RIBOFLAVIN-5'-PHOSPHATE
Explanation
Riboflavin was evaluated for acceptable daily intake (ADI) by the
Joint FAO/WHO Expert Committee on Food Additives in 1970 (see Annex,
Ref. 19). A toxicological monograph was published in 1970 (see Annex,
Ref. 20). Riboflavin-5'-phosphate has not been previously evaluated.
However, since riboflavin-5'-phosphate is rapidly hydrolysed to yield
riboflavin after ingestion, and riboflavin and riboflavin-5'-phosphate
are in metabolic equilibrium after absorption, the available toxicity
data are combined in a single monograph.
BIOCHEMICAL DATA
BIOLOGICAL ASPECTS
Riboflavin is essential for all animals and many microorganisms.
Riboflavin-5'-phosphate is the prosthetic group of flavoproteins
involved in general cell metabolism as hydrogen acceptors. It occurs
naturally throughout the plant and animal kingdom.
Riboflavin-5'-phosphate or flavin mononucleotide (FMN) is rapidly
dephosphorylated to free riboflavin by incubation with intestinal
mucosa or intestinal juice from rats (Christensen, 1969).
Lumichrome and lumiflavin were identified as metabolites of
riboflavin in the rat (Yang & McCormik, 1967); hydroxyethylflavin,
formylmethylflavin and an unknown metabolite were identified as
metabolites in male volunteers (West & Owen, 1969).
TOXICOLOGICAL STUDIES
Special studies on mutagenesis
Riboflavin and FMN were found not to be mutagenic to Salmonella
typhimurium strains TA-98, TA-100, 1535, 1537 or 1538 or to
Saccharomyces cerivisiae strain D4. Both suspension and plate
overlay tests were conducted and assays were done with and without
mammalian activation systems (Litton Bionetics, 1977a, 1977b).
Special studies on reproduction
Weanling male and female rats were fed daily doses of 10 mg of
riboflavin for 140 days. The animals were mated and normal litters
were obtained from the riboflavin and control groups. At three weeks
of age the offsprings of the first generation were fed daily with
10 mg of riboflavin. Daily feedings over periods of 140 days were
continued for three generations. There were no differences in the
development, growth, maturation, reproduction of treated and control
animals. Autopsies at the end of the test period did not show any
gross changes (Unna & Grislin, 1942).
A group of 13 female rats were fed diets containing 100 ppm
(0.01%) of riboflavin, for two weeks, and then bred. The diet was
maintained during gestation and lactation. Control rats received 4 ppm
(0.0004%) of riboflavin in the diet. The number of litters in the high
riboflavin group was less than control. The average birth weight,
number of young litter, and average weight at weaning was similar for
test and control animals. However, there was an apparent decrease in
the viability of the offspring in the high riboflavin group as a
result of the loss of one litter (Schumacher et al., 1965).
In another study group of young female rats (Wistar strain) were
fed diets containing 4 or 40 ppm (0.0004 or 0.004%) of riboflavin
during pregnancy and lactation. There were no significant differences
in the number per litter, mortality or weight gain of offspring, in
the groups (Le Clerc, 1974).
Acute toxicity
LD50
Compound Animal Route (mg/kg bw) Reference
Riboflavin Mouse i.p. 340 Kuhn & Boulanger,
1936
Monodiethanolamine
salts of FMN Mouse Oral 6 000 Randal, 1950
s.c. 800 Randal, 1950
i.v. 600 Randal, 1950
Rat Oral 10 000 Unna & Greslin, 1942
s.c. 5 000 Unna & Greslin, 1942
i.p. 560 Unna & Greslin, 1942
Dog Oral 2 000 Unna & Greslin, 1942
Sodium Rat Oral 10 000 Unna & Greslin, 1942
riboflavinate
s.c. 790 Unna & Greslin, 1942
i.p. 560 Unna & Greslin, 1942
The high oral LD50 in rats is probably due to poor absorption
from the gastrointestinal tract and the low solubility of riboflavin.
Parenteral administration of 0.6 g/kg bw leads to renal obstruction of
pelvis and collecting tubules with crystals of riboflavin and death
from renal failure and weight loss (Unna & Geslin, 1942).
Short-term studies
Rat
The monodiethanolamine salt of FMN was fed to groups of 10
weanling female Sprague-Dawley rats five days weekly for 29 weeks at
dose levels of 1, 4, 10 and 40 mg daily (approximately 5, 20, 50 and
200 mg/kg bw). No effects on growth or haematology were noted at the
5 or 20 mg/kg levels. At 50 mg/kg there was a slight decrease in
haemoglobin concentration. At 200 mg/kg, two rats died and the
surviving eight animals showed slight anaemia and decreased weight
gain (Randall, 1950).
Rabbit
Groups of four rabbits received 10 or 100 mg (5 or 50 mg/kg bw)
of monodiethanolamine riboflavin by intravenous or intramuscular
injection, five days per week for three weeks. One of the rabbits died
with evidence of renal damage following seven intravenous injections
at 50 mg/kg. No toxic effects were noted after intramuscular injection
(Randall, 1951).
Dog
Four dogs, 10 weeks of age were fed 25 mg/kg of riboflavin daily
over a period of five months. Growth was normal and no toxic effects
were observed. Autopsies at the end of the test period failed to
reveal any macroscopic changes in the organs (Unna & Grislin, 1942).
OBSERVATIONS IN MAN
When 5-500 mg of the sodium salt of FMN was administered orally
to human volunteers an increase in free riboflavin was noted in the
plasma and urine. However, absorption seemed to be due to a saturable
mechanism since an increase in urinary excretion did not occur at
doses greater than 50 mg (Stripp, 1965).
Increased absorption of FMN occurs in man if the substance is
given with a meal, probably due to increased intestinal transit time.
FMN and riboflavin are likely absorbed by a specific transport system
in the upper gastrointestinal system. FMN may be dephosphorylated
during absorption but then may be rephosphorylated in the mucosa,
transported to the liver where it is again dephosphorylated to
riboflavin, the form in which it travels in the circulation and is
excreted. Enterohepatic recycling of riboflavin may occur (Jusko &
Levy, 1967).
The recommended daily dietary allowances for man is
0.6 mg/100 Kcal. for persons of all ages engaged in normal activity
with a daily supplement of 0.3 mg for pregnant and 0.5 mg for
lactating women (U.S. Food & Nutrition Board, 1974).
The recommended therapeutic doses for treatment of prevention of
riboflavin deficiency are:
1968 BP and 1963 BPC 1965 USP XVII
Therapeutic dose 5-10 mg/day 10-15 mg/day
Prophylactic dose 1-4 mg/day 2 mg/day
Riboflavin has been administered in large doses for the treatment
of various clinical conditions. A seven-year-old patient with primary
hyperoxaluria was administered 4 g riboflavin/day, for nine days. No
toxic effects were reported (Shepard et al., 1960) In another study,
310 patients with psoriasis were administered oral daily doses of
10-60 mg (about 0.1-1.0 mg/kg) riboflavin-5'-phosphate or 20-1000 mg
(about 0.3-15 mg/kg) of riboflavin for periods up to 42 months. No
adverse effects were reported (Welsh & Ede, 1957).
Comments
Riboflavin is an essential nutrient in man and occurs widely in
plant and animal tissues. Riboflavin-5'-phosphate (FMN) is also
naturally occurring. Upon ingestion, FMN appears to be rapidly
hydrolysed to riboflavin plus phosphate. Riboflavin and riboflavin-5'-
phosphate are in metabolic equilibrium after absorption. Some evidence
indicates the absorption of FMN and riboflavin in man may be limited
by a saturable mechanism in the gastrointestinal tract. In this
monograph, safety data on riboflavin were considered supportive of the
safety of FMN. Normal reproduction performance was reported in a three
generation study in which rats received approximately 100 times the
normal daily requirement. Toxic effects have not been reported in
humans fed high levels of riboflavin.
EVALUATION
Estimate of acceptable daily intake for man
(Group ADI for riboflavin and riboflavin-5'-phosphate) 0-0.5 mg/kg
(expressed as riboflavin).
REFERENCES
Christensen, S. (1969) Studies on riboflavin metabolism in the rat. I.
Urinary and faecal excretion after oral administration of
riboflavin-5'-phosphate, Acta Pharmacol. Toxicol., 27, 37-40
Jusko, W. J. & Levy, G. (1967) Absorption, metabolism, and excretion
of riboflavin-5'-phosphate in man, J. Pharm. Sci., 56, 58-62
Le Clerc (1974) Influence de la teneur du regime alimentaire en
thiamine, en riboflavine et en vitamine B6 sur la teneur des
tissues de la ratte en lactation et des jeunes en ces memes
vitamines, Ann. Nutr. Aliment, 23, 111-120
Litton Bionetics, Inc. (1977a) Mutagenicity evaluation of FDA 75-76
riboflavin USP-FCC. Final report, LBI Project No. 2672,
Kensington, MD, 44 pp.
Litton Bionetics, Inc. (1977b) Mutagenicity evaluation of FDA 75-77
riboflavin-5'-phosphate sodium FDD 00146-17-8. Final report, LBI
Project No. 2672, Kensington, MD, 44 pp.
Randall, L. O. (1951) Chronic toxicity of mono-diethanolamine salt of
riboflavin monophosphoric acid ester dihydrate. Unpublished
report of Hoffmann-La Roche, Inc. Submitted in pursuance of Fed.
Register 38:28581 (October 15, 1973) in connection with the
review of over-the-counter vitamin, mineral and hematinic drug
products
Randall, L. O. (1950) Toxicity of mono-diethanolamine salt of
riboflavin monophosphoric acid ester dihydrate. Unpublished
report of Hoffman-La Roche, Inc. Submitted in pursuance of Fed.
Register 38:28581 (October 15, 1973) in connection with the
review of over-the-counter vitamin, mineral and hematinic drug
products
Schumacher, M. F., Williams, M. A. & Lyman, R. L. (1965) Effect of
high intakes of thiamine, riboflavin and pyridoxine on
reproduction in rats and vitamin requirements of the offspring,
J. Nutr., 86, 343-349
Shepard, T. H. II et al. (1960) Primary hyperoxaluria. III.
Nutritional and Metabolic studies in a patient, Pediatrics,
25, 1008-1017
Stripp, B. (1965) Intestinal absorption of riboflavin by man, Acta.
Pharmacol. Toxicol., 22, 353-362
U.S. Food & Nutrition Board, National Research Council (1974)
Riboflavin, Recommended dietary allowances, 8th ed. rev.,
National Academy of Sciences, Washington, D.C., pp. 68-69
Unna, K. & Greslin, J. G. (1942) Studies on the toxicity and
pharmacology of riboflavin, J. Pharmacol. Exp. Ther., 76,
75-80
Welsh, A. L. & Ede, M. (1957) An appraisal of the therapeutic
effects of riboflavin in psoriasis, Arch Dermatol., 76,
595-600
West, D. W. & Owen, E. C. (1969) The urinary excretion of metabolites
of riboflavine by man, Br. J. Nutr., 23, 889-898
Yang, C.-S. & McCormick, D. B. (1967) Degradation and excretion of
riboflavin in rats, J. Nutr., 93, 445-453