Snectronhotometric Determination of Diacetvl I
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I
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JOHN C. SPECK, JR. Kedaie Chemical Laboratory, Michigan State College, E n s t Lansing, M i c h .
With chroiiiotropic acid in sulfuric acid diacetj 1 gives a purple dye which is nearly indistinguishable from that obtained with formaldehyde under the same conditions. The formation of this color, which is attributed to the production of a mole of forrnaldeh>deby the sulfuric acid oxidation of a niole of diacetyl, has been utilized as the basis of a rapid spectrophotometric determination of diacetyl.
T
HE possibility of a spectropliotoliietric niet,hod of analysis for small concentrations of formaldehyde, based on the purple dye formed from this compound and chromotropic acid (4,5-dihydroxynaphthalene-2,7-disulfonic acid), was realized in the procedures developed by XacFayden (6) and Bricker (3). A4sshown in these investigations, and by the original qualitative observations of Eegriwe ( 3 ) , this rtwtion of formaldehyde nith chromotropic acid is highly specific. Its ana,lytical application is also subject to few interferences.
in order t o achieve maximum extinctions. Tlie developnic~nt procedure for t'he formaldehyde dye described by MacFayden, which eniploys a Ion-clr .wlfut.ic acitl concrntrat ion, gives loncr extinct.ion values.
1
A Beckman spectrophotometer (model DU) and 1-cm. Corex cells were used in obtaining all extinction data. The diacetyl used in these experiments was obtained from Eastman Kodak Co., Paragon Testing Laboratories, and Forest Products Chemical Co. The usual procedure for i b purification involved two distillations through a 45-cm. (18-inch) Stedman column, the middle fraction boiling a t 87.5-88' being taken each time. Technical chromotropic acid was obtained from Eastman Kodak Co. and Paragon Testing Laboratories. It was not purified, but the solutions were merely filtered before use. The reducing sugars which were examined as possible interfering substances were obtainrd from the Pfanstiehl Chemical CO. and were C.P. grade. For comparison of the absorption characteristics of the formaldehyde dye with that resulting from diacetyl, a weighed amount of freshly resublimed hexamethylenetetramine W&S hydrolyzed with 2 N sulfuric acid, as recommended by ?*lacFayden ( 6 ) , and the color development carried out according to Bricker's procedure. Thp effect of time of development of the color is shown in Table I. Thiq \vas detcrininetl by mairitairiirig the conditions of the above procodurc c*onst,ant ivith r h e pswpfion o f t inic. of heating in the boiling v-awr bath. The markrd c'ffcct of t tic sulfuric: acitl c o n w i l t ration, q l i n k i i in Table TI, was dc~tcvniinetisitnilarly.
PROCEDURE
To 1.0 ml.of solution containing 0.03 to 0.1 nig. of diacetyl per ml. in a test tube provided with a glass stopper is added 0.5 ml. of 10% chromotropic acid solution, and to this mixture are added gradually 5 ml. of 98% sulfuric acid. After mixing, the tube is heated in a water bath a t 100 O for 1 hour and cooled, and the contents are transferred to a 50-ml. volumet,ric flask and diluted t o the mark with distilled water. The extinction is then determined a t 570 mp; a blank containing all the reagents and treated in the same manner is employed as the reference solution. The size of the test tube used in the development of the color is not important. However, the level of the liquid in the tube should be below that of the water in the bath throughout, the period of heating. EXPERIMENTAL
I 350
I
400
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45 0
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500
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550
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600
WAVELENGTH, m u Figure 1. Absorption Characteristics
--dye (from 1.16 - - - -Diacetyl Formaldehyde dye (from tion)
X 10-6 M diacetyl molution) 1.13 X 10-6 Mformaldehyde molu-
Howwer, it has been observed in this laboratory that solut,ions of pure diacetyl give a color with chromotropic acid-sulfuric acid mixture, the absorption characteristics of which are nearly identical with those of the formaldehyde dye, as is shown in Figure 1. Moreover, the molar extinctions of the dyes, as based, respectively, upon moles of diacetyl and formaldehyde, are almost equivalent. These observations, other than implying a source of serious error in the cliromotropic acid method applied to certain formaldeh?-de estimations, have indicated a basis for a rapid spectrophotometric estimation of diacetyl. Accordingl>-,the analytical possibilities of the diacetyl reaction have been investigated. JIost of the previous nictliods iur dt,terniining diacetyl depend upon the formation of the diosinie arid its precipitation as the riic.l- as the riickelic diiiiethylglyoxime complex ( I d ) . Diacet-1 has also been determiried cqlorimetrically by means of the ferric dimethylglyoxime coniplcx (9) and as substituted quinosaliiies formed by its reaction with o-phenylenediaminps (8). .4 polarographic method Iiaa ? x ~ ndeveloped by Fulmcr et (11. ( 4 ) . In a preliminary application of tlw chromotropic acid-diacet.1 reaction t o the quantitative estimation of diacetgl, it was found that the procedure described by Rricker for formaldehyde gave excellent results with little alteration, and that the relationship of extinction (log l o / Z ) to concentration was nearly linear over the range of 0.01 to 0.1 mg. of diacetyl per milliliter of solution analyzed. It was merely necessary to increase the time of heating
Table I.
Effect of Time of Development
Time of Developinent, h h . 15
30 GO
Extinction (log Io/I) a t 570 mfi 0.133 0.146 0.148
Table 11. Effect of Sulfuric Acid Concentration Concentration of Sulfuric Acid Added i n Cglor Development, 98 80
GO
647
Extinction (log I o / I ) a t 570 ma 0.292 0.242 0.006.
ANALYTICAL CHEMISTRY
648 For the comparison of this proctdure with a standard nictliod, cliacetyl was precipitated from solution as the nickelous dimethylglyoxime complex; the precipitation conditions recommended by Wilson ( I S ) \Yere employed and tlie precipitates were xveighed. These gravimetric analyses were carried out on 10.0-ml. aliquots from t,he original diacetyl solutions, which were approximately 2%,diaeetyl. The spectrophotometric anal on solutions prepared by dilution of the original solutions either I to 25, 1 to 50, or 1 to 100. The values given in Table I11 correspond to the concentratioiis of t,he more dilute solutions. Those determinations designated by the subscript refer to analyses which \yere repeated after the original solutions had stood at, room temperature for one xveek. Obviously, the present method gives niucli better agreement with the values obtainetl with freshly prepared solutions than does the precipitation procedure. This, probably, is due to a nonprecipitation of polymerized diaeetyl as the nickelous complex, whereas some of the polymerized material is attacked by the chromotropic acidsulfuric acid mixture. The proposed method should give high results, therefore, in eases where some polymerization of the diacetyl has occurred. Although’the data in Table 111 indicate that the precision of the method is sonieivhat outside the limits imposed hy the spectrophot’ometer, the reproducibility of results obtained in the determination of the relationship of extinction to concentration was nearly that t o be expected of the iristrument over the range of extinctions covered. This precision is demonstrated in Table IV, in which are given the analyses oi solutioris of highly purifiid Forest Products’ diacetyl by means of a standard curve ohtairitd with highly purificd I’ai,agori Twting T,ahoratnrirs’ tliaret,yl.
Talde 11I. Iktermiiiation of lliacetyl Determination
Foluld Gravirnetricall\ M g . ’ml.
Pound Spectrophotornetricali \ .Wg./ml.
Table IV.
Precision Found Syectrophotomatricail!. nrg./l\n.
0.0192 0.0481 0.0871
0.020 0.047 0.08i
Table V.
Extinctions Extinction a t 570( rns lor K O / I I
Keducing Sugar Galactose
0.266
0,49!4
Glllcose .
.
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RCOCOR
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+
[O]
+
HZO
.. .
.
+ 2ItC02H
(2)
agents, it would hardly be espected to occw at a detectahlr rate in solutions in which the hydroxyl ion conrriitration is as low a:: that provided by concentrated sulfuric acid, iis it is probable t,hat this type of oxidation proceeds through the m-diketonc hydrate. ..\tteinpts to isolattl pyruvic acid, the other product iiitlicateti in Equation 1, from the risaction of sulfuric acid with diacetyl have nnt b t m successful, possihly because a t reasonable conccmtratioiis of diacetyl in sulfuric acid extensive polymerization and clec~c~iiipo~itiori of this substanre orc~ir, INTERFERENCES
.I11 t l i c s mhstancm that intilifere in thc forinaldehydr tltatt*riiiiiiatiori by the cliroinntropic acid method interfere in this tliacetyl estimation-for example, thr presence of methyl rthyl ketone produces a marked diminut,ion of the color developrd with diaretyl. Formaldehyde is itself an interference. Both hexoses and pentoses yield diacetyl as one of the products of acid rlcgrdation (7, li), and these substances give brown or purple rolors n-ith chromotropic acid-sulfuric acid mixture (3). Data indicating the extent of this reartion for tn-o 1im)sw are given in Table V. It is noteworthy that acetyl propioiivl, tlir. ncsst highcbr homolog of diacctyl. gives no purple color with cahromotropir acid ACKNOWLEDGMENT
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In order to ohtaim surli reproducibility betwcbu separate lots of diacetyl, it was necessary t,o perform rectifications with great care and to protect the purified material against moisture as well as to use it within a short time after distillation. Otherwise, low results were always obtained. The data shown in Table V, which were obtained with reduring hexoses in order to demonstrate their interference, are thr ext’inctions found upon treating 0.01 M solutions of the sugars as described in the above procedure. The color obtained with galactose was brown; however, the glucose color mas reddish purple, resembling very much the formaldehyde color. DISCUSSION
It must be concluded from the near-congruency of their spertra that the dyes produced from diacetyl and formaldehyde are very probably of the same structure. Moreover, the most logical explanation for this structural ideritit,y is that, under thest: conditions, one mole of diacvtyl is oxidized by the sulfuric acid with t8he production of one mole of formaldehyde, as is shown iii Equation 1. This is borne out by the extinction data and by the fact that non-oxidizing acids produce relatively little effcct. Although the “normal” oxidation of an aliphatic a-dikctone (Equation 2) proceeds with great facility with most osidizing
CH2=CCOCHc I’
bH
+ 2[0] +CHzO + CHsCOCOsH
(1)
The autlior wishes to txspres5 his ap,pl,t,ciation for helpful suggestions by C. D. Ball and F. R. Thkr. LITERATURE CITED
(1) Barnicoat, C. R., Analyst, 60, 653 (1934). (2) Bricker, C. E., and Johnson, H. R., INDENG.CHENI.,4 x 4 ~ .
ED.,17,400 (1945). (3) Eegriwe, E., 2. anal. Chem., 110, 22 (1937). (4) Fulmer, E., Kolfenbach, J . J., and Underkofler, L. A., IND. ENG. CHEM.,ANAL.ED.,16, 469 (1944). ( 5 ) Kunae, R., Mikrochemie, Festschr. von Hans Molisch, 1936,279. (6) MacFayden, D. M., J . B i d . Chtm., 158, 107 (1945). (7) Nodau, R., and bfatsui, K., Bull. Chem. SOC.Japau, 10, 4h7 (1935). (8) Pien. J., B a k e , J.. and Martin, R., Lait, 16, 119, 243 (1936); 17, 675 (1937). (9) Prill, E. X.,and Haminer, B. W., Zowa State Coll. J . Sci., 12, 385 (1938). (10) Schmallfuss, H., and Rethorn, H., 2 . I’ritefsuch. Lebensm. 70, 233 (1935). (11) Speck, J. C., unpublished work. (12) Y t o t z , E., and Raborg, J . , J . B i d . Chem., 150, 25 (1943). (13) Wilson, J. B., J . Assoc. Oficial Agr. C h e m . . 24, 655 (1941). R E C E I V E D Xovernber 19, 1947. The subject matter of this paper has been undertaken in cooperation with t h e Committee on Food Research of the Quartermaster Food and Container Institute for the .Irmed Forces. T h e opinions or conclusions contained in this report are those of t h e author. They are not t o be construed a s necessarily reflecting the views or endorsement of the K a r Department.
