140 Analyst, February, 1978, Vol. 103, $@. 140-148 Biacetyl Bis(4-phenyl-3-thiosemicarbazone) as a Reagent for the Spectrophotometric Determination of Copper A. G. Asuero and J. M. Cano Department of Analytical Chemistry, Faculty of Sciences, The University, Seville-4, Spain The synthesis, characteristics and analytical applications of biacetyl bis- (4-phenyl-3-thiosemicarbazone) (BBPT) are described. The reaction between copper(I1) and BBPT has been studied by spectrophotometry. The reddish orange 1 : 1 copper - BBPT complex (E = 12.7 x lo3 1 mol-1 cm-1 a t 485 nm and 8.2 x lo3 1 mol-1 cm-l at 530 nm) is formed at pH 1.8-11.9, in a solution containing 60% V/V of dimethylformamide. The effect of interferences was studied. A rapid and simple method for the spectrophotometric determination of copper in a white metal, in blende and in a waste water has beendevised.The method is compared with others proposed for the spectrophotometric determination of copper with thiosemicarbazone reagents. Keywords : Bzacetyl bis(4-phenyl-S-thiosemicarbaaone) reagent ; copper deter- mination ; sfiectrophotometry Thiosemicarbazones have been widely used for the spectrophotometric determination of metal ions and several papers have dealt with the use of dithiosemicarbazones as analytical reagents. The most studied have been derived from glyoxa1,l bipyridylglyoxal,2~3 cyclo- hexane-l,2-dione4-6 and ~yclohexane-l,3-dione.~ The introduction of the phenyl radical at the end of the thiosemicarbazide molecule is an excellent example of how group action in organic compounds can be modified to provide increased sensitivity ; the molar absorptivities of metal - thiosemicarbazone com- plexes are favourably influenced by this substitution, which also causes a shift of absorption peaks towards longer wavelengths.Some phenylthiosemicarbazones have been studied as reagents*-11 ; nevertheless, compounds with the bis(pheny1thiosemicarbazone) grouping have received very little attention. A search of the literature revealed that Niederschulte and Ballschmiter12 and Ball~chmiterl~ have studied some bis(pheny1thiosemicarbazones) derived from biacetyl and glyoxal, varying the phenyl substituent. However, the principal applica- tion reported by these workers for this type of reagent has been the separation of complex chelate mixtures by thin-layer chromatography on aluminium oxide or by liquid chromato- The use of biacetyl bis(4-phenyl-3-thiosemicarbazone) (BBPT) for the selective deter- mination of copper has been investigated, and this paper, which forms part of an investigation into the use of diphenylthiosemicarbazones as analytical reagents, describes the development of a simple procedure that has high sensitivity and selectivity.The determination of small amounts of copper in different materials is described. graphy. BBPT Experimental Synthesis of BBPT Phenylthiosemicarbazide (3.63 g) was dissolved in 100 ml of ethanol - water (1 + 1) andASUERO AND CAN0 141 the solution boiled under reflux. One millilitre of biacetyl dissolved in 20 ml of ethanol and two drops of glacial acetic acid were added and the mixture was refluxed for 1 h.The solution was allowed to cool to room temperature and the product was separated by filtration. The yellow powder obtained was washed with boiling ethanol and dried in a vacuum desiccator (yield 30%). The product had a melting-point above 300 "C and elemental analysis gave the following results: C 56.2, H 5.3, N 21.9 and S 16.8%; C,,H,,N,S, requires C 56.25, H 5.20, N 21.87 and S 16.67%. Apparatus A Unicam SP800 spectrophotometer was used for recording spectra in the ultraviolet and visible regions of the spectrum and a Coleman 55 (digital) instrument was used for measure- ments at fixed wavelengths. Quartz cells (1-cm path length) were used throughout the work. A Philips PW 9408 pH meter, with a glass and calomel electrode pair, was used for pH measurements.Throughout this paper pH is used to denote pH-meter reading and not the actual concentration of hydrogen ions in solution. A Metrohm E.1009 photometric titrator with a 4.0-cm glass cell was used in the deter- mination of the pK of the reagent. Reagents All solutions were prepared with analytical-reagent grade chemicals using distilled water. Biacetyl bis(4-~henyZ-3-thiosemicarbazone) stock solution. A 0.5% m/V solution was pre- pared in dimethylformamide. This solution was stored in an amber-glass bottle in a refrigerator. It was diluted to 0.033% for use in the spectrophotometric procedure. The stock solution was stable for several weeks when kept under these conditions. This solution was prepared by dissolving copper(I1) sulphate pentahydrate in water and was standardised by using a complexometric method with 1 -(2-pyridylazo)-2-naphthol as a metallochromic indi~at0r.l~ Sodium acetate trihydrate (105 g) and glacial acetic acid (100 ml) were diluted to 1 1 with distilled water.CoPper(I1) solz&on, 1 .OOO 3 mg mZ-1. B u f e r solution, PH 4.3. Dowex 50-X8 resin, sodium form, and Dowex 1-X8 resin, chloride form. Procedure Determination of copper in acetate medium To the copper solution (10-150 pg of copper) in a 25-ml calibrated flask, add 15 nil of 0.033% m/V BBPT solution in dimethylformamide, 1 ml of pH 4.3 acetate buffer and dilute to volume with water. Measure the absorbance a t 485 or 530 nm against distilled water. A reagent blank has negligible absorbance at these wavelengths.Determination of copper in moderately acidic medium To the copper solution (10-15Opg of copper) in a 25-ml calibrated flask, add 15 ml of 0.033% m/V BBPT solution in dimethylformamide. Adjust the pH to 2.0 & 0.05 with dilute hydrochloric acid and dilute to volume with water. Measure the absorbance at 485 or 530 nm against a reagent blank prepared simultaneously with the sample. Determination of copper in waste water, white metal and zinc blert.de Add 1 ml of 0.1 M EDTA to solutions of the first two materials and 2.5 ml of 0.1 M EDTA to solutions of zinc blende before adding the BBPT reagent solution, in order to prevent interferences by foreign ions. Calibration graphs were prepared by using standard solutions of copper(II), treated in the same way as in the recommended procedures. Results and Discussion Biacetyl BBPT Reagents BBPT has salubilitres in chloroform, methanol, ethanol, nitrobenzene and dimethyl-142 ASUERO AND CAN0 : BIACETYL BIS(4-PHENYL-3-THIOSEMICARBAZONE) Analyst, VOZ.103 formamide of 1.1, 0.2, 0.1, 1.2 and 31.3 g l-l, respectively. The solubility in water is less than 0.001 g 1-l. M) stored in darkness at low tempera- tures were stable for at least 1 week. A water - dimethylformamide medium (7 + 3 V/V) was used in the study of the reagent in order to prevent the precipitation of BBPT. In this medium a BBPT solution of 1.6 x M concentration shows maximum absorption at 343 nm, with a molar absorptivity of 4.24 x lo4 1 mol-l cm-l. Slow hydrolysis of the reagent to phenylthiosemicarbazide and biacetyl occurred in dilute solutions (1.6 x The rate of hydrolysis of the ligand increased at pH values below 4 and above 10, but decreased when the dimethylformamide to water ratio was increased. A bathochromic shift was produced at first in alkaline medium before the reagent underwent hydrolysis.A simultaneous potentiometric - photometric method15 was used in the determination of the ionisation constant when the average pK value was found to be 10.95. This behaviour may be caused by deprotonation of thiol groups. BBPT appears to be a tetradentate ligand with a convenient steric arrangement of its donor groups and contains a conjugated system of v-electrons connected with the donor system. A medium containing 60% of dimethylformamide and 40% of water was chosen for further experimental work.The main advantage of such a medium is that the chelates as well as the reagent were soluble in it at the concentration used in the photometric procedure. The characteristics of the most important RBPT complexes are shown in Table I. Dilute solutions in dimethylformamide (3.2 x M) at pH values of 6.0 and 10. The chelates of BBPT are uncharged. TABLE I CHARACTERISTICS OF BBPT COMPLEXES IN SOLUTION Metal ion Optimum pH Zn(I1) . . . . 5.5-9.5 Cd(I1) . . . . 6.2-10.7 Hg(I1) . . . . 4.2-9.7 Cu(I1) . . . . 1.8-11.9 Pb(I1) . . . . 6.5-10.5 Bi(II1) . . . . 5.9-7.2 Fe(II1) . . . . 4.8-7.5 Fe(I1) . . . . 4.5-7.1 Co(I1) . . . . 6.5-8.5 Pd(I1) . . . . 1.5-10.0 Ni(I1) . . . . 2.0-10.9 Amax./nm 440 430 420 485 530 440 450 400 400 600 400 428 582 405-420 Molar absorptivity/ lo4 1 mol-l cm-1 21.5 22.4 14.5 12.7 8.2 16.5 25.0 25.5 31.5 23.5 3.4 30.4 25.3 2.8 Colour of complex Yellow Yellow Yellow Reddish orange Yellow Orange Yellow Yellow Green Brown Green Study of Copper - BBPT System Addition of a solution of BBPT to a solution of copper(I1) ions produced a red - purple complex when the copper was in large excess; the colour changed progressively through red - purple to reddish orange as diphenylthiosemicarbazone was added in excess (Fig.1). The absorption spectra of solutions having constant contents of the reagent but increasing contents of the metal ion show an isosbestic point at 505 nm, but as the copper content of the solution exceeds the content of BBPT, the absorptivity curves no longer pass through this point.This shift indicates that a new species is being formed in the solution and also shows that the complex formation takes place stepwise. Stoicheio~netry of the complexes Job’s curves were plotted at different pH values and wavelengths (Fig. 2). At pH 10, 6.7 and 4.1 the curves intersected at 0.5 and 0.67 molar fraction of copper. This effect indicated the existence of two complexes in which the ratios of copper to BBPT were 1 : 1 and 2: 1, respectively. In acidic media, at pH 4.1 and 2.0, the ratio of metal to ligandFebrzcary, 1978 AS A SPECTROPHOTOMETRIC REAGENT FOR COPPER 143 differed from 1 : 1. This effect was attributed to competition of hydrogen ions for the reagent, which prevented the deprotonation of thiol groups. kkvelength/nm Fig.1. Absorption spectra of copper - BBPT complexes formed from various metal to ligand ratios (pH = 6.7; acetate buffer). Copper to ligand ratios: A, 1O:l (33 pg ml-l of copper); B, 1 : l (3.3 p g ml-l of copper) ; and C, 1 : 10 (3.3 pg ml-l of copper). Oxidation state of copper and structure complex previously mentioned with copper(I1). From experimental evidence it was concluded that the reagent forms the reddish orange The presence of ascorbic acid in the - Ratio [Cul /I[Cul + [BBPT] 1 Ratio [Cul /([Cul + [BBPT] 1 Fig. 2. Composition of copper - BBPT complexes by the con- (a) pH = 10: absorbance at A, 405 nm; (b) pH = 6.7 : absorbance a t A, 395 nm; (c) pH = 4.1 : absorbance at (6) pH = 2.0: absorbance a t tinuous variation method. B, 485 nm; and C, 545 nm.B, 485 nm; C, 560 nm; and D, 600 nm. A, 410 nm; B, 485 nm; and C, 540 nm. A, 485 nm; B, 540 nm; and C, 680 nm.144 ASUERO AND CAN0 BIACETYL BIS(4-PHENYL-3-THIOSEMICARBAZONE) Analyst, 'vd. 103 solution before adding the BBPT reagent did not alter the absorption spectra of the complex formed at high ligand to copper ratios. Hydrogen peroxide altered the absorption peak situated at 395 nm, because oxidising agents destroy the reagent (Fig. 3). The reddish orange complex was not retained on either a cationic or an anionic ion-exchange resin, indicating that it was uncharged. Wave I ength /n m Fig. 3. Influence of hydrogen peroxide (1 ml of 30% m/ V ) with time on the absorption spectra of copper - BBPT complex (2 pg ml-1 of copper; 15 ml of BBPT, 0.033% solution) : A, immediately; B, after 10 min; C, after 1 h ; D, blank; and E, blank after 1 h.pH = 6.7, acetate buffer. The exact configuration of a complex of this type, cc-diketone bis(thiosemicarbazone) - copper(II), has been established.l8~l7 It is apparent that BBPT acts as a tetradentate ligand; the co-ordination occurs by bonding from the first two nitrogen atoms of each phenyl- thiosemicarbazide residue and from the two sulphur atoms of the thiol groups so that three five-membered chelate rings are produced. This fact explains the greater stability of the reddish orange complex. When the ratio of copper to ligand was 10: 1, a new yellow complex appeared in the solution in the presence of ascorbic acid. The study of the copper - BBPT system when amounts of copper exceed the amounts of the reagent will be the subject of a future report.This forrnula has been previously suggested by Bahr.18 Injuence of $H When the pH was varied by addition of sodium hydroxide or hydrochloric acid, the absorb- ance remained constant over the pH range 1.8-11.9 (Fig. 4). The copper - BBPT chelate differed from cuproine analogues,lg in that the former was formed in moderately acidic as well as basic media. ~ - x - K ~ - x - x - - x - x - x ~ ~ - x - x - ~ cf 0.2 4 8 12 Ph Fig. 4. Influence of pH on the forma- tion of reddish orange copper complex. Absorbance measured at ( 0) 395 nm, ( x ) 486 nm and (a) 630 nm.Febrztary, 1978 AS A SPECTROPHOTOMETRIC REAGENT FOR COPPER 145 Temperature and colozcr development time When the temperature varied between 15 and 60 "C the absorbances remained constant within the limits of experimental error.The complex was stable for at least 24 h over the pH range 3.6-10 and it remained stable for at least 2 h at pH 1.9 (Table 11). The complex was formed immediately on addition of the reagent. TABLE I1 STABILITY OF COPPER - BBPT COMPLEX Absorbance of solution containing 2 pg ml-l of copper at pH 1.9 measured against distilled water. Timelmin f A 10 20 30 60 90 1 2 7 Absorbance a t 485 nm .. . . 0.436 0.442 0.447 0.454 0.452 0.452 Absorbance of blank . . .. . . 0.035 0.043 0.046 0.055 0.055 0.054 Absorbance difference , . * . . . 0.401 0.399 0.401 0.399 0.397 0.398 Absorbance at 530 nm 0.287 0.288 0.294 0.300 0.298 0.295 Absorbance of blank . . .. .. 0.029 0.029 0.033 0.040 0.042 0.040 Absorbance difference . , .. . . 0.258 0.259 0.261 0.260 0.266 0.265 Solvent extraction The complex can be extracted into chloroform, toluene, ethyl acetate, tributyl phosphate and benzene. The other metal - thiosemicarbazone complexes were also extracted by these solvents. Spectrophotometric Determination of Copper with BBPT Based on the experimental work two methods are proposed for the determination of trace amounts of copper. Beer's law is obeyed between 0.2 and 4 pg ml-l at 485 nm and 0.5 and 6 pg ml-l at 530 nm. The optimum concentration range, evaluated by Ringbom's method, is 1-3 pg ml-l of copper at 485 nm and 1 . 2 4 pg ml-l at 530 nm. The reddish orange complex gave values for the molar absorptivity of E = 12.710 x los 1 mol-l cm-l at 485 nm and 8.260 x lo3 1 mol-1 cm-1 at 530 nm.The sensitivities of the method according to Sandell are 0.005 and 0.0077 pg cm-2 at 485 and 530 nm, respectively. The relative error of the method is kO.19 and &0.27% at 485 and 530nm, respectively, when the pH is 6.2. At pH 2.0 0.05, the method gave a relative error (P = 0.05) of 0.27 and 0.38%. The photometric characteristics are similar in both instances. TABLE I11 EFFECT OF FOREIGN IONS ON THE DETERMINATION OF 50 pg OF COPPER AT pH 6.2 Amount tolerated/ Foreign ion pg ml-l SCN-, Br-, I-, BaOla-, PO,8-, citrate, tartrate, S,0,2- Cr(III), Mo(VI), W(VI), Tl(I), As(V), Se(IV), Mn(II), Al(III), C,042- 120 Pb(II), Zn(II), Cd(II), Hg(II), Pt(IV), Rh(III), Sn(I1) 2 000 200 Ce(IV), Th(IV), UOa(II), La(III), alkali and alkaline earth metals, Cr04a-, F- 20 V(V) Au (I1 I) 10 6 Ag(1).Bi(III), Pd(I1) 0.4 Ag(I), Pb(II), Hg(II), Au(II1) ; (SzOsa-, 2 ml of 0.26 N solution) Sb(V) ; (tartrate, 60 mg) Fe(II1) ; (F-, 180 pg d-l) 200 200 20146 ASUERO AND CAN0 : BIACETYL BIS(4-PHENYL-3-THIOSEMICARBAZONE) Analyst, VOZ. 103 TABLE IV EFFECT OF FOREIGN IONS ON THE DETERMINATION O F 50 pg OF COPPER AT pH 2.0 Absorbance measured at 530 nm. Amount tolerated/ Amount tolerated/ Ion added pg ml-1 Ion added pg ml-l Pd(II), Bi(II1) 1 C1-, Br-, I-, Se(IV), Sb(V) 5 SCN-, CH,COO-, Pt(IV), Sn(I1) 20 NO,-, phthalate, Hg(II), Fe(I1) 30 Bp0,2-, tartrate, 0 s (VIII) 80 citrate Cr( 111) 1 500 EDTA Rh( I1 I) 180 Pop- Mn(I1) 2 500 c10,- Pb(I1) 3 500 C,O,a- Zn(II), Cd(II), F- Al(III), Be(I1) 8 000 s20,2- As(V) and As(II1) 10 000 Mo(V1) > 700 Ti(IV), W(VI), Zr(1V) > 300 10 000 8 000 4 000 3 000 2 800 2 200 2 000 For the determination of 50 pg of copper by these methods, foreign ions can be tolerated at the levels given in Tables 111, IV and V.Silver and gold are reduced by BBPT to the elemental state and interfered at very low concentrations. The good results obtained by masking them with sodium thiosulphate were due to the fact that the amount of dimethyl- formamide contained in the samples prevented the decomposition of the metal - thiosulphate complexes. TABLE V ELIMINATION OF INTERFERENCES BY ADDITION OF MASKING AGENTS pH of solution 2.0. Amount tolerated/pg ml-I Without With masking Foreign ion masking agent agent Fe(II1) . . - 300 35 Ni(I1) .. - 250 Sb(V) .. 5 800 .. 5 2 000 2 000 1 000 ZF!) .. 3500 V(V) .. 20 50" - 200f. 1 30 1 000: Hg(I1) .. Au(II1) . . - >400f Fe(II1) . . - 800 Co(I1) . . - Bi(II1) . . 1 2 000 Sn(I1) . . 20 - * Masking agent EDTA; 2.5 ml of 0.1 M solution F- EDTA; 2.5 ml of 0.1 M solution EDTA; 2.5 ml of 0.1 M solution EDTA; 2.5 ml of 0.1 M solution EDTA; 2.5 ml of 0.1 M solution Tartrate; 0.25 g EDTA; 1 ml of 0.1 M solution EDTA; 2.5 ml of 0.1 M solution EDTA; 2.5 ml of 0.1 M solution S20a2-; 4 ml of 0.1 N solution S,0,2-; 4 ml of 0.1 N solution S2OS2-; 4 ml of 0.1 N solution * Absorbance measured directly after addition of reagent. t pH adjusted with dilute nitric acid. $ At pH 3.0. Applications The method has been applied to the determination of copper in industrial effluents from a sulphuric acid plant (pyrites process), a white metal and a zinc mineral.The waste water solution was filtered through an asbestos mat contained in a Gooch crucible. The average composition of the waste water analysed (seven samples) at pH 1.4 &- 0.1 was chloride 300, zinc 27.8, iron 53.1, chromium 0.05, copper 15.6, manganese 0.5, lead 0.05, arsenic 11.0 and calcium carbonate 534 pg ml-l. The copper content found by spectrophotometric determination was 15.6 0.1 pg ml-l (mean result of six determinations).February, 1978 AS A SPECTROPHOTOMETRIC REAGENT FOR COPPER 147 Zinc blende was treated first with a mixture of concentrated nitric acid and bromine (2 + 1 V / V ) and then with dilute nitric acid (1 + 1) and boiled to ensure complete dissolution of the are and to remove the oxides of nitrogen as well as the excess of bromine.The silicic acid that was precipitated was dehydrated and re- moved. White metal 8e had the following certificate composition: copper 4.57, lead 3.13, antimony 9.5, zinc 0.04, cadmium 0.14 and tin 82.6Zy0. The copper content found was 4.57 & 0.02%. A zinc blende I11 ore had the following certificate composition: zinc 52.99, lead 3.89, copper 0.12, tin 0.0018, mercury 0.039, arsenic 0.13, iron 6.59, manganese 1.45, cadmium 0.17, germanium 0.004, silver 0.005 and sulphur 30.04%. The copper content found was 0.124 These results are in exact agreement with those quoted in the certificates of analysis. The method is superior to the existing colorimetric procedure involving the use of sodium diethyldithiocarbamate,20 because it is less time consuming and involves fewer chemical manipulations.White metal was dissolved in aqua regia. Triplicate results were obtained in both instances. o.oo4~0. Conclusion Many methods are available for the determination of trace amounts of copper with thiosemicarbazone reagents (Table VI), but in our experience none of them is completely TABLE VI CHARACTERISTICS OF COPPER - THIOSEMICARBAZONE COMPLEXES Compound Picolinaldehyde thiosemicarbazone Picolinaldehyde phenylthiosemicarbazone Biacetylmonoxime thiosemicarbazone Biacetylmonoxime phenylthiosemicarbazone Thiophenaldehyde thiosemicarbazone Bipyridylglyoxal dithiosemicarbazone Cyclohexane- 1,2-dione dithiosemicarbazone Biacetyl diphenylthiosemicarbazone Optimum PH 8.9-1 0.7 8.5-10 8.5-9.5 8.5-9.7 4.5-8.2 7-9 4-10 1.8-1 1.9 Xmax./nm 410 400 345 360 372 390 467 485 630 Molar absorptivity/ 1 mol-1 cm-1 Reference 6 300 21 23 500 22 10 600 23 12 700 8 39 000 24 9 550 25 5 700 6 12 700 - 8 260 satisfactory. The great ability which the atoms of sulphur have for co-ordinating metal cations imposes a serious limitation on the use of these reagents, as it makes the establish- ment of selective methods of analysis difficult.This paper describes a study of the optimum conditions for a selective and sensitive spectrophotometric method for the determination of copper. We are grateful to Professor F. Pino for his interest and encouragement. 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. References Budesinsky, B. W., and Svec, J., Analytica Chim.Acta, 1971, 55, 115. Bahamonde, J. L., Bendito, D. P., and Pino, F., Talanta, 1973, 20, 694. Bahamonde, J . L., Bendito, D. P., and Pino, F., Analyst, 1974, 99, 355. Muiioz, J. A., Cano, J. M., and Pino, F., Infcidn. Quim. Analit. Pura ApZ. Ind., 1972, 26, 226. Muiioz, J. A., Cano, J. M., and Pino, F., An. Quim., 1976, 72, 392. Muiioz, J. A., Cano, J. M., and Pino, F., Quim. Analit., 1974, 28, 90. Berzas, J. J., Muiioz, J. 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F., “The Copper Reagents : Cuproine, Neocuproine and Bathocuproine,” IUPAC “Spectrophotometric Data for Colorimetric Analysis,” Butterworths, London, 1963, p. 154. Cano, J. M., and Pino, F., Talanta, 1972, 19, 1959. Ariza, J. G., PhD Thesis, University of Seville, 1976. Valcarcel, M., and Bendito, D. P., Infcidn. Quim. Analit. Pura A@. Ind., 1970, 24, 49. Muiioz, J. A., Cano, J. M., and Pino, F., An. Quim., 1973, 69, 251. Bahamonde, J. L., PhD Thesis, University of Seville, 1973. 1966, 88, 1845. G. Frederick Smith Chemical Co., Columbus, Ohio, 1958. Received June 3rd, 1977 Accepted August 12th, 1977 17. 18. 19. 20. 21. 22. 23. 24. 25.