摘要:

JUNE, 1968 Vol. 93, No. I107 THE ANALYS'T Editorial EVERY year the Society is faced with rising costs for the production of its journals, but this process is being considerably accelerated as a consequence of devaluation. In January appreciable increases in the prices of paper, envelopes and other materials came into effect. Printing costs were also increased, and a further rise is expected later this year. Inland postage will also cost more after September. With these rises it is necessary to couple increases in editorial costs and in general overhead expenses. Economies have been effected wherever possible, but even so only part of the increases can be absorbed. To meet this new financial situation Council has found it necessary to raise subscription rates for subscribers to the journals and for all grades of membership of the Society.A full list of the modified charges to subscribers operative from January lst, 1969, will be found on page xiii of the advertisement section. Members of the Society are being advised in the current issue of Proceedings of the revised subscription rates for all grades of membership also operative from January lst, 1969. Council regrets having to make a general increase in subscriptions both for subscribers and corporate members, but not to do so in the face of the present steep rise in costs would be both unrealistic and foolhardy, since the Society has no other appreciable source of income. However, it will be evident that the content of the Society's journals is also increasing, as would be expected from the present expansion of research in analytical chemistry, and such increases will, to some extent, offset the effect of new higher prices. All subscribers should note that it is necessary to renew subscriptions for 1969 and subsequent years with The Chemical Society, Publications Sales Office, Blackhorse Road, Letchworth, Herts. 353

ISSN:0003-2654    

DOI:10.1039/AN9689300353

出版商:RSC    

年代:1968

数据来源: RSC

摘要:

354 Anallyst, June, 1968, Vol. 93, +jb. 354-362 An Enzymatic Spectrophotometric Method for the Determination of “Oxypurines” (Hypoxanthine plus Xanthine) in Urine and Blood Plasma BY RONALD A. CHALMERS AND R. W. E. WATTS (The Medical Professorial Unit, St. Bartholomew’s Hospital, West Smithfield, London, E.C. 1) A quantitative method for the determination of “oxypurines” (hypo- xanthine plus xanthine) in the presence of relatively high concentrations of uric acid, as in urine and blood plasma, is described. The oxypurines are separated from the uric acid and, with plasma, from proteins, on a cation-exchange resin, eluted and oxidised enzymatically to uric acid by using xanthine oxidase (xanthine : oxygen oxidoreductase, E.C.1.2.3.2). The uric acid is then determined by the change in the value of E,,,, which occurs when it is oxidised to allantoin by the highly specific enzyme uricase (urate : oxygen oxidoreductase, E.C.1.7.3.3). The accuracy and reproducibility of the method have been evaluated and a comparison with other methods is included in the discussion. THE oxidation of hypoxanthine (6-oxypurine) and xanthine (2,6-dioxypurine) to uric acid (2,6,8-trioxypurine) by xanthine oxidase (xanthine : oxygen oxidoreductase, E.C.1.2.3.2) is the last step in uric acid bi0synthesis.l Blood and urine concentrations of hypoxanthine and xanthine are increased, and the uric acid concentration decreased, when xanthine oxidase is congenitally absent .2 The enzyme can be inhibited in vivo by 4-hydroxypyrazolo(3,4-d)- pyrimidine (allopurinol) and by 4,6-dihydroxypyrazolo(3,4-d)pyrimidine (oxypurinol) , which are structural isomers of hypoxanthine and xanthine, respectively.Allopurinol is used clinically to reduce uric acid production in, for example, g o ~ t , 3 , ~ , ~ and it is sometimes necessary to determine the concentration of xanthine and hypoxanthine in plasma and urine when xanthine oxidase inhibitors are administered, as well as in patients with congenital xanthine oxidase deficiency (xanthinuria) . This paper describes the relatively simple, but precise, methods that we have developed for the determination of the oxypurines in biological fluids; the term “oxypurine concentra- tion” refers, in the present context, to the concentration of hypoxanthine plus xanthine. EXPERIMENTAL Hypoxanthine and xanthine (the “oxypurines”) are separated from almost all of the uric acid that is present in the urine or plasma by a column of cation-exchange resin in the hydrogen form.With plasma, the oxypurines are also separated from the plasma proteins, thus obviating the necessity for de-proteinisation by precipitation, or for dialysis. Uric acid and proteins both pass through the column under the conditions used. The oxypurines are eluted, evaporated to dryness, and the residue dissolved in dilute sodium hydroxide solution. The oxypurines in the solution are determined by initial oxidation to uric acid by xanthine oxidase, and the uric acid so formed is determined by measuring the change in extinction at 292 nm (E,,J that occurs when it is oxidised to allantoin by uricase (urate : oxygen oxidoreductase, E.C.1.7.3.3).Traces of uric acid remaining in the solution are determined separately and subtracted from the value obtained for the oxypurine content. METHOD APPARATUS- Chromatographic colzcmns-All-glass chromatographic columns were fabricated by Howard Rawson Ltd., 22 Middle Street, London, E.C.l, to specifications (i) and (ii) for the analysis of urine and plasma, respectively. (i) 20-mm internal diameter, 50cm long, fitted with a B24 standard ground-glass joint at the upper end. (ii) 11-mm internal diameter, 20 cm long, fused at the upper end to a 30-cm length of 20-mm internal diameter tubing fitted with a B19 standard ground-glass joint. 0 SAC and the authors.CHALMERS AND WATTS 355 Both types of column had sintered-glass discs (porosity 1) and removable spring-loaded Micro $i$ettes-Constriction pipettes, to deliver 0.05 and 0.1 ml, with a quoted accuracy Rotary Jilm evaporator-A suitable rotary film evaporator, EU050 (Wright Scientific Ltd.) ; Spectrophotometer-A Unicam SP500 spectrophotometer with matched silica cells of 1-cm stopcocks at their lower ends.of k0-001 ml, were used; obtainable from H. J. Elliot Ltd., E-Mi1 Works, Glamorgan. obtainable through Gallenkamp Ltd. light path and 4-ml nominal capacity was used. REAGENTS- Analytical-grade reagents were used whenever possible. Xanthine oxidase-A suspension of milk xanthine oxidase in ammonium sulphate solution was used; obtainable from Worthington Biochemical Co., Freehold, New Jersey, U.S.A., through Cambrian Chemicals Ltd., 73, Cherry Orchard Road, Croydon.The stated activity of this material was 10 units per ml, one unit being that amount which forms 1 pmole of uric acid per minute at 25" C from 20 pg of hypoxanthine per ml; 0-05 M phosphate buffer at pH 7.5.6 Portions of the xanthine oxidase were diluted to 1 in 40 with "Tris" [2-amino-2- (hydroxymethyl) -1,3-propanediol or tris (hydroxymethyl) amino- methane] buffer, pH 8-2, before use, both the dilution and suspension being stored at -20" C when not in use. Uricase-This was supplied by Leo Laboratories Ltd., Hayes Gate House, Uxbridge Road, Hayes, Middlesex. The contents of four ampoules were dissolved in 2 ml of Tris buffer (pH 8.2) to give a solution containing 150 units per ml, which was stored at -20" C when not in use.One unit of uricase is the amount that oxidises 1 pmole of uric acid per minute to allantoin at 25" C in 0.1 M borate buffer (pH 9.4) containing 20 pg of uric acid per m1.6 Xanthine-Obtained from British Drug Houses Ltd. Hypoxmthine-Obtainable from Sigma London Chemical Co. Ltd., 12, Lettice Street, London, S.W.6. Uric acid-This was further purified by recrystallisation, as described by Liddle, Seeg- miller and Laster.' Ion-exchange resin-Dowex AG 50W x 12, analytical-grade cation-exchange resin (200 to 400 mesh, H+ form) ; obtainable from Biorad Laboratories, Richmond, California, U.S.A., through V. A. Howe & Co. Ltd., 46 Pembridge Road, London, W.11. The resin was washed with several portions of 6 N hydrochloric acid, transferred into the chromatographic column, and washed with de-ionised water until the eluate was neutral, before use.The used resin was regenerated in the same way. "T~is" bu@r working solution (PH &2), 0.05 M-A 0.2 M stock solution of Tris buffer was first prepared by dissolving 2.423 g of tris(hydroxymethy1)aminomethane in 100 ml of de-ionised water. The working solution was prepared by adding 22.5 ml of 0.1 N hydrochloric acid to 25.0 ml of 0.2 M Tris buffer stock solution and diluting to 100.0 ml with water. The solutions were stored at 4" C. Hydrochloric acid, 6 and 0.1 N. Ammonia solution, 5 and 1 N. Sodium hydroxide solution, 0-01 N. De-ionised water was used throughout the determinations and was prepared by passing distilled water through an Elgastat Deioniser (Elga Products Ltd., Buckinghamshire).COLLECTION AND PRESERVATION OF SAMPLES- Urine-Collect the urine in a polythene bottle containing 10 ml of toluene as preservative. Store 24-hour collections of urine at room temperature between voidings to avoid loss of uric acid by precipitation. If the analysis is not performed immediately, add saturated lithium carbonate (0.5 ml per 50 ml of urine) and store at -10" C; thaw the sample in a warm water bath and then cool to room temperature before analysis. Samples stored in this way showed no loss of uric acid or oxypurines over a period of 6 months. Blood-Collect 20 ml of venous blood into a heparinised syringe through a needle filled with heparin solution (5000 units per ml). Transfer the blood into a dry centrifuge tube,356 CHALMERS AND WATTS : AN ENZYMATIC SPECTROPHOTOMETRIC [Analyst, VOl.93 chill on ice immediately, and separate the plasma by centrifugation for 10 minutes at 760 g (Gave measured at the centre of the centrifuge tube) and 5" C. Plasma may be stored at -10" C and any film formed on thawing separated by centri- fugation. In most of the present work the determinations were made after an 8-hour fast by the patient. Immediate cooling of the blood and prompt separation of the cells and plasma are essential to minimise the effect of the liberation of hypoxanthine from the cells, which begins immediately blood is drawn. PROCEDURE AND CALCULATION URINE- Adjust the pH of 25.0 ml of urine to between 1.5 and 2 by dropwise addition of 6 N hydrochloric acid, with narrow-range pH papers.Pass it through a column (19 x 130 mm) of cation-exchange resin (Dowex AG 50W x 12, 200 to 400 mesh) in the hydrogen cycle, and wash the beaker and column with four 25-ml portions of water, discarding the effluent and washings. Elute the oxypurines from the column with 400ml of 1 N ammonia solution (this step can conveniently be carried out overnight by using a 500-ml separating funnel as a reservoir). Evaporate 100 to 150-ml portions of the eluate to dryness in a long-necked, 250-ml round- bottomed flask (Quickfit and Quartz, FR250/3U) at a temperature of about 35" C by using a rotary film evaporator. Dissolve the residue in 5 ml of 0.01 N sodium hydroxide, warming slightly if necessary, and transfer the solution into a 25-ml calibrated flask with the aid of further portions of 0.01 N sodium hydroxide, finally diluting to 25.0 ml with water.Determine the oxypurine content of this solution by measuring the change in the extinction (E) at 292 nm resulting from the enzymatic reactions described below. Cuvettes additional to the assay cuvette are required to correct for changes in optical density that result from factors other than the enzymatic reactions. (a) Assay cuvette-To 24ml of Tris buffer (0.05 M, pH 8.2) in a silica cuvette, add 0.1 ml of the test solution and 0-1 ml of dilute xanthine oxidase. Mix by covering the cuvette with Parafilm (Gallenkamp Ltd.) and inverting several times, and read the value of E292 at 1 to 2-minute intervals until the reaction is complete (5 to 10 minutes). Record the final value of E292 [reading (i)].Add 0.05 ml of dilute uricase, mix and read the value of E29, after 5 minutes, 30 minutes, and every 10 minutes thereafter until the reaction is complete (usually 40 to 50 minutes). Record the final value of E292 [reading (ii)]. (b) Enzyme bZank czwette-Mix 2.9 ml of Tris buffer and 0.1 ml of dilute xanthine oxidase and read the value of E292 at the same time intervals as the assay cuvette during the xanthine oxidase reaction, recording the final figure [reading (iii)]. Add 0.05ml of dilute uricase, mix, and read at the same time intervals as the assay cuvette during the uricase reaction, recording the final value of E292 [reading (iv)]. (c) Reference cuvette-Use Tris buffer to set the spectrophotometer to zero. Concurrently determine the residual uric acid content (if any) of the test solution by the following modification of the method of Liddle, Seegmiller and Laster.' (a) Assay czcvette-Mix 2.9 ml of Tris buffer and 0.1 ml of the test solution, and record the value of E292 [reading (v)].Add 0.05 ml of dilute uricase and read the value of E292 every 10 minutes until the reaction (if any) is complete, recording the final value of E292 [reading (vi)]. (b) Enzyme blank cuvette-Mix 3.0 ml of Tris buffer and 0.05 ml of dilute uricase and read the value of E292 at the same time intervals as the assay cuvette, recording the final value of E292 [reading (vii)]. (c) Reference cuvette-This is the same as for the oxypurine determination. CALCULATION- (i) Residual uric acid in cuvette. AE292 = reading (v) - [reading (vi) - reading (vii)] = A .June, 19681 METHOD FOR THE DETERMINATION OF “OXYPURINES” 357 (ii) Oxypurines + residual uric acid.The true initial reading for uricase reaction = reading (i) - reading (iii) = (a). The true final reading for uricase reaction = reading (ii) - reading (iv) = (b). :. AE292 (oxypurines + residual uric acid) = (a - b) = B. (iii) Oxypurines in cuvette. AE292 (oxypurines) = B - A . Therefore, oxypurines in cuvette ( ~ 0 . 1 ml of urine) = as uric acid. (iv) Thus, oxypurine content of urine x - X- mg per ml, as uric 1000 0.1 ‘ I (B - A ) x 3.05 1 = [- 0.0745 acid, mg per ml, expressed as hypo- 1000 xm 168 (B - A ) x 3-05 1 X - 0.0745 xant hine. The results are normally expressed as milligrams per 24 hours. PLASMA- Dilute 4.0ml of plasma, contained in a 50-ml beaker, to about 20ml with water and adjust the pH to between 1.5 and 2 by the dropwise addition of 6 N hydrochloric acid, with narrow-range pH papers. Pass the diluted, acidified plasma through a column (11 x 100 mm) of cation-exchange resin (Dowex AG 50W x 12, 200 to 400 mesh) in the hydrogen form, wash the beaker and column with four 25-ml portions of water, and discard the effluent and washings.Elute the oxypurines with 200 ml of N ammonia solution and evaporate the eluate as described for urine. Dissolve the dry residue in 4.0 ml of 0.01 N sodium hydroxide, warming slightly if necessary. Determine the oxypurine content of the solution by a similar procedure to that described for urine, with the cuvettes below. (a) Assay cuvette-To 1.9 ml of Tris buffer in a silica cuvette, add 1-0 ml of the test solution and 0.1 ml of dilute xanthine oxidase, and mix.Read the value of E292 at 1 to 2-minute intervals until the reaction is complete, recording the final value of E292 [reading (i)]. Add 0.05 ml of dilute uricase, mix, and read the value of E292 after 5 minutes, 20 minutes, and every 10 minutes thereafter until the reaction is complete, recording the final value of E292 [reading (ii)]. The procedures for (b), enzyme blank and (c), reference cuvette are the same as those described for urine. CALCULATION- 1.0 ml of plasma), is calculated as described in paragraphs (i) to (iii) above for urine. The oxypurine content of the cuvette, from 1.0ml of the test solution (equivalent to Thus, the oxypurine content of the plasma x 100 mg per 100 ml, as uric acid, 1 = [ 0-0745 1000 (B - A ) x 3.05 X - mg per 100 ml, as hypoxanthine.168 (23 - A ) x 3.05 X- 0.0745 1000 RESULTS Preliminary experiments showed that the oxypurine-containing fraction from blood plasma was protein free, the plasma proteins being washed completely through the resin, together with almost all (more than 99 per cent.) of the uric acid, with three 25-ml portions of water. The recoveries of xanthine and hypoxanthine from aqueous solutions, urine and plasma were determined and found to be satisfactory (Tables I and 11).358 CHALMERS AND WATTS : AN ENZYMATIC SPECTROPHOTOMETRIC [AutdySt, VOl. 93 In Table I, results axe corrected, where necessary, for the original urine oxypurine content, determined by the same method.Urine analyses were carried out with a 24-hour collection of urine from a normal subject. Uric acid and oxypurine content (expressed as uric acid) of the urine was 608 and 13 mg per 24 hours, respectively. In Table 11, results are corrected for the original plasma oxypurine content determined by the same method. These analyses were carried out with plasma from a normal subject, with uric acid content of 6.6 mg per 100 ml and oxypurine content of 0.04 mg per 100 ml, both expressed as uric acid. The plasma contained 65.5mg per ml of protein (determined by Warburg and Christian’s method) .8 The reading method was calibrated with standard oxypurine solutions and found to give rwults ranging from 98-5 to 101.8 per cent. w/w (mean 9943 per cent.)..v/0 50 100 150 200 250 300 350 400 ’ 0 mg per 24 hours (expressed as uric acid) The reproducibility of the method for the determination of the Fig. 1. oxypurines (hypoxanthine plus xanthine) content of urine The reproducibility of the methods for urine and plasma are shown graphically in Figs. 1 and 2, respectively. The standard deviations (Note 1) for the duplicate determinations on urine and plasma were 1-92 mg per 24 hours, with a standard error of the mean of k0-16 (70 paired observations in the range 6-3 to 420.5 mg per 24 hours) and 0-0092 mg per 100 ml, with a standard error of the mean of + l o 0 (40 paired observations in the range 0.016 to 1.157 mg per 100 ml), respectively. NOTE 1- Standard dmdation = 0 = dz where d is the difference between duplicate determinations, and Standard error of the mean =(I where u is the standard deviation.n is the number of pairs of determinations.* 4%TABLE I RECOVERIES FROM AQUEOUS SOLUTIONS AND FROM URINE The urine contained 608 mg of uric acid and 13 mg of oxypurine (expressed as uric acid) per 24 hours Oxypurine Oxypurine added per 25 ml of water or urine, mg found per 25 ml Number of of final solution, duplicate Total, expressed expressed as Experiment determinations Hypoxanthine Xanthine as uric acid uric acid, mg A A queous solutions- 1 to 6 6 2.506 0 3.097 3.093 7 to 12 6 0 2.379 2.628 2.650 Uvine- 1 to 4 4 2.615 0 3.1 1 3.097 5 to 8 4 0 2.532 2.80 2.795 9 1 0.252 0.253 0.59 0.61 10 1 0.765 0.760 1.77 1-82 11 1 1.258 1.266 2-96 2.94 12 1 2.62 2.63 5-91 5.87 TABLE I1 Recovery, Mean recovery per cent.w/w and range - 99.9 - 100.9 (98.2 to 101.1) (99.3 to 102.6) - 99-6 - 99.8 (98.4 to 102.9) (96.8 to 103.9) 103.3 - 102.8 99.3 101.2 99.3 (99.3 to 103.3) Over-all mean recovery and range 3 8 - E 100.2 z ! U 100.4 r cl tJ (98.2 to 102.6) (96.8 to 103.9) Z 5 ij 1: 0 r 0- RECOVERIES OF OXYPURINES ADDED TO PLASMA The plasma contained 6.6 mg of uric acid and 0-04 mg of oxypurine (expressed as uric acid) per 100 ml, and 66.5 mg of protein per ml x Ox ypurine Oxypurines added per 4.0 ml of plasma, pg foundper4*0-d Number of A of final solution, r duplicate Total, expressed expressed as Recovery, Experiment determinations Hypoxanthine Xanthine as uric acid uric acid, pg per cent. w/w 1 to 4 4 20-0 0 24.7 25-18 - 5 to 8 4 0 20.0 22- 1 2146 - 9 to 10 2 10.0 10.0 234 23.5 100.6 11 to 12 2 20.0 20.0 46.8 46.0 98.4 Over-all mean Mean recovery recovery and and range range 101.9 (101.2 to 102.9) 98.9 100.1 (97.3 to 101.8) 994 (97.7 to 101.7) (97-3 to 102.9) W860 CHALMERS AND WATTS : AN ENZYMATIC SPECTROPHOTOMETRIC 0.9 0.8 - n s 0-7 - .- L 3 0.6- W E I k 0.5 - 9) v - E 0 04- E! L 0.3 - 2 0.2 - 04 - [Analyst, VOl.93 0 0.1 0.2 0.3 0.4 0.5 0.6 0-7 0.8 0.9 mg per 100 ml (expressed as uric acid) Fig. 2. The reproducibility of the method for the determination of the oxypurines (hypoxanthine plus xanthine) content of plasma DISCUSSION The determination of “oxypurines” (hypoxanthine plus xanthine) in urine or blood plasma usually has to be undertaken in the presence of relatively high concentrations of uric acid.This could theoretically be overcome by determining the uric acid content before, and after, oxidation of the oxypurines with xanthine oxidase, the oxypurine content being represented by the difference between these two values. Such a difference method has been used for urine,1°~11~12 but, in our experience, this is only reliable if the oxypurine excretion is considerably increased. It is more satisfactory to remove the uric acid, and this is most con- veniently done chromatographically, rather than by preliminary incubation with uricase.l3J* In the latter procedure, the uricase must be completely destroyed by the alkalinisation of the reaction mixture before the oxypurines can be oxidised with xanthine oxidase, and the uric acid so formed finally determined by the further addition of uricase; this involves several adjustments of the pH of the reaction mixture, which introduces further potential sources of error.Chromatographic separation of oxypurines and uric acid can be carried out by anion or cation-exchange methods. The use of an anion-exchange resin for this purpose16 is more complicated, and the resin cannot be regenerated. In acidic solution, the oxypurines act as cations because of their ability to take up protons on to the C=N-C groups to form the cationic C=N-C group. This is inhibited where the C-N=C group occurs because of I H tautomerism between this configuration and C-N-C. One (hypoxanthine) and two (xanthine) nitrogen-containing groups are inhibited in this manner in the oxypurines, and nearly complete inhibition occurs with uric acid, which has three tautomeric groups.Hypo- xanthine and xanthine are thus retained on the cation-exchange resin, while the extremely weakly cationic uric acid is washed through by water (H,O+). A cation-exchange resin was chosen for the present method, as it is not only suitable for the determination of oxypurines in urine but also for their measurement in blood plasma. These compounds are removed + I OH I ll H OJune, 19681 METHOD FOR THE DETERMINATION OF “OXYPURINES” 361 directly from the plasma by the resin, and the plasma proteins washed through the column with the uric acid. Preliminary experiments showed that this method of de-proteinisation provides a high degree of accuracy compared with methods that involve the use of protein precipitants.Trichloracetic and perchloric acids are subject to co-precipitation errors, and have to be neutralised or removed before chromatography or enzymatic analysis. The method is also considerably simpler than methods that involve dialysis,12 which take longer and may require larger samples. The absorption spectra of xanthine, hypoxanthine and uric acid overlap, as shown in Fig. 3; thus xanthine, or a mixture of the two oxypurines, cannot be determined by direct oxidation to uric acid and measurement of the increase in E292, because of the contribution of xanthine to the initial value of E292. Oxidation of xanthine to uric acid is accompanied by two changes in E292, a rise caused by the formation of uric acid and a fall by the removal of xanthine; hence, the over-all rise in E292, which occurs during the reaction, is lowered because of the opposing changes taking place.This overlap could explain the low recoveries of added xanthine that some previous workers have reported.3 Xanthine oxidase also oxidises a wide range of purines, pteridines and aldehydes, and the oxidation of some of these in urine and plasma could be accompanied by changes in E292. 0.300 0.250 - c / - , I \ 0 Wavelength, rnp Fig. 3. Absorption spectra of A, hypoxanthine; B, xanthine; and C , uric acid (concentration is 20 pmolar) in Tris buffer (0.50 M, pH 8.2) These difficulties can be overcome by determining the uric acid formed from xanthine and hypoxanthine by xanthine oxidase. This is effected by measuring the decrease in E,, that occurs when uric acid is oxidised to allantoin by uricase, this enzyme being highly specific with respect to uric acid.16 Tris buffer (pH 8.2) was chosen for use as the reaction medium because, at this pH, xanthine oxidase is reacting at its optimum pH, and uricase is sufficiently active to allow the reaction to go to completion in a reasonable length of time.In addition, the extinction coefficient of uric acid at 292 nm is the same at pH 8.2 as at pH 9.4, normally used for the assay of uric acid,’ and no variation in this factor for the calculations is required. No difference was observed between recoveries of uric acid carried out by using Tris buffer (pH 8.2) and glycine buffer (pH 9.4). The use of a single buffer and pH for both enzymatic reactions avoids the necessity for adjusting the pH of the solution between successive stages of the reaction, which has made some previous methods cumbersome in operation.12~13s14s16 The presence of uric acid in the final oxypurine-containing solution is checked as a matter of routine to confirm the efficiency of the chromatographic separation.Although at very low concentration, when present, its determination is of particular importance when low concentrations of oxypurines are being determined, the amounts in these instances often being about the same. It has been shown1’ that allopurinol and its metabolite, oxypurinol, occur in the urine, and presumably the plasma, of patients undergoing treatment with allopurinol. Both of362 CHAL.MERS AND WATTS these compounds are xanthine oxidase inhibitors and might be expected to interfere with the enzymatic reaction in the assay procedure. Preliminary experiments showed that there is complete recovery of hypoxanthine added to the urine and plasma of a patient being treated with allopurinol and that suflticient xanthine oxidase is present to overcome any possible inhibitory effects of the drugs (Note 2).It has also been ~ h 0 ~ n l 7 that allopurinol is retained on Dowex 50W x 12 and eluted with the oxypurines, and that oxypurinol passes through the column with the uric acid. More than 90 per cent. of the allopurinol is excreted as its metabolite1’ and is, therefore, removed during the chromatographic separation pro- cedure; this reduces the possibility of enzyme inhibition during the assay.The addition of xanthine oxidase and uricase to solutions containing only allopurinoll0 and oxypurinol, in the present investigation, did not alter Emz; this excludes the possibility of error caused by reaction between the drugs and enzymes. NOTE 2- The recoveries of 2 and 4-pg amounts of hypoxanthine in the presence of about 5 pg of oxypurines were determined on the contents of the assay cuvettes during the measurement of the oxypurine content of urine from a patient who was receiving 600 mg of allopurinol per 24 hours. The recoveries obtained were 100 and 104 per cent., respectively, indicating that little or no inhibition of the enzyme had occurred. The present work has been concerned with the determination of hypoxanthine and xanthine together, but in certain studies it may be necessary to determine these purines separately.It would be expected that, with interference caused by uric acid removed, the oxypurine-containing solutions from the present chromatographic separation would be suitable for the separate determinations of hypoxanthine and xanthine by differential spectr~photometry,~~ and this is being investigated. CONCLUSION his int collect grants 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 16. 16. 17. The enzymatic spectrophotometric determination of the oxypurines (hypoxanthine $Zus xanthine) described above for the analysis of urine and blood plasma is accurate, reproducible and highly specific. The method has given better recoveries than those previously reported for other methods, is reasonably economic and requires a minimum of working time, and is suitable for routine determinations.The method may be extended by further study to enable the separate determinations of xanthine and hypoxanthine to be carried out. We are pleased to acknowledge our gratitude to Dr. Gertrude B. Elion, who kindly made information available to us in advance of publication, to Professor E. F. Scowen for :rest, and to the Governors of St. Bartholomew’s Hospital for their generous research Our thanks are also given to the staff of our metabolic ward for their help in the on of the numerous samples for this investigation. REFERENCES Wyngaarden, J. B., in Stanbury, J. B., Wyngaarden, J. B., and Fredrikson, D. S., Editors, “The Metabolic Basis of Inherited Disease,” Second Edition, McGraw-Hill Book Co. Inc., New York, 1966, p. 667. Watts, R. W. E., Engelman, K., Klinenberg, J. R., Seegmiller, J. E., and Sjoerdsma, A., Nature, 1964, 201, 395. Rundles, R. W., Wyngaarden, J. B., Hitchings, J. H., Elion, G. B., and Silberman, H. R., Trans. Ass. Amer. Physns, 1963, 76, 126. Klinenberg, J. R., Goldfinger, S., and Seegmiller, J. E., Ann. Intern. Med., 1965, 62, 639. Yii. T. F., and Gutman, A. B., Amer. J . Med., 1964, 37, 885. “Report of the Commission on Enzymes of the International Union of Biochemistry,” Pergamon Liddle, L., Seegmiller, J. E., and Laster, L., J. Lab. Clin. Med., 1959, 54, 903. Warburg, O., and Christian, W., Biochem. Z., 1941, 310, 384. Youden, W. J., “Statistical Methods for Chemists,” John Wiley and Sons Inc., New York; Chapman Watts, R. W. E., Watkins, P. J., Matthias, J. Q., and Gibbs, D. A., Brit. Med. J., 1966, i, 205. Goldfinger, S., Klinenberg, J. R., and Seegmiller, J. E., J. Clin. Invest., 1965, 44, 623. Klinenberg, J. R., Goldfinger, S., Bradley, K. H., and Seegmiller, J. E., Clin. Chem., 1967, 13, 834. Jarrgensen, S., and Poulsen, H. E., Acta Pharmac. Tox., 1956, 11, 223. Petersen, B. B., Jnrrni, J., and Jsrgensen, S., Scand. J. Clin. Lab. Invest., 1965, 17, 454. Simmonds, H. A., and Wilson, J. D., CZinica Chim. Acta, 1967, 16, 155. Keilin, D., and Hartree, E. F., Proc. R. SOL, 1936, Series B, 119, 114. Elion, G. B., Kovensky, A., Hitchings, G. H., Metz, E.. and Rundles, R. W.. Biochem. Pharmac., Received January 5th, 1968 Press, Oxford, 1961. and Hall Ltd., London, 1951, p. 16. 1966, 15, 863.

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DOI:10.1039/AN9689300354

出版商:RSC    

年代:1968

数据来源: RSC

摘要:

Analyst, June, 1968, Vol. 93, $9. 363-367 363 Extraction of Organochlorine and Organophosphate Insecticides from Lake Waters BY H. B. PIONKE, J. G. KONRAD, G. CHESTERS AND D. E. ARMSTRONG (Department of Soils, University of Wisconsin, Madison, Wisconsin 63706, U. S.A .) A quantitative method for the unified extraction of organochlorine and organophosphate insecticides contained in waters at microgram per litre concentrations is described. Recoveries of parathion-methyl, diazinon, malathion, azinphos-methyl, .)I-BHC, heptachlor epoxide, aldrin, dieldrin, endrin, @’-TDE, pfi’-DDT and methoxychlor added to seven lake waters and distilled water ranged from 93.9 to 102.4 per cent. Recoveries of hepta- chlor ranged from 81-0 to 88.0 per cent., apparently because of degradation of the insecticide in aqueous systems.METHODS for the extraction of insecticides from waters must essentially fulfil certain criteria. It is particularly important that the method is sensitive, that it quantitatively extracts all the insecticide present and is sufficiently rapid and simple to be applicable to routine analysis. The advent of gas - liquid chromatography, with either an electron-capture detector for organochlorine, or potassium chloride thermionic detectors for organophosphorus, insecti- cides, coupled with concentration techniques, has provided the necessary sensitivity for detecting these compounds in concentrations commonly found in naturally occurring waters. The criteria of quantitative extraction are necessary because of the heterogeneity of natural water with respect to the kinds and amounts of contaminants present that could alter insecticide extractability.The preferred methods for extracting organochlorine and organophosphate insecticides from waters have been solvent-extraction techniques because of their ready applicability to routine analysis. Recoveries of organochlorine insecticides by solvent extraction are usually high and, in many investigations, 85 to 103 per cent. of heptachlor,l DDT, aldrin, dieldrin and y-BHC192 in “spiked” water samples at pg per litre concentrations were recovered by extraction with hexane, diethyl ether or chloroform. In contrast, recoveries of organo- phosphate insecticides are generally lower and, in one instance, five successive extractions of waters containing pg per litre concentrations of parathion and diazinon with either a 1 + 1 mixture of light petroleum and ether or chloroform recovered only 90 per cent.of the added insecticide.2 Another worker achieved a more efficient extraction, in which 90 per cent. of parathion-methyl and parathion contained in waters at pg per Litre concentrations was recovered in a single hexane extraction.* However, there is little information available to indicate that an adequate extraction method for organophosphate insecticides in water has been obtained. Because the previously described batch extractions are less efficient in the ng per litre concentration range, continuous solvent-extraction technique^,^ or activated carbon filters,l s2 s6 are often used at this level of sensitivity to obtain detectable amounts of organochlorine or organophosphate insecticides.However, the use of activated carbon is limited because of suspected degradation or irreversible adsorption of organochlorine or organophosphate insecticides .1 0 SAC and the authors364 INSECTICIDES- yBHC (y-isomer of 1,2,3,4,5,6-hexachlorocyclohexane) was obtained from Hooker Electrochemical, Niagara Falls, New York, with a purity of greater than 99 per cent. Heptachlor (1,4,5,6,7,10,10-heptachloro-4,7,8,9-tetrahydro-4,7-methyleneindene) and heptachlor epoxide (the 2,3-epoxide of heptachlor) were obtained from Velsicol Chemical Corp., Chicago, Illinois, with a purity of 98.6 per cent. for heptachlor and 97.9 per cent. for hept achlor epoxide. Aldrin (1,2,3,4,10,l0-hexachloro-l,4,4a,5,8,8a-hexahydro-exo-l,4-eutdo-5,8-dimethanonaph- thalene), dieldrin (the 6,7-epoxide of aldrin) and endrin (the 1,4-endo-endo-stereoisomer of aldrin) were obtained from Shell Chemical Corp., New York, with purities of greater than 99 per cent.pp'-TDE [l,l-dichloro-2,2-di(4-chlorophenyl)ethane], also referred to as fi+'-DDD, was obtained from Rohm and Haas Chemical Corp., Philadelphia, Pennsylvania. The purity was not stated. pp'-DDT [ 1 ,l,l-trichloro-2,2-di(4-chlorophenyl)ethane] was obtained from Geigy Chemi- cal Corp., Ardsley, New York, with a purity of 99.9 per cent. @'-Met hox ychlor [ 1 , 1 , 1 -t richloro-2,2-di (4-met hoxyphen yl) et hane] was obtained from Geigy Chemical Corp., with a purity of greater than 99 per cent. Parathion-methyl (dimethyl (4-nitrophenyl phosphorothionate) was obtained from Shell Chemical Corp., with a purity of greater than 99 per cent.Diazinon (diethyl(2-isopropyl-6-methyl-4-pyrirnidinyl phosphorothionate) was obtained from Geigy Chemical Corp. labelled at the 4-position of the ring with carbon-14. Malathion (S- [ 1,2-di (ethoxycarbonyl) ethyl] dimethyl phosphorothiolothionate ] was ob- tained from Nuclear Chicago Corp., Chicago, Illinois, labelled with carbon-14 at the 2 and 3- positions of the succinic acid moiety. Azinphos-methyl (Guthion) { S- (3,4-dihydro-4-oxobenzo [dl - [ 1,2,3]-triazin-3-ylmethyl) di- methyl phosphorothiolothionate) was obtained from Chemagro Corp., Kansas City, Missouri, with carbon-14 labelling in the carbonyl group. PIONKE et al. : EXTRACTION OF ORGANOCHLORINE AND EXPERIMENTAL [Analyst, Vol.93 LAKE WATERS- Samples were chosen on the basis of wide variation in composition of the bottom sediments as described earlier.6 The sediments were removed by centrifuging the waters before analysis. INSTRUMENTAL- A Packard, model 7620, gas-liquid chromatograph was used for analysis of organo- chlorine insecticides and parathion-methyl. Gas-chromatographic conditions for chlorinated hydrocarbons were : carrier gas, nitrogen with flow-rate of 125 ml per minute; tritium-foil electron-capture detector 200" C, 50 volts; column, 2 metres long x 4-mm i.d., of 10 per cent. DC-200 on 60 to 80-mesh Gas Chrom Q; column temperature 195" C; inlet temperature 235" C ; outlet temperature 225" C. Gas-chromatographic conditions for parathion-methyl were: carrier gas, helium with flow-rate of 60 ml per minute; potassium chloride thermionic detector, 200" C, 300 volts; column, 2 metres long x 4-mm i.d., of 10 per cent.DC-200 on 80 to 90-mesh Chromosorb W; column temperature, 200" C; inlet temperature, 240" C; outlet temperature, 240" C. The instrument incorporates the use of glass columns and on-column injection to avoid sample degradation resulting from contact of the sample with metal surfaces. A Packard, model 3365, liquid-scintillation spectrometer was used for radioactivity measurements. PPO (2,5-diphenyloxazole) and dimethyl POPOP (1,4-bis- [2-(4-methyl- 5-phenyloxazole)]-benzene) were used as the primary and secondary fluors, respectively. For benzene extracts, a solution of 0.5 per cent.of PPO and 0.03 per cent. of dimethyl POPOP in toluene was used. Corrections for sample quenching were determined by external standardisation. SOLVENTS- were used for "spiking" and extracting the water samples. DESCRIPTION OF METHOD Benzene and acetone, re-purified by glass-distillation with a 3-ball Snyder column,June, 19681 ORGANOPHOSPHATE INSECTICIDES FROM LAKE WATERS 365 PROCEDURE- A method for the extraction of organochlorine and organophosphate insecticides from waters, with benzene as the extractant, was found to be quantitative, sufficiently rapid and simple for use as a routine procedure. The method is described as follows. A 250-ml water sample is extracted with 25 ml of benzene in a single extraction by shaking in a separating funnel for 2 minutes, and the separated benzene phase is analysed directly by gas-liquid chromatography with an electron-capture detector for organochlorine insecticides, or potassium chloride thermionic detector for organophosphates. If both organochlorine and organo- phosphate are present in the sample, simultaneous analysis can be accomplished by splitting the column effluent and by using a dual electron-capture potassium chloride thermionic detector.If emulsification is encountered, anhydrous sodium sulphate can be used to remove the water. RESULTS AND DISCUSSION ORGANOCHLORINE INSECTICIDES- The evaluation of the above extraction procedure for organochlorine insecticides was based on “spiking” water samples with a mixture of the insecticides as follows. Amixture of the insecticides in acetone (20 p1) containing y-BHC, heptachlor, heptachlor epoxide, aldrin, dieldrin, endrin, $9’-TDE, @’-DDT and methoxychlor was added to the 250-ml water samples.The “spiked” sample, contained in a separating funnel, was shaken end-over-end continuously for 12 hours to obtain maximum dissolution of the insecticides. TABLE I RECOVERY OF ORGANOCHLORINE INSECTICIDES FROM LAKE-WATER SAMPLES Recoveries of insecticides, per cent., at concentrations of- A I \ 3-28 pg per litre 6.40 pg 8.90 pg 32.6 pg per litre hepta- per litre chlor per litre per litre pp’- $p’- meth- 1 -60 pg 1-23 pg per litre 2-08 pg hepta- 3.98 pg 5-02 pg per litre per litre per litre Lake waters yBHC chlor aldrin epoxide dieldrin endrin TDE DDT oxychlor Distilled water . . . . 96.5 85.6 101.8 98.3 100.0 99-2 97.0 97.2 98.5 Allequash ... . 96.5 83.6 102.3 99.0 100-3 99.0 98.6 96.9 94.4 Content,. .. . . 96.3 81.0 100.2 96.2 99.0 98-9 96.6 96.7 97.1 Crystal . . .. . . 97-2 88.0 102.0 98.6 98-5 99.1 96.0 94.4 97.0 Pickeral . . . . 95-6 84.5 101.7 96.6 97.5 96.8 97.2 95.0 97.0 Plum . . .. . . 96-8 82.0 102.4 97.0 98-4 97.2 98.0 96.8 97-2 Tomahawk .. . . 97.5 81.7 102.1 96.6 99-0 98.7 98.3 95.3 98-5 Mean recoveries . . 96.7 84.6 101.8 97.4 98.7 98.4 97.3 96.1 97.3 Average range of dupli- cates . . .. . . 2.0 2.9 2.7 1.0 1.7 1.3 2-8 0-7 2.0 Standard deviation be- Little Arbor Vitae . . 97-1 82-0 101.8 96-8 97.0 98-1 97.0 96.3 98-6 tween water samples 0-60 2.60 0.62 1-03 1-12 0-92 0-89 1.02 1.36 With the exception of heptachlor, the recoveries of organochlorine insecticides from lake waters and distilled water ranged from 94.4 to 102 per cent.at pg per litre insecticide concentrations, as seen in Table I. These results are the average of duplicate determinations conducted at different times. The average range of recoveries between duplicate deter- minations i.e., the summation of the range between duplicates divided by the number (i.e., 8, of water samples) for each insecticide was 0.7 to 2.9 per cent. (including heptachlor). This error term combines the errors associated with extraction and the analytical technique, indicating that the method of extraction is reproducible even for heptachlor, which displayed low recovery values. A second error term, the standard deviation, relates the quantitative reliability of the method between lake-water samples.Average extractability ranged from 96.1 to 101.8 per cent. (except for heptachlor), with a standard deviation of 0-60 per cent. for y-BHC to 1.36 per cent. for methoxychlor. The average ranges and standard deviations were deemed sufficiently low to substantiate the general applicability of this method for con- sistently quantitative recoveries of the insecticides from water at pg per litre concentrations.366 [Artalyst, Vol. 93 Heptachlor recoveries were invariably low (81 to 88 per cent.), even when the “spiked” waters were re-extracted with benzene. The consistent reproducibility of heptachlor recovery from a given water, as shown by the low average range between duplicates (2.9 per cent.) and the absence of heptachlor in the second benzene extract, indicated that the low recovery of heptachlor was not caused by incomplete extraction.It seems more likely that the low recoveries were caused by heptachlor degradation during incubation in the water, as suggested by Lamar, Doerlitz and Law.l For heptachlor, the standard deviation between samples was greater than for the other insecticides, which is consistent with degradation losses. If degradation of heptachlor occurs in water, it is expected that degradation rates would vary among water samples because of differences in water composition. Because solvent evaporation was observed to cause large volatilisation losses of aldrin, acetone was chosen as the “spiking” solvent to avoid the evaporation step necessary for the removal of hexane or other water-immiscible “spiking” solvents.After “spiking,” the acetone concentration in the water samples was 80mg per litre, which was considered too small to affect insecticide extractability. The lake waters were chosen on the basis of differences in bottom sediment charac- teristics that are likely to affect the water composition. The sediment samples provided a range of organic matter (0.14 to 64.2 per cent.) and clay (4-4 to 36.1 per cent.) contents; the extent of oxidation of the organic matter as measured by methoxyl content and carbon- to-nitrogen ratio showed wide variability. Furthermore, benzene extracts of the lake waters when gas chromatographed showed no contaminants that would interfere with the detection of the nine organochlorine insecticides at pg per litre concentrations.The concentrations of organochlorine insecticides in water (Table I) were chosen to provide sufficient sample for the accurate evaluation of the extraction method without exceeding the maximum aqueous solubilities. Within these solubility limits, concentrations were selected to obtain about the same detector response for each insecticide, thereby facilitating simultaneous chromatographic analysis of the insecticides. Although the extraction method was not evaluated for insecticide concentrations below 1 pg per litre in water, the quantitative recoveries attained (assuming quantitative recoveries were achieved for heptachlor) are likely to be achieved at one tenth, or less, of the insecticide concentrations presented in Table I.If slight modifications are used to increase the sensitivity of the method, e.g., by extracting larger volumes of water, or by manually changing sensitivity settings of the gas chromatograph, sensitivities can be obtained that blanket the sensitivity range required for the determination of organochlorine insecticides in most natural waters.’ PIONKE et al. : EXTRACTION OF ORGANOCHLORINE AND ORGANOPHOSPHATE INSECTICIDES- The extractability of organophosphate insecticides from lake waters was evaluated on the same samples used for organochlorine insecticides, and the method of extraction used was the same. “Spiking” the waters with organophosphate was achieved in the following manner. The insecticide, in 1 ml of benzene, was added to 250 ml of water and the benzene evaporated by passing a stream of air over the water.Homogeneous solution of the insecti- cides was determined by removing three aliquots of an azinphos-methyl “spiked” water without shaking. Each aliquot was extracted with benzene and the carbon-14 activity of the extracts was 99.3 0.5 per cent. of the expected activity, indicating homogeneous distribution of the insecticide in the water. In a later investigation* it was confirmed by gas chromatography that organophosphate insecticide in benzene extracts of soil-insecticide systems remained in the intact form, indicating that organophosphates extracted from the lake water would not have undergone degradation. The extractability of parathion-methyl, diazinon, malathion and azinphos-methyl added to the seven lake-water samples and distilled water is shown in Table 11.With the exception of parathion-methyl, the average recoveries were 98-4, 99.7 and 99.5 per cent., with ranges of 97.6 to 99-2, 98.3 to 100.6 and 97.1 to 100.9 per cent. for diazinon, malathion and azinphos- methyl, respectively. The average recovery for parathion-methyl was 95.3 per cent ., with a range of 93.9 to 97-7 per cent. This lower recovery value for parathion-methyl probably reflects the higher inherent error in gas-chromatographic determination than in radioassay by liquid scintillation spectroscopy. However, parathion-methyl was determined satisfac- torily at a concentration of 38pg per litre in the lake water, and no concentration of theJune, 19681 ORGANOPHOSPHATE INSECTICIDES FROM LAKE WATERS TABLE I1 RECOVERY OF ORGANOPHOSPHATES FROM LAKE-WATER SAMPLES 367 Recoveries of insecticides, per cent., at concentrations of- A f \ 38 pg per litre 40 pg per litre 60 pg per litre 66 pg per litre Lake waters parathion-methyl* diazinon malathion azinphos-methyl Distilled water .. .. .. 97.7 99-2 10045 98.6 Allequash .. .. .. 96.1 98.4 1004 100.8 Content . . .. .. .. 96.3 98-7 99.3 100.9 crystal .. .. . . 94.4 99.2 99.6 100.6 Little Arbor Vitae . . .. 94.6 99.0 100.6 97.1 Pickeral .. .. .. - 98.1 99.6 98.9 Plum .. .. .. 97.6 98.3 98.7 Mean recoveries .. .. 96.3 98.4 99.7 99.6 Average range of duplicates . . 1.4 1.7 1.6 1.6 Standard deviation between Tomahawk .. .. .. 93.9 97.2 99.7 100.2 water samples . . .. 1.44 0.76 0.78 1.36 * Determined by gas chromatograph, other insecticides determined from carbon-14 activity.benzene extracts was found to be necessary. Concentrating benzene extracts is complicated by the thermal instability of organophosphate insecticides. Parathion-methyl was found to degrade when the benzene extracts were concentrated by evaporation at 80" C, and similar decomposition of other organophosphates would be expected to occur if a concentration step were used. Therefore, concentration of the benzene extract is not recommended prior to gas-chromatographic determination of organophosphate insecticides, unless investigations on the stability of the insecticide are conducted simultaneously. Statistical analyses of the extractabilities of organophosphates were conducted in a manner similar to that used for the organochlorine insecticides.In a l l cases the average range of duplicates and standard deviations were similar to those obtained for ~rganochlorine insecticides, and were well within the limits of statistical error, substantiating the general applicability of the prescribed extraction method for consistent quantitative recovery of organophosphate insecticides from lake waters in the pg per litre range. The method described here is a rapid, reliable procedure that is suited to routine analysis for the simultaneous extraction and determination of organochlorine and organophosphate insecticides from lake waters in pg per litre concentrations. This work was supported in part by the US. Department of Agriculture, ARS Contract No. 12-14-100-8164(41) and the U.S. Department of the Interior, OWRR Project No. B-016-WIS. REFERENCES 1. 2. 3. 4. 6. 6. 7. 8. Lamar, W. L., Doerlitz, D. F., and Law, L. M., Geol. Surv. Wut.-Supply Pup., 1965, 1818-B: 1. Teasley, J. L., and Cox, W. S., J . Amer. Wut. Wks Ass., 1963, 55, 1093. Warnick, S. L., and Gaufin, A. R., Ibid., 1966, 57, 1023. Kahn. L., and Wayman, C. H., Anulyt. Cham., 1964, 36, 1340. Rosen, A. A., and Middleton, F. M.. Ibid., 1959, 31, 1729. Lotse, E. G., Graetz, D. A., Chesters, G., Lee, G. B., and Newland, L. W., J. Envir. Sci. & Tecknol., Faust. S. D., and Suffet, I. H., in Gunther, F. A., Editor, "Residue Reviews," Springer-Verlag, Konrad, J. G., Armstrong, D. E., and Chesters, G., Agron. J., 1967, 59, 691. Received January Sth, 1968 1968, May, 2. Berlin, Gottingen and Heidelberg, Volume 16, 1966, p. 44.

ISSN:0003-2654    

DOI:10.1039/AN9689300363

出版商:RSC    

年代:1968

数据来源: RSC

摘要:

368 Analyst, June, 1968, Vol. 93, $$. 368-370 A Rapid Procedure for the Identification of Organochlorine Pesticides in Blood and Tissues BY C. E. ELIAKIS, A. S. COUTSELINIS* AND E. C. ELIAKIS (Department of Forensic Medicine and Toxicology, University of Athens, 48 Academy Street, Athens 146, Greece) A method is described for the rapid identification of chlorinated insecti- cides, such as aldrin, dieldrin, op’-DDT and pp’-DDT, in tissues and blood, without the necessity for a previous clean-up procedure of the extracts made from them. Two-dimensional thin-layer chromatography was used for this purpose. The plates were developed in hexane for one dimension and in cyclohexane for the other, and were sprayed with a solution of 0-5 per cent. silver nitrate in ethanol before exposure to germicidal ultraviolet light.The sensitivity of this method is 0.5pg, which is suitable for routine analysis. MANY papers have been published giving methods for the trace detection of chlorinated insecticides. These methods include two steps : an extraction and clean-up procedure for the extracts, and chromatography. The existing extraction and clean-up procedures were studied in an effort to reduce the time factor to a minimum and to develop an improved clean-up technique. Of the chromato- graphic procedures available, thin-layer chromatography, as developed by Kovacsl y2y3 and Abbott, Egan and Thom~on,~ has proved most useful for the semi-quantitative determination of the chlorinated insecticides, as it combines the advantages of rapidity and economy.Consequently, thin-layer chromatography is of particular interest to our laboratory, in which numerous toxicological analyses are carried out annually for the detection of traces of chlorinated insecticides in cases of poisoning that are attributable to the wide use made of these substances in agriculture. In these cases, the specimens examined are of varied nature, consisting frequently of viscera (liver) and blood in an already decomposed condition. As mentioned, the purpose of our laboratory was to find a method for the rapid identi- fication of chlorinated insecticides in tissues and blood. We have found that the use of two-dimensional thin-layer chromatography facilitates the clear identification of chlorinated insecticides such as aldrin, dieldrin of-DDT and +p’-DDT, without the necessity for a previous clean-up procedure of the extracts made from liver or blood.The use of two-dimensional thin-layer chromatography was considered valuable, because with viscera and blood in a decomposed condition, one-dimensional thin-layer chromato- graphy, without previous clean-up of the extracts, did not yield satisfactory results; this was because in most instances when chromatograms were developed with silver nitrate solution, in addition to the spots formed with the chlorinated insecticides, the whole length of the chromatogram course became dark, resulting in a decrease in the sensitivity. On the other hand, when examining blood we found that the extraction method described by Folch, Lees and Sloane Stanley,5 who had applied it to the extraction of total lipids, after being slightly modified by us gave satisfactory results with a short extraction time.The technique described below for the examination of blood is simple and rapid, and its sensitivity has been proved to be quite satisfactory in cases of poisoning. * Present address: Department of Pharmacology, School of Medicine, University of New Mexico, Albuquerque, N.M., 87106, U.S.A. 0 SAC and the authorsELIAKIS, COUTSELINIS AND ELIAKIS 369 EXTRACTION PROCEDURE- One millilitre of total blood was extracted with 6 ml of methanol by shaking the mixture for 30 minutes; 12 ml of chloroform were added and the mixture again shaken for another 30 minutes. The extract was filtered on a filter-paper and the filtrate made up to 20 ml with a mixture of chloroform - methanol (2 + 1 v/v).This solution was then shaken with one fifth of its volume, i.e., 4 ml, of water for 5 minutes. The mixture was then centrifuged and the chloroform layer separated and dried over anhydrous sodium sulphate. After evaporating the solution, the residue was dissolved in 1 ml of hexane and centrifuged at 3000 r.p.m. for 3 to 4 minutes. Following centrifugation, 0-2 ml of the hexane layer was used for chromatography. CHROMATOGRAPHY- Neutral plates (0.25mm thick) were prepared according to the method of StahP by using silica gel G. The plates were activated at 110" C for 1 hour. The spot was located at a distance of 1.5 cm from adjacent sides of the plate. The plates were developed in hexane, dried for half an hour at room temperature, and developed again in cyclohexane.In both systems the length of run was 12 cm from the start. During the run in both solvents, the tanks were saturated in the conventional way. A standard mixture of chlorinated insecticides was chromatographed for each dimension on a separate chromatoplate in the same chromatotank, with the same conditions and development time as before. The positions on the chromatoplate of the chlorinated insecti- cides examined had been previously determined by chromatographing each of them separately in two-dimensional co-ordinates with the same solvents. In addition, standard compounds, together with the sample, were chromatographed for each dimension on the same chromato- plate (Figs. 1 and 2). This was carried out because the RB values, which are influenced by various factors, are not constant.y = Cyclohexane y' = Hexane a = Dieldrin b = pp'-DDT c = op'-DDT d = Aldrin -1 a 0- Y I XI = Position of the standard spot for the development XU = Position of the standard spot for the development p-p' = Limits of chromatogram's development in both in the y' direction in the y direction systems Fig. 1. Two-dimensional thin-layer chromatography of standard compounds370 - I u‘ ELIAKIS, COUTSELINIS AND ELIAKIS Iy 1 - Y’ 0 Y = Cvclohexane d = Aldrin i‘ = Hexane a = Dieldrin b =PP’-DDT c =,op’-DDT XI = Position of the standard spot for the development in the y’ direction p-p’ = Limits of chromatogram’s development in both systems Fig. 2. chromatography Separation of organochlorine insecticides from blood by two-dimensional thin-layer The plates were then dried, sprayed with a solution of 0.5 per cent.silver nitrate in ethanol and exposed to germicidal ultraviolet light for 15 minutes. With the above method, 0.5 pg of aldrin, dieldrin and DDT can be identified as dark spots on a white background. The same chromatographic technique was used for liver, for which the extraction procedure described by Taylor, Rea and Kirby7 was used; the hexane layer was evaporated to 10 ml, 0.2 ml of which was chromatographed. The above-mentioned sensitivity was considered quite satisfactory for identifying traces of chlorinated insecticides in cases of acute poisoning. REFERENCES 1. 2. 3. 4. 5. 6. 7. Kovacs, M. F.. jun., J . Ass. Off. Agric. Chem., 1963, 46, 884. - , Ibid., 1964, 47, 1097. - , Ibid., 1966, 48, 1018. Abbott, D. C., Egan, H., and Thomson, J., J . Ckomat., 1964, 16, 481. Folch, S., Lees, M., and Sloane Stanley, G., J . Biol. Chem., 1957, 226, 497. Stahl, E., Editor, “Thin-Layer Chromatography : A Laboratory Handbook, ” Springer-Verlag, Taylor, A., Rea, R. E., and Kirby, D. R., Analyst, 1964, 89, 497. Received September Mth, 1967 Berlin, Heidelberg and New York; Academic Press Inc., New York and London, 1966.

ISSN:0003-2654    

DOI:10.1039/AN9689300368

出版商:RSC    

年代:1968

数据来源: RSC

摘要:

Analyst, June, 1968, Vol. 93, pp. 371-374 37 1 A Radiometric Procedure for the Micro Determination of Palladium and Iodide with Iodine131 BY MRS. USHA PURKAYASTHA AND H. P. MAITY (Nuclear Chemistry Division, Saha Institute of Nuclear Physics, 92 Acharya Prafulla Chandva Road, Calcutta, 9) A method is described for the micro determination of palladium and iodide in which the radioisotope iodine-131 is used as a tracer. The principle of the method depends on the formation of labelled palladium(I1) iodide by the addition of an excess of a solution of potassium iodide containing iodine-131 to the palladium, the palladium(I1) iodide being then carried by zirconium hydrogen orthophosphate, Zr(HPO,),, formed in the solution. From the loss of iodine-131 by the solution, the amount of palladium can be determined.By using the same principle, iodide can be determined if a known amount of palladium is added. The adsorption of palladium(I1) iodide by zirconium hydrogen orthophosphate has been found to be selective. Amounts of palladium in the range 4 x 10-8 to 2 x lO-'g have been determined so far, and errors are always within statistical fluctuations in activity measurements. The effect of the presence of some foreign ions has also been studied. The method is simple and can be completed within 30 minutes. THE determination of iodide as palladium(I1) iodide is well kn0wn.l Palladium is usually determined with dimethylglyoxime. An attempt has been made to determine palladium as palladium(I1) iodide by using iodine-131 as a radioactive indicator. The basis of the method is as follows.An excess of labelled potassium iodide solution is added to the palladium solution and the palladium(I1) iodide formed is carried with zirconium hydrogen orthophosphate. The radioactivity of the iodine left in solution is then measured. From the loss of radioactive iodine the amount of palladium can be calculated, assuming that all of the iodide is carried in the form of palladium(I1) iodide. By the same principle, micro amounts of iodide can be determined if a known weight of palladium, which is insufficient to combine with all of the iodide, is added after the addition of the tracer iodide. EXPERIMENTAL The canier-free iodine-131 used in this investigation was obtained from the Atomic Energy Establishment Trombay. The chemical reagents used, viz., palladium(I1) chloride, potassium iodide, zirconium nitrate and acids, were all of analytical-reagent quality. The procedure adopted in a typical determination is described below. A solution of carrier-free iodine-131 was thoroughly mixed with a solution containing a suitable amount of potassium iodide. A known volume of the mixed labelled solution containing two to four times the theoretical amount of iodide, as required by the equation was mixed with the solution of the palladium to be determined and a solution of zirconium nitrate, containing 0.5 to 1.0 mg of zirconium, then added. Orthophosphoric acid, equivalent to about ten times that required by the formula Zr(HPO,),, was added, dropwise, and the palladium( 11) iodide carried by the zirconium hydrogen orthophosphate formed.The mixture was thoroughly stirred and gently warmed until coagulation occurred. The volume of the solution at this stage was 12 to 14 ml, and the solution about 0.5 N with respect to sulphuric acid. It was then transferred into a 25-ml calibrated flask and the volume made up to the OSAC and the authors Pd2+ + 2KI -+ PdI2,372 PURKAYASTHA AND MAITY: A RADIOMETRIC PROCEDURE FOR THE MICRO [Artalyst, Vol. 93 mark. An aliquot was removed by pipette, with a filter-paper cap at its tip, and its activity measured with a liquid counter. From the loss of iodine-131 from the solution, the amount of palladium precipitated was calculated, assuming that zirconium hydrogen orthophosphate carries 100 per cent. of the insoluble iodide.For the determination of iodide, a solution containing the unknown iodide was thoroughly mixed, in a beaker, with carrier-free iodine-131. A solution containing palladium(I1) chloride, which was insufficient to combine with all of the iodide, was added, and the remaining operations were as previously described. TABLE I MICRO DETERMINATION OF PALLADIUM AND IODIDE WITH IODINE-131 Experiment No. 1 2 3 4 6 6 7 8 9 10 11 12 *13 14 15 16 17 *18 19 20 21 *22 23 *24 25 26 27 *28 29 30 *31 32 * 33 Experiment Nos. 1 to 3 4 and 5 6 to 33 14 to 18 19 to 24 25 to 28 29 to 31 32 and 33 Palladium taken, mg 4.55 2.28 1-14 0.100 0~0100 0~0100 0.00455 0.00273 0.001 14 0.000465 0.000228 0.000228 0.0001 14 0.001 14 0.001 14 0.001 14 0.00 1 14 0.00114 0-001 14 0-00114 0.001 14 0.001 14 2-28 2-28 0.001 14 0.00114 0-00114 0.00114 0.0 100 0~0100 0~0100 0~0100 0~0100 Palladium found, mg 4.67 2.2 1 1.1 1 0.0986 0.0101 0~0101 0.00455 0.00269 0~00110 0.000438 0.000223 0.000219 0.0000857 0.00113 0.001 11 0.001 11 0.00115 0.00 106 0.00115 0.00115 0.00112 0.000982 2.211 1.98 0.001 16 0.001 12 0.00115 0*000953 0.00973 0.00988 0.00890 0-00979 0.00880 Iodine, added as potassium iodide, mg 24.4 13.7 6-84 0.760 0.0760 0.0760 0.0367 0.0220 0.01 10 0.00367 0.00183 0.00183 0.001 10 0.01 10 0.0110 0.01 10 0.0110 0.0110 0~0110 0.01 10 0.0110 0.0110 13.7 13.7 0.0110 0.01 10 0.01 10 0.01 10 0.0760 0.0760 0-0760 0.0760 0.0760 Iodine found, mg 23.8 14-1 7-05 0.773 0.0758 0.0757 0.0367 0.0223 0.01 1 0.00381 0.00187 0~00190 0.00146 0.01 11 0.01 13 0.01 13 0.0109 0.0118 0.0109 0.0109 0.01 12 0-0128 14.1 15.7 0-0108 0.01 12 0.0109 0-0131 0.0783 0.0771 0.0854 0-0778 0.0865 Standard deviations of activity measurements , per cent.f 2-7 f3.1 f3.1 f2.0 f0.6 f 0.6 f 0-5 f 1-7 f 3.3 f 4.0 f 2-2 & 3.9 f3-6 f 1-3 f2.7 f 2.6 f 1.2 f 3.2 *1a1 f 1-4 f 1.7 f3-1 f 3.0 54.1 f 2.2 f 2.2 f 1.2 f 3.4 f 3.2 f 1.6 f 3.7 f2.7 k4.1 No carrier used. Amount of zirconium (as zirconium nitrate) used was 0.5 mg. Amount of zirconium (as zirconium nitrate) used was 1.0 mg. Amounts of nickel present were 0.011, 0-221, 0.554, 0-831 and 1-108 mg. Amounts of platinum present were 0.004, 0.008, 0.015, 0.023, 29.98 and 39-98 mg. Amounts of gold present were 0.0025, 0.005, 0.012 and 0.019 mg. Amounts of chlorine present were 2-20. 2.29 and 2-47 mg.Amounts of bromide present were 0-37 and 0-74 mg. * These results indicate that chemical errors are far greater than those caused by standard deviations in activity measurements. In these determinations, both palladium and iodide were calculated from the known iodine activity left in solution. Palladium was determined from knowledge of the amount of iodide added as potassium iodide, and the iodide determined from knowledge of the amount of palladium chloride added. It is evident from Table I that the errors in the determinationJune, 19681 DETERMINATION OF PALLADIUM AND IODIDE WITH IODINE-131 373 of palladium and iodide are equal in magnitude, but opposite in sign, because the values for both have been calculated from the same iodine activity left in solution in any operation.For the determination of palladium or iodide in the presence of foreign ions, a solution containing palladium or iodide was thoroughly mixed with the solution of the foreign ion and the palladium or iodide determined as already described, except that in the presence of gold and platinum, sodium sulphite (about 0.2g) was added to keep the medium in a reducing condition. To correct for any error caused by adsorption of iodide ions, a blank experiment was carried out without palladium, and this served as a standard for comparison. Experiments were also carried out to study the carrying of micro concentrations of various iodides on zirconium hydrogen orthophosphate (see Table 11). All of the results recorded in Tables I, I1 and I11 were obtained under the experimental conditions described above.TABLE I1 RESULTS WITH ZIRCONIUM HYDROGEN ORTHOPHOSPHATE AS A CARRIER OF VARIOUS IODIDES IN MICRO CONCENTRATION, WITH IODINE-131 AS A RADIOACTIVE INDICATOR Cations Iodine, added as Zirconium, contained in Uptake, Cations taken, potassium iodide, zirconium nitrate, per cent. x 10-6g x 10-6g mg Palladium . - .. 1.0 7.6 1.0 96.0 10.0 76.0 1.0 100.0 100.0 760.0 1.0 100.0 Silver .. .. 1.18 13.7 0.5 100.0 11.8 27.0 0.5 100.0 118.0 274.0 1.0 100.0 Thallium(1) . . .. 1.2 6.8 0.5 0.0 12.0 15.2 0.5 0.0 123.0 152.4 1.0 0.0 Copper(1) . . .. 1.0 5.4 0.5 0.0 10.5 54.8 0.5 0-0 105.0 648.0 1.0 0.0 Lead .. .. .. 1.1 2.7 0.6 0.0 11.5 27.4 0.5 0.0 115.0 274.0 1.0 0.0 Mercury(1) . . .. 1.3 1.5 0.5 0.0 13.0 15.2 0.5 0.0 131.0 152.0 1.0 0.0 Mercury(I1) .. . . 118.0 274.0 1.0 0.0 Cadmium .. .. 1.4 7.6 0.5 0.0 14.0 76.0 0.6 0.0 140.0 760.0 1.0 0.0 DISCUSSION Table I11 shows that the extent of adsorption of iodide ion, in a carrier-free state at a limited concentration, on zirconium hydrogen orthophosphate is negligibly small. Zirconium hydrogen orthophosphate takes up an appreciable amount of iodine activity in the presence of palladium. Table I shows that the iodide is carried as palladium(I1) iodide. Although TABLE I11 ADSORPTION OF VARYING AMOUNTS OF IODIDE IONS ON DIFFERENT AMOUNTS OF ZIRCONIUM HYDROGEN ORTHOPHOSPHATE IN 0-5 N ACID Zirconium taken, mg 0.5 0.5 0.5 0-5 1.0 Iodine, added as potassium iodide, mg 0.00 (without carrier) 0.0076 0.076 0.76 7.60 Iodide ion adsorbed, per cent.* 1.2 1-1 1.8 2.0 1.5 * Although the extent of adsorption is negligibly small, the blanks prepared, as mentioned in the Discussion, under identical conditions were used to give greater accuracy.374 PURKAYASTHA AND MAITY the adsorption of free iodide ion on zirconium hydrogen orthophosphate was negligibly small, blanks were made under identical conditions, so that errors caused by adsorption could be rendered insignificant (see Table 111).In Table I, typical results for the micro determination of palladium with iodine-131 are given, from which it is evident that palladium in the range 4 x to 2 x g can be determined with a fair degree of accuracy. It may be further observed that 2 x 10-6g of iodide can also be determined with the same degree of accuracy. Zirconium hydrogen orthophosphate can therefore be regarded as a reliable carrier in the system under investigation.The detemination of palladium has also been carried out in the presence of some inter- fering elements. It was found that it could be determined in the presence of about 750 times its weight of nickel and about twelve times its weight of gold and platinum, the latter metals being taken as their complex chlorides. It was also found that iodide could be determined in the presence of thirty times its weight of chlorine and five times its weight of bromine. The results in Table I1 show that only silver and palladium(I1) iodide are taken up quantitatively by zirconium hydrogen orthophosphate. In the presence of thallium(I), copper( I) , mercury( I), mercury( 11) , cadmium and lead, the recovery of palladium( 11) iodide was low, so that the determination of palladium in the presence of these elements was not successful.Palladium(I1) iodide probably formed complexes with the iodides of these metals. Adsorption of silver iodide on zirconium hydrogen orthophosphate has been re- ported in an earlier paper.2 It is of interest to note that silver and iodide can be determined in the presence of most of the iodides mentioned above by using zirconium hydrogen ortho- phosphate as a ~arrier.~ It will not be out of place to mention that cadmium is the only element in the group that interferes in the determination of silver by the m e t h ~ d . ~ In the present study, we observed that palladium(I1) iodide is adsorbed on zirconium hydrogen orthophosphate. The selective nature of this adsorption, which may be caused by anomalous mixed crystal formation, requires further study for its clarification. It is shown by experiment that the concentrations of palladium and iodide that can be determined are lower than 2 x 10-8 and 2 x lO-7g per ml, respectively. This method has various advantages over the classical procedures. It does not require more than half an hour to complete a full set of determinations ; filtration, washing and weighing are not required; and solubility factors, the use of which is necessary in radiometric analysis, are dispensed with by the application of a carrier. The authors express their thanks to Professor B. C . Purkayastha for his help and valuable suggestions. REFERENCES 1. 2. 3. Furman, N. H., Editor, “Standard Methods of Chemical Analysis,” Volume I, Sixth Edition, D. Van Nostrand Company Inc., Princeton, New York, Toronto and London, 1962, p. 616. Purkayastha, B. C., and Pai Verneker, V. R., J . Indian Chem. SOL, 1967, 31, 487. Purkayastha. Usha, and Maity, H. P., Indian J . AppZ. Chem., 1968, 31, in the press. First received August 22nd, 1966 Amended December 28th, 1967

ISSN:0003-2654    

DOI:10.1039/AN9689300371

出版商:RSC    

年代:1968

数据来源: RSC

摘要:

Analyst, June, 1968, Vol. 93, pp. 375-382 375 Studies of the Separation of Trace Metals by the Manganese Dioxide “Collection” Method Part 111.” The Behaviour of Copper and Zinc. Further Studies of the Behaviour of Lead and Tin: The Determination of Lead and Tin in Coppepbase Alloys BY C . M. PYBURN AND G. F. REYNOLDS? (Loughborough University of Technology, Loughborough, Leicestershire) Studies of the collection of copper and zinc by co-precipitation with manganese dioxide are reported. The collection of copper is shown to be small under all conditions studied; that of zinc is negligible. Further studies of the collection of lead and tin are presented, and a method for their separate and simultaneous determination is described. The virtual non-collection of copper and zinc is used as a basis for the application of these methods to the determination of lead and tin in copper-base alloys.The collection of lead and tin is made from a 0-008 M acidic solution containing ethanol to hold the tin in solution. After dissolution of the precipitate, tin and lead are determined together by polarography in acidic chloride solution. Lead is then determined alone by polarography in alkaline mannitol solution and the tin content found by difference. Results are presented and some aspects discussed. THE use of manganese dioxide, produced by the reaction of manganese(I1) ion with potassium permanganate, as a “collector” for the co-precipitation of metals was first described by Blumentha1,l who used it for antimony. He indicated that the procedure could also be applied to bismuth and tin.Kallman and Prestira2 showed that the precipitation of bismuth was incomplete in acidic solutions of concentration greater than 0.07 N. Previous work by one of us has shown that antimony and tin are collected over a wide range of acid concen- trations. Similar results have been obtained by Babko and Shtoka10.~ Several analytical applications have been reported, for example, the determination of antimony with Rhodamine B by MacNulty and Woollard,4 antimony in copper by Park and Lewis,s antimony and bismuth in anode copper by Yamazaki,B arsenic and tin in lead by Luke7 and antimony in cast iron by Rooney.8 No previous comprehensive study has, however, been made of the elements to which the procedure can be applied, or the con- centration range or conditions in which it is efficient for each species.The present work was undertaken to provide a comprehensive study, as mentioned above. Previous papers in the series have established conditions and concentration ranges for the “collection” of antimony, bismuth and tin,g and for tin and 1ead.lO In the present work, studies of copper and zinc have been made and further information on the behaviour and determination of lead and tin is presented. EXPERIMENTAL The “collection” procedure previously describedg Treat the sample solution, in a 400-ml beaker, with sufficient nitric acid solution to make the volume up to 200m1, add 5ml of 5 per cent. manganese(I1) sulphate solution and heat to boiling, then add 2-5 ml of 1.25 per cent. potassium permanganate solution, dropwise, and continue boiling for a further 2 minutes.Allow the solution to stand a t a was used in all experiments. * For details of Parts I and I1 of this series, see reference list, p. 382. t Present address: Shandon Scientific Co. Ltd., 65 Pound Lane, Willesden, London, N.W.10. 0 SAC and the authors376 PYBURN AND REYNOLDS: STUDIES OF THE SEPARATION OF TRACE METALS [Analyst, Vol. 93 temperature of 70” C for about 30 minutes and then filter off the manganese dioxide pre- cipitate with an 11-cm, Whatman No. 40 filter-paper. Wash the precipitate three times with 10-ml portions of the nitric acid solution used for the initial dilution; the last traces of precipitate are transferred from the beaker to the filter-paper by the first two washings.Two methods for the dissolution of the manganese dioxide have previously been des- cribed10; that in which a mixture of nitric acid and hydrogen peroxide is used was adopted for this work, as follows. Dissolve the manganese dioxide precipitate completely by allowing about 1 O m l of a hot mixture of 1.2 M nitric acid and 100-volume hydrogen peroxide (20 + 1) to run down the sides of the filter-paper. Wash the filter-paper with a further 15ml of the nitric acid- hydrogen peroxide solution and then with a small volume of water. Combine the solution and washings in a 400-ml beaker, then dilute to about 100 ml and boil gently to remove the bulk of the residual hydrogen peroxide and allow to cool. Bubble sulphur dioxide gas slowly through the solution to convert the last traces of hydrogen peroxide into sulphuric acid.Boil the solution to remove excess of sulphur dioxide, cool, transfer to a 250-ml calibrated flask and make up to volume with water. All determinations reported in this paper were made by d.c. or a.c. polarography. All polarography was carried out on a Cambridge pen-recording polarograph, in conjunction with which a Cambridge univector unit was used to provide a.c. facilities. All potentials quoted are on the European sign convention. According to this convention, the potential of the saturated calomel electrode is taken as +0.246 volt versas the normal hydrogen electrode. “COLLECTION” OF COPPER- A series of solutions, each containing 20 mg of copper and with initial acid concentrations within the range 1.2 to 0408 M, was prepared and subjected to the “collection” procedure described above.The manganese dioxide was re-dissolved with nitric acid and hydrogen peroxide, and made up to 250 ml in calibrated flasks, as already described. Twenty-five millilitre portions of each solution were placed in 50-ml calibrated flasks and made up to volume with 0.2 M sodium potassium tartrate. Aliquots were placed in polaro- graphic cells, de-oxygenated by passage of nitrogen gas for 5 minutes and subjected to d.c. and ax. polarography. Well shaped steps and sharp ax. peaks were obtained, with half- wave potentials -0.24 volt versus mercury pool. Concentrations of copper recovered were calculated by reference to a calibration graph. 8 Applied potential (volts versus mercury pool) Fig.1. Polarography of copper in 0.1 M sodium potassium tartrate fter collection on manganese dioxide: (a) d.c. polarographic step: (b) ax. polarographic wave A typical copper step and an a.c. wave are shown in Fig. 1. Copper recoveries are presented in Table I. The results in Table I show that the most efficient collection took place in a solution 0*008 M in acid. Further experiments were, therefore, made at this acid concentration, to study the efficiency of “collection” as a function of initial copper concentration. A series of solutions was prepared as before, but with an initial nitric acid concentration of 0.008 M andJune, 19681 BY THE MANGANESE DIOXIDE “COLLECTION” METHOD. PART 111 TABLE I EFFECT OF NITRIC ACID CONCENTRATION ON THE RECOVERY OF COPPER 377 Acid concentration, 1.2 1-2 0.12 0.12 0.06 0-06 0.03 0-03 0.015 0.015 0.008 0.008 0.004 0.004 0.002 0.002 M Copper * Added, Found, mg mg 20 0-088 20 0.080 20 0,088 20 0.088 20 0.095 20 0-096 20 0.080 20 0.10 20 0.075 20 0.075 20 0.60 20 0.48 20 0.18 20 0.18 20 0.18 20 0.076 Recovery, per cent.0-43 0.40 0.43 0.43 0.48 0.48 0.40 0.50 0-38 0-38 2.50 2.38 0.95 0.95 0.88 0.38 with various copper concentrations covering the range 5 to 100mg. These solutions were treated and polarographed exactly as already described. Copper recoveries are given in Table 11. TABLE I1 RECOVERY OF COPPER Copper & Recovery, Added, mg Found, mg per cent. 6 0.055 1.1 5 0.055 1.1 10 0.08 0.8 10 0.125 1.25 15 0.125 0.83 15 0.10 0.67 20 0.50 2.5 20 0.475 2.38 25 0-325 1.3 25 0.14 0.56 50 0.22 0.44 50 0.206 0.4 1 100 0-225 0.226 100 0.17 0.17 “COLLECTION” OF ZINC- A series of solutions, each containing 20 mg of zinc and with initial acid concentrations covering the range 1.2 to 0-008 M, was prepared and treated exactly as described for copper.It was found that polarography of these solutions yielded highly variable results because of the appearance of a step with a half-wave potential displaced from that expected for zinc. These experiments were repeated with various base electrolytes suitable for the determination of zinc, with similar results. Blank determinations were, therefore, made in the absence of zinc, and from the results obtained it was deduced that the interfering step was caused by residual manganese ion. Consideration was, therefore, given to the use of a base electrolyte that would remove the manganese interference by complex formation, which has no effect on the zinc half-wave potential.This was achieved by the use of M potassium hydroxide and 0.2 M mannitol.11 This completely removed manganese interference in the region of the zinc half-wave potential, which was -1.2 volts versus mercury pool. The results obtained with a fresh series of solutions, under these conditions, showed that only traces of zinc were “collected” over the whole range of acid concentration studied. The largest trace, however, occurred with solutions of initial acid concentration 1-2 M.378 PYBURN AND REYNOLDS: STUDIES OF THE SEPARATION OF TRACE METALS [Analyst, Vol. 93 Further experiments were carried out with a series of solutions with zinc concentrations in the range of 2 to 1OOmg in 1.2 M acid.No measurable amount of zinc was recovered, even at the highest zinc concentration studied. “COLLECTION” OF LEAD AND TIN- Previous workg has shown that tin is quantitatively “collected” from solutions of acid concentration 1.2 M, but that the efficiency of collection fell to 76 per cent. when the acid concentration was reduced to 0.008 M . ~ O It has also been shownlO that lead is not “collected” from solutions 1-2 M in acid, but that good recoveries are obtained from 0.008 M acid. In view of the low recoveries of copper and zinc reported above, it appeared that procedures could be developed for the determination of tin and lead in copper-base alloys. It was evident that straightforward procedures were feasible for the determination of tin in the presence, or absence, of lead, and for the determination of lead in the absence of tin’ DETERMINATION OF TIN- “Collection” was carried out, as already described, from a copper - zinc solution containing 1.2 M nitric acid.In view of previous results,1° dissolution of the precipitate with a mixture of nitric acid and hydrogen peroxide was not used, as this has been shown to cause oxidation of the tin to tin(1V) oxide. Instead, the filter-paper and precipitate were placed in a beaker and treated with 3 ml of 18 M sulphuric acid and 5 ml of 16 M nitric acid. The filter-paper was then destroyed by repeated evaporation, further amounts of nitric acid being added as necessary. After oxidation of all carbon to give a clear solution, careful evaporation, almost to dryness, was carried out.The residue was allowed to cool, 15ml of 36 per cent. hydrochloric acid were added and the solution diluted to about 50 ml. After heating for a few minutes, the residue dissolved to give a clear solution, which was cooled and transferred to a 250-ml calibrated flask. The flask was made up to volume with washings from the beaker. Fifty millilitres of the solution were transferred to a 100-ml calibrated flask, treated with 10 ml of 36 per cent. hydrochloric acid, 10 ml of N potassium chloride and 10 ml of 5 N sodium hydroxide before being finally made up to volume with water. An aliquot was transferred to a polarographic cell, de-oxygenated by passage of nitrogen for 5 minutes and polarographed for tin.The tin concentration was calculated by reference to a calibration graph. A series of standard copper - zinc solutions containing tin and lead were treated as Tin steps of good shape were obtained, the step heights of which were unaffected by described above. the presence of lead in the original solution. Recoveries are presented in Table 111. TABLE I11 RECOVERY OF TIN FROM SOLUTIONS CONTAINING 0.6 g OF COPPER AND 0.4 g OF ZINC Lead Added, mg 0 0 5 5 5 10 10 10 Tin 1 Added, mg Found, mg 2.6 2.4 2.5 ,2* 5 6.0 4-95 6-0 5-0 10.0 10.2 5.0 4.9 5-0 4.9 10.0 10.0 DETERMINATION OF LEAD (TIN ABSENT)- The “collection” and dissolution procedure used was exactly the same as that used for the studies of copper and zinc. The final solution, after dissolution of the precipitate with the nitric acid - hydrogen peroxide mixture, was made up to 250 ml in a calibrated flask.Fifty millilitres of this solution were transferred to a 100-ml calibrated flask, treated with 10ml of N potassium chloride and made up to volume with water.June, 19681 BY THE MANGANESE DIOXIDE “COLLECTION” METHOD, PART 111 379 An aliquot of the solution was transferred to a polarographic cell, de-oxygenated by passage of nitrogen for 5 minutes and polarographed for lead. The lead concentration was calculated by reference to a calibration graph. A series of copper - zinc solutions, containing lead, was treated by the above procedure. Good steps were obtained with half-wave potentials of about -0.5 volt versus mercury pool.Recoveries are presented in Table IV. TABLE IV RECOVERY OF LEAD FROM SOLUTIONS CONTAINING 0.6 g OF COPPER AND 0-4 g OF ZINC Lead P Added, m g Found, mg 2 2.04 2 2.00 5 4-85 5 4.90 10 9.90 10 10-10 DETERMINATION OF LEAD AND TIN- In order to attempt to develop a method for the simultaneous determination of lead and tin, with a single “collection,” a re-investigation of the behaviour of these two elements in 0.008 M acid was carried out. This arose as the result of observations that a white pre- cipitate formed on boiling tin solutions at this acidity, in the presence of manganese sulphate, before the addition of potassium permanganate. The formation of this precipitate was confirmed and it was removed and examined polarographically. It was found to contain about 77 per cent.of the added tin. It was evident from the above results that the “collection” of tin from 0 - 0 0 8 ~ acid, reported by Reynolds and Tyler,lo was not a true manganese dioxide collection, as the bulk of the tin was precipitated before the formation of the collector. Methods were, therefore, sought to retain the tin in solution at this acid concentration, Many complexing reagents were tried without success, but it was finally found that the presence of ethanol prevented tin precipitation. Furthermore, the ethanol did not react with the potassium permanganate and did not interfere with the “collection” process. The addition of 50 ml of ethanol to the initial solution was shown to be sufficient, and tin recoveries from these 0.008 M acidic solutions were almost 100 per cent.In the presence of ethanol, the nitric acid - hydrogen peroxide method for dissolution of the precipitate was satisfactory. A common procedure for the “collection” and dissolution of lead and tin could, therefore, be used. The procedure for simultaneous polarographic determination of lead and tin in copper- base alloys was based on a method described by Lingane.12 In this method, polarography is performed in acidic and in alkaline solutions. In the former, both lead and tin are reduced at similar half-wave potentials to yield a combined step, whereas in the latter, a lead step only is obtained. A series of “collections” from lead - tin solutions was made, as already described, but with the addition of 50ml of ethanol to the initial solution, after adjustment of the acid concentration to 0*008 M.After dissolving the precipitate and making up the volume in a 250-ml calibrated flask, two 50-ml aliquots were withdrawn and placed in 100-ml calibrated flasks. One of these was treated with 10 ml of 11 M hydrochloric acid, 10 ml of M potassium chloride and 10 ml of 5 M potassium hydroxide. The solution was then made up to volume with water. The other aliquot was treated with 10 ml of 5 M potassium hydroxide and made up to volume with water. Polarography of solutions prepared as above yielded combined lead - tin steps from the acidic aliquot and steps for lead alone from the alkaline solution. The latter, however, suffered interference from reduction of the residual manganese present. This series was, therefore, repeated with addition of 1 g of mannitol to each lead determination before making up to volume (see under “Collection” of zincll) .This eliminated the interference.380 PYBURN AND REYNOLDS: STUDIES OF THE SEPARATION OF TRACE METALS [AfidySt, VOl. 93 Initial recoveries were promising, but an unacceptably high percentage of erratic results occurred. This difficulty was overcome by increasing the amount of potassium permanganate added in the “collection” procedure, from the usual 2-5 ml of 1-25 per cent. solution to 5 ml of 1-25 per cent. solution. It was found, however, that this increase in permanganate con- centration caused manganese interference with the lead step in alkaline solution to recur and it was necessary to increase the amount of mannitol added to 3 g.Recoveries of lead and tin obtained from the series of solutions treated by the final procedure are given in Table V. TABLE V RECOVERIES OF LEAD AND TIN Lead & Added, mg Found, m g 5 4.95 5 4.65 5 4.70 5 4.96 5 4.50 10 9.95 10 9.80 Tin Added, mg 0 2.6 3.0 7.5 10.0 7.5 10.0 Found,. mg 0 2.6 2.86 7.73 10.0 7.80 10.20 In view of the satisfactory nature of the above results, a method was described for the determination of lead and tin in copper-base alloys. METHOD Place 1 g of the copper-base alloy sample (as drillings or turnings) in a 400-ml beaker and treat it with the minimum volume of 2 N nitric acid. Warm gently to complete dissolution, neutralise to litmus and cool. Add 200 ml of 0.008 M nitric acid, 5 ml of 5 per cent. man- ganese(I1) sulphate solution and 50 ml of ethanol.Heat to boiling, add 5 ml of 1.25 per cent. potassium permanganate solution, dropwise, and continue boiling for a further 2 minutes. Allow the solution to stand at 70” C for 30 minutes and then filter off the manganese dioxide precipitate through an 1 l-cm, Whatman No. 40 filter-paper. Wash the precipitate three times with 0408 M nitric acid solution, the first two washings being used to transfer the last traces of precipitate from the beaker to the filter-paper. Dissolve the manganese dioxide precipitate completely by allowing about 10ml of a hot mixture of 1.2 N nitric acid and 100-volume hydrogen peroxide (20 + 1) to run down the sides of the filter-paper. Wash the filter-paper with a further 15 ml of the nitric acid - hydrogen peroxide solution and then with a small volume of water.Combine the solution and washings in a 400-ml beaker. Boil gently to remove the bulk of the residual hydrogen peroxide. Allow to cool and bubble sulphur dioxide gas slowly through the solution to convert the last traces of hydrogen peroxide into sulphuric acid. Cool, transfer it into a 250-ml calibrated flask and make up to volume with water. DETERMINATION OF LEAD plus TIN- Place 50 ml of the prepared solution in a 100-ml calibrated flask, add 10 ml of 11 N hydrochloric acid, 10 ml of M potassium chloride and 10 ml of 5 M potassium hydroxide, then make up to volume with water. Transfer an aliquot to a polarographic cell, de-oxygenate by passage of nitrogen gas for 5 minutes and record a polarogram, versus the mercury pool as anode.Measure the height of the combined lead - tin step, which occurs at a half-wave potential of about -0-5 volt versus mercury pool. DETERMINATION OF LEAD- Place 50 ml of the prepared solution in a 100-ml calibrated flask and add 10 ml of 5 M potassium hydroxide and 3 g of mannitol. Transfer an aliquot to a polarographic cell, de-oxygenate by passage of nitrogen gas for 5 minutes and record a polarogram versus the mercury pool as anode. Measure the height Make up to volume with water.June, 19681 381 of the lead step that occurs at a half-wave potential of about -0.75 volt versus mercury pool. Calculate the lead content of the sample from a calibration graph prepared with standard solutions, or alloys, subjected to the full procedure.DETERMINATION OF TIN- Subtract the height of the lead step from the height of the combined lead - tin step. Calculate the tin content of the sample from a calibration graph prepared with lead-free standard solutions, or alloys, subjected to the full procedure. BY THE MANGANESE DIOXIDE “COLLECTION” METHOD. PART 111 TRIAL OF THE METHOD A series of solutions was prepared, each containing 0.6 g of copper and 0.4 g of zinc, and treated with varying amounts of standard lead and tin solutions. These solutions were subjected to the full procedure, as described above. The results obtained are given in Table VI. TABLE VI RECOVERY OF LEAD AND TIN FROM SOLUTIONS CONTAINING 0.6g OF COPPER AND 0-4g OF ZINC Lead & Added, mg Found, mg A 6.6 6.3 A 6.6 6.9 B 6-0 4.7 B 6.0 4.8 C 7.6 7.46 C 7.6 7.4 D 4.0 4.0 D 4.0 3.9 Tin t-,--7 Added, mg Found, mg 5.3 6-26 6.3 6.3 10.3 10.3 10.3 10.3 6.3 5.3 6.3 6.2 9.4 9.4 9.4 9.35 In view of the satisfactory results above, several determinations were made with samples of special copper-base alloys, the tin and lead contents of which had been determined gravi- metrically. The results of these analyses are presented in Table VII.TABLE VII RECOVERY OF LEAD AND TIN FROM COPPER-BASE ALLOYS Tin & 7 Present, Found, Present, Alloy per cent. per cent. per cent. Special alloy HC (1) 0- 6 0.62 0-26 (2) 0.6 0.6 1 0.26 Special alloy CA* (1) 4.45 4-40 2.2 (2) 4-45 4.35 2.2 Special alloy TU (1) 0.36 0.33 1.0 (2) 0.35 0.35 1.0 Special alloy TEN* (1) 2-05 2-00 3.5 (2) 2.06 2.00 3.5 0-6 g of sample used, instead of 1 g as specified.Lead - Found, per cent. 0.24 0.24 2-05 2.10 1.06 0-96 3.40 3.46 DISCUSSION The results presented in Tables I and I1 show that copper cannot be efficiently collected on manganese dioxide. It has also been shown that collection of zinc is negligible under all conditions studied. The optimum acid concentration for “collection” of copper was found to be 0.008 M, but even under these conditions the recovery of copper was only 2-5 per cent. of 20mg added. In general, an increase in copper concentration tended to increase the amount of copper recovered, but the percentage efficiency of the “collection” decreased sharply.382 PYBURN AND REYNOLDS Mannitol in alkaline solution is a valuable complexing reagent for manganese, as it forms a species that yields an anodic wave in the region of -0.2 volt versus mercury pool.Inter- ference by manganese in the region of 1 volt is therefore eliminated.ll This species also provides a valuable means of determining manganese in the presence of other species whose polarographic step would normally precede that of manganese. The inefficient “collection” of copper and zinc has been turned to advantage by the development of procedures for the determination of lead and tin in copper-base alloys. Tables I11 and IV show that good results are obtained for tin in the presence, or absence, of lead and for lead in “tin-free” alloys. The investigation of the tin precipitation, in the course of developing the method for the simultaneous determination of lead and tin, has provided interesting new information.It is evident that a species must be in true solution for effective “collection” to take place. Otherwise, presumably, the particle size is too large for efficient adsorption to occur on the manganese dioxide surface. Further fundamental studies are desirable. I t would appear, also, that “collection” in the form of simple ion species is desirable, as complexing reagents did not enhance the efficiency of collection, although they assisted in retaining the tin in solution. The mode of action of the ethanol is not fully understood. The method for the determination of lead and tin in copper-base alloys is shown to be reproducible within the range studied (Table VI) and to be applicable to alloys with a variety of tin and lead contents.In this method, a direct correction for the contribution of lead to the combined lead - tin step obtained in acidic solution is made by reference to the height of the lead step obtained in alkaline solution. This is not completely valid, as the diffusion coefficients, and hence the diffusion-current constants, will not be the same in the two media. However, Lingane has shown13 that the ratio of diffusion-current constants in acidic and alkaline solution is 1 : 1.036, so that the above assumption may be made without serious error. The accuracy of the results will, of course, be enhanced if the appropriate correction is made. The chloride content of the base electrolyte used for the determination of tin (1-2 N) is less than that generally recommended. A concentration of 5 to 6 N is more usual. This concentration was chosen to avoid possible loss of lead as chloride when both elements are present together. The results (Tables 111, V and VI) indicate that the present chloride concentration is adequate to give the chloro-complex of tin(1V) that is considered desirable for satisfactory polarography. 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. REFERENCES Blumenthal, H., 2 . analyf. Chem., 1928, 74, 33. Kallman, S., and Prestira, F., Ind. Engng Chem. Analyt. Edn, 1941, 13, 8. Babko, A. F., and Shtokalo, M. I., Zav. Lab., 1955, 21, 767. MacNulty, B. J., and Woollard, L. D., Analytica Chim. Acta, 1955, 13, 64. Park, B., and Lewis, E. J., Ind. Engng Chem. Analyf. Edn, 1933, 5 , 182. Yamazaki, Y., Bunseki Kagaku, 1957, 82, 619. Luke, C. L., Ind. Engng Chem. Analyt. Edn, 1943, 15, 626. Rooney, R. C., Analyst, 1957, 82, 619. Ogden, D., and Reynolds] G. F., Ibid., 1964, 89, 538. Reynolds, G. F., and Tyler, F. S., Ibid., 1964, 89, 579. Reynolds, G. F., and Shalgosky, H. I., Analyfica Chim. Acta, 1954, 10, 273. Lingane, J. J., Ind. Engng Chem. Analyt. Edn, 1946, 18, 429. -, “Electroanalytical Chemistry,” Interscience Publishers Inc., New York and London, 1953, p. 318. NOTE-References 9 and 10 are to Parts I and I1 of this series, respectively. Received November 17th. 1967

ISSN:0003-2654    

DOI:10.1039/AN9689300375

出版商:RSC    

年代:1968

数据来源: RSC

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Analyst, June, 1968, Vol. 93, pp. 383-387 383 The Spectrophotometric Determination of Titanium in Iron and Steel with Diantipyrylmethane BY J. A. CORBETT (Physical Metallurgy Section, Commonwealth Scientific 6. Industrial Research Organization, Baillieu Labmatovy, University of Melbourne, Australia) The use of diantipyrylmethane its a spectrophotometric reagent for the determination of titanium in iron and steel has been investigated. A rapid method is suggested in which the reagent, which permits the determination of titanium in a wide range of iron and steels, is used. Results are given for the analysis of stainless, high speed and mild steels, and cast irons. THE British Standards Institution has recently issued B.S. 1121 : Part 47 : 1966, which gives a method for the spectrophotometric determination of titanium in iron and steel, in which hydrogen peroxide is used to form a coloured complex.This method is similar to the American Society for Testing Materials method and, while quite satisfactory, is slow and tedious, particularly when molybdenum and vanadium are present. Several atomic- absorption methods1 s2 have been suggested but they have limited sensitivity. In seeking a more rapid method for general application, a study was made of various reagents, and diantipyrylmethane appeared to offer many advantages. The optimum conditions given in the literature under which diantipyrylmethane is used vary considerably. Polyak3 states that the optimum acid concentration, with respect to hydrochloric acid, lies within the range 1 to 4 M, and gives a development time of 40 to 50 minutes before measuring the absorbance. Later, Polyak4 introduced tartaric acid to keep niobium in solution when determining titanium in the presence of niobium, and allowed a development time of 2 hours.Jeffery and Greg~ry,~ when using 1 M hydrochloric acid, allow 3 hours for development. They also use tartaric acid when niobium is present and again develop for 3 hours. Lazareva and Lazarev6 conclude that the rate of formation of the coloured complex decreases as the hydrochloric acid concentration increases, but the complex formed is always the same. They consider that the optimum concentration, with respect to hydrochloric acid, is 0.25 M. They further show that when sufficient tartaric acid is present, a different complex is formed, with an absorption maximum at 326 to 330 nm, instead of 380 to 400 nm without tartaric acid.In methods of analysis given by the above workers, sulphuric acid is used in the prepara- tion of sample and standard solutions. The concentrations of sulphuric acid vary considerably and in some instances the amounts used are not specified at all. Two reactions are involved: first, the breakdown of titanium polyions to TiO+, and then the formation of the titanium oxide (TiO) - diantipyrylmethane complex. The de-poly- mensation rate of the titanium ions is slow and can be expected to be affected by the acid concentration and the presence of complexing agents, such as tartaric acid. The large excess of diantipyryImethane recommended (300 times the titanium concentration) suggests a low equilibrium constant for the titanium oxide - diantipyrylmethane reaction.The effects of the presence of other cations on the reaction have been studied by Pol~ak.~s* Nickel, chromium, copper, manganese and cobalt (11) do not react with diantipyrylmethane. Niobium, if present in high concentrations, suppresses the titanium - diantipyrylmethane reaction. Iron and vanadium do not interfere if reduced to their bi- and quadrivalent forms. Tungsten, in amounts exceeding 10 mg per 100 ml, precipitates with diantipyrylmethane, 0 SAC; Crown Copyright Reserved384 CORBETT : SPECTROPHOTOMETRIC DETERMINATION OF [ArtdJU!, VOl. 93 but smaller amounts do not interfere. Molybdenum, if reduced to the quinquivalent form, does not precipitate, but causes an increase in absorbance necessitating the addition of molybdenum to the calibration standards.’ Ascorbic acid is added to reduce iron, vanadium and molybdenum; the yellow titanium - ascorbic acid complex is destroyed by adding hydrochloric acid.3 EXPERIMENTAL Diantipyrylmethane (obtainable from Aldrich Chemical Co., Milwaukee, U.S.A.) was dissolved in various aqueous solutions containing acid; a molar solution of hydrochloric acid was found to be suitable.A stock titanium solution was prepared by dissolving high purity titanium metal in dilute sulphuric acid (1 + 3) and oxidising it with the minimum amount of dilute nitric acid (1 + 1). This solution was made up to volume so that the final sulphuric acid concen- tration was 2 M. To control the acid concentration, and also to avoid hydrolysis, dilutions were made of the stock solution with water immediately before use and the solutions were afterwards discarded.Solutions of diverse ions were prepared from high purity metals or analytical-reagent grade chemicals. RESULTS Tests were carried out with the conditions and reagents as recommended by other workers, viz., with 0 to 100 pg of titanium per 100 ml, 1 M hydrochloric acid, 0.006 M dianti- pyrylmethane and varying concentrations of sulphuric and tartaric acids. The following results were obtained. Beer’s law was not obeyed with amounts of titanium above 50 pg, suggesting that even a 300-fold excess of the reagent was not sufficient. Varying concentrations of sulphuric, hydrochloric and tartaric acids all had an influence on the development time before colour measurement and on the final absorbance.The wavelength - absorption plot was the same in all of the tests. The departure from Beer’s law was not serious, but, to achieve a reasonable precision, the variations in development time and absorbance would make it necessary to ensure that the calibration standards were treated in exactly the same manner as the test solution. Tests were carried out in which both the titanium and diantipyrylmethane concentrations were increased with the aim of increasing the breakdown rate of the titanium polyions. This was effected by reducing the final volume from 100 to 50 ml. To conserve the diantipyryl- methane, the excess was reduced to about 250-fold at the 1OO-pg titanium level.The results obtained, as shown in Fig. 1, indicate a considerable improvement in tolerance to variations in acid concentration and the presence of tartaric acid. Concentrations of hydrochloric acid up to 1 M give the same absorbance after 1 hour. Sulphuric acid in the concentrations tested, while not affecting the development time, caused a lower absorption. As the concentration of tartaric acid increases, the development time increases until, at 2.6 M, equilibrium has not been reached after 5 hours, but lower concentrations (0.01 M) can be accommodated. An improvement was also obtained in the plot for Beer’s law, although with amounts of titanium above 100pg Beer’s law was not obeyed. The addition of sodium or potassium sulphate in amounts up to 0.5 g per 50 ml in 0.5 M hydrochloric acid has no effect. Apparently, it is the hydrogen-ion concentration that causes the lower absorbance in the presence of sulphuric acid.Evidence of the formation of a titanium oxide - diantipyrylmethane - tartaric acid com- plex was obtained by using the conditions and reagents as described by Lazareva and Lazarev (viz., 0.25 M hydrochloric acid, 0.05 M diantipyrylmethane and 0.3 to 0.5 M tartaric acid). An absorption maximum was obtained at 326 to 330nm, but the complex slowly changed with time to the titanium oxide - diantipyrylmethane complex, with an absorption maximum at 380 to 400nm. The addition of ascorbic acid has no effect on the development time - absorbance curves shown in Fig. 1. No difficulty was experienced as a result of the formation of a titanium - ascorbic acid complex, probably because of the high acid concentration present when the ascorbic acid is added; the aliquot is later diluted with water to give the required acid concentration.June, 19681 TITANIUM IN IRON AND STEEL WITH DIANTIPYRYLMETHANE 385 The presence of 100mg of iron, 20mg of nickel, 20mg of chromium, 5 mg of copper, 5 mg of vanadium, 2 mg of manganese and 2 mg of tungsten ions per 50 ml of solution has no effect when ascorbic acid is present, and the absorption measurements are made with a reference solution containing the same amounts of these cations and ascorbic acid.Molybdenum, after reduction with ascorbic acid, was found to form a weak yellow complex with diantipyrylmethane (1 mg of molybdenum is equivalent to 1-5 pg of titanium).This interference was prevented by forming a complex of the molybdenum with tartaric acid. Tests showed that tartaric acid at a concentration of 0.013 M overcame interference from amounts of molybdenum up to 1Omg. Tungsten precipitates with diantipyrylmethane under the conditions used in these tests when more than 2 mg is present in the 50-ml aliquot. Most of the tungsten in steels can be rendered insoluble by treatment with a suitable acid, the amount remaining in solution in the final aliquot being less than 2mg. 0.3 E i f a . .. . . .. J L I I I I I Niobium is retained in solution by forming a complex with tartaric acid. However, it is preferable to restrict the amount of tartaric acid used, as shown in Fig. 1. As a result of these tests, it appeared possible to adopt a set of conditions that would permit the determination of titanium in a wide range of iron and steels. The reagents and conditions adopted were: 0.013 M tartaric acid, 0.01 M diantipyryl- methane, 0.5 M hydrochloric acid and 0.06 M ascorbic acid in a 50-ml volume, with a develop- ment time of 90 minutes.The complex is stable over the temperature range 18" to 30" C, and the extinction coefficient at 390 nm is 14,500. The standard deviation of the absorbance is 0.0025. When molybdenum and niobium are known to be absent the tartaric acid can be excluded and the development time reduced to 1 hour. The recovery of titanium from the acid-insoluble material may not be necessary with some samples.386 CORBETT : SPECTROPHOTOMETRIC DETERMINATION OF [Analyst, Vol.93 Table I shows the results obtained by using the method with standard samples of various steels. TABLE I RESULTS OBTAINED WITH THE METHOD DESCRIBED COMPARED WITH THE AVERAGE B.S. 1121 : PART 47 RESULT OBTAINED BY FIVE DIFFERENT LABORATORIES WITH THE STANDARD METHOD B.S. 1121 :Part 47 method Sample Type per cent. N.B.S. 170A B.O.H. 0.282 S.A.A. 101 High speed steel (chromium 4, molyb- 0.006 S.A.A. 106 Stainless steel (18/8) 0.81 S.A.A. 106 Austenitic iron (nickel 4, chromium 3 0.063 S.A.A. 108 Stainless steel (18/8) (niobium 1 per 0.014 S.A.A. 112 Stainless steel (18/8) (molybdenum 3 0.424 S.A.A. 114 Cast iron (vanadium 0-2 per cent.) 0.032 B.C.S. 322 Mild steel 0.043 S.A.A. 101 + 100 pg of titanium S.A.A. 112 denum 6 and tungsten 6 per cent.) and copper 6 per cent.) cent.) per cent.) S.A.A.106 + 10 mg of molybdenum - Recovery 106 pg = 100 per cent. Mean of 7 results = 0.426 per cent.; standard deviation = 0.003. (S.A.A. refers to Standards Association of Australia.) Diantipyrylmethane, method, per cent. 0.282, 0.282 0.006, 0.006 0-828, 0.824 0.060, 0.061 0.018, 0.017 0.424, 0.428 0-036, 0.037 0.046, 0-046 0.051 METHOD REAGENTS AND SOLUTIONS- Titanium sohtion-Dissolve O-lOOO g of high purity titanium metal in 50 ml of dilute sulphuric acid (1 + 3) and oxidise the titanium by adding nitric acid, dropwise. Cool, transfer the solution into a 1-litre calibrated flask, dilute to the mark with dilute sulphuric acid (1 + 9) and mix. Transfer 10 ml of this solution into a 100-ml calibrated flask, add 10 ml of tartaric acid solution (10 per cent.), make up to the mark with water and mix.This solution must be prepared freshly each day. Tartaric acid solution, 10 per cent. w/v-Dissolve 100g of tartaric acid in water and dilute to 1 litre. Ascorbic acid solution, 10 per cent. w/v-Dissolve 100 g of ascorbic acid in water and dilute to 1 litre. Diantipyrylmethane solution, 2 fier cent. w/v-Dissolve 2 g of diantipyrylmethane in dilute hydrochloric acid (1 + 9) and make up to 100 ml with dilute hydrochloric acid (1 + 9). PROCEDURE- Weigh 0.5 or 1 g of sample (Note 1) and transfer it into a 250-ml beaker. Add 20 ml of hydrochloric acid (sp.gr. 1-16 to 1*18), cover the beaker and digest until solvent action ceases, add 5ml of nitric acid (sp.gr.1-42) and evaporate the solution to dryness (Note 2). Add 20 ml of dilute hydrochloric acid (1 + 3) and heat to dissolve the iron salts. Filter the solution through an ll-cm No. 41 filter-paper and wash the paper with hot water. Remove the iron salts by washing the paper with 10 ml of dilute hydrochloric acid (1 + 1) and water (Note 3). Transfer the filter-paper into a platinum crucible, dry and ignite it at a temperature of 700" C. Cool, add several drops of dilute sulphuric acid (1 + 1) and 2 ml of hydrofluoric acid (40 per cent.), evaporate to dryness, ignite at 700" C, and cool. Fuse the residue with 1 g of potassium hydrogen sulphate and cool. Dissolve the melt by heating it with 10 ml of tartaric acid solution (10 per cent.), and add the solution to the original filtrate.Transfer two aliquots (Note 1) into 60-ml calibrated flasks, add dilute hydrochloric acid (1 + l), to make the final concentration 0.5 M with respect to hydrochloric acid (Note I), 1 ml of solution = 10 pg of titanium.June, 19681 TITANIUM IN IRON AND STEEL WITH DIANTIPYRYLMETHANE 387 and 5 ml of ascorbic acid (10 per cent.), mix, and allow the solution to stand for 10 minutes. To one flask, add 10 ml of diantipyrylmethane solution (2 per cent.), and make the solutions in both flasks up to the mark with water and mix. Allow the solutions to stand for 90 minutes and measure the absorbance of the coloured complex at 390 nm, with the blank as a reference solution, in cells of a suitable size (Note 1). Determine the blank value of the reagents concurrently with the test determination. CALIBRATION- Titanium in the range 0 to 0.07 per cent.-Transfer 0, 1.0, 2.0, 3.0, 4.0, 5-0, 6-0 and 7.0 ml of titanium solution (1 ml of solution = 10 pug of titanium) into 50-ml calibrated flasks, add 3 ml of dilute hydrochloric acid (1 + 1) and 5 ml of ascorbic acid solution (10 per cent.), mix, and allow to stand for 10 minutes.Add 10ml of diantipyrylmethane solution (2 per cent.), make up to the mark with water and allow the solutions to stand for 90 minutes. Measure the absorbance of the solutions at 390 nm in 2-cm cells, with the solution containing no titanium as a reference solution. Titanium in the range 0 to 1.20 #er cent.-Transfer 0, 2.0, 4.0, 6.0, 8-0, 10.0, 12-0, 14.0 and 150ml of the same titanium solution into 50-ml calibrated flasks and treat them as before, but measure the absorbance in l-cm cells. NOTES- 1. Appropriate weights and dilutions are listed below. Titanium range, per cent. 0 to 0.07 0 to 0.15 0 to 0-30 0 to 0.60 0 to 1.20 Weight of sample, Dilution, Aliquot, g ml ml 1.0 100 10 1.0 100 10 0.5 100 10 0.5 200 10 0.5 200 6 Hydrochloric acid (1 + 1) added, Cell size, mI cm 1.0 2 1.0 1 1.0 1 2.0 1 2.6 1 2. Additional nitric acid may have to be added to dissolve some steels. When this is necessary, evaporate the solution to dryness, dissolve the residue in 10 ml of hydrochloric acid (sp.gr. 1-16 to 1.18) and evaporate to dryness again. Continue as in Procedure. 3. If tungsten is present, wash the precipitate with ammonia solution (1 + 1). REFERENCES 1. 2. 3. 4. 6. 6. 7. Bowman, J. A., and Willis, J. B., Analyt. Chem., 1967, 39, 1210. Headridge, J. B., and Hubbard, D., Analytica Chim. Acta, 1967, 37, 161. Polyak, L. Ya., Zh. Analit. Khim., 1962, 17, 206. - , Ibid., 1964, 19, 1468. Jeffery, P. G., and Gregory, G. R. E. C., Analyst, 1965, 90, 177. Lazareva, V. I., and Lazarev, A. I., Zh. Analit. Khim., 1966, 21, 172. Polyak, L. Ya., Ibid., 1963, 18, 966. Received Junztury loth, 1968

ISSN:0003-2654    

DOI:10.1039/AN9689300383

出版商:RSC    

年代:1968

数据来源: RSC

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388 Analyst, June, 1968, Vol. 93, fifi. 388-393 The Use of a Non-absorbing Reference Line in the Simultaneous Determination of Platinum, Rhodium, Palladium and Gold by Atomic-absorption Spectroscopy BY P. B. ZEEMAN AND J. A. BRINK* (The Merensky Institute for Physics, The University of Stellenbosch, Stellenbosch, South Africa) A method for determining platinum, rhodium, palladium and gold simultaneously on a direct-reading spectrometer with a non-absorbing reference line is described. The concentration ranges of the working curves were as follows: platinum, rhodium and palladium 0 to 20 p.p.m. and gold 0 to 5 p.p.m. The detection limit for rhodium and palladium was 0-1 p.p.m., for platinum 0.5 and for gold 0.01 p.p.m. The reproducibility of the results was satisfactory. METHODS for the determination of the noble metals have already been described by Lockyer and Hames,l Strasheim and Wessels,2 Zeeman3 and others.A method is described here in which aqueous solutions of the noble metals were diluted with acetone before being sprayed into the absorbing flame. The advantages of the use of a non-absorbing reference line were previously discussed by Menzies.4 APPARATUS- A multi-element hollow-cathode lamp of the water-cooled type described by Zeeman and Butler6 was used as the light source. The cathode body was made of platinum metal 0-1-mm thick, while strips of palladium and rhodium were soldered with gold to the inside of the platinum cylinder, thus no base metals were introduced into the cathode, and a purer spectrum was obtained.The hollow-cathode lamp was connected to a glass vacuum system so that the carrier gas could be easily changed whenever necessary. A series of experiments, with neon, helium, argon, xenon and krypton as carrier gases, was carried out to investigate the purity of the spectrum in the region of the resonance lines of the noble metals. The results, which agreed with those of Jones and Walsh,6 showed that a neon-filled hollow- cathode lamp had the most satisfactory intensity and stability. The lamp operated under a pressure of 1.6 mm of mercury and 50-mA current, which was supplied by a stabilised d.c. power source. The absorption burner was of the type used by Zeeman and B ~ t l e r , ~ the various inlets of which are shown in Fig. 1. Light from the hollow-cathode lamp, C, is directed over the absorption flame, B, and focused on the entrance slit of the spectrometer, A, as described below.The Zeiss atomiser, H, sprayed the solution into the spray chamber, E, from where it proceeded to the flame. Propane - butane gas, under controlled pressure, is fed in through G. This gas can also be fed in at full pressure through F, to keep the flame burning when the introduction of organic solvent is stopped. The atomising air at 20 p.s.i. is introduced at M, and pre-heated at K to 200" C by a copper spiral inside a cylindrical electrical furnace. Extra air at a pressure of 0.4 kg per cm2 was introduced through L, and fed into the spray chamber through the normal gas inlet of the Zeiss atomiser. Further air was fed into the chamber through the normal condensed solution outlet of the spray chamber via N, because no liquid condenses when organic substances are sprayed as described above.This extra air is essential to obtain a clear soot-free flame when burning organic liquids. The pressure of this extra air is read on the gas-flow manometer, 0, and regulated by the valve, P, which is fed through inlet, Q. The light from the cathode was focused on the entrance slit by means of two quartz lenses. The first, a cylindrical lens of focal length 10 cm, was placed, with its axis horizontal, * Present address : National Physics Research Laboratory, Council for Scientific and Industrial Research, Pretoria, South Africa. 0 SAC and the authors. EXPERIMENTALZEEMAN AND BRINK 389 near the window of the hollow-cathode lamp.The second lens was spherical, with focal length of 30 cm, and was placed about halfway between the source and the slit, which were 120 cm apart. This arrangement just filled the grating with light. An entrance slit width of 30 p was used. The absorption burner was placed about 5 cm from the lamp. As a result part of the flame was focused on the entrance slit. t ‘F Fig. 1. Schematic drawing of the atomic-absorption equipment used (lettered parts of the apparatus are referred to in the text) The spectrometer used was a Bausch and Lomb, type 18000, direct-reading grating monochromator. The grating is in the Paschen mounting with radius 1.5 metres. Normally the instrument is fitted with R.C.A. 1P28 photomultipliers, but these were replaced by E.M.I.tubes, types 9601 B and 9526 B, for higher sensitivity. The power unit for the photomultipliers was modified so that the sensitivity of the five channels used could be varied separately. The instrument is equipped with automatic termination of the integration. The latter is terminated when a fixed voltage is reached on the integrating condenser of a pre- selected line. This results in a constant-intensity reading for one of the spectral lines, and is usually reserved for the internal standard line in emission analysis. In the present investi- gation it was coupled to the non-absorbing reference line. Five photomultipliers were used, one for the resonant line of each of the four noble metals and one for the reference line. PROCEDURE FOR TAKING MEASUREMENTS- In preliminary work photographic records were made with 35-mm film in the focal plane of the spectrometer.The spectra obtained were studied with the view of selecting the best non-resonant noble-metal line to be used as reference for absorption measurements. The platinum line 3042A was selected because it was reasonably intense and free from other interfering lines. A line belonging to one of the noble metals was chosen as reference line to simplify the analytical procedure. The resonance lines used were gold 2428 A, palladium 3404 A, platinum 2659 A and rhodium 3434 A. These lines were also used by Strasheim.2 In ordinary measurements the “100” reading is adjusted as best possible when the solvent is sprayed, and it is then assumed that the lamp’s intensity will stay constant while the standard or sample solution is sprayed afterwards.In the procedure described here, the emission of the hollow-cathode lamp is monitored during the whole period of every measurement by integrating the emission of the non-absorbing line. This applies to the period when the solvent is sprayed, as well as when the solutions are sprayed thereafter. Hence the uncertainty regarding the “100” reading during this latter period disappears. In the method finally adopted, 95 per cent. acetone was aspirated until the pre-set count for the non-absorbing line was reached, when the integration stopped automatically and the readings of the four noble-metal elements could be taken. Solutions containing the noble-metal standards or samples were then sprayed and the counts taken in a similar way.The counts obtained indicate the intensities or transmissions of the various lines. The sensitivity of the reference detector was adjusted so that the integration time was about 20 seconds. During this time 5 ml of solution were nebulised.390 ZEEMAN AND BRINK: NON-ABSORBING REFERENCE LINE IN THE [Analyst, VOI. 93 REPRODUCIBILITY OF THE HOLLOW-CATHODE LAMPS AND DETAILS OF THE ANALYTICAL PRO- The absolute intensities of the lines showed short-term fluctuations and also a slow decrease in intensity with time. This was caused by impurities gathering in the filler gas. When the ratios of the intensities of the different resonant lines to the intensity of the monitor were calculated, an improvement in both sources of error resulted.After a warming-up period of 45 minutes, the decrease in intensity of the resonance lines amounted to 3.5 per cent. every 5 minutes. The intensity ratios, however, decreased only 1 per cent. during 90 minutes. The ratios mentioned above were plotted as ordinates against time, and the curves obtained are shown in Fig. 2. They show that a warming-up time of 10 minutes is CEDURE- 0.61 I. I I I I I . I I I 1 I 0 5 10 I S 20 25 30 35 40 45 50 55 Time, minutes Fig. 2. Time variation of the intensity ratios for the palladium 3404 A. gold 2428 A. platinum 3042 A’ B’ platinum 3042 A’ various elements: A, platinum 2659 A . and D, rhodium 3434 A platinum 3042 A’ platinum 3042 A needed for the intensity ratios of the platinum and rhodium lines to reach stability, and 45 minutes for the palladium and gold.The improvement in the short-term fluctuations are shown in Table I. In agreement with Butler and Strasheim,’ significantly lower co- efficients of variation were obtained with the integrating method. The detailed procedure for calculating results is given here for gold. Let T be the number of counts on gold channel when 95 per cent. acetone is sprayed. Let Tm be the number of counts now observed on the monitor channel. Let T A ~ be the number of counts on the gold channel when the solution is sprayed. Let T’m be the number of counts now observed on the monitor channel. Let T’ be the number of counts that would have been observed on the gold channel if 95 per cent. acetone had been sprayed during this period. = constant. counts for line channel counts for monitor channel It was proved above that T T’ Tm Tm Therefore - = constant =I.T A ~ x 100 percent. T’ The percentage transmission for gold = - - - T ~ ~ ’ T ’ m x 100 per cent. Tm x 100 per cent. (from the above) Tlm T T/Tm But with the automatic stopping of the integration process, T‘m = Tm. Therefore percentage transmission for gold = - x 100 per cent. TAU TJune, 19681 DETERMINATION OF PLATINUM, RHODIUM, PALLADIUM AND GOLD 391 By a similar calculation the percentage transmission can be obtained for the other elements. It must be realised that although the percentage transmission is thus calculated simply by taking a ratio as proved, and the monitor count apparently plays no r61e, it did so in fact, as the periods of integration varied slightly, depending on the lamp operation.TABLE I SENSITIVITY AND REPRODUCIBILITY OF THE DETERMINATIONS Detection limit Resonance (99 per cent. wavelength, transmission), Element A p.p.m. Gold .. .. 2427.950 0.0 1 Platinum . . .. 2659454 0.5 Palladium . . . . 3404.580 0.1 Rhodium . . .. 3434.893 0.1 Coefficient of variation at 50 per cent. transmission Absolute intensities, Relative intensities, per cent. per cent. 4-6 2.3 3.8 1.4 3-1 1.6 5.1 2.4 A r \ STANDARDS- The standards used were prepared from “Specpure” (Johnson, Matthey & Co.) solutions, which were of the ammonium chloro type. The stock solution was obtained by diluting the smallest convenient volume of aqueous solution containing 8000 p.p.m. of noble metal with analytical-reagent grade acetone to give a concentration of 400 p.p.m.of noble metal, which was thus in a 95 per cent. acetone medium. The various standards were obtained by diluting this stock solution with 95 per cent. acetone solution. The concentration ranges used were as follows: platinum, rhodium and palladium 0 to 20 p.p.m., and gold 0 to 5 p.p.m. RESULTS- In the preliminary investigations, the various elements were determined separately. The working curves are given in Fig. 3. The curves for rhodium and palladium are non- linear throughout, that for platinum is linear, while the curve for gold is linear for concen- trations up to 1 p.p.m. and then bends downwards. Repeated determinations of the four elements were made and the detection limits are given in Table I. d o 0 5 20 n.- 540 ul .- 60 m 0 2 4 6 8 10 I2 I4 16 18 20 I,: p.p.m.--, Fig. 3. Working curves obtained when elements were analysed separately Standard solutions were then prepared containing the four elements in solution, and the same amcentration ranges as above were used. Prior to the final measurements, the influence of the various elements on one another was investigated in the normal way by varying the concentration of one element in a constant composition of the others. It was found that there was no interference from palladium and gold, which is in agreement with Strasheim and Wessels’ observations2; however, platinum was suppressed, and rhodium, which was suppressed below 5 p.p.m., was enhanced above 5 p.p.m. The influence mentioned above, however, was not serious as is to be expected at the392 ZEEMAN AND BRINK: NON-ABSORBING REFERENCE LINE I N THE [Artdyst, Vol.93 low concentrations involved. The working curves obtained, with the four elements simul- taneously in solution, do not differ much from those when the elements were treated separately (Fig. 4). With palladium and gold they were the same, while the absorption was suppressed with platinum, and for rhodium it was suppressed below 5 p.p.m. and enhanced for higher concentrations. The sensitivity and reproducibility were the same as those given in Table I. 0 2 4 6 8 10. I2 14. 16 I8 20 p.p.m.- Fig. 4. Working curves obtained when elements were analysed simultaneously The non-linear working curves for rhodium and palladium were caused by the emission of O-H bands in the flame.These bands are degraded to the red. The (0, 1) band has heads at 3428 A and 3432 A. Hence rhodium determinations are strongly influenced by these, and palladium to a lesser degree. DETERMINATION OF THE NOBLE-METAL ORES- The accuracy of the above method was tested by analysing four samples supplied by a gold mining company. The samples were dissolved and the noble metals extracted from the base metals to avoid interference effects that occur, especially when these metals are present in high concentration. The noble-metal content was then determined by the atomic- absorption method described above. The results are given in Table 11, together with the concentrations given by the mine laboratories, and those obtained by emission analysis (horizontal arc) of the same samples carried out in our laboratory.TABLE I1 RESULTS OBTAINED FOR ORE SAMPLES BY VARIOUS METHODS Sample 3A M 8 3F R1 3A M 8 3F R1 3A M S 3F R1 3A M 8 3F R1 Concentration by present method, Element p.p.m. Rhodium 121 42 68 79 Palladium 528 443 429 563 902 82 1 799 782 48 39 44 51 Platinum Gold Concentration given, p.p.m. 96 49 85 83 550 468 489 530 933 805 83 1 7 10 43 48 43 42 Concentration by spectroscopic method, p.p.m. 115 40 72 75 510 450 450 560 880 835 807 770 49 57 39 50June, 19681 DETERMINATION OF PLATINUM, RHODIUM, PALLADIUM AND GOLD 393 CONCLUSION The work described shows that a non-absorbing monitor line can be successfully used in atomic-absorption measurements. The agreement between the values obtained by the different analytical methods in Table I1 is satisfactory, especially as the spectroscopic methods were not then used on a routine basis. It also shows that the integrating method described can be a practical way of determining more than one element simultaneously. We wish to thank the South African Council for Scientific and Industrial Research for the grant without which it would have been impossible to carry out the research. REFERENCES 1. 2. 3. 4. 5. 6. 7. Lockyer. R., and Hames, G. E., Analyst, 1959, 84, 385. Strasheim, A., and Wessels, G., Appl. Spectrosc., 1963, 17, 65. Zeeman, P. B., NaudB, W. J., and van der Westhuysen, 0. A., Tydskr. Natuurwetenskap#e, 1964, Menzies, A. C., Analyt. Chem., 1960, 32, 898. Zeeman, P. B., and Butler, L. R. P., Afipl. Spectrosc., 1962, 16, 120. Jones, W. G., and Walsh, A., Spectrochim. Ada, 1960, 16, 249. Butler, L. P. R., and Strasheim, A., Ibid., 1965, 21, 1207. 4, 202. First received November 19th, 1964 Amended February 8th, 1968

ISSN:0003-2654    

DOI:10.1039/AN9689300388

出版商:RSC    

年代:1968

数据来源: RSC

摘要:

394 Afialyst, June, 1968, Vol. 93, +@. 394-397 The Determination of Metals in Wool by Atomic-absorption Spectroscopy* BY F. R. HARTLEY AND A. S. INGLIS (Division of Protekra Chemistry, CSIRO, Wool Research Laboratories, Parkville (Melbourne), Victoria 3052, Australia) Atomic absorption has been shown to provide a simple and precise method for the determination of chromium, copper, mercury, tin, zinc, calcium, strontium and barium in wool. The method of standard additions is used for calcium and a calibration-graph method for the other metals. Both methods have an accuracy and reproducibility of about f 3 per cent. The wool sample is hydrolysed in constant-boiling hydrochloric acid, and the calibration graphs are prepared by using wool hydrolysates containing standard amounts of metal to compensate for physical and chemical inter- ferences caused by hydrochloric acid, amino-acids and ions present in the wool.THE wide use of metal salts in the treatment of wool has led to a need for rapid and precise methods for the determination of metals in wool. Recently, aluminium in wool has been determined by atomic-absorption spectroscopy.l Because that work showed that the method offered advantages in speed, convenience and specificity over existing methods,2 the present study was undertaken with a view to analysing wool for chromium, copper, mercury, tin, zinc, calcium , strontium and barium by atomic-absorption spectroscopy. In our previous work, the wool sample was hydrolysed by heating it overnight with constant-boiling hydrochloric acid in a sealed tube at 110" C, and the hydrolysate aspirated directly into the flame of the atomic-absorption spectrophotometer.It was possible to com- pensate simultaneously for both physical and chemical interferences by using solutions of wool hydrolysates to prepare calibration graphs. This technique necessarily required the wool to be free from metal ions, and for all of the metals, except calcium, this condition was obtained by repeated extraction with an aqueous 0-5 M EDTA solution. With calcium, although extraction with 0.5 M EDTA solution initially removed a considerable proportion of calcium ions, a small constant amount remained after repeated extraction. The method of standard additions3 was, therefore, used for this metal. APPARATUS- The apparatus described previously1 was used.Because of the relatively large size of the sample (about 0-3g), advantage was rarely taken of the very high sensitivity possible with atomic-absorption spectroscopy and, for all of the metals, except tin and barium, a reduction in sensitivity was necessary. This was achieved either by using a less sensitive line than that normally used or by rotating the b ~ r n e r . ~ This technique was preferred to dilution because it is quicker, although for zinc and strontium a dilution technique was necessary because rotating the burner did not reduce the sensitivity sufficiently, and other resonance lines of suitable sensitivity were not available. The operating conditions, shown in Table I, are not, therefore, necessarily the most sensitive, but those found to give the best results for the samples under test.REAGENTS- All of the reagents used were of analytical-reagent grade. Stock solutions (about 0.1 M) were prepared by standard methods.6 The wool used for the calibration graphs was prepared by shaking scoured wool (30 g) gently for 3 days with 800 ml of EDTA (disodium salt) solution (0.5 M). After rinsing it thoroughly in de-ionised water, the extraction with EDTA solution was repeated once more. The absence of the metal under test was confirmed by wet ashing,2 freeze-drying to concentrate the sample and subsequent analysis. * The work reported here is part of that presented to the 6th Australian Spectroscopy Conference, Brisbane, August, 1967. 0 SAC and the authors. EXPERIMENTALHARTLEY AND INGLIS TABLE I INSTRUMENTAL CONDITIONS FOR THE DETERMINATION OF METALS IN WOOL 395 Lamp current, Element Fuel - oxidant mA Chromium Acetylene - airt 10 Mercury Coal gas - air 4 Tin Hydrogen - air 5 zinc Coal gas - air 8 Copper Coal gas - air 6 Calcium Acetylene - nitrous oxide (10 : 1)s 10 Strontium Acetylene - nitrous oxide (4 : 1)s 10 Barium Acetylene - nitrous oxide (2 : 1) f 10 Slit width, P 100 50 20 50 300 25 100 50 Wave- 4254.3 4289.7 2942.1 2536.5 2354-8 2138.6 4226.7 4607.3 5535.6 Burner length, 5 5 5 10 10 10 10 5 5 5 cm 1 Burner :otation, 40" 10" 0" 0" 10" 0" 0" 0" 0" 0" Approxi- mate sensitivity* 7.5 10.9 5.7 2.9 4.2 $ 0.0 15 0.055 0.20 3-8 46 * Sensitivity is the number of milligrams of element per litre of water necessary to give 1 per f An oxidising flame was used to reduce the background noise observed with the more 5 Ratio of height of red "feather" to height of inner blue cone.cent. absorbance. sensitive reducing flame. Water contained hydrochloric acid (0.01 N) to prevent hydrolysis of tin(I1) chloride. ANALYSIS OF SAMPLES- Calibration-graph method-Prepare the samples and standards by hydrolysis in constant- boiling hydrochloric acid, as described previous1y.l Method of standard additiorcs-Hydrolyse the sample, as before,l and withdraw, by pipette, three 1-ml samples from the hydrolysate. Dilute one of the samples to the volume necessary to give a transmittance of 80 per cent. To the others add different, but accurately known, amounts of calcium ion (about 0.6 and 1.2 p.p.m.) before making up the volume with constant-boiling hydrochloric acid.Aspirate the three samples consecutively into the atomic-absorption spectrophotometer, measure the absorbance values and repeat the readings twice more to eliminate the effect of fluctuations in the flame or instrument conditions. Determine the magnitude of the background absorbance by measuring the absorbance of the nearby 4201 A line that is not absorbed by calcium. The concentration of calcium in the hydrolysate can be calculated6 from the three corrected absorbance readings in two independent ways. RESULTS AND DISCUSSION SAMPLE SIZE- It was found that if the wool samples were too small sampling errors were introduced, probably because the uptake of metal ions was not completely uniform among the fibres. TABLE I1 REPRODUCIBILITIES AND CONCENTRATION RANGES OF THE ANALYSES FOR METAL IONS IN WOOL Metal Chromium Copper .. Mercury . . Tin . . Zinc . . Calcium . . Strontium Barium . . Reproducibility, per cent. .. & 3.4 .. f 2.3 .. & 3-0 .. f 3.5 . . f 2.4 .. f 3.5 .. f 3-0 .. f 3.0 Typical concentration range, * mg of metal per g of dry wool 0.7, to 29 0.4 to 8 6 to 120 0.56 to 11 0.2 to 4$ 0-3, to 71 0.3 to 61 0.5 to 10 Minimum possible concentration, t 0.03 0.09 0-4 0.002 0.06 0-04 0.5 mg of metal per g of dry wool 0.56 * This concentration range was obtained by using the instrumental conditions given in Table I. A higher concentration of metal in wool than the maximum given here can be accom- modated, by rotating the burner more, by using a less sensitive resonance line or by dilution.The values given were obtained by diluting 1 part to 100 parts (for zinc), to 50 parts (for calcium) and to 10 parts (for strontium). Obtained by using conditions for maximum sensitivity. $ The range depended on the dilution used.396 HARTLEY AND INGLIS: DETERMINATION OF METALS [ArtdySt, VOl. 93 Thus samples of about 0.1 g gave reproducibilities of about &8 per cent., whereas samples of about 0.3 g gave reproducibilities of about +3 per cent. (see Table 11). Larger samples did not give better reproducibility. PRESENCE OF SOLID MATTER- The method used to prepare the samples gave dark brown solutions containing finely suspended matter that arose largely from decomposition of the amino-acid, tryptophan. However, neither the average absorbance reading nor the meter fluctuation at this reading was altered after filtration through a Millipore filter (0.20 p).ACCURACY AND REPRODUCIBILITY- Wool was analysed independently for calcium, chromium, copper and zinc. The results, shown in Table 111, indicated that the agreement between atomic-absorption spectroscopy and the independent method was good. The reproducibilities of the analyses, determined by analysing the same sample of wool six times, were generally about +3 per cent. (Table 11). The concentration ranges of metal in wool for which the method is applicable are shown in Table 11. TABLE I11 COMPARISON OF ATOMIC-ABSORPTION AND ALTERNATIVE METHODS OF ANALYSIS Metal content, mg per g of dry wool Independent Atomic Metal Independent method of analysis method absorption Calcium .. Nitric acid extraction; EDTA titration 0.068 0.069 Copper . . Hydrochloric acid extraction; EDTA titration 8.0 8-1 Chromium . . Wet ashing; iron(I1) titration of chromium(V1) 9.8 10.0 Zinc . . . . Wet ashing; EDTA titration 5.5 5.5 EFFECTS OF DIFFERENT MEDIA- As in the deterrnination of aluminium in wool,l it was found that the absorbance values in water, constant-boiling hydrochloric acid and wool hydrolysate solutions were in each instance different. This was shown to be largely because of the different surface tensions and viscosities of the solutions, which emphasised the need for samples and standards to be prepared in identical media. This was achieved automatically with the method of standard additions, and with the calibration-graph method by preparing the standard solutions in wool hydrolysates. VALIDITY OF THE METHOD OF STANDARD ADDITIONS- The method of standard additions, as normally used, requires a linear relationship between absorbance and concentration.Although this relationship was not linear for calcium in water when a nitrous oxide - acetylene flame was used, it was linear in wool hydrolysates in the concentration range 0 to 8 p.p.m. of calcium. Willis4 has pointed out that the method of standard additions is based on the assumption that the interfering substance alters the absorbance of the added metal to the same extent as it does that of the original sample. This may not always be so, particularly when only a small amount of interfering substance is present, but this condition was established in the present work by making two separate additions to each sample, thus providing a check on the linearity of the analysis in the presence of the interfering material.Although errors caused by instrumental drift are minimised, by measuring in quick succession solutions that are effectively sample and standard, the method of standard additions involves an extrapola- tion, which makes it inherently less accurate than the calibration-graph method, which involves an interpolation. It is, therefore, essential to correct for the background absorbance, which is caused either by absorbance by the flame or scattering of the light by solid particles in the flame, by measuring the absorbance at a nearby wavelength that is known not to be absorbed by the metal ion under examination.Modulated atomic-absorption apparatus should be used to avoid measuring flame emission.June, 19681 IN WOOL BY ATOMIC-ABSORPTION SPECTROSCOPY 397 SCOPE OF THE METHOD- The results obtained confirm the conclusions drawn from the earlier work1 that metals in wool can be readily determined by atomic-absorption spectroscopy. The method should be equally suitable for other insoluble protein materials, such as hair and hide. The advantage of being able to use a hydrolysis technique to decompose the protein material, rather than the more lengthy ashing or extraction techniques will still apply. This procedure has the additional advantage, as far as the biochemist is concerned, that the hydrolysate could also be used for other determinations, such as those involved in an amino-acid analysis. The authors thank Mr. B. J. Wallace for assistance with the experimental work. REFERENCES 1 . 2. 3. Hartley, F. R., and Inglis, A. S., Analyst, 1967, 92, 622. “Methods of Test for Textiles,” British Standards Handbook, No. 11, 1963. Elwell, W. T., and Gidley, J. A. F., “ Atomic-Absorption Spectrophotometry,” Second (Revised) Edition, Pergamon Press, Oxford, London, Edinburgh, New York, Toronto, Sydney, Paris and Braunschweig, 1966, p. 74. Willis, J. B., Meth. Biochem. Analysis, 1963, 11, 1. Vogel, A. I., “A Textbook of Quantitative Inorganic Analysis including Elementary Instrumental Analysis,” Third Edition, Longmans, Green and Co. Ltd., London, 1961. David, D. J., Analyst. 1962, 87, 576. Received January 29th, 1968 4. 5. 6.

ISSN:0003-2654    

DOI:10.1039/AN9689300394

出版商:RSC    

年代:1968

数据来源: RSC

摘要:

398 Analyst, June, 1968, Vol. 93, pp. 398-399 The Rapid Determination of Cross-linking Agents in Dental Monomer BY J. F. DE FREITAS (Commonwealth Bureau of Dental Standards, Melbourne, A ustralza) A rapid method, involving the use of infrared spectrophotometry, is described for the identification and determination of the usual cross-linking agents that are mixed with methyl methacrylate in dental monomers. At selected wavelengths of maximum absorption, the absorbance of the cross- linking agent is proportional to its concentration in the mixture. By com- paring the absorbances of the unknown, and a mixture prepared with methyl methacrylate and the particular cross-linking agent, each diluted with carbon disulphide, the concentration of the cross-linking agent in the former can be readily calculated.THE acrylic monomer used with powdered polymer in the preparation of teeth, denture-base materials and various prostheses is nearly always basically methyl methacrylate, but is some- times blended with up to about 10 per cent. of a cross-linking agent to improve strength, hard- ness and resistance to solvents. On the other hand, too much can adversely affect bonding as, for example, between teeth and the denture base or when repairing dentures. Generally, the cross-linking material is allyl methacrylate or ethylene glycol dimethacrylate.1 These can be determined in the presence of methyl methacrylate by fractional distillation, bromina- tion2 or gas - liquid chr~matography.~ The first of these techniques is tedious and requires precautions against polymerisation and a relatively large sample ; the second needs prior knowledge of identity so that calculation, based on constants, can provide quantitative results; the third is a good approach, but if the special equipment is not in regular use the setting-up period is time consuming.The infrared spectra of the three monomers mentioned show similar absorption patterns. However, several areas of minor but characteristic absorption, mainly between 1300 and 800 cm-l, were found, for example, allyl methacrylate at 985, 1010 and 935cm-l and ethylene glycol dimethacrylate at 1045, 1245 and 885 cm-1. These allow identification of the respective agents and form the basis of the proposed method. METHOD APPARATUS- A Hitachi infrared grating spectrophotometer, model EP I-G2, together with a de- mountable cell with potassium bromide windows and containing a Teflon spacer (0.08 mm) were used for absorbance measurements.REAGENTS- was used as methyl methacrylate monomer. Kallodent denture acrylic liquid-The type without cross-linking agent (1.C.I.A.N Z.) Allyl methacrylate-Obtained from Sartomer Resins Inc., U.S.A. Ethylene glycol dimethacrylate-Obtained from Sartoiner Resins Inc., U.S.A. Carbon disulphide, analytical-reagent grade. TEST FOR THE OBSERVANCE OF BEER’S LAW- Each of the two cross-linking monomers was mixed with methyl methacrylate to make 0, 5, 10, 20 and 25 per cent. v/v solutions. One millilitre of each of these standards was diluted to 25 ml with carbon disulphide. With the aid of wavenumber and transmittance- scale expansion (both x 5), with air only in the reference beam, infrared spectra were recorded 0 SAC and the author.DE FREITAS 399 over the following ranges: allyl methacrylate solutions 1000 to 900 cm-l, maxima 985 cm-l; and ethylene glycol dimethacrylate solutions, 1100 to 1000 cm-l, maxima 1045 cm-1.By using the absorption maxima and base-line technique, the absorbances of the various concentrations of cross-linking agents were measured. The results are shown in Table I and confirm a linear relationship, of the type y = mx, between concentration in admixture and absorbance for each agent. TABLE I INFRARED ABSORPTION OF CROSS-LINKING AGENTS IN ADMIXTURE WITH METHYL METHACRYLATE Cross-linking agent, per cent. v/v 0 5 10 15 20 25 Methyl methacrylate, per cent.v/v 100 95 90 85 80 75 Absorbance P Ally1 Ethylene glycol methacrylate dimethacrylate 0 0 0.025 0.010 0.05 1 0.020 0.075 0.029 0.102 0.039 0.126 0.049 PROCEDURE- Identify the liquid as basically methyl methacrylate and the cross-linking agent present by examination of the infrared spectra, with particular reference to characteristic absorption peaks in the 1300 to 800cm-l region. Dilute 1 to 2 ml of the liquid to 25 ml with carbon disulphide. In the same way, dilute an equal amount of an accurately prepared blend of about the same concentration of the cross- linking agent in methyl methacrylate. Concomitantly determine the absorbances of both solutions in approximately 0-l-mm cells at the wavelength of maximum absorption: allyl methacrylate at about 985 cm-l and ethylene glycol dimethacrylate at about 1045 cm-l.Calcu- late the amount of cross-linking monomer by the formula C 2 where C is the percentage of cross-linking monomer in the standard, and A , and A , are the absorbances of the unknown and the standard, respectively, measured by the base-line technique. DISCUSSION In the concentrations normally present, inhibitors do not affect the determinations. Because of interfering absorption , some modern prosthetic liquids of the quick-curing or self-curing type containing modifiers may make it necessary to select alternative wavenumbers for the measurement of absorbances, but mixtures tested were found suitable for this method. A A8 ’ REFERENCES 1. 2. 3. Skinner, E. W., and Phillips, R. W., “The Science of Dental Materials,” Sixth Edition, W. B. Saunders Co., Philadelphia and London, 1967, p. 180. “Analytical Methods for the Acrylic Monomers,” Rohm and Haas Company, Special Products Department, Philadelphia, U.S.A., 1961. Koppang, R., Odont. Tidskr., 1967, 75, 253. Received November 24th, 1967

ISSN:0003-2654    

DOI:10.1039/AN9689300398

出版商:RSC    

年代:1968

数据来源: RSC