July 15, 1929
INDUSTRIAL AND ENGINEERING CHEMISTRY
droxide solution, each portion representing about 20 per cent of the volume of the oil. Several variations have been proposed for increasing the accuracy obtainable by this test, but almost without excep tion they tend to complicate the test, increase the time required, or interfere with the immediate comparison of the colloidal bromophenols. In certain special instances a little more refined technic has been employed and a somewhat higher degree of accuracy has been obtained at the expense of the time factor. The above procedure, however, was
decided upon as the best for the purpose in hand-the of phenols in the by-product coking system.
121
study
Acknowledgment
Grateful acknowledgment is made to Erich h u e for making check analysis relative to establishing the accuracy of the method. Literature Cited (1) Rose and Sperr, Am. Gas Assocn. MOnlhl7, 2, 117, 328 (1920). (2) Skirrow, J . SOC.Chem. I n d . , 27, 58 (1908).
Determination of Nitrate Nitrogen in Tobacco' Hubert Bradford Vickery and George W. Pucher BIOCHEMICAL LABORATORY, CONNECTICUT AGRICULTURAL EXPERIMENT STATION,
N E W HAVEN, CONN.
A modification of the Jones method for determining HE nitrate content of absolutely and r e l a t i v e l y nitrate nitrogen is described which is especially adapted plant tissue is usually greatly reduced, and no espefor the investigation of tobacco or its extracts. The determined by titracial precautions are necessary method consists in the preliminary quantitative retion of the ammonia produced to carry out exactly duplicate moval of nicotine by steam distillation of a suspension after subjecting an aqueous distillations of sample and suspension of the tissue to the *of the tobacco in an alkaline solution. Nicotine may be blank. A determination of determined in this distillate and nitrate in the residue action of a reducing agent. the nicotine content may be by reduction with acid and reduced iron powder. It is Owing to the presence of ammade on the steam distillate. necessary to conduct a blank determination omitting monia, or of substances yieldSteam distillation with alkali the reducing agent, but the relative magnitude of this ing it, in most tissues, it is removes any p r e e x i s t i n g blank is small and it is not highly variable. Data are ammonia from the sample, necessary to conduct a congiven illustrating the wide variability of the proportion trol determination in the aband also hydrolyzes and reof nitrate nitrogen in tobacco and its dependence on sence of the reducing agent, moves most, a t least, of the the type of nitrogenous fertilizer employed in growing and the nitrate content is amide nitrogen together with the Plantcalculated from the difference some ammonia of-other secin the amounts of ammonia ondary origin-e. g., from found. arginine. The occasional use of urea as a fertilizer for toThe wide variation in the nitrate content of samples of bacco introduces a possibility that this substance may be normal tobacco renders the determination of this constituent present in the sample. Although the writers have not yet a matter of considerable importance. Unfortunately, how- found urea in tobacco extracts, control experiments with ever, accurate analyses are difficult, since by the usual extracts to which urea had been added have shown that it is methods a considerable proportion of the nicotine present decomposed to ammonia and removed during the alkaline in the sample comes over during the distillation of the am- distillation. Control experiments also showed that the premonia from alkaline solution. A method for the determina- liminary distillation had no effect upon the nitrate present tion of nitrate in tobacco was therefore sought which should in, or added to, a tobacco extract and that added nitrate be free from inaccuracy due to the volatility of nicotine. could be recovered quantitatively. Many methods have been described for the reduction of Identification of Substance Yielding Ammonia nitrate to ammonia as a step in the quantitative estimation of nitrate nitrogen. Sbydell and Wicher (6)*investigated a As a preliminary step i t was thought essential definitely number of these methods and concluded that the most satisto establish the identity of the substance occurring in tobacco factory results were secured when the reduction was carried out in an acid solution. Methods for the analysis of the which, on reduction, yields ammonia. Accordingly a sample nitrate content of fertilizers, based on this procedure, have of an extract of fresh tobacco leaves was distilled with steam been in common use for several years. The presence of such in the presence of an excess of calcium hydroxide until the ingredients as cyanamide or urea in fertilizers introduces a nicotine was removed. The residue was evaporated to drypossibility of error which can be controlled only by properly ness, powdered, and extracted in a Soxhlet apparatus with conducted blank experiments, and Jones (6) has described a 95 per cent alcohol, in which calcium nitrate is soluble, until modification of the acid reduction method which can be the diphenylamine reaction for nitrate could no longer be applied in such cases. His method has recently been em- obtained upon the residue in the extractiou thimble. The extract was evaporated, taken up in cold water, filtered from ployed for the analysis of tobacco (8). The present method is a modification of the Jones method. a little fat, and treated with nitron acetate solution. Nitron I n order to avoid the high blank determinations due to the nitrate crystallized almost immediately and, when recrystalpresence of nicotine, this substance is removed from the lized from water, decomposed a t 259-261' C. (not corrected). sample by steam distillation from an alkaline solution. This Busch (3) gives the decomposition point as 260' C. The dehas several advantages. The blank determinations are both composition point of a mixture of this preparation with nitron nitrate prepared from the pure potassium salt was 1 Received April 3, 1929. The expenses of this investigation were 261" C. The sample of tobacco leaf extract so treated conshared by the Connecticut Agricultural Experiment Station and the Carnegie tained 0.149 gram of nitrate nitrogen, as indicated by the Institution of Washington, D. C. * Italic numbers in parenthesis refer to literitture cited at end of article, iron reduction method described below. The nitron ni-
T
ANALYTICAL EDITION
122
trate secured weighed 0.374 gram, equivalent to 0.140 gram of nitrate nitrogen, a yield of 94 per cent. Method
The material, either dried, fresh leaf, or cured tobacco, is ground to a fine powder and mixed. Two 5-gram samples are transferred to Kjeldahl flasks; 30 cc. of water, a small piece of paraffin, and a few angular quartz pebbles are added and the mixture is shaken; 5 cc. of 50 per cent sodium hydroxide are added and the flasks are immediately connected to an apparatus arranged for distillation in a current of steam. The distillate from each flask is collected in a 1-liter beaker containing a suitable quantity of 0.1 N hydrochloric acid, 100 cc. of water, and a few drops of methyl red solution. The adapters of the condensers are dipped below the surface of the acid in the receivers and a moderate current of steam is admitted to the flasks. Micro burners beneath are turned on full until the volume of the contents has been reduced to 10 to 20 cc. The current of steam is then increased and the micro burners so adjusted as to maintain the volume of fluid in the flasks approximately constant. Distillation is continued until 800 cc. have passed over (30 to 45 minutes). Titration of the distillates gives a measure of total volatile base (nicotine ammonia unidentified volatile bases). The distillates are subsequently used for the determination of nicotine by the silicotungstic acid method (1). The flasks are removed from the still, the inlet tubes and walls are rinsed down, and 25 cc. of water are added to each; 15 to 16 cc. of 1:l sulfuric acid are then added and the flasks are shaken. To one flask 3 grams of reduced iron powder (weighed to *0.2 gram) of known nitrogen content are added; the flask is rotated and then allowed to stand until ' t h e reaction has moderated. Funnels are placed in the necks of both flasks to serve as condensers and the contents are slowly heated and then boiled gently for 5 minutes with occasional shaking; 200 cc. of cold water and a few angular quartz pebbles are added to each flask and the contents are made alkaline by the addition of 30 to 35 cc. of saturated sodium hydroxide solution. The flasks are a t once connected to a Kjeldahl dishillation apparatus and the contents distilled into 0.1 N hydrochloric acid until approximately 150 cc. of distillate have been collected. The difference between the acid consumed in the titration of the two distillates, further corrected for the titration value of the ammonia contained in the reduced iron powder, gives the nitrogen equivalent of the nitrate in the original 6-gram sample.
+
+
Vol. 1, No. 3
determine the ammonia in these solutions by the use of permutite (4),since nicotine is not taken up by this reagent. The sum of the alkalinity due to the nicotine and that of the ammonia, when deducted from the total titration figure, invariably leaves a small quantity which we attribute to the presence of traces of as yet unidentified volatile bases. This subject is under investigation. The application of Folin's permutite technic to the determination of ammonia and amide nitrogen in tobacco is to be described in a forthcoming paper. Table I-Analysis
of 5-Gram Samples of Dry Tobacco for Nitrate Nitroeen and Nicotine -
zXPT. 0.1 N HC1 AFTER 0.1 N HC1 REDUCTION BLANK
cc. 45
6.33 6.45 46 12.25 13.50 13.89 13.40 48 5.50 5.95 51 30.00 30.80 30.20 53 24.30 24.20 23.10 55 9.42 10.00 9.68 83 4.60 4.29 85 5.10 5.35 42 5.35 Extract = 5 grams 5.01 44 5.32 Extract = 5 grams 5.42
CC. 1.80" 1.74 1.80 1.60 1.70 1.70 1.80 1.80 1.95 1.80 1.95 1.85 1.90 h
1.50 1.50 1.60 1.40 1.47 1.40 0.60 0.61 0.53 0.60
NO8
NITROGENNICorlNE Per cent 0.11 0.12 0.28 0.32 0.32 0.31 0.087 0.099 0.77 0.80 0.77 0.61 0.61 0.58 0.21 0.22 0.21 0.067 0.064 0.085 0.094 0.13 0.12 0.12 0.12
Per cent 1.96 1.98 2.06 2.01 1.50 1.48 1.75 1.66 1.96 1.92 1.94 1.95 1.22 1.23 2.22 2.26 2.01 1.81
Discussion of Method
Because of reactions yielding a little ammonia which occur when the sample is heated with acid, the blank determination conducted as described cannot be entirely avoided. It is apparent from Table I, however, that the blank is remarkably constant in magnitude and is small in relation to the nitrate determination. The steam distillation of the nicotine can hardly be conducted without the aid of an antifrothing material. Paraffin has proved to be satisfactory, although traces of it distil over. EXTRACT DRYSAMPLE The steam distillate is acidified with 3 cc. of dilute hydroSAMPLE NO8 chloric acid immediately after titration, is then concenNicotine nizzien Nicotine nitrogen trated in vucuo to about 100 cc., transferred quantitatively Per cent Per cent Per cent Per cent through a filter into a 200-cc. flask, and made to volume. 0.32 0.32 A One-quarter of this solution is a convenient aliquot for the 0.31 0.31 A 0.15 0.14 B determination of nicotine by the silicotungstic acid method 0.11 0.12 C 0.78 0.73 when the nicotine content of the tobacco is over 1 per cent. D 1.39 1.20 E The titration of the steam distillate gives a figure from which, after subtracting a quantity of acid equivalent to the Table I11 gives a number of nitrate determinations by nicotine determined gravimetrically, the non-nicotine volatile bases may be calculated. It has been found possible to the Jones method, together with determinations on the
I N D U S T R I A L AND ENGINEERING CHEMISTRY
July 15, 1929
123
same samples by the present modification. Full titration data are shown for the first four samples.
Considerable experience with this modification of the Jones method has shown that in duplicate experiments the blank determinations should check within 0.5 cc. 0.1 N acid while Table 111-Determinations of Nitrate Nitrogen in Tobacco by Jones 0.2 cc. is usual. With tobacco samples containing 0.1 per Method a n d b y Modified Method cent or more of nitrate nitrogen duplicate determinations I TONES METHOD 1 MODIFIEDJONESMETHOD should agree within 5 per cent of themselves. With samples 0 1 N 0.1 N of lower nitrate content a slightly greater variation may be HC1 0.1 N NOa HCl 0.1N NOa O’’ SAMPLE with HCl nitrowith HCl nitro- 7;:-admitted. These conclusions are exemplified by the data of reduc- blank gen reduc- blank gen base Table I. tion tion To illustrate the wide variation in nitrate content of tobacco cc. cc. Per cent cc. cc. Per cent cc. grown with different fertilizers, the data in Table I V are 13.55 1.85a 0.31 21.4 given. 4.73 1.30 0,079 16.6 5.23 1.44 0.090 17.0 It is of particular interest to observe that the proportion 15.15 1.00 0.35 5.4 of nitrate nitrogen in the field crops is lowest in the tobacco 0.09 0.25 grown on sodium nitrate fertilizer. This, of course, is due to 0.20 0.13 the rapid leaching of this substance from the soil under field conditions. The nitrate in the hot-house crop on poa 0.6 cc. to be added to this figure for blank on reduced iron powder used. b Data obtained by Analytical Department of Connecticut Agricultural tassium nitrate fertilization, where no leaching could occur, Experiment Station. was unusually high. The hot-house crop grown without fertilizer contained no nitrate whatever, even the delicate Table IV-Nitrate-Nitrogen a n d Nicotine-Nitrogen Content of Tobacco Grown under Application of Different Nitrogenous Fertilizers qualitative tests being negative. Nitrate nitrogen may (Results calculated on air-dry basis) therefore vary from zero to over 1 per cent of the air-dry TOTAL NO2 NICOTINE NICO- weight of the tobacco leaf and is the most highly variable FERTILIZER NITROGEN NITROGEN NITROGEN TINE nitrogenous constituent of this plant that we have yet encountered. % % % % Although this method is chiefly designed for the investiFIELD CROPS-CURED TOBACCO None 2.54 0.113 0,321 1.86 gation of tobacco, there are many reasons why it may prove C&&eed meal 3.19 0.296 0.346 2.00 advantageous for the determination of nitrate in other plants, 2.63 0.093 Sodium nitrate 0.258 1.49 4.00 0.779 Ammonium sulfate 0,294 1.70 particularly where these contain high proportions of amide 3.71 0.598 0,335 1.98 Urea 2.79 0.213 Calcium nitrate 0.336 1.99 nitrogen.
1
I
~~
Castor pomace
2.45
HOT-HOUSE CROPS-FRESH
None Cottonseed meal and castor pomace Potassium nitrate
2.04 5.2 4.9
0.121
0,352
2.04
0.141
0.816 2.30 2.21
LEAF
0.000
1.19 0.722
Literature Cited 0,398 0.382
While the-agreement between the two methods is fairly close, it is clear that the variation is irregular and in several cases is 20 per cent or more of the quantities measured.
(1) Assocn. Official Agr. Chem., Methods, 1925,p. 66. (2) Bailey, Nolan, and Mathis, Connecticut Agr. Expt. Sta. Rept., 1927, p. 338. (3) Busch, Ber., 88,861 (1905). (4) Folin, J. Bid. Chem., 29,329 (1917). (5) Jones, IND.ENG.CHEM.,19, 269 (1927). (6) Seydell and Wicher, Z . angew. Chem., 24, 2046 (1912).
A New Volumeter Using Water as the Measuring Medium’ J. A. Fries 137 PARKAvE., STATECOLLEGE, PA.
HE chemist or research worker, especially in dealing with gases, frequently needs to know the exact volume of a particular set of apparatus, as well as the volume of reagents used and the displacement values of other filling materials, etc., so that he can at the end of an operation compute the residual contents and dispense with the final sweeping-out process. But, aside from the measuring by means of filling with some liquid, there is apparently no convenient method for such volume determinations. The method presented herein is simple and accurate, and uses water as the reagent or measuring medium. The volumeter is constructed of materials found in most laboratories, and simply by coupling in different sizes of additional water containers it can be made to suit almost any size of apparatus. Further, the method is applicable to any Condition of vapor pressure from nothing to that of complete saturation in the presence of liquid water.
T
1
Received April 5, 1929.
Underlying Principle
Assume that a flask or any other apparatus containing air is connected air-tight to the upper end of a tube, such as a buret, which is filled with water to a definite height, the outlet end being open to the air. If, after the pressure of the gases in the apparatus has been made equal to that of the atmosphere, the tube is opened, the water in the tube will empty only to within a certain distance from the outlet end. The vertical distance of the column of water remaining in the buret represents a condition of equilibrium for the total volume of the particular apparatus plus connections, and the atmospheric pressure. Thus the change of water level in the buret, from zero down, is a measure of the total expansion of the air volume due to a definite change in pressure, and this change in pressure is indicated by the height of the water column from the water level to the outlet end of the tube. The barometric pressure minus the pressure of this water column equals the pressure
