WHO/Food Add./24.65 FAO Nutrition Meetings Report Series No. 38A SPECIFICATIONS FOR IDENTITY AND PURITY AND TOXICOLOGICAL EVALUATION OF SOME ANTIMICROBIALS AND ANTIOXIDANTS The content of this document is the result of the deliberations of the Joint FAO/WHO Expert Committee on Food Additives which met 8-17 December 1964a a Eighth Report of the Joint FAO/WHO Expert Committee on Food Additives, Wld Hlth Org. techn. Rep. Ser., 1965, 309; FAO Nutrition Meetings Report Series 1965, 38. POTASSIUM NITRITE CHEMICAL NAME Potassium nitrite EMPIRICAL FORMULA KNO2 MOLECULAR WEIGHT 85.11 DEFINITION Potassium nitrite after drying contains not less than 95% of KNO2. It conforms to the following specifications. DESCRIPTION Small, white or slightly yellow, deliquescent granules or rods. USE As a colour fixative in fish products and in pickling and curing meat, sometimes in combination with sodium nitrite and with potassium and sodium nitrates. IDENTIFICATION TESTS A. Solubility: Water: Very soluble Ethanol: Sparingly soluble B. In neutral, concentrated or moderately concentrated solutions of potassium nitrite, sodium bitartrate TS slowly produces a white, crystalline precipitate which is soluble in ammonia TS and in solutions of alkali hydroxide or carbonates. C. Potassium nitrite yields brownish red fumes when treated with diluted mineral acids or acetic acids. PURITY TESTS Arsenic: Not more than 3 mg/kg. Lead: Not more than 10 mg/kg. Heavy metals: Not more than 20 mg/kg. Dissolve 2 g in water, and dilute to 40 ml. For the control, add a volume of standard lead TS equivalent to 0.02 mg of lead to 10 ml of this solution and use the remaining 30 ml for the sample. Evaporate both solutions to dryness on the steam bath. To each residue add 5 ml of hydrochloric acid, and again evaporate to dryness. Dissolve the residues in 20-ml portions of water, adjust each to a pH between 3.0 and 4.0, dilute to 40 ml and add 10 ml of hydrogen sulfide TS to each. The colour in the sample solution, measured in a 50-ml Nessler tube, is not darker than the control solution, similarly measured. pH of a 5% solution: Dissolve 10 g in 200 ml of carbon dioxide- and ammonia-free water: the pH of this solution is between 6.0 and 9.0. ASSAY Dissolve 1.000 g, previously dried over silica gel for 4 hours, in water and make to 100.0 ml. Pipette 10 ml of this solution into a mixture of 50.0 ml of 0.1 N potassium permanganate, 100 ml of water and 5 ml of sulfuric acid, keeping the tip of the pipette well below the surface of the liquid. Warm the solution to 40°C, allow it to stand for 5 minutes and add 25.0 ml of 0.1 N oxalic acid. Heat the mixture to about 80°C and titrate with 0.1 N potassium permanganate. Each ml of 0.1 N potassium permanganate is equivalent to 4.256 mg of KNO2. SODIUM NITRITE CHEMICAL NAME Sodium nitrite EMPIRICAL FORMULA NaNO2 MOLECULAR WEIGHT 69.00 DEFINITION Sodium nitrite after drying contains not less than 97% of NaNO2. It conforms to the following specifications. DESCRIPTION White or slightly yellow, hygroscopic granules, powder, or opaque, fused masses of sticks. It is deliquescent in air. USE As a colour fixative in fish products and in pickling and curing meat, sometimes in combination with potassium nitrite, or potassium and sodium nitrates. IDENTIFICATION TESTS A. Solubility: Water: 1 g dissolves in 1.5 ml Ethanol: Sparingly soluble B. Sodium nitrite, after conversion to chloride or nitrate, yields with uranyl zinc acetate TS a golden-yellow precipitate, which forms after several minutes' agitation. C. Sodium nitrite yields brownish red fumes when treated with diluted mineral acids or acetic acid. PURITY TESTS Loss on drying: Not more than 0.25% by drying over silica gel for 4 hours. pH of a 5% solution: Dissolve 10g in 200 ml of carbon dioxide- and ammonia-free waters: the pH of this solution is between 6.0 and 9.0. Arsenic: Not more than 3 mg/kg. Lead: Not more than 10 mg/kg. Heavy metals: Not more than 20 mg/kg. Dissolve 2 g in water and dilute to 40 ml. For the control, add a volume of standard lead solution equivalent to 0.02 mg of lead to 10 ml of this solution and use the remaining 30 ml for the sample. Evaporate both solutions to dryness on the steam bath. To each residue add 5 ml of hydrochloric acid, and again evaporate to dryness. Dissolve the residues in 20-ml portions of water, adjust each to a pH between 3.0 and 4.0, dilute to 40 ml and add 10 ml of hydrogen sulfide TS to each. The colour in the sample solution, measured in a 50-ml Nessler tube, is not darker than the control solution, similarly measured. ASSAY Dissolve 1.000 g, previously dried over silica gel for 4 hours, in water and make to 100.0 ml. Pipette 10 ml of this solution into a mixture of 50.0 ml of 0.1 N potassium permanganate, 100 ml of water and 5 Ml of sulfuric acid, keeping the tip of the pipette well below the surface of the liquid. Warm the solution to 40°C, allow it to stand for 5 minutes, and add 25.0 ml of 0.1 N oxalic acid. Heat the mixture to about 80°C and titrate with 0.1 N potassium permanganate. Each ml of 0.1 N potassium permanganate is equivalent to 3.450 mg of NaNO2. Biological Data Biochemical Aspects Following absorption of nitrites, the most important biochemical reaction that occurs is the conversion of haemoglobin to methaemoglobin. There is some controversy concerning the molar ratios involved in this reaction. Making an extreme assumption, it may be stated that 1 g of sodium nitrite could convert as much as 1855 g of haemoglobin to methaemoglobin.1 The sub-acute hazard of nitrites rests on the amount of methaemoglobin formed and on the ability of the body to reconvert this methaemoglobin back to haemoglobin. Acute toxicity Animal Route LD50 Reference (mg/kg body-weight) Mouse oral 220 2 Mouse-female oral 175 3 Rat-female oral 85 3 Many cases have been reported of accidental poisoning resulting from the presence of sodium nitrite in food products. From this information it is possible to deduce that the oral lethal dose in man varies from 0.18 to 2.5 g, the lower figures being those for children and old people.4,5,6,7 Sodium nitrite has been used for therapeutic purposes as a vasodilating agent in dosages of 30-120 mg. Short-term studies Rat and cat. Rats were fed a sodium nitrite supplement for a period up to 168 days. One rat received a total of 167 mg of sodium nitrite in 121 days. This represents 93 ppm in the daily diet. No effects on growth or on the weights of important organs were noted. In a similar experiment with cats, one animal received a total of about 4100 mg of sodium nitrite during a period of 105 days. This represents approximately 390 ppm in the daily diet. No effects on the growth rate or on the weight of important organs were noted. No histopathological examination has been reported on any animal fed with nitrite. Long-term studies Rat. The continuous administration of sodium nitrite in the drinking-water at the rate of 100 mg/kg body-weight daily over the whole life span and in 3 successive generations (95 rats) resulted in spite of the high dosage (67% of the acute LD50) in only a slight inhibition of growth (10-20%) and in a shortening of the median life span from 740 to 640 days. Reproduction was normal. Neither the blood picture nor the organs showed any ill effects. The number of tumours observed in the test group (1 thymoma and 1 hepatoma) was not greater than in the control group. Cumulative toxic effects were not observed.9 Since nitrous acid reacts easily with secondary amines yielding dialkylnitrosamines which are highly toxic and produce cancer of the liver in rats, additional experiments have been made in which nitrite was administered together with diethylamine. Thirty hybrid rats were given sodium nitrite in the drinking-water at a daily dosage of 100 mg/kg body-weight and diethylamine in the food at the rate of 500 mg/kg body-weight daily. The same slight shortening of the median life span (to 625 days) was observed as in the experiments with nitrite alone, but there were no tumours.9 Comments on experimental studies reported Though in the long-term studies cited only a slight inhibition of growth occurred, the dose causing this effect appears to give the best approximation to the threshold dose level. Evaluation Level causing no significant toxicological effect in the rat From consideration of the long-term studies it can be concluded that this level will be somewhat below 100 mg/kg body-weight per day. Estimate of acceptable daily intakes for man mg/kg body-weight Unconditional acceptance 0-0.4 Conditional acceptance 0.4-0.8 Comment It is recommended that if possible sodium nitrite should be used mixed with common salt, the amount of nitrite in the salt being not more than 0.6%. Food for babies should not contain added nitrite. Further Work Considered Desirable 1. The estimation of nitrite in vegetables. 2. Chronic toxicity experiments in non-rodent species. References 1. Lehman, A. J. (1958) Quart. Bull. Ass. Food Drug. Off., 22, 136 2. Riemann, H. (1950) Acta pharmacol. (Kbh.), 6, 285 3. United States Food and Drug Administration (Unpublished data) 4. Schrader, G. & Gessner, O. (1943) Samml. Vergiftungsf., 13, 101 5. Greenberg, M., Birnkrant, W. B. & Schiftner, J. J. (1945) Amer. J. publ. Hlth., 35, 1217 6. Schmidt, H., Stick, W. & Kluge, F. (1949) Dtsch. med. Wschr., 74, 961 7. Naidu, S. R. & Venkratrao, P. (1945) Calcutta med. J., 42, 79 8. Tarr, H. L. A. & Carter, M. H. (1942) J. Fish. Res. Board, Can., 6, 63 9. Druckrey, H. (1963) Arzneimittel-Forsch., 13, 320
See Also: Toxicological Abbreviations Potassium nitrite (ICSC) POTASSIUM NITRITE (JECFA Evaluation)