Table I. Comparison of Metallic Sodium Analyses in Three Mineral Oil Slurries" of Sodium Hydride Total Na Present as Metal, % b Butyl Bromide Hz Evolved on
Titration Hydrolysis No. of Av. X O . of Av. Av. detns. dev. Av. detns. dev. 25.0 1 .. 24.6 1 ,. 19.0 4 0 . 1 22.8 1 .. 29.9 4 0 . 1 31.4 4 1.2 Solids constitute about 10% of each slurry. Based on HC1 titration for total sodium also.
magnetic stirring bar. The system is evacuated and about 15 ml. of ammonia are condensed into the tube. The contents are warmed to about the boiling point of the solution, the nitrogen sweep is turned on, and the buret is attached to the top of the adapter. The solution is then titrated to a n end point characterized by a change from dark blue to colorless. Titration of Metallic Sodium in Sodium Hydride Slurries. The slurry t o be sampled is shaken thoroughly
and about 5 ml. are immediately transferred to a centrifuge tube and vl-eighed. Hexane ( 2 5 ml.) is then added t o the centrifuge tube and the contents are centrifuged. As much clear liquid as possible is carefully removed, and 1 ml. of hexane is added. A small stirring bar is then placed in the tube and the tube is connected to the adapter. The contents are cooled with liquid nitrogen, the system is evacuated, and about 15 ml. of ammonia are condensed into the tube. The titration is then carried out as described above. RESULTS AND DISCUSSION
The titration of sodium in ammonia in the absence of mineral oil was readily accomplished. Butyl bromide in hexane, made up to be equivalent to 2.00 mg. of sodium per ml. of solution, was found by actual titration to be equivalent to 2.02 mg. of sodium per ml. with an average deviation of 0.04 mg. per ml. based on nine analyses. The end point of the titration was sharp, one drop of titrant changing the solution from blue to colorless. Reproducible results were obtained on sodium hydride slurries when the
sample was first centrifuged and the major portion of the mineral oil removed. This prevented the solids from becoming coated by a viscous, semisolid mineral oil from which metallic sodium is only s l o ~ l yextracted. Comparative analytical data obtained on three sodium hydride slurries are summarized in Table I. Vhile more data, covering a wider concentration range, would be desirable, it is evident that this method has a broad range of applicability. The poor agreement between methods is not unexpected because an error of 1% in the hydrogen evolved on hydrolysis results in about a 2% error in metallic sodium. The presence of sodium hydroxide could also explain the discrepancies. LITERATURE CITED
(1) Budrieth, L. F., ,Kleinberg, J., "Non-
aqueous Solvents, p. 109, Wiley, New York, 1953. (2) Ethyl Corp., Chemical Research Laboratory, private communication. RECEIVED for review August 5, 1957. Accepted July 3, 1958.
Inorganic Nitrate, Nitrite, or Nitrate-Nitrite Rapid Colorimetric Determination of Microgram Quantities in Aqueous Solution F. L.
FISHER, E. R. IBERT, and H.
F. BECKMAN
Departments of Agronomy, Oceanography, and State Chemist, Agricultural and Mechanical College of Texas, College Station, rex.
,Rapid colorimetric methods have been developed for the quantitative determination of nitrate, nitrite, and both nitrate and nitrite ions using 2% brucine hydrochloride and either a 17 or 50% sulfuric acid solution. The methods are adapted for analysis of water, soil extracts, and other aqueous solutions where nitrogen concentration i s low. The brucine method for nitrates is more r a p i d and easier to use than the phenoldisulfonic acid method. Results obtained b y the two methods show excellent agreement. In analyzing soils and irrigation waters (brucine method) for nitrate and nitrite content, no interfering ions have been encountered.
T
accurate determination of nitrates or nitrites has long been a difficult but important analytical problem. The total amount of these two anions can be determined by phenolHE
1972
e
ANALYTICAL CHEMISTRY
disulfonic acid (?'), but the method is time-consuming in spite of many proposed niodifxations of the original procedure. The modification proposed by Stanford (9) is rapid when dealing with samples high (10 or more p.p.m.) in nitrate or nitrite content. -4rapid colorimetric method for the determination of nitrates and nitrites, proposed by Winkler (10) and cited by Yoe (la),Snell and Snell (a), and No11 (6),employs brucine instead of phenoldisulfonic acid. Yoe ( l a ) states that this method can be used to detect nitrate ions only, even in the presence of nitrites, and his method was further modified by Wolf (11). R o r k by Peech (6) and Joham (4) indicated that the brucine method could be used successfully on either soil or plant extracts to detect the total nitratenitrite content. Peech (6),in his quick test procedures, found close agreement between results by the brucine and phenoldisulfonic acid methods. H o w
ever, attempts to use these methods as routine laboratory procedure failed to give consistent results. This inconsistency is in agreement with Greenberg et al. ( 2 ) who say that published procedures using brucine give extremely erratic results. They have proposed a modified brucine procedure for determining nitrates only; nitrites ' must be removed as an interferin, ion. The method as described does afford one possible improvement of the method proposed in this paper-that is, pretreatment for removal of chlorine (Clz). To date the authors have not encountered chlorine as a problem, and chlorides do not interfere. REAGENTS AND STANDARD SOLUTIONS
Concentrated sulfuric acid (free of nitrate and nitrite). Brucine hydrochloride. Dissolve 2 grams of brucine hydrochloride in 100 ml. of distilled water or 0.05N hydrochloric acid.
I
Standard nitrate solution. Dissolve 0.6071 gram of dried sodium nitrate in 1 liter of distilled water for a stock solution containing 100 p.p.m. of nitrogen. Standard nitrite solution. Dissolve 0.4929 gram of dried sodium nitrite in 1 liter of distilled water for a stock solution containing 100 1xp.m. of nitrogen.
Table I. Effect of Brucine Hydrochloride Concentration on Color intensity 1 1\11. of Reagent Nitrogen Added as 0.5% 1.0% 2.0% 4.0% ' NO,, y Added nitrogen recovered, % 100 5 66 92 100 100 15 74 100
STANDARD CURVES
Nitrate Standard Curve. Place 30 ml. of the 100-p.p.m. nitrogen standard (NaNOJ in a 1-liter volumetric flask. Dilute aliquots of 1, 2, 3, 4, 5 , and 6 ml. of this solution to 15 ml., and proceed as indicated in Procedure
2
A.
Place 30 ml. of the 100-p.p.m. nitrogen standard (NaX0.J in a 1-liter volumetric flask. Dilute aliquots of 1, 2, 3. 4, 5 , and 6 ml. of this solution t o 15 ml., and proceed as indicated in Procedure B.
51: 1
Nitrite Standard Curve.
4: 1
3:l
111
211
EXPERIMENTAL RESULTS
The effect of brucine hydrochloride on color intensity with 5 and 15 y
1 5 micrograms of n i t r o g e n as nitrite
A
7 . 5 m i c r o g r a m s of n i t r o g e n as nitrate plus
1j m i c r o g r a m s of nitrogen os
llltrate
RATIO O F H20 TO CONC. H2SO4
Figure 1. Effect of water-acid ratio on nitrate-nitrite color development when brucine hydrochloride i s added before sulfuric acid
PROCEDURES
Procedure A, for detection of total nitrogen present as the nitrite or nitrate anions. Place a n aliquot of the sample to be analyzed containing not more than 15 y of nitrogen in a 50ml. Erlenmeyer flask and bring the total volume to 15 ml. with distilled n-ater. Add 1 ml. of 294, brucine reagent and mix; add 5 ml. of concentrated sulfuric acid with continuous mixing. Allow the mixture to stand at least 3 minutes (this develops nitrite color), then add 10 ml. of concentrated sulfuric acid, mixing continuously (this develops nitrate color). Immediately place in a dark chamber (3) and allow to cool to 30" C. before reading with a colorimeter. Interpolate nitrogen concentrations from a nitrate standard curve. Procedure B, for detection of nitrite ions only (even if nitrate ions are present). Place a n aliquot of the sample containing not more than 15 y of nitrogen as nitrite in a 50-ml. Erlenmeyer flask and make the total volume 25 ml. with distilled water. Add 1 ml. of 2y0 brucine reagent and mix. Then add 5 ml. of concentrated sulfuric acid with continuous mixing. Immediately place in a dark chamber, allow to cool to 30" C., and interpolate nitrogen concentration from a nitrite standard curve. Procedure C, for nitrate ions only (nitrites must be absent). Place an aliquot of the sample containing not more than 15 y of nitrogen as nitrate in a 50-nil. Erlenmeyer flask and dilute to 15 ml. with distilled water. Add 1 ml. of 2% brucine reaqent and mix, then add 15 nil. of concentr,zted sulfuric acid with continuous mixing. Place in a dark chamber and allow to cool to 30" C. Interpolate nitrogen concentration from a nitrate standard curve.
t:
I
rr;
E 2o Io
T---il
loo
\ I
-
s
-
D E G R E E S CENTIGRP.DE
c
Figure 3. Effect of temperature on nitrate-nitrite color development when brucine hydrochloride i s added after sulfuric acid
t: z
a-
c 40-
u
m
15 micrograms as n i t r a t e
0 0
11 55 m m ii cc rr oo gg rr aa m m ss o o ff n n ii tt rr oo gg ee nn as nltrlte
of n i t r o g e n
V
c:
k 20A
7 . 5 micr as n 7 . 5 micr as n
I
5 1
4 1
o g r a m s of n i t r o g e n itrate plus o g r a m s of n i t r o g e n ltrlte I
I
I
3 1
2 1
1 1
R A T I C O F H 2 O TO CONC. H2S04
Figure 2. Effect of water-acid ratio on nitrate-nitrite color development when brucine hydrochloride i s added after sulfuric acid
of nitrogen as nitrate present is shown in Table I. One milliliter of 1% brucine hydrochloride will furnish enough brucine to react with 5 y of nitrogen but not 15 y. One milliliter of 2% reagent will give maximum color intensity even xhen 15 y of nitrogen are present. Samples containing nitrate, nitrite, and nitrate-nitrite were prepared and color was developed using various water-sulfuric acid ratios. These data (Figure 1) indicate that a water-acid ratio of 5 to 1 (volumc ratio) will give maximum color development from nitrite ions, vhile at this water-acid ratio no color will develop in solutions containing only nitrate ions. Color can be developed from either or both anions by increasing the acid concentration to 5oYob. Increasing the acid concentration to 85% results in the saine color
development. The data in Figure 1 were obtained by folluning Procedure A. For data in Figures 2 and 3, the water-acid ratio x i s established, then the brucine reagent added. The addition of acid before the brucine reagent reduces the sensitivity of the method, which accounts for the difference in transmittance between Figures 1 and 2. The data shown in Figure 3 illustrate the effect of temperature on color development when 50% acid is used. To control temperature, the sulfuric acid was added and the desired temperatures were established with a water bath. Then the brucine reagent was added and all samples were put on a steam bath for 10 minutes. The samples were nest cooled to room temperature, and the per cent transmittance was read a t 410 mF. This wave length was found to be the point of maximum absorption (Beckman DK-2), which is in agreement \vith No11 (6). DISCUSSION
Data obtained during these investigations indicate that color intensity is dependent upon the amount of nitrate or nitrite present, the sulfuric acid concentration, the order of addition of sulfuric acid and brucine reagent, the masiinuin temperature the solution attains, arid the amount of brucine. The use of chloroform as a carrier VOL. 30, NO. 12, DECEMBER 1958
1973
Table II. Nitrogen as Nitrate Found in Water Extracts of Soils
Soil No.
pH of Soil
54-71 54-76 55-42 55-60 54-3 54-12
5.6 5.4 6.5 6.5 7.8 7.9
Sitrogen Found, P.P.M. Phenoldisulfonic Brucine acid method method 27 28 17 8 8 35
27 30 18 9 8 35
Table 111. Determination of NitrateNitrite Content of Irrigation Waters and Per Cent Recovery of Added Nitrate (Procedure A)
Sample
Total Solids, P.P.M.
.536 540 551 557 568 584 585
3476 2053 66 1 1049 757 487 4241
NO? Recovery Present, of Added P.P.M. XOI, 70 0.20 6 84 0 73 0 20 0 32 0.20 1.59
103 107 104 102 104 95 106
for the brucine (6, 6, 8 , 12) proved hazardous as the solution heated rapidly when the sulfuric acid was added, causing rapid volatilization of the chloroform n-ith possible loss of solution and resulting in poor reproducibility. Dissolving the brucine in concentrated sulfuric acid as described by Kolf (11) gives reproducible results. This mixture develops a yello~vcolor on standing due to nitrogenous vapors in the air. A stabk reagent can be prepared by dissolving brucine hydrochloride in distilled water or 0.05S hydrochloric acid. This reagent will slowly develop a pink color and should be discarded after about 1 month. Nitrate ions will not develop color in a solution with a n acid concentration below 25% while nitrite ions will develop maximum color when the acid concentration is 25% or less. An acid concentration of 33% produced a slight color development with nitrate ions. These results are in direct contrast to previous reports as to selective determination of the anions (6, 8, 12). With an acid-water ratio of 1 to 1 or greater, the development of color due to the presence of nitrates is strong and reproducible. Color development from nitrites was markedly affected by temperature (Figure 3). For maximum color development, the temperature must be kept below 70” C. The lack of color development from nitrites in Figure 2 is due to the increased temperature
1974
ANALYTICAL CHEMISTRY
associated with the more concentrated acid solutions-for example, a maximum temperature of 110” C. was obtained by mixing water and concentrated sulfuric acid a t a ratio of 1 to 1 with the automatic pipet arrangement employed. Water-acid ratios of 5 to 1 and 3 to 1 produced temperatures of approximately 70” C. and 100” C., respectively. S o color will develop when a sample containing only nitrite ions in 50% acid is heated to boiling (about 145” C.) before the brucine is added. The addition of brucine to a boiling 50% acid solution containing nitrates will develop color of about a n equal intensity as when the temperature is controlled a t 110’ C. This treatment, boiling a 1 to 1 11-ater-acid solution, might be a method for the direct detection of nitrate ions, even if nitrite ions are present in the sample; but more n-ork is needed to clarify this procedure. As color development by the brucine method is subject to several factors, devising a routine method hecomes somen-hat difficult. A variation in factors will affect the intensity of color developed by a standard amount of nitrate or nitrite. One method of handling this is always to run three points on a standard curve with each set of samples. Operating in this manner and Jvith a variable wave length photometer one can superimpose the standard curve on a predetermined standard curve. The standard curre should be established under low light intensities as this enhances color development, All measurements should be carefully taken, as the final solution is not made to volume. APPLICATION
The nitrate content (Table 11) of a series of soils was determined by the phenoldisulfonic acid ( 1 ) and the proposed brucine method (Procedure C) with excellent agreement. The soils ranged in p H from 5.4 to 7.9, with some having a free lime content of 10% or more. As suggested by No11 ( 5 ) , no interfering ions m r e encountered when determining the nitrate content of soils by the brucine method. The brucine method as developed has been used routinely by undergraduate students in the soil chemistry laboratory with very good success and with a considerable sal-ing of time compared with the method formerly used. Table I11 gives the nitrate content of several irrigation waters that have been processed by the State Chemist of Texas. The samples selected were from wells and streams from all parts of Texas and exhibited a wide range in total
dissolved solids. The recovery of added nitrate is high. CONCLUSION
Data obtained indicate that color development by brucine with either anion closely follows Beer’s law when the nitrogen concentration in a 15ml. aliquot ranges from 0 to 1 y per ml. This gives a range up to 15 y of nitrogen that can be detected with a precision t o &2%, if the outlined procedures are followd. Data obtained show that it is possible with this bne reagent to detect nitrite ions only (Procedure B) or both nitrite and nitrate ions (Procedure L4), which, in turn, makes it possible to calculate, by difference, the amount of nitrate ion present in a mixture containing both anions. Procedure C, which can be used when nitrates only are present, is especially adapted to the analysis of soils or any other highly aerated sample. Technique is of great importance, so care must be exercised in all steps of the outlined methods, and even then some variation in color intensity will occur. This variation can be compensated for by use of a variable wave length spectrometer (Bausch R: Lomb Spectronic 20). ACKNOWLEDGMENT
The authors wish to evpress their appreciation t o Ellen Cranford and June Cooper for their faithful services in the laboratory while these methods were being developed. LITERATURE CITED
(1) iissoc. Offic. ilgr
Chemists, Kashington, D. C., “Official Methods of Analysis,” 7th ed., p 535, 1950. (2) Greenberg, A. E., Rossum, J. R., 310skowitz, Y., Villarruz, P. A , J . A m .
W a t e r Works Assoc. 50, 821 (1958). (3) Hatfield, W. D., Division of Water,
Se-ivage, and Sanitation Chemistry, 110th meeting, ,4CS, Chicago, Ill., September 1946. (4) Joham, H. E., Plant Physiol. 26, 76 (1951). (5) Soll, C. A, IND.EAG CHEM.,ANAL. ED. 17, 426 (1945). (6) Peech AI., English, L , Soil Sci. 57, 167 (1944). ( 7 ) Prince, L , Ibid., 59, 47 (1945).(( (8) Snell, F. D., Snell, C. T., Colorimetric Methods of Analysis,” T’ol. 11, pp. 798-9, Van h’ostrand, Sew York, 1949. (9) Stanford, G , Hanway,- J., Soil Scz. Soc. Am. Proc. 19,74 (1935). (10) Winkler, L. S . , Chem. Zt9. 23, 454 (1899); 2 5 , 586 (1901). (11) K o l f , B., IND.ENG.CHEXI.,ilsa~. ED.16,446 (1944). (12) Poe, J. H., “Photometric Chemical Analysis,” Vol. I, p. 318, Kiley, S e w York, 1928.
RECEIVEDfor revietv July 5, 1957 Accepted July 21, 1958.
