` oo~9 ~'~~- `©F'PT .:). o - oo 33 

Submission 

For Fulfilment of Data Requirements 

for Biuret 

Under TSCA Section 4 

1. Chemical Name: Biuret 

2. CAS Number: 108-19-0 

3. Industry Sponsor: The Fertilizer Institute 

William Herz 

820 First Street NE, Union Ctr Plaza Ste 430 

Washington, DC 20002 

Tel : (202) 515-2706 

-r. 

tr. 

CJ 

t 

9. Date of Submission : July 2007 a - 

10. Comments : Biuret is not manufactured or processed for distribution i 

commerce as chemical substances per se and, relative to tl~e, ---1=--i 

fertilizer industry, has no commercial purpose separate from the = ~~ 

, substance mixture or article of which it may be a part (40 CFR 

710.4(d). 

Biuret relies on data presented for Urea (CAS 57-13-6; 

Screening Information Data Set (SIDS) of OECD High 

Production Volume Chemicals Programme (1994; see attached 

documents). Both Biuret and Urea are enzymatically hydrolyzed 

to ammonia and carbon dioxide in the environment and in 

mammalian systems. 

BIURET 

U 
p. 2 

BIURET 

CONTENTS 

1 IDENTITY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 

1 .1 Identification of the Substance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 

1 .2 Physico-Chemical properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .6 

2 GENERAL INFORMATION ON EXPOSURE. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 

2.1 Environmental Exposure and Fate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 

2 .1 .1 Sources of Environmental Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 

2.1 .2 Photodegradation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 

2.1 .3 Stability in Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 

2.1 .4 Transport between Environmental Compartments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 

2 .1 .5 Biodegradation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7 

3 HUMAN HEALTH HAZARDS. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 

3.1 Effects on Human Health . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 

3 .1 .1 Toxicokinetics, Metabolism and Distribution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .."""""""""""""" 9 3 .1 .2 Acute Toxicity 8 

3.1 .3 Repeated Dose Toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .~.~.~.~. . . . . . . . . . . . . . . . . . . . . . . . . . . . .'. 

3.1 .4 Mutagenicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .12 

3.1 .5 Toxicity for Reproduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .13 

4 HAZARDS TO THE ENVIRONMENT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .13 

4.1 Aquatic Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .13 

5 RECOMMENDATIONS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .14 

6 REFERENCES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .15 
p. 3 

BIURET 

Tables 

Table 1 Summary of physico-chemical properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 

Table 2 Summary of acute toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9 

Table 3 Summary of repeated dose toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .12 
Submission 

for Fulfilment of Data Requirements 

for Biuret 

Under TSCA Section 4 

1 IDENTITY 

1.1 Identification of the Substance 

CAS Number: 108-19-0 

Molecular Formula: C2H5N302 

Structural Formula: 

O 

NH2 

O 

NH---\ 

NHZ 

Molecular Weight: 103 

Synonyms: Imidodicarbonic diamide 

Biuret 

Allophanamide 

Allophanic acid amide 

Allophanimidic acid 

Carbamylurea 

Dicarbamylamine 

Isobiuret 

L-101 

Urea, (aminocarbonyl)-

Ureidoformamide 
p. 5 

BIURET 

1 .2 Physico-Chemical properties 

Table 1 Summary of physico-chemical properties 

Property Value Reference 

Physical state 

Melting point 188-190°C Aldrich, 1978, 2000 

Boiling point 193 °C at 1013.25 hPa Merck, 1976 

Relative density 1 .467 at -5 °C Merck, 1976 

Vapour pressure 1 .02E-5 mm Hg Epiwin, v3 .20 

Water solubility 8.777E+4 mg/L YALKOWSKY, and DANNENFELSER, 1992 

Partition coefficient noctanoUwater (log value) 

-1 .3988 Epiwin, v3 .20 

2 GENERAL INFORMATION ON EXPOSURE 

Biuret is not a commercial product for the members of The Fertilizer Institute (TFI); it is formed 

incidentally during the production of fertilizer grade urea at high temperatures (TFI, 2007). The 

existence of biuret was first identified by Wiedemann in 1848 after heating urea nitrate to 150° -

170°C. (Redemann et al . 1958). It has long been recognized as a component of urea fertilizers 

(Webster et al ., 1957) . The amount of biuret increases with increasing temperatures and some biuret 

is formed even when a solution of urea in water is heated above 50° C. Current levels of biuret in 

fertilizer grade urea are slightly less than 1 .0% wt with a recent range of 0.84 to 1 .04%. 

Some urea fertilizer manufactures reported the manufacture and import of biuret and urea reaction 

products associated with the manufacture and import of urea fertilizer in TSCA Inventory Update 

Rule (IUR) Form U submissions including the 2002 reports for activities in 2001- EPA Form 

Numbers 6511017 and 65011017. In 2003, TFI, petitioned EPA for a partial reporting exemption 

for fertilizers and certain fertilizer materials including biuret and urea reaction products from 

inventory update reporting requirements proposed for 2006 and beyond. In November 2003 and 

December 2004, EPA issued letters to TFI concluding that biuret and urea reaction products with 

formaldehyde are "impurities" as defined in 40 CFR. § 720.3 and therefore were not subject to 2006 

IUR reporting. The applicable provision of the now final regulation is § 710.50(c) which defines 

manufacturing a chemical substance "in a manner described in § 720.30(g) or (h)" of the regulations 

as an activity for which reporting is not required. 

A similar regulation, § 710.30(c), applies to 2002 IUR reporting. Some of the urea fertilizer 

manufacturers 2002 IUR reports included chemical substances for which it is now clear, based on 

EPA's November 2003 and December 2004 interpretation letters, that reporting was not required . 

These urea fertilizer manufacturers revised and resubmitted 2002 Form U Reports removing biuret 

(CAS No. 108-19-0) and urea reaction products with formaldehyde (CAS No. 68611-64-3). 

Other producers sell biuret as an FDA-approved non-protein nutrient supplement for ruminants. 

Biuret is considered an impurity of urea with no commercial purpose for reporting under TSCA 

Section 8 Inventory Update Reporting (IUR), is not manufactured or processed for distribution in 

commerce as a chemical substances per se and, relative to the fertilizer industry, has no commercial 

purpose separate from the substance mixture or article of which they may be a part (40 CFR § 

710.4(d). 
p. 6 

BIURET 

Based on the well established mechanisms for enzymatic hydrolytic degradation in the environment 

and in the mammalian gut of biuret to urea (and then to ammonia and carbon dioxide), data from 

urea (UNEP Publications, date not specified) are used to provide additional information for biuret. 

As stated in the UNEP document, "urea is an important endogenous product of mammalian 

metabolism. This may partly explain why it has not been rigorously studied with toxicological tests. 

Nevertheless, urea appears to cause little or no toxicity to most mammalian species (ruminants are 

an exception) and humans at reasonable dose levels . Further, The EPA has indicated that urea 

poses no risk concerns within the limits of the existing tolerance exemptions 

(http://www.epa. ov/oppsrrdl/reregistration/urea/), urea is approved for several human therapeutic 

uses (h=://216 .239.51 .104/search?q=cache_p1264Z8FFIJ :www.fda . gov/cder/ogd/rld/ 12154s24.pdf+FDA+category+C+urea&hl=en&ct=clnk&cd 

=1 & 1=us ) and is considered "Generally Recognized As Safe" (GRAS) as affirmed by the U. S. 

Food and Drug Administration for use in food (21 C.F .R. § 184.1923 Urea. Title 21 - Food and 

Drugs). 

2.1 Environmental Exposure and Fate 

2.1 .1 Sources of Environmental Exposure 

2.1 .2 Photodegradation 

The overall OH Rate Constant was = 3 .0000 E-12 cm3/molecule-sec with a half-life = 3 .565 days 

(Epiwin, v3 .20) . 

2.1 .3 Stability in Water 

Stability on water studies have not been conducted with biuret. Modeling indicates that the 

hydrolysis rate will be extremely slow (tl/2 > 1 year) (Epiwin, v3 .20) . 

2.1 .4 Transport between Environmental Compartments 

The EQC Level III Fugacity model (Epiwin, v3 .20) was used to evaluate the fate, transport and 

distribution of biuret between environmental matrices. Level III fugacity modelling, using loading 

rates of 1000 kg/h each for air, soil and water, shows the following percent distribution when it is 

released simultaneously to all three compartments: Air = 4.83E-6%; Soil = 60.9%; Water = 39%; 

Sediment = 0.713%. 

2.1 .5 Biodegradation 

Standard ready biodegradability studies have not been located for biuret. Biuret does not 

accumulate in soil and is mineralized within a single growing season . Biuret is not retained in soil 

(Orgata, year not specified) and is easily leached (Gadet et al ., 1959). Nishihare et al . (1965) found 

an enzyme, biuret-hydrolyzing enzyme, capable of decomposing biuret into ammonia, carbon 

dioxide and urea. Jensen (1966) found several widespread species of soil fungi and bacteria capable 

of degrading biuret . Jensen also reported that biuret at a concentration of 100 ppm was almost 

completely nitrified after two to three months. Several studies have concluded that the rate of biuret 

mineralization depends on soil composition, temperature, and biuret concentration (Mithyantha et 

al ., 1977) . Epiwin (v3.20) modeling suggests biuret is biodegradable, but may not meet the criteria 

for ready biodegradability . 
p. 7 

BIURET 

Urea also is ultimately biodegradable in inherent biodegradability test (OECD 302 B). In semicontinuous activated sludge (SCAS) urea is biodegraded on the average of 93-98% in 24-hour 

cycle. The main mode of degradation is enzymatic mineralization . In soil and water urea is expected 

to biodegrade fairly rapidly to ammonia and bicarbonate if temperature is not too low. Ammonia is 

a volatile gas in alkaline solutions. In natural waters, most ammonia appears in the form of 

ammonium, from which nitrogen is oxidized as the result of bacterial action, forming nitrite and 

nitrate. According to US-EPA PCGEMS-model hydrolysis of urea is expected to be extremely slow 

(T1/2 > 1 years) (UNEP, date not specified) . 

3 HUMAN HEALTH HAZARDS 

Data from urea (UNEP Publications, date not specified) are used to provide additional information 

for biuret. 

3.1 Effects on Human Health 

3.1 .1 Toxicokinetics, Metabolism and Distribution 

It has been well established that in the rumen, biuret and urea are hydrolyzed by microflora 

enzymes to ammonia and simple carbon compounds. These hydrolysis products can then be used to 

synthesize amino acids. The complete hydrolysis and deamination of protein involves numerous 

enzymes. Biuret is decomposed by biuretase to urea, and then to ammonia and carbon dioxide. 

Biuret is more slowly hydrolyzed than urea (Fonnesbeck, et al, 1975), thus accounting for the lower 

toxicity profile of biuret (reduces the toxic accumulation of ammonia in blood plasma). Biuret has 

not been shown to accumulate in rats, chickens or sheep. 

Urea is extremely soluble in water and oral doses are rapidly absorbed and distributed in humans. 

Urea is a normal human body constituent and is constantly being produced through amino acid and 

protein metabolism where urea is formed through a cyclic mechanism. 

3.1 .2 Acute Toxicity 

The acute toxicity of biuret has been studied extensively, although most studies have been 

conducted in livestock from the 1950's to the 1970's . The reliability of the studies with biuret are 

generally considered to be = (4) not assignable due to the inability to access the original studies. 

Fonnesbeck et al (1975) provides a review of many of these studies with biuret, and some are 

provided in the attached dossier for biuret and in Table 2. Biuret is of low acute oral toxicity. 

The acute toxicity by urea is well delineated by the oral route. Toxicity is low in mammals other 

than ruminants, especially cattle, and sheep, in which the rumen micro-organisms contain urease 

activity and metabolize urea to ammonia at a high rate . In mice and rats, urea is of low toxicity even 

by the subcutaneous and intravenous route (UNEP, date not specified) . Examples of specific 

toxicity values published for urea are provided for comparison in Table 2 . 

Despite the assignment of RL =4, the studies provide a weight of evidence regarding the lack of 

acute toxicity of biuret, as well as its lower toxicity when compared to urea. 
p. 8 

BIURET 

Table 2 Summary of acute toxicity 

Species Value (mg/kg bw) RL/Reference 

Biuret 

Sheep No effect at 4000 4/Swart and Lynch, 1972 

Sheep No effect at 680 4/Mieske et al., 1955 

Sheep No effect at 6900 4/Clark et al ., 1963, 1965; Fonnesbeck et al ., 1975 

Sheep Toxicity at 4700 4/Hatfield et al ., 1969; Fonnesbeck et al ., 1975 

Urea 

Sheep Oral LD50 = 510 Not specified/FDA, 1978 

Rat Oral LD50 =14300 

(M); = 15000 (F) 

Not specified/Sato et al ., 1978 

3.1 .3 Repeated Dose Toxicity 

Repeated dose toxicity studies with biuret have been conducted by the oral route. No wellconducted repeated dose toxicity studies on urea were located; oral chronic/carcinogenicity 

screening studies and repeated dermal exposure studies have been published for urea (LTNEP; no 

date specified) . These studies are summarized below and in Table 3. 

Male rats (6/group) were fed a basal diet supplemented with biuret at levels of 0, 0.2, 0.4, 0.6, 0.8 

and 1 .0% for 30 days (Berry et al ., 1956; RL=2). Animals were observed for signs of toxicity and 

body weights were recorded after 30 days . Based on the lack of toxicity noted in these animals a 

second experiment was initiated. Animals from groups 1 (control), 3 (0.4%) and 4 (0 .6%) were 

used in the second experiment. The control group was continued as previously described. Biuret 

was added to the basal diet at the rate of 5.0% and 10.0% and fed to groups 3 and 4, respectively . 

This feeding experiment was continued for 140 days to observe any long range effects of a high 

biuret diet . On the 35th and 98th days, one randomly selected animal from each of the three groups 

was killed and subjected to gross necropsy. There were no significant differences in growth rates 

among the groups . The appetites were normal, all animals appeared thrifty throughout the duration 

of the experiment, and no illnesses or deaths occurred. There were no histological findings. Since 

biuret is chemically related to urea it might be expected that the reaction of the rat to biuret would 

be similar to its reaction to urea. Observation of a higher water intake among the biuret-fed groups 

was noted. 

Groups of ten pullets were provided one of four dietary treatments (basal, urea, biuret, and soybean) 

(two groups of 10 within each treatment) (Berry et al ., 1956; RL=2). Group 1 (control) was fed a 

milo-soybean meal basal ration calculated to contain 14.5% crude protein. Groups 2 and 3 were fed 

the same basal diet except 1 % of the milo of each ration was replaced with 1 % feed urea or 1 % 

purified biuret, respectively, which raised the calculated crude protein equivalents to 17.3%. Group 

4 was fed the basal diet with enough milo replaced by soybean oil meal to raise the calculated crude 

protein value to 17 .3%. The duration of the study was not reported . Feed consumption was about 

5% lower for the biuret and urea supplemented groups than for the control and soybean oil meal 

supplemented groups . The body weight gain of the birds receiving the added soybean oil meal was 

significantly higher than that of the basal, urea and biuret groups . The inclusion of biuret or urea 

had no statistically significant effect on body weight gain. 

Three groups of 9 lambs were fed for 84 days with rations consisting of 42.5% cottonseed hulls, 

10% blackstrap molasses, and 40-45% ground milo (Berry et al., 1956; RL=2). The various crude 

protein supplements were 7.5% cottonseed meal (Group 1), 1 % cottonseed meal plus 1 % urea feedurea (Group 2), and 1 % cottonseed plus 1 % crude biuret (Group 3) . All three rations were 
calculated to supply equal levels of dry matter, total digestible nutrients and crude protein. In order 

to prevent differences related to palatability of the rations, the total intake of each was adjusted to 

that of the lowest consuming group. This was accomplished by hand feeding twice daily. Daily 

observations of water intake and urination were monitored. Average weight gain was calculated at 

study termination. One lamb from each test group was killed and a gross necropsy conducted. The 

biuret group was the slowest, the urea group intermediate and the control group the fastest in 

cleaning up the rations. Therefore, feed consumption of the biuret-fed group controlled the feed 

intake of the other groups . Daily observations indicated greater water intake and urination for the 

non-protein nitrogen (NPN)-fed groups, these conditions being more apparent for the biuret group 

than for the urea group. An examination of the internal organs showed no changes among the 

lambs from the three treatment groups . 

Five groups of steers were fed a high-roughage, low-concentrate wintering ration in dry lot for 84 

days (Berry et al ., 1956; RL=2). The basal ration was made up of 36.5% Atlas silage (hay basis), 

24% cottonseed hulls, 6% alfalfa leaf meal, 11% blackstrap molasses, and from 13% to 20% milo 

grain and 1 % minerals (bonemeal and salt). The crude protein supplements used were 7.5% 

cottonseed meal (Group 1), 1 % feed-urea (Group 2), 1 % crude biuret (Group 3), 1 .5% a mixture of 

crude biuret and feed-urea (Group 4), and 1 .25% of a urea-fatty acid complex (Group 5). The total 

intake of each group was adjusted to that of the lowest consuming group. This was accomplished 

by hand feeding. Each ration was calculated to supply equal amounts of dry matter, total digestible 

nutrients, and except for Group 4, protein equivalent. Group 4 was 1 .4% higher in crude protein 

equivalent. The urea-fatty acid complex was 75% urea and 25% cottonseed foots. Animals were 

observed for signs of toxicity and average weight gain was calculated at study termination. The 

average gains made by the control group and the urea group (Group 2) were not significantly 

different. However, when the urea was replaced with the crude biuret (Group 3), a significantly 

lower gain resulted. This group also consumed the least feed and therefore, restricted the intake of 

the other groups . When additional urea was included with the crude biuret (Group 4), the average 

gain was statistically significantly. Thus, simply adding urea overcame the adverse effects of 

biuret . Although Group 4 gained slightly more than the group fed urea alone (Group 2), the 

difference was not significant. The increase in daily gain may suggest that contrary to the results 

obtained with Group 3, the biuret nitrogen was utilized to some extent by the ruman bacteria. The 

group fed urea-fatty acid complex (Group 5) gained essentially the same as the urea fed group 

(Group 2) . Based on this single study it appears that urea fed as a urea-fatty acid complex is 

efficiently utilized . 

Biuret is an FDA-approved (Federal Register . Rules and Regulations. 21 CFR Part 573 . Federal 

Register 68(99) :27904 . 2003) NPN supplement for ruminants and a good deal of data exists for 

toxicity and nutritive dosages of urea/biuret combinations for these animals (Fonnesbeck et al ., 

1975). Many of these studies are summarized in the attached dossier; however, the RL of these 

studies are considered to be (3), as per se they do not provide useful hazard information for the 

purposes of this program, or (4), as sufficient details regarding methods and results were not 

available. Generally, plant proteins stimulate the most efficient animal growth and milk production . 

Animal response is satisfactory, however, when biuret and other NPN products provide part of the 

nitrogen requirements . When low quality forage diets deficient in nitrogen were fed to ruminants, 

response to feed grade biuret was very good. Supplementation with biuret stimulated good 

response from animals receiving medium and high energy diets. Biuret is an ideal supplement 

because ruminant animals may be fed more biuret than urea. The low solubility of biuret allows it to 

remain longer in the rumen. Biuret is hydrolyzed more slowly to ammonia and carbon dioxide in 

the rumen than urea and thereby eliminates toxic accumulation of ammonia in blood plasma. This 

is cited as the primary reason biuret is readily consumed by animals at high levels . Biuret is 

comparatively non-toxic even when supplying nitrogen at levels several times the requirements of 

the animal . This enlarges the margin of safety permitting feeding of higher concentrations of biuret 
p. 10 

BIURET 

in the diet than of other commercial NPN products. Biuret exhibits no undesirable palatability 

characteristics . Its low solubility and slow degradation rate reduces the hazard of self-feeding on a 

continuing or intermittent basis. 

No well-conducted repeated dose toxicity studies on urea were located (UNEP, no date specified) . 

Chronic toxicity and carcinogenicity screening studies in mice and rats fed with 4500, 9000 or 

45000 ppm in diet (up to about 6750 mg/kg body weight/day for mice and about 2250 mg/kg body 

weight/day for rats) did not uncover any treatment-related toxic syndromes in the various organs 

studied (Fleischman et al ., 1980). Neither was any weight depression noted at terminal necropsy for 

animals of either sex or species at any dose levels . Thus the NOAELs were about 6750 mg/kg body 

weight/day for mice and about 2250 mg/kg body weight/day for rats . 

Repeated dose toxicity studies with rats by skin application over 4 weeks and 25 weeks were 

conducted using urea ointment at 10%, 20% and 40% concentrations, and no consistent treatmentrelated toxic effects were found. The ointments were applied on a 20 cm2 area of the back skin but, 

unfortunately, the quantities of urea applied were not given. Therefore no value for NOAEL by 

dermal route can be determined. It can be concluded, however, that the repeated dose toxicity of 

urea by dermal route was low (UNEP, no date specified) . 
p. 11 

BIURET 

Table 3 Summary of repeated dose toxicity 

Study Design Result RL/Reference 

Biuret 

Rats (M); diet : 0, 0.2, 0 .4, 0.6, 0.8 and 1 .0% - 30 d NOAEL = 1 % 2/ Berry et al., 1956 

Pullets ; diet : 0, 1% urea, 1% biuret, or soybean oil - NOAEL = 1% 2/ Berry et al ., 1956 

exposure time not specified biuret 

Lambs; diet: 7 .5% cottonseed meal (Group 1), 1% NOAEL =1% biuret 2/ Berry et al ., 1956 

cottonseed meal plus 1% urea feed-urea (Group 2), 

1 % cottonseed plus 1 % crude biuret (Group 3) - 84 d 

Steers; diet : 7.5% cottonseed meal (Group 1), 1% NOAEL =1% biuret 2/ Berry et al., 1956 

feed-urea (Group 2), 1% crude biuret (Group 3), 1 .5% 

a mixture of crude biuret and feed-urea (Group 4), 

and 1 .25% of a urea-fatty acid complex (Group 5) - 

84 d 

Urea (*) 

Pullets; diet : 0, 1 % urea, 1 % biuret, or soybean oil - NOAEL = 1 % urea 2/ Berry et al ., 1956 

exposure time not specified 

Lambs; diet : 7.5% cottonseed meal (Group 1), 1% NOAEL=1% urea 2/ Berry et al., 1956 

cottonseed meal plus 1% urea feed-urea (Group 2), 

1 % cottonseed plus 1 % crude biuret (Group 3) - 84 d 

Steers ; diet: 7.5% cottonseed meal (Group 1), 1% NOAEL =1%urea 2/ Berry et al ., 1956 

feed-urea (Group 2), 1% crude biuret (Group 3), 1 .5% 

a mixture of crude biuret and feed-urea (Group 4), 

and 1 .25% of a urea-fatty acid complex (Group 5) - 

84 days 

Rat or mouse; diet : 4500, 9000 or 45000 ppm - 12 m NOAEL (mouse) ca. = Not 

6750 mg/kg bw/day ; (rat) specified/Fleischman 

ca. = 250 mg/kg bw/day et al ., 1980 

Rat; dermal : 10%, 20% and 40% - 4 or 25 w NOAEL not identified. Not specified/ Not 

Repeated dose toxicity specified; UNEP, date 

"low" not specifed 

(*) The Berry et al ., 1956 studies presented in Table 2 were not included in the SIDS documents for Urea. 

3.1 .4 Mutagenicity 

Mutagenicity studies were not located for biuret . Urea has been tested for mutagenicity in vitro and 

in vivo. The mechanism of genotoxicity of urea is probably non-specific (e.g . difference in osmotic 

pressure across the cell membrane) (UNEP, no date specified) . Similar effects are expected for 

biuret. 

In vivo Studies 

Urea was clastogenic for mammalian cells in vivo but the interpretation of this clastogenic effect is 

limited by the usage of a single extremely high dose level (Chaurasia and Sinha, 1987). 

In vitro Studies 

Urea has been negative in several appropriately conducted bacterial mutagenicity tests (Ishdate, et 

al, 1981 ; Shimizu et al ., 1985; Mortelmans et al ., 1986). Urea caused DNA single strand breaks in 

mammalian cells in mouse lymphoma cells (Garberg et al ., 1988) but not rat hepatocytes (Sina et 

al ., 1984). Urea was clastogenic in a Mouse Lymphoma TK Locus Assay; but only at 

concentrations much beyond the physiological range (about 50-100 higher concentrations than 
found in human blood) (Wangenhaim. and Bolcsfoldi, 1988). The positive genotoxic findings may 

depend on secondary cellular effects (e.g . difference in osmotic pressure across the cell membrane) 

at high concentration which might not take place under in vivo conditions. The maximally effective 

concentration or production of chromosome aberrations in Chinese hamster fibroblast cells was 266 

mmolar, a very high concentration suggesting a low clastogenic potential (Ishidate and Odashima, 

1977). Chromosomal aberrations were detected in 20% of metaphases of Chinese hamster fibroblast 

cells at urea concentration of 13 g/L (216 mmolar) indicate a low clastogenic potential (Ishdate et 

al . 1981) . The mechanism of genotoxicity is probably non-specific (e.g . difference in osmotic 

pressure across the cell membrane). 

3 .1 .5 Toxicity for Reproduction 

Standard reproductive toxicity studies have not been conducted with biuret or urea. Urea is an 

important endogenous product of mammalian metabolism . This may partly explain why it has not 

been rigorously studied with toxicological tests. Nevertheless, urea appears to cause little or no 

toxicity to most mammalian species (ruminants are an exception) and humans at reasonable dose 

levels (UNEP, no date specified) . 

Effects on Fertility 

The studies cited under repeated dose toxicity for urea did not indicate any toxic effects on the 

reproductive organs of mice and rats . No NOAEL can be given for the reproductive/developmental 

toxicity of urea because appropriate studies are lacking (UNEP, no date specified) . 

Developmental Toxicity 

Several non-standard studies with pregnant livestock (primarily cows) have indicated that biuret has 

no adverse effect on weight gains of cows or nursing calves, mild production or composition 

(Behrens, 1971 ; Iwata et al, 1956; Fonnesbeck et al ., 1975 ; Raleigh, 1969). 

No adequate teratogenicity/developmental toxicity studies of urea with mammals were located. 

According to one rat study, 50 g/kg body weight/day administered by gavage in two doses 12 hours 

apart for an average of 14 days did not cause outstanding (external) teratogenicity ; the mean birth 

weight of the newborn was lower but the litter size greater. Injection of urea into the air sack of 

eggs shows that urea is toxic to the development of chick embryo (UNEP, no date specified) . 

4 HAZARDS TO THE ENVIRONMENT 

4.1 Aquatic Effects 

Acute aquatic toxicity studies were not located for biuret ; data on urea is expected to be 

representative of biuret based on its degradation to urea, carbon dioxide and ammonia. Urea has 

generally low acute ecotoxicity to organisms. The degradation product of urea, ammonia, is known 

to be toxic to all vertebrates. In neutral and acidic conditions, however, ammonia exists in the form 

of ammonium. Instead of toxic effects, more emphasis should be put on the indirect influence of 

urea to the environment via eutrophication and the pollution risk of urea to groundwater, when urea 

is used as a fertilizer and a deicer agent in airports . Also the long-term influence of urea use e.g . soil 

acidification after the long-term fertilization and ammonia emissions to air can have influence to the 

whole ecosystem and should be considered (UNEP, no date specified) . 
p. 13 

BIURET 

Acute Toxicity Test Results 

Urea is not acutely toxic to fish, algae or aquatic invertebrates . Examples of published data include 

an LC50 for 96 hours > 9100 mg/L for Barilius barna (Dodriyal and Bahuguna, 1988; a toxicity 

threshold for 192 hours > 10000 mg/L for Scenedesmus quadricauda (Bringmann and Kuhn, 1978) 

and an EC50 for 24 hours > 10000 mg/L for Daphnia magna (Bringmann and Kuhn, 1982). 

5 RECOMMENDATIONS 

Additional testing is not required for biuret . The existing or modeled data for biuret in combination 

with published data on urea provides a complete profile for this substance. 
p. 14 

BIURET 

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