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Complexometric determination of citric acid with copper
0039-9140/85$3.00+ 0.00 Pergamon Press Ltd
Talanro,Vol. 32, No. 2, pp. 153-154, 1985
Printed in Great Britain
COMPLEXOMETRIC DETERMINATION ACID WITH COPPER
OF CITRIC
E. SZEKELY Institute for Chemistry & Chemical Technology, The Institutes for Applied Research, Ben-Gurion University of the Negev, P.O. Box 1025, Beer-Sheva 84110, Israel (Received
13 April 1984. Revised
14 June 1984. Accepted
14 September
1984)
Summary-A
method is presented in which citrate is determined by complexation with excess of copper( followed by back-titration with EDTA. Small amounts of citrate can be determined by this method in the presence of large amounts of most organic acids present in natural products. Interfering reductants are removed by treatment with permanganate, amino-acids with nitrous acid, and metal ions with a cation-exchanee resin. The indicator used for end-point detection is Szechromotrope, 4,4’-sulphonyl-bis(azob&zene)
dichromotropic
acid
The use of 4,4’-sulphonyl-bis(azobenzene) dichromotropic acid as a metallochromic indicator for calorimetric and complexometric determination of small amounts of copper(H) was reported earlier.’ This paper presents a method in which this indicator is used in the determination of citrate by addition of excess of copper(H) and back-titration with EDTA. Citric acid can be determined by this method with high precision in the presence of other acids if interfering amino-acids are removed by treatment with nitrous acid, reductants by treatment with permanganate, and metal ions by treatment with a cation-exchange resin. The use of copper(I1) for complexation and determination of citric acid has been reported before,*-* but our method has advantages of simplicity and reliability, can be performed without expensive instruments, and can also be used for micro-amounts of citrate. The indicator was synthesized in our laboratory by diazotizing 4,4’-sulphonyldianiline and coupling it with chromotropic acid. It has been recommended as a reagent for the calorimetric determination of boron.’ The indicator (C,2H,,N,0,8Na,) is a brownish-red powder, easily soluble in water to give a pink solution with molar absorptivity 5.8 x IO4 l.molee’ .cm-’ at &,,,,, = 538 nm. A blue copper complex forms instantly at pH 6.8-7.6 (molar absorptivity 4.2 x IO4 1. mole-’ .cm-’ at A,,,,, = 384 nm). Aqueous solutions of the reagent and its complexes are stable. No blue colour develops with other uni-, bior tervalent metal ions. The reagent is commercially available under the trade name Szechromotrope, from Gammatest Ltd, Ben-Gurion University of the Negev, Beer-Sheva 84110, Israel. EXPERIMENTAL Reagents Aqueous indicator solution, 0.1%. Copper sulphate solution, O.OlM, standardized titration.
by EDTA
Standard O.OlM and O.OOlM EDTA. Buffer solution. Ammonium chloride (lM)/sodium
acctate (lM), adjusted to pH 7.5 with dilute ammonia solution or hydrochloric acid. Tetra-amminocuprate(If) solution. Prepared by mixing ml of O.OlM copper sulphate with 40 ml of the buffer.
10
Procedures Detection and estimation. Transfer about 5 ml of water into a small flask or large test-tube, add 0.25 ml (5 drops) of amminocuprate solution and 1 drop of indicator solution and mix. Add nearly neutral sample solution slowly from a graduated pipette, continuously shaking the vessel until the colour changes from blue to violet, which corresponds to the presence of about 0.1 mg of citric acid in the volume of sample solution added. A colour change to pink indicates the presence of larger amounts of citric acid. Repeat the estimation with a more dilute sample solution if a colour change is observed after addition of only a few drops of sample. Interfering substances must be removed beforehand. Determinations. (A) Transfer an amount of sample solution containing l-8 mg of citric acid into an Erlenmeyer flask and dilute it to about 100 ml. Add exactly 5 ml of O.OlM copper sulphate, 2 drops of indicator solution, and if necessary O.lM ammonia or hydrochloric acid until the colour turns blue (pH 6.8-7.6). Add 5 ml of buffer solution and titrate the excess of copper with 0.01 M EDTA until the colour changes to pink. (B) Transfer an aliquot of sample solution containing 0.2-3 mg of citric acid into an Erlenmeyer flask and dilute it to about 50 ml. Add exactly 2 ml of O.OlM copper sulphate, 1 drop of indicator solution, and if necessary 0. I M
ammonia or hydrochloric acid until the colour becomes blue (uH .. 6.7-7.6). Add 2 ml of buffer solution and titrate the excess of copper with O.OOlM EDTA until changes to violet, then slowly until it becomes
RESULTS AND
the colour pink.
DISCUSSION
Interferences
Only a few of the organic acids present in natural products interfere in the detection, estimation and determination of citrate ions by the procedures above. Up to 1000 ppm of tartaric, succinic, malic, hydroxyacetic, lactic, salicylic and quinic acids have no influence on the precision, nor do phosphoric or
154
SHORT
COMMUNICATIONS
nitric acids or univalent metal ions. Determinations performed in the presence of up to 5 g of glucose and up to 20 ml of ethanol in 100 ml of sample solution showed no bias. Interference by reducing acids such as ascorbic, oxalic, fumaric and sulphurous was prevented by the addition of O.lN permanganate. Metal ions were removed by treatment with a cationexchange resin. Interference from amino-acids was eliminated by addition of sodium or potassium nitrite to acidified solutions. The reliability and precision were determined by repeated analysis. An accurately prepared 20-mg/ml citric acid stock solution was prepared with anhydrous citric acid dried at 110” to constant weight; a lo-ml aliquot of this solution was diluted, neutralized and made up to 200 ml: 1 ml = 1 mg of citric acid. Various mixtures were also analysed. Sample I. A 5-ml aliquot of the 1-mg/ml solution was diluted to about 100 ml and analysed by procedure (A). In 5 determinations an average of 2.40 ml of EDTA solution was used for titration of the excess of copper, with a maximum deviation of kO.02 ml (0.8%). The amount of citric acid was calculated from citric acid = (5 - a) x 1.92 mg where a = ml of O.OlM EDTA used (found: 4.99 mg). A 5-ml aliquot of the 1-mg/ml solution was diluted accurately to 100 ml and a lo-ml portion of this solution was titrated with O.OOlM EDTA by procedure (B). In 5 determinations an average of 17.40 ml of EDTA was used, with a maximal deviation of fO.1 ml (0.6%). The amount of citric acid was from citric acid = (20 - a) x 0.192 calculated mg = 0.499 mg. Sample II. Citric acid, 0.02 g; malic acid, 0.50 g; tartaric acid, 1.00 g; glucose 10.0 g; ethanol, 20 ml; water to 100 ml. Samples of 10, 25, and 50 ml were taken. The lo-ml sample was diluted to about 50 ml, treated by procedure B, and titrated with O.OOlM EDTA (14.9 ml). Citric acid found =0.98 mg. The 25- and 50-ml samples were diluted to about 100 ml and titrated with O.OlM EDTA. Citric acid found =2.53 mg (25-ml sample) and 4.90 mg (50 ml). Sample III. Citric acid, 0.05 g; calcium chloride, 1.O g; magnesium chloride, 0.50 g; urea, 5.0 g; water to 100 ml. A 50-ml portion was treated with Amberlite IR 120 cation-exchange resin, neutralized, and diluted to 100 ml. Three aliquots (25 ml each) were diluted to about 100 ml, treated by procedure A and titrated with O.OlM EDTA (1.76, 1.80 and 1.78 ml). Citric acid found =6.18 mg, corresponding to 49.4 mg in 100 ml of sample III. Sample IV. Citric acid, 0.50 g; ascorbic acid, 0.25 g; oxalic acid, 0.05 g; water to 100 ml. Ten ml were
diluted to about 50 ml and 1 ml of 0.1 N hydrochloric acid was added, and then 0.1 N permanganate (dropwise). The excess of permanganate was removed by adding 1 drop of 0.1 N thiosulphate. The solution was then treated with the cation-exchange resin, neutralized, and diluted to about 100 ml, treated by procedure A, and titrated with O.OlM EDTA (2.40, 2.42 and 2.42 ml). Citric acid found =4.79 mg, corresponding to 0.479 g in sample IV. Sample V. Citric acid, 0.50 g; lactic acid, 5.0 g; glutamic acid, 1.O g; water to 100 m. To 25 ml of this solution 5 ml of concentrated hydrochloric acid and 0.5 g of sodium nitrite were added. The solution was mixed and heated to about 70” until no more gas was evolved. The neutralized solution was diluted to 100 ml. Three 5-ml aliquots of the sample solution were diluted to about 100 ml, treated by procedure A, and titrated with O.OlM EDTA (1.74, 1.72 and 1.74 ml). Citric acid found =6.28 mg, corresponding to 0.502 g in sample V. Though there are several indicators for complexometric determination of copper( which can also be used in our method, the indicator recommended has several advantages. Murexide, though widely used, is less stable, the colour change from yellow to violet is less sensitive, and it is more difficult to maintain the desired pH. The nitrosochromotropic acid indicator”.” gives a colour change comparable to that of our indicator, but its solution, which has to be freshly prepared for each assay, is less stable. Also, two different pH ranges were recommended by the authors (pH 7.25-8.80 and 5.8-6.5) without explanation. REFERENCES
1. E. Szekely, M. Flitman
2. 3. 4. 5. 6.
7. 8.
and M. Leon, Proc. Israel Chem. Sot., 42nd Meeting, 1972, 94. G. Graffman, H. Domels and M. L. Strater, Fetten, Seifen, Anstrich., 1974, 76, 218. M. Hamon, A. Hoppendt and M. Guemet, Ann. Pharm. Franc., 1972, 30, 595. M. F. El-Taras and E. Pungor, Anal. Chim. Acta, 1976, 82, 285. A. Olin and B. Wallen, ibid., 1983, 151, 65. A. Pierre and S. Braley, Lair, 1983, 63, 623. H. L. Nours, J. F. Le Meur and C. Bourgeois, Sci. Aliments, 1982, 2, 483. M. T. M. Zaki and R. Alquasmi, Z. Anal. Chem., 1981,
306,400. 9. E. Szekely, Proc. Annual Meeting Israel Chem. Sot., 41st Meeting, 1971, 13. 10. S. Schwarzenbach and H. Flaschka, Complexometric Titrations, 2nd Ed., Methuen, London, 1969. 11. C. S. Panda and T. S. Srivastava, Z. Anal. Chem., 1962, 184, 248. 12. A. B. Sen and T. S. Srivastava, ibid., 1962, 187, 401.
Talanro,Vol. 32, No. 2, pp. 153-154, 1985
Printed in Great Britain
COMPLEXOMETRIC DETERMINATION ACID WITH COPPER
OF CITRIC
E. SZEKELY Institute for Chemistry & Chemical Technology, The Institutes for Applied Research, Ben-Gurion University of the Negev, P.O. Box 1025, Beer-Sheva 84110, Israel (Received
13 April 1984. Revised
14 June 1984. Accepted
14 September
1984)
Summary-A
method is presented in which citrate is determined by complexation with excess of copper( followed by back-titration with EDTA. Small amounts of citrate can be determined by this method in the presence of large amounts of most organic acids present in natural products. Interfering reductants are removed by treatment with permanganate, amino-acids with nitrous acid, and metal ions with a cation-exchanee resin. The indicator used for end-point detection is Szechromotrope, 4,4’-sulphonyl-bis(azob&zene)
dichromotropic
acid
The use of 4,4’-sulphonyl-bis(azobenzene) dichromotropic acid as a metallochromic indicator for calorimetric and complexometric determination of small amounts of copper(H) was reported earlier.’ This paper presents a method in which this indicator is used in the determination of citrate by addition of excess of copper(H) and back-titration with EDTA. Citric acid can be determined by this method with high precision in the presence of other acids if interfering amino-acids are removed by treatment with nitrous acid, reductants by treatment with permanganate, and metal ions by treatment with a cation-exchange resin. The use of copper(I1) for complexation and determination of citric acid has been reported before,*-* but our method has advantages of simplicity and reliability, can be performed without expensive instruments, and can also be used for micro-amounts of citrate. The indicator was synthesized in our laboratory by diazotizing 4,4’-sulphonyldianiline and coupling it with chromotropic acid. It has been recommended as a reagent for the calorimetric determination of boron.’ The indicator (C,2H,,N,0,8Na,) is a brownish-red powder, easily soluble in water to give a pink solution with molar absorptivity 5.8 x IO4 l.molee’ .cm-’ at &,,,,, = 538 nm. A blue copper complex forms instantly at pH 6.8-7.6 (molar absorptivity 4.2 x IO4 1. mole-’ .cm-’ at A,,,,, = 384 nm). Aqueous solutions of the reagent and its complexes are stable. No blue colour develops with other uni-, bior tervalent metal ions. The reagent is commercially available under the trade name Szechromotrope, from Gammatest Ltd, Ben-Gurion University of the Negev, Beer-Sheva 84110, Israel. EXPERIMENTAL Reagents Aqueous indicator solution, 0.1%. Copper sulphate solution, O.OlM, standardized titration.
by EDTA
Standard O.OlM and O.OOlM EDTA. Buffer solution. Ammonium chloride (lM)/sodium
acctate (lM), adjusted to pH 7.5 with dilute ammonia solution or hydrochloric acid. Tetra-amminocuprate(If) solution. Prepared by mixing ml of O.OlM copper sulphate with 40 ml of the buffer.
10
Procedures Detection and estimation. Transfer about 5 ml of water into a small flask or large test-tube, add 0.25 ml (5 drops) of amminocuprate solution and 1 drop of indicator solution and mix. Add nearly neutral sample solution slowly from a graduated pipette, continuously shaking the vessel until the colour changes from blue to violet, which corresponds to the presence of about 0.1 mg of citric acid in the volume of sample solution added. A colour change to pink indicates the presence of larger amounts of citric acid. Repeat the estimation with a more dilute sample solution if a colour change is observed after addition of only a few drops of sample. Interfering substances must be removed beforehand. Determinations. (A) Transfer an amount of sample solution containing l-8 mg of citric acid into an Erlenmeyer flask and dilute it to about 100 ml. Add exactly 5 ml of O.OlM copper sulphate, 2 drops of indicator solution, and if necessary O.lM ammonia or hydrochloric acid until the colour turns blue (pH 6.8-7.6). Add 5 ml of buffer solution and titrate the excess of copper with 0.01 M EDTA until the colour changes to pink. (B) Transfer an aliquot of sample solution containing 0.2-3 mg of citric acid into an Erlenmeyer flask and dilute it to about 50 ml. Add exactly 2 ml of O.OlM copper sulphate, 1 drop of indicator solution, and if necessary 0. I M
ammonia or hydrochloric acid until the colour becomes blue (uH .. 6.7-7.6). Add 2 ml of buffer solution and titrate the excess of copper with O.OOlM EDTA until changes to violet, then slowly until it becomes
RESULTS AND
the colour pink.
DISCUSSION
Interferences
Only a few of the organic acids present in natural products interfere in the detection, estimation and determination of citrate ions by the procedures above. Up to 1000 ppm of tartaric, succinic, malic, hydroxyacetic, lactic, salicylic and quinic acids have no influence on the precision, nor do phosphoric or
154
SHORT
COMMUNICATIONS
nitric acids or univalent metal ions. Determinations performed in the presence of up to 5 g of glucose and up to 20 ml of ethanol in 100 ml of sample solution showed no bias. Interference by reducing acids such as ascorbic, oxalic, fumaric and sulphurous was prevented by the addition of O.lN permanganate. Metal ions were removed by treatment with a cationexchange resin. Interference from amino-acids was eliminated by addition of sodium or potassium nitrite to acidified solutions. The reliability and precision were determined by repeated analysis. An accurately prepared 20-mg/ml citric acid stock solution was prepared with anhydrous citric acid dried at 110” to constant weight; a lo-ml aliquot of this solution was diluted, neutralized and made up to 200 ml: 1 ml = 1 mg of citric acid. Various mixtures were also analysed. Sample I. A 5-ml aliquot of the 1-mg/ml solution was diluted to about 100 ml and analysed by procedure (A). In 5 determinations an average of 2.40 ml of EDTA solution was used for titration of the excess of copper, with a maximum deviation of kO.02 ml (0.8%). The amount of citric acid was calculated from citric acid = (5 - a) x 1.92 mg where a = ml of O.OlM EDTA used (found: 4.99 mg). A 5-ml aliquot of the 1-mg/ml solution was diluted accurately to 100 ml and a lo-ml portion of this solution was titrated with O.OOlM EDTA by procedure (B). In 5 determinations an average of 17.40 ml of EDTA was used, with a maximal deviation of fO.1 ml (0.6%). The amount of citric acid was from citric acid = (20 - a) x 0.192 calculated mg = 0.499 mg. Sample II. Citric acid, 0.02 g; malic acid, 0.50 g; tartaric acid, 1.00 g; glucose 10.0 g; ethanol, 20 ml; water to 100 ml. Samples of 10, 25, and 50 ml were taken. The lo-ml sample was diluted to about 50 ml, treated by procedure B, and titrated with O.OOlM EDTA (14.9 ml). Citric acid found =0.98 mg. The 25- and 50-ml samples were diluted to about 100 ml and titrated with O.OlM EDTA. Citric acid found =2.53 mg (25-ml sample) and 4.90 mg (50 ml). Sample III. Citric acid, 0.05 g; calcium chloride, 1.O g; magnesium chloride, 0.50 g; urea, 5.0 g; water to 100 ml. A 50-ml portion was treated with Amberlite IR 120 cation-exchange resin, neutralized, and diluted to 100 ml. Three aliquots (25 ml each) were diluted to about 100 ml, treated by procedure A and titrated with O.OlM EDTA (1.76, 1.80 and 1.78 ml). Citric acid found =6.18 mg, corresponding to 49.4 mg in 100 ml of sample III. Sample IV. Citric acid, 0.50 g; ascorbic acid, 0.25 g; oxalic acid, 0.05 g; water to 100 ml. Ten ml were
diluted to about 50 ml and 1 ml of 0.1 N hydrochloric acid was added, and then 0.1 N permanganate (dropwise). The excess of permanganate was removed by adding 1 drop of 0.1 N thiosulphate. The solution was then treated with the cation-exchange resin, neutralized, and diluted to about 100 ml, treated by procedure A, and titrated with O.OlM EDTA (2.40, 2.42 and 2.42 ml). Citric acid found =4.79 mg, corresponding to 0.479 g in sample IV. Sample V. Citric acid, 0.50 g; lactic acid, 5.0 g; glutamic acid, 1.O g; water to 100 m. To 25 ml of this solution 5 ml of concentrated hydrochloric acid and 0.5 g of sodium nitrite were added. The solution was mixed and heated to about 70” until no more gas was evolved. The neutralized solution was diluted to 100 ml. Three 5-ml aliquots of the sample solution were diluted to about 100 ml, treated by procedure A, and titrated with O.OlM EDTA (1.74, 1.72 and 1.74 ml). Citric acid found =6.28 mg, corresponding to 0.502 g in sample V. Though there are several indicators for complexometric determination of copper( which can also be used in our method, the indicator recommended has several advantages. Murexide, though widely used, is less stable, the colour change from yellow to violet is less sensitive, and it is more difficult to maintain the desired pH. The nitrosochromotropic acid indicator”.” gives a colour change comparable to that of our indicator, but its solution, which has to be freshly prepared for each assay, is less stable. Also, two different pH ranges were recommended by the authors (pH 7.25-8.80 and 5.8-6.5) without explanation. REFERENCES
1. E. Szekely, M. Flitman
2. 3. 4. 5. 6.
7. 8.
and M. Leon, Proc. Israel Chem. Sot., 42nd Meeting, 1972, 94. G. Graffman, H. Domels and M. L. Strater, Fetten, Seifen, Anstrich., 1974, 76, 218. M. Hamon, A. Hoppendt and M. Guemet, Ann. Pharm. Franc., 1972, 30, 595. M. F. El-Taras and E. Pungor, Anal. Chim. Acta, 1976, 82, 285. A. Olin and B. Wallen, ibid., 1983, 151, 65. A. Pierre and S. Braley, Lair, 1983, 63, 623. H. L. Nours, J. F. Le Meur and C. Bourgeois, Sci. Aliments, 1982, 2, 483. M. T. M. Zaki and R. Alquasmi, Z. Anal. Chem., 1981,
306,400. 9. E. Szekely, Proc. Annual Meeting Israel Chem. Sot., 41st Meeting, 1971, 13. 10. S. Schwarzenbach and H. Flaschka, Complexometric Titrations, 2nd Ed., Methuen, London, 1969. 11. C. S. Panda and T. S. Srivastava, Z. Anal. Chem., 1962, 184, 248. 12. A. B. Sen and T. S. Srivastava, ibid., 1962, 187, 401.