Toxicants Occurring Naturally in Foods. (1966)

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180 A. G. VAN VEEN manufacture bongkrek, poisonings are much more common than usual. Outbreaks of bongkrek poisonings were reported at the end of the last century. It was found that a newly discovered bacterium, Pseudomonas cocovenenans, produces two poisonous substances. One, called bongkrek acid, is especially interesting. In the bongkrek itself, dissolved in residual fat, this poison is very stable and is seldom, if ever, decomposed during heating. It was found that even small quantities cause hypoglycemia in human consumers and in experimental animals (e.g., monkeys, rabbits, and rats). It has been found in animals that the glycogen of liver and muscles is mobilized so that a hyperglycemia precedes the hypoglycemia after which the victim dies, either within a few hours or after 1 or 2 days. Glucose injections only prolong life by a number of hours. In spite of these outbreaks of poisoning, people go on consuming this palatable and cheap food because they think that the poisonings are due to evil spirits or something comparable. The same bacteria do not produce bongkrek acid in soybeans and peanut presscake. Bongkrek acid has the properties of an unsaturated fatty acid with about 29 carbon atoms.!7 The pharmacological activity of the preparations depends on the optical rotation. In the purified state the product is extremely unstable. It is interesting to note that the substance is also an effective anti- biotic for the Rhizopus fungus!® on which it acts in much the same way as on human beings and experimental animals, i.e., it upsets glycogen metabolism. It has been shown by Berends and co-workers that bong- krek acid disturbs oxidative phosphorylation; oxidation of pyruvate and malate was inhibited.!” It has been found that if the coconut presscake before inoculation with the fungus is treated with some acid, for example, with leaves from a certain variety of Oxalis (which is available everywhere in central Java), the pH gets low enough (approximately 5.5) to inhibit the growth of Pseudomonas cocovenenans, although the fungus grows extremely fast at this pH. However, it is not easy to convince the population that by the simple method of adding oxalis leaves to bongkrek the health hazard would be reduced. Ackee The ackee fruit (Blighia sapida) of Jamaica, which is known in Nigeria as “isin,” grows on a tree 15 to 25 feet high. Even the unripe fruit is attractive, having a bright pinkish red outer pod. The ripe fruit splits

SOME UNUSUAL FOODS 18] open and then shows two or three light yellow arilli in the bottom of which are embedded the hard black seeds. The fruit is an extremely popular food. A branch of the ackee tree with leaves and fruit is to be found on the Jamaican “Independence” 5d stamp. It is said that in general the housewife is aware of the health hazard of eating the arilli of the unripe fruit, but nevertheless poisonings occur every now and then when the fruit is not carefully selected. The ackee is usually boiled, but because the arilli are very delicate, the cooking time is usually rather short—15 to 20 minutes. The cooking water is said to be used for other dishes. Sometimes ackee is fried in coconut oil. In many public places it is sold ‘‘ready to eat,”’ although such places as supermarkets do not seem to carry it. It has been found that the fruit contains a water-soluble substance, hypoglycin, a-amino-§-methylene-cyclopropanyl-propionic acid,!® that can cause acute hypoglycemia, which seems to be responsible for the “vomiting sickness” that has been reported from Jamaica so often. Jelliffe and Stuart published an excellent review on this disease!® and at the same time reported on the treatment with intravenous glucose in- jections of five young patients all having low blood sugar levels. Al- though one patient died, the others recovered. This is rather different from the bongkrek poisoning mentioned elsewhere in which hypo- glycemia is also found, but in which glucose injections usually are not effective. The authors report that “vomiting sickness” usually occurs in winter when food is relatively scarce and poverty and “‘subnutrition” are frequent. The mortality is high, death usually occurring after 12 hours. It should be kept in mind that the term “‘vomiting sickness”’ is used rather loosely in the country and that sometimes other products may be the cause of similar poisonings.?! Recently, it has been suggested that the ackee toxin (hypoglycin) may act as a riboflavin antimetabolite.20 REFERENCES 1. E. Sergent, “‘Les cailles empoisonneuses dans la bible, et en Algerie de nos jours,” Arch. Inst. Pasteur Algerie, 19 (No. 2), 161 (1941). 2. A. G. van Veen and A. J. Hyman, “On the Toxic Component of the Djenkol- bean,” Geneesk. Tydschr. Nederl. Indie, 73, 991 (1933); 76, 840 (1936). 3. A. G. van Veen and H. E. Latuasan, “The State of Djenkolic Acid in the Plant,” Chronica Naturae (Indonesia), 105, 288 (1949). 4. M. D. Armstrong and V. du Vigneaud, *““A New Synthesis of Djenkolic Acid,” J. Biol. Chem., 168, 373 (1947).

182 nu r 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. A. G. VAN VEEN L. Lewin, Uber Piper methysticum (Kawa), August Hirschwald, Berlin (1886). W. Borsche and M. Gerhardt, “Untersuchungen iiber die Bestandteile der Kawa- Wurzel, I,’’ Chem. Ber., 47, 2904 (1914); W. Borsche and B. K. Blount, “*Unter- suchungen iiber die Bestandteile der Kawa-Wurzel, XIII,” Chem. Ber. 66, 803 (1933). G. Madaus, Lehrbuch der biologischen Heilmittel, G. Thieme, Leipzig (1938). A. G. van Veen, “‘Over de bedwelmende stof uit de kawa-kawa of wati plant (Piper methysticum),’’ Geneesk. Tydschr. Nederl. Indie, 78,1941 (1938); “*Isolation and Constitution of the Narcotic Substance from Kawa-Kawa (Piper me- thysticum),” Rec. Trav. Chim., 58, 521 (1939); “On the Isolation of the Soporific Substance from Kawa-kawa or Wati,” Proc. Roy. Acad. Sci. Amsterdam, 41, 855 (1938). . K. Morris, “Le Leucaena glauca (tamarin sauvage) faisant tomber les poils des animaux qui s’en nourrissent,” Rep. Pharm., 9, 364 (1897); ref. Tropenpfian- zer (1897), p. 169. A. G. van Veen, “Lamtoro en Kaalhoofdigheid, Natuurw. Tydschr. Nederl. Indie, 101, 55 (1941). D. Kostermans, “‘Lamtoro-zaden (Leucaena glauca) en Kaalhoofdigheid,” Geneesk. Tydschr. Nederl. Indie, 80, 2959 (1941); “‘Notes on Mimosine,” Rec. Trav. Chim., 65, 319 (1946); “The Structure of Mimosine,” Rec. Trav. Chim., 66, 93 (1947). R. Adams and J. L. Johnson, “‘Leucenol (VI), A Total Synthesis,” J. Am. Chem. Soc., 71, 705 (1949). Jung-Yaw Lin and Kuo-Hang Ling, “Isolation and Identification of Mimosine,” J. Formosan Med. Assoc., 60 (7), 657 (1961); “Studies on the Mechanism of Toxicity of Mimosine,” J. Formosan Med. Assoc., 61 (10), 997 (1962). W. Montagna and J. S. Yun, ‘“‘The Effects of the Seeds of Leucaena glauca on the Hair Follicles of the Mouse,” J. Invest. Dermatol., 40, 325 (1963). W. K. Mertens and A. G. van Veen, “Die Bongkrekvergiftungen in Banjumas,” Meded. Dienst. Volksgez. Nederl. Indie, 22, 209 (1933); A. G. van Veen and W. K. Mertens, “Uber das Toxoflavin,” Rec. Trav. Chim., 53, 257 (1934); “Uber die Giftstoffe der Bongkrekvergiftungen,” 54, 398 (1935); “Der Einfluss der Bongkreksaure auf den Kohlehydrat-stoffwechsel,” Arch. Neerl. Physiol., 21, 73 (1936). A. G. van Veen, “Bongkrek Acid, A New Antibiotic,” Doc. Neerl. Indon. Morbis Tropicis, 2, 185 (1950). W. Welling, J. A.Cohen, and W. Berends, “Disturbance of Oxidative Phosphory- lation by an Antibioticum Produced by Pseudomonas cocovenenans,”” Biochem. Pharmacol., 3, 122 (1960). C. H. Hassal, K. Reyle, and P. Feng, “Hypoglycin A,B: Biologically Active Polypeptides from Blighia sapida,’? Nature, 173, 356 (1954); E. C. De Renzo, K. W. McKerns, H. H. Bird, W. P. Cekleniak, B. Coulomb, and E. Kaleita, ‘“‘Some Biochemical Effects of Hypoglycin, Biochem. Pharmacol., 1, 236 (1959). D. B. Jelliffe and K. L. Stuart, ‘Acute Toxic Hypoglycemia in the Vomiting Sickness of Jamaica,” Brit. Med. J., 1,75 (1954). H. C. Fox and D. S. Miller, ‘“*Ackee Toxin: A Riboflavin Antimetabolite,” Nature, 186, 561 (1960). “Vomiting Sickness,” in Farmer's Food Manual, Jamaica Agr. Soc., Kingston (1957), p. 347.

ROSEMARIE OSTWALD and GEORGE M. BRIGGS Toxicity of the Vitamins The physiological effect of any food agent or drug depends on the animal species involved, on the amount of the substance administered and its route of administration, and on a number of variables con- cerning the specific individual, such as, age, sex, nutritional state, and presence of disease. The vitamins are nontoxic in the amounts and combinations ordinarily found in food sources and as normally eaten in foods. These compounds, however, are now available in pure or highly concentrated forms. They may be purchased more or less indiscriminately, either singly or in a variety of combinations and dosages, as “‘food supplements.” They are prescribed by physicians for prophylactic purposes in the treatment of specific vitamin deficiencies and of various other conditions and diseases. Because of the easy availability of vitamin preparations and their wide and increasing use it seems pertinent to make more widely known the available information concerning the effects of these compounds when used in greater than “normal” amounts. Most of the vitamins do not produce any ill effects in amounts many times greater than those likely to be ingested even as pills, powders, or drops. However, caution should be exercised in the case of some of the fat-soluble vitamins, particularly A, D, and K. This section reviews the pharmacological effects of vitamins with emphasis on clinical data. Results of animal experiments are included where relevant. No attempt has been made to cover all the literature reviewed by Molitor and Emerson,! Robinson,? Bicknell and Prescott,3 Sebrell et al.,4 Spies et al.,5 and Meyler.® 183

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186 ROSEMARIE OSTWALD AND GEORGE M. BRIGGS The data in Tables 1, 4, and 5 are compilations of the literature and represent results of investigations under different experimental condi- tions and of varying degrees of reliability. They should therefore not be taken at face value or applied to other circumstances, and should be checked for additional details by referring to the original work. FAT-SOLUBLE VITAMINS* Vitamin A One of the first cases of chronic hypervitaminosis A in man was reported in 1944 by Josephs.’ Since then, reports of at least 50 cases—most of them infants and small children—have appeared in the literature (see references 8-13). The symptoms of chronic toxicity in infants have been described as anorexia, growth failure, irritability, skin lesions, alopecia, and skeletal lesions accompanied by pain and bony swellings (hyperostosis). Enlargement of liver and spleen occurs, and occasionally an increased tendency to bleeding has been observed.!2 The only consistent change in blood constituents is a rise in vitamin A levels. Few patients showed levels below 500 international units (IU)/ml. (The normal range is 50-150 IU/ml.!3) A rise in serum alkaline phosphatase generally occurred. The symptoms in adults were similar to those seen in children, but were generally milder. Structural bone changes are less likely to occur in adults. Menstrual alterations, exophthalmos, and pigmenta- tion of the skin have been reported.!4 Disturbances of the central nervous system observed in adults and bulging fontanelles seen in infants are believed to have resulted from increases of the cerebrospinal fluid pressure. !5 The symptoms of acute vitamin A poisoning in children are vomiting, drowsiness, and bulging of the fontanelle as described by Marie and See.!6 In adults, severe headaches, vertigo, abdominal pain, vomiting, and diarrhea have been reported.!7 Cessation of the intake of excessive amounts of vitamin A in cases of acute or chronic toxicity is followed by a rapid and complete recovery in adults. Features such as anorexia, bone pain, and pruritus have disappeared within 1 to 3 days. Other effects, the result of tissue changes, disappeared more slowly. In * A summary of the recommended dietary allowances or estimated minimum requirements, therapeutic or prophylactic doses, and effects of large doses of the fat-soluble vitamins in man are given in Table 1.

VITAMINS 187 children, however, the skeletal lesions may be irreparable, and full longitudinal bone growth may not be attained.!8 In experimental animals, the most characteristic symptoms are spon- taneous fractures and internal hemorrhages. Excessive intake of vitamin A during pregnancy often leads to characteristic malformations in the young.!9-2! Jt is interesting that many of the symptoms of hyper- vitaminosis A are similar to those observed in vitamin A deficiency, for example, disturbances in the blood clotting mechanism,!2:22 skin disorders,3 congenital malformations,4 and increases of the cere- brospinal fluid pressure and hydrocephalus.> For an excellent review of the biochemistry and pathology of hypervitaminosis A_ see reference 26. The only known cases of vitamin A poisoning from naturally oc- curring food sources were reported by Rodahl and Moore27-28 and by Knudson and Rothman.!’ These outbreaks occurred when arctic explorers ate polar bear livers (in spite of Eskimo warnings). Such livers have been shown to contain up to 20,000 IU of vitamin A/g. Very high levels of vitamin A have since been reported in livers of seals, sharks, percomorph fishes, and whales.29 The chronic poisoning of children resulted from overdosing with high-potency vitamin A preparations. The adults in charge either failed to distinguish between cod liver oil (850 IU/ml) and oleum percomorphum (60,000 IU/ml) or between drops and teaspoons as measures of dosage, or they believed that if a small amount of a preparation is good for a child, a larger amount should be better. In adults, chronic poisoning has occurred most frequently when high doses of vitamin A were taken for prolonged periods as treatment for acne or other skin disorders. Sometimes the dose prescribed by the physician was very high; more often the patient, not aware of the hazard, increased the dose or prolonged the treatment beyond that prescribed. The efficacy of massive doses of vitamin A in the treatment of acne or similar skin disorders, in the absence of a vitamin A deficiency, is questionable.!2.!330 Furthermore, there is no basis for the belief that this vitamin will protect against colds. The review by Nieman and Klein Obbink?* indicates that, for adults, 1 million IU 1s a toxic dose. A dose of 100,000 IU/day for 6 months resulted only in a rise of serum vitamin A level, while 600,000 IU/day produced most of the symptoms mentioned above. From these data the chronic toxic dose for children 1 to 3 years of age was calculated to be about 100,000 IU/day, with a latent period of 6 months. More recent

188 ROSEMARIE OSTWALD AND GEORGE M. BRIGGS reviewers!3 1 report acute toxicity for infants at a dose of 350,000 IU and chronic toxicity symptoms at doses from 50,000 IU/day for children and 90,000 IU/day for adults after 12 months. Since the recommended daily allowance (RDA) of vitamin A for infants is 1,500 IU and for adults 5,000 IU, these chronic toxic doses are about 20 and 30 times the RDA, respectively. It should be pointed out that sensitivity to excess amounts of vitamin A varies widely among individuals and even in a single individual at different times.3233 This phenomenon may be partially explained by results from animal studies which showed that the absorption of vitamin A from the intestinal tract varies among individuals, and that other components of the diet as well as certain physiological variables modify the effects of large amounts of vitamin A. For instance, it has been shown that in rats, cortisone administration exacerbated its effect,34 while toxic amounts of vitamin D or moderate amounts of vitamin E or K rendered excess amounts of vitamin A harmless.!2.35 36 Vitamin A Precursors 8-Carotene and other carotenoids occurring widely in foodstuffs of plant origin are converted to vitamin A by man and other animals. Intake of large amounts of these foodstuffs may lead to hyper- carotenemia as distinct from hypervitaminosis A, the increased carotene content of the blood being accompanied by a yellow discoloration of the skin called xanthosis.26 Administration of 0.1 percent 6-carotene in the diet of four successive generations of rats over their life-spans and of 100 mg/kg of body weight/day to dogs for 3 months produced no toxic effects. These doses represent 16,000 IU of vitamin A/day/rat during the rat’s life-span and 160,000 IU of vitamin A/kg of body weight/day/dog. Signs of hypervitaminosis A were completely absent.3’ Fifteen human subjects given daily doses of B-carotene equivalent to 100,000 IU of vitamin A for 3 months showed no increase of serum vitamin A levels or any symptoms of vitamin A toxicity.38 The subject of hypercarotenemia has been reviewed.’ No reports of clinical hypervitaminosis A due to the ingestion of excess amounts of carotene have been reported, although an increase of serum lipids and a decrease of the basal metabolic rate have been observed in both condi- tions. Toxic effects of 7,000 IU of carotene*? and of large amounts of carrots,4° reported earlier, were probably the result of contamination with vitamin A, in the first case, and of other toxic factors in the second. A recent study of carotenemia in Ghanaians,‘! the result of high levels

VITAMINS 189 of carotenoids in the diet, found that the very high serum carotene levels were accompanied by very low levels of serum vitamin A. The authors suggest that high carotene intake may inhibit mobilization of liver vitamin A stores. Carotenemia is not accompanied by further pathological symptoms, and disappears completely when the ingestion of excess carotenoids is discontinued. More cases of hypervitaminosis A than of vitamin A deficiency occur today in the United States. A greater awareness of the potential dangers, by physicians and patients alike, could completely eliminate these con- ditions. One helpful step in this direction would be adequate warnings on the labels of vitamin A concentrates. Vitamin D Toxic effects have been observed in man and animals when vitamin D was ingested in excessive amounts. The symptoms of vitamin D toxicity in man are anorexia, nausea, vomiting, diarrhea, headache, polyuria, and polydipsia (see reference 5, page 399 and references 42 and 43). Calcium levels are increased in serum and urine, and calcium is dee posited in soft tissue. The primary target is the kidney, but deposits also appear in heart, lung, large blood vessels, and other organs. These deposits can lead to hypertension, renal failure, and death. In the early stages of chronic toxicity, the bones may show accelerated calcification of the provisional zone of calcification, while in more advanced cases interference with cartilage growth and demineralization of bone occur. A hypochromic normocytic anemia with azotemia has been reported.44 (For reviews of the effects of hypervitaminosis D in experimental animals see references 45 and 46.) The clinical symptoms of vitamin D toxicity are the result of resorb- tion of calcium salts from bones, increased intestinal absorption of calcium and phosphorus, increased urinary calcium, and increased levels of blood and tissue citrate.47.48 Some evidence has been reported, in studies with man, that vitamin D increases intestinal absorption of calcium only in cases when calcium metabolism is disturbed, as in rickets, osteomalacia, and hypoparathyroidism. It occurs only secondarily to increased urinary calcium excretion in cases of over- dosage of vitamin D in normal individuals.4849 Recent work with experimental animals indicates that the primary biochemical lesion may be at the level of cell mitochondria, particularly those of the renal tubules (see references 50 and 51, page 330).

190 ROSEMARIE OSTWALD AND GEORGE M. BRIGGS The clinical effects of vitamin D toxicity are reversible if administra- tion is stopped in time. A diet low in calcium and measures designed to decrease serum calcium levels, such as treatment with EDTA, phytic acid, or cortisone will speed recovery.48 52-53 The sensitivity to vitamin D varies greatly among species and among individuals within a species. In rats on a diet high in calcium and low in phosphorus, | IU of vitamin D per day produces a physiological effect by increasing the level of serum calcium considerably. The borderline toxic dose, as measured in growing rats on a diet balanced in calcium and phosphorus, has been found to be 300-700 IU per day.® Toxic effects have been observed in chicks when 100,000 IU of vitamin D2 per day were administered. This is a thousand times the dose necessary to maintain normal mineral responses in this species, i.e., 100 IU per day. In pigs, a supplement of 250,000 IU of vitamin D per day produced growth retardation and other toxic effects. The normal requirement for pigs is about 150 IU per day.°>5 The vitamin D tolerance of human adults appears to vary con- siderably.56 For example, cases of toxicity have been reported after 25,000 IU per day, after 50,000 IU per day for a few weeks, and after over | million IU per day for 4 years.42:57-59 RDA for adults is 400 IU. On the basis of linear growth in childhood, infants can be very sensitive to the effects of vitamin D; growth has been said to be retarded by the administration of only 1,800 IU per day.®! 62 A German study showed that while a daily dose of 1,000—1,500 IU to each of 300 infants pro- tected all of them from rickets, it was dangerously high for 84 percent of them.® On the basis of these data, the minimum toxic dose is cal- culated to be five to ten times that recommended for optimal growth and rickets prevention.® © This statement appears to be controversial, however, since the Committee of the American Academy of Pediatrics has set the toxic dose of vitamin D at 1,000-3,000 IU/kg of body weight/day for several weeks or months—a figure many times that quoted above. Individual variability in sensitivity to vitamin D may result in part from the influence of certain physiological states and nutrients on the effects of vitamin D. Hypo- and hyperparathyroidism will alter calcium and phosphorus metabolism, thereby influencing the effects of added vitamin D. Diseases interfering with fat absorption, and therefore with the absorption of vitamin D, are obviously important in this connection. Studies with rats indicate that pregnancy protects the mother against vitamin D toxicity.°7 On the other hand, some authors warn against large intakes of calcium and vitamin D for pregnant women because

VITAMINS 191 of possible damage to susceptible embryonic tissue from calcification.® Hypothyroidism, with its associated elevated serum cholesterol level, has been reported to lead to increased sensitivity to vitamin D.§9 There is evidence of an interrelationship between vitamin D toxicity, hypercholesteremia, and hypercalcemia, with associated calcifying arteriopathies such as arteriosclerosis. Increased serum cholesterol levels were found in hypervitaminotic rabbits.7°7! Simul- taneous feeding of excess vitamin D and excess cholesterol to rabbits led to more atheromatosis than when either was fed alone.72 The production of atherosclerosis in animals by a diet rich in vitamin D has been reported frequently.49:7!:73-75 Elevated serum cholesterol levels have been reported in some cases of hypercalcemia in children’6-77 but not in others.7”8 The level of calcium and phosphorus intake is obviously important in relation to the effects of excess vitamin D. Among other dietary components, the amount and kinds of fat in the diet influence the absorption of vitamin D. High levels of vitamin A have been reported to afford some protection, in rats, from the effects of excess doses of vitamin D,’9 as does choline.3°.8! Linoleic acid appears to counteract the toxic effects of vitamin D in essential fatty acid- deficient animals.82.83 This, however, is probably a nonspecific stress effect.84 While normal individuals react to excess doses of vitamin D with varying degrees of sensitivity, normal amounts of vitamin D may be at least partly responsible for certain diseases in hypersensitive individuals. Idiopathic hypercalcemia of infancy is a disease that, in mild cases, resembles vitamin D toxicity and in severe cases is associated with physical and mental retardation, osteosclerosis, renal and cardiac damage, and early death. In studies of this disease, there was usually no evidence that abnormally high amounts of vitamin D had been administered, and in some cases the calculated intakes were less than 1,000 IU per day. No cases have been reported in exclusively breast-fed infants. Epidemiological studies, however, implicate vitamin D in the etiology of this disease. During 1953, 1954, and 1955, 100 cases per year were reported from Britain, but only 12 cases from the United States. During that period, the milk in Britain was fortified to an uncer- tain extent in excess of the minimal limit, then stipulated to be 1,600 IU per quart. In the United States, milk was then and is now fortified with 400 IU per quart. Cases in England decreased markedly after 1957 when vitamin D fortification of milk and of infant cereals was reduced and an average content of 600 IU per quart of milk was pre- scribed.53:65.77.85.86 Other possible etiologies for this disease have been

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VITAMINS 193 proposed, e.g., disturbance of cholesterol metabolism leading to sterols with vitamin D-like or toxic effects’6 and derangement of calcium metabolism.®7 A hypersensitivity to ingested vitamin D has also been proposed as cause for some cases of sarcoidosis*® 88 and as contributing to develop- ment of atherosclerosis‘? 6.89 and of some cases of gastric ulcers, the latter a result of the effect of vitamin D on the acid-secreting cells of the gastric mucosa. 9° Preformed vitamin D occurs primarily in foodstuffs of animal origin, but it is present in appreciable amounts only in the livers of certain fishes and in animals that feed on them (Table 2). It is formed in the animal body by ultraviolet irradiation of the provitamins ergosterol, obtained from plant sources, and dehydrocholesterol, obtained either from animal sources or by endogenous synthesis from cholesterol. There is no evidence that either ingestion of even very large amounts of foods containing these precursors or excessive exposure to ultraviolet light leads to vitamin D toxicity. With the possible exception of the hypersensitivity mentioned above, most cases of vitamin D poisoning have occurred through the intake of excess doses of vitamin D con- centrates. Adequate warnings against this danger, both by the pre- scribing physician and through labels on the preparations, would be very desirable. Large doses of vitamin D are prescribed in the treatment of rickets, rheumatoid arthritis, tuberculosis, asthma, scleroderma, and other diseases. Its efficacy, except in the case of rickets, is today considered doubtful.58 A chemical derivative of vitamin D, dihydrotachysterol (AT 10), which is used in the treatment of hypoparathyroidism, can also lead to vitamin D poisoning. Both vitamin D and AT 10 should be taken in therapeutic doses only under close supervision by a physician. The minimal human requirement for vitamin D is not well estab- lished. The latest RDA® are 400 IU per day for infants, children, and pregnant or lactating women. Healthy adults are presumed to receive vitamin D or its precursors sufficient for their needs from their food and normal exposure to sunlight. In the United States most of the commercially available milk, except dry skim milk, is fortified to con- tain 400 IU of vitamin D per quart. Many breakfast cereals, proprietary infant foods, and other foods are also fortified with vitamin D (Table 2). When supplementing an infant’s or child’s diet it is therefore necessary to estimate the intake of vitamin D from all sources in order to avoid overdosages.*! For instance, it is possible for an infant receiving 1.5

194 ROSEMARIE OSTWALD AND GEORGE M. BRIGGS pints of fortified milk, 1 ounce of fortified cereal, and 1 teaspoonful of fortified cod liver oil to have an intake of about 4,000 IU92 ten times his RDA (Fortification levels referred to are those common in England around 1955). An intake not exceeding 1,000 IU per day has been recom- mended because this level seems to be best suited to protect most children against rickets, caused by a lack of vitamin D, and against hypercalcemia, caused by an excess. The situation of the elderly adult with respect to vitamin D is not clear. On the one hand, there is a tendency with aging to osteoporosis, which leads primarily to an increase in the resorbtion of bone.* This demineralization has reportedly been slowed in certain individuals with the aid of a diet rich in calcium and, perhaps just as important, addi- tional vitamin D.95 On the other hand, hypercholesteremia, athero- sclerosis, and other calcifying diseases associated with increasing age may be exacerbated by intakes of large amounts of calcium and vitamin D, especially in certain hypersensitive individuals. The optimum dose of vitamin D and calcium for this age group is still under discussion.%-*? Vitamin K Vitamin K was previously believed to be innocuous at any dose level. Studies with experimental animals and clinical experience, however, have shown that this is not always the case. The effects produced depend on the particular preparation used and the route of administration. The oral acute LDso in mice was found to be 0.2 g/kg of body weight for phtiokol* and about 0.5 g/kg for menadione, Synkayvite, or its sodium-bisulfite salt. No lethal effect up to 25 g/kg was observed for vitamin K, (reference 3, page 694). Similarly, chronic toxicity in rats was found at 0.35 g/kg of body weight for phtiokol and 0.5 g/kg for menadione, but with the same dose of vitamin K; for 30 days no adverse effects were observed.%-99 Toxicity from subcutaneous administration of water-soluble analogues of vitamin K is three to five times that resulting from oral doses (reference 3, page 694). The production of hyperthrombinemia by oral doses of menadione (5 mg/kg, rabbit; 10 mg/kg, dog; and 20 mg/kg, rat) have been reported.!©° In rats, intra- muscular injection of a water-soluble analogue produced hemo- globinuria,! classical hemolytic anemia,!°2 and hepatocellular damage.!® On the other hand, no hemolysis was produced in rats either by intraperitoneal administration of vitamin K; or by oral administra- tion of water-soluble analogues.!1 * See Table 3 for chemical names for the different vitamin K preparations.

VITAMINS 195 TABLE 3 Some Common Vitamin K Preparations—Their Names, Analogues, Solubilities, and Routes of Administration COMMON OR SOLUBLE ROUTES OF TRADE NAMES CHEMICAL NAMES IN ADMINISTRATION ® Vitamin Ky 2-Methyl-3-phytyl-1 , 4-naphtho- Fat IM, IV, or Oral (phylloquinone, quinone phytonadione, Konakion Mephyton) Phtiokol 2-Methyl-3-hydroxy-1,4-naphtho- Water quinone Vitamin K2 2-Methyl-3-difarnesyl-1 , 4- Fat naphthoquinone Vitamin K3 2-Methyl-1, 4-naphthoquinone Fat Oral, IM (menadione, menaphtone) Hykinone Menadione sodium or potassium Water Oral, SC, IM, IV bisulfites or sulfonates Menadiole 2-Methyl-1 , 4-naphthohydroquinone Various trade Menadiole sodium or potassium Water All routes names di- or tetraphosphates, sulfonates, and bisulfites «IM, intramuscular; IV, intravenous; SC, subcutaneous. A recent study with intraperitoneal injections of Synkayvite dem- onstrated hyperbilirubinemia in newborn rats at 20 mg/kg of body weight and hemolysis at 80 mg/kg. In adult rats, 80 mg/kg was required to produce a rise in bilirubin, and 160 mg/kg was required to produce signs of hemolysis.!% The chronic toxic effects in animals result from injury to the circu- lating red cells and aplastic anemia.99!05 Hypertrophy of the hypophysis,!% respiratory depression,!°? and focal hemorrhages! have been reported. Vitamin K has been used in the prevention and treatment of hemor- rhagic disease of the newborn and in the treatment of certain other hemorrhagic conditions. It has been found effective only if the condi- tion resulted from a prothrombin deficiency, caused either by over- dosage with anticoagulants such as dicoumarol, or by a relative vitamin K deficiency resulting from malabsorption or inadequate supplies of bile salts. In adults, considerable amounts of vitamin K are synthesized in the intestinal tract. In newborn infants, the intestinal flora is not well

196 ROSEMARIE OSTWALD AND GEORGE M. BRIGGS established and, therefore, adequate amounts of vitamin K may be lacking. Prematurity and anoxia increase the susceptibility to hemor- rhage. The routine administration of a vitamin K preparation either to women in labor or to the newborn as a prophylactic measure against hemorrhage has been both advocated and questioned (reference 5, page 375 and reference 60). The validity of evidence both as to the efficacy and safety of supplementing diets of pregnant women with vitamin K has also been questioned, and the U.S. Food and Drug Administration (FDA) has barred the use of menadione in vitamin capsules and other food supplements for pregnant women because of possible dangers when used in excessive amounts. The evidence for toxic effects of excess amounts of vitamin K in humans, including infants and mothers, has been reviewed (see reference 92, page 318 and references 109-111). From this, it appears that a total dose of 10 to 30 mg of a water-soluble analogue is often associated with unconjugated hyperbilirubinemia and subsequent kernicterus. The effects are observed more often in premature infants than in full-term infants and when given parenterally rather than orally. The mechanism of the hyperbilirubinemia is uncertain. Some authors believe that an increase in hemolysis is involved ;!!2 others consider the possibility of hepatocellular toxicity!!3 or an inhibition of glucoronide formation with subsequent accumulation of nonconjugated bilirubin.!!4 Hyper- thrombinemia in man as a result of excess vitamin K has only rarely been reported.!°9 Nausea and vomiting occurred when 3 mg/kg of body weight of menadione were given, and severe dermatitis has been re- ported to occur frequently when the pure drug comes in contact with the skin (see reference 92, page 319). In some cases, a hypothrombinemia in response to large doses of vitamin K has been observed, especially in patients with liver diseases.!!5 Effects of the different vitamin K compounds vary. Vitamin Ky, administered either intravenously or orally, decreased prothrombin time to safe values in cases of overdosages with anticoagulants, while water-soluble analogues proved ineffective as antidotes (see references 51, page 434; 92, page 315; and 116). On the other hand, there is little difference between the action of vitamin K, and that of Synkayvite, given either orally or intravenously, in the treatment of hemorrhagic disease of infants. In cases of severe liver damage, however, administra- tion of any vitamin K by any route may not produce a prothrombin effect (see reference 92, page 319). The minimal requirements of vitamin K for infant, child, mother, or normal adults are unknown. It is known, however, that 1-2 mg of

VITAMINS 197 compounds exhibiting vitamin K activity will correct deficiency symp- toms in most cases. A daily allowance for vitamin K has not been established because of the wide but inconsistent distribution of the vitamin in the diet and because of the variability of need, which depends on such factors as synthetic activity of the intestinal flora, absorption capacity of the individual, and the state of the individual’s hepatic functions (reference 5, page 368). The spread between effective preven- tive or therapeutic doses and those producing toxic effects is wide enough to present no hazard if these compounds are used wisely. The Food and Nutrition Board of the National Academy of Sciences—National Research Council® has recommended a single oral dose of 1.0-2.0 mg or a parenteral dose of 0.5—1.0 mg of vitamin K, or of the newer water-miscible preparations for the prevention of hemor- rhagic disease in infants. The Council on Drugs of the American Medical Association!!7 has recommended that not more than a single dose of a water-soluble vitamin K analogue equivalent to 1 mg of menadione be given for the same purpose. Vitamin E Vitamin E is a relatively nontoxic substance. Toxicity produced in animals resulted in symptoms such as decreased thymus weight and increased adrenal weight in rats,!!8 hyperthyroid effects in rats and rabbits,!!9 adrenal degeneration and growth inhibition in chickens,!20 and sensitization and various tissue lesions in guinea pigs.!2!.122 The major reactions to toxic levels in most species, however, were in the reproductive organs of both males and females. These have been de- scribed for mink,!2 voles,!%4 and rats, !18 125 .126 Tocopherol toxicity is usually considered not to occur in man (reference 92, page 293). Beckman,!27 in his exhaustive review, states that he has never seen a case of hypervitaminosis E even though he has given 300 mg per day, both by mouth and parenterally, for many months to a number of patients. His review shows that disturbances of reproductive functions occurred at dosage levels that led to the recom- mendation not to use more than a total of 4 to 12 g of a-tocopherol. A heroic volunteer!28 took a total of 296 g of a-tocopherol over a period of 93 days. He observed a doubling of serum tocopherol levels and a transient creatinuria. Clinical symptoms were chapping, cheilosis, angular stomatitis, frequent gastronintestinal disturbances, and vague generalized muscle weakness. All symptoms disappeared 2 weeks after tocopherol intake had been discontinued. Another author reports doses

198 ROSEMARIB OSTWALD AND GEORGE M. BRIGGS of 40 g per day to have had no ill effects.'29 Untoward symptoms, including hypoglycemia and depression of prothrombin levels, have been reported, however, at doses of 300—1,500 mg per day.!30—133 Thus man apparently has a considerable tolerance for tocopherol, and creatinuria may represent an early index of metabolic malfunction. It is interesting to note that the two best-established effects of vitamin E toxicity, disturbances of reproductive function and creatinuria, are also symptoms of vitamin E deficiency. Vitamin E treatment has been attempted for many diseases in the hope that it might be effective in curing human diseases similar to those seen in animals with a vitamin E deficiency. A critical analysis of these attempts suggests a well-founded evidence for tocopherol therapy only in the following conditions:!4 intermittent claudication, fat malabsorp- tion syndromes, and supplementation of diets for certain newborn infants and of diets containing large amounts of unsaturated fats. Tocopherol administration has been reported helpful also in certain anemias in infants with protein-calorie malnutrition.°5 Beneficial effects in the treatment of habitual abortion and of stasis ulcers have not been definitely established. The human dietary requirement for vitamin E is unknown. The mean intake in the United States has been estimated to be 14 mg per day.1% Recent work with a group of human subjects indicates an increased need for vitamin E when high levels of unsaturated fatty acids are ingested, for example, in certain diets recommended for the control of serum lipid levels in people prone to atherosclerotic heart disease.!37 Horwitt!37 recommends doses of 5 mg per day to as much as 30 mg per day, depending on the tissue and dietary levels of linoleic acid. The same levels are suggested by the NAS-NRC Food and Nutrition Board.6° WATER-SOLUBLE VITAMINS The water-soluble vitamins, in which we include the B group, ascorbic acid, and choline, are nontoxic in the amounts available from ordinary food sources. They are also innocuous in the amounts generally used for the prevention or correction of a specific deficiency. They do, how- ever, have pharmacological and toxic effects when administered in very large amounts as “drugs” or pharmacologic agents. The earlier literature has been reviewed by Molitor and Emerson,! Robinson,?

VITAMINS 199 Bicknell and Prescott,3 Sebrell et al.,4 and Nutrition Reviews.!38 Tables 4 and 5 summarize some of the available data. Some of these vitamins exert certain pharmacological, toxic, or nonspecific effects. These observations are discussed below. TABLE 4 Water-Soluble Vitamins:* Recommended Daily Allow- ance or Estimated Minimum Requirements, Therapeutic Doses, and Effects of Very Large Doses in Man? THERAPEUTIC EFFECTS OF VERY VITAMIN RDA‘ DOSES? LARGE DOSES Biotin 300 ug* 75-300 pg/day,IM — Pantothenic 10 mg* 20-40 mg/day, IM 100 mg, IV, NE* acid Pyridoxine 1-2 mg* 200-400 mg, IV 200 mg, IV, NE! Riboflavin 0.6 mg/ 6 mg, OS — 1,000 cal 6-25 mg, IM Thiamine 0.4 mg/ 50 mg/day, SC, IM 500 mg, IV, NE* 1,000 cal 10 mg, OS 10-100 mg, par, side effects¢ Niacin or 6.6 mg 150-500 mg/day, OS 100-300 mg, OS nicotinamide N-equiv/ 2-6 g/day, OS 20 mg, IV* side effects 1,000 cal 100 mg, IV 2-300 mg, OS, flushing¢ Folic acid 0.1-0.2 mg* 15-20 mg, OS 400 mg, OS, 5 mo, NE* 5 mg, OS 10 mg, OS, 5 yr, NE Br 3-5 ug* 10-15 ug/day, 1 mg/day, IM, 10 days, NE* SC, IM 500 pg /wk, IM!74 1 mg/day, IV, 3 days¢ 1 mg/day, OS, 3-5 yr, NE’ Ascorbic acid 70 mg 100-500 mg/day, OS 6 g/day, OS, NE* 0.5-1.0, IV, NE 1 g/day, OS, side effects!38 Choline — — 50 mg/day, 7 days, NE « The data are compilations of the literature and represent results of investigations under differen: experimental conditions and of varying degrees of reliability. For references, see bibliographies of the individual vitamins. » Abbreviations used: T, toxic effects observed; NE, no effect observed; OS, oral; IM, intramuscular; IV, intravenous; SC, subcutaneous; par, parenteral administration; wk, weeks; mo, month; yr, years (duration of treatment). ¢ Recommended Dietary Allowances as set by the NAS-NRC. Starred items are “‘estimated daily intakes” or ‘“‘estimated minimum requirements.” € Values quoted vary widely, depending on whether treatment was prophylactic or therapeutic, what condition was treated, and how far disease was advanced. For bibliography see references 1, 2, 3, and 5 of the main text. ¢G. Glass, D. Lee, H. Skeggs, and J. Stanley, ‘“‘Hydroxocobalamin 3. Long-Acting Effects of Mas- sive Parenteral Doses on Vitamin Bi: Blood Levels in Man,” J. Am. Med. Assoc., 183, 425 (1963). fC. Conley and J. Krevans, ‘‘New Developments in the Diagnosis and Treatment of Pernicious Anemia,”” Ann. Intern. Med., 43, 758 (1955). ¢ P. McIntyre, R. Hahn, J. Masters, and J. Krevans, ‘‘Treatment of Pernicious Anemia with Orally Administered Cyanocobalamine (Vitamin Bi:)," Arch. Intern. Med., 106, 280 (1960).

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