Document Q15q0YbGvmmdLm688NYNLpwE

41120 SCIENTIFIC AMERICAN H9. 500,000 JUN 1975 How the Liver Metabolizes Foreign Substances ,Among the most significant of the liver's chemical transformations are the inactivation of drugs the detoxification of environmental pollutants and the activation of chemicals that can cause cancer by At(nllnh Kappa* and Alvito P. Alvorea he intensity and duration of the text the liver through the large portal Taction of moit drugs is determined in large part by their rate of me vein, which ramifies into fine channels through which the blood perfuses slow tabolism, If nothing else happenedlytoamong the liver cells. Here nutrients a drug aftor It entered the body and and other foreign substances are re reached its target organ, for example, It moved, metabolized, in sente cases might continue to act indoihiitoly- Some stored and then released into the gen thing does happen, however: most drug! eral circulation. Amino adds, for exam are transformed into Inactive substance! ple, are made Into proteins and other and then excreted. The biotransforma nitrogenous compounds; glucose is con tion can occur in any of several tissues verted into glycogon and stored, to be and organs. Some drugs are transformed converted back into glucose and released chemically In the intestine, some in the as required. And drugs and other toxic lung, t)>o kidney or the skin. Dy far the substances are detoxified. Nut every greatest number of these chemical re thing is metabolized on the first passago actions are carried out In the liver, which of blood through the liver, of course; metabolism not only drugs but also most drugs, for example, are given in doses of live other foreign chemicals to which such that a sufficient amount of the drug the body Is exposed. Btotratufocmattan moves through the Uver to its site of ac in the Uvor Is therefore a critical factor tion and it transformed later, on return not only in drug tlterepy but also in de visits to the liver [see Ulunration on page fending the body against the toxic effects 2$J, The Uver alio produces bile, which of a wide variety of environmental chem Is a secretion that aids in the digestion icals such as insecticides, herbicides, of fats when It is released into the small dyes, food preservatives end a number intestine and is also a vehicle for the of substances that are suspected of in exaction of transformed substances and ducing cancer. The central step In the other waste products of metabolism. metabolism of most of these agents in The blotranfCormation of drugs and volves an oxidation reaction mediated by other foreign compounds In tho iivor is a complex of enzymes that has coma un accomplished by several remarkable en der intensive study in recent years in our zyme systems that can metabolize a wide laboratory at the Rockefeller University variety of structurally unrelated drugs, Hospital and In other laboratories. toxio agents and environmental pollu tants, which aider the body primarily rP!\o Uver it the Urgett organ (n the through ingestion but also through the " body (It weighs about three pounds In lungs and the skin. The enzyme systems an adult) and has divers* functions. It are built into the membranes of the en serves, first of all, as the primary receiv doplasmic reticulum of the liver cells, a ing depot, clteinicnl-processing plant and network of interconnected channels that distribution center for almost everything is present in the cytoplasm of most ani tlutt enters the body through the walls mal cells. There are two kinds of endo of the alimentary canal. All the blood plasmic reticulum, rough snd smooth, that has absorbed digested food and and they differ in both form and func other cubstanoM from the intestines trv- tion. The surfaces of the rough mem branes are studded with ribosomes, small granules that Isaiulate tlie genetic code into the sequences of amine acids that constitute proteins. The smooth mem branes have no riboaomes, In the Uver a major function of both kinds of mem brane ia to assemble the enzymatic com plexes that transform foreign substances snd then to serve as the sit* of those transformations. Unlike someother cellu lar subsystems, the endoplasmic reticu lum cannot ba separated from cells aa an intact structure. If Uver cclla are ho mogenized and then centrifuged, the tubular reticulum breaks up and bits of the membranes are sealed off to form the tiny vesicles, or sacs, called micro tomes. The microsomal fraction thus ob tained from liver cells is a convenient natural source of enzymes for laboratory studies of Uver-cell mcteltoUsm. I'lmgl and other foreign compound* ^ are metabolized in the liver by a rather small number of reactions: oxida tions, reductions, hydrolyses and con jugations. Their essential effect Is to convert lipophilic, or fat-soluble, com pounds into hydrophilic, or water-solu ble, ones. The hydrophilic compounds are the more readily removed from the blood by the kidneys and excreted. Oxidation accounts for most of the transformations, largely because there are so msny different weyi in which compound can be oxidized [ate (frustra tion on pages 26 and 39V The alkyl itAn chains of barbiturates and some other drugs, for example, are oxidized to form alcohols. In the case of compounds In corporating aromatic rings, Including polycyclic hydrocarbons (such as those in clgaretto smoke) and many drugs, a hydroxyl group 1* Inserted Into the ring. 22 HUNS 036338 accomplished as a result o? the estab lishment of certain new rural practices. Tito first of these is related to house hold economics at the production-team level. Individual household members reech'o a sharo (in money and produce) of tlio annual oamings of their team; the thnro it proportional to tho number of work points each has earned during the yonr. For example, a 70-yoar-old grand mother in a suburban Shanghai com mune told mo that sho and nino of tho other 10 members of the household worked regularly in tho Reids. Tho only nonproduccr was a five-year-old child. Tho family received a monthly advance of tho equivalent of $80 against its |oint annual earnings. In 1973 its share of tho (cam's incomo after operating expenses and taxes were deducted was $2,100, or about $200 per person. Withholding for taxes and expenses and to provide funds for reinvestment absorbs about half of tho gross incomo of a commune. For example, the man* ogor of tho suburban Rainbow Bridge Comimmo in Shanghai told us that 30 porcent of tho commune's gross incomo was required to meet the cost of its agri cultural endeavors. Another 11 peroent went into a fund for capital reinvest ment, 4 porcent was paid to the govern ment as tax and 3 porcent wont into a welfnro fund for the support of new mothers and the elderly. That left 52 percent of the gross income for distribu tion to individuals in proportion to tlie work ]>oints oamod. Such a work-point system simulta neously accomplishes two objectives. It provides a strong incentive for increased ovorall output in order to maximise the amount that is dividod among the indi viduals in a household. At the same time it cncouragos householders and produc tion loams alike to kcop their numbers front increasing, so that the maximum earnings are divided into tho fewest pos sible sharos. Tho latter constraint, of course, places tho disadvantages of pop ulation growth squarely before each in dividual. This ocoitomic consideration must generate substantia! social pressure in favor of small families. Other growth-limiting factors are at work. As one example, it is said that im proved health care hat greatly reduced infant mortality, thus undercutting (he validity of the traditional view that many births are needed to ensure the survival of oven one offspring to maturity. Anolhor example Js the current program of providing care for the aged, one of the uhjoctivos of the 3 percent welfare with holding reported by .the Shanghai com mune. We visited one such residence for LIVESTOCK IN CHINA are compared according to rategorier; eitimatu, made by the UJ. Department of Agriculture, are (or 1949 (gray) and 1972 (color). Sheep and goatt are largely confined to grasslands and mountains In tho west and northweit. That la alao true of range cattle, lumped together here with Urge dreft anlmaltt hornet, mulaa, donkey* end water buffaloes. Pigs are by fer the most numerous of all tho domestic mammals in China. the elderly, maintained by a commune near Peking. Normally, we were told, each family carnd for its own elders. Nonetheless, some individuals inevitably lost their families or became separated from them; that was the case with the 75 residents we saw. They were encouraged to engage in handicrafts; meanwhile they were housed, clothed and fed at commune expense, and a clinic attached to the residence looked after their health needs. Custodial care of this kind, if it were widely practiced, would diminish the validity of another traditional view that favors multiple births: the view that one needs to have many children in or der to ensure one's welfare in old age. A third growth-limiting factor that may even now be undergoing a crucial test is the continued existence of private landholdings throughout rural China. We were told that when the commune system was established in tho late 1950's, between 5 and 7 percent of the arable land in each commune was set aside for private use. The land was par celed out at a fixed rate: each adult in a household was assigned one 150th of a hectare. The household children re ceived private land too, but only the two eldest children in each household were eligible. If a household had more than two children at the time of the land assignment, or if more children arrived later, those children received no share. In such a family there would simply be less private land per person. In its early days the allocation system probably had relatively little effect on rural family planning. Today, nearly a generation later, the system must act as a powerful social force favoring small families. In summary, with respect to agricul tural production in China now and in the near future our group was inclined to ac cept the government's assettiou that self sufficiency was achieved in 1971, when some 250 million metric tons of rice, wheat and other major foodstuffs were harvested. Furthermore, given a contin uation of the present aggressive and co ordinated effort it seems that China wilt be able to achieve substantial increases in agricultural production over the next decade. Whether the increase for any particular crop will bo as little as 20 per cent or as much as 50 percent will prob ably provo to bo a function of present yields. For example, rice yields are al ready high, so that an increase of 50 per cent in the annual harvest will be much harder to achiove thou a 50 percent in crease in the maize or sorghum harvest. /^hiiia nonetheless faces serious long. term problems. If the country's re markable agricultural advanco is to con tinue, two of these problems must soon bo resolved. The first problem lies in tlte current Chinese policy that emphasizes applied and decentralized agricultural research for the sake of Immediate in creases in food production. Although the short-term benefits of this policy are ob viously important, the policy must be complemented in the neni future by sim ilar emphasis on more basic scientific in vestigation. The second problem has to do with the government's remarkably compre hensive efforts to retard the growth of China's population. Only if this policy meets with success will the Chinese peo ple continue to receive the benefits of in creased agricultural production and the accompanying advances in living stan dards that so many of China's people now seem to expect. 2i MCNS 086339 I 'S 0 * ENDOPLASMIC RETICULUM ( a rut llw call U enlarged 111,000 diameter* Ib this electron micrograph wade bp Edward S. Reynold# el (he Harvard Madical Seheel. Moat of iba blotranifermellen* of (erl|n substance* taka place la thia irw f mam* branona tabules. The "roegh" endoplaimie reticulum Is covered wUH tlbatooM, tKa ilmlvm Ib which pietriM ara synthesised i h appear* hare a* fairly linear doable wembranat Madded with black data. The "smooth" endepUamle reticulum lack* rlbeaawe* end farm* a ware branching, tabular oatwackt tbaca are pe&ahaa el to between tbe mitochondria, tbe finajralDad gray ebjeeta, it right. * V * y*\ SMOOTH ENDOPLASMIC RETICULUM proliferates whan tbe synthesis af tbe ensyme tyttemi It contain* I* atlmulated by drug*. Thla micrograph, elao made by Reynold* and reprodnred at the ame trale ai the one at the tap of tbe page, la a( a cell from the liver of a rat that wa* treated with phenebarbltaL The drug atirau* lalca the ayntWiI* of cytochrome P4S0, an ensyme located In the endoplaamic reticulum membrane*. Tbe cytoplasm ef the cell bee expanded because of a striking Increete la tbe smooth endoplee* mlc reticulum membranes, which bow All moM af the *paca tween the mare disported mitochondria. Ensyme Induction by drug can significantly affect the metabolism, and Ibar iba tlvlty, of other drugs that are metabolised by iba i - 23 HONS 086340 lair In other cases alkyl groups'are removed from eithor nitrogen or oxygen atoms, amino groups are removed or sulfoxides arc formod. Reduction and hydrolysis aro also catalyzed by liver enzymes, but theso reactions are less common than oxidntiou, j Conjugation of a chemical is combina tion with some natural constituent of the body such as the glucose derivative glu curonic acid, tho amino acid glycine or iho tripeptido glutathione. In the pres ence of tho appropriate enzyme theso natural agents can combine readily with compounds that have carboxyl (COOH), tulfhydryl (SH), amino (NHg) or hy droxyl (OH) groups. Some drugs have these groups when they are In their ac tive form and are handled in the liver by conjugation; for most drugs, however, conjugation is a second step that comes after metabolism by oxidation, reduction or hydrolysis. Almost without exception the conjugated compound is devoid of pharmacological (or any biological) ac tivity. Clearly the enzyme systems that cata lyse these reactions were net invented by the mammalian organism in order to oope specifically with drugs or novel pollutants. Their basic physiological role has presumably been to metabolize en dogenous substrates: substances nor mally present in tho body. For exam ple. liver microsomal enzymes oxidize steroid hormones, cholosterol and fatty acids. The products of these oxidations may then be conjugated (with glucuron ic acid, for instance) and excreted. Bill- LIVER Is a primary site ef metabolism of oubstances entering the body ihrouih tha alimentary tract. All the fine blood vessels that absorb nutrients and other substances through tho wall of tha Intestines coma together and eater tha liver throofh tha portal vein. The liver's supply of oxygenated blood from the heart enters through the hepatie artery. After passing through the sinusoids of the liver (see fffiutreifon on page ff) the blood la collected by tho hepotle veins, which feed Into the vena cava. The liver secretts bile, which is collected by the bile duct, stored In the gall bladder and emptied Into the duodenum, the first segment of the small Intestine. 24 HONS 086341 robin, product of the oxidation of heme, tho rod pigment of hemoglobin, U an example. It 1 normally prepared for excretion by glucuronlde conjugation. Tho rato of formation of gjucuronides is generally low in nowborn infants, howovor, because of a deficiency of tho en zyme glucuronyl transferase. As a result bilirubin mny not bo conjugated and oxcrotod at an adequate rate. Excessive amounts of it can then accumulate and eauso grave damage to the brain, a con dition called kemictorus. It has boon apparent for some time that tho human fetus and the newborn infant are far more sensitive than adults to many drugs. A number of drugs can pass across the placenta, so that obste tricians nood to exert care in administer ing them to an expectant mother. Bar biturates or morphine given to a woman during childbirth can bo stored in the infant's tissues and cause respiratory de pression and occasionally death. The ex planation for the sensitivity of infants to drugs has emerged from a number of re ports in recent years on the maturation of tho capacity to metabolize and con jugate drugs. Theso studios make it clear tliat the capacity to oxidize and conju gate it negligible in the mammalian fetus and nowborn animal and increases after birth at rate tliat varies with the spe* cios, tho typo of reaction and the drug. Impaired drug metabolism Increases the intensity and duration of drug action. For examplo, newborn mico treated with doso of tlie hypnotic drug hoxoberbital equivalent (o 10 milligrams per kilogram of body wolght sicop more than six hours, whereus adult mice given 10 times as large a dose sleep loss than an hour. rPho adverso effect of drugs on new^ born infants as a result of inefficient metabolic conversion is illustrated by the striking "gray-baby syndrome" In infants that may come a few days after treat ment with the antibiotic chlorampheni col; abdominal distention, respiratory difficulty, cyanosis (blue skin color as a result of Insufficient oxygenation of the blood) and shock. The condition is ap parently the result of deficient metabo lism of the antibiotic and deficient con jugation with glucuronic add. In adults about 90 porccnt of the antibiotic is excretod in tho urino in the form of con* jugeted metabolites within 24 hours after it Ims been orally administered; only a small amount is excreted un changed. In comparison a 10-day-old Infant In one study oxcroted less than 50 porcent of the drug in 24 hours. Ratos of drug metabolism are very dif GENERAL CIRCULATION INTESTINE PATHWAY l drii| ikil U trani(orrod in lh* livar it ihowo ch*aMiclly. The drag (co/ortd arrows) enters the liver through the large portal rein, peteee into the general circulation, hot Its affect on the target organ end eventually returns ta the Hvar. Ob each passage through the liver a fraction of the drug ie converted, usually iuto inactive urtitii lUea (block tnom). The metabolites may be carried by the bile into the Inteetinee for excretion or may pose through the circulation to the kldneye, to be excreted into the urine. ferent in different species, and the effec tive dosage varies accordingly. The anti inflammatory agent phenylbutazone is metabolized slowly in man; its half-life in the plasma averages about three days. In the horse, the dog, the rabbit, the rat and the guinea pig, however, the drug Is metabolized much more quickly and the half-life ranges from three to six hours. (Knowledge of the rapid rate of metabo lism of phenylbutazone in the horse is of practical importance because the drug is administered to treat arthritic conditions in racehorses.) A dose of hexobarbital (adjusted for the body weight of the ani mal) that makes mice sleep for an aver age of 12 minutes puts rabbits to sleep for 49 minutes, rats for 90 minutes and dogs for 315 minutes. When the enzy matic oxidation of hexobarbital by mi crosomes from the liver of these animals was measured, the fastest oxidation was carried out, as expected, by microsomcs from the mouse; the rates of oxidation by inicrosomes from the rabbit, the rat and the dog were proportionately lower. Dif ferences in the rates and patterns of drug metabolism in the various animals seem to explain most of the species differences In effect, but there may also be differ ences in the distribution of the drug, the response of the target tissues and excre tion. Even among human patients there are marked individual variations in the metabolism of drugs that are handled primarily by microsomal enzymes. The variability causes some patients to me tabolize a drug so quickly that tlierapolitically effective blood and tissue lev els are difficult to achieve and others to metabolize the drug so slowly that they suffer toxic effects. It can therefore be difficult for the physician to predict just what dosage of some drugs will provide a safe and therapeutic effect in an in dividual patient. Very largo individual differences have been noted in the me tabolism of the ooumarin-derivative an ticoagulant drug Dicumarol; the halflifo can vary from seven to 74 hours. That makes it hard to predict how much of the drug will provide the desired an ticoagulant effect in a patient. Marked variations have also boon observed in the metabolism in man of phenylbuta zone and of diphenylhydantoin, a drug that is administered to control epileptic seizures. Genetic factors may play an impor- MGNS 086342 25 Iiint role in the mctnlxilism of drugs, hr lllliot S. Vcsdl of tho Pennsylvania Stnto University College of Medicine has shown. The Inrgo indivtdunl differences In Dienmnro) hnlMife persist to some ex tent when frnlcrnnl twins arc compared but almost disappear in identical twins, mul similar results have been repotted for oilier drugs studied in twins. The imtilnlxTCulosis drug isoniaz.ld has been the suhjeet of A number of investigations covering different populations. Isonia/.id is mctulmUml in man primarily by nn Acetylation rcnelion catalyzed by the enzyme N-acetyl transferase. Soon after the drug was introduced it became apparent that individuals vary in their ability to acotylalo it. The distribution of the acetylation rates for n group of subjects tends to bo bimodal, thut is, ]iloMing the rate against the number of patients exhibiting oncli rato yields a curve with two peaks. Apparently there are two classes of individuals: those who ucclylntc Jsoninzid rapidly and excrete tho drug primarily at ncotylisoomzid and (Ikiso who ncctylale and excrete tho drug more slowly and excroto more of it in an unchanged form. Tlie frequency of rapid inactivators is about 90 porccnt in a population of Eskimos or of Japanese, whereas among both whitos and blacks in North America there are about as ninny slow inactivators as there ore rapid Olios. A dour relation has now been estab lished between this variation in acety lation rato and the Incidenco of itoniazid toxicity. Several studies hove shown that tho slow inactivators are more suscep tible to a toxic cfToct of isonlazid: a dis order of the peripheral nerves caused by a specific vitamin Dg deficiency resulting from an interaction of the drug and the vitamin. rphe fast indication that microsomal * enzymes were special kinds of com plexes was developed some 20 years ago by Ccrnld C. Mueller and James A. Mil ler at tbu University of Wisconsin. They discovered that compound# known as amimntzo dyes could be oxidized (spe cifically, N-dcmcthylated) by liver microsomcs ami that the oxidation required molecular oxygen (02) and tho coenzymo NADP1I (reduced nicotinamide-adenine dinuclcotidc phosphate). Soon afterward Itcrnnrd B. Hrodia's group at the Na tional Heart and Lung Institute showed that there was a similar requirement for the oxidative metabolism of a number of dmgs. In 1957 Howard S. Mason of tho University of Oregon Medical School proposed that such oxidations aro cata lyzed by a class of "mixed-function o\idaso*": enzyme complexes that require oxygen and NADPII, are nonspecific (they catalyze tho oxidation of different kinds of compounds) and catalyze the consumption of a molecule of oxygen for each molecule of the drug or other sub strate, with ono atom of oxygen appear ing in the metabolized substrate and tho other atom usually combining with two hydrogen atoms to form water. The key enzyme of these oxidases is cytochrome P-450. A cytochrome is a complex of protein and heme, the ironcontaining ring structure that is the oxy gen-binding component of hemoglobin. Like hemoglobin, tho various cyto chromes serve 0 bind oxygen, which they deliver to their substrates in such processes as cell respiration. Cytochromu T-450 gets its designation from the fact that in the reduced form it binds carbon monoxide and then absorbs light most intensely at a wavelength of 450 nano meters. The ainplitudo of the absorbance peak is the basis of quantitative studies of the enzyme. In tho mixed-function oxidases cytochrome P-450 serves as tho terminal oxidase: it accepts electrons passed along by several intermediates, binds oxygen and then delivers the oxy gen to oxidize its substrate and (usually) produce water (sec illustration on page 30], In addition to NADPII the system includes the enzyme cytochrome P-450 reductase. Another heme protein, cyto chrome b6, is present in liver microsomes and may participate in drug oxidations, but its precise function is not dear. Nor is it known just how the various components of the mixed-function oxi dase complex arc arrayed in tho tulndar membranes of the endoplasmic reticu lum. It appears that the reductase and cytochroma bt are on the outside of the membranes, with cytochrome P-450 in the deeper layers. The enzymo gltieuronyl transferase, which catalyzes tlie most important conjugation reaction, is also present within the membranes. The drug or other fat-soluble compound to bo transformed is presumably bound and IJVF.lt LOBULE, ill* functional unit f liver iImuc, is defined by {tranches of the portal vein, the hepatic artery, lha bile duct and lympltatle vessels, which run together through the lUsue, oulllnini lha lobutas. (The lymphatics ara not shown hore.) Tho venous blood, with its nutrients and other ingested substances, and the oaygonited arterial blood enter the fine sinusoids and perfuse the liver cells, which carry out the metabolic funrllene dUrmaed In ihe text. The blood drains into a central vein in earh lebolo and ihence to sublobular and hepatie veins. The liver cells also secret* bile, which is collected by bile canaliculi that feed into bil* duct ules and eventually Into the main bile duct. This two-dimension al view of part of a lobule it highly simplified and diagrammatic. 26 HONS 086343 metabolized by the mixed-function oxi dase system tlvnt contains cytochrome JM50 and then is often converted into a highly water'Soluble compound by conjugation with glucuronic add; tho transformed product passes into tho lu* men. or central chatmd, of tho tubular membranes and Is excreted from tho coll into tho bile or the bloodstream. Hie cytochrome P-450 mixed-func tion oxidnso system has come to be rec ognised ns having a central role In tho body's defense against chemical agents, whether they aro normal body constitu ents or aro iulroducod from tho environ ment. It is now clear that the system is responsible for the detoxification of many of tho potentially harmful environ mental pollutants. Tho system is highly inducible, that is. its activity can be greatly Increased by exposure to a wide variety of environmental agents and drugs that act as substrates for tho sys tem. Such chcmiculs stimulate the syn thesis of cytochrome P-450 and other comi>oncnts of tho complex. More than 200 storoid hormonos, drugs, insecti cides, carcinogens and other foreign chemicals are now known to stimulate drug motnboliim in experimental ani mals, and many of thorn have been sltown to do tlto samo thing in man. On tlto other bond, somo substances (such as load and other heavy metals) Inhibit tit mixed-function oxidase system, al though ihoy are fower in number and lest dlvorse than tho inducers. The P-450 system is alio tlto site of much competi tive Interaction among drugs and other chemicals that aro undergoing transfor mation. LIVER TISSUE HOMOGENIZED rpho consequence* of this inducibility, * Inhibition and competition have im portant implications in drug therapy. Pationts aro often given several drugs at tho same time. Certain combinations can hovo unpredictable and often undosirablo effects if one drug Inhibits or stim ulates the metabolism of another or competes with it. For example, phenylbutnaon*, tho ooumarin anticoagulants and chloramphonlcol compete with the ntotabolio Inactivation of tolbutamide, a drug givon to roduce the blood-sugar lovol in diabolict; tho competition can lend to excesslvo tolbutamide activity and thus to sorious hypoglycemia, or low blood sugar. PhmvbarbUal it a primo example of a drug that has a different effect: enzyme induction. When rats are treated with phcnobarbltal, there can be a three- to fourfold increaso in the microsomal con tent of cylochromo P-450 and a two- LIVER MICR090ME5, the membrane structures that contain moat ot the liver entymee ensued In detoxification, are isolated by spinning homogenised liver tissue at successively higher speeds,'which produce successively higher gravity (g) forces, In e centrifuge. Homogenised liver tissue U centrifuged for 14 minutes at 600 g; a pellet ot dense materiel, primarily whole cells, cell debris and cell nuclei, collects et the bottom of tho tubo. Tho supernatant, or liquid portion, is centrifuged more strongly, isolating lese denee structures such es mitochondria snd piaeos of membrane. When supernatant 11 is spun at vary high f forces In an uitracentrifuie, the microtomes are eeparated Irons the cytosol, or cell fluid. MONS 086344 (old Inoroaie In the reductase, end drugs ueh as methndono and ethylmorphine ere metabolized three or (our times as (aat. In accordance with thoso experi mental observations chronic adminis tration of phcnobnrbitel to patients decreases tlio effects of many drugs ' by hastening their inactivation. Sedative dotes of the drug reduce the concentra tion in the blood plasma o( phenylbuta zone, the anelgesio antipyrine and tha coumarin anticoagulants; it also de creases their pharmaoological actions. Anticoagulant therapy is particularly sensitive to dosage. A patient who is sat isfactorily maintained on a given coumtrfn-drug dosage while he is being given phenobarbital as a sedative may hemor rhage when the phenobarbital is with drawn because the JM50 systom is no longer being induced and the anticoagu- ADMINISTERED ORUQ8 TRANSFORMED PRODUCTS ADMINISTERED DRUGS REDUCTION II SCNTOOAAeiTAL SIDE-CHAIN OXIDATION I j ^CHCH,-CHCH, <S`^.'cs' X J AROMATIC HYDROXTLATIOM -UNO--lAMTAL I I. ON,--CM, NOEALKYLATtON 9 CM "x. L, " OH,----CM,--C----C---- CM,---- CM---- N. k ^--Ky", MO^H HOCM^CMNHCCMCt CHkOAAMAHtNIOOl O-DEALKYLATION 4* HCHQ DEAMINATION TT SULFOXIDE FORMATION TCM, + V,gO, SAllCVVIC ACID OIUCUAONC AClO + MOOCCM, CM,--CM,--cm,--n; OMkOtWnOMftIMl CM,--CM,--CM,---- SOfAH, IAFAHUMM BIOTRANSPORMATION8 carried eat in the liver Include exideties, redartlea, hydrolysii and eenjnfatlen, example* ef which are given here. The molecular site* ef each reaction are Indicated In color. In few cates traniformaUeo convert! an Inactive farm ef a draf (each ae Preateiil) into an active Ions (ralfenilaalde), bet iamI el the reactions lead to Inactivation, Conjnittlon with a ail- MONS 086345 font is therefore being inactivated more slowly. Aloohol is converted into eoe(alde hyde largely In the Uver, perhaps to somo extent by the mixed-function oxi dase system: heavy drinkers are found to have an increased concentration of TRANSFORMED PRODUCTS T1 MOCKtCMNHCeM&l COOH HOCHjCKjNlCW), or-oc + ural substance sueb as lucuronie acid often fellows Beubolisas ef draft end feelliletee escretlen. Net all the byproducts are ibewa. cytochrome P-450. The habitual con sumption of alcohol therefore stimulates (he metabolism of a wide variety of drugs. This helps to explain why heavy drinkers are less effected than other peo ple by barbiturates and other sedativeswhen thoy are sober, A single very large dose of alcohol taken together with an other drug, on the other hand, inhibits the drug's metabolism, presumably by competing with the drug lor the ap propriate enzymes. This effect, in addi tion to the depressant effect of alcohol on the central nervous system, helps to explain the enhanced sensitivity to bar biturates and other sedatives of a person who bus been drinking heavily. The syn ergistic actions of alcohol and sedatives in the bruin cun cause death. The inducibility of microsomal en zymes by drugs suggests a form of ther apy for certain conditions in which normal body constituents ordinarily me tabolized by such enzymos are present in excessive amounts. Long-term admin istration of phenobarbital, for example, cau lower the concentration of bilirubin, the pigment that produces Jaundice, in the blood of patients with chronic ob struction of bile flow in the liver. The excessive bilirubin levels that are nor mally observed in infants after birth can also be markedly reduced if the mother is given a small dose of phenobarbital for a number of days before delivery, Presumably the drug crosses the placen ta and stimulates the conjugating en zyme system diet is ordinarily tkm to develop in the fetus end the newborn infant. Tfalogenatedhydrocarbon insecticides * such as DDT are potent stimulators of drug and steroid sex-hormone me tabolism in mammals and in birds; the breakdown of sex hormones explains in part tho devastating effects of DDT on reproduction in some bird populations. The minimum exposure to DDT that will stimulate the metabolism of pento barbital and decrease its hypnotic action in experimental animals is one that re sults in concentrations of from 10 to 15 micrograms of DDT per gram of fat; that is a level commonly found in human fat tissues. Among the other insecticides that induce microsomal enzymes in ex perimental animals are chlordane, aldrin and dieldrin. It is interesting that piperonyl butoxide, a synergist that was added to insecticides to Inhibit the enzymatic defenses Insects had developed against DDT and its chemical relatives, also in hibits the activity of the microsomal en zymes in the mammalian liver. ThQl^dorinated biphenyls (PCB's) constitute another class of environmen tal pollutants that huvo been shown to induce microsomal enzymes. The PCB's ore lubricants, heat-exchange fluids. Insulators, plasticizers for paints and plastic compounds and a major com ponent of the lens-immersion oil used in microscopy. Whereas some of the con sumer-product applications have recent ly been curtailed, tho immersion oils are still handled dally by many laboratory workers. PCB's have been found in tlie tissues of numerous bird and Ash species and in human (at and milk, although the route of entry into tho human body has not been accurately determined. In re cent experiments wc have been able to show that the application of pure PCB's or of microscope immersion oil to tho skin of experimental animals in very small amount* (one mtaroliiei) causes a marked increase in mixed-function oxi dase activity and reduces the pharma cological effect of zoxazolamine, a mus cle relaxant, and of hexobarbital In the live animal. These Andings suggest that trivial skin exposure to chomicals can have significant and perhaps harmful biological effects in man. A number of chemicals to which hu man beings are regularly exposed have been identified as chemical carcinogens, that is, they cause cancers when they are applied to the skin of experimental animals or otherwise administered to them. Clearly a factor that inhibits or stimulates the metabolism of such com pounds may affect the development of human cancers. Benzpyrene, benzan thracene and similar polycyclic aromat ic hydrocarbons are among the most ubiquitous carcinogens: they are present in tobacco smoke, in polluted city air and in charcoal-broiled and smoked foods. The polycyclic hydrocarbons are metabolized by a mixed-function oxidase enzymo that in this case is called aryl hydrocarbon hydroxylase because of the particular oxidation It catalyzes. Like other microsomal oxidations, this one requires NADPH and molecular oxygen, but the terminal oxidase of the system induced by the polycyclic hydrocarbons is somewhat different from the oxidase induced by drugs. The catalytic proper ties of the cytochrome are changed, as are its spoctral properties: the absorb ance maximum of the complex of carbon monoxide with the reduced cytochrome is at 448 nanometers rather than at 450, and so the enzyme is designated cyto chrome P-448. Apart from tit* polycyclic hydrocarbons, only one other class of compounds has so far been noted to in duce the formation of cytochrome P-448: we have found that the PCB's induce 29 MONS 036346 ton>o of the newly identified cytochrome long with cytochrome P-450. Thif rug- gotti tike possibility, for which there is developing evidence In animals, that the POP's too may have carcinogenic prop erties. , Tho aryl hydrocarbon hydrbxylase system has boon the subject of intoncive investigation as a possible link in the causation of somo cancers. Tho reason is (lint rnthor than detoxifying its polycy clic-hydrocarbon substrates it seems to mako somo of them more toxic: interme 3 diates of polycyclic-hydrocarbon metab olism such as epoxides are more active in tl>e malignant transformation of tis sue-cultured cells than the parent prod ucts are. Moreover, the enzyme system for carcinogen metabolism is available at many sites In the body that are ex posed to poiycyolio hydrocarbons and is induced not only in the liver but also in the gastrointestinal tract, the kidneys, the skin and the lungs. Cigarette smok ing markedly induces aryl hydrocarbon hydroxylase activity even in the human placenta, as Allan H. Conney, Richard M. Welch and their colleagues showed at the Wellcome Research Laborato ries. Little or no such enzyme activity was found in placentas from nonsmokers. We have investigated the cytochrome P-448 system In human skin and have found that there is marked variability in the hydroxylase activity of different skin samples end that incubation of skin in tissue culture with die polycyclic hydrooarbon benzanthracene induces more enzyme activity. A skin biopsy is easy to do, so that assaying skin aryl hydro*, ytaso In the presence or absence of polycyclic hydrocarbons may provide a convenient test of individual differ ences in the capacity to metabolize cer tain environmental carcinogens. The possibility that induction of aryl hydro carbon hydroxylase is of considerable significance in chemical carcinogenesis is suggested by the results of some re cent experiments performed by Cottfried Kellermann and his associates at the University of Texas, They found that the indudbllity of such hydroxylase activity in lymphocytes is significantly greater in cigarette smokers who have lung cancer than it is in healthy non smokers. OXIDATION OP A DRUG by tho onaymo cytochrome P-450 U vieuaiiaed bare a *ik|bbb> tial prorate. Tha oniyme (/) Is a cample* of a pratsin and tha e*y|en-bindin| compound heme, which contain* an Iran atom that i* initially In tha ferric (Fe* + + ) fora (open circle). The cytochrome bind* the drug (1). Then (3) the enzyme eytoebrome P-450 reduc tase, utilising the ceensyme NADPH, reduce* the iron of the homo to tho ferrous (Fe+ * ) form (block dot), in which it con bind a moleeule of oxygen (4). It supplies one atom of oxygon to oxidise the drug and one generally to form water, in tho process reverting to its oxidised form (5). The drug, oxidised and in most cases inactive, is thereupon released (4). It is clear tiiat the liver's mixcd-func1 lion oxidase system Is implicated in a number of processes affecting human health. The ability of a drug or other foreign substance to stimulate the me tabolism of another drug by the system may explain some of the adverse drug reactions observed in clinical practice. Drug interactions have been well docu mented In anticoagulant therapy; more research is required to explore other drug interactions, since most patients are given several drugs at the same time. The ability of environmental pollutants to modify drug action is now under ac tive investigation. It is clear that insecti cides, for examplo, stimulate drug-metabollzing enzymes and that heavymetal substances such as lead and methyl mercury inhibit tho enzymes; tho clinical significance of these effects may bo appreciable in populations that are exposed occupationally to such agents. Finally, recently acquired evidence seems to indicate that the Induction of aryl hydrocarbon hydroxylase in human tissues by polycyclic hydrocarbons may play a significant role in chemical car cinogenesis. Research on this biological action of carcinogens may provide im portant leads toward predicting the sus ceptibility of individuals to oertain kinds of chemically induced cancer. 30 MONS 086347 1 MIKNOBARDITAL Induce* cytochrome P-451 md tbua itlmulalea lb* iMtiboilim of drug# (nidlml by that ertayme. The curve* (b/0 tbow ihi U|bvcbmlMBN ipwwuii will* peek el 454 * ler*, cheracterblle ef reduced cytoehremo P-450 compleiad with urbee Bwuidi, the mwl umi ef identifying end quantifying the anayma. In e ret treated with pbaoobarbltal tbe cytochrome P* 4$B content of mlerowmoi (cofered curve) < Mbatentlelly higher than that ef microtome* from an entreated ret (block curve). More of the teat dreg etbylmorpblno I* therefore mctebeUaed (com ier) by liver tlaaue from a pbenobarbUal-troatod vet (colored bar) then by the aome amount ef lltaue from an untreated ret (gray). Similarly, phenebarbtlal apeed* up metabolic ef the drug antipy Hne in men (right) i antlpyrine level in pleama fella faetar after pheneberbltal treatment (colored curve) thou ordinarily (Mach). CARCINOGENS induce e different apeciea of cytochrome, cytorhreme P-441. Tbe curver (left) compare the ebaorhance ef micro hum from tbe liver ef a rat treated with the carcinogen methylrholenlhrene (color) with the abaerbance of control microcomea (blech) | the induced eytoehreme baa a peek el 448 nanometer*. Methylrholanlhrene enhance* metabellam ef another carcinogen, benspyrana (canter): the mierotomea of Irootod rat* prodoco more metabolite (color) than control mlcroaomo* (grey) do. Similarly, treating human akin In llaauo culture with tbe carclnegen beneeu' ihracene stimulate* akin-cell metebollam of benapyreno (right). MONS 086348 I