ANALYTICAL
VOLUME5 NUMBER 5
EDITION
Industrial Chemistry AND ENGINEERING
SEPTEMBER 15, 1933
PUBLISHED B Y T H E A M E R I C A NCHEMICAL SOCIETY E. HOWE,EDITOR HARRISON
Photometric Investigation of Nessler Reaction and Witting Method for Determination of Ammonia in Sea Water HENRYE. WIRTHAND REX J. ROBINSON, Univgsity of Washington, Seattle, Wash.
B
UCH (S),in his spectro-
APPARATUS Four Nessler reagents have been prepared and photometric studies of the color developed by them at low ammonia The Zeiss Pulfrich (gradation) the Witting method for concentrations in distilled water and in sea photometer was used. I n order the determination of free amwater has been investigated photometrically. to obtain significant variation in monia in o c e a n w a t e r , found light absorption in the s m a l l I n distilled water, all four reagents give a that the Nessler reagents were range of ammonia concentrainsensitive to concentrations less definite nonsensitive region, varying between tions studied, photometer tubes than 0.05 mg. nitrogen per liter 0.00 to 0.03 and 0.00 to 0.08 mg. approximately 30 cm. long and (in d i s t i l l e d water). It was Treadwell’s reagent, in sea water, does not 2.5 cm. in diameter were contherefore deemed necessary to have a nonsensitive region. It is therefore structed. The per cent light add a known quantity of amtransmitted by the nesslerized possible to determine as little as 0.003 mg. of monia if a lesser a m o u n t was ammonia solutions as compared originally present. Urbach ( 7 ) ammonia nitrogen in sea water without resorting with d i s t i l l e d water is read also has made a photometric into the artifice of adding known amounts of nidirectly on the i n s t r u m e n t . vestigation of the Nessler reagent trogen. The sensitivity of the Treadwell reagent Distilled water is p r e f e r a b l e in distilled water and like Buch increases with increasing chlorinity. With the as a reference standard since it found that Beer’s law holds, but is a f f e c t e d less by light and Treadwell reagent it has been shown that Beer’s did not try concentrations less oxygen than a standard made of than 0.2mg. per liter-the region law does not apply for concentrations of ammonia ammonia-free water plus of g r e a t e s t i n t e r e s t to the nitrogen less than 0.02 nag. per liter. N e s s l e r r e a g e n t as used by oceanographer. Wattenberg (8), There is no error due to adsorption in the Urbach. Since t h e light using his direct nesslerization Witting method for the determination of ammonia a b s o r p t i o n of t h e N e s s l e r method, did not find this minic o l o r is in t h e v i o l e t end of mum sensitivity in sea water and in sea water. the spectrum, a s U r b a c h has was able to estimate 0.005 mg. s h o w n , the 5-43 f i l t e r p r o or less of ammonia nitrogen per liter. Cooper (4) with Wattenberg’s method but a modified vided with the instrument was used. This filter has an reagent was able to distinguish visually between 0.000 and effective range of 200 to 250 p p and transmits light with an 0.003 mg. The authors ( 5 )using an entirely differentreagent average wave length of 430 p p , The general form of the curves was verified by means of the S-47 filter, which has an were unable to determine less than 0.02 mg. It was believed that the inconsistent results previously average wave length of 470 p p , The tubes containing the mentioned were due to the varying sensitivities of the Nessler ammonia and distilled water solutions were always interreagents employed, and in the Witting method to partial changed to avoid errors owing to unequal illumination of the adsorption of ammonia during the precipitation of the two sides of the photometer. Each reading was checked interfering ions, If adsorption does take place, the Witting by another observer so that a t least four readings were method is useless. This is of importance, as this method averaged for each sample reported in the various figures. is otherwise the better for the determination of ammonia in sea water having a large plankton content. Plankton, REAGENTS AND SOLUTIONS which is believed to cause a turbidity in the sea water standNessler reagents were made according to the directions ards of the Wattenberg method, is removed in the Witting procedure by occlusion in the precipitate of barium sulfate, given by ( A ) Ringer and Klingen; ( B ) Raben; (C) Standard Methods of Water Analysis; and (0)Treadwell. All calcium carbonate, and magnesium hydroxide. 293
ANALYTICAL EDITION
294
chemicals ‘were freed of ammonia, and ammonia-free water was used a t all times. RINGER AND KLINGEN (8). Add slowly 35 grams of potassium iodide in 100 ml. of water to 16 grams of mercuric chloride in 300 ml. of water until a permanent precipitate remains. Add slowly 200 grams of potassium hydroxide in 600 ml. of water t o the above solution. RABEN(9). Add 50 grams of potassium iodide in 50 ml. of boiling water t o a boiling solution of 25 grams of mercuric chloride in 300 ml. of water. Filter the cooled solution into 300 ml. of water containing 150 grams of potassium hydroxide, and then dilute to a liter. STANDARD METHODS OF WATER ANALYSIS(1). To 50 grams of potassium iodide in 35 ml. of cold water add a saturated mercuric chloride solution until a slight precipitate persists. Add 400 ml. of 9 N potassium hydroxide and dilute to one liter. TREADWELL (6). Dissolve 115 grams of mercuric iodide and 80 rams of potassium iodide in enough water t o make 500 ml. Ad8 500 ml. of 6 N sodium hydroxide.
Vol. 5 , No. 5
LIGHTABSORPTION BY
NESSLER COLORIN DISTILLED WATER
The variation of light absorption in distilled water with varying ammonia content was first determined one hour after nesslerization for each of the four Nessler reagents (Figure 1). Time of color development was also determined for a concentration of 0.1 mg. ammonia nitrogen per liter (Table 11). TABLE11. TIME OF DEVELOPMENT OF NESSLERCOLORIN DISTILLEDWATERCONTAINING 0.1 MG. AMMONIANITROGEN PER LITER REAQENT Ringer and Klingen Raben StandardNethods Treadwell
PERMEABILITY IN PERCENTAFTER: 0 94.0 91.0 40.0 34.0
0.5 84.0 86.0 29.0 17.5
(Time in hours) 2 26 32.3 23.0 79.0 33.0 27.0 26.0 1 6 . 0 17.2
1
44.0 83.0 29.0 17.0
34 24.5 30.0 23.5 17.5
50 22.2 25.5 22.0 18.0
The concentrations are compared in the following table:
The Treadwell and Standard Methods reagents showed practically complete color development a t the end of 30 TABLEI. CONCENTRATIONS OF VARIOUSNESSLER REAGENTS minutes, whereas the development was complete only after CONCENTRATION Hg HYDROXIDE REAQENT (as K%HgL?) CONCENTRATION5 and 10 hours in the case of the Ringer and Klingen, and 0.059 M 3.6 N Ringer and Klingen Raben reagents, respectively. The color intensity was un0.092 M 2.7 N Raben changed for 50 hours, but was exposed to the photometer 0 . 0 9 M (approx.) 3.6 N Standard Methods 0.25 M 3.0 N Treadwell light only during readings. Urbach found that color started These reagents were selected because they have a l l been t o fade after 30 minutes, but his samples were illuminated continuously. used in the determination of ammonia in sea water, The It might be thought that all four Nessler reagents would reagent of Urbach gives approximately the same concentragive the same curve if sufficient time were allowed for comtion as Raben’s when 1 ml. is used per 100 ml. of water plete color development. This was not the case. After instead of 2 ml. per 100 ml., as is usual with the other reagents. 24 hours the form of the curves remained unchanged, while Wattenberg (9) suggests the use of a Nessler reagent (attributed by him to Treadwell) which is approximately 5 N the nonsensitive region was reduced by only 0.01 mg. in the in sodium hydroxide and 0.2 M in mercury. The authors case of the Ringer and Klingen and Raben reagents, It was thought desirable to compare these results with were unable to prepare such a reagent, as a copious prethose in sea water which had been diluted with the Witting cipitate was obtained when alkali was added. reagents. To accomplish this the amount of ammonia actually present in the distilled water wa6 only 2s/28 of that indicated on the graph.
MILL IGRAN
F NtTROGW PFR LITrR
FIGURE1. SENSITIVITY OF NESSLER REAGENTS ( A , RINGERAND KL7NGEN; B , RABEN; C, STANDARD METHODS; D , TREADWELL) IN DISTILLED WATER USING“S-43” FILTER The barium chloride (200 grams of barium chloride-dihydrate per1iter)and sodium hydroxide-sodium bicarbonate (200 grams of sodium hydroxide and 69 grams of sodium bicarbonate per liter) solutions were freed of ammonia by partial evaporation. Ammonia-free sea water was prepared by adding half a volume of distilled water to sea water and evaporating to the original volume. From this the desired chlorinities were obtained, either by further evaporation or dilution as the case might be.
ACCURACY OF WITTINGMETHOD To 500 ml. samples of ammonia-free sea water (C1 = 14.4 parts per thousand) 0.000, 0.0025, 0.005, 0.015, 0.025, 0.04, and 0.06 mg. of nitrogen as ammonia was added, the sample well shaken, and the interfering ions precipitated with 16 ml. of barium chloride solution and 32 ml. of sodium hydroxide-sodium bicarbonate. After standing one week, the clear supernatant liquid was siphoned, and 125 ml. portions were nesslerized with 2.5 ml. of reagent (Standard Methods and Treadwell). The color intensity was evaluated by means of the photometer after 30 minutes. A 3500-ml. sample of the same water was treated with proportionate amounts of barium chloride and sodium hyhydride-sodium bicarbonate solutions. The clear liquid was siphoned and sufficient ammonia added to 125-ml. portions to give the same concentrations as above. The permeabilities are given in Figure 2. Attempts were made to speed the settling of the precipitate by centrifugalizing, since it is desirable to analyze the samples immediately, but turbidities were always obtained upon nesslerization. Evidently the precipitation is complete only on standing for several days. If I is the value of the scale reading (permeability in per cent) and 10 is the value of I corresponding to zero ammonia concentration, then -log I/Io plotted against the ammonia content should give a straight line if Beer’s law holds for these dilutions. This has been done in Figure 3 for the Treadwell reagent, in distilled water, and sea water of chlorinities 10.0, 14.4, and 18.0 parts per thousand. The plotted values for sea water were those obtained after first precipitating with barium chloride and sodium hydroxide-
September 15,1933
INDUSTRIAL AND ENGINEERING CHEMISTRY
sodium bicarbonate solutions and then adding known quantities of ammonia. To determine the ammonia content of sea water, the following procedure was used: To 250 ml. of the sample in a 300-ml. bottle add in succession 10 ml. of barium chloride solution and 20 ml. of sodium hydroxide-sodium bicarbonate, shaking after each addition. With chlorinities less than 17 parts per thousand, smaller quantities of reagents were used (C1 = less than 11.0 parts per thousand, 5 ml. BaClz; C1 = 11.0-13.9 parts per thousand, 6.5 ml. BaClz; C1 = 14.0-16.9 parts per thousand, 8.0 ml. BaC12, with twice as much NaOH-
295
The Standard Methods reagent shows a definite but shorter nonsensitive region in sea water, but its use is excluded by a turbidity which always appears a t ammonia concentrations of 0.1 mg. nitrogen per liter or higher, which is not the case with the Treadwell reagent. The Ringer and Klingen and Raben reagents are not applicable, as they develop their color too slowly, usually give a turbidity, and have large nonsensitive regions in sea water as well as in distilled water. Two Nessler reagents which had been prepared for 19 months were also tested, one of which, the Treadwell reagent, was much less sensitive and dependable in distilled water, but gave satisfactory results in sea water. The other, a Standard Methods reagent, was slightly more sensitive (distilled water) after standing, but was not tested in sea water. The above Standard Methods reagent was the one used by the authors (6) in the determination of ammonia in sea water by the Wattenberg method. As the minimum sensitivity for this reagent is 0.02 mg. of nitrogen per liter, the inability of the authors to detect less than this quantity is thus explained. Moreover, it should be noted that this minimum sensitivity likewise applies to the determination of Kjeldahl and albuminoid nitrogen in water so that a zero blank does not necessarily mean there is no ammonia, but merely that 0.02 mg. or less of nitrogen per liter is present. r
MILLIGRAfl OF NITROGEN PEff LITCR
FIGURE2. ACCURACYOF WITTING IN SEAWATER USING“S-43” METHOD FILTER (C, STANDARDMETHODS D, TREADWELL REAGENT) REAGENT; A Precipitated before addition of NHa 0 Precipitated after addition of NHs
NaHCOa in each case). This eliminated difficulties encountered in obtaining clear solutions on nesslerization, which were believed to be due to too great an excess of reagents. After standing for a t least 3 days, the supernatant liquid was siphoned and the ammonia determined photometrically. This procedure was tested on eighteen samples taken in Hood Canal, April 29 to 30, 1933, and gave excellent results in every case. Three duplicates, which were allowed to stand for 25 days, showed 0.010 mg. more ammonia nitrogen than those tested after 10 days, indicating a probable decomposition of organic matter in the alkaline solution. This would be the greatest increase normally expected, since these samples were especially rich in plankton. The necessity of finishing the determination as soon as possible after sampling is, however, evident.
DISCUSSION It is apparent from Figure 1 that the various Nessler reagents have different characteristics a t low ammonia concentrations, but uniformly show a minimum sensitivity. The form of the curve depends apparently on the concentration of the constituents in the reagent. As far as results in distilled water are concerned, the Standard Methods reagent is preferable, since it has a shorter nonsensitive region. The Treadwell reagent has too great a color of its own, and in addition, samples nesslerized with it always give a precipitate of mercuric iodide on standing. I n sea water, however, the Treadwell reagent gives far better results. I n the chlorinities investigated no limiting sensitivity was apparent, but there was a very definite change in the log curve (Figure 3) a t N = 0.02 mg. per liter in every case. In addition, the sensitivity of the reagent apparently increases with increasing chlorinity.
MILLIGffAfl
OF NITROGW FfR WTLR
FIGURE3. APPLICABILITY OF BEER’S LAW TO NESSLERREACTIONUSING TREADWELL REAGENTAND SMALL CONCENTRATIONS OF AMMONIA IN DISTILLED WATER(D)AND SEA WATER OF VARYINGCHLORINITY (C = 10, B = 14.4, A = 18 PARTS C1 PER THOUSAND)
Figure 2 shows that there is no significant adsorption of ammonia on the precipitate, as in most cases the values by the two methods agree within 0.005 mg. The Witting method can therefore be used in sea water with little error arising from this source. Furthermore, it is possible to obtain an accurate standard curve by addition of known quantities of ammonia to sea water from which the interfering ions have previously been removed. Figure 2 also shows that, with the Treadwell reagent, it is possible to determine directly as little as 0.003 mg. of ammonia nitrogen per liter without resort to the artifice of adding 0.05 mg. of nitrogen to each sample. It has been observed by the authors, and also by Braarud and Klem (2) that artificial ammonia-free sea water as prepared in the above experiments gives a slight color with the Nessler reagent which is not obtained with naturally occurring ammonia-free water. Where possible, therefore, it is desirable to make standards from untreated ammonia-
ANALYTICAL EDITION
296
free sea water, which in the Puget Sound region is most apt to be found in the surface waters. LITERATURECITED (1) Am. Pub. Health Assoc. N. Y., Standard Methods of Water Analysis, 7th ed., 1933. (2) Braarud, T., and Klem, A., Hvalradets Skrifter Norska Viden8 k ~ p s - A k ~ dOslo, . 1, 50 (1931). (3) Buch, K., R a p p . proc. verb. reunion. conseil perm. intern. explor. mer., 53, 36 (1929).
Vol. 5 , No. 5
(4) Cooper, L. H. N., J . Murine Biol. Assoc. United Kingdom, 18, No. 2, 719 (1933). ( 5 ) Robinson, R. J., and Wirth, H. E., J. conseil intern. exploration mer. . Accepted for publication. (6) Trcadwell and Hall, “Analytical Chemistry,” 6th ed., Vol. I, Wiley, 1927. (7) Urbach, C., Mikrochemie, 11, 50 (1932). (8) Wattenberg, H., Ann. der Hydrog. Murit. Meteorol., 59, 95 (1931). (9) Wattenberg, H., R a p p . proc. verb. reunion. conseil perm. intern. ezplor. mer., 53, 108 (1929). RWEIVDDJuly 3, 1933.
X-Ray Method for Quantitative Comparison of Crystallite Orientation in Cellulose Fibers WAYNEA. SISSON~ AND GEORGEL. CLARK,Department of Chemistry, University of Illinois, Urbana, Ill.
T
HAT the orientation of Since these planes are parallel to The relation between orientation of the crystalthe cellulose structural the long axis of the crystallites lites or micelles and various properties of cellulose units varies w i d e l y in ($4, 66), each plane a t angle 0 fibers is pointed out and a n x-ray method for different fibers and in the same to the x-ray beam (fulfilling the quantitatively comparing the orientation is fiber (1, 8) is well known, and its conditions of the Bragg equation described. The method is based upon the asinfluence upon the physical and nX = 2d sin 0) w i l l d i f f r a c t chemical properties of the fiber x-rays in the limiting case, upon sumption that the distribution of the crystallites has been discussed by many 002 diffraction ring a t right the around the pencil of x-rays is proportional to investigators (Ic5, 23), particuangles to the long axis of the the distribution of intensity around the 002 larly with reference to the degree crystallite. The diffraction ring diffraction ring. Intensity measurements are of m e r c e r i z a t i o n , t e n s i l e registered on the photographic made with a microdensitometer equipped with a s t r e n g t h , classification, elasfilm is thus a summation of the ticity, and dyeing properties of individual diffractions from all rotating stage. cotton ( 5 , 6 , 1 9 , 3 1 ) ; the swellthe diffracting crystallites, and a The distribution of the crystallites is calculated comparison of the intensity dising, elasticity, tensile strength, from the intensity values and the orientation is tribution around this ring gives ability to take dyes, resistance expressed by distribution curves which may be to e n z y m a t i c decomposition, a measure of the distribution of differentiated f r o m one another by statistical the crystallites around the pencil gloss, creasing resistance, refractiveindex, and extension of rayon of x-rays. For a complete picmethods. The data obtained may be used to (3, 5 , 7 , 9,11,18,19, 21,22,28); ture of the orientation in any study the structure of the fiber or to predict sample it is necessary to obtain and the density, tensile strength, physical and chemical properties which are an x-ray pattern with the sample e x p a n s i o n , and shrinkage of anisotropic. Typical data for three grades inclined at various angles wood ( I , 17, 32). of cotton are presented to show the sensitiveness The use of x-rays in studying to the x-ray beam. However, this orientation and differentifor comparing the orientation of the method. of different fibers, one pattern ating b e t w e e n the v a r i o u s taken w i t h t h e x - r a v b e a m textce fibers, especially rayon, has been described by Clark ( 5 ) , Mark (20), and others. perpendicular to the fiber axis is sufficient. A perfectly uniThese investigators, however, used a visual method of com- formly intense 002 diffraction ring, such as that obtained with paring the x-ray patterns which is essentially qualitative in the x-ray beam perpendicular to the cellulose sheet, implies nature. It is the purpose of this paper to describe briefly that the long axes of the crystallites have a random orientaan improvement over the visual method, which enables a tion in the plane of the sheet. If the intensity is concenquantitative comparison to be made, and to point out the trated into localized maxima, as is the case for the x-ray beam perpendicular to the ramie fibers, then the crystallites appossible applications of the method. The improved method is based on a comparison, for differ- proach a parallel arrangement along the fiber axis. ent samples, of the relative intensity distribution along the EXPERIMENTAL circle on which the 002 interference maxima are localized. Although the significance of cellulose fiber diagrams has been The intensity measurements are made on a microdensitomedeveloped by Polanyi and Weissenberg (26, 27) and the rela- ter of the “photograph wedge” type, the principle of which tion between crystal orientation and sharpness of the localized was first described by Hartman (14) and more recently by maxima is explained in textbooks on x-rays (2, 4), the basis Vasil’ev (36). It is manufactured by the Gaertner Scientific of the method may be explained briefly as follows: Corporation and involves some novel features. The wedge Figure 1 shows typical x-ray patterns of a cellulose sheet used is not strictly a photographic wedge but a dyed gelatin and a bundle of ramie fibers. I n both patterns the most wedge between glass plates. This is essential in order to give intense interference ( A ) line) is due to the 002 planes (2.4). the necessary linear relation between density and scale readings. To make up for the lack of grain of this type wedge a 1 Senior Textile Foundation Fellow.
