J . Sci. Fd Agric. 1975, 26,879-885
Spectrophotometric Estimation of Benzoic and Sorbic Acids H. Zonneveld Sprengev Instituut, Wageningen, The Netherlands (Manuscript received 2 July 1974 and accepted 19 September 1974)
A spectrophotometricmethod is described for estimations of benzoic and sorbic acids in materials with background absorption in the ultraviolet region. The estimation can be done when both preservatives are present in equal or nonequal ratios. The coefficients of variation (average standard deviation) are 0.231.98% for benzoic acid and 0.40-1.58 % for sorbic acid. The recovery factors are 95.5-103% for benzoic acid and 81.5-89% for sorbic acid.
1. Introduction It is known that after extraction with ether some natural substances of biological material may interfere with the spectrophotometric estimation of benzoic acid and to lesser degree with that of sorbic acid. This inconvenience was found in our laboratory during the determination of benzoic acid in apple juice and fresh sauerkraut. Likewise, Gantenbein and Karaszl obtained results too high when estimating low concentrations of benzoic acid (200 pg/ml) and sorbic acid (40-50 ,ug/ml) in orange and apple juices. This was probably due to the background absorption of the juices. Furthermore the method described by Roos and Versne12 (isoabsorption point) could be used only when there was an equal ratio between both acids. Therefore a method was sought that would eliminate background absorption and allow estimations to be made of these preservatives, whether they were present in equal or non-equal ratios.
2. Experimentala 2.1. Apparatus and reagents 1. Spectrophotometer Beckman, DB-GT, and Beckman Ten Inch Lab. Pot. Recorder. 2. Centrifuge tubes of pyrex glass, 10 ml calibrated to an accuracy of 0.1 ml, adapted, to be closed hermetically with a lockable screw cap provided on the inside with a disc (Figure 1). This disc prevents leakage of the mixture of diethyl ether and petroleum ether (MDEPE, 6). Other plastic material was not used since plasticisers would be extracted. 3. Centrifuge tubes of pyrex glass, 10 ml, calibrated to an accuracy of 0.1 ml without screw cap to be closed with polyethylene caps. a Trade names mentioned are given for information only and do not imply endorsement either by the author or by the Institute. 1 879
H. Zonneveld
880
Teflon disc Lockable screw cop
Figure 1. Details of the centrifuge tube (2) described in section 2.1. (Apparatus and reagents).
4. Benzoic acid solutions: (a) stock solution: Benzoic acid p.a. Merck (50 mg) was dissolved in absolute ethanol p.a. Merck (50 ml); and (b) standard solutions: the stock solution was diluted 100 times with absolute ethanol, i.e. 10 pg/ml. One ml (10 pg) and 2 ml(20 pg) of the diluted stock solution were separately poured into a calibrated centrifuge tube (3) and made up to a final volume of 5.0ml with absolute ethanol p.a. They constituted the standard solutions. 5. Sorbic acid solutions. Stock and standard solutions were prepared as in (4). 6. A mixture of diethyl ether and petroleum ether (MDEPE). Diethyl ether, p.a. Merck was mixed with petroleum ether (40-60 "C)p.a. Merck (1 : 1 v/v). 7. Diluted sulphuric acid: 95-97 % sulphuric acid, p.a. Merck, was diluted five times with distilled water. 8. Oxidation mixture: 3.50 ml of distilled water, 1.50 ml of 95-97 % sulphuric acid diluted 1.5 times with distilled water, and 1.50 ml of potassium dichromate solution (3.4 g in 100 ml distilled water) were mixed successively into a centrifuge tube (2) before use. After that, the liquid was brought to room temperature by holding the tube for a few minutes in a beaker with tap water.
2.2. Preparation of standard curve Spectra of the standard solutions of benzoic and sorbic acid were determined relative
Estimation of benzoic and sorbic acids
881
to absolute ethanol p.a. Merck. Quartz cuvettes, 10 mm light path with grooved cover plates of Teflon were used (Hellma, No. 100, West Germany). The X max for benzoic acid was at 225 nm and for sorbic acid at 255 nm. The absorptions obtained at 225 nm were respectively for 10 and 20 pg benzoic acid in 5 ml ethanol 0.184 and 0.368. The absorptions at 255 nm for 10 and 20 pg sorbic acid were respectively 0.480 and 0.960. A straight line may be observed, of the absorption values as a function of the above mentioned concentrations, i.e. the Lambert-Beer law was valid between these absorption values. All analyses were arranged so that they would fall within the said straight line.
2.3. Preparation of samples Sauerkraut: A representative part (200 g) of the substrate was pressed (Braun Multipress MP 50, West Germany). The sauerkraut juice obtained (1 10 ml) constituted the sample-solution. Extra jam (strawberry): The substrate was homogenised in a Waring blendor. A part of the homogenate (40g) was diluted with distilled water to 100ml. The filtrate constituted the sample-solution. Apple juice: The juice constituted the sample-solution. For determining recovery factors benzoic (250 pg) and sorbic (500 pg) acids were added as sodium salts to 5 ml of the sample solutions.
2.4. Procedure Sample solution (5.0ml) was poured into a calibrated centrifuge tube (2). After acidification with 0.4 ml of diluted sulphuric acid (7, 5.0 ml of MDEPE (6) were added. The tube was hermetically closed and shaken for 25 min at 150 strokes/min (Mechanical horizontal shaking machine, platform type Edmund Biihler, type SM2, Tubingen, West Germany), and centrifuged at 1600 g for 15 min at room temperature. Afterwards the water layer was quickly removed with a long pipette. Immediately, half the amount of MDEPE extract (about 2.0 ml) was carefully poured into a centrifuge tube (2) containing about 150 mg sodium sulphate anhydrous and tightly closed. This tube contained untreated MDEPE extract. The other half (about 2.0ml) of the MDEPE extract was transferred to another centrifuge tube (2) containing 6.5 ml of freshly prepared oxidation mixture (8). After closing the tube tightly the suspension was shaken for 90 min a t 90-96 strokes/min. Afterwards the water layer was carefully removed and the MDEPE extract poured into another tube (2) containing 150mg sodium sulphate anhydrous and closed hermetically. This last tube contained oxidised MDEPE extract. After drying the oxidised MDEPE extract (as well as the untreated MDEPE extract) for a few minutes by swivelling gently, 0.20 ml of the dehydrated solution were poured into a centrifuge tube (3) and evaporated with a cold air stream. The residue was dissolved in absolute ethanol (5.0 ml) and the spectrum recorded relative to absolute ethanol. The spectrum of the untreated MDEPE extract was used for the estimation of sorbic acid and the spectrum of the oxidised MDEPE extract for benzo-ic acid.
H. Zonneveld
882
2.5. Calculation Using 0.20 ml of dehydrated MDEPE extract: For benzoic acid: Apple juice : Extra jam:
A 50 x __ 0.184-pg’m1 A
125 x _ _--Pglg 0.184
A Sauerkraut: 0 . 2 5 ~x -- = pglg 0.184
For sorbic acid: The calculations were the same as for benzoic acid but the number 0.184 must be replaced by 0.480. A for sorbic acid signifies the absorption of the h max pertaining to the spectrum of the oxidised MDEPE extract (at 255-253 nm); A for benzoic acid signifies the absorption of the h max pertaining to the spectrum of the oxidised MDEPE extract (at 225-223nm); a signifies the number of ml sauerkraut juice obtained after pressing 200 g of sauerkraut; 0.184 signifies the absorption value at 225 nm of the standard solution (10 pg/5 ml absolute ethanol) of benzoic acid; 0.480 signifies the absorption value at 255 nm of the standard solution (10 pg/5 ml absolute ethanol) of sorbic acid. Factor 50 was obtained as follows: 1 5.0 ml MDEPE extract ( = pg/ml apple juice) x 10 pgx5 ml apple juice 0.20 ml MDEPE extract
Factor 125 was obtained as follows: 5.0 ml MDEPE extract 0.20 ml MDEPE extract
pg
--
100 ml suspension 1 _____ x (= pg/g jam) 5 ml sample soln. 40 g jam
Factor 0 . 2 5 ~was obtained as follows :
a ml sample soln. 1 ___ 5.0 ml MDEPE extract X x 10 pg x ____--__ 5 ml sample soln. 200 g sauerkraut 0.20 ml MDEPE extract (= pg/g sauerkraut) 3. Results and discussion
Table 1 shows that the results with a recovery factor of 95.5-103.0% for benzoic acid were satisfactory. A recovery factor of 8 1.5-89.0 % for sorbic acid was accepted, as a part was lost probably by autoxidation or oxidation by other substances.3~4 The additions of benzoic and sorbic acids as sodium salts were made to the sample solutions, i.e. before acidification and extraction. This meant that the substances were added practically at the start of the procedure. In other words the recovery factors are representative for the whole method. According to the Dutch Jam and Lemonade Decree no benzoic or sorbic acid may be added to apple juice, fresh sauerkraut and “extra” jam (Gersons, personal
Average of six estimations. Average of four estimations.
100.0 27.5 125.0
Expected
X
0.69 0.23 1.98
s, k CV, f
96.Sa 1.70 27.4* 0.47 1 2 3 9 3.96
2
s=(Cd2/n- l)ll2; CV (coefficient of variation)= : x 100 (%).
b
a
Apple juice (pglml) Sauerkraut (pg/g) Strawberry jam (pg/g)
Sample
Found
Benzoic acid
96.8 99.8 99.0
95.5-100.0 98.0-102.0 96.0-103.0
100.0 55.0 125.0
84.8a 2.44 46.9’ 0.79 106.6b 3.15
s,
X&
( %>
(%) Expected
Found
Extreme values of recovery
Mean recovery
Recovery factor
(%) 84.8 85.3 85.3
1.00 0.40 1.58
Mean recovery
81.5-87.0 83.5-87.0 84.0-89.0
Extreme values of recovery ( %)
Recovery factor
CV, If:
Sorbic acid
Table 1. Estimation and recovery factors of benzoic and sorbic acids in various samples. Both compounds were together in the sample
H. Zonneveld
884
X (nm)
Figure 2. Extraction by MDEPE (5.0 rnl) of apple or sauerkraut solutions (5.0 nil). MDEPE-extract (0.20ml) was evaporated and the residue dissolved in 5 ml of absolute ethanol and spectrum recorded in respect of absolute ethanol. A, Apple juice (or sauerkraut juice). Untreated MDEPE-extract. Note background at 240-200 nm. B, Apple juice (or sauerkraut juice). Oxidised MDEPE-extract. Interfering background was removed. The same as above but to apple juice benzoic and sorbic acids were added in a proportion of 1 : 1 o r 2 : 1: C, Untreated MDEPE-extract. Sorbic acid (20 pg) at 255 nm. D, Oxidised MDEPE-extract. Benzoic acid (20 pg) at 227 nm. E, Untreated MDEPE-extract. Sorbic acid peak (7.5 pg) at 255 nm. F, Oxidised MDEPE-extract. Benzoic acid (15 pg) at 226 nm.
A
0.00
0.4 0
/
\
0
o*20* 200
220-
240
260
280
300
320
A (nrn)
Figure 3. The same as Figure 2 but to sauerkraut juice benzoic and sorbic acids were added in a ratio of 1 : 2. A, Untreated MDEPE-extract. Sorbic acid peak (20 pg) at 255 nm. B, Oxidised MDEPE-extract. Benzoic acid peak (10 pg) at 227 nm.
Estimation of benzoic and sorbic acids
885
communication). However these products were employed because they have a background in the ultraviolet region (240-210 nm), caused mainly by plienolic compounds.5 Likewise benzoic acid is strongly absorbed in this region. By using the oxidation mixture described here, the background was removed and the main part of sorbic acid was oxidised (Figures 2 and 3). Benzoic acid remained intact for 30 min after the oxidation and its maximal absorption at 225 nm could be measured. In addition the diagrams given in this text, show that benzoic and sorbic acids can be estimated whether present in equal or unequal ratios. Furthermore the known interference of vanillin1 with the spectrophotometric estimation of benzoic acid could be avoided by the oxidation treatment described. To apple juice (5.0 ml), vanillin, sorbic and benzoic acid (250 pg of each), were added. The oxidation process removed vanillin and other interfering substances, allowing the estimation of benzoic acid (recovery factor 100.5-104.3 %) and sorbic acid (recovery factor 85.8-88.3 %).
Acknowledgements The author thanks Dr N. Gorin and Ir W. Klop for valuable discussions.
References 1. Gantenbein, W. M.; Karasz, J. J. Ass. O f . analyt. Chem. 1969,52, 738. 2. Roos, J. B.; Versnel, A. Dt. Lebensmitt. Rdsch. 1960, 56, 128. 3. Pekkarinen, L. Z. Leber2smittel-unters. u. -Fousch. 1968, 139,23. 4. Rajama, J.; Makela, P. J. Chromat. 1973, 76, 199. 5 . Harborne, J. B. In Methods in Polypheno1 Chemistry, 1964, p. 13 (Pridham, J. B., Ed.) Oxford, Pergamon Press.
Spectrophotometric Estimation of Benzoic and Sorbic Acids H. Zonneveld Sprengev Instituut, Wageningen, The Netherlands (Manuscript received 2 July 1974 and accepted 19 September 1974)
A spectrophotometricmethod is described for estimations of benzoic and sorbic acids in materials with background absorption in the ultraviolet region. The estimation can be done when both preservatives are present in equal or nonequal ratios. The coefficients of variation (average standard deviation) are 0.231.98% for benzoic acid and 0.40-1.58 % for sorbic acid. The recovery factors are 95.5-103% for benzoic acid and 81.5-89% for sorbic acid.
1. Introduction It is known that after extraction with ether some natural substances of biological material may interfere with the spectrophotometric estimation of benzoic acid and to lesser degree with that of sorbic acid. This inconvenience was found in our laboratory during the determination of benzoic acid in apple juice and fresh sauerkraut. Likewise, Gantenbein and Karaszl obtained results too high when estimating low concentrations of benzoic acid (200 pg/ml) and sorbic acid (40-50 ,ug/ml) in orange and apple juices. This was probably due to the background absorption of the juices. Furthermore the method described by Roos and Versne12 (isoabsorption point) could be used only when there was an equal ratio between both acids. Therefore a method was sought that would eliminate background absorption and allow estimations to be made of these preservatives, whether they were present in equal or non-equal ratios.
2. Experimentala 2.1. Apparatus and reagents 1. Spectrophotometer Beckman, DB-GT, and Beckman Ten Inch Lab. Pot. Recorder. 2. Centrifuge tubes of pyrex glass, 10 ml calibrated to an accuracy of 0.1 ml, adapted, to be closed hermetically with a lockable screw cap provided on the inside with a disc (Figure 1). This disc prevents leakage of the mixture of diethyl ether and petroleum ether (MDEPE, 6). Other plastic material was not used since plasticisers would be extracted. 3. Centrifuge tubes of pyrex glass, 10 ml, calibrated to an accuracy of 0.1 ml without screw cap to be closed with polyethylene caps. a Trade names mentioned are given for information only and do not imply endorsement either by the author or by the Institute. 1 879
H. Zonneveld
880
Teflon disc Lockable screw cop
Figure 1. Details of the centrifuge tube (2) described in section 2.1. (Apparatus and reagents).
4. Benzoic acid solutions: (a) stock solution: Benzoic acid p.a. Merck (50 mg) was dissolved in absolute ethanol p.a. Merck (50 ml); and (b) standard solutions: the stock solution was diluted 100 times with absolute ethanol, i.e. 10 pg/ml. One ml (10 pg) and 2 ml(20 pg) of the diluted stock solution were separately poured into a calibrated centrifuge tube (3) and made up to a final volume of 5.0ml with absolute ethanol p.a. They constituted the standard solutions. 5. Sorbic acid solutions. Stock and standard solutions were prepared as in (4). 6. A mixture of diethyl ether and petroleum ether (MDEPE). Diethyl ether, p.a. Merck was mixed with petroleum ether (40-60 "C)p.a. Merck (1 : 1 v/v). 7. Diluted sulphuric acid: 95-97 % sulphuric acid, p.a. Merck, was diluted five times with distilled water. 8. Oxidation mixture: 3.50 ml of distilled water, 1.50 ml of 95-97 % sulphuric acid diluted 1.5 times with distilled water, and 1.50 ml of potassium dichromate solution (3.4 g in 100 ml distilled water) were mixed successively into a centrifuge tube (2) before use. After that, the liquid was brought to room temperature by holding the tube for a few minutes in a beaker with tap water.
2.2. Preparation of standard curve Spectra of the standard solutions of benzoic and sorbic acid were determined relative
Estimation of benzoic and sorbic acids
881
to absolute ethanol p.a. Merck. Quartz cuvettes, 10 mm light path with grooved cover plates of Teflon were used (Hellma, No. 100, West Germany). The X max for benzoic acid was at 225 nm and for sorbic acid at 255 nm. The absorptions obtained at 225 nm were respectively for 10 and 20 pg benzoic acid in 5 ml ethanol 0.184 and 0.368. The absorptions at 255 nm for 10 and 20 pg sorbic acid were respectively 0.480 and 0.960. A straight line may be observed, of the absorption values as a function of the above mentioned concentrations, i.e. the Lambert-Beer law was valid between these absorption values. All analyses were arranged so that they would fall within the said straight line.
2.3. Preparation of samples Sauerkraut: A representative part (200 g) of the substrate was pressed (Braun Multipress MP 50, West Germany). The sauerkraut juice obtained (1 10 ml) constituted the sample-solution. Extra jam (strawberry): The substrate was homogenised in a Waring blendor. A part of the homogenate (40g) was diluted with distilled water to 100ml. The filtrate constituted the sample-solution. Apple juice: The juice constituted the sample-solution. For determining recovery factors benzoic (250 pg) and sorbic (500 pg) acids were added as sodium salts to 5 ml of the sample solutions.
2.4. Procedure Sample solution (5.0ml) was poured into a calibrated centrifuge tube (2). After acidification with 0.4 ml of diluted sulphuric acid (7, 5.0 ml of MDEPE (6) were added. The tube was hermetically closed and shaken for 25 min at 150 strokes/min (Mechanical horizontal shaking machine, platform type Edmund Biihler, type SM2, Tubingen, West Germany), and centrifuged at 1600 g for 15 min at room temperature. Afterwards the water layer was quickly removed with a long pipette. Immediately, half the amount of MDEPE extract (about 2.0 ml) was carefully poured into a centrifuge tube (2) containing about 150 mg sodium sulphate anhydrous and tightly closed. This tube contained untreated MDEPE extract. The other half (about 2.0ml) of the MDEPE extract was transferred to another centrifuge tube (2) containing 6.5 ml of freshly prepared oxidation mixture (8). After closing the tube tightly the suspension was shaken for 90 min a t 90-96 strokes/min. Afterwards the water layer was carefully removed and the MDEPE extract poured into another tube (2) containing 150mg sodium sulphate anhydrous and closed hermetically. This last tube contained oxidised MDEPE extract. After drying the oxidised MDEPE extract (as well as the untreated MDEPE extract) for a few minutes by swivelling gently, 0.20 ml of the dehydrated solution were poured into a centrifuge tube (3) and evaporated with a cold air stream. The residue was dissolved in absolute ethanol (5.0 ml) and the spectrum recorded relative to absolute ethanol. The spectrum of the untreated MDEPE extract was used for the estimation of sorbic acid and the spectrum of the oxidised MDEPE extract for benzo-ic acid.
H. Zonneveld
882
2.5. Calculation Using 0.20 ml of dehydrated MDEPE extract: For benzoic acid: Apple juice : Extra jam:
A 50 x __ 0.184-pg’m1 A
125 x _ _--Pglg 0.184
A Sauerkraut: 0 . 2 5 ~x -- = pglg 0.184
For sorbic acid: The calculations were the same as for benzoic acid but the number 0.184 must be replaced by 0.480. A for sorbic acid signifies the absorption of the h max pertaining to the spectrum of the oxidised MDEPE extract (at 255-253 nm); A for benzoic acid signifies the absorption of the h max pertaining to the spectrum of the oxidised MDEPE extract (at 225-223nm); a signifies the number of ml sauerkraut juice obtained after pressing 200 g of sauerkraut; 0.184 signifies the absorption value at 225 nm of the standard solution (10 pg/5 ml absolute ethanol) of benzoic acid; 0.480 signifies the absorption value at 255 nm of the standard solution (10 pg/5 ml absolute ethanol) of sorbic acid. Factor 50 was obtained as follows: 1 5.0 ml MDEPE extract ( = pg/ml apple juice) x 10 pgx5 ml apple juice 0.20 ml MDEPE extract
Factor 125 was obtained as follows: 5.0 ml MDEPE extract 0.20 ml MDEPE extract
pg
--
100 ml suspension 1 _____ x (= pg/g jam) 5 ml sample soln. 40 g jam
Factor 0 . 2 5 ~was obtained as follows :
a ml sample soln. 1 ___ 5.0 ml MDEPE extract X x 10 pg x ____--__ 5 ml sample soln. 200 g sauerkraut 0.20 ml MDEPE extract (= pg/g sauerkraut) 3. Results and discussion
Table 1 shows that the results with a recovery factor of 95.5-103.0% for benzoic acid were satisfactory. A recovery factor of 8 1.5-89.0 % for sorbic acid was accepted, as a part was lost probably by autoxidation or oxidation by other substances.3~4 The additions of benzoic and sorbic acids as sodium salts were made to the sample solutions, i.e. before acidification and extraction. This meant that the substances were added practically at the start of the procedure. In other words the recovery factors are representative for the whole method. According to the Dutch Jam and Lemonade Decree no benzoic or sorbic acid may be added to apple juice, fresh sauerkraut and “extra” jam (Gersons, personal
Average of six estimations. Average of four estimations.
100.0 27.5 125.0
Expected
X
0.69 0.23 1.98
s, k CV, f
96.Sa 1.70 27.4* 0.47 1 2 3 9 3.96
2
s=(Cd2/n- l)ll2; CV (coefficient of variation)= : x 100 (%).
b
a
Apple juice (pglml) Sauerkraut (pg/g) Strawberry jam (pg/g)
Sample
Found
Benzoic acid
96.8 99.8 99.0
95.5-100.0 98.0-102.0 96.0-103.0
100.0 55.0 125.0
84.8a 2.44 46.9’ 0.79 106.6b 3.15
s,
X&
( %>
(%) Expected
Found
Extreme values of recovery
Mean recovery
Recovery factor
(%) 84.8 85.3 85.3
1.00 0.40 1.58
Mean recovery
81.5-87.0 83.5-87.0 84.0-89.0
Extreme values of recovery ( %)
Recovery factor
CV, If:
Sorbic acid
Table 1. Estimation and recovery factors of benzoic and sorbic acids in various samples. Both compounds were together in the sample
H. Zonneveld
884
X (nm)
Figure 2. Extraction by MDEPE (5.0 rnl) of apple or sauerkraut solutions (5.0 nil). MDEPE-extract (0.20ml) was evaporated and the residue dissolved in 5 ml of absolute ethanol and spectrum recorded in respect of absolute ethanol. A, Apple juice (or sauerkraut juice). Untreated MDEPE-extract. Note background at 240-200 nm. B, Apple juice (or sauerkraut juice). Oxidised MDEPE-extract. Interfering background was removed. The same as above but to apple juice benzoic and sorbic acids were added in a proportion of 1 : 1 o r 2 : 1: C, Untreated MDEPE-extract. Sorbic acid (20 pg) at 255 nm. D, Oxidised MDEPE-extract. Benzoic acid (20 pg) at 227 nm. E, Untreated MDEPE-extract. Sorbic acid peak (7.5 pg) at 255 nm. F, Oxidised MDEPE-extract. Benzoic acid (15 pg) at 226 nm.
A
0.00
0.4 0
/
\
0
o*20* 200
220-
240
260
280
300
320
A (nrn)
Figure 3. The same as Figure 2 but to sauerkraut juice benzoic and sorbic acids were added in a ratio of 1 : 2. A, Untreated MDEPE-extract. Sorbic acid peak (20 pg) at 255 nm. B, Oxidised MDEPE-extract. Benzoic acid peak (10 pg) at 227 nm.
Estimation of benzoic and sorbic acids
885
communication). However these products were employed because they have a background in the ultraviolet region (240-210 nm), caused mainly by plienolic compounds.5 Likewise benzoic acid is strongly absorbed in this region. By using the oxidation mixture described here, the background was removed and the main part of sorbic acid was oxidised (Figures 2 and 3). Benzoic acid remained intact for 30 min after the oxidation and its maximal absorption at 225 nm could be measured. In addition the diagrams given in this text, show that benzoic and sorbic acids can be estimated whether present in equal or unequal ratios. Furthermore the known interference of vanillin1 with the spectrophotometric estimation of benzoic acid could be avoided by the oxidation treatment described. To apple juice (5.0 ml), vanillin, sorbic and benzoic acid (250 pg of each), were added. The oxidation process removed vanillin and other interfering substances, allowing the estimation of benzoic acid (recovery factor 100.5-104.3 %) and sorbic acid (recovery factor 85.8-88.3 %).
Acknowledgements The author thanks Dr N. Gorin and Ir W. Klop for valuable discussions.
References 1. Gantenbein, W. M.; Karasz, J. J. Ass. O f . analyt. Chem. 1969,52, 738. 2. Roos, J. B.; Versnel, A. Dt. Lebensmitt. Rdsch. 1960, 56, 128. 3. Pekkarinen, L. Z. Leber2smittel-unters. u. -Fousch. 1968, 139,23. 4. Rajama, J.; Makela, P. J. Chromat. 1973, 76, 199. 5 . Harborne, J. B. In Methods in Polypheno1 Chemistry, 1964, p. 13 (Pridham, J. B., Ed.) Oxford, Pergamon Press.
