in the body of the parent birds and in the eggs there were stored enough insecticides to stop the little turning wheels of oxidation on which the supply of energy depends?
It is no longer necessary to guess about the storage of insecticides in the eggs of birds, which obviously lend themselves to this kind of observation more readily than the mammalian ovum. Large residues of DDT and other hydrocarbons have been found whenever looked for in the eggs of birds subjected to these chemicals, either experimentally or in the wild. And the concentrations have been heavy. Pheasant eggs in a California experiment contained up to 349 parts per million of DDT. In Michigan, eggs taken from the oviducts of robins dead of DDT poisoning showed concentrations up to 200 parts per million. Other eggs were taken from nests left unattended as parent robins were stricken with poison; these too contained DDT. Chickens poisoned by aldrin used on a neighboring farm have passed on the chemical to their eggs; hens experimentally fed DDT laid eggs containing as much as 65 parts per million.
Knowing that DDT and other (perhaps all) chlorinated hydrocarbons stop the energy-producing cycle by inactivating a specific enzyme or uncoupling the energy-producing mechanism, it is hard to see how any egg so loaded with residues could complete the complex process of development: the infinite number of cell divisions, the elaboration of tissues and organs, the synthesis of vital substances that in the end produce a living creature. All this requires vast amounts of energy—the little packets of ATP which the turning of the metabolic wheel alone can produce.
There is no reason to suppose these disastrous events are confined to birds. ATP is the universal currency of energy, and the metabolic cycles that produce it turn to the same purpose in birds and bacteria, in men and mice. The fact of insecticide storage in the germ cells of any species should therefore disturb us, suggesting comparable effects in human beings.
And there are indications that these chemicals lodge in tissues concerned with the manufacture of germ cells as well as in the cells themselves. Accumulations of insecticides have been discovered in the sex organs of a variety of birds and mammals—in pheasants, mice, and guinea pigs under controlled conditions, in robins in an area sprayed for elm disease, and in deer roaming western forests sprayed for spruce budworm. In one of the robins the concentration of DDT in the testes was heavier than in any other part of the body. Pheasants also accumulated extraordinary amounts in the testes, up to 1500 parts per million.
Probably as an effect of such storage in the sex organs, atrophy of the testes has been observed in experimental mammals. Young rats exposed to methoxychlor had extraordinarily small testes. When young roosters were fed DDT, the testes made only 18 per cent of their normal growth; combs and wattles, dependent for their development upon the testicular hormone, were only a third the normal size.