No.Weight.Caffein per kiloResults.Remarks.Kilos.Gram.4713.600.144SurvivedVomiting observed.5512.75.200do.Stiffness of muscles; no other symptoms.567.95.200Found dead next day5213.60.147Survived576.50.230do.Vomited after 1 hour; convulsions after 1 hour and 45minutes.3923.10.120do.Increased frequency of respiration, thirst, loss of appetite, vomited rest of day when he drank water, salivation, restlessness, passed feces frequently.4811.50.174do.Vomiting observed.4812.00.200Found dead next dayNo vomiting observed. Second dose was given 8 days after first.5413.40.200SurvivedVomiting observed.4913.15.152Found dead next day3814.50.120do.Symptoms after 1½ hours: Dog had a temperature of 104° F. before caffein was given; vomited 3 hours after caffein was fed.1810.80.145do.
Table 13.—Subcutaneous injection; dogs.(Series B.)
No.Weight.Caffein per kiloResults.Remarks.Kilos.Gram.629.300.161SurvivedRestlessness and vomiting one-half hour after injection.61A14.00.160Found dead next day6312.00.150SurvivedRestlessness 1 hour after injection.6414.00.150do.Restlessness and thirst 45 minutes after injection.597.20.160Died 1 hour and 20 minutes after injection.Marked restlessness, but no convulsion.6114.60.143SurvivedSymptoms observed in 1½ hours.
Table 14.—Administration by mouth to puppies.(Series C.)
No.Weight.Caffein per kilo.Results.Remarks.Kilos.Gram.83.100.354Found dead next dayVomited in 4 hours after feeding; restlessness, loss of appetite, thirst, incoordination of muscles, convulsions.93.15.350SurvivedMuscular incoordination and stiffness, restlessness, vomited 1 hour after caffein was given.101.60.350Died in 3 hoursConvulsion; no vomiting.111.26.200Found deadSalivation; convulsions.121.28.200SurvivedNo symptoms.151.20.250do.Subcutaneous injection.163.50.300Died in 1 hourConvulsions 45 minutes after caffein was fed.
The object of this study was to ascertain the effect of repeated dosage when caffein was given daily or at longer intervals. The experiments were tried on rabbits and on dogs. As in the experiments on acute intoxication, the animals were under observation for some time in the laboratory before the administration of caffein was begun in order to ascertain the presence or absence of abnormality. The relation of diet to toxicity received some attention, but the question was not studied exhaustively in the present investigation.
Full-grown adult as well as young rabbits were employed. The diet consisted either of carrots or of oats; water was given ad libitum. The rabbits were kept in metal cages in a well-lighted and well-ventilated room. Unnecessary handling or any other procedure tending to fatigue or to cause discomfort to the animals was very carefully avoided, since we had found that such treatment was likely to decrease the resistance of the rabbit to caffein. The caffein was administered by feeding by mouth and through a stomach tube, or by the subcutaneous method. In a good many cases it was given daily, in some at longer intervals.
Series A.
The experiments of this series formed a preliminary study for the purpose of testing the effect of moderate doses. One decigram of caffein per kilo was given daily for several days; when administered at longer intervals the dose was increased to 150 to 200 mg per kilo. It was found that the smaller doses did not produce any symptoms; even the weights of the animals were not influenced. Doses of medium size given on two successive days were likewise without any noticeable effect (Nos. 182, 183, 123, 101). When a third dose of this size was given within 48 or 24 hours it proved fatal (Nos. 123, 182, and 183). Exceptionally, however, moderately large doses (for rabbits) may be given for three consecutive days without fatal issue, as in rabbit No. 101. When given at intervals of two to three days, larger doses, as may be seen from the protocols, can be administered without causing acute death (Nos. 173, 181, 201).
The results of the tests of this series point to the absence of any accumulation and to the possible elimination of moderate doses of caffein and its products of decomposition within 24 hours or thereabouts. When the doses are larger the time of its elimination is apparently longer, as shown by the fact that repetitions of the dose the next day may be fatal, but when a longer interval is allowed it may be given without causing death. It will be observed that only one rabbit of this series survived, but it was extremely emaciated. This condition has been observed in a number of cases after caffein had been given for several days. Even when the drug was withdrawn the animals continued to lose weight. This may be explained by the condition of the gastro-intestinal canal as found at autopsy. The presence of inflammation of the mucous membrane of the stomach and intestines, with ulceration of the mucous membrane of the pylorus in one of the rabbits (No. 173) of the series, in all probability caused diminution or loss of appetite, which of itself would tend to cause loss of flesh and strength and finally death. Protocols of the experiments follow.
Rabbit 173.Carrots were fed from October 1 to 18 and oats for the remainder of the experiment.
Date.Weight.Caffein per kilo.Date.Weight.Caffein per kilo.Grams.Gram.Grams.Gram.Oct. 91,9800.141Oct. 162,0050.220Oct. 111,905.190Oct. 181,845Oct. 131,930.207Oct. 201,740.230
October 21: Paralysis of posterior extremities.
October 22: 9 a. m., found dead.
The urine was examined before and after the administration of caffein. No symptoms were observed after the administration of caffein (5 doses in 11 days), nor was albumen or sugar found in the urine after any of the experiments on this rabbit.Autopsy: Pyloric mucosa exhibited several ulcers; small intestines showed slight inflammation; liver deeply congested; kidneys showed marked inflammation of cortex; other organs practically normal.
Rabbit 181.Diet, carrots September 29-October 17, then oats.
Date.Weight.Caffein per kilo.Date.Weight.Caffein per kilo.Grams.Gram.Grams.Gram.Oct. 41,4250.105Oct. 111,3700.175Oct. 51,450.100Oct. 131,385.180Oct. 61,270.100Oct. 161,345.200Oct. 71,210.100Oct. 171,030Oct. 81,375.130Oct. 181,230Oct. 91,305.153Oct. 201,215
Rabbit was markedly emaciated and weak. No albumen or sugar found in the urine as a result of caffein feeding.
Rabbit 182.Diet of carrots from September 29.
Received caffein subcutaneously as follows:
Date.Weight.Caffein per kilo.Date.Weight.Caffein per kilo.Grams.Gram.Grams.Gram.Oct. 41,7650.100Oct. 81,685.135Oct. 51,880.100Oct. 9.150Oct. 61,750.100Oct. 111,605.174Oct. 71,710.100
October 12: 11 a. m., 23 hours after caffein was given, convulsions with recovery; rabbit died at 1.30 p. m. No sugar was found in the urine at any time after the administration of caffein. Albumen was present only in one specimen.
Rabbit 183.Diet of carrots from September 29.
Received caffein subcutaneously as follows:
Date.Weight.Caffein per kilo.Date.Weight.Caffein per kilo.Grams.Gram.Grams.Gram.Oct. 41,3850.100Oct. 81,3100.153Oct. 51,460.100Oct. 91,390.142Oct. 61,385.100Oct. 111,390.187Oct. 71,240.122
October 12: 9 a. m., found dead. No albumen was found in the urine. Only one sample contained sugar.
Rabbit 123.White, female.Diet, oats.
Received caffein subcutaneously as follows:
Date.Weight.Caffein per kilo.Date.Weight.Caffein per kilo.Grams.Gram.Grams.Gram.Apr. 142,35042Apr. 202,126141Apr. 162,25090Apr. 211,965152Apr. 172,32586Apr. 221,876160
Rabbit died 30 minutes after last injection of caffein.Autopsy: Stomach exhibited marked inflammation of mucosa. Slight enteritis. Liver and kidneys were deeply congested and dark colored.
Rabbit No. 101, white male.Diet, oats.
Received caffein subcutaneously as follows:
Date.Weight.Caffein per kilo.Date.Weight.Caffein per kilo.Grams.Gram.Grams.Gram.Mar. 182,0250.100Mar. 241,815.166Mar. 191,970.100Mar. 251,830.185Mar. 202,009.100Mar. 261,710.176Mar. 221,855.100Mar. 291,734.219Mar. 231,738.114Apr. 11,606.224
April 5: Found dead.Autopsy: Marked inflammation of gastric mucosa. Considerable enteritis affecting the whole extent of the intestines; liver congested and friable; kidneys deeply congested in cortical and medullary portions; spleen congested, but of normal size; lungs and heart normal.
Four days, 0.1 per kilo; 10 doses in 14 days.
Rabbit 201.Diet of carrots begun October 1; October 19, oats.
Subcutaneous injections as follows:
Date.Weight.Caffein per kilo.Date.Weight.Caffein per kilo.Grams.Gram.Grams.Gram.Oct. 91,0000.150Oct. 161,065.225Oct. 111,015.180Oct. 18850Oct. 131,065.187Oct. 20890.111
Under observation six hours October 20; no symptoms.
October 23: Died; was much emaciated but did not show any symptoms; emaciation set in when caffein was withdrawn; urine never contained sugar or albumen; symptoms observed after second dose only.
Series B.
The question whether caffein is cumulative in the rabbit, suggested in the preceding experiments, was the subject of further investigation in Series B. Caffein was given by mouth or subcutaneously. Carrots formed the exclusive diet, a measured amount being given. The rabbits were kept under observation for two weeks, except Nos. 370 and 373, records of which were made only for four days before the administration of caffein was begun. Caffein was given by mouth in experiments of Groups I and III. Rabbits 292, 293, and 295 received daily 20 cc water by mouth for four days previous to the administration of caffein, while in the rabbits of Group II the caffein treatment was preceded by the injection of 0.8 per cent salt solution subcutaneously. The object in both cases was to ascertain whether or not the method of the administration of caffein has any influence on the animal, but observation made from day today failed to show any effect of such treatment. About 1 decigram of caffein per kilo was administered daily, with occasional intermissions. Later in the course of the experiment the doses were increased, 0.15 gram per kilo being the maximum dose given. Rabbit 293 died after the third dose with symptoms of typical caffein poisoning. The administration of the same dose of caffein was continued 10 days longer in Nos. 292 and 295. It was omitted on the seventh, fourteenth, and fifteenth days of the experiment. On the eighteenth day of the experiment the dose was increased to 150 mg per kilo and was repeated 2 days later. No. 295 was found dead the next day. No. 292 survived. Rabbits 313 and 315 may be considered together, as they were treated alike in every respect. The initial dose of 100 mg caffein per kilo was finally increased to 122 mg. After the twelfth dose the emaciation was well marked and the rabbits were very weak. No. 313 was found dead 2 days, and No. 315 3 days, after the last dose of caffein was given. It should be remarked in this connection that symptoms of caffein poisoning were never observed in these rabbits. Death was not due, therefore, primarily to caffein, but the rapid loss of flesh and strength observed during the last few days suggests that it was due to malnutrition apparently brought about by caffein.
The results obtained by subcutaneous injection of caffein are given in the table as Group II. The initial dose of 100 mg per kilo was injected daily. No. 298 died after the second dose. Nos. 223 and 296 received this amount daily for 6 days. An intermission of 2 days followed, at the end of which the same dose was given again. The next day it was increased to 150 mg per kilo, but no effect was observed; 48 hours later this dose was repeated. No. 223 was found dead, but its mate survived. Symptoms of acute caffein intoxication were not observed in any of these rabbits. It would seem, therefore, that caffein is not cumulative. This supposition, however, appears somewhat contradictory in view of the fact that out of the eight rabbits of this series six died, nor could any cause of death be ascribed other than caffein. Also the first results of Experiments 293, 370, and 373 might be considered as indicating that cumulation, though to a moderate extent, does take place, since in these cases reflexes developed after the drug was given for some time. But this view is contradicted by the results of Experiment 371, in which 150 mg per kilo given 5 days after the daily dosage of caffein was suspended likewise caused increased reflexes. Cumulation, therefore, does not account for the effects noted in the other rabbit. It will be observed that rabbit No. 370, as well as Nos. 371 and 373, had diarrhea for several days. It is quite possible that the weakened condition rendered the rabbits more sensitive to the action of the drug. This is made highly probable by the observations recorded in the experiments on acute intoxication with caffein in which death occurred after small doses. In such cases some pathological condition was often disclosed by the autopsy. The results of this series corroborate, therefore, those of Series A, and indicate again the absence of cumulative action. The results obtained are in all probability due to malnutrition and other conditions brought about by congestion of the viscera and consequent injury to the gastro-intestinal canal.
Table 16.—Chronic caffein intoxication of rabbits; Series B on cumulation.
Group I.Group II.Group III.Data.No. 292.No. 293.No. 295.No. 296.No. 223.No. 298.No. 315.No. 313.Diet (grams carrots in 2 days)1,0001,000975930905880355300Caffein administered (cc) and weight (grams):Mar. 51,4101,4701,0451,0401,070955770770Mar. 71,4151,3601,1401,0901,0951,000715690Mar. 91,3501,2701,0701,0001,0551,005655665Mar. 111,5051,4651,1901,2301,2851,250755760Mar. 161,5801,4601,2301,1651,1701,145730745Mar. 171,5151,4151,0801,0401,1151,105720685Mar. 191,5651,5701,2801,1951,2351,220710735Mar. 21776441,5851,5301,2651,1501,2151,260755700Mar. 22776(1)(1)(1)441,4401,3151,1751,1001,0451,150675635Mar. 23776(1)(1)(1)441,3351,1401,1101,1451,1901,230715700Mar. 2476(1)(1)(1)441,310(2)1,0901,1151,1701,250680650Mar. 2576(1)(1)(1)441,3751,0351,1251,2151,215695685Mar. 267641,2551,0951,1051,1551,150675695Mar. 27Mar. 28765.566441,3551,1151,1201,1601,155595685Mar. 2976665441,3851,1501,1551,165955695675Mar. 307666441,3301,0751,0351,095Dead.630610Mar. 317666441,3251,1701,1101,140690605Apr.(1)7666441,3351,0501,0501,120625620Apr. 27666441,3901,1251,0901,155695625Apr. 3..200...Apr. 41,3001,0051,1051,080585580Apr. 57666441,3851,0901,1301,090655630Apr. 697.57.581,260..1,0101,0501,110560530Apr. 7(3)Apr. 897.57.581,2601,0001.0901.965Dead.Apr. 9Survived.Dead.Surv.Dead.Dead.1On these days 5ccof salt solution was administered subcutaneously.2Dead Mar. 23.3Found dead 9 a. m.
1On these days 5ccof salt solution was administered subcutaneously.
2Dead Mar. 23.
3Found dead 9 a. m.
Table 17.—Chronic intoxication of rabbits, series B, Group IV, on cumulation.
RABBIT, 370.Date.Weight.Carrots.Water.Urine.Caffein by stomach.Symptoms.Grams.Grams.cc.Mg per kilo.Aug. 72,15545050280Aug. 82,03045025185Aug. 92,1052900275Aug. 102,09545030335Aug. 112,1054506536050Aug. 122,1254506522050Aug. 132,1203502526550Aug. 142,1704503527575Aug. 152,175350(?)20075Aug. 162,1703606525075Aug. 172,17531035170100Aug. 182,09518040285100Severe diarrhea.Aug. 192,120400(?)285125Do.Aug. 202,120400(?)310125Better.Aug. 212,12040070250125Do.Aug. 222,04040045265150Diarrhea bad.Aug. 232,03037035220150Diarrhea better.Aug. 241,95021540120150Do.Aug. 251,8851953560200Reflexes.Aug. 26Found dead at 9.RABBIT, 373.Aug. 72,24045050230Aug. 82,15015030300Aug. 92,1202050150Aug. 102,15045015245Aug. 112,195450528550Aug. 122,1604506532550Aug. 132,1203004519050Aug. 142,1954504026575Aug. 152,2153503520075Aug. 162,2053104522575Aug. 172,24040040265100Aug. 182,25535030320100Aug. 192,115185(?)170125Severe diarrhea.Aug. 202,11528035195125Diarrhea better.Aug. 212,05017575115125Slight diarrhea.Aug. 222,06018075130150Aug. 232,00520075125150Reflexes.Aug. 241,99020075150150Slight diarrhea.Aug. 251,95025555132175Severe diarrhea.Aug. 261,87020580140NoneDo.Aug. 271,83020050140do.Do.Aug. 281,95040025265do.Slight diarrhea.Aug. 291,8254000315do.Very weak and in poor conditionAug. 301,85010140do.Aug. 311,835RABBIT, 370.Aug. 72,24045050300Aug. 82,26045050225Aug. 92,310430(?)300Aug. 102,29545050305Aug. 112,3204505033550Aug. 122,2804507040050Aug. 132,3003507025550Aug. 142,2654255515475Aug. 152,2602504012575Aug. 162,29515570Lost75Aug. 172,18010570120100Severe diarrhea.Aug. 182,15012570100100Diarrhea better.Aug. 192,075210(?)192100Diarrhea severe.Aug. 202,07528070180100Do.Aug. 212,16526050225NoneDiarrhea better.Aug. 222,10540050275do.Aug. 232,0803000145do.Diarrhea severe.Aug. 242,10525015245do.Do.Aug. 252,05532010176150Reflexes.Aug. 262,04019075250150Died at 1 p. m., without having showed any symptoms other than reflexes.
Series C.
The subjects used in these experiments were rabbits of medium size and were apparently young or at any rate were not very old. The series was planned for the study of the possible effect of diet on the toxicity of caffein when given for some time, and therefore oats were substituted for carrots, which had been fed in the previous work, as already stated. Caffein was given by mouth in the usual way, in 1 per cent solution, 100 mg. per kilo daily. Fourteen rabbits were used for these tests. Their weights were recorded daily and observations made at frequent intervals during the day.
The only change noticed in all of the experiments of this series was progressive loss of weight which set in from 3 to 8 days after the administration of the drug was begun. The duration of life varied considerably. No. 382 died after the first dose. No. 389 lived 2 days, No. 386, 3 days, and No. 385, 5 days; No. 390 lived 7 days and No. 404 lived 20 days after the administration of caffein was begun. The duration of life in all the others was from 11 to 16 days. The findings at autopsy are interesting and suggestive as regards the possible explanation of the effects of repeated dosage of caffein. In eight of the rabbits there was involvement of the mucous membrane of the stomach or intestines or of both. Since the same condition of the gastro-intestinal canal was observed in previous experiments with caffein when injected subcutaneously, the mere passing of the tube into the stomach is obviously not the cause of this condition. The fatal outcome due is therefore, as was suggested above, to inanition brought about by the condition of the gastro-intestinal canal. Moreover parallel experiments carried out on rabbits in the same way with alcohol survived this treatment much longer. Obviously then the passing of the soft rubber catheter is not the cause of this condition of the gastro-intestinal canal nor the diet. Rabbits were fed oats exclusively for several months in this laboratory and thrived. The presence of pneumonia in the other rabbits of this series may be regarded as accidental, as it is inconceivable that one or two doses of caffein, as was the case in Nos. 382 and 389, could predispose the lungs to infection. The results of these experiments therefore are in harmony with those of the preceding two series, indicating that caffein does not accumulate in the body, and that the toxicity of caffein, whether of the single dose or of repeated doses is the same,on a diet of carrots or of oats. These results also show that caffein is much more toxic with repeated dosage. As stated in the historical part of this bulletin the same view was held by Gourewitch.28
Rabbit 386.Belgian female.
Given 1 cc of 1 per cent caffein for each 100 grams, through stomach tube.