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Factors Affecting the Nitrate Content of Vegetable and Fruit Foods
Factors Affecting the Nitrate Content of Vegetable and Fruit Foods T. Bodiphala1 and D. P. Ormrod 2 Faculty of Agricultural Sciences University of B.C., Vancouver 8, B.C.
Abstract Th nitrate contents of a number of canned foods, baby s hozen foods and vegetables were determined. Sp~nach, food, d several fruits were found to generally have the hIghest ~et~ ~content. Nitrate was partially transferred to the liquid nl~ e of foods consisting of a solid immersed in liquid and po also transferred to blanching water. The rate of transfer ;fasni~rate from solid to liquid varied for diff~rent fo~ds and diff rent plant parts. Nitrate was not lost durmg coo~mg but "t :te reductase activity was quickly destroyed by cookmg. The Dl r of nitrate fertilizers on soil markedly increased nitrate content use . ,m t h at SOl. '1 of plants growmg
Resume Les teneurs en nitrate ont ete dosees dans des conserves aJimentaires des aliments de bebe, des aliments congeles et dans des Ieiumes. Les teneurs les plus elevees ont ete observees dans les epinards, les betteraves et plusieurs fruits. Le nitrate a passe partiellem~nt a la. partie. Iiquide des .ali~ents c?mposes de constituants sohdes en ImmerSIOn dans un hqUIde, et il a passe aussi a l'eau de bJanchiment. Le taux de transfert rlu nitrate du solide au \iquide a varie pour differents aliments et differentes parties de la plante. La cu;sson n'a pas affecte Ie nitrate mais a detroit rapidement I'activite de la reductase de nitrate. L'application de nitrate au sol a augmente considerablement la teneur en nitrate des plantes cultivees sur ce sol.
Introduction The presence and amounts of nitrates in foods has been noted and tabulated by Richardson (1907) and Wilson (1949). Nitrates are natural constituents of plant material since they are the main source of nitrogen required for growth. The roots of most plants absorb nitrogen from the soil in the form of nitrate. It must be reduced to ammonia before it can be incorporated into the organic nitrogenous compounds of the plant. Nitrate will accumulate if the rate of nitrate reduction is less than that of nitrate uptake. A most significant trend is the increased use of nitrogen-containing fertilizers which, in general, results in increased nitrate content of plant materials. Public concern has been aroused over the possible health hazard of high levels of nitrates found in some foods, and on the use of these foods in infant diets. The term "nitrate toxicity" as commonly used, is actually "nitrite toxicity" resulting from nitrite which is produced following the reduction of nitrate to nitrite by microbial action prior to ingestion or within the gastrointestinal tract. The nitrate content of a food is. thus only an index of the amount of nitrite which mIght be formed. Both nitrate and nitrite freely traverse the gastrointestinal wall into the blood stream. Nitrite, but not nitrate, oxidizes the ferrous iron of the r';d blood pigment hemoglobin and causes methemoglo?lllemia (Wright and Davison, 1964). Nitrates can ~terfere with normal iodine metabolism of the thyroid 1 pPresent address: Food and Drug Control Division. Ministry of ubllc Health. Bangkok. Thalland. 2 Present address: Department of Horticultural Science, University Of Guelph. Guelph, Ontario. 3 Pwaj)er presented at the Annual Conference of the C.l.F.T., lndsor. Ontario, June 8-10. 1970. J. lust. Can. Techno!. Ailment. Vo!. 4, No 1. 1971
gland and result in a reduction in the liver storage of vitamin A (Bloomfield and Welsch, 1961). Widely differing values for nitrate content of individual foods have been reported. Many factors probably contribute to this variability. The present study . was conducted to obtain more information on the nitrate content of vegetable and fruit foods and the effects of various factors on nitrate level.
Experimental Methods Samples of canned fruit and vegetables, baby foods, frozen vegetables and fresh vegetables were obtained from markets in the Vancouver area. Some veO"etables were also grown in the greenhouse. <=> Canned food was opened and drained to separate the liquid and solid portion. The liquid portio.n was used for determination of nitrate content dIrectly while the solid portion was blended to make a puree with water in the ratio of one unit solid to three units water. Strained baby food was used directly for ana1.)7sis. Frozen food was thawed, drained and separated into liquid and solid portions for analysis as for canned food. Fresh vegetables were cut into small pieces and blended with water in the ratio of one unit solid to four units water. Greenhouse grown plants were handled the same way. The sodium salicylate method of nitrate analysis as described by Muller and Widemann (1955) ,~as used. 'fhis method was found to be generally superIOr to the rapid method recommended by Woolle:y et al (1960) the phenol-disulfonic acid method descrIbed by Barnett (1954), and the nitrate-ion electrode method suggested by Mayers and Paul (~968).. . . The effect of time of blanchIng In bOIlIng water was determined. The variability in nitrate in different plant parts was also determi~ed. as w~ll as the effect of nitrate addition to the SOlI In whIch plants were growing. Nitrate reductase activity in plant tissue was measured by the method described by Evans and Nason (H)53) as modified by Hageman and Flesher (1960). The effect of blanching time on nitrate reductase activity was determined.
Results and Discussion Analysis for nitrate in foo.ds showed that ther<> is much variation between speCIes (Table 1). Among samples of baby food, one sample o~ beet was high.est in. nitrate. Fruit samples also had a hIgh concentratIon of nitrate. The beet sample was much higher than the 0.068 mg. nitrate nitrogen/gm. food maximum suggested for infants by Simon (1966). Most other samples also exceeded this level. The. high levels of nitrate found in fruit products are In contrast to relatively low values found by other investigators. Kamm et al (1965) reports a range of 0 to 0.007 mg/ in 22 samples of infant fruit foods. 6
Nitrate content of foods.
Table 1
Table 3
Nitrate nitrogen in mg.!gm. sample
Foods
1"
Baby food Bean Beet Carrot Corn (mixed with other ingredients Peas Peas and carrot Spinach (mixed with other ingredients) Squash Mixed vegetables Apple sauce Apricot Peach Plum Frozen vegetables
.041 .055 .070
.038 .024 .070
.113 .200 .046 .253 .185 .195 .123
.062 .138 .043 .231 .201 .185 .117
Peas Market vegetables
.075
.090
Bean Spinach Greenhouse vegetables
.138 .165
.070 .135
.212 .043 .027
.198 .053 .033
Beet Bean Spinach
The distribution of nitrate between solid and liquid portions of frozen food and canned food. Nitrate-nitrogen in mg.!gm. sample
Foods
1"
2
1
2
.102 .200 .053 .268 .465 .495
.078 .180 .043 .284 .435 .417
.086 .233 .090 .411 .460 .236
.054 .188 .086 .429 .432 .258
.140 .240
.186 .280
.040 .300
.055 .303
" Sample. Each sample was from a separate container obtained at a different time.
7
2 .157 .230 .110 .110 .075 .060 .068 .050 .035 .034 .045 .042
1
2
.243 .320 .230 .200 .165
.2 .2
.~
.26
.058 .081 .070 .070 .085
.o~ .0 .O~
.03 .03
transfer -of nitrate from the solid to the liquid portio (Table 3). There was a striking difference betwee spinach and bean. In spinach the nitrate analysi indicated a doubling of apparent total nitrate in th solid and liquid portion with iust one minute of blanch ing followed by a decline with longer heating time. T, bean, in contrast, the total nitrate content of soli~ and liquid did not vary from the initial total. In ad~ dition , the nitrate content of the blanching water .. 01. spinach became much higher than that remammg ID the solid portion while in bean the concentration was about equal in the two portions. Kilgore et al. (196~~} found that the nitrate in blanching water of turD! greens, collards and beets increased with increase i blanching time. This was not evident in the presen~ experiments. . II The distribution of nitrate in plant parts dlffere~ amonO' species (Table 4). There was little differencei betwe~n parts of the bean plant while beet roots were higher in nitrate than stems or leaves and young pe· tioles were highest in nitrate in spinach. Nitrogen fertilizer use may increase the nitrate content of tissue very markedly (Table 5). In the present ex·. periment nitrate was increased ten to twenty times by the addition of only 0.3 gm. nitrate-nitrogen per pot of four spinach plants. Spinach was found to have nitrate reductase ac, Table 4
Liquid
Solid
1" .202 .067 .063 .056 .064 .050 .135 .075 .078 .066 .070 .082
-
" Sample number. Each sample was of different plant material obtained on different days.
Nitrate was frequently higher in the liquid portion than in the solid portion of canned foods and in the liquid portion of thawed frozen foods (Table 2). Because nitrate salts are water soluble, much of the nitrate existing in the solid portion of canned food would be expected to diffuse to the liquid portion. A notable exception was bean which exposes less surface area to the liquid than does spinach. Among canned vegetable foods the nitrate content was highest in spinach. According to Wilson (1949), 1 gm. of po~s. sium nitrate ingested at a single meal may be tOXIC. Consuming 17 ounces of solid portion or 10.7 ounces of liquid from this spinach could therefore be harmful. Blanching and cooking resulted in the rapid
Canned Bean Beet (whole) Peas Spinach Apricot (halved) Peaches (sliced) Frozen Bean Spinach
0 1 3 5 10 20 0 1 3 5 10 20
Bean
" Sample number. Each sample was from a separate container obtained at a different time.
Table 2
Liquid
Solid
Foods
Spinach
-..
Nitrate-nitrogen in mg.!gm. sample
Blanching time min.
2 .125 .150 .083
.163 .350 .075
The distribution of nitrate between solid attI liquid portion during blanching of vegetables.
Nitrate in different plant parts. Nitrate-nitrogen in mg.!gm. sample
Plant Parts
Root Stalk or stem Petioles -old -young Leaves -old -young Fruits -old -young
Spinach
Beet
Bean
1"
2
.030 .038 .042 .037 .053 .033 .016
.019 .029 .030 .040 .039 .042 .014
1
2
.110 .080
.090 .084
.039
.041
1
2
.088 .060 .160 .073 .078
.138 .058 .120 .038 .065
.034
.044
" Laboratory duplicates. Can. Inst. Food Techno!. J. Vo!. 4, No. I, 1971
Table 5
_ Nitrate in different parts of spinach, with and without nitrogen fertilization. Nitrate-nitrogen mg/gm.
Parts of Plants
Stem Petiole Leaves Floral parts
No fertilizer
Average
sample With fertilizer
2
Ave.
1
.086 .204 .134
.140 .194 .120
.113 .199 .127
2.531 3.271 1.234
.042
.036
.039
.721
times increased by
Ave.
fertilization
2.303 3.129 1.428
2.467 3.200 1.331
21.08 16.08 10.55
.821
.771
19.99
2
o Laboratory duplicates. Fertilizer was made of 5 ml. of 1MCa(NOs). and 5 ml. of IMKN0 3 in a litre of water. Totally each pot of 4 plants received 0.3 gm. nitrate-nitrogen .
tivity capable of producing nitrite. Enzyme activity was reduced to twenty percent by blanching for two seconds and eight seconds blanching completely inactivated the enzyme. Therefore, blanching or cooking spinach for a short time will stop nitrate reductase activity.
J. lnst. Can. Techno!. AIlment. Vo!. 4, No 1, 1971
References Bloomfield. R A.. and Welsch, C. W. 1961. Effect of dietary nitrate on thyroid function. Science 134:1690. Barnett, A. F. G. 1954. Determination of nitrate in silage, in Silage Fermentation. Butterworths Scientific Publications, London. Evans, H. G., and Nason, A. 1953. Pyridine nucleotide-nitrate reductase from extracts of higher plants. Plant Physio!. 28:233. Hageman, R. H., and Flesher, D. 1960. Nitrate reductase activity in corn seedlings as affected by Ilght and nitrate content of nutrient media. Plant Physio!. 35:700. Kamm, L., McKeown, C. G., and Smith. D. M. 1965. New colorimetric method for the determination of the nitrate and nitrite content of baby foods. J. Assoc. Offic. Agric. Chem. 48:892. Kilgore, L., Stasch. A. R, and Barrentine, B. F. 1963. Nitrate content of beets, collards, turnip greens. J. Amer. Dietetic Assn. 43 :39. Mayer~, R. J. K., and Paul, E. A. 1968. Nitrate ion electrode method for soil nitrate-nitrogen determination. Can. J. Soil Sci. 48:369. Muller, R, and Widemann, O. 1955. Die bestimmung des nitrat-ions in wasser. Vour Wasser. 22:247. Richardson, W. D. 1907. The occurrence of nitrate in vegetable foods. in cured meats and elsewhere. J. Amer. Chem. Soc. 29:1757. Simon, C. 1966. Nitrite poisoning from spinach. Lancet 1 :872. Wilson, J. K. 1949. Nitrate in foods and its relation to health. Agron. J. 41:20. Wright, M. J., and Davison, K. L. 1964. Nitrate accumulation in crops and nitrate poisoninF; in animals. Adv. in Agron. 16:197. Woolley, J. T., Hicks, G. P., and Hageman, R. H. 1960. Rapid determination of nitrate and nitrite in plant material. J. Agri. and Food Chem. 8 :481. Received June 23, 1970.
8
Abstract Th nitrate contents of a number of canned foods, baby s hozen foods and vegetables were determined. Sp~nach, food, d several fruits were found to generally have the hIghest ~et~ ~content. Nitrate was partially transferred to the liquid nl~ e of foods consisting of a solid immersed in liquid and po also transferred to blanching water. The rate of transfer ;fasni~rate from solid to liquid varied for diff~rent fo~ds and diff rent plant parts. Nitrate was not lost durmg coo~mg but "t :te reductase activity was quickly destroyed by cookmg. The Dl r of nitrate fertilizers on soil markedly increased nitrate content use . ,m t h at SOl. '1 of plants growmg
Resume Les teneurs en nitrate ont ete dosees dans des conserves aJimentaires des aliments de bebe, des aliments congeles et dans des Ieiumes. Les teneurs les plus elevees ont ete observees dans les epinards, les betteraves et plusieurs fruits. Le nitrate a passe partiellem~nt a la. partie. Iiquide des .ali~ents c?mposes de constituants sohdes en ImmerSIOn dans un hqUIde, et il a passe aussi a l'eau de bJanchiment. Le taux de transfert rlu nitrate du solide au \iquide a varie pour differents aliments et differentes parties de la plante. La cu;sson n'a pas affecte Ie nitrate mais a detroit rapidement I'activite de la reductase de nitrate. L'application de nitrate au sol a augmente considerablement la teneur en nitrate des plantes cultivees sur ce sol.
Introduction The presence and amounts of nitrates in foods has been noted and tabulated by Richardson (1907) and Wilson (1949). Nitrates are natural constituents of plant material since they are the main source of nitrogen required for growth. The roots of most plants absorb nitrogen from the soil in the form of nitrate. It must be reduced to ammonia before it can be incorporated into the organic nitrogenous compounds of the plant. Nitrate will accumulate if the rate of nitrate reduction is less than that of nitrate uptake. A most significant trend is the increased use of nitrogen-containing fertilizers which, in general, results in increased nitrate content of plant materials. Public concern has been aroused over the possible health hazard of high levels of nitrates found in some foods, and on the use of these foods in infant diets. The term "nitrate toxicity" as commonly used, is actually "nitrite toxicity" resulting from nitrite which is produced following the reduction of nitrate to nitrite by microbial action prior to ingestion or within the gastrointestinal tract. The nitrate content of a food is. thus only an index of the amount of nitrite which mIght be formed. Both nitrate and nitrite freely traverse the gastrointestinal wall into the blood stream. Nitrite, but not nitrate, oxidizes the ferrous iron of the r';d blood pigment hemoglobin and causes methemoglo?lllemia (Wright and Davison, 1964). Nitrates can ~terfere with normal iodine metabolism of the thyroid 1 pPresent address: Food and Drug Control Division. Ministry of ubllc Health. Bangkok. Thalland. 2 Present address: Department of Horticultural Science, University Of Guelph. Guelph, Ontario. 3 Pwaj)er presented at the Annual Conference of the C.l.F.T., lndsor. Ontario, June 8-10. 1970. J. lust. Can. Techno!. Ailment. Vo!. 4, No 1. 1971
gland and result in a reduction in the liver storage of vitamin A (Bloomfield and Welsch, 1961). Widely differing values for nitrate content of individual foods have been reported. Many factors probably contribute to this variability. The present study . was conducted to obtain more information on the nitrate content of vegetable and fruit foods and the effects of various factors on nitrate level.
Experimental Methods Samples of canned fruit and vegetables, baby foods, frozen vegetables and fresh vegetables were obtained from markets in the Vancouver area. Some veO"etables were also grown in the greenhouse. <=> Canned food was opened and drained to separate the liquid and solid portion. The liquid portio.n was used for determination of nitrate content dIrectly while the solid portion was blended to make a puree with water in the ratio of one unit solid to three units water. Strained baby food was used directly for ana1.)7sis. Frozen food was thawed, drained and separated into liquid and solid portions for analysis as for canned food. Fresh vegetables were cut into small pieces and blended with water in the ratio of one unit solid to four units water. Greenhouse grown plants were handled the same way. The sodium salicylate method of nitrate analysis as described by Muller and Widemann (1955) ,~as used. 'fhis method was found to be generally superIOr to the rapid method recommended by Woolle:y et al (1960) the phenol-disulfonic acid method descrIbed by Barnett (1954), and the nitrate-ion electrode method suggested by Mayers and Paul (~968).. . . The effect of time of blanchIng In bOIlIng water was determined. The variability in nitrate in different plant parts was also determi~ed. as w~ll as the effect of nitrate addition to the SOlI In whIch plants were growing. Nitrate reductase activity in plant tissue was measured by the method described by Evans and Nason (H)53) as modified by Hageman and Flesher (1960). The effect of blanching time on nitrate reductase activity was determined.
Results and Discussion Analysis for nitrate in foo.ds showed that ther<> is much variation between speCIes (Table 1). Among samples of baby food, one sample o~ beet was high.est in. nitrate. Fruit samples also had a hIgh concentratIon of nitrate. The beet sample was much higher than the 0.068 mg. nitrate nitrogen/gm. food maximum suggested for infants by Simon (1966). Most other samples also exceeded this level. The. high levels of nitrate found in fruit products are In contrast to relatively low values found by other investigators. Kamm et al (1965) reports a range of 0 to 0.007 mg/ in 22 samples of infant fruit foods. 6
Nitrate content of foods.
Table 1
Table 3
Nitrate nitrogen in mg.!gm. sample
Foods
1"
Baby food Bean Beet Carrot Corn (mixed with other ingredients Peas Peas and carrot Spinach (mixed with other ingredients) Squash Mixed vegetables Apple sauce Apricot Peach Plum Frozen vegetables
.041 .055 .070
.038 .024 .070
.113 .200 .046 .253 .185 .195 .123
.062 .138 .043 .231 .201 .185 .117
Peas Market vegetables
.075
.090
Bean Spinach Greenhouse vegetables
.138 .165
.070 .135
.212 .043 .027
.198 .053 .033
Beet Bean Spinach
The distribution of nitrate between solid and liquid portions of frozen food and canned food. Nitrate-nitrogen in mg.!gm. sample
Foods
1"
2
1
2
.102 .200 .053 .268 .465 .495
.078 .180 .043 .284 .435 .417
.086 .233 .090 .411 .460 .236
.054 .188 .086 .429 .432 .258
.140 .240
.186 .280
.040 .300
.055 .303
" Sample. Each sample was from a separate container obtained at a different time.
7
2 .157 .230 .110 .110 .075 .060 .068 .050 .035 .034 .045 .042
1
2
.243 .320 .230 .200 .165
.2 .2
.~
.26
.058 .081 .070 .070 .085
.o~ .0 .O~
.03 .03
transfer -of nitrate from the solid to the liquid portio (Table 3). There was a striking difference betwee spinach and bean. In spinach the nitrate analysi indicated a doubling of apparent total nitrate in th solid and liquid portion with iust one minute of blanch ing followed by a decline with longer heating time. T, bean, in contrast, the total nitrate content of soli~ and liquid did not vary from the initial total. In ad~ dition , the nitrate content of the blanching water .. 01. spinach became much higher than that remammg ID the solid portion while in bean the concentration was about equal in the two portions. Kilgore et al. (196~~} found that the nitrate in blanching water of turD! greens, collards and beets increased with increase i blanching time. This was not evident in the presen~ experiments. . II The distribution of nitrate in plant parts dlffere~ amonO' species (Table 4). There was little differencei betwe~n parts of the bean plant while beet roots were higher in nitrate than stems or leaves and young pe· tioles were highest in nitrate in spinach. Nitrogen fertilizer use may increase the nitrate content of tissue very markedly (Table 5). In the present ex·. periment nitrate was increased ten to twenty times by the addition of only 0.3 gm. nitrate-nitrogen per pot of four spinach plants. Spinach was found to have nitrate reductase ac, Table 4
Liquid
Solid
1" .202 .067 .063 .056 .064 .050 .135 .075 .078 .066 .070 .082
-
" Sample number. Each sample was of different plant material obtained on different days.
Nitrate was frequently higher in the liquid portion than in the solid portion of canned foods and in the liquid portion of thawed frozen foods (Table 2). Because nitrate salts are water soluble, much of the nitrate existing in the solid portion of canned food would be expected to diffuse to the liquid portion. A notable exception was bean which exposes less surface area to the liquid than does spinach. Among canned vegetable foods the nitrate content was highest in spinach. According to Wilson (1949), 1 gm. of po~s. sium nitrate ingested at a single meal may be tOXIC. Consuming 17 ounces of solid portion or 10.7 ounces of liquid from this spinach could therefore be harmful. Blanching and cooking resulted in the rapid
Canned Bean Beet (whole) Peas Spinach Apricot (halved) Peaches (sliced) Frozen Bean Spinach
0 1 3 5 10 20 0 1 3 5 10 20
Bean
" Sample number. Each sample was from a separate container obtained at a different time.
Table 2
Liquid
Solid
Foods
Spinach
-..
Nitrate-nitrogen in mg.!gm. sample
Blanching time min.
2 .125 .150 .083
.163 .350 .075
The distribution of nitrate between solid attI liquid portion during blanching of vegetables.
Nitrate in different plant parts. Nitrate-nitrogen in mg.!gm. sample
Plant Parts
Root Stalk or stem Petioles -old -young Leaves -old -young Fruits -old -young
Spinach
Beet
Bean
1"
2
.030 .038 .042 .037 .053 .033 .016
.019 .029 .030 .040 .039 .042 .014
1
2
.110 .080
.090 .084
.039
.041
1
2
.088 .060 .160 .073 .078
.138 .058 .120 .038 .065
.034
.044
" Laboratory duplicates. Can. Inst. Food Techno!. J. Vo!. 4, No. I, 1971
Table 5
_ Nitrate in different parts of spinach, with and without nitrogen fertilization. Nitrate-nitrogen mg/gm.
Parts of Plants
Stem Petiole Leaves Floral parts
No fertilizer
Average
sample With fertilizer
2
Ave.
1
.086 .204 .134
.140 .194 .120
.113 .199 .127
2.531 3.271 1.234
.042
.036
.039
.721
times increased by
Ave.
fertilization
2.303 3.129 1.428
2.467 3.200 1.331
21.08 16.08 10.55
.821
.771
19.99
2
o Laboratory duplicates. Fertilizer was made of 5 ml. of 1MCa(NOs). and 5 ml. of IMKN0 3 in a litre of water. Totally each pot of 4 plants received 0.3 gm. nitrate-nitrogen .
tivity capable of producing nitrite. Enzyme activity was reduced to twenty percent by blanching for two seconds and eight seconds blanching completely inactivated the enzyme. Therefore, blanching or cooking spinach for a short time will stop nitrate reductase activity.
J. lnst. Can. Techno!. AIlment. Vo!. 4, No 1, 1971
References Bloomfield. R A.. and Welsch, C. W. 1961. Effect of dietary nitrate on thyroid function. Science 134:1690. Barnett, A. F. G. 1954. Determination of nitrate in silage, in Silage Fermentation. Butterworths Scientific Publications, London. Evans, H. G., and Nason, A. 1953. Pyridine nucleotide-nitrate reductase from extracts of higher plants. Plant Physio!. 28:233. Hageman, R. H., and Flesher, D. 1960. Nitrate reductase activity in corn seedlings as affected by Ilght and nitrate content of nutrient media. Plant Physio!. 35:700. Kamm, L., McKeown, C. G., and Smith. D. M. 1965. New colorimetric method for the determination of the nitrate and nitrite content of baby foods. J. Assoc. Offic. Agric. Chem. 48:892. Kilgore, L., Stasch. A. R, and Barrentine, B. F. 1963. Nitrate content of beets, collards, turnip greens. J. Amer. Dietetic Assn. 43 :39. Mayer~, R. J. K., and Paul, E. A. 1968. Nitrate ion electrode method for soil nitrate-nitrogen determination. Can. J. Soil Sci. 48:369. Muller, R, and Widemann, O. 1955. Die bestimmung des nitrat-ions in wasser. Vour Wasser. 22:247. Richardson, W. D. 1907. The occurrence of nitrate in vegetable foods. in cured meats and elsewhere. J. Amer. Chem. Soc. 29:1757. Simon, C. 1966. Nitrite poisoning from spinach. Lancet 1 :872. Wilson, J. K. 1949. Nitrate in foods and its relation to health. Agron. J. 41:20. Wright, M. J., and Davison, K. L. 1964. Nitrate accumulation in crops and nitrate poisoninF; in animals. Adv. in Agron. 16:197. Woolley, J. T., Hicks, G. P., and Hageman, R. H. 1960. Rapid determination of nitrate and nitrite in plant material. J. Agri. and Food Chem. 8 :481. Received June 23, 1970.
8