Simultaneous Determination of Benzoic Acid and Its Monohydroxy Isomers by a Spectrophotometric Method Mariana Mantel" Soreq Nuclear Research Center, Yavne, Israel
Mariana Stiller Weizmann lnstitute of Science, Rehovot, lsrael
A spectrophotometric method for the simultaneous determination of benzoic acid and its ortho, meta, and para hydroxy isomers is described. The four acids are determined without any prior separation by measuring the absorbance of their mixture at 300, 260, and 225 nm and that of the Fe(lll) complex of o-hydroxybenroic acid at 525 nm. The concentrations of the individual acids are calculated by solving a set of two simple equations or by a graphical method. An error of f5-10% was obtained.
The determination of each of the acids in a mixture of benzoic acid a n d its ortho, meta, a n d para hydroxy isomers has been of interest for m a n y years. M o s t of the methods published require the preliminary separation of the individual acids by techniques such as paper chromatography ( I ), ion-exchange (2, 3 ) , thin-layer chromatography (TLC) (4-6), or gas chromatography of their methyl ether derivatives (7, 8). The separated acids are t h e n quantitatively evaluated b y spectrophotometry ( 4 ) . The method described in the present paper permits t h e simultaneous direct determination of the four acids without a n y prior separation. The method is based o n measurem e n t s of t h e absorbance of the mixture of acids at 300, 260, and 225 n m and of the Fe(II1) complex of o-hydroxybenzoic acid at 525 nm. T h e calculation of t h e individual concentrations is described. EXPERIMENTAL Reagents and Standard Solutions. The following stock soluM benzoic acid and 0 - , m - , and p-hytions were prepared: droxybenzoic acids, in distilled water, using the respective Fluka analytical grade reagents; 0.1 M iron(II1) solution in dilute hydrochloric acid by dissolution of metallic iron. Apparatus. Absorbance readings were taken with a Beckman DU spectrophotometer and quartz cells of 10- and 20-mm light path. A Radiometer 25 pH meter was used for pH measurements. Calibration curves. (1): Separate calibration curves for each acid at 300, 260, and 225 nm were obtained as follows. Except for the two cases described below, 0.5 to 7 ml of each of the standard stock solutions were adjusted to pH 2 and diluted to 25 ml with distilled water. The absorbances of these solutions were measured at the three wavelengths with 10-mm light path cells. For the calibration curves of p-hydroxybenzoic acid at 260 nm and benzoic acid at 225 nm, 0.5-2.5 ml of the stock solutions were treated in the same way. (2): The iron(II1)-salicylic acid complex calibration curve was obtained by treating 0.5 to 5 ml of the low3M stock solution of o-hydroxybenzoic acid (salicylic acid) as described below under Procedure for the determination of salicylic acid. Procedure. One ml of 0.1 M iron(II1) solution was added to an aliquot of the mixture of acids. The pH was adjusted to 2, the mixture diluted to 25 ml with distilled water, mixed well, and allowed to stand for at least half an hour. The absorbance of the complex was measured at 525 nm (with 20-mm light path cells) vs. a reagent blank. A separate aliquot was adjusted to pH 2 and diluted to 25 ml with distilled water. The absorbance of this solution was measured at 300, 260, and 225 nm with 10-mm light path cells. Calculation of Results. 1 ) o-Hydroxybenzoic acid The 712
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amount of o-hydroxybenzoic acid present in the sample was calculated from the absorbance of its Fe(II1) complex at 525 nm. 2 ) m-Hkdroxybenzoic acid. The amount of m-hydroxybenzoic acid was calculated from the absorbance at 300 nm. The absorbance due to the amount of o-hydroxybenzoic acid found in (1)was read from its standard calibration curve at this wavelength and subtracted from the total reading. The remainder corresponds to the absorbance of m-hydroxybenzoic acid (Table I). 3 ) p-Hydroxybenzoic and Benzoic Acids. The amounts of p hydroxybenzoic acid and benzoic acid were calculated from the absorbance readings at 260 and 225 nm. The absorbances at these wavelengths corresponding to the amounts of 0 - and m-hydroxybenzoic acids calculated in (1) and (2) were read from the respective calibration curves and deducted from the total absorbance. The remaining absorbance corresponds to the sum of benzoic and p-hydroxybenzoic acids. The amount of each acid was calculated either algebraically (a) or graphically (b). a) The respective amounts were calculated by solving the following set of 2 equations:
where A2fio and AZz5= net absorbance calculated at 260 and 225 nm, respectively; X and Y = molar concentrations of p-hydroxybenzoic and benzoic acids, respectively; e6;o and tf5 = absorption coefficient of p-hydroxybenzoic acid at 260 and 225 nm, respectively (see Table I); and et6' and cgZ5 = absorption coefficient of benzoic acid at 260 and 225 nm, respectively (see Table I). b) A graphical method is recommended as an alternative to (a) (see Figure 1).A nomograph was constructed as follows. Two vertical axes represented the absorbance readings at 225 and 260 nm. On these two axes, the absorbances measured at the two wavelengths corresponding to a given amount of one of the acids were recorded. A straight line was drawn through the two points and extended until it intersected the X-axis. At this point, a vertical line was raised. The procedure was repeated for the other acid and a second vertical line was raised. (These two verticals will serve as concentration scales.) The two lines which connect the respective absorbances of each acid at the two wavelengths are now extended at their other end until they intersect the last pair of verticals. The distance from the X axis to these points of intersection represented the respective amounts of benzoic and p-hydroxybenzoic acids. A concentration scale was obtained by dividing this section of the verticals into equal parts. Figure 1 shows such a nomograph constructed from the absorbance readings obtained for 10 wg/ml of benzoic and 10 fig/ml of p-hydroxybenzoic acid. For an unknown mixture of the acids, the net absorbances obtained at 260 and 225 nm were plotted on the vertical axes representing absorption. A line was drawn through these 2 points and extended until it intersected the concentration scales. The concentrations of the respective acids were read at the points of intersection.
RESULTS AND DISCUSSION Ultraviolet Spectra. T h e absorption spectra of t h e four acids of interest were determined in t h e uv region (210-340 n m ) at different pH values. B y comparing these spectra (which agreed well with those previously found b y other workers (9)),i t was concluded that pH 2 is the most suitable for t h e simultaneous determination of t h e 4 acids. A t
Acid
E
E
N
m
5
Table I. Absorption Coefficients at pH 2 Wavelength
C
-
N N
e
Benzoic o-Hydroxy rn-Hydroxy p-Hydroxy
22s
260
12250 5270 5520 2340
828 307 522 14000
300
0 3400
2500 0
Table 11. Analysis of Mixtures of Acids Concentration % Taken
, I I1 I11 IV
38 10 20 50
Found
m
P
b
0
m
P
b
25 52
22
15
36
20
14
10 10
30
24 40 30
11 22
26 55 8 8
13 25 42 31
10
47
15
31 9
this pH, it is possible to choose 3 wavelengths for which t h e differences in t h e values of the absorption coefficients are maximum. Table I summarizes the absorption coefficients calculated a n d Figure 2 shows the absorption spectra obtained a t p H 2. As may be seen, benzoic and p-hydroxybenzoic acids d o n o t absorb a t all a t 300 nm. I t follows t h a t the absorbance reading a t this wavelength is t h e sum of t h e readings d u e to 0 - a n d m-hydroxybenzoic acids. Favorable, too, is the fact t h a t t h e absorption coefficients of these two acids are similar in magnitude. This allows t h e subtraction of t h e a m o u n t of o -hydroxybenzoic acid, calculated according to its Fe(II1) complex, from the total reading, to obtain the a m o u n t of m-hydroxybenzoic acid without introducing significant errors. T h e other two wavelengths, 225 a n d 260 n m , were chosen since, a t each, t h e absorption coefficient of one of t h e two acids (benzoic a n d p-hydroxybenzoic) is much higher t h a n t h a t of t h e others. T h u s , it is possible in this case, too, t o deduct the absorbances due to 0 - a n d m-hydroxybenzoic acid a t these two wavelengths from t h e total reading, witho u t introducing significant errors. I t also permits t h e use of a graphical solution since, a t each wavelength, t h e absorption of one acid is much higher t h a n t h a t of t h e other (-5 times a t 225 nm a n d -17 times a t 260 n m ) . Iron(II1)-o-Hydroxybenzoic Acid Complex. Fe(II1) solutions produce, with salicylic acid, a complex whose color varies from violet in a n acid medium to yellow in a n alkaline one. This complex has been extensively used for t h e quantitative determination of salicylic acid (10) as well as of Fe(II1) (11 ), Bertin-Batsch (12) studied this complex by spectrophotometric methods a n d found it t o have a 1 : l composition,
Figure 1. Nomograph constructed for the calculation of the concentration of benzoic and p h y d r o x y b e n z o i c acids based on the absorbance readings (10-mm light path cells) for 10 k g / m l benzoic and 10 p g / m l p h y d r o x y b e n z o i c acid
J
1
1
1 b
'02-). S h e also found t h a t 0Beer's law is obeyed for p H values lower t h a n 3 a n d a concentration ratio of R/Fe(III) = 10. T h i s same ratio has been used for t h e determination of iron by Scott (13) a n d other workers ( I f ) . For the determination of salicylic acid a n excess of iron was used (14). In t h e present work, we too used a n excess of Fe(II1) a n d found t h a t Beer's law was obeyed in t h e range 3-30 lg/ml. An absorption coefficient of 1680 was obtained which agreed well with t h e value of 1630 obtained by Bertin-Batsch (12). Results. Several mixtures containing varying concentrations of t h e four acids were prepared a n d analyzed by t h e present method. T h e results obtained are shown in Table 11. As may be seen, an error of &5-10% was obtained for in-
'1 lI o t
:
2
T
-
06
Waveknglh ( n m l
Figure 2. A and 6.Absorbance spectra at pH 2 B = benzoic acid, 0 = o-hydroxybenzoic acid, M = mhydroxybenzoic acid, P = phydroxybenzoic acid. A . 210-280 nm, concentration of acids = 2.10-4 M, 5-mrn cells. 8.250-340 nm, concentration of acids = 2.10-4 M, 10-mm cells ANALYTICAL CHEMISTRY, VOL. 48, NO. 4, APRIL 1976
713
dividual results and a recovery of &5% for t h e sum of t h e acids. T h e present method was applied t o t h e study of t h e deiodination products of 1311-labeled o-iodobenzoic acid produced by radiolysis under t h e influence of different catalysts. I t is known t h a t benzoic and hydroxybenzoic acids are among t h e products of this radiolytic deiodination. T h e extent of deiodination was determined by measuring t h e radioactive iodide originating from t h e labeled acid. T h e interference of t h e unreacted iodobenzoic acid with t h e ahsorhance measurements was estimated as follows. T h e absorption coefficients of iodobenzoic acid were determined a t 300, 260, a n d 225 nm. T h e following values were ohtained: t = 720 a t 300 nm; 1140 at 260 n m and 10 000 a t 225 nm. Before applying t h e method for t h e calculation of o-, m-, p - hydroxybenzoic acids and benzoic acid, as described above, t h e absorbances corresponding t o t h e concentration of t h e unreacted iodohenzoic acid, were deduced from t h e total readings a t t h e respective wavelengths. T h e same solutions were also analyzed by paper chromatography ( I ) ; very good agreement was found between t h e two methods.
tives. T h e advantages of t h e method are its simplicity and rapidity. T h e procedure consists essentially of four spectrophotometric readings which can he carried out in about 40 minutes.
LITERATURE CITED (1) M. Mantel and E. Kushnir, J. Chromatogr., 17, 624 (1965). (2) N. E . Skelly and W. E. Crummett, Anal. Chem., 11, 1681 (1963). (3) Wataru Funasaka, Kaseimi Fujimura, and Sue Kushida, J. Chromatogr.. 64, 95 (1972). (4) E . Ludwig and U. Freirnuth, Nahrung, 9, 751 (1966). (5) S. Y. Pinella. A. D. Falco, and G. Schwartman, J. Assoc. Off. Agric. Chem., 49, 829 (1966). (6) J. E . Sinsheirner and G. 0. Breault, J. Pharm. Sci., 60, 255 (1971). (7) E. R. Blakiey. Anal. Biochem., 15, 350 (1966). (8) C. M. Williams, Anal. Biochem., 11, 224 (1965). (9) L. Lang, Ed., "Absorption Spectra in the Ultraviolet and Visible Region," Academic Press, New York, 1961. (10) A. C. Kelly, J. Pharm. Sci., 59, 1053 (1970). (11) F. D. Sneli and C. Snell. "Colorimetric Methods of Analysis," 1949, 3rd ed., Van Nostrand, London. (12) C. Bertin-Batsch, Ann. Chim., 7, 481 (1952). (13) R. 0. Scott, Analyst(London),66, 142 (1941). (14) V. Das Gupta, J. Pharm. Sci., 61, 1625 (1972).
CONCLUSION A method is described for t h e simultaneous determination of benzoic acid a n d its three isomeric hydroxy deriva-
RECEIVEDfor review September 8, 1975. Accepted December 8,1975.
Extraction and Spectrophotometric Determination of Vanadium(V) with N-[p-N,N-DimethylaniIino)-3-methoxy-2naphtho]hydroxamic Acid Shahid Abbas Abbasi Deparfment of Chemistry, lndian lnstitute of Technology, Bombay-400 076, lndia
On the basis of available information on the methods of determination of vanadium(V) by hydroxamic acids, the title reagent was modeled and developed as a selective and sensitive reagent for vanadium(V). The chloroform solutions of the reagent extract vanadium rapidly (5 2 min) from 2-6 M hydrochloric acid solutions as a violet complex which is measurable spectrophotometrically at 570 nm ( E = 1.2 X lo4 1. mol-' cm-'). The color system which contained vanadium and the reagent in the molar ratio 1:2, obeys Beer's law in the range 0.15-8.5 ppm of vanadium. The method of determination has a maximum standard deviation of f0.02 and tolerates the presence of several milligrams of 50 foreign ions. Vanadium was determined in ilmenite, rock phosphate, and steels.
+
N-Phenylhenzohydroxamic acid ( P B H A ) has been extensively used in analytical chemistry (1-3) as a reagent which is fairly sensitive and selective for t h e liquid-liquid extraction and spectrophotometric determination of vanadium(V). T h e advantage with P B H A is t h a t its vanadium complex attains maximum color intensity ( 6 = 4650) ( I ) in highly acidic (2.8-4.3 M HC1) solutions and can he readily extracted into various organic solvents such as chloroform, benzene, and carbon tetrachloride. T h e quickness and convenience of t h e P B H A method ( I ) has prompted many a t tempts t o find analogues of P B H A which are more sensitive and selective reagents for vanadium t h a n PBHA, note714
ANALYTICAL CHEMISTRY, VOL. 48, NO. 4, APRIL 1976
worthy among them being N-p-tolyl-2-thenohydroxamic acid ( t = 5700 in CHC13)(4), N-phenylcinnamohydroxamic acid ( e = 6300 in CHC13)(5) and N-phenyl-3-styrylacrylohydroxamic acid ( t = 7500 in CHC13)(6). Various claims of improved selectivity are made with t h e above-mentioned reagents. T h e N-phenyl-2-naphthohydroxamicacid proposed recently ( 7 ) does not appear t o be a significant imacid if provement over AJ-phenyl-3-styrylacrylohydroxamic judged by t h e sensitivities and tolerance limits of t h e methods using t h e two respective acids. From a n extensive literature search on the sensitivities and selectivities of various hydroxamic acid methods ( I , 4-13), it was noticed t h a t the presence of strong electrondonating substituents in t h e para position of t h e N-phenyl fragment of PBHA, a n increase in conjugation a t t h e henzo site and introduction of electron-donating groups ortho t o t h e functional -C=O group helps in enhancing t h e sensitivity and selectivity of a hydroxamic acid towards vanadium(V) relative t o P B H A (I). T h e studies of Cassidy and
