Folic Citrovorum factor

In humans and rats, early investigations showed that large doses of folic acid resulted in increased excretion of a substance that stimulated the growth of P. cerevisiae and that was presumed to be citrovorum factor.45 This response is now also associated with other reduced folates. 

Excretion of thiamine appears to vary from individual to individual,23 and some other data are available regarding the other better-known B vitamins.24 The differences in the excretion of nicotinic acid-like compounds strongly suggest the existence of individual pattems.25,26 The urinary excretion of vitamin B12, folic acid, and the citrovorum factor by different individuals, even on controlled diets, was found to vary through rather wide ranges (2- to 9-fold) though the study was not concerned with individual differences and individual patterns were not established. 27... 

Johnson and colleagues made a provocative observation in the course of exploratory preclinical toxicological studies of vincristine, namely, that folinic acid (Leucovorin citrovorum factor 5-formyl-5,6,7,8-tetrahy-drofolic acid) was able to protect mice from the toxicity of high doses of vincristine lb). Vincristine, at a dose of 2.5 mg/kg administered intravenously, resulted in a mortality of 90% over a period of 30 days, but treatment with folinic acid lowered the mortality to 25%. The protection against vincristine toxicity did not occur when folic acid was substituted for folinic acid. A report has appeared (45) indicating that there is no specific protective effect of folinic acid against vincristine toxicity in mice and that the protection can be observed by comparable treatment with isotonic saline solution. As discussed in Section Vll, there is not conclusive evidence that folinic acid is able to ameliorate vincristine toxicity in humans (46). 

Inhibition of nudeobase synthesis (2). Tet-rahydrofolic acid (THF) is required for the synthesis of both purine bases and thymidine. Formation of THF from folic acid involves dihydrofolate reductase (p. 274). The folate analogues aminopterin and methotrexate (amethopterin) inhibit enzyme activity. Cellular stores of THF are depleted. The effect of these antimetabolites can be reversed by administration of folinic acid (5-formyl-THF, leucovorin, citrovorum factor). Hydroxyurea (hydroxycarbamide) inhibits ribonucleotide reductase that normally converts ribonucleotides into deoxyribonucleotides subsequently used as DNA building blocks. 

D16. Doctor, V. M., In vitro studies on the conversion of folic acid to citrovorum factor. J. Biol. Chem. 222, 959-968 (1956). 

C20H23N7O7, Mr 473.45, pale yellow crystals +3 H2O, mp. 248 -250 °C (decomp.), [a]n +16.8° (aqueous hydrogen carbonate). F. occurs in various microorganisms in which it acts as a growth factor e. g., for the lactic acid bacterium Leuconostoc citrovorum, therefore, it is also known as citrovorum factor. F. functions as an active 1-C unit (cf. folic acid) and formyl group transfer agent in the metabolism of folic acid tetrahy-drofolic acid. F. is accessible by synthesis and its calcium salt is used therapeutically as an antidote for folic acid antagonists such as methotrexate. 

Folic Acid (Pteroylglutamic Acid) and Citrovorum Factor... 

Another member of the folic acid group of vitamins, citrovorum factor or folinic acid, is also found in natural materials, both in free and combined form. Citrovorum factor is believed to be a metabolically active form of folic acid and is formed in the body from folic acid. The citrovorum content of foods is largely unknown. 

In vitro studies indicate that folic acid may be converted to citrovorum factor by liver slices and that this conversion is aided by ascorbic acid. Vitamin B12 may also have a role in the conversion of folic acid to citrovorum factor, and both vitamin B12 and ascorbic acid may stimulate synthesis of folic acid in the body. Relationships between vitamin B12 and folic acid are complex both function in hematopoiesis, probably in nucleoprotein synthesis, and in the metabolism of labile methyl groups and choline. 

Citrovorum factor stimulates hematologic improvement, similar to that induced by folic acid, in sprue, nutritional macrocytic anemia, macrocytic anemia of pregnancy, pernicious anemia and megaloblastic anemia of infancy. Whether this factor is more active than folic acid has not been determined. Neither folic acid nor citrovorum factor will prevent hematologic relapse or the development of neurologic lesions in pernicious anemia. Folic acid is helpful at times in the therapy of macrocytic anemia associated with cirrhosis of the liver. 

Bussi et al. (1953) and Cox (1953) divided pernicious anemia serum into protein and nonprotein fractions by ultrafiltration. The inhibitor was usually in the ultrafiltrate. Cox (1953) found that the protein fraction, rid of this inhibitor by ultrafiltration, had maturing properties. Microbiological assays showed that this fraction contained not only bound vitamin B12 but bound forms of the folic acid group of substances including citrovorum factor (Cox, Hornsby et al., unpublished). 

To this effect would be added the antianemic effect of the folic acid and citrovorum factor contained in the liver. Folic acid does not increase the absorption of vitamin B12 (page 183). The unlikelihood that it potentiates the hemopoietic effect of the vitamin is considered on page 183. 

Liver extracts for parenteral use were given in amounts 40 to 80 times less than those required when the same extract was administered orally. The amounts of folic acid and citrovorum factor in a parenteral dose thus became negligible by comparison. There is now no doubt, however, that vitamin Bi is fully effective in pernicious anemia and subacute combined degeneration of the cord without the need for accessory factors such as folic acid and citrovorum factor. 

The Citrovorum Factor is a growth factor for Leuconoatoc citro-vorum. It may be the active form of folic acid. Synthetic citrovorum factor (folinic acid, leucovorin) is formyl tetrahydrofolic acid. Conversion of pteroylglutamic acid to the citrovorum factor is believed to be helped in vitro and in vivo by ascorbic acid (see pages 186 and 188). 

In an untreated case of pernicious anemia, the folic acid content of the liver was found to be present as such, whereas in normal livers the citrovorum factor predominated (Girdwood, 1952a). It is not yet certain, however, whether B12 aids in the conversion of pteroylglutamic acid to the citrovorum factor. In any case, this is not likely to be the chief role of vitamin B12. 

Treatment of subacute combined degeneration with masmve doses of citrovorum factor did not prevent neurological deterioration (Ungley, unpublished), and it appears very unlikely that this compound will protect the nervous system in pernicious anemia. Treatment with folic acid or citrovorum factor has no effect on the serum Bw levels of patients with pernicious anemia (Mollin and Ross, 1953b). 

X. The Interrelationship between Vitamin Bij, Folic Acid, THE Citrovorum Factor, and Ascorbic Acid... 

This subject is discussed in reviews by E. L. Smith (1951,1954), Welch and Heinle (1951), Vilter and Mueller (1952), Welch and Nichol (1952), and Nieweg et al. (1954). Both vitamin B12 and the folic acid group of substances (which includes citrovorum factor) are concerned in the formation and transfer of one-carbon intermediates and in the synthesis of... 

May et al. (1950a,b, 1951) originally considered that ascorbic acid was necessary for the conversion of pteroylglutamic acid to citrovorum factor. In monkeys fed on diets deficient both in ascorbic acid and folic acid, megaloblastic anemia developed. This could be relieved by ascorbic acid, by folic acid in large doses, or by small doses of citrovorum factor. Later May et al. (1953) reported that ascorbic acid was not required for the conversion of folic acid to citrovorum factor. The severe deficiency of folic acid compounds occurring in scorbutic monkeys was probably due to nonspecific factors operating in scurvy. 

Nevertheless, others have shown that ascorbic acid aided the conversion of folic acid to citrovorum factor in rat liver slices (Nichol and Welch, 1950) and that the urinary excretion of citrovorum factor by rats or human beings given pteroylglutamic acid was increased by the administration of ascorbic acid (Anker et al., 1950 Welch et al., 1951a,b). The role of vitamin Bij and other hemopoietic factors for hemopoiesis in monkeys is discussed by May et al. (1952a,b, 1953) and by S. C. Smith and Elvehjem (1951). 

Very rarely an anemia, thought to be pernicious, is associated with a deficiency of folic acid severe enough to prevent the initial response to vitamin Bis. Ungley (1952) has described two such cases, and Cox and Ungley (unpublished) have seen a third. These three patients did not respond initially to vitamin Bis until another hemopoietic agent such as folic acid or citrovorum factor bad been supplied. Thereafter and in... 

Wallerstein et al. (1953b) found a severe biochemical deficiency of ascorbic acid in pernicious anemia. They followed reticulocyte responses to Bi2 or folic acid without, and then with, ascorbic acid. They concluded that ascorbic acid may play an adjuvant role in hemopoiesis, probably by enhancing the production of citrovorum factor. 

Many cases are refractory to vitamin B12 (Israels and Sharp, 1950 Tuck and Whittaker, 1950). Some of the cases studied by Cox et al. (unpublished) have responded poorly or not at all to vitamin B12 (Fig. 9). In all the refractory cases so far reported, the patients have responded to folic acid or to the citrovorum factor, but not all attained normal blood values, even after prolonged treatment. In patients undergoing treatment with folic acid, cerebral, spinal, or neural disorders occasionally develop sometimes these can be arrested or alleviated by administering vitamin Bi2. Cloake et al. (1954) describe 18 patients with a variety of neurological disorders associated with steatorrhea. None had all the essential characteristics of true subacute combined degeneration of the cord. Results of treatment with vitamin B12 or other B vitamins varied unaccountably. 

Fig. 9. Case of malabsorption syndrome. No response to 10 jug. vitamin Bij either before or after administration of folic acid and citrovorum factor. Good response to 40 mg. citrovorum factor with increase in body weight (Cox, E. V., and Ungley, C. C., unpublished).

In two cases reported by Davidson and Girdwood (1951) the anemia responded to the citrovorum factor. The dosages used were not appreciably less than those commonly used for folic acid, e.g., see Isra s and Da Cunha (1952). Our experience in six patients has been similar (Cox... 

Response to folic acid and citrovorum factor should not be taken to mean that the cause was necessarily a deficiency of these nutrients the fallacy of such reasoning became apparent from experiences with folic acid in Addisonian pernicious anemia. It is not even certain that all megaloblastic anemias of pregnancy are of nutritional origin (Thompson and Ungley, 1961). 

In most instances the primary deficiency is in folic acid, or there may be a combined deficiency of folic acid and of ascorbic acid (May et al., 1950a Zuelzer and Ogden 1946a,b Zuelzer, 1946, 1947 Aldrich and Nelson, 1947). The citrovorum factor was effective in doses as small as 75fig. daily (Woodruff et al., 1951). As might be expected, some infants failed to respond to vitamin B12 examples are given by Luhby and Wheeler (1949) and Woodruff et al. (1949, case 3). 

Zuelzer and Rutzky (1953) conclude that the majority of infantile megaloblastic anemias are due essentially to a deficiency of folic acid rather than to that of vitamin B12. Some infants show an incomplete response to vitamin B12 followed by an adequate response to folic acid or the citrovorum factor. In such patients there is probably a combined deficiency. 

Since vitamin B12 may fail, the proper course in treating infants with megaloblastic anemia is probably to give foUc acid, which is always effective. As there may be an associated deficiency of ascorbic acid and vitamin B12, these agents may be administered too (May et al., 1950a). The possible role of these two substances in the conversion of folic acid to citrovorum factor is discussed on pages 186 and 188. 

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