Anal. Chem. 1980, 52, 857-860
a57
Determination of Phenol in Polluted Air as p-Nitrobenzeneazophenol Derivative by Reversed Phase High Performance Liquid Chromatography Kazuhiro Kuwata,
Michiko Uebori, and Yoshiaki Yamazaki
Environmental Pollution Control Center, 62-3, 1 Chome, Nakamichi, Higashinari-ku, Osaka City 537, Japan
derivative in a complex mixture is also described.
Phenol In polluted airs, such as urban air, Industrial emission, auto exhaust, and tobacco smoke, was collected by using a fritted bubbler with 10 mL of 0.06% sodium hydroxide solution and determined by reversed phase high performance liquid chromatography via derivatlzation with p-nitrobenzenedlazonlum tetrefluoroborate in aqueous medium at pH 11.5. The analytical column used was packed with Polygosll 60-5 CIB (20 cm X 4.6 mm 1.d.) and the mobll phase was 85% methanoVl5 % water. The p-nitrobenzeneazo derivative of phenol could be thoroughly separated from other phenolic members. The detection limit of phenol was 0.05 ppb for 150 L of a gas sample. Phenol vapor in the low ppm or ppb levels could be determined with less than 3 % relative standard deviation.
EXPERIMENTAL Reagents. All of the chemical reagents used were special grade from Wako Pure Chemical Industries (Osaka, Japan) and Tokyo Kasei Kogyo (Tokyo, Japan). Methanol was liquid chromatographic grade from Wako Pure Chemical Industries. p-Nitrobenzenediazonium tetrafluoroborate (NBDATFB) as a derivatizing agent was prepared by the standard method (11,12). The stock solution containing 500 mg/L of phenol was made with distilled water. Lower concentrations were made by appropriate dilution of the stock solution. The 0.1% NBDATFB solution was prepared by dissolving 0.1 g of NBDATFB in 100 mL of distilled water. The buffer solution for pH 11.5 was Menzel buffer which was an aqueous solution containing 0.51% of sodium bicarbonate and 0.21% of sodium carbonate. The sampling solution used was 0.06% sodium hydroxide aqueous solution. Apparatus. A Waters Associates (Milford, Mass.) ALC/GPC 244 liquid chromatograph equipped with a U6K injector and an ultraviolet absorbance detector adjusted to 365 nm was employed. The analytical column used was a 20 cm X 4.6 mm i.d. stainless steel tube, slurry-packed with Polygosil60-5 CIS(Machery-Nagel Co., Diiren, West Germany). The mobil phase was 85% methanol/l5% water, and the flow rate was 1.1mL/min. The column pressure was 1500 psi. Analytical Procedure. Five to one hundred and fifty liters of an air sample were bubbled at 1 to 2 L/min through a fritted bubbler (30 mL) with 10 mL of the sampling solution. In the case of tobacco smoke, 120 mL of smoke for 4 puffs generated by the method described in the previous report (13) were bubbled with a carrier of 1 L/min of clean air. One milliliter of the buffer solution and 3 mL of the NBDATFB solution were added to the sample, and the sample was brought up to 20 mL with distilled water. After 15 min, 2 to 40 p L of this solution were analyzed by LC. The identification of phenol was made by retention time, and the quantitation was performed by peak height. For further confirmative identification or purification of the NBA derivative of phenol in the sample, the sample solution was subjected to the following extraction technique. A part or all of the sample solution was adjusted to pH 4 with 0.1 N hydrochloric acid, and extracted with 2 mL of carbon tetrachloride. The extract was removed, and washed with 2 mL of distilled water. Two milliliters of 0.1% sodium hydroxide solution were added to the extract. The solutions were shaken, and then centrifuged. Two to ten microliters of the aqueous layer were analyzed by LC.
Phenols have received much attention as odorous substances in air pollution studies since the compounds are often observed in polluted airs such as industrial emission, auto exhaust, and even ambient air. Such traces of phenol vapors are usually determined by spectrophotometry ( I , 2) or by gas chromatography (GC) ( 1 , 3 ) . However, only the total amount of phenolic compounds is determined by spectrophotometry unless each member is separated, and phenols such as those found in ambient air are difficult to determine by GC because of its insufficient response. Because phenols react with reactive substances and because they decompose with time, prior t o analysis for phenols it is preferable to derivatize them to stable compounds and then to analyze for the derivatization products. Determinations of phenols as the 2,4-dinitrophenyl derivatives ( 4 , 5 ) ,the pentafluorobenzyl derivatives ( 5 ) ,the 2,6-dinitro-4-trifluoromethyl derivatives ( 5 ) , heptafluorobutyryl derivatives (6) and the pentafluorophenyldimethylsilyl derivatives (7) by GC are reported, but fail to achieve satisfactory recoveries of the phenols from samples. Phenols quantitatively react with p-nitrobenzenediazonium tetrafluoroborate (NBDATFB) in a weak alkaline solution a t room temperature to form p-nitrobenzeneazo (NBA) derivatives of phenols. T h e phenol derivatives can be determined by spectrophotometry if they are separated (8-10). Our goal was t o develop a convenient method for routine analysis of phenol in polluted airs that satisfied the following requirements. (i) Sample analysis should be repeatable within a reasonable time. (ii) Phenol should be separated from other phenolic materials. (iii) Phenol vapor should be accurately detected a t the ppb level. (iv) The method should be applicable in an aqueous medium in which phenol vapor is easily trapped. In this article, a convenient analytical technique which fulfilled the requirements cited above is presented to determine traces of phenol in polluted airs, such as urban air, industrial emission, auto exhaust, and tobacco smoke. Phenol is determined as the NBA derivative of itself by reversed phase high performance liquid chromatography (LC). A technique for confirmative identification and purification of the NBA 0003-2700/80/0352-0857$01 .OO/O
RESULTS AND DISCUSSION Phenols couple with NBDATFB to form the red NBA derivatives in a weak alkaline aqueous solution (8). The reaction between phenols and NBDATFB normally occurs a t the para position in the phenols. If the para position is blocked, the reaction occurs a t the ortho position in the phenols instead. T h e reaction mechanism for phenol is as follows:
C
1980 American Chemical Society
858
ANALYTICAL CHEMISTRY, VOL. 52, NO. 6 , MAY 1980 x 1r2
Table I. Effect of Alkali on Storage of the NBA Derivative of Phenol
-
absorbance of sample (3.36 pglmL as phenol)= X l o - * storage 0.03% NaOH 0.1% NaOH time, day added added
U.0
1.5
1'
0 1 2
3 8
, /y
----.----+o--
4 5
M
0
60 a
REACTlLW TIE, MIN
Figure 1. Effect of pH on the derivatization of phenol with NBDATFB. Injection volume of sample onto the column, 2 pL. 400 pg of phenol in 10mL: ( l ) p H 1 0 . 9 , ( 2 ) p H 1 1 . 3 , ( 3 ) p H 11.5,(4)pH 11.8. 5 0 p g of phenol in 10 mL: (5) pH 10.9, ( 6 ) pH 11.3, (7)pH 11.5, (8) pH 11.8
t
0
1.10
1.07 1.11 1.07
1.07
1.10
1.10
1.09 1.10 1.07
A volume of 2 pL was injected onto the column.
Table 11. Specification of Analytical Columns for the NBA Derivatives of Phenolic Compounds column
mobil phase
specification
Column pressure, 1500 psi Polygosil 60-5 C,, 85% methanol/ Separation, excellent (Machery-Nagel, 15% water without tailing Retention time, 4.00 West Germany ) 20 cm X 4.6 mm 1.1 mL/min min for phenol i.d. Sensitivity, 0.05 ng as phenol at 2 of S/N ratio
xu-' , q'O
1.09 1.09
1
2
3
4
5
6
7
M RATIO ff NBDATFB TO ml Figure 2. Mole ratio of NBDATFB to phenol required in the quantitative derivatization. Injection volume of sample onto the column, 2 pL. Concentration of phenol: (1) 100 pg/mL, (2) 80 pg/mL, (3) 40 pg/mL, (4) 20 pg/mL
In this experiment, the derivatization of phenol for LC analysis was examined for appropriate pH of the reaction medium by using a 5-mL sample containing 50 to 400 pg phenol and bringing the final volume up to 10 mL. Aspects of the derivatization of phenol under several conditions (pH 10.9 t o 11.8) are shown in Figure 1. The reaction rapidly proceeded in the p H range 10.9 to 11.6. The absorbance of the NBA derivative of phenol became stable within 15 min after the reaction began. In the medium of pH 10.9, however, the absorbance was not very stable and a precipitate was formed in the medium as time passed. The absorbance reduced to 68% in 24 h in the medium of pH 10.9, whereas 98% of the absorbance was observed in 24 h a t pH 11.5. In the medium of p H above 11.8, the reaction rate was slow. As a result, the suitable p H in the reaction medium proved t o be about 11.5. The absorbance of the NBA derivative of phenol could be stabilized by adding 0.03 t o 0.1% of sodium hydroxide t o the medium. Table I shows that the absorbance remains constant a t least for a week in the alkaline aqueous solution. T h e mole ratio of NBDATFB to phenol required in the quantitative reaction was examined under several concentrations of phenol. The results are shown in Figure 2. The quantitative derivatization was performed with the mole ratio of NBDATFB to phenol being between 3:l and 4:l. Three analytical columns on LC were useful for separation of the NBA derivatives of phenol. The specification of the columns are shown in Table 11. A Polygosil60-5 CI8 column was used for the purpose. Figure 3 shows a liquid chromatogram of the NBA derivatives of phenolic compounds obtained by the Polygosil60-5 CIB column. Relative standard
RiChrosorb RP 18 (5 P ) (Merck, West Germany) 2 0 cm X 4.6 mm :-I
I.U.
Column pressure, 2000 pSi 85% methanol/ Separation, excellent 15% water without tailing Retention time, 4.70 min for phenol 1.1 mL/min Sensitivity, o.05 ng as phenol at 2 of S/N ratio Column pressure, 2000 psi
Nucleosil C,, (5 p ) 85%methanol/ Separation, excellent (Machery-Nagel, 15% water without tailing West Germany) Retention time, 6.30 20 cm X 4.6 mm 1.1 mL/min min for phenol i.d. Sensitivity, 0.15 ng as phenol at 2 of S/N ratio
4
5
i
1 2
7
I
1
I
I
I
0
5
10
15
FETLVlIcN TIE, Mltl
Figure 3. Liquid chromatogram of the NBA derivatives of phenolic compounds. Conditions: mobil phase, 85 % methanoVl5 % water: pressure, 1500 psi; flow rate, 1.1 mL/min. Phenols: 50-400 ng. (1) Phenol, (2) rn-cresol, (3) o-cresol, (4) a-naphthol, (5) 3,5-xylenol, ( 6 ) 2,3-, 23-,2,6-xylenol, (7) p-cresol, (8) P-naphthol, (9) 3,4xylenol. (10) 2,4-xylenol
ANALYTICAL CHEMISTRY, VOL. 52, NO. 6, MAY 1980
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Table 111. Collection Efficiency and Accuracy of Determined Values of Phenol in t h e Air Samples sampling flow rate, L/min
Amount of phenol trapped in the bubbler,c pg .~ Sample l a Sample 2a Sample 3a _ _ ~ _ _ _ B-1 B-2 B-1 B-2 B-1 B-2
run
3
8.78 8.89 8.7 3
n.d. n.d. n.d.
58.6 57.9 56.9
n.d. n.d. n.d.
67 3 686 666
n.d. n.d. n.d.
1
8.85
2
8.38
3
8.38
n.d. n.d. n.d.
58.5 57.6 57.1
n.d. n.d. n.d.
686 688 659
n.d. n.d. n.d.
1
1.0
2
2.0
859
________
8.67
t
0.230
57.8
5
0.70
676 * 2.2
0.451
t
0.0120
3.01
i
0.036
35.2
average amount of phenol trapped i S.D., pg average concentration of phenol S.D., ppmb
z
2.66
R.S.D., %
1.20
t
0.63
1.79
A 5-L sample volume was taken in each run. Concentration at 25 "C. B-1: first bubbler, B-2: second bubbler, n.d.: not detectable, S.D.: standard deviation, R.S.D.: relative standard deviation. a
Table IV. Phenol in Polluted Air sample
phenol found
urban air, ppb' 1-3 1-3 1-3 1-3
L.
L
5
10
0
5
pm, pm, pm, pm,
Aug. 3 0 , 1 9 7 9 , sunny Sept. 1 7 , 1 9 7 9 , cloudy Sept. 20,1979, sunny Sept. 2 1 , 1 9 7 9 , sunny
1.01
0.55 0.96 0.69
industrial emission, ppmb Casting
No. 1 2
3 4
0.353 0.644 0.918 0.208
auto exhaust, ppmb 5
19
0
5
13
L JL
0
5
10
0
5
10
FfTINI[xI TIS, YIN
Confirmative identification of phenol in samples by the extraction technique. Arrow marks show peaks for phenol. (1) Initial sample, (2)sample purified by the extraction technique. (A) Urban air, 0.69 ppb; (B) auto exhaust, 0.320 ppm; (C) tobacco smoke (Peace), 436 Kglcigarette Figure 4.
deviations of the retention times for the compounds were less than 0.1% more than on a day when the analytical conditions were not changed. T h e NBA derivative of phenol could be thoroughly separated from those of other phenolic members. The extraction technique was useful for further confkmative identification of phenol and minimization of background in environmental and complex samples. In an acidic medium at p H 4 to 5, more than 99.8% of the NBA derivative of phenol extracted into the carbon tetrachloride layer. T h e derivative in the organic layer could be quantitatively extracted into an alkaline aqueous solution though complete extraction was not obtained. T h e higher concentration of sodium hydroxide in t h e aqueous layer might give a better extraction, but 0.1% of sodium hydroxide seemed t o be the maximum while still ensuring protection of t h e analytical column. When 0.1% sodium hydroxide solution was used, the extraction recoveries for 10 to 50 gg of phenol were 72.2 to 72.4% and their relative standard deviations were less than 1.6%. The recoveries for
Toyota Engine VG-20
No. 1
2 3
0.295 0.320 0.233
tobacco smoke, pg/cigarettec Hi-lite(Japan) Seven-star (Japan) Cherry (Japan) Peace (Japan)
317 323 312
436
a A volume of 150 L of air was sampled at the Environmental Pollution Control Center, Osaka City. Sample volume was 1 0 L. Average value in 3 runs.
300 to 500 ng of phenol corresponding to urban air levels decreased to 58 to 60% and the relative standard deviations were approximately 5%. Figure 4 shows that the extraction technique is useful for identification of phenol in urban air, auto exhaust, and tobacco smoke. This technique was applicable to phenol only for quantitative determination. The minimum detectable quantity of phenol (a response to twice the noise level) by LC via the derivatization was 0.05 ng. The sensitivity was 30 times higher than that achieved without the derivatization by LC. The ultraviolet absorbance detector used was adjusted to 254 nm. An excellent linear calibration graph could be obtained over the range of 0.5 to 160 ng as free phenol, where relative standard deviations of the determined values were less than 3%. At concentrations near the detection limit between 0.05 to 0.1 ng. the calibration graph curved, but still the relative standard deviations were 10 to 15%. No phenol peak was observed in blank tests being carried out by using 150 L of phenol-free urban air purified through a tube packed with Molecular Sieve 13X granules. When the minimum amount of phenol was 0.05 ng, the es-
860
ANALYTICAL CHEMISTRY, VOL. 52, NO. 6, MAY 1980
0
2
4
6
8
1
0
EIEVICN T I E , MiN
Figure 5. Typical liquid chromatograms of the NBA derivative of phenol isolated from polluted airs. Arrow marks show peaks for phenol. (A) Urban air (at Environmental Pollution Control Center), 0.96 ppb; (B) industrial emission (casting),0.918 ppm; (C) auto exhaust, 0.295 ppm; (D) tobacco smoke (Seven-star),323 Fglcigarette
timated detection limit of phenol vapor was 0.05 ppb for 150 L of a gas sample at 25 "C. Effects on the analysis of a number of organic substances other than phenolic compounds were apparently not observed in most cases since organic substances having ultraviolet absorbance below 300 nm of wavelength were not detected a t 365 nm where the NBA phenol derivatives were measured. In the case of an extremely polluted gas such as tobacco smoke, the extraction technique was useful to minimize the background as shown in Figure 4. For investigations on the collection efficiency in the sampling system and on the accuracy of the analysis, 50 L of sample air containing 0.45 to 35 ppm of phenol were prepared in polyester bags. Five liters were repeatedly bubbled a t 1 and 2 L/min through two fritted bubblers with 10 mL of the sampling solution, followed by analysis. Table I11 reports that under these conditions phenol can be thoroughly trapped in
the first bubbler, and that traces of phenol in gas samples can be accurately determined by the method presented. For validation of the collection efficiency of phenol vapor a t urban air levels, similar tests were carried out by sampling 150 L of urban air a t 2 L/min. No phenol peak was detected from the second trap. Phenol in urban air, auto exhaust, industrial emission and tobacco smoke was determined. Table IV reports analytical data of phenol in several sources, and Figure 5 shows typical liquid chromatograms of the NBA derivative of phenol isolated from the sources. For urban air, 100 L or a higher volume of air should be sampled to obtain reliable data for extreme traces of phenol. In this study 150 L of ambient air were sampled. The daytime concentrations of phenol in summer urban air in Osaka were usually 0.5 to 1.5 ppb. The concentrations of phenol during a sunny day tended to be higher than those during a cloudy day. The concentrations of phenol in industrial emission were different from source to source. In auto exhaust, 0.2 to 0.4 ppm of phenol was observed under the standard operation. Considerably high concentrations of phenol were observed in tobacco smoke. The regular sized cigarettes seemed to generate 300 to 450 pg/cigarette of phenol. In closing, it is suggested that the analytical method by LC might be extended to phenolic compounds other than phenol in polluted airs if suitable conditions for the derivatization and identification of each member are examined.
LITERATURE CITED Van Haverbeke, L.; Herman, M. A. Anal. Chem. 1979, 5 1 , 932-936. Ramstad, T.; Armemtrout, D. N. Anal. Chim. Acta 1978, 102, 229-231. Hoshika, Y.; Muto, Y. Bunseki Kagaku 1978, 27, 273-278. Cohen, I. C.; Nwcup, J.; Ruzicka, J. H. A,; Wheals, B. E. J . Chromatogr. 1969, 4 4 , 251-255. (5) Seiber, J. N.; Crosby, D. G.; Fouda, H.; Soderauist. C. J. J . Chromatoor. 1972, 73, 89-97. (6) Lamparski, L. L.; Nestrick, T. J. J Chromatogr. 1978, 156, 143-151. (7) Francis, A. J.; Morgan, E. D.; Poole, C F. J . Chromatogr. 1978, 167. 111-1 17. (8) Leibnitz. E.: Behrens, U.: Riedel. L: Gabert. A. J . Prakt. Chem. 1960. 1 1 , 125-132 (9) Teodorescu, V I Toma. C , Deceanu. T I Rev Chm (Bucharest) 1975, -26 - 605-607 ..- - .. . (IO) Teodorescu, V.; Popescu, R.; Glodeanu, E. Rev. Chim. (Bucharest) 1978, 29, 586. (11) Cheronis, N. D.; Entrinkin, J. E. "Semimicro Qualitative Organic Anaslysis"; Crowell: New York.,, 1957; p 237. (12) The Chemical Society of Japan. A Course in Experimental Chemistry", 3rd ed.; Marusen: Tokyo, 1967; Vol. XVI, p 326. (13) Kuwata, K.;Uebori, M.; Yamazaki, Y. J , Chromatogr. Sci. 1979, 17, 264-268. (1) (2) (3) (4)
.
RECEIVED for review October 29, 1979. Accepted January 18, 1980.
