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T H E JOURNAL OF IlVDUSTRIAL A N D ENGINEERING CHEMISTRY
hours of lassitude. A rabbit weighing about 2 kg. was fed 2 g. of the pure product in a bread paste and 2 4 hrs. later received a subcutaneous injection of 6 cc. of 1 2 . 5 per cent alcoholic solution of the drug. Beyond a sedative action, this high dosage of the benzyl succinate had no apparent effect. The animal was kept under observation for 2 wks. and developed no untoward symptoms. Feeding tests upon guinea pigs and rabbits have demonstrated the surprising fact t h a t a guinea pig may eat at least I O g. of berhzyl succinate per kilogram of b o d y weight without harm and t h a t rabbits may eat a t least 6 g. per kilogram of body weight and show no bad results. Comparative subcutaneous injections of the benzyl succinate and the benzyl benzoate show the former t o be less toxic t h a n t h e latter, and inasmuch as the benzyl group-which is recognized as the significant group-makes up a far greater portion of the molecule, the succinate should prove t o be a valuable therapeutic agent. By analogy, for any condition where benzyl benzoate is indicated, benzyl succinate should prove applicable, with the added advantages of ease of administration, safety of retention, freedom from nausea a n d after-intestinal disturbances, and greater benzyl strength. Many clinical cases are proving the t r u t h of the above statement daily. The writer has compiled a number of such cases which it is hoped will prove t h e basis of a future paper on the subject. ATROPINE SULFATE FROM DATURA STRAMONIUM1 By H. W. Rhodehamel and E. H. Stuart RESEARCH DEPARTMENT ELI LILLYS: Co., INDIANAPOLIS, INDIANA
Datura stramo?zium, commonly known as Jimson weed, grows in almost all parts of the United States and Europe. Atropine was first isolated from this plant in 1833 b y Geiger and Hesse,2 but hyoscyamine is t h e chief alkaloidal constituent. The latter is isomeric with atropine, and is converted into i t during the process of extraction and purification. The percentage of total alkaloids varies froin 0.15 t o 0.6 per cent in the dried plant, the variation depending mainly on the age and vigor of growth of the plant. The moisture content of the whole plant is from 75 t o 85 per cent. Assays made of the plant in flower showed the distribution of alkaloid t o be as follows: Stems below the first f o r k . , ....................... Stems above the first f o r k . . ....................... Seed pods without seeds.. ......................... Seeds Leaves
........................................... ..........................................
Per cent 0.054 0.069 0.054 0.45 0.414
The comparatively small amount of alkaloids occurring in Datura s t r a m o n i u m made their commercial extraction impracticable until the discovery in 1913 by J. U. Lloyd t h a t under certain conditions fuller’s earth would adsorb alkaloids from dilute solutions of 1 Presented before the Division of Medicinal Products Chemistry a t the 60th Meeting of the American Chemical Society, Chicago, Ill., September 6 to 10, 1920. 2 A n n . , 6 . 43; 6 , 44; 7 , 269
1701.
13,
NO.3
their salts. The entire green plant was ground, percolated with acidulated water, and t o the percolate Lloyd’s reagent was added. After drying, the Lloyd’s reagent contained b y assay from 2 t o 2 . 5 per cent alkaloidal material. I n this manner a considerable concentration of the alkaloids was effected. COMMERCIAL E X T R A C T I O N O F T H E ALKALOID
The stramonium plant was harvested frpm about the middle of July t o the first hard frost, which usually occurred during October. The entire plant, except the roots, was ground in the green state, packed in large wooden tanks, and percolated with water containing 0.2 per cent sulfuric acid and 0.5 per cent formaldehyde. Maceration was allowed t o continue 3 days, and the rate of percolation controlled so t h a t about 300 gal. each day were obtained from a 4000-gal. tank. Fifteen hundred gallons of the dilute atropine sulfate solution were treated a t one time with Lloyd’s reagent. The minimum amount of the latter necessary for maximum adsorption was controlled by assay of the percolate prior t o its addition. To insure adequate agitation, air was blown into the mixture for about 20 min. T t required about 12 t o 14 hrs. for the Lloyd’s reagent t o settle, after which the exhausted percolate was decanted, and the precipitate drained and thoroughly dried a t a temperature of about 50’ C. Percentage Alkaloidal Alkaloidal of Total Lloyd’s Percolate Material Material Alkaloidal Reagent Collected per 100 Cc. in Percolate Material Added Portion Gallons Grams Pounds Removed Pounds Taizk 1’ l . . . . . 1266 0.0123 1.30 10.75 60 1.34 0.0107 2 . .... 1500 11.08 60 1500 0.0075 0.94 7.77 45 3..... 0.0042 0.53 4.38 20 4 . . . . . 1500
TOTAL .....................
-
- -
4.11 33.98 Assay of extracted drug gave 0.07 per cent alkaloid.
185
Tank 22 l..... 2... .. 3..... 4.....
1600 900 1500 950
0.03676 0.0319 0,0028 . 0,0006
TOTAL ....................
4.91 2.40 0.35 0.05
-.
59.28 28.98 4.23 0.57
I
7.71 93.06 Assay of extracted drug gave 0.024 per cent alkaloid. T a n k 33 3.14 0.0251 25,99 l . . . . . 1500 1.79 1500 0.0143 2..... 14.81 0.0071 0.89 3 . . . . . 1500 7.35 0.23 1500 0.0018 1.36 4.....
-.
230 110, 16 3
-
359
150 90 45 12
-
TOTAL ..................... 6.05 50.01 297 Assay of extracted drug gave 0.054 per cent alkaloid. Average Percentage of S o . of Tanks Packed in Alkaloid Present 10 JLtb 40.69 11 August 69.03 September 58.89 8 7 October 39.02 Average for the year 51.9 per cent 1 Packed July 15 with 26 000 lbs. of green stramonium. Moisture 85 per cent. Assay o f dried sample gave 0.31 per cent alkaloidal material. From the assay the tank contained 12.09 Ibs. alkaloidal material, Moisture 2 Packed August 23 with 15,500 lbs. o f green stramonium. 85 per cent. Assay of dried sample gave 0.356 per cent alkaloidal material. From the assay the tank contained 8.28 lbs. alkaloidal material. a Packed October 4 with 26,000 Ibs. o f green stramonium. Moisture 85 per cent. Assay of dried sample gave 0.31 per cent alkaloidal material. From the assay the tank contained 12.09 lbs. alkaloidal material.
Results obtained on a great number of tanks would be of no especial interest. The percolation records of three tanks have therefore been selected t o show, first. the poor adsorption obtained when using young drug harvested in July; second, the best adsorption from August drug; and third, the decrease in adsorption in October drug.
T H E J O U R N A L O F I i Q D U S T R L A L A N D E N G I N E E RI iVG C H E M I S T R Y
Mar.. 1921
D E T E R MI N A T I 0 P; 0 F ADS 0 R B E D A L X A L O I D
The adsorbed alkaloidal material was determined as follows: Two grams of the Lloyd’s reagent containing the aIkaloid were placed in a glass-stoppered bottle, and 40 cc. ether and 3 t o 5 cc. 16 per cent ammonia water were added. The contents were thoroughly agitated, and after settling the ether was decanted. The ether extraction was repeated six times, and the extracts combined, filtered, and evaporated t o dryness. Ten cc. 0.05 N sulfuric acid were added, and the excess acid titrated with 0.02 N sodium hydroxide solution, using cochineal as the indicator. Determination of the alkaloid remaining in the Lloyd’s reagent after extraction was made in the same way, except t h a t a larger sample was used. The reagent after extraction always contained about 0.2 per cent alkaloidal material which could not be economically removed. Percentage of Total Lloyd’s Alkaloidal Alkaloidal Material Material in Alkaloidal Reagent Added per 100 Cc. Percolate Material Portion Grams Pounds Removed Pounds Barrel 11 l . . . . . 36 0.01721 0.0517 41.87 2.6 2 . . . . . 36 0.0085 0.0255 20.68 1.3 3 . . . . . 36 0.0046 0.0138 __ 11.18 0.7 TOTAL 0.0910 73.73 4.6 Assay of exhausted drug gave 0.02 per cent alkaloid. 4 6 lbs. Lloyd’s reagent recovered. Assay 1.99 per cent. 74.09 per cent of the alkaloidal material was removed by the Lloyd’s reagent. Barye1 21 0.0401 32.50 2.0 l..... 36 0.01336 1.3 0.0261 21.16 2..... 36 0.0087 0.9 0.0180 __ 14.60 3...,. 36 0.0060 TOTAL 0.0842 68.26 4.2 .%say of exhausted drug gave 0.048 per cent alkaloid. 4.2 lbs. Lloyd’s reagent recovered. Assay 1.97 per cent. 67.00 per cent of the alkaloidal znaterial was removed by the Lloyd’s reagent. Bawel 3 3 l..... 36 0.0148 0.0445 36. OS 2.25 2..... 36 0.0077 0.0231 18.73 1.15 __ 6.81 0.4 3..... 36 0.0028 0.0084 TOTAL.. 0.0760 61.59 3.8 Assay of exhausted drug gave 0.036 per cent alkaloid. 3.9 Ibs. Lloyd’s reagent recovered. Assay 2.05 per cent. 64.74 per cent of the alkaloidal material was removed by the Lloyd’s reagent. Barrel 44 36 0.0127 0.0381 30.89 1.9 I..... 0.0048 0.72 2 . . . . . 36 0.0144 11.68 3..... 36 __ 8.27 0.80 E 2 0.0034 TOTAL 0.0672 50.84 3.12 Assay of exhausted drug gave 0.032 per cent alkaloid. 3.25 lbs. Lloyd’s reagent recovered. Assay 1.97 per cent. 51.84 per cent of the alkaloidal material was removed by the Lloyd’s reagent. 1 T h e menstruum was water containing 2 5 g. of bleaching powder and 10 cc. of sulfuric acid per gallon. 9 The menstruum was water containing 8 ccI of cresol and 7 cc. of sulfuric acid per gallon. 3 The menstruum was water containing 30 cc. of 40 per cent solution of sodium bisulfite and 10 cc. sulfuric acid per gallon. 4 The menstruum was water containing 18 cc. formaldehyde solution and 7 cc. sulfuric acid per gallon. Percolate Gallons
...................
...................
.................
-
-
-
-
...................
Differences in age of t h e plant and condition in growth caused some variations in the extraction of the alkaloidal material. The plants harvested in July were about 2 f t . high and just beginning t o flower. The alkaloidal content of the young plants was 0.1 per cent higher t h a n those gathered in August, and considering this, the July plants should have yielded more atropine per pound of drug than did the August plants. This, however, was not the case. Taking into consideration the amount of alkaloid obtained and the amount actually present in t h e plant, the average yield for July was 28.5 per cent lower t h a n t h a t obtained in August. Since the drug after extraction showed only negligible amounts of alkaloid, i t was assumed t h a t a considerable proportion of
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alkaloid was decomposed during percolation. Examination of the exhausted drug showed no evidence of decomposition resulting from bacteria or molds. Possibly the loss was due t o a n enzymic action, although this point was not gone into. X considerable number of small experiments were made with the attempt t o increase the yield of alkaloid. A menstruum containing acetic hydrochloric or sulfuric acid in various concentrations gave no better results and showed no marked difference. Somewhat better results were obtained, however, when other preservatives were substituted for the formaldehyde. Results on four of these experiments are given below. With the usual formaldehyde preservative 50.84 per cent of the total alkaloidal material was extracted; with sodium bisulfite as the preservative 61.59 per cent; with cresol 68.26 per cent; with bleaching powder 73.73 per cent. A number of barrels were packed September 20 with green stramonium, each containing 260 Ibs. Moisture 80.8 per cent. Assay of dried sample gave 0.24 per cent alkaloidal material. Accordingly, the drug in each barrel contained 0.1236 lb. of alkaloidal material. Percolated a t the rate of 12 gal. per day. EXTRACTION
O F T H E A L K A L O I D F R O M LLOYD’S R E A G E N T
I n a general way the alkaloidal material was obtained from the Lloyd’s reagent by first adding water, then making the mixture alkaline, and subsequently extracting with an organic solvent. I t was very essential t o make the mixture alkaline with a base t h a t would not decompose the alkaloid. Ammonia and lime water were found t o be the best for this purpose. Three methods were used for the extraction. First, ammonia water and ether; second, lime water and ether; and third, lime and alcohol. The extractions were made in a revolving drum. METHOD 1-To 300 lbs. of Lloyd’s reagent containing the alkaloid, 35 gal. of 16 per cent ammonia were added. This gave a rather thick paste, and on revolving the drum t h e material was carried part way up the sides, then fell back, and in this way was thoroughly mixed with t h e ether. Each extraction was continued for 45 min. The results were as follows: Extraction 1. . . . . . . . . . 2.......... 3. . . . . . . . . . 4
.........................
5 ......................... 6 ......................... 7 ......................... 8 ......................... 9 ......................... 10 .........................
Ether Used Gal. 200 130 120 120 120 150 140 120 120 120
Alkaloidal Material Removed 02.
13.5 7.1 9.4
8.2 5.3 8.0 6.1 4.9 4.1 3.8
An assay of the extracted Lloyd’s reagent gave 0.45 per cent alkaloid, showing t h a t 22.5 per cent of the alkaloidal material still remained in it. N E T H O D 2-Same as Method 1. except t h a t in place of 16 per cent NH3 40 lbs. of CaO in 35 gal. of water were added. An assay of the extracted Lloyd’s reagent gave 0.29 per cent alkaloid, showing t h a t 13 per cent of the alkaloidal material still remained in it.
T H E J O U R N A L OF I N D U S T R I A L A N D ENGINEERING C H E M I S T R Y
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Extraction 1 .2 3. 4 5
........................ ........................ ....................... ........................ ........................ 6 ........................ 7 ........................ 8 ........................ 9 . . ...................... 10 ........................
Alkaloidal Material
Ether Used Gal. 160 140 145 160 150 150 150 150 150 150
Removed 02.
14.22 9.41 8.62
5 7.66 .80 5.80 5.44 2.89 5.33 4.41
METHOD %-To 300 lbs. Lloyd’s reagent (assay 2.36 per cent) 40 lbs. CaO were added and the mixture extracted for about 3 hrs. with t h e following volumes of 80 per cent alcohol: 80 Per cent Alcohol Used
Gal.
Alkaloidal Material Removed 02.
45.44 17.61 13.68 8.92 7.42
An assay of the extracted Lloyd’s reagent gave 0.18 per cent alkaloid, showing t h a t 7.6 per cent of the alkaloidal material still remained in it. T h e following is of interest in connection with the alcohol extraction of the alkaloidal material from Lloyd’s reagent. A mixture of 10 g. of Lloyd’s reagent with 1.3 g. of slaked lime was extracted for 14 hrs. with 50 cc. of alcohol of the following strengths: Strength of Alcohol Per cent 100 90 80 70 60
50
T h e volume of follows:
80 per cent alcohol was varied as
Alcohol Used cc. 20 30 40 ..
50
80
100
Alkaloidal Material Removed Per cent 0.00 25.42 41.25 41.25 33.82 23.50
Alkaloidal Material Removed Per cent 26.8 32.6 35.0 41.25 43.6 47.0
PURIFICATION OF THE CRUDE ATROPINE
The alkaloidal material was extracted from the Lloyd’s reagent with 95 per cent alcohol, using lime t o obtain the proper alkalinity. The extractions were acidulated with acetic acid and the solution concentrated first t o 12 per cent, and then under diminished pressure t o 2 per cent of its original volume. This procedure was sufficient t o convert all the hyoscyamine into atropine. After neutralization with ammonia, the solution was allowed t o stand over night and filtered. A test portion of the filtrate was shaken with ether. If a n emulsion resulted, t h e solution was ‘diluted about one-fourth and returned t o the vacuum still. Distilling the neutral liquid, and again filtering, usually prevented the troublesome emulsion with ether. Ammonia was added until the solution was alkaline and the atropine alkaloid extracted with ether. After evaporation of the ether, t h e alkaloid was carefully dried a t about 3 5 ” C. The dried alkaloid was dissolved in ethyl alcohol in t h e proportion of one ounce of alkaloid t o two fluid ounces of solvent, and the solution almost neutralized
Vol. 13, No. 3
with sulfuric acid, using cochineal as indicator. After filtering i t was evaporated on the water bath t o a thin sirup, and t o this sirup, while still warm, acetone was added almost t o the point of precipitation of t h e atropine sulfate. On cooling, the atropine sulfate crystallized. If not sufficiently pure the crystals were dissolved in alcohol and recrystallized as outlined above. The acetone was evaporated from the mother liquor, and the alcoholic solution of atropine sulfate poured into a large volume of water. From this the alkaloid was extracted with ether, and if not of sufficient purity t h e process already outlined was repeated. AN INVESTIGATION OF THE U. S. P. ASSAY FOR PHOSPHORIC ACID AND SOLUBLE PHOSPHATES‘ By A. E. Steam, H. V. Farr and N. P. Knowlton MALLINCKRODT CHEMICAL WORKS, ST. LOUIS, MISSOURI
I n routine analysis of samples of phosphoric acid in this laboratory, it was noted t h a t , although aliquots from the same solution when assayed according t o the directions given in the U. S. Pharmacopeia2 gave concordant checks, i t was difficult t o obtain check results when two different samples were weighed out and made u p t o volume, unless the size of t h e sample happened t o be nearly the same in both cases. Briefly, the method is t o transform the acid t o t h e disodium salt b y neutralizing with NaOH t o a phenolphthalein end-point, precipitate with a n excess of standard silver nitrate solution, bring the solution t o neutrality t o litmus with ZnO, and determine the excess AgN03. Calculations were made of the error introduced by the actual volume occupied by the precipitate. This error, assuming a specific gravity of I for t h e precipit a t e (the value is given as between 7 and 8 ) , adsorption of water t o the extent of one mole per mole of phosphate, the presence of the equivalent of 50 cc. 0.1N salt, and an equal volume occupied by the excess of ZnO added, was shown t o have a maximum possible value of 0.5 per cent, and more reasonable assumptions reduced this error t o 0 . 0 8 per cent on a 90 per cent sample. This small error by no means explained t h e large discrepancies of 5 t o I O per cent met with a t times. A few preliminary experiments seemed t o indicate t h a t the results were influenced very markedly by the size of t h e sample. T h e larger the am0 sample, the lower were t h e results obtained. I n the filtrates, after the Ag3P04 had been filtered off, a test with ammonium molybdate showed the presence of significant quantities of phosphate which t h e silver, though present in considerable excess, had failed t o carry down. The importance of this particular method may be realized when we recall t h a t i t forms the basis for t h e assay not only of phosphoric acid and t h e alkali phosphates, but also of many hypophosphites, such a s those of Ca, Na, K, Mn, “4, etc. The method is not confined t o t h e U. S. P., but is found in t h e N. F. and even in the “New and Non-official Remedies.” 1 Presented before the Division of Medicinal Products Chemistry, a t the 60th Meeting of the American Chemical Society, Chicago, Ill., S e p tember 6 to 10, 1920. 2 Ninth Revision, p. 21.
