A S d L YTICAL EDI T I O S
4
trations of acid and alkali, so that the colors given cannot be recommended for such conditions. The mixtures of indicator and dye were developed for use in titrating Kjeldahl distillates and hence were not subjected to hydrogen-ion concentrations more acid than that equivalent to pH 1.0 or more alkaline than that equivalent to pH 7.0.
J-ol. 2 , s o . 1
of the mixture of indicator and dye from that of the indicator alone. Methylene blue or guinea green gave the best results Tvith methyl red. Guinea green or indigo carmine gave the best results Lvitli sodium alizarin sulfonate.
Conclusions
Literature Cited
The use of several blue greenish blue dyes methyl red and with sodium alizarin sulfonate has been found to effect an increase in the sensitivity with which the end point can be determined in titrating acids and bases. The most satisfactory proportions in which to combine indicator and dye are given, the criteria being to obtain the sharpest end point possible and not to change the pH range
(1) Am. Assocn. Cereal Chem., “hfethods for the Analysis of Cereals and Cereal Products,” p . 19, Lancaster Press, 1928. (*) Cohn, J , Gen, Physiol,, 6, 697 (1922), (3) Hallstrom, Ber., 38, 2288 (1905). (4) Hickman and Linstead, J . Chem. SOL.,121, 2502 (1922). (’) Kirschnick7C ’ z e m ~ - Z t s318. ~ 960 (lgo7). (6) Luther, I b i d . , 31, 1172 (1907). ( 7 ) hfoerk, A ~J , ,Phar,n,, 93,675 (1921), (8) Salle, J . Injec/ecliozis Diseases, 38, 293 (1926).
Measuring the Toxicity of Insect Fumigants’,’ A. L. Strand3
A review of the methods used for establishing the H E search for new mac i e n t s , the ratios Letween relative toxicities of insect fumigants is presented. terials useful as insect the toxicities of fumigants in It is shown that the greatest error in these methods fumigants h a s p r o c o m p a r i s o n with sulfur dirises from the attempt to determine minimum lethal gressed rapidly during the last o x i d e taken as a standard. concentrations. The same method was used few years. Much valuable inA method of measuring relative values by comparing by Moore (12) but with some formation has been a s s e in concentrations which kill 50 per cent of the test insects bled and from this have come variations. The concentrain a period of 5 hours has been investigated. These several compounds which are tion of a fumigant, expressed concentrations may be designated as the 5-hour median in gram molecules, required proving of economic imporlethal concentrations. The method appears to possess to kill 5 house flies in 400 tance. Although the methgreater possibilities for accurate work on fumigants minutes was taken by Moore ods for d e t e r m i n i n g t h e than those now in general use. a s t h e measure of toxicity. relative toxicities of chemical compounds have served The experiments were carried very well for pointing out those of outstanding value, the out in liter flasks a t room temperature. Tattersfield and use of many of the data for establishing the general Roberts (23) followed the method of Moore, but also used principles underlying the study of t,he subject is question- a “mean toxic concentration,” the mean of the upper and able, because of the differences in these methods and the lower concentration (death and recovery) values. The same method, but with still greater increase in the selection of criteria for the indication of toxicity. One of the factors measured, and one on which the great majority time of exposure was used by Neifert and his eo-workers of workers seem to agree, is the very one that may coiitribut’e ( 1 4 ) , mho express their criterion of toxicity as “the minimost to inaccuracies in interpretation. Hence a compari- mum percentage concentration which consistently causes son of t,he results of several workers is impossible if close 100 per cent mortality after exposure of 24 hours.’’ The estimates are desired. It would seem worth while, then, to experiments were performed in glass vessels under temperareview briefly what these methods have been and to discuss tures varying from 21” to 32” C. Fleming’s method ( 7 ) possible improvemenB based mostly on findings in the meas- is similar, but his experiments were made a t 26.5” C. Roark and Cotton (6, 18, 19) used the minimum lethal concentraurement of toxicity already made. tion, but with the variation of having the test insects in the Previous Work flasks covered by 250 cc. of wheat. With one exception, then, all of these investigators determined the minimum Among the first important papers on insect funiigant,s amount of a substance required to kill 100 per cent of the having to do with the establishment of relative values is insects in a fixed period of time. that of McClintock, Hamilton, and Lowe (10). These Holt ( 9 ) took the average time to kill 100 per cent of the workers determined the Concentrations of subst,ances required test insects a t varying concentrations. Barnes and Grove ( 2 ) to kill various insects in 1 hour. Exceptions were made in determined the time, temperature, and gas concentration the case of pyridine and nicotine for which %hour exposures necessary to produce death, calling the time so taken t o kill were allowed. They also expressed their results as coeffi- insects the “lethal period.” Keifert and Garrison (13) made * Received June 24, 1929. Presented before the Division of Agricul- comparisons on the basis of the time to obtain 100 per cent lethal doses a t different concentrations. Bertrand and tural and Food Chemistry at the 75th Meeting of the American Chemical Society, St. Louis, Mo., April 16 to 19, 1928. Rosenblatt ( 3 ) compared the vapors of several substance* 2 Published with the approval of the Director as Paper 843 of the by exposing insects to different concentrations, usually for Journal Series of the Minnesota Agricultural Experiment Station. periods of 1 hour, and taking into account the time for the 8 This paper constitutes one section of a thesis presented t o the Graduinsects to recover. Strand (21) used 100 per cent kills a t ate School, University of Minnesota, in partial fulfilment of the requirements different intervals of time and plotted these against temfor the P h . D . degree.
T
January 15, 1930
ISDL-STRIAL A S D ESGISEERISG CHEMISTRY
perature when the concentration mas constant and against concentration when the temperature was constant. Coleman ( 6 ) describes a procedure for determining the toxicity of hydrocyanic acid gas and indicates that the results are to be based on time and concentration as factors. Some of the results referred to by Coleman are given by Woodworth ( 2 5 ) , who, in a discussion of “the theory of toxicity,” has drawn curves to shorn ‘[toxic phases” in which time, percentage mortality, and concentration are represented. The concentrations, however, increase according to a geometrical ratio and are not expressed in absolute units but only relatively. hllison (1) presents a curve showing the time-concentration relationship for hydrocyanic acid and also a 3-dimensional diagram to sum u p toxicity factors in which time, concentration, and percentage mortality appear. M7e have referred here only to experiments carried out on a laboratory scale and have purposely omitted the citation of the many toxicity experiments designed for the most part as practical fumigations. If we look outside the field of insect fumigation, still other methods for estimating relative toxicity under very analogous conditions may be found. Among these is the method of Powers (16), who tested the toxicity of many inorganic salts to goldfish. H e considers the survival time and the concentration. The reciprocal of this relationship he calls the velocity of fatality. Thus he expresses toxicity in the form of a n equation which takes into account the threshold of concentration and the rate at which lethal action takes place. Likewise, Carpenter (d), interested also in the action of soluble metallic salts on fish, found that the survival time a t 1 different concentrations closely follows the equation K = 1
t
log G., where K is a constant dependent on the toxic substance employed and t is the survival time in minutes. Both Powers’ and Carpenter’s experiments are based on the survival times of individual fish, that is, on 100 per cent mortality. llarcovitch (11) has used this formula for estimating the toxicity of stomach poisons t o mosquito larvae. Which Factors Shall Be Varied a n d Which Held C o n s t a n t i n Measurement of Relative Toxicity?
It is evident that much of the diversity in methods for determining the relative toxicities of fumigants arises from discrepancies between workers in the factors which shall be varied and those which shall be held constant. Altogether, we believe we are safe in saying that the majority favor holding constant in all tile experiments the duration of exposure, the temperature, and, although little is said about it, the percentage of relative humidity. The period of exposure is really a function of the dosage, that is, a known concentration when allowed to act for a known time produces a certain dosage within the body of the insect. If the concentration is t o be varied, it would seem unnecessary to vary the period of exposure. By holding the time constant fumigation experiments are put on much the same basis as those in which other contact insecticides are applied, namely, contact sprays, where the dose given a n insect is achieved almost a t once and there is no time factor other than that for the physiological action of the chemical. The time after the application of the insecticide when the percentage of mortality is calculated, of course, is very important in all cases. This leaves as the only other factors’ to be considered the concentration and the percentage mortality to be used to It is important t o consider the kind of insect used when comparing the results of different workers. One fumigant may outrank another when used against one particular species and fall far below it when tested against other kinds of insects.
5
indicate equivalent concentrations. How concentration shall be expressed makes little difference. Whether or not there is justification for comparing fumigants on a gram-molecular basis, which can do little more than favor the high-boilingpoint, high-molecular-weight compounds, is apart from the present discussion. If the results of investigations in other classes of insecticides or recent pharmacological work on higher animals are to be considered, there can be no doubt but that the general use of the 100 per cent lethal dose idea in fumigation experiments constitutes their greatest error. Tattersfield and Morris (22) have presented a statement by R. A. Fisher, statistician a t the Rothamsted Experiment Station, in which this factor is considered from the theoretical standpoint. It is in part as follows: The relation between the concentration and the probability of death could theoretically be determined by exposing a large number of insects to the action of the insecticide a t each concentration. The number of insects required, however, increases enormously if we wish to explore in this manner the region in which the probability of death is high. In as many as 99 per cent of the insects killed, the accuracy of the comparison between two insecticides would depend upon the comparatively few insects which survived, and to compare them with any accuracy many thousands of insects would have to be used * * * For a given number of insects the most accurate comparison can be made when the Concentrations are such that about 50 per cent perish. The region between 25 per cent and 75 per cent can be fairly easily explored, It is for this reason that the preliminary examination of chemical substances should be made by a comparison of concentrations required to give a mortality of 50 per cent.’ * * The direct comparison of mortality when the probahility of survival is very small would seem to be beyond the scope of accurate laboratory investigation.
Tattersfield and Morris accordingly showed that duplicate experiments on the toxicity of various contact sprays to aphids are in closest agreement when the 50 per cent death points are compared. On the other hand, the minimum lethal concentrations show extreme differences, notwithstanding the fact that fairly large numbers of insects were employed. Richardson and Smith ( 1 7 ) realized this when they compared various sprays on the basis of the concentrations that would kill all but about 5 per cent of the aphids upon which they were experimenting. hlarcovitch (11) used the survival time for 50 per cent of the test organisms in his application of the Carpenter ( 4 ) formula, but does not show how he was able to calculate the exact time at which survival was attained. The obvious way t o do this would appear t o involve observations over a n indefinite period, and, in the case of fumigants where a recovery period is necessary, such a procedure seems entirely unsuitable. Such time-concentration curves t o be of real value should be constructed from a series of concentrationmortality curves for different time intervals. Trevan (24) took the method of Shackell (20) to show in a most excellent manner the error in toxicity measurements if minimum lethal doses are considered. I n fact, he doubts the existence of such doses as ordinarily interpreted in the toxicological work covering higher animals. Trevan also suggests the comparison of conc$ntrations which divide the animals used into two equal groups, all the individuals in one being killed and all in the other surviving. To this 50 per cent mortality concentration he has applied the name “median lethal dose.” This same general idea, of course, had already been expressed by Osterhout (1.5)when he stated that the best time to take as a criterion in toxicity experiments would be when the organism was “half dead.” H e showed that the death curve approaches the axis asymptotically, forming a region where the exact moment of death cannot be determined. The percentage mortality curve for different concentrations of a fumigant or other contact insecti-
ASALYTICAL EDITIOS
6
cide approaches the 100 per cent mortality axis in a similar manner and is accordingly as difficult t o interpret as the exact moment of death of a single organism, For the foregoing reasons, a study of the method by which 50 per cent mortality concentrations of fumigants are c o n pared v a s undertaken. Experimental Method
Erlenmeyer flasks of 6.4 liter capacity when closed with paraffined rubber stoppers were used. The unstoppered flasks were placed in a constant temperature and humidity cabinet where air was rapidly circulating for 2-3 hours before starting the experiment. A temperature of 25' C. and a relative humidity of 40-50 per cent were chosen as constituting "normal" conditions. The stock culture from which insects (Triboliunz c o n f u s ~ m Duv.) were taken for the experiments did not fluctuate more than from 19" to 25' C. and during several hours immediately prior to each experiment the culture mas held at 2 5 O C. Trevan ( 2 ; ) showed that the use of much less than 30 test animals for each concentration brings in a considerable error and that, on the other hand, the use of more than this number contributes little additional accuracy. For this reason a n effort was made to place 30 beetles in each cage, the actual number, however, varying from 25 to 38, with an average of about 30.6. The cages were made of bolting cloth, were cylindrical in shape, and were closed a t the ends with paraffined stoppers. They were suspended with thread so as to remain 4 or 5 inches above the bottoms of the flasks. The liquid substances were measured into the flasks with pipets graduated to 0.01 and 0.002 cc. The hydrocyanic acid gas was generated from calcium cyanide. The exact concentrations were determined a t the conclusion of the experiment by washing out the flask with dilute sodium hydroxide and titrating with 0.01 S silver nitrate. After a consideration of the time-concentration curves of Moore (12) and of Strand (Zf), a period of 5 hours was selected as the standard period of exposure. The more commonly chosen period of 24 hours was considered to be much longer than necessary, and, as several trials must often be made before attaining concentrations that will produce progressive increases in the percentage of mortality from zero to nearly 100 per cent, the shorter period would facilitate the work. The range of concentrations covered is, of course, far below that which would be used in practical fumigation. The highest concentration, roughly, would be the miniinurn concentration permissible in a successful practical fuinigation, a minimum that would have to be present for 5 hours during the course of the fumigation over and in addition, usually, to that lost by adsorption, etc. This minimum would appear to be more useful than the determination of one that would have to be maintained throughout the entire period of the usual 24-hour fumigation. The insects, after being removed from the fumigation chambers were held in shallow glass dishes a t room temperature for 2-4 days follawing the experiments. I n several lots of 30 unfuniigated insects held in the same manner over t h e same periods the iibrmal mortality was so low as to be practically negligible, The percentage of mortality of the fumigated insects \vas recorded 12 to 24 hours after the conclusion of the experinlent and then checked on the second, third, or fourth days, depending on the nature of the fumigant used. With hydrocyanic acid gas, hydrogen chloride, ammonia, chloropicrin, acetic acid, and ethylene dichloride there was little or no change in the mortality percentage after 1 Hydrogen chloride, ammonia, and acetic acid are included here merely f r o m t h e standpoint of t h e method employed. T h e wrlter's Interest I n measuring their toxicity came from a conzideration of some factors in insect respiration t o be discussed elsewhere.
T'ol. 2, s o . 1
24 hours. K i t h carbon disulfide a somewhat longer period was required to obtain the fullest possible recovery of the insects. I n no case did we find recovery after the third or fourth day and in this respect we cannot confirm the work of Hanilin and Reed (8). Experimental Results
The results with hydrocyanic acid, chloropicrin, ethylene dichloride, and carbon disulfide, all common fumigants, together with hydrogen chloride, acetic acid, and ammonia are presented in Table I. The Roman numerals in the table refer to curves drawn for the data and given similar designations in the graphical representations in Figure 1, Table I IKSECTS
INSECTS
hlORC o i i c s . LIVIKGDEAD TALITY
M g . per
Per cenl
liter
INSECTS
0
0 0 3.2 17.2 30.0 56.3 68.8 72.4 79.3 97.0 96.7 96.6 96.9 96.9 100.0
a
21 23 32 25 28 31 31 30 C."
HYDROGEN CHLORIDE I
0.72 1.44 2.16 2.88 3.60 4.32 5,04 6,76
29 26
a
0 0 0 0 0
3 4 26 33 29 30 32 32
9.7 13.3 83.9 100.0 100.0 100.0 100.0 100.0
H Y D R O C E S CHLORIDE I1
0.72 1.44 1.80 2.16 2.52 2.52 2.88 3.24
36
23 4 1 0 2 1 0
0 8 24 32 34 31 33 32
0 25,g 85.7 97.0 100.0 93.9 97.0 100.0
HYDROGEN CHLORIDE I11
0.72 1.08 1.44 1.80 2.16 2.52 2.88
30 32 28 10 6 2 0
0 0 2 21 28 26 35
0 0 6.8 67.7 82.3 92.9 100.0
H Y D R O G E S CHLORIDE IV
1.15 1.44 1.73 2.02 2.31 2.60 2.88 3 17
30 23 16 0 2 0 2 0
1,esM. L. c.
14 30 28 31 29 29
0 23.3 46.7 100.0 93.3 100.0 93.6 100.0
CHLOROPICRIN I
2.60 3.88 5,li 6.47 7.76
32 5 4 0 0
1 26 25 32 30
3.0 83.9 86.2 100.0 100.0
CHLOROPICRIN I1
2 3 3 4 4 5 5
56 08 59 10
61 13 64 6 1.5
30 28 26 23 19 6 2 1
1 2
; 12 26 30 29
Per cent
liter
E T H Y L E N E DICHL< O F:IDE
0 9 18 22
AIORTALITY
M g . per
HYDROCYANIC ACID
0.35 30 0.41 30 0.46 30 0.52 24 21 0.56 0.62 14 0.71 10 8 0.67 0.78 6 1 0.83 1 0 89 1 0.86 1 1.09 1.06 1 1.15 0 0 . 6 0 7 11.L.
INSECTS
LIVINGDEAD
CONCN.
3.2 6.8 13.3 23.3 38.7 81.3 93.8 96.7
19.63 31 30 23.55 27.48 26 31.40 20 35.33 14 39.25 5 2 39.25 43.18 2 47.10 0 33.0 M.L. C.
0 0 3 11 15 23 2i 32 35
0 0 10.3 35,5
51.7 82.1 93.1 94.1 100.0
ACETIC ACID 1
7.81 15.62 23.42 31.23 39 04 46.85
29 14 1 0 0 0
1 13 28 27 30 31
3.3 48.1 96.6 100.0 100.0 100.0
ACETIC ACID 11
10.93 29 12.49 27 14.05 27 18 15.62 17.18 10 3 18.74 20.30 2 1 21.86 1 6 . 0 hI. L. C.
2 3 3 14 20 28 29 29
6.9 10.0 10.0 43.: 66. t 90.3 93.6 96.7
CARBON D I S U L F I D E I
49.06 52.99 56.91 60.84 64.76 28.69 82.61 76.54
27 23 19 13 7 2 1 0
2 7 9 14 23 29 29 29
6.9 23.3 32.9 51.9 76.7 93.6 96.7 100.0
CARBON DISULFIDE II
49.06 26 52.99 24 56.91 25 15 60.84 64.76 4 68.69 4 72.61 0 76.54 0 60.85 bf. L.C.
4 6 9 14 29 24 32 31
13.3 20.0 26.5 48.3 87.9 85.7 100.0 100.0
AMMONIA I ~
~~
27 23 19 9 2
5.30 6.36 7.42 8.48 9.53 10.59 11.65 12.71
2 7 12 19 24 27 26 30
0 0
0
6.9 23.3 38.1 67.9 92.3 100.0 100.0 100.0
A M M O X I A I1
5.30 6.36 7.42 8.48 9.53 10.59 11.65 12.71 1 . 9 3 M.
CHLOROPICRIS I l l
0 0 2 56 30 12 9 4 3 0s 27 20 6 7 3.59 27 2.5 8 8 23 4 10 43 3 13 4.61 17 8 7.5 2 s 5.13 4 96 S 30 1 5 64 93.3 2s 6.15 2 4.70 M. L. C . 0 hI. L. C. = Median lethal concentration.
28 19 28 13 6 1 2 1
L.
1 12 4 18 25 27 29
c.
30
3.4 38.7 12.5 58.1 83.3 96.4 03.6 96.8
Discussion of Results from Standpoint of Method of Determining Relative Toxicity
When the percentages of mortality are plotted as abscissas and concentrations of the fumigant as ordinates, an S-shaped curve is obtained (Figure 1). This is a typical frequency curve and gives the probabilities of death a t the different concentrations. The best range for the comparison of concentrations would be over that part of the curve where the mortalities vary from about 30 to 70 per cent. It is evident that within this range there would be three concentrations suitable for our measurements, as previou’sly called to our attention by Trevan (24) in the case of the toxicity of cocaine to mice.
First, there is the true arithmetical mean. For the hydrocyanic acid curve this would be 0.626 mg. per liter calculated as shovn in Table 11. Table I1 DIfF I N PERCENTAGE MORTALITY LIEAN
MIDPOIKT 0 35 0.45 0.55 0.65 0 75 0.85 0.95 1.05
Discussion of Curves in Figure 1 H Y D R O C Y ~ N I CAcrD-The interesting part of this curve, from the standpoint of toxicity measurements, is toward the 100 per cent mortality axis. Here practically 97 per cent kills were obtained at concentrations varying from 0.83 to 1.09 mg. per liter before a 100 per cent mortality was obtained a t 1.15 mg. per liter. This demonstrates very nrell how the variability of only 3 per cent of the insects can increase the minimum lethal dosage or concentr:ttion by more than three-fourths of the total range of concentrations required to carry the mortality from 0 to 97 per cent. Thus no point on the 100 per cent axis could be designated as the minimum lethal concentration unless it was meant that this
Hydrogen Chloride Acetic Acid Curves for Various F u m i g a n t s from Which t h e Median L e t h a l Doses, or C o n c e n t r a t i o n s , for t h e Confused Flour Beetle Were Calculated
Ammonia Figure 1-Concentration-Mortality
COSCS. INTERVAL M g . Der i i l e v 0.3-0.4 0.4-0.5 0.5-0 6 0.6-0.7 0.7-0. E 0 8-0 9 0 9-1.0 1.0-1.1
7
I S D r S T R I A L , 4 5 0 E S G I X E E R I S G CHEMISTRY
January 15, 1930
cf) 0 11 37 28 16
5.5 1.5
1.0 __
100.0
ZX = 0.626 mg. per liter
(ix) 0 4.Q5 20 35 18.20 12.00 4.675 1.425 1,050
___ 62.65
z”f Second, we could take for our comparison the mode or that class which vould show the concentration a t which the highest percentage of insects was killed; in other words, the concentration shon-n by a point on the curve representing the maximum slope. For hydrocyanic acid this would be approximately 0.55 mg. per liter. And third, that concentration which killed 50 per cent of the insects could be taken as our measure of toxicity. As this seems to be the ideal criterion for tovicity measurements in general, it could very well be adopted for the comparison of the toxicities of fumigants. Following the suggestion of Treran ( Z X ) , we will call this dose the “median lethal dose,” or perhaps in the case of fumigants it would be better named the “median lethal concentration.”
concentration depending on the variability of the test insects would be a t times much above what was actually required. I n ot,her words, it’ could not be done without admitting inaccuracy. CHLoRoPIcRIs-The curves for chloropicrin show how closely the 50 per cent mortality concentrations agree when the same experiment is repeated. Curves I1 and I11 are drawn through the last two sets of data for chloropicrin in Table I, but the data from the first set are represented merely as crosses. CARBON DrscmIDE-The two curves fall fairly close together. Here again, however, the median let,hal concentrations from the two sets of data vary by approximately 0.75 mg., whereas the 100 per cent concentrations differ by 3.95 nig. per liter. FUMIGAST Hydrocyanic acid Hydrogen chloride Chloropicrin Ammoni:i Acetic acid Ethylene dichloride Carbon disulfide
Table I11 :-HOUR ~ I I Z U I A N LETEALC o x s . .rig. g e r liter 0 60i 1.68
;,;;
16 or) 33 00 60,s:
COCFFICII~NT 1.0000 0 3618 0.1291 0.0763 0 0379 0 018k 0.0039
AinfosIa-In Curye I a concentration t o kill 100 per cent of the insects n-as obtained at, 10.59 m g . per liter. On the other hand, in t!ie second experiment (Curve 11) a concentration of 12.71 mg. per liter failed t o kill 100 per cent. In fact, the 100 per cent mortality v a s not reached in the second experiment a t all. Considering that 12.71 nig. per liter is
8
A S A L Y T I C d L E DI TI 011-
almost u p to the 100 per cent mortalit,y mark, there is a spread betiTTeen i t and the 100 per cent dosage on the ot,her curve of 2.12 mg. The t’mo concentrations required to kill 50‘per cent of the insects vary by only 0.37 mg. per liter. With hydrocyanic acid gas as a standard, the median lethal concentrations and coefficients are as given in Table 111. Literature Cited (1) Allison, I O W QStare Coll. J . Sci., 2, 243 (1928). (2) Barnes and Grove, M e m . Dept. Agr. India, 4, 166 (1916). (3) Bertrand and Rosenblatt, Comfit. rend., 168, 911 (1919). (4) Carpenter, Brit J . Exptl. Bid.,4, 378 (1927). (5) Coleman, J . Econ. Entomol., 4, 528 (1911). (6) Cotton and Roark, IND.EXG.CHEM..2 0 , 380 (1928). (7) Fleming, N. J. Agr. Expt. Sta., Bull. 410 (1925). (8) Hamlin and Reed, J . Econ. Enlomol , 20, 400 (1927).
1-01, 2! s o . 1
(9) Holt, Lancet, 190, 1136 (1916). (10) McClintock, Hamilton, and Lowe, J . A m . P u b . Health A s s o c n . , 1, 227 (1911). (11) Marcovitch, Term. Agr, Expt, Sta,, Gull. 139 (1928), (12) Moore, J . A ~ YReseavch, . 9, 371 (1917). (13) Keifert and Garrison, U. S. Dept. Agr., BUU. 893 (1920). (14) Neifert et. a l , , I b i d . , (1926). (15) Osterhout, J . B i d . Cliem., 13, 67 (1915). (16) Powers, Illinois Biological Monograph, Vol. IV, p. 1 (1917). (17) Richardson and Smith, U. S. Dept. Agr., Bull. 1160 (19233). (18) Roark and Cotton, IND.END.CHEM.,20, 512 (1928). (19) Roark and Cotton, I b i d . , 21, 135 (1929). (20) Shackell, J . Phermacol., 25, 275 (1925). (21) Strand, hlinn. 4 g r . Expt. Sta., Tech. Bull. 49 (1927). (22) Tattersfield and Morris, Bull. Entomol. Research, 14, 223 ( 1 9 2 4 ) (23) Tattersfield and Roberts, J . Agr. Sci., 10, 199 (1920). (24) Trevan, Proc. Roy. S O L . B101, , 483 (1927). ( 2 5 ) Woodworth, J . Econ. Enlorno!., 8 , 509 (1915).
Use of Protective Colloids in Colorimetric Determination of Certain Metals as Lakes of Dyes‘ Walter E. Thrun VALPAKAISO ~ X I V E R S I T Y ,VILPARAISO,IXD.
HE use of adsorption indicators for the detection and determination of certain metals (aluminum ( 4 ) , beryl-
T
lium ($?),and magnesium (2,S, 4 ) ) has been the subject of many papers in recent years. For purposes of this report the lakes are divided into two classes. The first class includes those lakes that can exist when the color of the unadsorbed dye is negligible or almost discharged by a change in hydrogen-ion concentration of the solution in which they have been formed. T o the second class belong those lakes that are in suspension in solutions of excess dye which are deeply and possibly differently colored than the lakes. Obviously the first class is more suitable for quantitative work, provided color intensity and adsorption are nearly equal. The use of protective colloids t o keep the lakes in solution is a disadvantage rather than an advantage when lvorking with lakes of the second class. The work here reported, therefore, has been done on lakes of the first class. Yoe and Hill (6) made a few observations on the effect of starch as a protective colloid for the aluminum lake of aurintricarboxylic acid, but did not recommend it. The same investigators (6) reported that glycerol keeps the aluminum lake of alizarinsulfonic acid in suspension, but that the color intensity of the lake formed under the given conditions is decreased. Thrun ( 4 ) found that 1 cc. of 1 per cent gum arabic solution can keep the aluminum lake of auiintricarboxylic acid indefinitely in solution in a volume of 100 cc. and that it does not change the color intensity of the lake. It is the purpose of this paper to call attention to the value of gum arabic and. particularly, of starch glycerite as protective colloids for dilute lake solutions. Experimental The gum arabic solutions were prepared by dissolving clean lumps of the gum in water with the aid of heat. The starch glycerite solutioiis were prepared by shaking the jelly* with water and filtering off the undissolved residue. To obtain the results here given each lake was prepared Received August 5, 1929. T h e u. 8. P. directions for making starch glycerite jelly are as follows, “Starch 10 grams, water 20 cc., glycerol 70 cc. Triturate the starch with the water until a homogeneous mixture is obtained. Then gradually add this to the glycerol contained in a porcelain dish and heat t o about 140° C. Continue the heat, keeping it below 1 4 4 O C . , constantly stirring the mixture until a translucent jelly is formed.” 1
3
according to the directions given in the literature in two series, using different quantities of the salts of the metals which furnish the adsorbing hydrous oxides. The series containing the colloid was compared with the series that contained no colloid. Results LkLUHIhXJ31 1, \hE O F z4URIliTRICARBOYYLIC .ICID I ;)-Gum arabic is effective. Starch glycerite is not effective. a t least in small quantities. ALCMIKUNLAKEOF ALIZARINSULFOSIC ACID (6)-Guni arabic in small quantities is ineffective. If approximately 7 cc. of a 2 per cent solution of the gum is added, before the lake solution is made acid, and the solution made u p to 50 cc. in a tall Kessler tube, upon standing several weeks part of the lake settles toward the bottom while the remainder iiae; toward the top. Starch glycerite solution (1-2 cc. per 50 cc.) will keep the lake in solution unless heat is used during the lake formation. BERYLLIUM LAKEOF CURCURJIIN(2)-Starch glycerite (1 cc. per 12 cc. final volume) causes a very decided increase in color intensity. The presence of this colloid makes the Kolthoff method for detecting and determining small quantities of beryllium more sensitive and accurate. ~ I A G N E S ILAKE V M OF CURCURMIN (2)-Starch glycerite (1 cc. per 12 cc.) retards the rate of settling of the lake. The lake which is formed in the presence of the colloid remains for days. TT hile the unprotected lake gradually disappears. Other lakes not mentioned specifically in the literatuic. such as the alizarinsulfonic acid lake of the ferric hydrous oxide, n-ere found to be kept in solution by starch glycerite. Lakes of oxides other than aluminum oxide formed bv the adsorption of aurintricarboxylic acid are kept in solution by gum arabic. I n general, lakes have a tendency to settle out if the excess or unadsorbed portion of dye is present in relatively low concentration. Under such conditions the protective colloid. are valuable. Starch glycerite loses its effeCtiVeneS8 if the Solution containing i t is heated to a n y extent. This is due to the hydrolysis of the c o m p o u n ~which is the effective colloid. Judging from its effect On the curcurmin lakes Of and magnesium, starch glycerite seems capable of lengthen-
