Retention time of this myrcene peak was identical with that of linalool, since the dehydration occurred a t the end of the column. If the boric acid section had been placed a t the front of the column, the dehydration would have occurred a t the beginning of the run and the retention time would have corresponded to myrcene. Therefore, glacernent of the boric acid section apparently is critical, particularly with compounds prone to dehydration. Whether tertiary alcohols will be retained or dehydrated will depend on the eabe of dehydration. The tertbutyl alcohol mas retained on the column, while the easily dehydrated linalool emerged as myrcene. Linalool can be dehydrated during gas chromatographic analysis if the support is acidic ( 2 ) . Tertiary alcohols do not
form triborate esters (3). Since the tert-butyl alcohol was retained by the boric acid column, one can speculate that the mono- or diborate ester was formed. To show that a boric acid-treated column can be useful, a sample of peppermint oil was analyzed by gas chromatography with and without the boric acid section. The chromatograms are shown in Figure 2. The upper chromatogram was made without the boric acid section. The major peak, corresponding to the retention time of menthol, has been completely removed by the column treated with boric acid. Also, the shoulder on the peak a t about 9 minutes, which was probably caused by neomenthol, was removed. Smith and Levi (4) identified an alcohol, 3-octanol, in peppermint oil; this may
account for the peak which disappears a t approximately 4 minutes. LITERATURE CITED
(1) Guenther, E., "The Essential Oils," Vol. 11, p. 793, Van Kostrand, New
York, 1949.
( 2 ) Mitzner, B., ANAL.CHEM. 36, 242
(1964). (3) Scattergood, A., Miller, W. H., Gammon, J., Jr., J . A m . Chetn. SOC. 67,2150 (1945). ( 4 ) Smith, D. M . , Levi, L., J . A g r . Food Chem. 9, 230 (1961).
Philip Morris, Inc. P. 0. Box 3-D Richmond 6, Va.
ROBERTM. IKEDA DONALD E. SIMMONS JAMES D. GROSSMAN
Division of Agricultural and Food Chemistry, 145th Meeting, ACS, New York, September 1963.
Quantitative Determination of 0 - and p-Dihydric Phenols in Presence of Monohydric Phenols by Use of a Phosphotungstic Acid Reagent SIR: All dihydric phenols and most monohydric phenols are oxidized by the Folin-Ciocalteu phenol reagent ( I I ) . To simplify product analysis, initial investigations on the mechanism of the Elbs peroxydisulfate oxidation ( 4 ) were conducted on two phenols which do not reduce this reagent. In the Elbs oxidation, hydroquinone and catechol monosulfates are produred. Their hydrolysis products, the free dihydric phenols, are, of course, determinable with the FolinCiocalteu reagent. During further studies on the Elbs oxidation in which phenols of widely varying structure were used (a), it became desirable to develop a general and simple method for the quantitative determination of 0- and p-dihydric phenols in the presence of a large excess of monohydric phenol. The phosphomolybdic acid reagent is reduced by all except highly deactivated monohydric phenols and was hence useless. A suitable reagent was found, however, in the phosphotungstic acid reagent of Folin and Denis (12). This reagent was originally developed for the determination of uric acid. However, in qualitative tests of the scope of the reagent, Fohn and Denis found that, in addition to uric acid, a positive reaction was given by 0- and p-dihydric phenols and p-aminophenols. There was no reaction with most monohydric phenols. Since 1912, the reagent has been used solely as a uric acid reagent and as such has undergone many modifications ( 5 , 6, 9 , 13, 1 5 ) .
h st'udy of reagents for the qualitative analysis of phenols has appeared recently (18). d number of t,he reagents described appear suitable for qualitative detection of 0- and p-dihydric phenols. However, the phosphotungstic acid reagent appeared most convenient for quantitative work. EXPERIMENTAL
Reagents. Phenols were purchased from the -4ldrich Chemical Co., Eastman, or Matheson Coleman & Bell, and were redistilled or recrystallized before use. 2,5-Dihydroxypyridine was synthesized by the persulfate oxidation of 2-hydroxypyridine (3). 3,4-Dihydroxypyridine was synthesized by the persulfate oxidation of 4-hydroxypyridine (16) and gave the following data. Crystals from water, m.p. 239.5" to 240.5' C. [lit., 239.5" to 240" C . 2eamlr ( 7 ) ] , e*:-%' 11,750; e,,, HCI 5950; 242mg eo I N HCI 3950. Ferric chloride complex, 535rnF HCI 1480 [compare (S)]. .\rial. calcd. for CsH5N02: C, 54.05; H, 4.54; N, 12.6. Found: C, 54.0; H, 4.5; N, 12.4. The diacetyl derivative was prepared and recrystallized from ethyl acetate, m.p. 137" to 137.5" C. [lit., 138.5" to 140" C. (7)l. 2,3Pyridinediol was prepared by alkali fusion of 3-hydroxypyridine (1 4 ) . Kudernatsch reported formation of the 2,5-isomer. The major product is the 2,3-isomer ( 1 7 ) ,although small amounts of the 3,4-isomer were detected on paper chromatograms of the mother liquors. The phosphotungstic acid reagent was prepared according to the procedure of Folin (IO). Other reagents were
sodium carbonate solution saturated at 24' C., 20y0 urea solution, and 120/, sodium cyanide solution ( 8 ) . General Procedure. An appropriate sample of t h e phenol was made u p t o 1 . O ml. with water. Five milliliters of water or 20% urea were added, followed b y 1.0 ml. of the phosphotungstic acid reagent. Color was developed b y t h e addition of 3.0 ml. of either sodium carbonate or sodium cyanide solution. The alkaline solution was mixed a t once and the absorbance determined as a function of time in a Klett-Summerson colorimeter with the 66O-mp filter. When cyanide was used, the absorbance was read after 30 to 40 minutes. The absorbance, ;1, is expressed in Klett units, K ; 5OOK, = A. A l l l operations were carried out a t room temperature (24" C. + 3"). Procedure for Yield Determination in Elbs Peroxydisulfate Oxidation. Each phenol, 0.005 mole, was dissolved in 25 ml. of 1.74X KOH. Five milliliters of 0.10051 K2S2OS were added and the volume was made up to 50 ml. with additional KOH. The reaction mixtures were allowed to stand a t room temperature until persulfate was no longer detectable ( 4 ) . One-milliliter aliquots of the reaction mixtures were removed and heated in a water bath at 100" C. for 30 minutes with 1.0 ml. of 6 S HC1. The hydrolysates were cooled and then made up to 12.5 or 25 ml. with water. Onemilliliter aliquots were analyzed using the procedure already de,wrihed. The high concentration of KC1 in these solutions resulted in the a1)l)earanre of a precipitate approximately 3 minutes VOL. 36, NO. 1 1 , OCTOBER 1964
2189
300r
I
’
1
I
2 3
I
3
4
I.
‘I
I
I
I
t
I
followed by first-order decay. The recorded figures represent extrapolated zero-time readings for the first class and steady-state readings for the second class. The decay behavior for three representative phenols is shown in Figure 1. Comparison of Results under Different Assay Conditions. Virtually identical results were obtained (Tahle I) in t h e presence or absence of urea. T h e addition of urea prolongs the time during which meaninqful readings may be taken by delaying the time of appearance of a precipitate. The use of sodium cyanide in place of sodium carbonate as base has three effects: blue color develops more slowly, it decays more sloa ly, and absorptivity is increased. These advantages, however, must be weighed against the inconvenienceq involved in using concentrated solutions of this highly poisonous material. I n addition, the intensification of color given by cyanide is greater for the 2,5-isomer than for the 3,4-isomer. This is a disadvantage for thp assay of mixtures of these isomerq. The sodium carbonateurea method gives more nearly equiyalent color yields for the t n o isomers. Use of sodium carbonate is most generally convenient in the presence of urea. However, the decay curveq of the color produced by gentisic acid @,5dihydroxybenzoic acid) did not follow simple first-order kinetics. For determination of this compound, the cyanide method was used. Beer’s law was obeyed for all three assay methods up to about K , = 600. Relationship between Color Yield and Structure. The following generalizations emerge from the results. Only highly activated monohydric phenols reduce the reagent under the conditions described. Among those
,
5 6 7 8 9 1 0 1 1 12 MINUTES
Figure 1 . Determination of color yield for three phenols b y the sodium carbonate procedure showing differences in the decay behavior of the color produced A. 0.2 p o l e of 2,5-pyridinediol. grnole of 3,4-dihydroxynitrobenzene. and 0.4 pnole of p-dihydroxybenzene
B. 0.2 C.
0.2
after sodium carbonate was added if the assay was conducted without urea. RESULTS AND DISCUSSION
Quantitative. Quantitative assays i n the absence of cyanide ions were complicated by the more or less rapid decay of the blue color with time. This point, however, presented no real difficulty since the decay in general followed first-order kinetics. Two classes of phenols were recognized according to the decay behavior of the blue color. For the first class, first-order decay began as soon as readings could be taken (within 20 seconds). For the other class, the absorbance was constant for some time,
Table I.
Color Yield for 0 . 2 pMole of Various 0 - and p-Dihydric Phenols as Function of Structure of Phenol
Numbers are given in K , units for 0 . 2 pmole of each phenol. Precision is estimated at *37i Method Compound Na2COS ?;azCOa urea S a C N urea 225 250 205 2,3-Dihydroxypyridine 225 -22,50 230 2,5-Dihydroxypyridine
+
3,4-I)ihydrox
2,3-Dihydroxgz?$racid 2,5-Dihydroxybenzoic acid 3,4-Dihydroxybenzoic acid 2,s-Dihydroxynitrobenzene 3,4-Dihydroxynitrobenzene 2,5-Dihydroxyhenzaldehyde 3,4-TXhydroxybenzaldehyde
2,5-Dihydroxychloro benzene 3,4-Dihydroxychlorobenzene
p-Dihydroxpbenzene o-Dihvdroxybenzene 2.5-Dihydroxytoluene 3.4-Dihydroxytoluene a ?;onlinear plots.
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ANALYTICAL CHEMISTRY
, . .
...
4 0 ...
230 140 200 210 130 120 100 155 90 125
+
-50a
-140”
-80
800
203 170
375
220
270 360 210 800
195 148
452 720
100 144-
350
190 180 146 116 186
416 690 370 690
- 0 4 -02
0.0
0 2 0 4 06 06
Vd Figure 2. Color yield for 0.2 pmole of five monosubstituted p-dihydric phenols as a function of the Hammett un+ constants. K, values determined b y the sodium carbonate-urea method Substituents are: 4.-H; 5.-CHa
1 .-NOp;
2.-CHO;
3.-CI;
giving a positive qualitative test are the pfollowing: o-methoxyphenol; methoxyphenol; 2,4,6-trimethylphenol; 2,6-di-t-butyl-4-methylphenol;2.6-dit-butylphenol; 2,6-dimethylphenol. The molar color yield is, however, 1017. For p-methoxyphenol, for example, K,O.* ! ~ m o l s = 10 (sodium carbonate method). -111 p - and o-dihydric phenols teqted reduce the reagent. m-Dihydric phenols do not. Within the group of monosubstituted p-dihydric phenols, there is a rough correlation between the Hammett substituent constant up+ (1.9) and the molar color yield when sodium carbonate is used as the dewloping agent (Figure 2). !Yo similarly clear correlation emerges from the data for the class of o-dihydric phenols examined. The factors determining this relationship presumably involve the stability of the reagent in alkali, the stability of the dihydric phenol in alkali, the stability of the reduced reagent in alkali (which is, in turn, a function of the nature of the phenol in question), and the rate of reaction of the reagent with the phenol. Application of Method to Determination of Yield in Elbs Peroxydisulfate Oxidation. Second-order rate constants have been reported for the reaction of a number of monosubstituted phenols with persulfate ( 2 ) . Yield of product had been determined. however, only for two of these: 2-hydroxypyridine and o-nitrophenol. The significance of the rate conbtants for the remaining phenols was, therefore, in doubt.
The present method was used to determine the yield of dihydric phenol from the reaction of persulfate ions with a series of monosubstituted phenols representative of those used in a previous qtudy ( 2 ) . A tenfold excess of phenol was used under the conditions previously dewribed ( 2 ) . The o- and m-isomers of the nitro-, halo-, and alkylphenols gave yields of dihydric phenol in the 60 to 75y0 range. The yield of dihydric phenol from the p isomers was, however, only in the 15 to 30% range. The yield of dihydric phenol from p-substituted phenols is too low to allow conclusions derived from their kinetic behavior in the persulfate oxidation to be used with much assurance. The yields determined here are in agreement with the low isolated yields reported for a number of p subbtituted phenols ( I ) . Some other reaction must he taking place in these cases. Preliminary experiments with polychlorophenols have shown substantial formation of chloride ions during the reaction with persulfate.
The other products have not, however, been characterized. ACKNOWLEDGMENT
The authors thank Carol K. Fritz, who made the analyses. LITERATURE CITED
(1) Baker, W., Brown, N. C., J . Chem. SOC.1948, 2303. (2) Behrman, E . J., J . Am. Chem. SOC. 85, 3478 (1963). (3) Behrman, E. J., Pitt, B. M., Ibid., 80. 3717 11958). (4) Behrman, E.'J., Walker, P . P., Ibid., 84, 3454 (1962). ( 5 ) Benedict, S. R., J . B i d . Chem. 51, 187 (1922). ( 6 ) Benedict. S. R.. Hitchcock. E . H.. Zbid.. 20. 619 11916). (7) Bidkel,' A. F . , J . Am. Chem. Soc. 69, 1805 (1947). (8) Brown, H., J . Bwl. Chem. 158, 601 (1945). (9) Folin, O., Ibid., 86, 179 (1930). (10) Zbid.. 106. 311 (1934). ( 11) Folin, O., 'Ciocalteu, V., Zbid., 73, 627 (1927). (12) Folin, O., Denis, W., Ibid., 12, 239 (1912). ~
13) Jackson, H., Jr., Palmer, W. W., Ibid., 50, 90 (1922). 14) Kudernatsch,. R.,. Monatsh. 18. 613 (1897). 15) Morris, J. L., Macleod, A. G., J . B i d . Chem. 50, 55 (1922). 16) Pitt, B. M., Behrman, E. J., Cancer Research Institute, Sew England Deaconess Hospital, Boston, Mass., unpublished data, 1958. (17) Shickh, 0. von, Binz, A , , Schulz, A,, Her. 69, 2593 (1936). (18) Smith, B., Chalniers Tek. Hogskol. Handl., .\a 263 (1963). (19) Stock, L. ?VI., Brown, H. C., A d v . Phys. Org. Chem. 1, 35 (1963). E . J. B E H R Y A K ~ 31.X . D . GO SWAMI^ Department of Biological Chemistry Harvard Medical School and Cancer Research Institute New England Deaconess Hospital Boston, Mass. 02215 This investigation was supported by U. S. Public Health Service Grant A567 and by U. S. Atomic Energy Commission Contract AT(30-1)-901 with the Sew England Deaconess Hospital ( S Y O 901-3). 1 Present address, Dept. of Chemistry, Brown University, Providence, R.I. Present address, Laboratoire de Chimie Biologique, 96 Blvd. Raspail, Paris 6, France.
Titrimetric Determination of Bis(disu bstitutedphosphiny1)alkanes and (Trisubstituted phosphine) Oxides SIR: Phosphoryl-containing compounds are widely used as solvent extractants. I n spite of this, only one general method has been reported for the quantitative determination of the phosphoryl group. Wimer (6,3 ) has reported thp potentiometric titration of trimethy1phoq)hine oxide, trioctylphosphine oxide, and triphenylphosphine oxide, using perchloric acid in dioxane as the titrant and acetic anhydride as the solvent. I n the present work, the general applicability of this type of titration for the determination of a number of different phosphorylcontaining compounds was tested. EXPERIMENTAL
Reagents. A 0 . 1 s titrant was prepared by disqolving 9 ml. of 70% perchloric acid in glacial acetic acid, adding 25 ml. of acetic anhydride, and diluting to 1 liter with glacial acetic acid. T h e solution mas allowed to i t a n d for 24 hours prior to use so t h a t t h e acetic anhydride would have time to remove the water present. Perchloric acid in dioxane can also be used as the titrant. However, reagent grade dioxane requires purification before it can be used a' the diluent. The titrant n as standardized potentiometrically against primary standard pota-sium acid phthalate dissolved in glacial acetic acid.
Apparatus. The titrations were carried out using a Beckman Model G pH meter with calomel and glass electrodes. T h e aqueous bridge in a sleeve-type calomel electrode was replaced with a saturated solution of anhydrous lithium perchlorate in acetic anhydride. A fiber-type calomel electrode with a n aqueous salt
bridge can be used but the potential readings are somewhat unstable. Procedure and Results. Samples (0.5 to 2 mmoles) were dissolved in 100 ml. of acetic anhydride, stirred with a magnetic stirrer, and titrated with 0.1S perchloric acid in a glacial acetic acid-acetic anhydride mixture. Three basic types of phosphoryl com-
Table I. Titration of Phosphoryl Compounds" Compound % Purity 1. Tri-n-octyl hosphine oxide 92.8 100.0 2. Tris( 2-ethy~hexy1)phosphineoxide 3. Di-2-ethylhexylmethylphosphineoxide 8t.3 4. Triphenylphosphine oxide c 5. Di-n-octylphosphine oxide 6. Di-n-butyl n-butylphosphonate c 7. Di-n-heptylphosphinic acid 8. Bis(di-n-hexy1phosphinyl)methane 100,6 100.5 9. Bis(dicyclohexylphosp1iinyl)methane 97.6 98.2 10. Bis(di-2-ethylbutylp hosphiny1)methane 99 3 11. Bis(di-2-ethylhexy1phosphinyl)methane 12. [(Ili-n-octylphosphinyl)(diphenylphosphinyl)] methane 13. Bis(di-n-hexylphosp hiny1)ethane 14. Bis(di-n-hexylphosphinvl) ropane 15. Bis( di-n-hexylphosphinil )gutane 16. Acetonyldi-n-hexylphosphine oxide 17. Phenacyldi-n-hex 1 hosphine oxide 18. PhenacyldiphenyGgosphine oxide Compounds 1, 2, 4, and 6 were obtained commercially and were analyzed without purification: all other compounds were synthesized in the Ames Laboratory and were purified before being titrated. Titrant was perchloric acid in a glacial acetic acid-acetic anhydride mixture. Poorly defined end point. X o t titratable.
VOL. 36, NO. 1 1 , OCTOBER 1964
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