Analyst, May, 1978, Vol. 103, pp. 521-524 52 1 Analytical Met hods Com mittee REPORT PREPARED BY THE IRON SUB-COMMITTEE General Method for the Determination of Iron with 4,7-Diphenyl-l,lO-phenanthroline (Bathophenanthroline) Keywords Iron determination ; 4,7-diphenyl-l, 10-phenanthroline ; batho- phenanthroline ; spectrophotovnetry The Analytical Methods Committee has received and approved for publication the following Report from its Iron Sub-committee. Report The constitution of the Sub-committee responsible for the preparation of this Report was : Mr. A. G. Hill (Chairman), Professor E. Bishop, Dr. L. E. Coles, Dr. E. J. McLauchlan, Mr. D. W. Meddle, Mr. M. J. Pater, Mr. C. A. Watson and Mr. C. Whalley, with Mr. P. W. Shallis as Secretary. Introduction The Iron Sub-committee of the Analytical Methods Committee has earlier recommended a general method for the determination of iron with 1,lO-phenanthro1ine.l The work was carried out at the request of the British Standards Institution, acting on behalf of the International Standards Organisation (ISO), who needed a general method that could be used in all specifications that require the determination of iron content.After considering the available reagents and methods, the Sub-committee concluded that the use of 1,lO- phenanthroline, a reagent already widely used in IS0 specifications for the determination of iron, offered adequate sensitivity for the purpose. A standardised version of this method was evaluated and was recommended to IS0 for use as a general method. The Sub-committee then proceeded to the second phase of its programme, which was to carry out investigations that would lead to the recommendation of the best general method for determining iron.Several reagents are available that will form coloured complexes with iron that can be extracted into an organic solvent, and of these perhaps the best known and most widely used is 4,7-diphenyl-l , 10-phenanthroline (bathophenanthroline) . This reagent was recommended for the determination of iron by Smith et aL2 and, since its intro- duction, has been proposed by many different workers for determining iron in a variety of sample materials. The bright red complex of bathophenanthroline with iron(II), which is formed in the pH range 2-9, has a molar absorptivity of about 22 000, which is approximately twice that of the 1,lO-phenanthroline - iron(I1) complex in aqueous solution. Moreover, as the iron(I1) - bathophenanthroline complex can be extracted into an organic solvent and concentrated, it offers considerable advantages in terms of sensitivity over the use of 1 , 10-phenanthroline.Interferences in the method are few and the major problems that can be encountered have been summarised previ~usly.~ The Sub-committee therefore decided to investigate the bathophenanthroline method as it was so widely used and recommended. It was realised that other reagents, such as the disodium salt of 4,7-diphenyl-l,1O-phenanthrolinedisulphonic acid, FerroZine and 1,3,5- triazine, had been recommended for the determination of iron, but that these were, a t best, unlikely to be appreciably better than bathophenanthroline and were in any event much less widely available.Atomic-absorption spectrophotometry is another technique that it was considered could possibly provide a general method for determining iron, and some work done by the Sub- committee is the subject of a separate r e p ~ r t . ~522 ANALYTICAL METHODS COMMITTEE : DETERMINATION OF IRON Analyst, VoZ. 103 Experimental The bathophenanthroline method as recommended by Cluley and Newman3 was selected by the Sub-committee for investigation, except that ascorbic acid was to be used as the reducing agent as it can be obtained with a lower iron content that can hydroxylammonium chloride. It was suggested to the Sub-committee that propylene carbonate was a satisfactory alternative solvent to chloroform for the iron(I1) - bathophenanthroline complex, and it was decided to investigate this at the same time.A collaborative test was arranged in which the nine solutions already distributed to the collaborators for use in the work on the 1,lO-phenanthroline methodl were to be used. Each laboratory was asked to carry out at least two determinations of the iron contents of the nine solutions by the method given in Appendix I and also by virtually the same method but with propylene carbonate as the extraction solvent instead of chloroform. As the use of chloroform is now restricted in some laboratories, two of the collaborators also carried out some work by the method given in Appendix I but with l,l,l-trichloroethane as the extraction solvent.Results arid Discussion The results of the determination of iron at three different levels in each of the three sample materials are shown in Table I and indicaie that extraction of the iron(I1) - bathophen- anthroline complex into chloroform provides a good general method for the determination of iron. Only one set of results is given in Table I for the work in which the iron(I1) - bathophenanthroline complex was to have been extracted into propylene carbonate, as nearly all members found this to be unsatisfactory under the conditions employed. Of the two laboratories that carried out some work in which the coloured complex was extracted into l,l,l-trichloroethane, one prepared only a calibration graph covering the range 5-60 pg of iron, which was satisfactory and from which it was concluded that the sensitivity when using l,l, 1-trichloroethane was very similar to that when using chloroform.The other TABLE I DETERMINATIONS OF IRON IN SOLUTIONS OF TECHNICAL MATERIALS BY THE RECOMMENDED METHOD USING BATHOPHENANTHROLINE AND EXTRACTION INTO CHLOROFORM Sample Aluminium sulphate Ammonium sulphate Laboratory 2 3 4 5 6 7 72 Intra-laboratory Inter-laboratory11 , 1 Value A B C Mean*/vg mi-' 2.185 3.802 6.708 R.S.D.,? yo 1.80 1.80 0.35 hleanlyg ml-l 2.247 4.008 7.036 R.S.D., % 1.56 1.50 0.68 Mean/vg ml-1 2.121 3.854 7.004 R.S.D., yo 5.17 3.16 4.25 Mean/vg ml-1 2.168 3.790 6.667 R.S.D., % 0.73 0.33 0.45 hlean/yg ml-I 2.104 3.825 6.712 R.S.D., yo 0.41 0.81 0.30 Mean/vg ml-l 2.045 3.834 6.834 R.S.D., yo 0.177 2.83 0.53 Mean/vg ml-1 2.258 8 4.052 5 7,183 5 R.S.D., yo 0.127 0.216 0.373 Mean/wg ml-I 2.247 4 4.069 2 7.205 0 R.S.D., Yo 0.251 0.094 0.132 Mean R.S.D.,§ % 1.48 1.34 0.88 Mean/vg ml-1 2.172 3.904 6.919 R.S.D., yo 3.58 3.01 3.14 , 1 A B C 1.505 3.984 5.418 2.50 0.89 0.57 1.500 4.102 5.595 0.94 0.40 0.99 1.487 3.992 5.456 3.57 3.29 4.60 1.496 3.907 5.356 2.27 0.09 0.33 1.553 3.956 5.502 0.74 0.24 0.73 1.547 3.939 5.602 1.90 0.92 1.12 1.554 1 4.138 7 5.590 5 0.237 0.062 0.177 1.572 4 4.174 7 5.665 9 0.081 0.527 0.113 1.53 0.80 1.08 1.524 4.024 5.511 2.07 2.49 1.69 Sodium tetraborate ---- A B C 1.301 2.742 5.678 0.48 2.33 1.20 1.172 2.925 5.914 0.54 0.43 0.92 1.148 2.583 5.712 3.70 6.80 3.40 1.112 2.745 5.686 1.70 0.65 0.16 1.169 2.806 5.698 1.50 0.26 0.65 1.258 2.769 5.623 0.0 0.26 0.65 1.197 3 2.788 2 5.907 7 0.188 0.082 0.064 1.220 0 2.835 3 5.996 3 0.131 0.111 0.296 1.03 1.39 0.87 1.197 2.774 5.777 5.10 3.50 2.41 Instru- ment H700 SP 1800 SP 500 SP 1750 SP 500 DRGT Cary 16 Cary 16 * Mean of three separate complete experiments.3 Extraction into propylene carbonate carried out in parallel with the chloroform method and a t the same time. The high photometric accuracy, wide scale, logarithmic amplification and close thermostatting (0.02 "C) of the Cary 16 instrument warrant the additional significant figure for laboratory 7. f The average of the relative standard deviations within individual laboratories. 11 The unweighted mean of all 24 results for each sample, with the over-all R.S.D. as dehed in the second footnote.No results have been rejected. Relative standard deviation expressed as a percentage = 100 x standard deviationlmean.May, 1978 WITH 4,7-DIPHENYL-1 ,lO-PHENANTHROLINE (BATHOPHENANTHROLINE) 523 laboratory applied the method with extraction of the coloured complex into 1,1, l-trichloro- ethane to the determination of iron in the nine circulated solutions; the results are given in Table 11. TABLE I1 DETERMINATION OF IRON IN SOLUTIONS OF TECHNICAL MATERIALS BY THE RECOMMENDED METHOD WITH EXTRACTION INTO 1 ,l , 1-TRICHLOROETHANE The figures in parentheses are the means from Table I. Iron found/pg ml-I , I Aluminium sulphate Ammonium sulphate Sodium tetraborate - I 7 A B C A B C A B 2.29 3.90 7.01 1.66 3.98 5.71 1.16 2.84 2.12 2.25 (2.17) (3.90) (6.92) (1.52) (4.02) (5.61) (1.20) (2.77) C 5.82 (5.78) Recommendation The Sub-committee recommends that the method given in Appendix I should be used for the determination of iron when the highest sensitivity is required.It is also recommended that if necessary chloroform can be replaced with confidence by l,l,l-trichloroethane as the extraction solvent. APPENDIX Recommended General Method for the Determination of Iron with Bathophenanthroline Object diphenyl-1 ,lo-phenanthroline (bathophenanthroline) is described. A general spectrophotometric method for determining iron involving the use of 4,7- Principle Iron(I1) forms a bright red complex with bathophenanthroline within the pH range 2-9. Any iron(II1) present is reduced to iron(I1) with ascorbic acid. The coloured complex is extracted into chloroform (or 1,1, l-trichloroethane) and its absorbance is measured at the maximum at about 533 nm.Reagents Ascorbic acid solution, 100 g 1-1. Acetate bufer solution, 1 M in acetate (see Note 1). Prepare freshly each week. Dissolve 68 g of sodium acetate tri- hydrate and 28.6 ml of glacial acetic acid in 800 ml of water, adjust the pH to 4.6, if necessary, with sodium hydroxide solution or acetic acid, add 1 ml of ascorbic acid solution and dilute to 1 1 with water. Transfer the solution into a separating funnel, add 10 ml of batho- phenanthroline solution, mix and set aside for 15 min. Extract by shaking vigorously with 10-ml portions of chloroform successively until the solution is colourless. Discard the extracts, extract with two further 10-ml portions of chloroform and also discard the extracts.Store the solution in an iron-free glass or polyethylene container. Dissolve 0.332 g of 4,7-diphenyl- 1,lO-phenanthroline in 1 1 of absolute industrial methylated spirit. Store the solution in an iron-free glass bottle. Bathophenanthroline solution, approximately 0.001 M. Ethanol. Chloroform (or 1 ,1, l-trichloroethane) . Perchloric acid, 60% m/m. Ammonia solution, 5 M. Standard iron solution, 1 mg ml-l. Industrial methylated spirit, 74 0.p. Analytical-reagent grade. Dissolve 8.65 g of ammonium iron(II1) sulphate in 50 ml of concentrated nitric acid and dilute to 1 1 with water.524 ANALYTICAL METHODS COMMITTEE Dilute standard iron solution, 10 pg ml-l. Dilute 10.0 ml of the standard iron solution to 1 1 with water; this solution must be freshly prepared.Procedure Transfer 10ml of the sample solution containing between 0.25 and 1OOpg of iron (see Note 2) into a 100-ml Squibb’s-type separating funnel, add 1 ml of 60% m/m perchloric acid and 5ml of ascorbic acid solution, mix and add 10ml of bathophenanthroline solution. Set aside for 5 min, then add 2 ml of 5 M ammonia solution and 10 ml of buffer solution, mix and set aside for a further 10 min. Add 10 ml of chloroform (see Note 3), shake the funnel vigorously for 30 s, allow the layers to separate and run the chloroform layer into a 25-ml calibrated flask containing 1 ml of ethanol. Extract similarly with two further 5-ml portions of chloroform, run the extracts into the same 25-ml calibrated flask and dilute to the mark with ethanol.Measure the absorbance of this solution in a 10-mm cell at 533 nm against the reagent blank. Preparation of Calibration Graph Transfer appropriate portions of the dilute standard iron solution covering the range 0-100 pg of iron into separate Squibb’s-type separating funnels. Dilute the contents of each funnel to 10 ml with water and proceed with each as described for the sample, beginning a t “. . . add 1 ml of 60% m/m perchloric acid . . .” Construct a graph of iron content, in micrograms, against absorbance. NOTES- When necessary, the acetate buffer solution can be replaced by a citrate or tartrate buffer solution without further modification to the method. By altering the final volume and also the path length of the spectrophotometer cell used, the method can be applied to other ranges of iron contents, as indicated in Table 111. In each instance the figure in line (a) is the amount of iron, in micrograms, that will give an absorbance reading of about 0.004 and that in line (b) is the amount of iron, in micrograms, that will give an absorbance reading of about 1.6. 1. 2. TABLE I11 APPLICATION OF METHOD TO DIF:FERENT RANGES OF IRON CONTENTS Final volume/ml 5 10 25 50 100 5 (a) 0.1 0.2 0.5 1 .o 2.0 10 (a) 0.05 0.1 0.25 0.5 1 .o 7- A 3 Cell path length/mm (b) 40 $10 200 400 800 (b) 20 4,O 100 200 400 20 (a) 0.025 0.05 0.13 0.25 0.5 (b) 10 20 50 100 200 (b) 5 10 25 50 100 40 (a) 0.013 0.025 0.06 0.13 0.26 3. In all instances where chloroform is referred to in the method, it can be replaced if desired by 1,1,1-trichloroethane. References 1. 2. 3. 4. Analytical Methods Committee, Analyst, 1978, 103, 391. Smith, G. F., McCurdy, W. H., jun., and Diehl, H., Analyst, 1952, 77, 418. Cluley, H. J., and Newman, E. J., Analyst, 1963, 88, 3. Analytical Methods Committee, Analyst, 1978, 103, in the press.