1214
ANALYTICAL CHEMISTRY
(13) (14) (15) (16) (17) (18) (19) (20)
Dombrowski, A., Mikrochemie, 28, 136 (1940). Dubsky, J. V., Chem. Ztg., 40, 201-3 (1916). Elek, A., IND.ENG.CHEM.,ANAL.ED.,20, 51 (1948). Elving, P. J., and McElroy, W.R., Ibid., 13, 660 (1941). Emich, F., Monatsh., 13, 76 (1892); 15, 378 (1894). Epstein, F., 2. anal. Chem., 46, 771 (1907). Heron, A. E., A n a l y s t , 72, 142 (1947). Hornung, E. C., and Hornung, 11. G., .i.v.ir.. CHEM.,19, 688
(21) (22) (23) (24)
Ingram, G., A n a l y s t , 73, 548 (1948). Israelstam, S. S.,AXAL.Cmni., 24, 1207 (1952). Kainz, G., Mikrochemie, 35, 569 (1950). Kemmerer, G., and Hallett, L. T., I n d . Eng. Chem., 19, 173
(1947).
(1927). (25) Kirner, W. R., IND.ESG. CHEM..ANAL.ED., 7, 366 (1935). (26) Lacourt, d.,Chang, C. T., and Vervoor, R., B u l l . SOC. chim. Belges, 50,67 (1941). (27) Lindner, J.,Ber., 59,2561 (1926). (28) Ibid., 65, 1702 (1932). (29) Lindner, J., “hfikro Massanalytische Bestimmingen des Kohlenstoffes und Wasserstoffes,” Berlin, Vetlag Chemie, 1935.
(30) Millin, D., J . Soc. Chem. I n d . ( L o n d o n ) ,58, 215 (1939). (31) Muller, von E., and Barck, H., Z . anorg. Chem., 129, 310 (1929). (32) Xeumann, B., Panzner, H., and Goebel, E., 2. Elektrochem., 34,703 (1928). (33) Iiiederl, J. B., and Whitman, B., Mikrochemie, 11, 274 (1932). (34) Roger, R., and Mackay, W. B., J . SOC.Chem. I n d . ( L o n d o n ) , 5 4 , 4 6 (1935). (35) Sabatiet, P., and Senderens, J. B., Compt. rend., 114, 1429-76 (1892); Bull. SOC. chim., 7, No. 3, 502 (1892); Ann. chim. et phys., 7,348 (1896). (36) Steacie, E. W. R., and Johnson, F. 11. G., Proc. R o y . SOC. ( L o n d o n ) , A112,542 (1926). (37) Stragand, G. L., and Safford, H. W., A ~ ~ A ICHEM., ,. 21, 625 (1949). (38) Titov, E. AI., Zavodskaya Lab., 9,853 (1940). (39) Wagman, D. D., and Rossini, F. D., J . EeseamhNatl. BUT.Standards, 3 2 , 9 8 (1944). (40) White, E. T., and Wright, G. F., Can. J . Research, 14B, 427 (1936). R E C E I V E Dfor review June 26, 1951. Accepted April 16, 1932.
Infrared Absorption Spectra of Porphyrins CHARLES W.CRIVEN’, KURT R. REISSIIIANW, ~ N HERMAN D I. CHI“ U S d F School of Aviation Medicine, Randolph Field, Tex. THILE hemoglobin metabolism during acclimatization t o altitude was being studied, changes in the excretion of porphyrin were observed ( 5 ) . The usual chemical processes for isolation and identification are laborious and time-consuming.
sorption spectrum of various porphyrins might give additional information on the chemical structure of these compounds. EXPERI.MENTA L
The absorption spectra were determined with a Baird doublebeam instrument equipped with a sodium chloride prism and sample cell. T h e cell thickness was 0.1 mm. and carbon tetrachloride was the solvent used throughout. Crine was obtained from the bile fistula of dogs during altitude acclimatization. I n these animals the bile duct emptied directly into the ureter, so that pigment loss through the feces was avoided. Extraction and partition according t o the procedure of Dobriner ( 2 ) gave the following fractions: amisture of bile pigments and protoporphyrin, deuteroporphyiin, and a mixture of coproporphyrins I and 111.
: Y
I
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,
180-
170-
-
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160-
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8 150(3 I!
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-
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P..C-C-COO*
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30
40
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WAVE LENGTH, MICRONS
Figure 1. Composite Curves for S o m e Metabolic Products of Respiratory Pigments Furthermore, losses of pigment are difficult t o avoid ( 8 ) . The utility of infrared spectrophotometry in the analysis of mixtures of closely related compounds suggested that such a n approach might be profitable. Also, i t \vas hoped that the infrared ab-
* Present address, Department of Physiological Chemistry. Ohio State University, Columbus, Ohio. 2 Present address, Department of Medicine, rniversity of Kansas Medical Scbool, Kansas City, K a n .
Purified preparations of bilirubin (Eastman Kodak Co., recrystallized), coproporphyrin I (generously supplied by Samuel Schwartz, department of medicine, Gniversity of Minnesota), and coproporphyrin I11 (generously supplied by C. H. Gray, biochemistry department, King’s College Hospital, London), were also analyzed. The absorption spectra and the structural formulas according to Dobriner and Rhoads ( 3 ) are shown in Figure 1. As can be seen, the most prominent absorption band in each case is evident between 3.35 and 3.45 microns, with secondary bands in each case at a slightly longer wave length (3.45t o 3.60 microns) and, again, a t 6.8 microns. These bands are probably attributable to the methyl groups which were present in all compounds stud-
V O L U M E 24, NO. 7, J U L Y 1 9 5 2 ied. The 3.4-micron band is characteristic of carbon-hydrogcn stretching in alkane groups and the 6.8-micron band of carbonhydrogen bending ( 1 ) . This latter band is also characteristic of the pyrrole nucleus which is present, in each preparation dcscribed. Strong absorption was also noted between 5.7 and 5.8 microns and a t 8.5 microns a i t , h every preparation except the deuteroporphyrin extract. Bilirubin showed characteristic bands a t 7.9 and 10.7 microns, not found in any other preparation. These bands may be related t o the unfolding of the molecule in bilirubin. S o differentiation could be detect,ed betn-een coproporphyrins I and 111. Varying amounts of coproporphyrin I11 !yere dissolved in carbon tetrachloride and the transmittance was determined a t 5.72 microns. .it least for the range of investigation, a 3traightline relationship exists between concentration and the 1op:iiithm of transmittance (Figure 2 ) .
1215 Gray et al. ( 4 )determined the infrared absorption curves for the methyl esters of coproporphl-rins I and I11 in the 7.8- to 11.4micron range (using the S u j o l technique) and reported a difference between the two spectra. However, attempts in this laboratory t o differentiate pure preparations of coproporphyrins I and I11 by infrared spectrophotometry proved unsuccessful. The failure of deuteroporphj-rin to absorb strongly in the 5 . i and 8.5-micron range may he of value in identifying this pigment. LITERATURE CITED
(1) (2) (3) (4)
Coltrup, N. B., J . Optical SOC.Bm., 40, 307 (1950). Dobriner, K., J . B i d . C h e m . , 113, 1 (1936). Dobriner, K., and Rhoads, C. P., Phv8iol. Rcc., 20, 416 (1940). Gray, C. H., Seuherger, -L,and Sneath, P. H. A , , Biochem. J . , 47,
87 (1950). ( 5 ) Reissmann, E(. R., and Chinn, H. I., unpublished data.
RECEIVED for review S u g u s t 13, 1951. Accepted March
11, 1952.
High-Alkaline Colorimetric pH Determination ROBERT H. M. SIMON General Engineering Laboratory, General Electric Co., Schenectady, N. Y .
El O R
detection of changes in p H in the high-alkaline range where most p H meters are not wholly reliable, a colorimetric method may advantageously be used. This method employs a n indicator dye in which the change in light transmittance TTith p H over a portion of the visible spectrum is great enough t o permit calibration in p H units when properly incorporated n i t h a suitable light-sensitive device. The p H range about 12.5 wis selected for test purposes because i t is of particular interest to the tanning industry. Several dyes which changed color in this range were prepared in aqueous solution. These included Acyl Blue and Parazo Orange (IT’. A. Taylor and Co.), 0.5% Poirrier’s Blue and 0.1% Tropacolin-0
(&illliedChemical 6: Dye Corp.), and saturated sodium indiyodisulfonate (practical grade, Eastman Kodak Co.). Parazo Orange was used in the commercial solution. The commercial solution of Acyl Blue was concentrated tenfold by evaporation of the solvent. Tumerous dyes are hnonn t o change color about p H 12.5. The above dyes were selected because their color changes orcur over a comparatively narrow p H band, and because they appeared, from initial tests, to be soluble and free of reactions which 7 form precipitates with the solutions to be used. L Four test solutions were prepared and measured with a p H meter (Beckman &lode1 11 with high-pH sealed glass and calomel
WAVELENGTH I N MILLIMICRONS
Figure 1.
Light Transmittance Change with pH of Parazo Orange
WAVELENGTH I N MILLIMICRONS
Figure 2.
Light Transmittance Change with pH of Tropaeolin-O
