Chromatographic Separation and Identification of Photographic Developers JAMES H. PAKNELL
AND JATtIES E. L U V A L L E Technical Operations, Znc., iirtington, Mass.
course of a study of reactions of photographic developers, I-it became necessary to devise methods for the separation and of the developers and their oxidation products. v
THE
identification Knowledge of the reactions of developers was seriously limited by a dearth of information on such separations. A method based on paper chromatography was evolved which should be of value as it has the advantages of simplicity and specificity. However, it is probably applicable only to substances which are moderately stable to air.
Table I. R/ Values of Some Photographtic D e v e l o p e r s Hydroquinone disulfonate p-Methylaminophenol monosulfonate Hydroquinone monosulfonate Quinone monosulfonate p-Aminophenol hydrochloride p-Phenylenediamine hydrochloride p-Methylaminophenol sulfate Hydroquinone
0.00 0.09 0 15 0 34 0 43 0 5.5 0 61 0 80
layer is separated and used. .4s spray reagent, a 2% solution of ammoniacal silver nitrate gives the highest sensitivity. A 5% solution of phosphomolybdic acid, while less sensitive, produces chromatograms which do not darken with time. Best results are obtained when 10 y amounts of developers are used, although 1 y can be detected. Table I shows R/ values obtained by descending developmenton JVhatman S o . 2 paper using butanol-acetic acid-water solvent at room temperature (22’ C.) where the solvent front moved 40 cm. Previous published work available gives the datum only for hydroquinone (2), of the substances tabulated, with an RJ value of 0.88 obtained under similar conditions.
R I,
ACKNOWLEDGMENT
& 0.01
f 0 01 f 0 02 4= 0 03 i 0 03 f 0 02 i 0 02
This investigation was sponsored by the Air Research and Development Command under Air Force Contract KO.AF18 (600)-3i1. LITERATURE CITED
Bate-Smith, E. C., Biochem. SOC.Symposia (Cambridge, Enol.), 3, 62 (1949). (2) Block, R. J., “Paper Chromatography,” p. 124, New York. dcademic Press, 1952. (1)
The method employs standard chromatographic equipment and a solvent composed of butanol, acetic acid, and water ( I ) , mixed in the volume proportion 4 to 1to 5, fromwhich the organic
RECEIVED March 19, 1953. Accepted July 2, 1953.
Spectrophotometric Determination of Glycerol as Sodium-Cupri-Glycerol Complex FRANK SPAGNOLO Research Laboratories. National Lead Co., Brooklyn, !V. Y .
investigation of the analysis of glycerol-containing mateIrials, the literature revealed a method by Bertram and Rutgers (1)based on the formation of the sodium-cupri-glycerol comv
AN
plex and its subsequent iodometric determination. A later method published by Whyte ( 4 ) describes the determination of glycerol by spectrophotometric measurement of the blue-colored sodiumcupri-glycerol complex. On analyzing various glycerol-containing materials in the writer’s laboratory, it became desirable to further increase the sensitivity of the spectrophotometric method for application to smaller working quantities of glycerol than employed by Whyte, and to test the application of such a method to products not previously analyzed by the spectrophotometric method. Accordingly, a modified procedure was developed whereby 1. Sensitivity is increased by use of absorption cells of longer light path. 2. Titration of the glycerol solution with cupric chloride reagent to a faintly perceptible turbidity is eliminated by use of a definite amount (6.0 ml.) of reagent and a working aliquot of solution t o contain 17 to 65 mq. of glycerol for the color development. This modification in effect reduces operating variables between analysts. 3. The method can be applied t o products not previously examined, such as glycerol esters and resinous vehicles.
Condensers, water-cooled, to fit Erlenmeyer flasks. Buret, for delivering the cupric chloride reagent. pH meter or Universal-type pH paper. REAGENTS
hqueous sodium hydroxide solution containing 21.0 grams of sodium hydroxide ( 3 ~ 0 . 2gram) per 100 ml. Ethyl alcohol, 95%, reagent grade or formula 3A. Cupric chloride reagent. Dissolve 10.0 grams (fO.l gram of cupric chloride dihydrate) reagent in 95% ethyl alcohol and dilute to 100.0 ml. Benzene, reagent grade. Dissolve 66.0 grams of reagent Potassium hydroxide, 1.0 J\-. grade (85%) potassium hydroxide in 95% alcohol and dilute to 1 liter. Hydrochloric acid, reagent grade. Ethyl ether, reagent grade. Glycerol, reagent grade. Determine purity by the American Oil Chemists’ Society’s specific gravity method No. E.4-7-50, and apply the necessary gravimetric correction. PRODEDURE
Preparation of Standard Calibration Curve. Pipet into 100ml. volumetric flasks various volumes of an aqueous stock solution of glycerol to contain 17 to 65 mg. of glycerol. (At least 5 or 6 aliquots should be taken for the calibration.) Add distilled water to bring the volume to 10.0 ml. Add 10.0 ml. of 21% sodium hydroxide solution, and 60 ml. of ethyl alcohol. rldd slon-ly from a buret, 6.0 ml. of cupric chloride reagent with vigorous swirling. Stopper, shake vigorously for 2 minutes, and dilute to volume with ethyl alcohol. Transfer a well-mixed portion of the colored mixture to a 50ml. centrifuge tube, stopper, and centrifuge a t a relative centrifugal force (r.c.f.) of 420 for 10 minutes. (R.c.f. = 28.6 n * ~ , where n = 1000 X r.p.m., and r = distance in inches from the center of rotation to the center of gravity of the solution.) The time required for sedimentation can be decreased by increasing
APPARATUS
The Beckman Model DU spectrophotometer, equipped with 5.0-cm cells. Volumetric flasks, lOO-ml., 200-ml., and various other sizes. Volumetric pipets various sizes. Separatory funnefs, 250-ml., pear-shaped. Centrifuge, equipped with 50-ml., rubber-stoppered tubes. Erlenmeyer flasks, 200-ml., glass-stoppered, 24/40$, with pourout lip. 1566
V O L U M E 2 5 , NO. 10, O C T O B E R 1 9 5 3 the centrifugal force. Transfer the clear supernatant liquid to the spectrophotometer cell and determine the absorbancy a t 635 mp against a reagent blank similarly prepared using 10.0 ml. of water in place of the glycerol sample. Analysis of Aqueous or Alcoholic Solutions. The method of sampling and predilution will depend upon the nature of the sample and the glycerol content. Acidic or basic samples are first neutralized with hydrochloric acid or sodium hydroxide, and adjusted to suitable volume before taking an aliquot for analysis. In any case, the procedure of sampling and dilution should be such that volumetric errors are kept a t a minimum. Pipet accurately an aliquot of the neutral sample to contain 18 to 64 mg. of glycerol (maximum volume 10.0 ml.), transfer to a 100-ml. volumetric flask, and adjust to a 10.0-nil. volume with distilled water. Proceed with the color development and measurement of absorbancy as previously outlined. Determine the glycerol concentration in the aliquot taken for analysis by referring to the calibration curve, and calculate the glycerol content of the sample as follows:
% Glycerol where C
.i.b’. TV
=
= =
=
2 x B.F. 1000
x
100
W
concentration of glycerol in the aliquot taken, expressed as milligrams prr 100 ml. of the final colored solution aliquot factor weight of sample in grams, taken for analysis
Analysis of Glyceryl Esters and Resinous Vehicles. Weigh accurately into a 200-ml. Erlenmeyer flask sufficient sample to yield 0.36 to 1.28 grams of glycerol upon saponification (10.0gram maximum). Add 10 ml. of benzene and warm gently if necessary to diisolve the sample. Add 60 ml. of 1.0 S alcoholic potassium hydroxide, attach a condenser, and reflux for 1.5 hours. Cool to room temperature, xash down the condenser and joint with a few milliliters of water, and tranqfer to a 250-ml. separatoiy funnel with the aid of water from a wash bottle. Dilute with water to approsinlately 150 ml., add 8 ml. of concentrated hydrochloric acid, and cool to room temperature. Extract with successive, 15-ml. portions of ether until a colorless ether extract is obtained, ombining the ether extracts in a third separatory funnel. Wa h the combined ether extracts with four successive 5-ml. portions of nater, adding each waqh to the main aqueous phase. Seutralize the final aqueous ?\tract 1% ith the 21 % ’ sodium hydroxide solution. Transfer quantitatively to a 250-ml. beaker with the aid of water, and warm on the steam bath to expel dissolved ether. Cool to room temperature, transfer quantitatively to a 200-ml volumetric flask, and dilute to volume. Take a 10.0ml. aliquot and proceed with the color development as previously outlined. If the final solution is not clear, filter a portion through No. 40 Whatman paper, and take a 10.0-ml. aliquot of the filtrate for the color development as previously described. Calculate the glycei ol content, expressed as grams of glycerol liberated upon saponifying 100 grams of sample, as follows:
% Glycerol where C
=
Tt’
=
cx
2.0
= ___
w
concentration of glycerol in the 10.0-ml. aliquot, expressed as milligrams pel 100 ml. of the final colored solution weight of sample in grams, taken foi analysis EXPERIM EYTA L
The blue sodium-cupri-glycerol complex exhibited a rather broad absorption curve, with a maxima at 630 to 635 mp. This is in agreement with the earlier data of Whyte (4). A stock solution of C.P. glycerol was made up by accurately weighing from a dropping bottle 0.8 gram of glycerol into a 100ml. volumetric flask, and diluting to volume with distilled water. The purity of the glycerol was previously determined b y specific Table I. Glycerol, Mg. 17.20 24.55 40.92 49.11 65.50
Calibration Data Absorbancy a t 635 XI# 0.175 0.288 0,543 0.689 0.950
156’2 gravity, and the appropriate gravimetric correction was applied. Suitable aliquots were then taken for preparing the calibration curve (Table I). It will be noted that, although Beer’s law is not obeyed, a linear relationship is obtained for glycerol concentrations of 17 to 65 mg., which is satisfactorily reproducible under the conditions specified. All readings were obtained at constant slit width (0.066 mm.) in preparing the calibration curve, and for all subsequent analyses. When working with concentrations of glycerol of less than 17 mg. for color development, the readings were not reproducible; they were highly erratic. Various solutions of glycerol were carefully prepared and analyzed by the spectrophotometric method, and by the periodateiodometric method of Reinke and Luce ( 2 ) . The results are shown in Table 11. Table I11 shows the results obtained on glyceryl esters and various commercial resinous vehicles. I n the latter case, the periodate method was applied to the aqueous glycerol extract obtained as described in the spectrophotometric procedure.
Table 11. Analysis of Glycerol Solutions
Glycerol Solution I n dilute hydrochloric acid I n methanol I n dilute sodium hydroxide I n water
Glycerol Found, % Spectrophotometric method
% Error,
Glycerol Present, Theory, 7c
Periodate method
Spectrophotometric Method
23.0 3.9
22.3 3.7
22.5 3.8
-2.18 -2.56
10.9 51.6
10.8 50.7
10.8 51.0
-0.92 -1.16
~
_
Table 111. Analysis of Glyceryl Esters and Resinous Vehicles
Sample Alkali refined linseed oil Triacetin Oil modified alkyd resin solution A Oil mpdified alkyd resin solution
varnish Ester w i n Glyceryl rnonoricinoleate
Glycerol Found, % -~ 7% Error, SpectroSuectrophotometric photometric method Method
Glycerol Present, Theory, 70
Periodate method
10.5 42.1
10.3 39.9
10.6 41.3
+ 1 15 -1.90
..
11.9
12.3
....
..
.. 25.0
7.88
8.03
3.54
3.49
4.01 5.59
4.28 5.81
24.4
25.7 Av. % error
..,.
..
.... =
+2.80 1 1 8
Dehydrated castor oil.
Various polyhydroxy compounds were tested with the reagent. Ethylene glycol and 1,Qpropylene glycol formed a blue color and therefore would interfere with the determination of glycerol, Pentaerythritol, diethylene glycol, dipropylene glycol, trimethylene glycol, and polyethylene glycol did not form any color with the reagent and therefore will not interfere if present. Schoorl (3) has stated that a slight excess of cupric chloride results in the incomplete formation of the colored complex, and that a large excess yields low results. The data shown in Table \’ indicates that only a slight excess of cupric chloride results in incomplete formation of the complex. However, under the specific conditions of the modified procedure, using 6.0 ml. of cupric chloride, and wherein excesses of 400 to 1800% are incurred, a straight-line calibration and reproducible results are obtained on 17- to 65-mg. quantities of glycerol. Whyte (4)adds the reagent until a persistent slight turbidity is obtained, then adds 1 ml. in excess. Others have stated that the blank is unstable and must be examined immediately. Since it was desirable in this case to add a constant amount of reagent
_
ANALYTICAL CHEMISTRY
1568
to shorten working time and to minimize operating variables between analysts, a series of experiments were performed to determine the effect of various excess quantities of reagent on the stability of the blank and colored solution. STABILITY OF BLANK. The absorbancy of blank solutions versus distilled water were measured a t various time intervals, each blank solution containing a different amount of cupric chloride reagent. The results are shown in Table IV. STABILITY OF COLORED COMPLEX.The effect of excess reagent on the stability of the colored complex was investigated. Table V shows the results obtained. The absorbancy of each colored solution was measured by using in the reference cell a blank prepared simultaneously, and to which was added an equal amount of cupric chloride reagent as indicated in the table.
Table IV.
Effect on Absorbancy at 635 Mg of Varying A m o u n t s of Cupric Chloride
Cupric chloride added, ml. Time in min. after centrifuging 1-5 15 30 Decrease in absorbancyafter 15min. ~
0.7
0.9
2.3
4.1
6.0
6.0
0.053 0.060 0.062
0.053 0.060 0.061
0.040 0.044 0.048
0.032 0.036 0.037
0,038 0.039 0.041
0.041 0,042 0.044
0.007
0.007
0.004
0,003
0,001
0,001
~~
Table V. Effect on Absorbancy at 635 M p of Excess Cupric Chloride Reagent on Stability of Complex Sample 1 2 Glycerol taken, mg. 36 36 Cupric chloride rea0 7 0.9 gentadded,rnl. Equivalent % excess reagent presenta 5 35 Time in min. after centrifuging 1-5 0.430 0.495 15 0.445 0.493 30 0.470 0.492 Variations in absofbancy after 15 min. 0.015 0,002 0 Assuming CaHa(0H)r CuCln.
3 36
4 36
5 36
6 25
7 17
2 3
4 l
6 0
6 0
6.0
248
515
800
1200
1800
0 . 5 0 8 0.482 0.474 0.300 0.171 0.509 0.481 0.475 0.301 0.172 0 . 5 1 4 0 . 4 8 4 0 . 4 7 8 0 . 3 0 1 0 173 0.001
0.001
0.001 0 . 0 0 1
0.001
Under these conditions, little variation, if any, is encountered if readings are taken within the first 15 minutes after centrifugation. In view of the increased sensitivity of the modified method, it can be advantageously applied to those cases where only relatively small quantities of sample are available, or where the glycerol content is too low for determination by other methods. ACKNOWLEDGMENT
CONCLUSIONS
Gilder the conditions prescribed, optimum stability of the blank solution was obtained by the use of 6.0 ml. of cuprir chloride reagent. Optimum stability of the colored complex was observed a t 35 to 1800% excess reagent levels, whereas stability with only a slight escess of reagent wag very poor. According to the modified procedure, 400 to 1800% excesses are encountered, depending on the glycerol concentration. In view of these results, it was not desirable to test the effect of greater quantities of reagent than 6.0 ml.
The author wishes to express his appreciation to hlrs. hl. hi. Agarwal of the Brooklyn Laboratories for her aid in obtaining the experimental data. LITERATURE CITED
R., Rec. traz. chim., 57, 681 (1938). (2) Reinke, R . C., and Luce, E. N., IND.ENG.CHEM.,ANAL.En.. (1) Bertram, S. H., and Rutaers,
18,244 (1946). (3) Schoorl, K.,Pharm. Weekblad, 7 6 , 777 (1939). (4) Whyte, L. K., Oil & Soap, 23, 323 (1946). RECEIVED for review March 20, 1953. Accepted June 17, 1953.
Determination of Small Amounts of Niobium and Tantalum Using Radioisotope Tracer Technique THOMAS F, BOYD AND MICHAEL GALAN Industrial Test Laboratory, Philadelphia Naval Shipyard, Naval Base, Phihdelphia 12, Pa.
an investigation of a colorimetric method ( 1 ) for the D determination of niobium and tantalum in austenitic steel, i t was desirable to know how completely these elements were preT
RixG
cipitated a t various steps of the analyeis. An outline of the method investigated which is essentially a modification of that of Thanheiser (2) follows. ANALYTICAL PROCEDURE
Two grams of sample are dissolved in a mixture of concentrated hydrochloric and nitric acids. Thirty milliliters of perchloric acid (70%) is added, and the solution is boiled until the chromium is oxidized t o chromic acid. Two-hundred milliliters of hot water, 10 ml. of concentrated hydrochloric acid, and 50 ml. of saturated sulfurous acid are added, and the solution is boiled for 3 minutes. Paper pulp is added, and after standing 15 minutes on the steam bath, the solution is filtered through S o . 40 Whatman paper. The paper is mashed 15 times with hydrochloric acid (2%). The paper and precipitate are ignited a t a low temperature in a platinum crucible until the precipitate is free of carbon. Three milliliters of concentrated hydrofluoric acid and 5 ml. of 1 to 1 sulfuric acid are added, and the solution is heated t o fumes of sulfur trioxide until the volume is about 1.5 ml. (Tungsten and molybdenum interfere with the subsequent colorimetric determinations and are removed if present.) The contents of the
cooled crucible are washed into the original beaker containing 5 ml. of boric acid (4%) by a stream of hot hydrochloric acid (2y0), using about 100 ml. Fifty milliliters of saturated sulfurous acid is added, and the solution is boiled 10 minutes. Paper pulp is added, and after standing 15 minutes on the steam bath, the solution is filtered through Xo. 40 Whatman paper and washed 15 times with hot hydrochloric acid (2%). The paper and precipitate are ignited a t a temperature of 1000" to 1050" C. The melt is fused with potassium bisulfate and dissolved in saturated ammonium oxalate solution. The niobium is determined colorimetrically in a portion of the solution, after adding dehydrated phosphoric and sulfuric acids and hydrogen peroxide (30%). The tantalum is determined colorimetrically on another portion, after adding phosphoric and pyrogallic acids. APPLICATION O F RADIOISOTOPE TECHNIQUES
Niobium Loss in First Hydrolysis and Precipitation. The steps of the operations described are shown in Figures 1 and 2. Triplicate 2-gram samples of Bureau of Standards sample 1234 (containing 0.75% niobium and 0.02% tantalum) were dissolved and taken to fumes as in the analytical procedure. Two tenths milliliter of a solution of niobium sulfate, containing niobium-95 (V.S. Atomic Energy Commission No. 41F), equivalent to 1 mg. of niobium with an activity of approximately 0.06 microcurie was added, The niobium sulfate solution was made by fusing niobium oxide (NbzOs)with a small amount of potassium bisul-
