ISSN:0003-2654    

DOI:10.1039/AN96186FX037

出版商:RSC    

年代:1961

数据来源: RSC

ISSN:0003-2654    

DOI:10.1039/AN96186BX039

出版商:RSC    

年代:1961

数据来源: RSC

ISSN:0003-2654    

DOI:10.1039/AN96186FP181

出版商:RSC    

年代:1961

数据来源: RSC

ISSN:0003-2654    

DOI:10.1039/AN96186BP191

出版商:RSC    

年代:1961

数据来源: RSC

摘要:

SEPTEMBER, 1961 THE ANALYST Vol. 86, No. 1026 Analytical Methods Committee REPORT PREPARED BY THE MEAT PRODUCTS SUB-COMMITTEE Nitrogen Factors for Pork THE Analytical Methods Committee has received the following Report from its Meat Products Sub-committee. The Report has been approved by the Analytical Methods Committee and its publication has been authorised by the Council. REPORT The Meat Products Sub-committee was appointed by the Analytical Methods Committee in 1955 to succeed the Meat Extracts Sub-committee, which had been in existence since 1948 under the chairmanship of Mr. G. Taylor. It was reconstituted with wider terms of reference than its predecessor, and its new title indicated the widening of the scope of its work. Its first Chairman was Dr. H. G. Rees, who had been Honorary Secretary of the old Sub-committee, but, on his resignation in 1956, he was succeeded by Dr.S. M. Herschdoerfer; its constitution is as follows: Mr. S. Back, Mr. P. 0. Dennis, Mr. J. R. Fraser, Dr. R. A. Lawrie, Dr. A. McM. Taylor and Mr. E. F. Williams (deputy, Mr. H. C. Hornsey), with Dr. C. H. Tinker as Secretary. The following have also served on the Sub-committee: Miss E. M. Chatt (October, 1955, to June, 1960), Mr. C. D. Essex (June, 1955, to January, 1960) and Dr. H. Amphlett-Williams (June, 1955, to January, 1961). The Terms of Reference are- “(a) The determination of the meat content of products containing meat; (b) the determination of the constituents of meat and meat products. NOTE-The term ‘meat products’ to include hydrolysed protein and, if found necessary, fish In the first instance, the Sub-committee reviewed the alternative methods for the determination of the meat content of manufactured products-namely, that of Stubbs and More,l in which the nitrogen content is determined, and that of Osman Jones, based on the determination of starch content.The Sub-committee considers Stubbs and More’s method to be more reliable and suggests that, as a further check, a determination of starch by the method of Fraser and Holmes2 may be used. There was evidence that the nitrogen factors published by Stubbs and More in 1919 for the different types of meat (pork, beef, mutton, etc.), or even the modified factor of 3-6 for pork (recommended by the Analytical Methods Commit- tee in 1940),s were not altogether valid for present-day use.The Sub-committee therefore reviewed all data relating to the determination of nitrogen factors published over the last 30 years. Jackson and Jones: on the basis of 21 analyses of lean pork, including 10 of “mixed meat,” derived an unweighted mean for the water-to-protein ratio of 3.4 to 1, corresponding, on the basis of a nominal ash content of 1 per cent., to 3.60 per cent. of nitrogen in the fat-free meat. Steiner,5 quoting results from a private communication, states that ‘‘ . . . an examina- tion of the analytical data for various cuts of pork has shown that the protein percentage 557 pastes.”558 ANALYTICAL METHODS COMMITTEE : [Vol. 86 in the fat-free portion of the meat is distributed approximately normally, with mean value 21.9 per cent.” (equivalent to 3.50 per cent.of nitrogen) “and standard deviation of 2.76 per cent.” Reith, Hofsteede and Langbroek‘j examined the meat from 19 Dutch animals, all meat muscle being “cut out, divided into 50-g pieces and mixed in the same quantitative relation as exists in the animal.” The average nitrogen factor from these analyses was 3.40. In analyses by Marshall,’ the nitrogen content (on a fat-free basis) of the whole of the lean meat from one side of pork ‘ I . . . calculated by proportion was 3.39 per cent.; a higher figure of 3.43 per cent., which is less accurate, is obtained by direct averaging of the analyses.” In his second series of analyses, for which another carcass was used, “. . . the nitrogen content of the lean meat on a fat-free basis, calculated both by proportion and by direct averaging, was 3.43 per cent.” A circulars issued by the American Meat Institute Foundation quotes a large number of figures; these, however, refer partly to analyses reported in 1928 and include many types of meat not used in Britain for manufactured products.EXPERIMENTAL AND RESULTS In view of the paucity of published data, the Sub-committee appealed to various meat product manufacturers and meat research organisations in this country and abroad for information on the analyses of meat products, with particular reference to the nitrogen contents of pig carcasses and different types of cuts. Nearly 1200 results, ranging from 3-0 to 4.0 per cent. and including values for individual muscles, named cuts and unspecified mixtures of pork meat, were submitted to the Sub-C~rrimittee.~ These results, together with corre- sponding values from the literature, are shown in Fig.1, from which it is apparent that there are considerable differences between the nitrogen contents of the fat-free meat in the different cuts of the animal. The Sub-committee considered the possibility of calculating the value for a nitrogen factor for the whole carcass from these figures, but thought this to be too dependent on arbitrary assumptions. Apart from the uncertainty associated with such assumptions, a further limitation arises from the fact that most of the analytical results were obtained from small samples of the relatively large cuts of meat involved, thereby introducing the possibility of sampling errors.In view of these difficulties, two particular series of tests were carried out, at the request of the Sub-committee, in which the entire cuts of meat were comminuted and thoroughly mixed before sampling. These tests were planned to give an automatic weighting of the analytical figures in relation to the proportionate amounts of the cuts in the carcass. In each instance, one side was completely boned out, very finely comminuted, mixed and used as sample for determining the average nitrogen cjontent of all the edible meat; the other side was divided into shoulder, middle and leg cuts, each of which was boned out, comminuted and analysed. Gristle, fat, rinds, ears, snouts, tails, gullets, tongues and offal were excluded, so that only the carcass meat-the so-called “usable meat”-was employed for the analytical work.A single sample from each mix was taken and analysed in duplicate, each observation recorded being the arithmetical average of these two analyses. The results from the analyses of 18 carcasses are shown in Table I. In the first series of tests, 18 carcasses were divided into left and right sides. Each sample was analysed for fat, nitrogen and water contents. TABLE I NITROGEN CONTENTS FOUND I N PORK MEAT FROM 18 CARCASSES cut Average nitrogen Standard deviation Coefficient content (fat-free basis), of a single observation, of variation, Side A - Shoulder .. .. . . Middle . . .. .. .. Leg . . .. .. .. % 3.37 3.66 3-52 Side B- All edible meat from whole side 3-45 % 0.08 0.07 0.06 % 2.3 1.9 1.7 0.075 2.2 In the second series of tests, in order to determine the average nitrogen content of all edible meat, i.e., lean and fatty tissue, but not offal, one side from each of a further 35 carcasses was completely boned out, and the meat was very finely comminuted and mixed.Two samples of each lot, i.e., each carcass, were taken, and a single analysis was made on each sample, eachSeptember, 19611 NITROGEN FACTORS FOR PORK 559 0 320 3 D Live weight, Ib Fig. 2. Relationship between nitrogen content of fat-free meat and live weight of animal: 0 , results found by laboratory F; x, results found by laboratory E; 0, average results (see Table 11) observation recorded being the arithmetical average of these two determinations. The over- all average nitrogen content (fat-free basis), for the 53 carcasses was 3.47 per cent., with a standard deviation of individual observations of 0.12 per cent.The series of results forming the basis of this assessment was obtained from a number of pigs ranging in weight from 147 to 320 lb. A significant regression of the nitrogen content (fat-free basis) on live weight was apparent (see Fig. 2). Another laboratory, examining pigs of live weight 180 to 200 lb, found an average value of 3.36 for 6 carcasses (whole meat) as the nitrogen factor. These results are also shown in Fig. 2 and are seen to be on the same regression line. The relationship between the nitrogen factor and the live weight is summarised in Table 11. TABLE I1 RELATIONSHIP BETWEEN NITROGEN FACTOR AND LIVE WEIGHT These results are for the total edible meat from 59 citrcasses Live weight of animal, Nitrogen content (fat-free basis), lb % 140 to 159 160 to 179 180 to 199 200 to 219 220 to 239 240 to 259 260 to 279 280 to 299 300 to 320 3-26 3.32 3.37 3-43 3-47 3.50 3.52 3-55 3-65 RECOMMENDATION After due consideration of the types of meat normally used in the manufacture of comminuted products, and of the use of pigs of varying live weight, the Sub-committee recommends an average nitrogen factor of 3.45 as the best compromise for general use.560 1.2. 3. 4. 5. 6. 7. 8. 9. ANALYTICAL METHODS COMMITTEE REFERENCES [Vol. 86 Stubbs, G., and More, A., Analyst, 1919, 44, 125. Fraser, J . R., and Holmes, D. C., Ibid., 1958, 83, 371. Analytical Methods Committee, Ibid., 1940, 65, 257. Jackson, F. W., and Jones, O., Ibid., 1932, 57, 562. Steiner, E. H., Ibid., 1948, 73, 15. Reith, J. F., Hofsteede, M. J. N., and Langbroek, W., J . Sci. Food Agric., 1955, 6, 317. Marshall, C. R., Analyst, 1955, 80, 776. Maroney, J. E., and Landmann, W. A., “Moisture and Fat in Fresh Meat Materials as Related to Sausage Formulations,” Circular No. 49, American Meat Institute Foundation, Chicago, 1959. Personal communications from- Danish Meat Research Institute, Roskilde, Denmark. N.V.H. Hartog’s Fabrieken, Oss, The Netherlands. J. Sainsbury Ltd. T. Wall & Sons Ltd. Federal Meat Industry Research Institute, Kulmbach, Germany. Low Temperature Research Station, Cambridge.

ISSN:0003-2654    

DOI:10.1039/AN9618600557

出版商:RSC    

年代:1961

数据来源: RSC

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560 ANALYTICAL METHODS COMMITTEE [Vol. 86 Analytical Methods Committee REPORT PREPARED BY THE ME.AT PRODUCTS SUB-COMMITTEE Nitrogen Content of Rusk Filler THE Analytical Methods Committee has received the following Report from its Meat Products Sub-committee. The Report has been approvled by the Analytical Methods Committee and its publication has been authorised by the Conncil. REPORT The Sub-committee responsible for the preparation of the Report was constituted as follows: Dr. S. M. Herschdoerfer (Chairman), Mr. S. Back, Mr. P. 0. Dennis, Mr. J. R. Fraser, Dr. R. A. Lawrie, Dr. A. McM. Taylor and Mr. E. F. Williams (deputy, Mr. H. C. Hornsey), with Dr. C. H. Tinker as Secretary. Although various types of cereal filler, e.g., rusk, bread and rice, can be used in the manufacture of sausages and some other meat products, the general practice nowadays is to use rusk, and this Report deals only with the correction to be made for nitrogen in this type of filler.No correction for nitrogen is made when the filler consists of potato starch* or corn flour (ie.? maize starch). In 1952, the Analytical Methods Committee1 recommended that, in calculating the results of analyses, a figure corresponding to 13 per cent. of the dry carbohydrate plus the crude cellulose should be deducted from the result for total nitrogen to allow for nitrogen present in the cereal filler. However, in view of the wide range of nitrogen contents of flours, this figure of 2 per cent. would appear to correspond to rusk flour of the lowest protein content. Values for analyses of rusks recently submitted by rusk manufacturers indicate nitrogen contents ranging from 1.9 to 2.6 per cent.(calculated on dry carbohydrate). In this event, the Sub-committee recommends as the best compromise a figure of 2.3 per cent. as the correction for nitrogen in the rusk: filler; this figure corresponds to the average composition of culinary flour. The Sub-committee also recomends that this figure should be reviewed periodically, in view of changes that occur in milling practices. REFERENCE 1. Analytical Methods Committee, Analyst, 1952, 77, 643. * This is known in the trade as farina and should not be confused with potato flour.Nitrogen content, yo 3.2 3.4 3.6 3.8 40 42 I 1 I I I I I 1 1 I 1 1 2 Analyses b a d on samples - Hog Hog Ham trimmings, lean Sow Gammon I Head Meat Laboratory A Laboratory D Laboratory D Laboratory F Laboratory A Laboratory A 21 Cheeks SOW Hog I Cheek meat Laboratory A Laboratory D Laboratory D Butt (neck) Shoulder Jackson and Jones Marshall Laboratory D Laboratory D Laboratory F Laboratory F Laboratory F Laboratory F Laboratory A Laboratory E Laboratory B Laboratory C Laboratory C --+---- sow sow Hog E: Unspecified Bladebone Forequarter Female Subscapularis muscle Extensor carpi radialis muscle Lateral head triceps muscle I Laboratory F Laboratory F Laboratory F Laboratory F Laboratory F Laboratory F Laboratory F Laboratory F Laboratory F Laboratory F Marshall Laboratory A Laboratory E Laboratory E Laboratory E Laboratory E Laboratory E Laboratory E Laboratory F Jackson and Jones Marshall Laboratory B Laboratory B Marshall Laboratory A Laboratory D Laboratory D Laboratory D Laboratory C Laboratory C Middle Neck end Sow 8: Centre cut Sow‘ Gilt Leg end Sow Gilt Hog Hog Unspecified i 9 i I i7j I isi I i9i I i 9 i I Loin lean Fore Male Middle Male Hind Male Female Female Female Back “Kotelett, normal” “Kotelett.lean” Belly Diaphragm muscle tissue Sow !:fpecified Longissimus dorsi muscle (thoracic) Longissimus dorsi muscle (lumbar) (3) I I Leg sow Marshall Laboratory F Laboratory F Laboratory F Laboratory F Laboratory A Laboratory D Laboratory D Laboratory D Laboratory D Jackson and Jones Laboratory E Laboratory A Laboratory B Laboratory C Laboratory B Laboratory B Laboratory C Laboratory C Laboratory C Laboratory C n I I 1 I I I I ~ I I I Hog Gilt Unspecified I sow Hindquarter Female Psoas major muscle Adductor muscle Semi membranosus muscle Deep digital flexor muscle Superficial digital flexor muscle dectus femoris muscle Sartorius muscle Mixed Meat Trimmings from cutting room Pork. Unspecified Muscle of unknown origin Fat Jackson and Jones Laboratory A Laboratory A Laboratory B Laboratory A Analyses of whole comminuted cuts - Shoulder Laboratory F Middle Laboratory F Leg Laboratory F Whole sides. 180 to 200 Ib. live weight Laboratory E Laboratory F 140 to 320 Ib. live weight I I I I I I I 1 - I I 1 I I 1 1 I 1 I 1 3.2 3.4 3.6 3.8 4.0 42 Nitrogen content, yo Fig. 1. Nitrogen contents of various cuts of meat. Horizontal lines represent the range of .nitrogen contents, short vertical lines indicate the average values

ISSN:0003-2654    

DOI:10.1039/AN9618600560

出版商:RSC    

年代:1961

数据来源: RSC

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September, 19611 HARRISON, PALFRAMAN AND ROSE 561 Detection, Determination and Identification of Furfuraldehyde in Hydrocarbon Oil BY R. B. HARRISON, J. F. PALFRAMAN AND B. A. ROSE (Department of Scientific and Industrial Research, Laboratory of the Governmtwt Chemist, Clement’s Inn Passage, Strand, London, W.C.2) A colour test for detecting furfuraldehyde in hydrocarbon oil, a quanti- tative spectrophotometric method for determining from 1 to 10 p.p.m. of furfuraldehyde in hydrocarbon oil (by formation of the addition compound with aniline acetate) and a gas-chromatographic identification of furfuraldehyde after extraction from hydrocarbon oil as the aldehyde - bisulphite compound are described. IN accordance with Statutory Instrument No. 1127, 1948, made under the Motor Spirit (Regulation) Act, 1948, chemical markers were added to petrol intended for commercial use.The markers included a red dye so that visual inspection of the petrol was sufficient to show for which class of road users it was intended. Petrol in the tank of a vehicle could be sampled by the police, and a colorimetric test performed at the roadside for one of the markers; if this test was positive, a sample was taken for further tests in the laboratory to establish the presence of “red” petrol. Quantitative work was not necessary, and although some was carried out no reports on it were published. The same problem is now posed by “diesel oil” and “gas oil,” which are often identical in composition, but have different uses. Diesel oil, or DERV, is used as a fuel for diesel-engined road vehicles and, as such, attracts duty as a road fuel; gas oil is used in gas manufacture and as a fuel for stationary diesel-engined machinery and therefore attracts no road-fuel duty.It has been decided to mark oils of this type in order to prevent their misuse as road fuel. The markers had to be selected with care, and five requirements had to be satisfied- (;) A rapid and simple colorimetric test, with the minimum of apparatus, for use at (ii) An accurate quantitative method of determination for laboratory use. (iii) Positive identification of the added chemical. (iv) Each marker added must not interfere with the detection, determination and identification of any other added marker. (v) The marker must have sufficient solubility in a hydrocarbon solvent to permit concentrates to be prepared in volumes convenient to mark several thousands of gallons of oil at a time.Furfuraldehyde fulfilled these conditions and was chosen as one of the markers. It should be mentioned here that furfuraldehyde is used in the refining of heavy oils, but the recovery processes used by the oil companies are highly efficient. It is in the manufacturers’ interest to remove traces of furfuraldehyde, as these cause discoloration due to oxidation. In more than twenty samples of gas and diesel oils obtained from various different companies and depots throughout the United Kingdom, furfuraldehyde was found in only one, and then at the level of 0.1 p.p.m. the roadside. DETECTION OF FURFURALDEHYDE An ideal roadside test is one in which an impregnated paper is used, thereby obviating the necessity for glassware and solutions. Aniline acetate was found to be the most sensitive c o h r reagent for furfuraldehyde, but was unsuitable for impregnating paper, owing to its instability in the absence of excess of acetic acid.However, if spots of the aniline acetate reagent solution were placed on the strip of filter-paper just before immersion for a few seconds in an oil containing furfuraldehyde, a red colour was produced; this colour reached maximum intensity in about 15 seconds and then faded. By this method, 2.5 p.p.m. of562 HARRISON, PALFRAMAN AND ROSE : DETECTION, DETERMINATION AND [VOl. 86 furfuraldehyde were just detectable in a pale oil and 10 p.p.m. in a dark oil. As the required limit.was 1 p.p.m. in pale or dark oils, this technique was abandoned in favour of that described below. METHOD REAGENT- of glacial acetic acid. PROCEDURE- Place 1 ml of aniline acetate solution and. 15 ml of the suspect gas oil in a test-tube (6 inches x 0.75 inch), invert the tube three times, allow the layers to separate, and observe the colour of the lower layer. A bright red colour indicates the presence of furfuraldehyde; a negative test results in a yellow-brown lower layer. With dark-coloured oils, the result is sometimes easier to see if the tube is emptied, when some of the acetic acid phase adheres to the glass, leaving a red film if furfuraldehyde is present. With this test, concentrations of furfuraldehyde down to 1 p.p.m. can be detected in pale or dark oils, equivalent to a dilution of one part of marked oil with nine parts of unmarked oil.The lower limit in a pale oil is 0.2 p.p.m., which results in a pink colour ratiher than bright red. The colours produced persist for several minutes. Aniline acetate solzction-Dissolve 100 ml of analytical-reagent grade aniline in 900 ml Store the solution in a dark-coloured bottle. DETERMINATION OF :FURFURALDEHYDE Several methods for determining furfuraldehyde in oil have been described,l t o 5 and we have modified that of Milner and Liedemad to give a rapid method having the required accuracy and involving use of the minimum amount of sample. The method proposed is designed for the range 1 to 10 p,p.m. of furfuraldehyde, but can be extended to determine higher concentrations by dilution of the oil containing the furfuraldehyde.Difficulty was experienced in extracting the furfuraldehyde completely from the oil, yet maintaining the volume of solution sufficiently small for the production of a colour having an intensity suitable for spectrophotome tric determination. It was therefore decided to measure the colour of the Schiff’s base produced in the reversible reaction between fur- furaldehyde and aniline acetate in sitw in the oil. As aniline acetate in glacial acetic acid is immiscible with gas oil, a diluent was needed, and toluene was used instead of benzene on account of the toxicity of benzene vapour. When the diluted gas oil was mixed with the aniline acetate solution, a red colour developed, increas- ing to a maximum and then fading; this process was followed with a spectrophotometer.The maximum optical density attained was a function of the concentration of furfuraldehyde in the gas oil, and the time within which the maximum was reached was also dependent on the concentration of furfuraldehyde, being about 2 to 3 minutes for concentrations of 1 to 10 p.p.m. and 1 to 2 minutes for concentrations below 1 p.p.m. METHOD REAGENTS- Standard furfuraldehyde solution-From a stock solution containing 0.5 per cent. w/v of freshly distilled furfuraldehyde in gas oil prepare a solution containing 10 p.p.m. of fur- f uraldehyde. Aniline acetate solution-Prepare as described above. PROCEDURE- By pipette, place 5.0 ml of gas oil in a 25-ml calibrated flask, and add 7 to 8 ml of toluene.(Larger aliquots of gas oil cannot be used, owing to immiscibility with the reagent.) From a pipette having an automatic filler add 10.0 ml of aniline acetate solution, and dilute to the mark with toluene. Invert the flask, shake to mix the contents, and rapidly transfer a few millilitres to a parallel-sided 1-cm cell of a Unicam SP600 spectrophotometer or similar instrument. Set the wavelength scale at 520mp, follow the optical density until it attains the maximum, and record this figure. Plot a graph of optical density against concentration of furfuraldehyde from measurements made on pairs of aliquots, cine being treated with reagent and the other diluted with toluene as blank. Use as blank solution 5-0 ml of th.e same gas oil plus 20.0 ml of toluene.September, 19611 IDENTIFICATION OF FURFURALDEHYDE IN HYDROCARBON OIL 563 DISCUSSION OF THE METHOD The anilii,e acetate solution exhibits slight absorption a t 520 mp, the concentration used in the procedure described above giving, when fresh, an optical-density reading of 0.004 (equivalent to 0.07 p.p.m.of furfuraldehyde). This reagent darkens to a yellow colour within 24 hours and must therefore be freshly prepared each day, so that, when the blank value just mentioned is used, the correction is within the limits required. The use of the same amount of gas oil in the blank automatically compensates for any colour in the oil. LIMITS- If furfuraldehyde is dissolved in toluene instead of gas oil and toluene is used as blank, the graph of optical density against concentration of furfuraldehyde in the range 1 to 10 p.p.m.is linear and passes close to the origin. With a solution of furfuraldehyde in a gas oil and an equivalent amount of the same gas oil in the blank, a similar plot gives a graph following the above within 0.1 p.p.m. up to 6 p.p.m. of furfuraldehyde and then deviating from it, the maximum deviation being 0.7 p.p.m. at the 10 p.p.m. level. This deviation is probably caused by the difference in the effects of gas oil and toluene on the equilibrium of the reaction, the gas oil appearing to inhibit the decomposition of the Schiff’s base. To attain an accuracy within 0.1 p.p.m., therefore, the range 1 to 6 p.p.m. of furfuraldehyde should be used; dilution of the gas oil with an equal volume of toluene will bring any result between 6 and 10 p.p.m.within this range. Repeatability is within 0.1 p.p.m. in the range 1 to 10 p.p.m. of fur- furaldehyde. STABILITY OF FURFURALDEHYDE IN OIL- Twelve gas oils having different colours and compositions from various sources were “dosed” to the level of 10 p.p.m, of furfuraldehyde; they were analysed immediately and at fortnightly intervals for 3& months to check the stability of the marker in the oils at this concentration. Some of the results are shown in Table I, from which it can be seen that the concentration of furfuraldehyde does not vary significantly over 3 months, by which time it is estimated that any particular batch of oil leaving a refinery will have been used. The results also show the effects of the gas oils on the equilibrium, the apparent initial concentra- tions of furfuraldehyde being in the range 9.3 to 10.1 p.p.m.It must be emphasised that these results are outside the range of greatest accuracy and were intended only to check the stability of the marker at this concentration. TABLE I VARIATION IN CONCENTRATION OF FURFURALDEHYDE WITH TIME Oil No. 1 2 3 4 5 6 7 8 9 10 11 12 Concentration of furfuraldehyde found- initially, after 14 days, after 38 months, A f \ p,p.m. p.p.m. p.p.m. 10.0 10.2 10.8 9.5 10.1 10.6 9.6 9.9 10.1 9.3 10.1 9.8 9-7 10.2 10.4 10.1 10.0 9.8 10.1 10.6 10.8 10.1 10.1 10.1 10.1 10.2 10.4 9.3 9.3 9.5 10.1 10.2 10.5 9.9 10.0 10.1 INTERFERENCE- Milner and Liedermanl stated that the reaction was specific for the furfuraldehyde structure ; methylfurfuraldehyde and hydroxymethylfurfuraldehyde reacted, but absorbed only slightly at 520 mp.These workers found that formaldehyde, acetaldehyde, propion- aldehyde and crotonaldehyde did not interfere, and we confirmed this.564 HARRISON, PALFRAMAN AND ROSE : IIETECTION, DETERMINATION AND [VOl. 86 IDENTIFICATION OF FURFURALDEHYDE In the event of a prosecution, positive evidence of the identity of furfuraldehyde may be required. As the ultra-violet spectrum shows no fine structure, and infra-red analysis is impracticable because of the small amount of furfuraldehyde present in a sample of marked oil, a gas-chromatographic technique was evolved. The detection of furfuraldehyde in the sample proved to be impossible, owing to the complexity of the chromatogram, so an extraction method based on formation of the aldehyde - bisulphite compound was tried, as in Gent, Pomatti and Levin's method.2 METHOD APPARATUS- Petroleums for distilling petroleum products boiling below 370" C (IP123/58).cent. w/v of squalane on Celite 545 (100 to 1.20 mesh). Distillation assembly-Use an apparatus of the type recommended by the Institute of Gas chromatograph-Fitted with an ionisation detector. Use a column containing 20 per REAGENTS- Diethyl ether, peroxide- free. Sodium hydroxide solution, 10 per cent. w / v , aqueous. Sodium bisulphite solution, 10 per cent. w/v, aqueous. Sodium sulphate, anhydrozts. PROCEDURE- Concentrate the furfuraldehyde by separatlely collecting the first 60 ml of the 200 ml of distillate from the standard distillation carried out as recommended by the Institute of Petroleum (method C).6 Transfer to a 100-ml separating funnel, and shake for 5 minutes with 10 ml of the 10 per cent.solution of sodium bisulphite. Allow the layers to separate, and run the lower aqueous layer containing the furfuraldehyde - bisulphite compound into a second 100-ml separating funnel. To this solution add 10 ml of the 10 per cent. solution of sodium hydroxide to liberate the furfuraldehyde, and shake for 2 minutes with 5ml of peroxide-free diethyl ether. Dry the neck of the separating funnel, pour the ether extract through the neck of the funnel into a suitable stoppered receptacle, add a little anhydrous sodium sulphate, andset aside for about 15 minutes. Inject about 50p1 (1 drop) of the anhydrous ether extract on to the top of the gas- chromatographic column, and record a chromatogram.Immediately after the chromatogram is complete, record a chromatogram for either pure furfuraldehyde or a suitable reference liquid whose retention time has been adopted as standard; in this laboratory, toluene is used as standard. Allow to separate, and reject the lower layer. Suitable operating conditions when a Pye argon chromatograph is used are- Column temperature-50" C. Detector voltage-2000 volts for the sample; 1000 volts for the standard. Amplifier sensitivity- x 10. Argon input ~5ressure-7 to 10 lb per sq. inch. Chart speed-9 inches per hour. RE s u LTS The retention time of furfuraldehyde on squalane at 50" C was found to be 1.06 relative to toluene (1.00), and, as these retention times were so close, it was considered desirable to ascertain whether or not the presence of toluene in gas oil could interfere with the identi- fication of furfuraldehyde.Six samples of oil, treated as shown in Table 11, were examined, with the results indicated. Toluene, when present in these samples, gave a peak similar in size to that for fur- furaldehyde, and this would represent a concentration of toluene in the sodium bisulphite extract of the order to be expected from the solubility of toluene in water. It was found that a second extraction with sodium bisulphite solution of the diluted ether extract was sufficient to remove any toluene present after the first extraction.September, 19611 IDENTIFICATION OF FURFURALDEHYDE IN HYDROCARBON OIL 565 DISCUSSION OF THE METHOD In order to detect the minimum concentration of furfuraldehyde in the ether extract, a large sample was injected, thereby deliberately overloading the column with ether.As the peak for ether emerges almost immediately and is widely separated from the peak for furfuraldehyde, the resultant broadening of the peak for ether does not cause interference. The concentration of furfuraldehyde in the ether solution was over thirty times that in the original sample of oil, and the minimum concentration detectable was found to be approxi- mately 30 p.p.m. in the ether extract, corresponding to 1 p.p.m. in the original oil. It is therefore possible to identify furfuraldehyde in a mixture of one part of marked oil with nine parts of unmarked oil.Peroxide-free diethyl ether is used to avoid any stray chromatographic peaks, which may occur with analytical-reagent grade ether. Extraneous peaks (notably for toluene) were observed on the chromatograms from time to time, probably caused by contamination of the TABLE I1 IDENTIFICATION OF FURFURALDEHYDE I N PRESENCE OF TOLUENE Furfuraldehyde Oil No. added, p.p.m. 9 5 12 Nil 12 Nil 13 5 13 Nil 13 6 Toluene added, Compound identified % 3 Furfuraldehyde and toluene 3 Toluene 3 Toluene 3 Furfuraldehyde and toluene Nil Nil Nil Furf uraldehyde glassware used in the extraction, which must be kept clean. Toluene is particularly objec- tionable, as its retention time under the conditions used is similar to that for furfuraldehyde and the peaks may overlap if toluene is present in large amount; it is, however, unlikely to be present in gas oil. Chromatograms have been recorded for n-but yraldehyde, n-hexaldehyde, n-heptaldehyde and 5-methylfurfuraldehyde, none of which gives a peak capable of interfering with the identification of furfuraldehyde. This paper is published with the permission of the Government Chemist, Department of Scientific and Industrial Research. REFERENCES 1. 2. 3. 4. 6. 6. Milner, 0. I., and Liederrnan, D., Anal. Chem., 1955, 27, 1822. Gent, L. L., Pomatti, R. C., and Levin, H., Ibid., 1954, 26, 413. Woelfel, W. C., Good, W. Il., and Neilson, C. A., Petroleum Engr., 1952, 24, C42. Javes, A. R., Proc. Amer. Petroleum Inst., 1949, 29M, 39. Borrow, A., and Jefferys, E. G., Analyst, 1956, 81, 598. “IP Standards for Petroleum and its Products,” Nineteenth Edition, The Institute of Petroleum, Received April 18th, 1961 London, 1960, Part I.

ISSN:0003-2654    

DOI:10.1039/AN9618600561

出版商:RSC    

年代:1961

数据来源: RSC

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566 HARRISON AND HEAYSMAN : METHOD 5 FOR DETECTION, DETERMINATION [Vd. 86 Methods for the Detection, Determination and Identification of Quinizarin in Hydrocarbon Oil BY R. B. HARRISON AND L. T. HEAYSMAN (Department of Scientific and Industrial Research, Laboratory of the Government Chemist, Clement’s Inn Passage, Strand, London, W.C.2) A colorimetric method for detecting quinizarin and a spectrophotometric method for its identification and determination in concentrations from 0.2 to 2.0 p.p.m. in hydrocarbon oil are described. IN connection with the marking of heavy oils to distinguish between duty-paid oil for road vehicles (DERV) and duty-free oil for other purposes, quinizarin was chosen as one of the markers, as it fulfilled the necessary conditions stated by Harrison, Palframan and Rose in their paper on furfura1dehyde.l DETECTION OF QUINIZARIN Quinizarin is 1,4-dihydroxyanthraquinone ; a search of the literature2 s3 14 produced the information that it is soluble in ethanol, ether, benzene, alkali or sulphuric acid and gives a blue colour with aqueous solutions of alkali, alkali carbonate or ammonia, but no method of determination was found.A rapid test for quinizarin in solution can be carried out by dipping the end of a chromato- graphic column of alumina into the solution. Quinizarin is adsorbed in a band well defined by the formation of a red lake. With a solution in gas oil, however, the combined effects of the colour of the oil and the low concentration of quinizarin made this technique of little value. The method described below for the roadside detection of quinizarin depends on the colour produced in alkaline solution.When a gas oil containing 2 p.p.m. of quinizarin is shaken with an aqueous solution of sodium hydroxide of any concentration between 1 and 30 per cent. w/v, quinizarin is extracted from the oil and produces a blue-violet colour in the aqueous phase. Stability of the colour increases with increasing concentration of sodium hydroxide, and is sufficient for a visual test for up to 30 minutes when a 1 per cent. w/v solution of alkali is used. METHOD REAGENT- Sodium hydroxide solution, 1 per cent. w/:v, aqueous. PROCEDURE- Pour 15 ml of the gas oil into a stoppered test-tube (6 inches x 0.75 inch), and shake for approximately 15 seconds with 2 @ of the sodium hydroxide solution. Allow the dis- persion to separate into two layers (this may take 2 or 3 minutes), and observe the colour of the lower aqueous phase; a blue-violet colour indicates the presence of quinizarin.The test works well for the chosen concentration of marker (2 p.p.m.) and for a dilution of one part of marked oil with one part of unmarked oil. For a dilution of one part of marked oil with four parts of unmarked oil, the colour is not easy to see in a pale oil and impossible to see in a dark oil. The sensitivity of this test. can be increased by using a larger volume of oil, but the same volume of sodium hydroxide solution. DETERMINATION OF QUINIZARIN An attempt was made to utilise the blue-violet colour produced with alkali as the basis of a quantitative spectrophotometric method, but several difficulties were encountered.Straightforward extraction of quinizarin from a marked gas oil by an aqueous solution of sodium hydroxide, with n-butyl alcohol to prevent emulsification, resulted in a solution that was not optically clear, owing to entrained droplets of oil. Acidification of this extract, subsequent extraction with diethyl ether, chloroform or light petroleum and then re-extractionSeptember, 1961 J AND IDENTIFICATION OF QUINIZARIN IN HYDROCARBON OIL 567 from the organic solvent with aqueous sodium hydroxide did not solve the problem. Centri- fugation of the separated aqueous layer gave an optically clear solution, which, when spectro- photometrically examined, was found to have an absorption maximum at 560 mp, but the colour was not completely stable; further, different gas oils gave different colours varying from blue to blue-violet.During work on the problem of positive identification, the ultra-violet - visible spectrum of a 0.001 per cent. w/v solution of quinizarin in cyclohexane was plotted with an automatic recording spectrophotometer. A characteristic curve was produced having five peaks a t 463, 476, 487, 508 and 521 mp; those peaks at 508 and 521 mp were extremely well defined. It was found that this method could be applied to the determination of quinizarin in marked samples of gas oil, thereby avoiding the difficulties mentioned above and giving results of greater accuracy than those obtained when the coloured aqueous solution was examined spect rophot ome t rically .METHOD REAGENTS- Sodium hydroxide solution, 5 per cent. wlv, apeow. Hydrochloric acid, concentrated. Cyclohexane, spectroscopically pure. n-Butyl alcohol. PROCEDURE- Shake 50.0 ml of the gas oil suspected to contain quinizarin with 5 ml each of the sodium hydroxide solution and n-butyl alcohol in a 100-ml separating funnel for 45 seconds. When the two layers have separated (this may take several minutes), run the aqueous phase into a second 100-ml separating funnel, and wash it through with about 2 ml of distilled water. Extract the gas oil with a further 5ml of the sodium hydroxide solution by shaking for 45 seconds, allow the layers to separate, run the aqueous layer into the second separating funnel, and wash through with water as before.Repeat with a further 5 ml of the sodium hydroxide solution, and, after separation, add the aqueous layer to the previous extracts. Quinizarin forms a blue-violet colour with sodium hydroxide, and the third extract should be colourless; if it is not, extract with further 5-ml portions of the sodium hydroxide solution until a colourless extract is obtained. Acidify the combined extracts by adding 1 ml of concentrated hydrochloric acid for each 5 ml of sodium hydroxide solution used. When the solution has cooled, add, by pipette, 10.0 ml of spectroscopically pure cyclohexane, and shake for 30 seconds. After separation of the two layers, run the aqueous layer to waste, and use a clean dry pipette to transfer the amber-coloured cyclohexane solution to a stoppered parallel-sided 1-cm cell of an Optica automatic recording spectrophotometer.With pure cyclohexane as blank, plot the absorption spectrum from 420 to 540mp; use the peak at 521 mp for determining quinizarin, as this is the most pronounced peak and is also furthest away from any possible interference from components of the gas oil. Prepare a standard curve by using a 0.001 per cent. w/v solution of recrystallised quinizarin in spectroscopically pure cyclohexane. DISCUSSION OF THE METHOD Complete absorption due to components of the gas oil carried over in the separations usually occurs at wavelengths shorter than 420mp, but there is no interference with the five peaks produced by quinizarin. Since the quinizarin used to mark gas oils is of the quality normally available on the market and not specially purified, a more realistic result is obtained if an average absorption figure is taken as standard. Accordingly, 0.01 per cent.w/v solutions of quinizarin from different sources were prepared in cyclohexane and used to mark a gas oil at the level of 2.0 p.p.m. The 0.01 per cent. w/v solutions of quinizarin were also diluted to 0.001 per cent. w/v, and these solutions were used directly in the spectrophotometer. The aliquots of marked gas oil were extracted by the proposed procedure, and the optical densities of the cyclohexane extracts at 521 mp were compared with those of the 0-001 per cent. w/v solutions of quinizarin; the results were- Quinizarin sample No. . . .. .. .. .. 1 2 3 4 Optical density of 0.001 yo w/v solution in cyclohexane .. 0-244 0.240 0.235 0-253 Optical density of quinizarin extracted from gas oil . . 0.243 0.240 0.235 0.253568 HARRISON AND HEAYSMAN METHODS FOR DETECTION, DETERMINATION [Vol. 86 from which it can be seen that the extraction process is satisfactory. The mean optical density of the 0.001 per cent. w/v solutions of quinizarin is 0.243, and there is a Linear relationship between optical density and concentration of quinizarin. In Table I are Listed the results obtained when the proposed method was applied to different oils marked at the level of 2.0 p.p.m. of quinizarin; the mean figure of 0.243 for the optical density at 521 mp was taken as being equivalent to 2.0 p.p.m. of quinizarin. A pale oil has a colour equal to or less than NPA 1, and a dark oil has a colour equivalent to NPA 3.5 TABLE I RECOVERY OF 2 P.P.m.OF QlJINIZARIN FROM VARIOUS OILS Oil No. Colour of oil 1 Pale 2 Dark 3 Dark 4 Pale 5 Pale 6 Dark 7 Dark 8 Dark 9 Dark 10 Dark 11 Pale Optical density at 521 m p 0.237 0.253 0.258 0.237 0.237 0.248 0.253 0,253 0-257 0.257 0.237 Quinizarin content found, p.p.m. 1.95 2-08 2.12 1-95 1-95 2-04 2.08 2.08 2.1 1 2.1 1 1.95 DISCUSSION OF RESULTS The repeatability of the method is within 1 per cent. The result of an extraction from a pale oil is within 2 per cent. of the correct amount, but the result of an extraction from a dark oil may be high by up to 8 per cent. This was found to be caused by absorption from components of the gas oil carried over in the separations. All gas oils absorb slightly in the range 460 to 540mp, and the darker the oil, the greater is the absorption. In the laboratory, where the unmarked oils were available for blank determinations, a correction could be applied, and this brought all the results to within 2 per cent.of the correct amounts. However, in practice, unmarked oil is not available, and no method of producing a satis- factory blank from the marked oil has been found. When this is taken into account, together with the fact that samples of quinizarin from all the manufacturers in the United Kingdom varied in purity by 4 per cent. from the mean value, the proposed method will in the worst circumstances give a result within 10 per cent. of the amount present; for a pale-coloured oil, the result will be well within 5 per cent. of the amount present.During the extraction, the colour in the sodium hydroxide solution may change from blue-violet to blue or fade completely, but this does not affect the determination. The method is satisfactory for concentrations of from 0.2 to 2.0 p.p.m. of quinizarin in gas oil and therefore can be used for a mixture of one part of marked oil diluted with nine parts of unmarked oil. The stability of quinizarin at a concentration of 2 p.p.m. in gas oil is good; determinations at the time of preparation of a dilution and 3 months later gave identical results. IDENTIFICATION OF QUINIZARIN The five peaks at 463, 476, 487, 508 and 521 mp produced by the spectrophotometric analysis are used to identify quinizarin. Other dihydroxyanthraquinones absorb in this region, but they can be easily distinguished from one another.The 1,2- and 1,5- isomers have only two peaks in this region, and these occur between 400 and 440 mp. The absorption spectrum of the 1,8- isomer is similar to that of quinizarin; it has five peaks, each occurring at a wavelength about 50mp shorter than the corresponding peak for quinizarin, but the peaks near 508 and 521 mp are not at all well pronounced, as they are for quinizarin. Meek and Watson3 reported on the spectrophotometric curves of six other polyhydroxyanthra- quinones, none of which can be confused with that of quinizarin. It is possible to see the typical structure of the spectrophotometric curve even when the extraction is made from a gas oil containing 0.2 p.p.m. of quinizarin, i.e., a dilution of one part of marked oil with nine parts of unmarked oil.September, 19611 AND IDENTIFICATION OF QUINIZARIN IN HYDROCARBON OIL 569 This paper is published with the permission of the Government Chemist, Department of Scientific and Industrial Research. REFERENCES 1. 2. 3. 4. 5. A.S.T.M., Tentative Method D155-45T. Harrison, R. B., Palframan, J. F., and Rose, €3. A., Analyst, 1961, 86, 561. “The Merck Index of Chemicals and Drugs,” Seventh Edition, Merck & Co. Inc., Rahway, New Meek, D. B., and Watson, E. R., J. Chem. Soc., 1916, 109, 544. Radt, F., Editor, “Elsevier’s Encyclopaedia of Organic Chemistry,” Series 3, Elsevier Publishing Received April 18th, 1961 Jersey, 1960, p. 893. Co., Amsterdam, 1946, Volume XIII, p. 527.

ISSN:0003-2654    

DOI:10.1039/AN9618600566

出版商:RSC    

年代:1961

数据来源: RSC

摘要:

September, 19611 AND IDENTIFICATION OF QUINIZARIN IN HYDROCARBON OIL 569 Determination of Diquat Residues in Potato Tubers BY A. CALDERBANK, CLARE B. MORGAN AND S. H. YUEN (Plant Protection Ltd., Jealott’s Hill, Bracknell, Berks.) A specific method is described for determining diquat (1,l’-ethylene- 2,2’-bipyridylium dibromide monohydrate) in potato tubers with a sensitivity of 0.01 p.p.m. It depends on measurement of the light absorption of reduced solutions of diquat after concentration and purification by cation-exchange chromatography. Correction for extraneous background absorption from natually occurring plant substances is made by using a “base line” method of calculation. The method has proved extremely reliable for determining traces of diquat in potato tubers harvested from crops sprayed in the field.DIQUAT, 1 ,l’-ethylene-2,2’-bipyridylium dibromide monohydrate (I), formed by quaternising 2,2’-bipyridyl with ethylene dibromide, is the active ingredient of Reglone, a new herbicide and desiccantlJ recommended in the United Kingdom for destroying potato haulm. It is one of a group of quaternary bipyridylium salts whose herbicidal activity depends on their reduction in photosynthesising plant t i s ~ u e . ~ After application of diquat to the foliage of potato plants, death of the leaves is rapid, and the plant is completely desiccated when the crop is harvested 10 to 14 days later. Although diquat is known to be translocated within the plant, initial experiments indicated that only minute amounts of the compound were transported to the tubers.However, for the purpose of assessing possible hazards to the consumer, it was essential to confirm this by precisely determining concentrations present in the tubers after recommended field treatments. It was therefore essential to develop a reliable, sensitive and specific method for determining the compound. Diquat carries two positive charges, is highly polar and cannot be extracted into water- immiscible organic solvents. However, owing to its cationic nature, it is retained on a cation- exchange resin when a dilute aqueous solution is allowed to percolate through the resin. The diquat can be displaced from the resin by a high concentration of an inorganic cation ( e g . , H+, Na+ or Ca2+) and collected in a much smaller volume of solution.Diquat in solution is easily reduced4 with sodium dithionite, by transfer of one electron, to give a water-soluble and relatively stable free radical having an intense green colour.670 CALDERBANK, MORGAN AND YUEN : DETERMINATION [Vol. 86 3 c \ A Wavelength, r n l Fig. 1. Absorption spectra of an aqueous solution of diquat: curve A, before reduction; curve B, after reduction Solutions of the free radical exhibit a sharp absorption peak at 378 mp (see Fig. 1) ; this peak is of a greater intensity (6 = 28,000) than that of unreduced diquat at about 310 mp (e = 19,200), and spectroscopic measurement of reduced solutions provides a sensitive method for determining the compound. When potato-tuber extracts are subjected to cation-exchange chromatography under the conditions necessary for concentrating diquat, the resulting effluents invariably contain small amounts of cationic plant constituents.These also exhibit ultra-violet absorption 1 I 300 400 Wavelength,, rnp Fig. 2. Absorption spectra of ion-exchange effluents from hydrolysates of 1 kg of potato tubers. Broken-line curves are for tubers containing no diquat and full-line curves for tubers containing 0-4 p.p,m. of added diquat : curves A and B, before reduction; curyes C and D, after reductionSeptember, 19611 OF DIQUAT RESIDUES I N POTATO TUBERS 57 1 both before and after reduction, but the intensity of this absorption is relatively less and more linear in the region of 378 mp than at 310 mp (see Fig. 2). For this reason, as well as for the additional sensitivity attained, it is more advantageous to determine diquat in such a solution by making measurements at the longer wavelength after reduction.Correction for the small amount of absorption from plant constituents can then be mathematically applied by using the “base-line” method of calculation first introduced6 for the analysis of vitamin A. This procedure has the advantage of obviating the need for analysing control samples from untreated plants. Other methods of determining diquat, e.g., by spectrofluorimetry,e are inapplicable when small concentrations of the compound are to be determined in potato-tuber extracts, owing to interference from naturally occurring cations. The reduction method has the additional advantage of complete specificity for the herbicide.EXPERIMENTAL EXTRACTION OF DIQUAT FROM POTATO TUBERS- Diquat is highly soluble in water and is adsorbed from aqueous solutions on to many materials, including starch, activated carbon and soils. It seemed likely, therefore, that any diquat translocated to the tubers would be adsorbed on the starch or might even be occluded within the granules while deposition of the starch was proceeding. It was therefore clear that hydrolysis of the starch would be necessary in order to free any adsorbed diquat residues before the determination could be attempted, and this was confirmed by experiment. Hydrolysis of a potato-tuber macerate by N sulphuric acid showed that hydrolysis of starch, as measured by the increase in concentration of reducing sugar,’ was substantially complete after boiling for 12 hours. Hydrolysis of cell-wall polysaccharides probably con- tinued beyond this stage, and there was a gradual increase in the ease of filtration of the hydrolysate after more prolonged boiling.A 5-hour period of boiling had the advantage of completely hydrolysing hemicellulosic material, leaving a filterable residue and ensuring maximum release of any diquat residues. There was no detectable loss of diquat in N sulphuric acid heated under reflux for this length of time. CONCENTRATION OF DIQUAT ON CATION-EXCHANGE RESIN- The acid hydrolysate of potato tubers is first neutralised with calcium carbonate to remove the large excess of hydrogen ions from solution before passage through the ion-exchange resin. Ethylenediaminetetra-acetic acid (EDTA) is also added in order to remove competing metallic ions that have high affinity for the resin, and the solution is made slightly alkaline.This procedure was found to result in a much decreased background light absorption in the final effluent containing diquat. Zeo-Karb 225 (containing 8 per cent. of divinylbenzene) was the best of the complete range of Permutit cation-exchange resins examined. Diquat could be eluted from this resin by a small volume of solution containing a high concentration of Hf (eg., 5 N hydrochloric acid) or other cations, such as Na+ or Ca2+. Reduction of diquat by sodium dithionite was not possible in acid or in the presence of Ca2+ ions, but proceeded satisfactorily in a saturated (approximately 6 M) solution of sodium chloride, which was found to be an efficient eluting agent for diquat.Recovery of diquat from the resin is reproducible, although not quantitative. Complete recovery can be attained only by collecting a relatively large volume of effluent, but, by limiting the volume to 25 ml the lower recovery (approximately 80 per cent.) is offset by the gain in sensitivity over that which would be obtained if a larger volume of effluent were collected. Intermediate washing of the resin with dilute acid before the final elution with sodium chloride solution was found to remove a large proportion of the interfering plant material without eluting the diquat, thereby considerably increasing the sensitivity of the method. When diquat is adsorbed on the resin from pure solution, the recovery under these conditions is 80 to 90 per cent.When diquat is added to a clear filtered potato-tuber hydroly- sate, which is then allowed to pass through the resin, the recovery in the sodium chloride effluent is 60 to 70 per cent. This suggests that further loss of diquat occurs owing to its non-quantitative retention by the resin in the presence of a relatively large volume (1 litre) of potato-tuber hydrolysate. The cationic naturally occurring plant substances possibly compete with diquat cations for negative sites on the resin.572 CALDERBANK, MORGAN AND YUEN DETERMINATION [Vol. 86 Reduction of diquat by sodium dithionite was found to take place as effectively in 6 M sodium chloride as in water, but maximum absorption was at 379 instead of 378 mp.The light absorption of the potato-tuber constituents eluted by 6 M sodium chloride is much less in the region of 379 mp than at 310mp (where unreduced diquat has its absorption peak). The difference in sensitivity is clearly demonstrated in Fig. 2, which shows the absorption curves of the same mixtures before and after reduction. When sodium chloride effluents were reduced, it was necessary to neutralise the small concentration of acid present by using excess of alkali in the sodium dithionite reagent solution, and, after investigating different relative concentrations of reagents, initial difficulties experienced with the instability of the reduced product were overcome by using a Oh1 per cent. w/v solution of sodium dithionite in 3 N sodium hydroxide. Under these conditions, reduced diquat is stable for a period sufficient to permit accurate measurements of absorption, Solid sodium dithionite can be kept under absolutely dry conditions without change, but in slightly moist air it is rapidly oxidisecl.When stored in a closed bottle for several months, it gradually loses its reducing power, and it was found to be necessary to use frequent supplies of fresh material and also to reduce standard solutions of diquat with each experiment. As well as being oxidised by atmospheric !oxygen, aqueous solutions of sodium dithionite slowly decompose to form thiosulphate and bisulphite. Decomposition is retarded in alkaline solution, and, under the conditions selected for the determination (k, reduction of 10 ml of the diquat solution by 2.0 ml of a 0.1 per cent.W/V solution of sodium dithionite in 3 N sodium hydroxide, the excess of dithionite present was sufficient to continue reduction of a solution containing 5 pg of diquat per ml for up to 3$ hours, after which the concentration of dithionite was too low to effect complete reduction. It was therefore considered advisable to carry out reductions within 1 to 2 hours of preparing the alkaline dithionite reagent solution. We found it necessary to correct for the slight absorption at 379mp contributed by both sodium chloride and alkaline dithionite solutions, and this was done by measuring the solution of reduced diquat against a blank :mixture containing these reagents placed in the reference cell. The product of reduction of diquat in 6 M sodium chloride was stable for 1 hour when measured against an alkaline dithionite - 6 M sodium chloride blank prepared at the same time.However, small errors were incurred if the reduced diquat was measured against a fresh blank or a blank prepared a long time previously, as the diluted dithionite solution remained stable for only about 20 minutes. For this reason, it was found advisable to prepare a fresh blank at the same time as each diquat solution and to take readings 1 minute after mixing the solutions (or as soon after as Fossible) in order to minimise any slight errors due to different rates of decomposition of the dithionite in the blank and diquat solutions. When 10-ml aliquots of solutions of diquat in water or 6 M sodium chloride were reduced by 2.0-ml portions of a 0.1 per cent.w/v soluticln of sodium dithionite in 3 N sodium hydroxide and the optical densities at 378 or 379 mp of the resulting solutions were measured in 1-cm cells against similar blank solutions after 1 minute, the relationship between optical density and concentration was linear over the range 1 to 5 p g of diquat per ml. Beer’s law was obeyed, but the gradient of the line varied slightly on different occasions, owing to changes in the reducing power of the dithionite. For concentrations of diquat below 1 pg per ml, greater sensitivity was attained by using 4-cm cells. Under these conditions, the relationship between optical density and concentration was linear down to 0.2 pg of diquat per ml, and a concentration of 0.1 pg per ml was just detectable.With a 4-cm light path, a concentration of 1 pg of diquat per ml gave, after reduction, an optical density of approximately 0-32. The optical densities at 379 m p of ion-exchange effluents from control (untreated) potato tubers increased on reduction with the dithionite reagent solution, often to the extent of about 50 per cent. Hence, certain naturally (occurring plant substances, which are concen- trated on the resin and eluted with the diquat, are reducible to give a product or products whose absorption at this wavelength is greater than that of the unreduced forms. In mixtures containing diquat it is therefore impossible to ascertain the contribution of the plant material by measuring the optical density at 379 mp before and after addition of the dithionite solution, in spite of the fact that unreduced diquat has zero absorption at this wavelength. Equations were derived for calculating the absorption at 379 mp contributed by reduced diquat present in ion-exchange effluents also containing plant substances that contribute REDUCTION AND DETERMINATION- CORRECTION FOR BACKGROUND ABSORPTIONSeptember, 19611 OF DIQUAT RESIDUES IN POTATO TUBERS 573 background absorption to an unknown and variable extent.This method is based on that described5 for determining vitamin A in mixtures, assuming that the background absorption is linear in the narrow wave-band chosen, and it entails accurate measurement of the intensity of absorption at three wavelengths. From these values are calculated the contribution of the pure compound in the mixture (after having measured the relative optical densities of the compound alone at these wavelengths).The background absorption of reduced effluents from potato tubers was found to be sufficiently linear in the range 375 to 385 mp to permit application of this method, and equa- tions were derived in two ways to correct the composite absorption for the unknown amount of background material mixed with the reduced diquat. For equation (l), the wavelength of maximum absorption for reduced diquat (379 mp) was selected, together with supplemen- tary wavelengths (375 and 385 mp) on either side of the absorption peak. For equation (2) were used 379 mp and supplementary wavelengths of 375 and 383 mp, at which the optical densities of reduced diquat in sodium chloride solution were found to be the same.The necessary constants for the optical-density ratios of pure diquat were determined by reducing solutions of diquat in 6 M sodium chloride (10 ml) with a 0.1 per cent. w/v solution of dithionite in 3 N sodium hydroxide (2ml). The optical densities at the four wavelengths mentioned above were accurately measured, and, from an average of ten determinations, it was found that- I<, = K2 = K, = DERIVATION OF EQUATION (1)- If E379, E3, and E3, are the observed optical densities of the mixture at 379, 375 and 385 mp, respectively, and b379, b,,, and b3,, are the background contributions to these figures, then (E379 - b37g) , (E,,, - b,,,) and (E385 - b385) will be the contributions of diquat to the observed absorptions at these wavelengths.Since these contributions are for diquat only, then- E379 - b379 =-= K, = 1.260 , . .. .. . . (i) E375 - b375 . (ii) Since the background absorption is linear- b37g = 379 m + c b,,, = 375 m + c b385 = 385 m + c where m and c are constants for the straight line. By eliminating the constant c from these expressions- b,,, = b,,, - m (379 - 375) b,, = b,,, - m (379 - 385) Substitution of these values for b,,, and bas5 in equations (i) and (ii) and solving for b3,, gives- The corrected value for the absorption of diquat at 379mp is given by- and therefore b,,, = 2.28 E375 + 1.52 E385 - 2.79 E379. E,,, (corrected) = E,,, - b379 E,,, (corrected) = 3.79 E1,, - 2.28 E,,, - 1.52 E385 . . .. * . (1)574 CALDERBANK, MORGAN AND YUEN : DETERMINATION [Vol.86 DERIVATION OF EQUATION (2)- If E,,, and E,,, are the observed optical densities of the mixture at 375 and 383mp, at which wavelengths the absorptions of reduced pure diquat in 6~ sodium chloride are equal, then, from the similar triangles GHK and FHJ in Fig. 3- KH GK GK JH -FJ AL ---- -- GK is therefore equal to AL.KH - JH = (E375 - E383) f83 383 - 37g) 375 = & (E,,, - E3=) Now the absorption contributed to the mixture by’diquat is given by (E383 - HN) at 383 mp and (E,,, - GM) at 379 mp, i.e.- from which- KM = 2.985 E383 - 3.973 E,,, + 1.986 But- E,,, (corrected) = E,,, - KM - GK and substitution for KM and GK gives- E,,, (corrected) = 4.97 E,,, - 2.49 (&, + ES3) . . .. * - (2) Since differences between observed optical densities are involved in these equations, it is clear that the precision of measurement must be greater than that required in the final result.Measurements are made at wavelengths where the absorption curve falls steeply, Wavelength, rnp Absorption spectrum of reduced diquat in ion-exchange effluent. Curve AOB represents the observed results and is a summation of the curves for the absorptions of reduced diquat (CED) and back- ground material (FGH), both of which are unknown. The absorption of reduced diquat is the same a t 376 and 383 mp, i.e., line CD is horizontal, and line AB therefore represents the gradient of the background absorption Fig. 3.September, 19611 OF DIQUAT RESIDUES IN POTATO TUBERS 575 and it was considered advisable to re-adjust the wavelength settings between determinations on duplicate samples.This, in conjunction with the use of equations (1) and (Z), gives four results for each sample, and these results serve as a check on each other and on any errors in measurement. Excellent replication has been obtained between duplicate observed optical densities when using this method, and the agreement between the corrected absorptions calculated from the two equations is usually within 10 per cent. Obviously, the limiting factor in the accuracy of the method is the degree of linearity of the background absorption. This has been examined for numerous samples of potato tubers of Majestic, King Edward and Redskin varieties, and for all samples the absorption was nearly linear in the range 375 to 385 mp under the conditions described for the deter- mination.Any significant deviations from linearity should be detected immediately by increased differences between the results calculated from the two equations. If the difference is large, it is advisable to use control samples of plant extract alone in order to detect any serious deviations from linearity, since, in this event, the equations would be invalid. In fact, when the equations were applied to effluents from untreated tubers, spurious low results were found in a few instances, corresponding to apparent-diquat contents of less than 0.01 p.p.m. in the tubers. The accuracy of results calculated by using equations (1) and (2) has been confirmed by (a) applying the equations to solutions of pure diquat in 6 M sodium chloride, which gave results of 100 per cent., and (b) adding known amounts of diquat to ion-exchange effluents containing plant material from untreated tubers, which gave results of 92 to 100 per cent.The method of calculation described has the particular advantage of rendering un- necessary the analysis of control samples from untreated tubers. Not only does this decrease the amount of labour required, but it also eliminates substantial errors, since variation in the chemical composition of biological samples is such that control and treated samples rarely have exactly the same content of interfering material. It also provides a method directly applicable to commercial samples when no controls are available. METHOD Diquat is extracted from potato tubers by boiling them with dilute sulphuric acid, and the extract is neutralised and passed through a column of cation-exchange resin.Diquat is retained on the resin and finally eluted by a small volume of sodium chloride solution. It is determined by measuring the optical density of the reduced solution in the region of 379 mp, a correction being made for the irrelevant background absorption. APPARATUS- Macerator-A Top-Drive macerator obtained from Townson and Mercer Ltd, was used. Boiling JEasks-Flasks of capacity 2 litres fitted, by means of standard ground-glass Tubes for cation-exchange columns-Glass tubes, 40 to 50 cm long and 9 to 10 mm internal Automatic-feed or flow-control devices joints, with reflux condensers. 'diameter, with taps (25-ml burettes are suitable).are advantageous. REAGENTS- acid to about 1 litre of water, allow to cool, and dilute to 2 litres. Spectrophotometer-A Unicam SP500 or SP600. Sulphuric acid, 18 N-Cautiously add, with stirring, 1 litre of concentrated sulphuric Hydrochloric acid, 2 N-Dfiute 175 ml of concentrated hydrochloric acid to 1 litre. Sodium chloride solzction, approximately 6 M-Shake 400 g of analytical-reagent grade Ethyzenediaminetetra-acetic acid, disodium salt. Capryl alcohol. Cation-exchange resin-Use Permutit Zeo-Karb 225 (52 to 100 mesh), containing 8 per cent. of divinylbenzene, in the sodium form. Pack the column by adding approximately 3 g of the resin to about 15 ml of water in the tube, so that the settled column, supported on a pad of glass-wool, is 5 cm in length. Pack a small pad of glass-wool above the resin, allow the water to run away, and pass successively through the column at 2 to 5 ml per minute 25 ml each of 6 M sodium chloride and water; the column is then ready for use.Keep the resin covered with water or solution at all times. Use a freshly prepared column of new resin for each test. sodium chloride with a litre of water until the solution is saturated.576 CALDERBANK, MORGAN AND YUEN : DETERMINATION [Vol. 86 Sodium dithionite solution, 0.1 per cent. w l v , in 3 N sodium hydroxide-This reagent is unstable; it should be prepared immediately before use and on no account used after it has been prepared for longer than 1+ hours. STANDARD SOLUTIONS OF DIQUAT- Stock solution, 250 p.p.m.-Dissolve 25 mg of pure diquat in 6 M sodium chloride, and Solution A , 10 @.$.m.-Dilute 10 ml of stock solution to 250 ml with 6 M sodium chloride. Solution R, 2.5 p.p.m.-Dilute 25 ml of solution A to 100 ml with 6 M sodium chloride.Solution C, 1.5 p.p.m.--Dilute 15 ml of scllution A to 100 ml with 6 M sodium chloride. Solution D , 1.0 @.p.m.-Dilute 10 ml of solution A to 100 ml with 6 M sodium chloride. Solution E , 0-5 p.p.m.-Dilute 5 ml of sol.ution A to 100 ml with 6 M sodium chloride. These solutions are stable under normal laboratory conditions, but must not be exposed dilute to 100ml with 6~ sodium chloride. to direct sunlight. EXTRACTION AND CONCENTRATION OF DIQUAT-- Take about 2500g of tubers a t random from the sample provided, wash free from soil, and remove surplus water with a dry cloth.Cut each tuber into four approximately equal segments, and reject two opposite segments from each tuber. Cut the remaining segments into small pieces (approximately 1-cm cubes) with a chipping machine or knife, thoroughly mix the pieces, and weigh out a 500-g portion for the determination. Place approximately half of the weighed portion into the macerator jar (since this will not accommodate the whole 500 g), add 150 ml of water and 5 ml of 18 N sulphuric acid, macerate for 3 minutes, and transfer the macerated material to a 2-litre boiling flask, Treat the remainder of the 500-g portion in the same way, and add the macerated material to the contents of the boiling flask. Rinse the macerator jar with 40ml of water, transfer the rinsings to the 2-litre flask, and then add 34 ml of 18 N sulphuric acid and a few drops of capryl alcohol.Swirl the flask, support it on an asbestos-coated gauze over a tripod, attach a reflux condenser, and gradually heat with a bunsen burner until the contents of the flask boil gently. Swirl the flask occasionally to prevent local overheating and charring, and allow the solution to boil steadily. Boil under reflux for 5 hours, and allow to cool (the mixture can be left overnight at this stage). Quantitatively transfer the contents of the flask to a 3-litre beaker, and neutralise the excess of acid by adding 50g of powdered calcium carbonate in small portions; keep the suspension well stirred during the addition. (Add a further few drops of capryl alcohol during the addition of calcium carbonate to decrease the amount of froth formed.) Add 10 g of anhydrous sodium carbonate, again in small portions, and stir to dissolve. Pour the suspension on to a Whatman No.5 filter-paper supported in a 16-cm Buchner funnel, apply suction, and collect the filtrate in a 2-litre filter flask. Suck the residue dry, and wash it successively with two 100-ml portions of water; allow the first portion to be sucked through completely before the second is added. Return the clear filtrate to the original beaker, add 5 g of EDTA, and stir to dissolve. With use of a pH meter, adjust the pH of the solution to between 7.1 and 7.5 by adding small amounts of anhydrous sodium carbonate (a total of about 1.5 g will be needed). Quantitatively transfer the solution to a ].-litre separating funnel supported above the previously prepared cation-exchange column.Adjust the rate of flow to between 7 and 8 ml per minute, and allow all the solution to percolate through the column. Then, at the rate of about 2 ml per minute, allow 25 ml of water, 50 ml of 2 N hydrochloric acid and 25 ml of water (in that order) to percolate through the column. (The process can be discontinued overnight at this stage, provided that the resin is covered Ldh a small volume of water.) Run the water from the column, and elute the diquat by passing 25 ml of 6 M sodium chloride through the column at about 1 ml per minute. Collect the effluent in a 25-ml calibrated flask, and adjust the volume to 25 ml with effluent. DETERMINATION OF DIQUAT- By pipette, transfer a 10-ml aliquot of effluent to a 25-ml stoppered cylinder, add 2.0 ml of sodium dithionite solution, and mix.Within 15 minutes of adding the sodium dithionite, measure the optical densities of the solution at 375, 379, 383 and 385 mp in 4-cm glass cellsSeptember, 196 13 OF DIQUAT RESIDUES IN POTATO TUBERS 577 with a spectrophotometer ; use as reference solution a mixture of 10 ml of 6 M sodium chloride and 2-0 ml of sodium dithionite solution, prepared at the same time. Record the observed optical densities at these wavelengths (E375, E3791 E,,, and E3=). Measure in a similar manner, concurrently with each series of determinations, the optical densities at 379 mp of 10-ml aliquots of standard diquat solutions B, C, D and E, each reduced with 2-0 ml of sodium dithionite solution; use a reference solution prepared as indicated above.(Standard solutions containing higher concentrations of diquat must be prepared if the optical density recorded for the sample solution is greater than that of standard solution B). From these readings, construct a graph relating optical density at 379mp to concen- tration of diquat in parts per million. CALCULATION- tion by means of equations (1) and ( 2 ) . be For each solution tested, correct the optical density at 379 mp (E,,J for irrelevant absorp- Let the optical density corrected by equation (1) and by equation (2) E”379. E’379 = 3.79 E379 - 2.28 E375 - 1.52 E385 E”379 = 4.97 E379 - 2.49 (E375 + EMS) These equations are based on the assumption that the irrelevant absorption is linear, or nearly so, with respect to wavelength over the range 373 to 385 mp.The optical density of reduced pure diquat in 6 M sodium chloride is the same at 375 mp and 383 mp, and the difference between the observed optical densities at these wavelengths, E375 and E,,,, there- fore indicates the gradient of the irrelevant absorption. Let the mean of E’379 and E’’379 be E,,, (mean corrected), and, from the previously constructed calibration graph, ascertain the concentration of diquat present in the final effluent corresponding to this value. Let this concentration be Y p.p.m., and calculate the concentration of diquat in the sample (see Note) from the equation- loo x Y Volume of effluent, ml Weight of sample, g Diquat content, p.p.m. = X Recovery, yo 25 100 500 Recovery, yo Recovery, yo - 5Y - NOTE-The amount of diquat present in the effluent does X Y not represent the total amount in the sample taken for analysis, as &ere are losses a t the various stages of thk assay.The amount therefore has to be corrected by applying a factor dependent on the percentage recovery. Before the method is routinely applied, each operator must carry out a series of determinations on samples prepared by adding diquat a t the appropriate concentration to unhydrolysed macerates of control potato tubers in order to ensure that a reproducible technique has been acquired. Also, during the routine analysis of samples, recovery experiments should be carried out in the same way on each of the varieties of potato being tested. DISCUSSION OF RESULTS I t should be understood that the 1 ,l’-ethylene-2,2’-bipyridylium ion is determined, but concentrations are expressed in terms of the monohydrated dibromide.LIMITS OF DETECTION- The weight of potato-tuber sample taken for each individual determination is 500g, and diquat can be detected in ion-exchange effluents at a minimum concentration of about 0.2 pg per ml. Twenty-five millilitres of effluent are collected, so that this minimum repre- sents 5 4 p g of diquat, equivalent t o a detection limit in the tubers of 0.01 p.p.m. Further sensitivity below this level is ultimately limited by the concentration of interfering cationic compounds present in the effluent. RECOVERY- was 59 per cent. chopped potato tubers from untreated plants before the determination. Diquat is normally obtained pure as a crystalline pale-yellow monohydrate.In these laboratories, the average recovery in a series of experiments by two operators This is shown in Table I, which records recoveries of diquat added to[Vol. 86 578 CALDERBANK, MORGAN AND YUEN : DETERMINATION TABLE I RECOVERY OF UIQUAT ADDED TO 500-g FORTIONS OF UNTREATED POTATO TUBERS (MAJESTIC VARIETY) Diquat recovered by operator -4 Diquat recovered by operator R A I 7 w 7 Equation (1) used Diquat added, r-A-, 0.03 1 65 {::-: 1 ;::;; 62 54 0.054 56 0.096 < 0.051 54 0.057 59 0.054 56 0.065 68 0-127 66 (. . . 60 Average . . p.p.m. P.P.m- % - - 0.048 - - i 0.192 Equation (2) used w-7 P.P*m* % 0.026 55 - - - - 0.056 59 0.049 52 0-051 53 0-045 47 0.054 56 0.052 54 0.06 1 64 0-118 61 - - - - 56 Equation (1) used & P*P.m* 70 0.029 60 0.026 54 0.033 69 0-065 68 0.054 56 0.058 60 - - - - - - __ - 0.124 65 0-120 62 0-117 61 62 Equation (2) used 0.028 58 0-025 52 0-03 1 65 0.063 66 0.048 50 0.054 56 - - - - - - - I 0.119 62 0.111 58 0.105 55 58 \ Y --A -___ Mean average recovery, % .. . . . . 59 Statistical analyses of these results showed that (a) the mean difference between operators was not significant, (6) the difference between results obtained from the two equations was consistent, but small when compared with the difference between experiments and (c) the standard error of a single determination was k5.3 per cent. FIELD TRIALS- During 1959, diquat was used for destroying potato haulm in extensive field trials at ten independent sites in the United Kingdom in order f o assess the residual levels in the harvested tubers.In these experiments the rates of application per acre were from 18 to 4 lb of diquat in 20 gallons of water. All treatments were replicated four times, and 10-lb samples of tubers were collected at random from each experimental plot. At least two of the four replicate samples were analysed by the proposed method, and recovery experiments were carried out concurrently on the varieties testled. Some of the results are summarised in Table 11. It was found that the diquat contents of the tubers increased with the rate of application, although not in direct proportion. At the commercially recommended rates, Le., '1.5 to 2 lb of diquat per acre, the residues were less than 0-05 p.p.m.For control tubers, the apparent-diquat contents obtained by application of the two equations were of the order of 0.01 p.p.m, No correction for this was madle in the results for the treated tubers, which were all less than 0.1 p.p.m. In the recovery experiments with 0.048 to 0.24 p.p.m. of added TABLE I1 DIQUAT RESIDUES FOUND IN POTATO TUBERS AFTER FIELD TRIALS IK 1959 The figures in brackets are the numbers of determinations made. All results have been corrected for recovery of 59 per cent. Residue found after application of- Variety of potato no diquat, p.p.m. Arran Pilot . . 0.015 (2) Redskin . . . . 0-008 (2) Over-all mean and Majestic . . . . 0.012 (3) Record . . . . 0.010 (2) standard error . . 0.011 f 0.002 (9) 1-5 lb of diquat per acrc, p.1j.m. 0.027 (5) 0.015 (3) 0.023 (5) 0.019 (5) 0.022 f 0.002 (18) 2 lb of diquat 4 lb of diquat per acre, per acre, p.p.m.p.p.m. 0-025 (4) - 0.020 (2) - 0.034 (4) 0.091 (4) 0.023 (4) 0-038 (4) 0.026 f 0-002 (14) 0.051 f 0.014 (8)September, 19611 OF DIQUAT RESIDUES IN POTATO TUBERS 579 diquat, the mean recovery (k the mean standard error) based on twenty-four determinations was 58 +_ 2.2 per cent., which compared favourably with that shown in Table I. There appeared to be no significant difference in the recoveries obtained for different varieties of potatoes. In 1960, further experiments on potatoes were carried out on l-acre plots at twelve sites, with the object of comparing applications of 2 lb of diquat in small (20 gallons) and large (100 gallons) volumes of water per acre; results for diquat found in the tubers are shown in Table 111.From these results it is clear that, at the recommended rate of application of diquat, there was no significant difference between the residues found in tubers sprayed with the solutions of small and large volume. Untreated controls had apparent-diquat contents of the order of 0.005 p.p.m., whereas treated tubers contained residues of about 0.04 p.p.m. TABLE I11 DIQUAT RESIDUES FOUND IN POTATO TUBERS AFTER APPLICATION OF 2 lb OF DIQUAT PER ACRE IN 1960 FIELD TRIALS The figures in brackets are the numbers of determinations made. All results have been corrected for recovery of 59 per cent. Residue found after application of diquat in- Residue found p.p.m. Variety of potato in control, King Edward .. . . 0.005 (4) Majestic . . .. .. 0.004 (2) Dunbar Standard . . .. 0-004 (2) Redskin . . . . .. 0.005 (1) Over-all mean and standard error . . .. . . 0.004 f 0.001 (9) r 1 20 gallons of water, 100 gallons of water, p.p.m. p.p.m. 0.041 (7) 0.043 (9) 0.039 (5) 0.037 (5) 0.019 (2) 0.041 (2) 0.036 (3) 0-033 (3) 0.037 & 0.006 (17) 0.040 & 0-005 (19) METABOLISM- In experiments with plants grown in boxes and sprayed with l*C-labelled diquat it was shown that the total radioactivity in the tubers could be completely accounted for as un- changed diquat, residues of which were small (0.05 p.p.m.); the amount of any radioactive metabolite of diquat translocated to the tubers was therefore negligible. This evidence, together with the absence of detectable metabolic products of labelled diquat in the foliage, indicates that hazards to the consumer arising from the formation of toxic metabolites of diquat may be discounted. APPLICATION OF METHOD TO OTHER CROPS- With slight modifications to the procedure for extraction and processing, the method was found to be applicable to the determination of diquat in other crops, including ginned cotton seeds, onions and peas. Provided that the diquat can be successfully extracted from the crop, the main criterion of applicability of the method is linearity of the light-absorption curve in the region 375 to 385 mp for reduced naturally occurring plant compounds in the ion-exchange effluent. We thank Mr. J. H. Dunn for his part in the early stages of the development of the method and for helpful discussions. Dr. S. H. Crowdy, Mr. G. Douglas and Mr. M. G. Ashley planned the field trials, and Mr. Ashley supervised the subsequent analysis of the field samples and collated the results. The technical assistance of Mrs. Dorothy Pettifer is also gratefully acknowledged. REFERENCES 1. Brian, R. C . , Homer, R. F., Stubbs, J., and Jones, R. L., Nature, 1958, 181, 446. 2. Calderbank, A., Agric. Vet. Chemicals, 1960, 1, 197. 3. Homer, R. F., Mees, G. C., and Tomlinson, T. E., J . Sci. Food Agric., 1960, 11, 309. 4. Homer, R. F., and Tomlinson, T. E., Nature, 1959, 184, 2012. 5. Morton, R. A., and Stubbs, A. L., Analyst, 1946, 71, 348. 6. Freed, V. H., and Hughes, R. E., Weeds, 1959, 7, 364. 7. Somogyi, M., J. Biol. Chem., 1945, 160, 61. Received March 28th, 1961

ISSN:0003-2654    

DOI:10.1039/AN9618600569

出版商:RSC    

年代:1961

数据来源: RSC

摘要:

580 PIERCE AND PECK: A RAPID METHOD FOR [Vol. 86 A Rapid Method for Determining Indium by Neutron Activation BY T. B. PIERCE AND P. F. PECK ( U. K . Atomic Energy Authority, Atomic Energy jPeseavch Establishment, Harwell, Didcot, Berks.) A method is described for determining indium in complex mixtures by neutron-activation analysis. The indium is rapidly isolated from other active components by passage through a column consisting of dithizone and an organic solvent retained on cellulose acetate. By this means, the high potential sensitivity afforded by the measurement of the 54-minute lleIn nuclide can be conveniently exploited. NEUTRON-ACTIVATION analysis is an extremely sensitive method for the quantitative deter- mination of many elements1 Nevertheless, the activity induced in a complex mixture during irradiation is likely to be due to a number of nu.clides, and care must be taken to ensure that the radiation from the element to be determined can be distinguished from all other activity present. It is sometimes possible to carry out measurements without prior chemical separa- tion by correcting for interfering radiation after analysis of the decay curve of the sample, by excluding unwanted radiation with use of ga.mma-ray spectrometry or by choosing irradi- ation and decay times so that, at the time of measurement, the activity of the sample is due solely to the element being determined. Usually, however, chemical separation is effected to achieve radiochemical purity, and a k:nown amount of inactive carrim is first added to avoid microchemical manipulation and to permit assessment of the chemical yield.If this separation is lengthy and the half-life of the activity being measured is short by corn- parison, sensitive determination may be precluded. Further, the more complicated the separation technique, the greater will be the likelihood of low over-all chemical yield, and associated with this will be reduced sensitivity. It is therefore desirable that the method of separation should be as rapid and simple as possible. A sensitive method for determining indium is by measuring the activity of the 54-minute lleIn nuclide, which could be expected to give an ultimate sensitivity of 5 x 10-l2g after irradiation for one half-life in a flux of 10l2 neutrons per sq. cm per second (if an efficiency of counting of 10 per cent.and 10 counts per minute above background are assumed2). However, in view of the relatively short half-life of the isotope, samples must be rapidly processed if the high potential sensitivity of the method is to be exploited. The usual techniques for separating indium from complex mixtures, such as rocks or minerals, involve a number of precipitation and extraction ~ t e p ~ , ~ , ~ but it was thought that the separation could be considerably simplified by using a column consisting of a solution of a chelating agent retained on a solid support. EXPERIM.ENTAL COLUMN MATERIAL- Many chelating agents used extensively in two-phase systems for extracting metals are hydrophobic and cannot be used in solid form as column material, since the rate of formation of the metal complex in the absence of organic: solvent is exceedingly slow. Nevertheless, solutions of these substances can be used as stationary phase for column operation if they are retained on a solid support that holds them firmly enough to prevent them from being stripped off the column by passage of an aqueous phase and yet does not impede reaction between metal and complexing agent.Silica gel has been used to retain solutions of dithizone in carbon tetrachloride or chl0roform,4~~ but, as careful pre-treatment is necessary to obtain a gel of satisfactory form and purity, the possibility of using other support materials was investigated. It was found that a number of polymers could be “gelled” sufficiently with some organic solvents to permit appreciable retention of the solvent without causing the separate particles to coalesce.The best of the polymers investigated, cellulose acetate, retained dithizone dissolved in a mixture of chloroform and carbon tetrachloride so firmlySeptember, 19611 DETERMINING INDIUM BY NEUTRON ACTIVATION 581 that the support could be made into a slurry with an aqueous phase or mechanically packed into a column without stripping. Only when high pressures were used to increase the rate of flow did the organic solution leave the support. The ability of commercially available cellulose acetate to retain chloroform - carbon tetrachloride was found to vary considerably from sample to sample. Some cellulose acetates were capable of holding little organic solvent, which resulted in decreased extraction of the metal and poor separation factors, even under optimum conditions; the weight of organic solvent on a satisfactory support was at least three times the original weight of the cellulose acetate.Studies with indium tracer indicated that the element could be quantitatively extracted from a variety of aqueous solutions buffered to a pH of approximately 5 if the indium solution was passed down a dithizone column of the type discussed above. The complex formed was sufficiently stable to permit the column to be washed with a number of aqueous phases of different compositions without dissociating the indium dithizonate, but N hydrochloric acid eluted the indium quantitatively. ACTIVATION OF SAMPLES CONTAINING INDIUM- The possible errors and limitations of neutron-activation analysis for determining indium have been discussed elsewhere2 and will not be reconsidered here.The effectiveness of the separation of indium in a state of radiochemical purity from complex mixtures (a number of rocks and a meteorite) by means of a dithizone column was investigated. The purity of the indium activity isolated from the irradiated samples was assessed by comparing the gamma-ray spectrograms and the decay curves with those of standards prepared from irradiated pure indium foil. The samples of rock used were a fayalite ferrogabro from the Skaergaard Intrusion, East Greenland (specimen collection No. E.G. 4327), the standard dia- base W-l from Centerville, Virginia, and the standard granite G-l from Westerly, Rhode Island ; the meteorite was the coarse octahedrite Canyon Diablo.IRRADIATION- About 100mg of meteorite drillings or powdered rock were accurately weighed and sealed into silica tubes (4 mm internal diameter). A standard solution of indium in dilute nitric acid was prepared from pure indium foil, and portions of this solution, each containing about 10 pg of indium, were also weighed and sealed into similar silica tubes. Samples and standards were then packed together and activated in a flux of approximately 10l2 neutrons per sq. cm per second for 1 hour. Analysis of the samples of meteorite and rock was begun 5 and 15 minutes, respectively, after their removal from the reactor. PREPARATION OF SUPPORT- Purified dithizones was dissolved in a mixture of equal volumes of chloroform and carbon tetrachloride to give a solution that was almost saturated.Cellulose acetate (16- to 22-mesh, in flake form) was placed in a beaker and stirred as dithizone solution was added to it until a little free liquid appeared at the bottom of the beaker. Surplus solvent was removed by stirring the support in a stream of air until a free-running powder was obtained, and this was then made into a slurry with a 2 N sodium acetate - hydrochloric acid buffer solution (pH 5 ) that had previously been equilibrated with chloroform. The slurry was poured into a chromatographic tube of the normal type, and the cellulose acetate was held in position by placing a sintered-glass disc on top of the column. The satisfactory functioning of a column material of this type is dependent on the retention of an appreciable amount of organic solvent by the cellulose acetate, and great care must therefore be exercised not to over-dry the support before preparing the slurry.The columns used were approximately 2 cm in diameter and 20 cm in depth; rates of flow were normally 15 to 25 ml per minute. SOLUTION OF SAMPLES- Samples of rock were dissolved after being either heated to fumes with a mixture of hydrofluoric, perchloric and nitric acids or sintered with sodium peroxide. With both procedures, it was necessary to precipitate and remove all silica before passing the solution through the column; this avoided separation of silica during passage down the column, which could decrease or even prevent flow of liquid.The more complicated method of solution, involving sintering with peroxide, is described under “Procedure.”682 PIERCE AND PECK: A RAPID METHOD FOR [Vol. 86 Samples of the meteorite were easily dissolved in a mixture of hydrochloric and nitric acids. REAGENTS- nitric acid. METHOD Indium carrier solution-Prepare a solution containing 4mg of indium per mlin dilute Sodium peroxide, powdered. Ammonia solution, sp.gr. 0.880. Perchloric acid, 72 per cent. Sodium acetate, 2 N. Acetate bufer solution-Adjust the pH of 2 N sodium acetate to 5 by adding hydrochloric Hydrochloric acid, sp.gr. 1.18. EDTA solution-Prepare a 5 per cent. w/v solution of the disodium salt of ethylene- Perchloric acid solution, dilute-Saturate 0.01 N perchloric acid with chloroform. Hydrochloric acid solution-Saturate aqueous N hydrochloric acid with chloroform. Oxine solution, 5 per cent.w/v, in 96 per ct:nt. ethanol. Ammonium acetate solution, 2 per cent. w/v, aqueous. PROCEDURE- Transfer the irradiated sample of rock from the silica tube used for the irradiation to a nickel crucible containing sodium peroxide, mix the two powders intimately, and heat in an oven at 480” Ifi 10” C for 10 minutes. Remove the crucible from the oven, empty its contents carefully into a beaker containing 5ml of carrier solution, wash out the crucible, and add the washings to the contents of the beaker. Add a few drops of hydrochloric acid, sp.gr. 1.18, to obtain a clear solution, warm, and then add ammonia solution, sp.gr. 0.880, to precipitate hydroxides. Separate the precipitated hydroxides from the supernatant liquid by centrifugation, discard the liquid, arid dissolve the precipitate in the minimum amount of 72 per cent. perchloric acid.Transfer the solution to a beaker, and heat until fumes of perchloric acid are evolved to precipitate all silica. Adjust the pH of the contents of the beaker to about 5 by adding 2 N sodium acetate, and filter through glass-wool into a 100-ml separating funnel. Shake the filtrate with a few millilitres of chloroform to saturate the aqueous phase, discard the excess of chloroform, and allow the aqueous phase to run through a column of dithizone. Wash the column with 50 ml of acetate buffer solution, 100 ml of E:DTA solution and 100 ml of dilute perchloric acid, and elute the indium with the N hydrochloric acid.Collect 150 ml of eluate as soon as hydrochloric acid begins to leave the column. Make the eluate alkaline with ammonia solution to precipitate indium hydroxide, boil to complete the precipitation, spin in a centrifuge, and reject the supernatant liquid. Dissolve the precipitated indium hydroxide in the minimum amount of hydrochloric acid, sp.gr. 1-18, dilute to about 25m1, and add 3ml of oxine solution. Warm to about 60” C, and add ammonium acetate solution to precipitate indium oxinate. Spin in a centrifuge, discard the supernatant liquid, and wash the precipitate with hot 5 per cent. ethanol. Make the precipi- tate into a slurry with a little 96 per cent. ethanol, transfer the slurry to a weighed aluminium counting tray, and remove the ethanol by evaporation under an infra-red lamp.Weigh to determine the chemical yield, and measure the activity of the sample with a scintillation counter. acid, sp.gr. 1.18, and saturate with chloroform. diaminetetra-acetic acid in water, and saturate with chloroform. TREATMENT OF STANDARDS- Transfer the standard to a 100-ml calibrated flask with a little 60 per cent. v/v hydro- chloric acid, and dilute to the mark with de-ionised water. Add a suitable aliquot of this solution to 5 ml of carrier solution, then add 2 .ml of 72 per cent. perchloric acid, and heat until fumes of perchloric acid are evolved to achieve exchange between active and inactive indium. Dissolve the residue in water, and continue as described under “Procedure,” beginning at “Make the eluate alkaline with ammonia solution. .. .” A separate series of experiments showed that the gamma-ray spectrum and specific activity of indium oxinate prepared by this method were similar to those obtained when theSeptember, 19611 DETERMINING INDIUM BY NEUTRON ACTIVATION 583 indium standard and carrier were precipitated as hydroxide and then treated as described under “Procedure,” beginning a t “Separate the precipitated hydroxides from the super- natant liquid. . . .” DISCUSSION OF THE METHOD After irradiation for 1 hour, the proportion of the total activity of a sample that was due to indium was extremely low. Consequently, it is probable that any failure of the subsequent procedure to isolate indium from all other active elements would lead to a signifi- cant contribution from impurities to the final measured activity.Differences would also be observed between the gamma-ray spectra and half-lives of the indium separated from the samples and from the standards. ‘0406 MeV 1, Energy. 2 Fig. 1. Typical gamma-ray spectra : curve A, standard: curve B. sample No. E.G. 4327 There are certain differences between the extraction of a metal from an aqueous phase by dithizone retained on a column of cellulose acetate and by dithizone dissolved in an organic solvent, but the over-all pattern of extraction appears to be similar. For example, the dithizonates most stable to acid in liquid-liquid systems are formed by metals such a s silver, mercury and palladium, and these require the most concentrated acid for elution from a column of dithizone.Elements likely to interfere in the final determination and which are extracted from the aqueous phase a t pH 5 during the procedure and subsequently eluted by N hydrochloric acid are zinc and cadmium. The activity induced in these elements after irradiation for 1 hour is considerably less than that of the indium, and any interference from zinc and cadmium is further reduced by washing the column with EDTA solution, which decreases the ratio of both zinc and cadmium to indium on the column. No trace of zinc was observed in the final measured activity, even for the diabase W-1, in which the ratio of zinc to indium exceeded 1000 to 1. The indium contents found when the proposed method was applied to various samples of rocks are shown below, each result being the mean of three or more determinations. Sample .. .. .. . . E.G. 4327 w-1 G- 1 Canyon Diablo Average indiumcontent, p.p.m. 0.152 3- 0.001 0.055 0.003 0.025 t 0.002 0-012 0.001 Decay curves of the indium derived from samples and standards indicated that the indium had been isolated in a radiochemically pure state, and this was confirmed by the gamma-ray spectrograms, for which the peak-height ratios calculated for the 0.406-, 1.09- and 1.27-MeV gamma rays were always similar for both standards and samples; examples of typical gamma-ray spectrograms are shown in Fig. 1. The indium contents found by the proposed method compare favourably with those previously reported for E.G. 4327, W-1 and G-l (0-169 i 0.004, 0-064 0-003 and 0.026 k0.002 p.p.m., respectively), which were obtained by measuring the llsIn activity after sep- aration by a more complicated procedure involving precipitation and solvent-extraction stages.584 SIMS: FORMATION OF HETEROPOLY BLUE BY SOME REDUCTION [Vol.86 The chemical purity’of the separated indium was not investigated, because of the relatively large amounts of carrier added, but the final precipitate of indium oxinate was always yellow, even for the meteorite, which contained more than 90 per cent. of iron. Over- all chemical yields depended on the rate of flow of the liquid phase through the column during extraction and elution and on the amount of solvent retained by the solid phase, but yields were at least 50 per cent. and sometimes better than 85 per cent. The time taken for a complete determination was dependent on the time needed for solution of the sample, but, if a rapid result was desired, the indium in the eluate from the column could be precipitated immediately as oxinate, the precipitation as hydroxide being omitted. This permitted counting to be started less than 30 minutes after the beginning of the separation (treatment with chloroform at pH 5 ) . However, this led to lower yields, and the hydroxide precipitation is normally included. We thank Mr. A. A. Smales and Mr. D. Mapper for helpful discussion. REFERENCES 1. 2. 3. 4. 5. 6. Sandell, E. B., “Colorimetric Determination of Traces of Metals,” Third Edition, Interscience Received February 24th, 1961 Jenkins, E. N., and Smales, A. A., Quart. Rev., 1956, 10, 83. Smales, A. A., Smit, J. van R., and Irving, IS., Analyst, 1957, 82, 539. Irving, H., Smit, J. van R., and Salmon, L., Ibid., 1957, 82, 549, Pierce, T. B., Anal. Chim. Ada, 1961, 24, 146. Pierce, T. B., and Peck, I-’. F., J . Chromatograjhy, in the press. Publishers Inc., New York, 1959, p. 170.

ISSN:0003-2654    

DOI:10.1039/AN9618600580

出版商:RSC    

年代:1961

数据来源: RSC