South Dakota State University Open PRAIRIE: Open Public Research Access Institutional Repository and Information Exchange

Electronic Theses and Dissertations

1927

A Study of Atropa Belladonna, (Linne) Grown in South Dakota

Floyd J. LeBlanc

Follow this and additional works at: https://openprairie.sdstate.edu/etd

Part of the Pharmacy and Pharmaceutical Sciences Commons

Recommended Citation LeBlanc, Floyd J., "A Study of Atropa Belladonna, (Linne) Grown in South Dakota" (1927). Electronic Theses and Dissertations. 5656. https://openprairie.sdstate.edu/etd/5656

This Thesis - Open Access is brought to you for free and open access by Open PRAIRIE: Open Public Research Access Institutional Repository and Information Exchange. It has been accepted for inclusion in Electronic Theses and Dissertations by an authorized administrator of Open PRAIRIE: Open Public Research Access Institutional Repository and Information Exchange. For more information, please contact [email protected]. A STUDY O ATROPA BE�LADON A, (Linne) GRO\lN IN SOUTH DAKOTA

By

FLOYD J. B. S. 1924

A Thesis Submitted to the Faculty of e South Dakota State College of gricu ture and echanic ts June 192? In P.artia Fulfi e oft e Req irements or t e Degree of ster of cience In Phar1acy

COll GE rs OA O'TA AT u;;i])ICI /\� G RD�N

DI JISIQJt 0..., P.' IU.:ACY SO ¥TH D�KOTA T�TE CL 'G

Tio It!jC

'" The econo ic im ortc ce of r nn l lon b eco nized beca �e or the valuable al -�101d i C n le nc . eloni;. to he 01· f hie is C r:1... o...,ed or a ut 1 -oo

,c-. c.:. •- I man,. - ..._h f aleo e.:..n of economic i ... ort nee. t tr ca .em., of A :!�ic .1 :ure in K an ...en r Li e

.l L 9 6 t\ consta ..1 eviat� ns from the no varie .,, of Atro a el c:.d nn L. h re been gr·o m. The� re tr �c �ell�dunna v r. fl va and tr a B_l ad nn var. in er l t of the norm vari t

I' part..., the C :, e _l.... _o_ nna ) n in ot r o:.: nt ... n OU tedl� 1 a been tl orou hl.,r tudi d in o:: the _i i be pr n e h r . 1 e or C a. elo c.,Il 1a e a_l rec i.ed con- .a.. e ti n ir p e� en t c.� a i era r • f r C o en t .., n in

n

c n en of n·e r le

0 i r o f.. ., both of ich contai n alkaloids. Then, too, t e met od of extraction o t e l raloids ra not consi d rdas accurate a it s uld be, and ith thi s t ou ht in mind var·ous alkaloi dal olvents and met ods re-e tried. One of t e c ief dif iculti es in tle extracti on of t e alkaloids and in the obtaini ng of t em in a pure cond·tion is t e chlorophyll iliic· is alra· present in the leaves. Several methods for teeli mination of t is im uri ty Yere also atte pted. To make tis 1ork more complete and to bring out the structura c ract ri ti cs o Belladonna more in detai l a n ber of photogr�phs and micro-photograp s ·ere ta en. The technique o� taking micro-photo raphs as vell as the developing and printin of the plates found in tris thesis required con idera le ime to :or out. The tropa Belladonna experi mented 1ith 1as ro min t e dicinal d Poi son us Plant Garden of t e Di i �on of 1 r c , out D �ota tate College. HISTORY Bellad onna , more commonly ca lled Dea dly or Sleeping ightshade, is a perennial herb found na tive in the ,rnods of centra l and southern Europe, sia Minor, and Persia. It is largely cultivated in �ngland and Germany and at the present time Bel ladonna is grovm in various parts of the United States verJ succes sfully. Other common syno­ n�rms for Bell&donna are: Dea th's Herb, Banewort, Mekil­ ·iort, Poison Black Cherry, Da ft-(Dway-)berry, and Dwal·e. The derivation of tropa Belladonna is as follows: A tropa - L. fr. Gr. A 1'" Po 1't' o s ,. .!_ , priv. , -r Pt. rre: z. r , to turn, not to turn, to be inflexible, one of tne yt olog­ ical fates, 7hose office it was to cut the thread of life. Belladonna - L. bella, bea utiful, plus donna , a lady. The berrie 0 vere formerl used b r Italian la dies a_ a cosmetic and to dila te t·_e pu..9ils of t eir e... - es, t us ivin t:iem a striki.gl andsome appear·nce. rom kno ·led 6e hat 1e are able t gain from the arlie ....t of ,r j_ tin 1e kno r tl at Belladonna · a no, to t e ancient Its ham is in a ore or le s re trict d area of ce tral urope. It did no mi ra te as man ot er s Jee of t· ...e olanac e fa:c: il T . Be ad n. a d t prouerties of Bell donna fere probab_ kno 1n to e early ed ·cine men. he kne� tat i oul cure certain di e �es, alleriate pain, ad even ca e de<-i, en ive -3- in large amo unts. This knowledge \Yas of course made use of to bev.,ilder and hold in avre the mass of people. Wha t may have been Belladonna Q,ppears many times in the early stories of the bards, told and sung for generations before we have any written records. Then slowly there appeared written records but these are generally the old stories and myths hanued do wn from centuries before. Various species of t e Solanaceae fa m­ ily are mentioned giving their u0 es at th at time. Bella­ donna, alth ougl not kno ·m as Belladonna in early times 1 seems to be included in the list. Hippo crates, the Father of edicine, while introducing no neu drugs recorded much t.iat had gone before him. Drugs of the Solanaceae family are mentioned in his writings. Aristotle grev the solanums in his garden. Dioscorides .rites of several specie of solanums iving descriptions and uses. Plin � tell� of t ree species of solanums, one probab ....y being t e Bella­ donn of toda . Galen, too deQcri be tree pecies of solan s. A little later :e find medicine pract·ced by t e men s 1ith little progress made. Then in 1438 e ave t .e in ention of printin and soon after t e Herbals ere printed and bee e popu ar. The name Be lladonna doe n t appear but ram t e de script�ons and use given for s veral pec·e s of t_e �olanums it a er· ently 1 o m b o er na e . o from ti brie outline 7e see -4- that Belladonna a ..., vie know it was also known to the ancients although it was not used by them to any great extent. In ab out 1504 Belladonna ca�e into use as a cosmetic and for poisoning -urposes. Because of the slow course of action and t e ob cure symptoms it was a favorite with professional poisoners of the �iddle Ages. Belladonna received little medical notice until 1860 when it was introduced a_ an anodyne. The two chief alkaloids of Belladonna are atroDine and hyoscyamine. A small a.mount of sco:polamine is also found. Belladonnine, an alkaloid devoid of mydria�ic porer, is stated to occur in small amcunts. Atropine a� fir�t obtained from tle root of Datura tra.1-onium in 1831 by ein. Geiger and Hesse discovered atro�ine al- ost s·multaneou0 l7 :i� ein, but from the root of Belladonna. T eir material "1as anal... zed by Liebig ,J ...o assigned to it t e empirical formula no in use, C17H23-O3. The ease o tran for a ion o o c mine into atrop�ne had c �ed inve t· atoro to onder if tropine existed pr - f rmed in pl t . ill (Pictet d Biddle 16) believes � e 1 n t co mo t vosc�amine. He y (21) s, sa It iz probable t at thi a aloid (atro�ine) never n occur in 0 _an races in so lanae eou..,; plants. The in se e al chemi a d �n a:?:"ticu ar C ana C u·te ( ic et a d Biddle 16) eem to prove otherwise and it is a generally accepted fact now that atropine and hyoscyarnine both exist in the plant, hyoscya­ mine occuring more �bundantly in the older parts. Upon extraction of the alkaloids the hyos cyamine pres ent in the plant is at least partly changed to atropine by con­ tact with the alkalies. Atropine is the tropeic acid es ter of a ba�e, tropin, and is is omeric with hyoscyamin; hyoscyamin being levoro­ tory, while atropine is racemic. Commercially atropine is obtained by treating hyos­ c amine extracted from plants vdth a dilute alkali. It undergoes isomerisation to atropine. Atropine ic hydro ly zed when treated with dilute acids or alkalies and even 1it_ water. Tropine and tro ic acid are formed a follo7s: c17H23�o3 plus HOH plu� CgH1o03. The fact t1at t e hydrolysis of atropine produces tro ine ic po se se a coholic functions ad tropic

cid c ows tat t e alkaloi it elf mu t be an e ter. T follo •·ing formula repre entEi a tr opine: CH2 -- C C 2 C6H5 :c I 3 fH--0--CO --t- CH2--CH--CH2 C 20H uc ere · t for est bli hing t.e con titution of tro ine . d 0 C ine is due Ladenbur , erling, and -6- illstatter. Atropine has been made synthetically by ill£tatter. '7ills ta tter 's Synthesis of tropine is given

fillstatter's Synthesi� of Tropine

0 0 0 o\ II II II I I CH C-CH 2 2 -c- -ca-o-c CH2- C �-OH I I OH CH2 Ca( OH)?> I CH2----CH2 CH2 CH2 Sube ic Acid

Heat

:2·-- -c CH2 II I .. OH CHI 2 CH __ nH 2 � �2-- -2 Suber loxime Suber one Reduce I with H do en ,J,

}.

C 2-C-- CH2 CH2--cH����cH I l stive ffi2 CH2 atio� bH 2 I I c-...2 - C 2--CH2 C 2 C 2 --c::2 Sb� lamine C clo- e_pte e

-7- l (1) Alk. KOH CH -- CHBr- -CHBr CH2--yrH -- C11 H 2 I CH3/ CH3 CH teus CH2 I < I CH CH2-- CH2--cH2 (2) (CH3)2NH CL2--CH2--- 2 Dimethyl!mine cycloheptene Cycloheptene dibromide Exhaustive ethylation Br t I CH CH CH CH-- H--CH • II CH Bri2 CHBr � I r,t T Thiele Reacti n � v C1!2 H2 2 CH2 -- CH2 -- CH2 .L. C... o- e � ine

Q,uinoline Reduce ---· ===-=- CH CH CH 2 CHI CHI 2 +HBr CH ) Br JH r II j 11 CH E---C 2--C CY C Cycle Heptatri ne Structure turned (Tropili ene) (Rotated) I (CH3)21-H t 1� • KOH

---,CH---

la ·noc o epte e

-8- CH <;rn CH CH2--iCH---CH2 2 I 2 CH3 I ( CE:3 / '-CH3 CH2 Bromine '------N-Br CH2 Br I •igrates to CH3/ I CH2 CH CHBr Nitrogen CH2--CH--- CHBr Bromotropanemethyl­ a:rnmonium Bromide 1-HBr_ CH --CH CH 2CH I . I 2 < r CH 3 I CH2-- CH ·cH Tro:pidine Methyl Bromide

CH2 CI H C! H2 CH3-N HO-C-H CHI b CH2 H2 Tropine This product did not respond to actions of Tropine (OH) on ·wrong side of carbon --spacial con:'i ·uration.

H2--1H--�m2 CH2--yH---1H2 H - C O Zn+HC.1. CH3- H--C-OH 3 I I ) I I CH2 -- CH -- CH2 CH2-- CH H2 ropinone Tropine yn esis of Trop ic Acid La e_ bur and ug imer from cetophenone

C6H5 /OC2H5 / / v Cl CH3 C o e ne -3 V C H _;;/ 1 0 C H5 1 6 5 C6H5 JC2H5 6 / / (�· Cl C- C # ( �6i. / CH OI HO-CH2 OOH 2 CH3 COOH Atropic Acid Ethyl Phenyllactic l Acid

H Tropic cid Lade bur in 1883 performed the lat step br con- de ing t e acet,rl derivetive of Tropic Acid Chloride ·ath Tropine by eans of dilute HCl.

/C6H5 !I V > HOOC -C -H H20: -o CH3 -CO-CH3 Tropic Ac d �C!l5 0 / c II 6 5 Cl C -C--H CH2-- 0 - -CH3 II

cet�1 deri tive of Tropic C.u C lo . de

-10- Hydrolysi s

-11- ROOT STRUCTURE The Belladonna grovm in South Dakota, has a large, fleshy, cylindrical, tapering root, .5 to 2 cm. in thicknes.J, and 8 to 12 inches or more in length, from which a few branches arise. Externally the roots are br owni.... h -nhi te, internally creamy-white. A cross-section of the root sho 1s a structure similar to t at of the stem wit h the exception that we have in the stem more lignified tissue. The ma in root also contains a arge vascular strand which, of course, is not found in t e stem but in its place a distinct pith. In the root is found first a la-er of cork cell�. Immediately beneath the cork cells occur a large number of parench atous cells, the major ity of them being filled dth starch grains, both simple and compound. However, a fe contain sp enoidal micro.. cr ....tals of calcium OJ(..alate. Ju t above the ca b�um la er i� found a mal l amount of sie e t·ssue. Belo t e cambi la r numerous fibro- va cul bundles cont,inin tr cleae ur rounded b some ood f"ber occur . The m dullar ra s are d�otinct, con- i of 1 '-"eou a enc tou ... ce ls. e center of

.C'. e sec ...r b ndle s . e ir ...., rear r 0 a lo .ed 0 gro nor.a ly, the .&'. r ar ro 0 a fer 1·zed lot, t root ...ra m on .:.c no fl er Q,d bee allo ed 0 develop, -12- and t e root from )1 nt c f a eecond year s rov·t � ov:ed

ractical y the rone struct e. Ho 1ever-, the root from a plan of the ec nd er' s better eveloped o fibers.

ST TRU.J.1.URE

T ... e structure of the s em for te o�t art agree

.:.tl that of the root, exce_ t t at in the E. te� 'I fino mo_e s portin tissue nd a pit c __,m:posec of 1 r e irre u l" ren c'_ ce_ls.

Th te C .L.r cture proper begin.,;; i.L.h an 0 er i h .L er V a i cu ie f llo ed b .. a 0 ex 0 arenc cells nc: an c:nd odermis ich t be a filled ro s st rcn gr" .:.ns. Ben ath l i e e r1:.:: f nd i ! t ... n llf a d onl ....li li i d. - nd a fe:; 1 le t ....e . 1 con .... i t .._ 0 us tr C e 00 ib r inn e seve 1 b s er . C n 0 e 0 1 r ·e r p n C 11 n one V .

av en re br adl o a e d t a e e b ro 7 C in

0 . in · e ur c ce 00

G the margins entire, and the petioles short. On the under side may be seen, chiefly along the veins, a fe1.1 hairs con­ taining tro or more eel s. The upper surface of the leaf is dark greenish wbile the lower surface is more of a gray­ ish green. Stomata are found on both the upper and lower surfaces but more nmerous u�on the lower surface . A number of whitish sp ots also appear on the under side of the lea·--·, locating t e cells containing calcium oxalate. The veins of the first order diverge from the midrib at an angle of about 45 degrees. In a cross ·section of Bellad onna leaf re find the epidermis r.Lt. a slig tly t ic_�ened outer cutinized wall. The sto ata are el ongated and surrounded by guard or neighboring cells. Immediatelr belo� t· e epidermis on e upper side onl: is found one layer of p lisade cel_s filled vit chloro�hyll grains. Below t' e palisade cells i found t e meso_hyll parenc yma in ich are cells con­ tain�n t e ine sand er� tals o calc�um oxalate.

-1 - I

Portion o cros section of root shorin umerou t rch grains in pa.re c yma t, t rc1; Pr, Pcrenchyma

PL II

Po rtio OI eras e tio- t rougl ro ot C , CO r - C el l 6 ; t, re ; r, P nc

ood r r, r C

"ddle .... tio Tr, che orti no on itu�i 1 � cti n tr g de elo 0 r, e . J ar, P no ------TR

------PAR

/ PL V

ortio o c oss ec•io o st i C 1, i r C d e -- s

-- C.

--TR.

I PAR. PL I orti o l ngit din 1 ection _rou ste i ; Co C llenc r n i V r, r chea

PLJ VI I orti no� cros sec tio n t ro i rib O- 1 f e is; Pr, _, Tr che e ------EP

--TR

- - -s CULTUR L D LOP · T The seed from whic t e Atropa Bella onna (1926) grown were coll cted in the edicinal d Poi onou Plant Garden in the f 11 of 1925. The maj ori t-, of experts on Bell donna culture no a ree tat Bell· donna, to be successful sh ould be started in the green house. On February 10, 1926 to flats of Be_ladonna seed �ere so n. The seed had not been selected or treated in any ray to hasten germination. The soil u ed .a a bl ck dirt mixed rith a little sa d . The seeds e e covered to a dept of ab out 1/8 of an inch d 1atered frequently. glas as placed over the top of t flat o ind uce sveat�n . Ger ination a rat er sporadic, t e first plant appearin in seventeen da and in t etree da follo ri et e fir t germinat·ons both fl t developed fa ir stan d. However, not over 10 to 15 p rcent o� t e seed r ·nated. Tepla_ t er allo ed to r in ·n t e 1 or inal fl t until rch 30 'I. en t ey r'iere trans nt d but till kept in the green house. Due to t e late er of frost t e Belladon a v-a ot tran s 1 d e garde until 13. Lig ...t frost occu red a ter 13 but t e Bell onna id not see o be injured in n b t m. e 1926 B lladonna ra ro _ on ive differ 1 s . o lo . umb er 1, 1 o e d to ro 1 nor c...ll · umber 2, -15- grown on plot fe t · lized vi th aN0 3 four hundred pounds to t e acre; number 3, plants of second year growth ; n ber _ , pla t grown in shade; and n ber 5, plants allo ed to gro nor ma 1 excep t that tl flo -., buds le1 e pie ed off as soon a� they appear d. Al l of the Belladonna plants set out in t e garden developed lo ly at i st, due to adve rse weat er con­ dit·on , t e spring ·nd early summer being very dr- . Later, howeve r, cond · �ons vere mor fa ar able an d the plant s developed rapidly. Th e Be la donna on lot nun ber l de e o e normally in all res ect . Plot number 2, ich as e rtilized, developed fa t rad obtain d a lar er gr�,t . Th leaves ere muc la rge tan t ose gro or - al lJ ad th e root�, tables atta h d , t - tain ed a av ra0e ro t of t ree time tat of t root 0 t plants g ro m norma lly. Th e plants of' se co d year's gro7th on plot number 3 de lo ed r pidl ad attained li htly larg r i et n o e of fir t y ar. Teleav e e 1 r er and t e oo , a per ttac ed t�ble� , er ed rem five to ix i 1 i t o t o e fro a fir t ear gro • (In o rder o obtain p ants of a second r s ro-1 an ber 0 -l:' pla nt,.. of a i st ea ' s gro th tr·mm ed of fa ia e t d cro cove ed Ii t dan ra s. 0 er pla 0 te e plot ere du na. to e in t e cold fr e . -16- In the spring the gra0s was removed from the plants allowed to remain ou ts ide but onl:r one or two survived the free zing winter weather. The plants kep t in the co ld frame vere transp lanted in the spring and started a vigorous growth. On plot number 4 pla nts grovm in the shad e did not develop near as rapidly as any of the others. The foliage wa s decided ly th�nner and the root development smaller th an roots on anv other p lot. Plot number 5 contained tr e plants fro. rhich flower buds were picked as soon as they appeared. In ievers (2 6) stud y of the Daturas he found the al_ aloidal content to be �nc rea ed . The le aves formed ,ere larg er and tie roots atta ined an average size of tl ree times th�t found on plants al lowed to develop normally. In pursuing t e li�eratur available on Belladonna it ha be en ::ound tha t different i ves tigat ors vary �idely on Bell adonna culture. ome claim Belladonna gro s best in tj1 e shade; ot ers claim t� direct ray 0 t e sun are neces sar, for full development. ome recommend tae use of one ertilizer and ome anot e�. fe fi nd a hig .er a aloidal conte- t in Belladonna gro in Les had e, vl ile ot ers find Bel l adonna ro n in e s un os esses a igher perce t of al loids . H. D l uard (3�) fi nds Belladonna grown i � e sun yields or al� loids . H. St ouffer (35) as Be llad na r -1? - in partially shaded beds i ric er in al loid� than th t gro m in sun. ome invec tig ators find tat t e cl im ate and weat er a ect al .. loi dal onte nt I il o t er say tat sea onal co ndition ma ..e only a small var · ation. ccording to ill er and ed ( 33) , r. o rev iewed re ort on rican Belladon ·�, t.e al alo·dal value o t e dr ug may range from zero percent to 1. 32 perce tin t e leaf and from .1 to . 86 perce tint e ro ot. A. F. ievers (2 ?), Bu re au of Pant I ndustry , U. epar ent of ric- ltu re ha� done muc JO rk on B lla- dn nna and t po s· bi�ity of incr t e alKaloidal co nte nt. The follo- -i g paragraph is ta en .... rom one of io articles . " uch ha been said ad 1ritten re ard·ng t in- oil adcl · m te on t rodu t· o c a loids a d t. e cone u �on re c d ave va ried gr t-Y · In vie + of t gr a variation e_ isti in indiv idual lants as 0 he rom und e i e ti cal con ·tion� a reg ds o i 1 a. d c 1 · ,. te , it mu t be p 1 in at li tl i an i - por ta c can b a ta ed to ex rime t to o 1 e in- flu ce of the � acto s u t·1 at e oi la t c· . b secured Olb t e m� nimtLm amo u to ind i i ual

0 ·at· on. (59 plant t en or experi t. Leav co 1 cted t various ta Al lo� dal conte. t ra d r a a a o . 2?? to . 8 1 pre tin 1 11 d �ro 3 · 9 t . 792 perc tin 9 2. ) umerou in t cs are - 18- fo und in literat ure wher differenc es in the alkalo·d­ a conte_ t of Bell· donna plant..., w ic are c1a·m d to be t e res ults of fertili�er or clima tic c nuition are c n­ siderab le le0 s tan var i tion found i t e 59 plan�L here tudied. There is no doubt t eir en viro 1ental fac tors ave so me influe nce but the extent of su ch i �lu nee c n not be studied under pr sent condi tion� it an de ree o� ce ta · n y. • Tle Bellado nna leave used in connection rl th tis or ,ere coll cted on Se� tember 15 , 1926 , prea on ·1 ire .. es ... re.cks and dried at a temperature of 60° C. for t 1en -four hours. They rnr- t ie n coar e ., -round and st red i te 1 o nd b �in po�der cans. Te rat� e1 e coll cted on ept ember 2 5 , dried and to ed in t esme manner a� t e 1 a s . In r i · n ellado nna commerc iall r an f ctors ould nec ,.,ssc :. 1 .. ave to be co i ered. e yielu of 1e� ves a d rootc- per ac re to geti r 7i t ... t eir , � loidal conte t, 1 et er hi h or lo ·1 , rnuld o cour e be decidin f tor nd d �e ine in iic O- thefi ve ·a tri d t Be 1 donna o ld be ro fortebe� t re ult �. T e � 0 fol 0 ng c..tes, mb er s I, I, s on1 e l 10 ill on t var 0 plot , al 0 t root g UC re er root dev lop e . t on t pl nt r 0 s t e r .:.1 · zed ot C p "'e 0 roo -19-- grown normally. The root of plo.nts ro m on lot on w ic ... no rui ts ·1eJ.e al lo .ed to develop an d t 1e roo ts of a second year'� gro 1t sho 1ed as gre tor gre te development than t ose of the fer tilized plo t. No pictures o th ese ar sl o, BELLADONNA ROOTS 1 g 2 6

1 eigh t of roots fro m plants allowed to gro norm- ally. Collected 9/25/26 . 1. 60 gm. 2. 72 gm . 3 . ?5 gm . 4. 68 gm. 5. 138 gm . 6. 127 gm .

7. 90 gm. 8. 73 gm. 9. 91 gm . 10. 107 gm.

11. 109 gm. 12 . 101 gm . 1 3 . 101 gm . 14.. 98 gm. 15. 128 gm.

16. 114 gm . 17. 32 gm. 18. 153 gm. 93 19. gm. 20. 111 gm . verage 11eight 97. 05 gram s . -21- BELLADOlTI�A ROOTS 1 9 2 6 eight of roots from plot fertiliz ed vd th Na.N03 40 0U. to the acre. Collected 9/25/ 26 . 1. 4?4 gm . 2. 220 gm. 3. 385 gm. 4. 22? gm. ·5. 243 gm . 6. 352 gm.

7. 31 4 gm . a. 183 gm . 9. 250 gm . io. 285 gm . 11. 224 gm . 12 . 462 gm . 13 � 247 gm. 14 . 18 5 gm . 15. 216 gm. Average weig ht 28 4 . 46 grams .

- 22- B DO OOTS 1 9 2 6

ei�ht of root from la nt s of sec ond ye r gro1th.

e roots of fi rst year pla nt be ing rept over ii nte r an d transplanted in garde n in spring . Collected 9/25/ 6. 1. 37 2 gm. 2. 58 0 gm. 3. 521 319 gm. 5. 830 gm. 6. 594 g . ve e ei t 536 gr .ms.

-23- BELL.AD01U\fA RC OTu 1 9 2 6 Weight s of roots from pla nt s from which flovrers ha d been picked and no fruits alloned to develop. Collected 9/25/26. 1. 304 gm. 2. 441 gm. 3. 265 gm. 4. . 283 gm. 5. 341 gm. 6. 294 gm. ?. 151 gm.

80 306 gm.

- 9. 34 0 gmo 10 . 342 gm. 11. 308 gm. 12 . 224 gm. 13 . 18 4 gm. 14. 196 gm. 6 15. 25 gm. 16. 307 gm . Average weight 283. 87 grams o re_·e The plants from -.ih i ch these weights ob tained t ere selecte d from t e average growth of e year.

-24- t4.:>NI I = SllNI, Ot:

+> +> f.◄ 0 ...-i

en f.◄ "O Q) � 0 ') ...-i •r-i n..

...-i

-25- tr B 11 0 t ro :>a

• and Do ir t V ear r th C / r a Bell on 0 i l a.f vel e t o rt · ize lot

t trop lad onn Deve t nder nor al co

I

t Be d n l n on e t · · z

A UDIDAL A S YS

Unif orm samples of the dried Belladonna le v.es and roots vere taken, ground, and run thro ugh a number 60 sieve , and placed in paper ca rton . The assa s of the 1924 and 1925 Bella donna le es were made according to the alkaloi dal a s .. y in the U. s . P. IX ; also, the Bella donna fruits of 1925 . Some of thes e assays were c eck ed in 1926 by the new U. S . P. X metho d of alkaloidal assay and the results obtained com­ pared closely. The assays of t e 1926 leaves , roots, and fr1it ere made according to the new U. S. P. X met od, using the s ecial type percola tor recommended . everal samples er also run by th U. S. P. IX method and re- ults c ec ed ver, closely. In goin throug dth the regula r a sa process the et er-c loroform ( 3 to 1) e_ tr ct 'le al 7 s ha e . 01.t i ti1 portions of 10 cc eac of an /2 H2 04 solution. T i a fund to t oroughly extractal l of t e a -alo ids. The ac· d ashin s ier f ·lter d . de dee dly a a- li -+- ' e i ammonia 7a t r (1 %) ; transferre d a separa- or: funnel a ...d ha e. out �i th 4 portions of 1 C e ch C lorof is a found to extract all of t e al a- lo · o. T e C lor form extract a :plac d on a ire uze e ab ra diator and allo ed t e a orate un ·il 0 1 2 or -26- 3 cc. remained. The te .perat re did not exce d 4 - v O de rees C . It aL the n al lo ve d to ev orate at ro om temper�ture . The atropine t time s cry talli zed out in the be autiful, c .ar cteri .... tic cry tals of that al k - loid n at ot er time fo rmed onl what loo ked like r inou mas � , hav in t e ch�rt cteric tic al aloi a odor. T ...e 1 \. loid 1er di olved in 5cc. o ne tr 1·zed al-

co ol (95%) and ten 5 cc. of N/10 H2 04 added \. th cochine· 1 as an ind ic tor, and the exce ac id titrate d Iitl /50 KOH . t t·me durin t e a in out proceu ul ion er formed . The be t met od f nd or bre kin t _e e .u sion a..., to heat entl on a ste bath . T e fruit 'I. en d1 i eel ,a foun to conta in con�id- er bl e amount of a ub st ce re mbl in a ixed oil. ss a f t _ _ rui t re run on ample con ainin t e oi , al o on ro 1 l ic t e oil h been e tr-- c te d . he latte r hig er 1 conten ir ic tin tat t e oil in rf red i h proce s. Ten ram 0 t e d i d B lad nn f "t e e pl ce in a percol to r and pe tr " le etl e e n� er 1 tion a lo ed to oceed un til all of t e oil h d be en extrac t a. . e n f' it t en drie d n, t e re ular s roce carr out. e et r-c ,Lloroform ex- ct of e l r e ro oil a cl r·, t one con- �7- / taini ng th e oil m cloudy. In s ak i g out �Tit N 2 H 2so4 no el !lul ion vas formed yii th ether-c lor form ex tract of sample mi nuc:• t e oil, but sample vi t oil in r ad ily fori ed e1 ulaions � ich were hard to bre k . Th e ac id "ashin s of th e oi l free sam le were colc1·le s and cle ar ifh ile the acid ashing s of the one contai ning oil were cl ar but of a decided pi nkish co lor . Th e final chl oroform extract of bot :Vas clear .

The subst ne e resemblinu a fixed oil wa obtaine d by vapo rating t e petroleu m et er. It a of dark G� n- ish color, med·um thick, adh· v·n som at mus ty od or , re eJ bling t -t of ranc·d waln ut . The individual vari tion in t e al � loidal 8o n tent of Bel lauonna varies gre atly. Th e diffe ·cnce in t e p rcent of a _aloid in �he leaves in 1924 , 1925, and 1926 ma be due pa rtly to tl e var yin ea t1 r co n ition in th e years in v ic it was gr o· m. At the pre sent t·me B lladonna pl ts icl will not o individual variati ons have not been found. Belladonna rom on t e lot 7.iiC. a shaded o ed no 0 r a- er a kalo idal cont . t tan tat gro i t di - ct r· s of �e sun. al alo" da_ c ntent va ra ti- c 1_� t e am o er, plants gro T in t sh de did or � not develop n arl� as fa t , d� d not large, and did not eve op t e oliage found on plants gro �n t sun. -28- Te yield of leave s on plants ro wn n direct sun ra s ould be muc gr r- ter and a t ...e perc en t of al aloids i..., the s e it seem · advis ble to gro I on uns· aded p lo ts.

On t plot fert· 1�zed 1it NaN03 t e u rcen t o al- ka loid in the le ave · na t e sa ea on - e plot 'l ich were not ferti li zed . Therefore , NaNo3 di d not inc reabe ti e al alo_dal conte. t in ti e 1 aves but di d inc reaoe t alkaloi dal content in t· e root above t· t grff:rn on a plot ,. c va._ not fe rtil ·zed. The le aves of plant� of a second y ar' s grovt sow d a lo ve al alo· d 1 content t an le· ve c of a fi rst ea ,' ro�t ; t roots fte se ond e · r ' ro ,tr: .:;; o · ng a dec ide d increa e in al lo " ds. Leaves from plants of no mal rowt1 coll cted after a sl ig t fros t s.o,ed onl a very sl i ght dec rease in al- k loidal content. Leaves co-1 cted from t s e plan s afte r h av ro t o.ed a deci ed d cr ea e i al rloida con t . In 1926 one plot of pl nt w· ro m and no lo er or frui t allo,e d to de ve o . T, e al�aloi al co t �t of e le ve s aver ,ed . l o h.:. tan thos e gro . norma 1 . TeB el do a root r no_ a-lY ho red a lo pe - c � o �l 'aloids ed to e root of a econd ar ro t· . root on t plot and th plot o. ·ch no blo � m o d to de v lo . ile t e -29- leaves of th e second ye ar plan te contained a lo� percent of aD�aloid s as compar ed with le ave� of a fir0 t ye ar grovth, th e ro ots of the tw o plants showed ju t the re ­ ve rse re cultE as to al£aloidal conte nt. The leave s of th e fe r tili zed plot sh o ;1 ed no gre ater aL aloidal content than th os e grown norma lly but the root� of plants on the fertilized plot showed a decided increase in alkaloid�. Both th e le aves and roots on �lot from which blossoms were picked showe d a gre ate r aUcaloidal conte nt than an y other plot. Th e following table s giv e th e exact resul ts of all of the alk aloidal a says of 1924, 1925, and 1926. The ash an d moisture conte nt of repre se ntative sample is also given .

-30- DAL ASSAYS , iiSH AJ:D J.:OI STURE

Be lladonna Leaves - 1924

.'" n.lkaloids ;o Alkaloids sh oisture U. S. P. IX U . S . P. X

o rr:1al r;rowt h • -:r:0410 ll. 4L.. 6 • 5810

Shaded r.:rovrt h • 419 /0

.3572;o 11. 73;u 6 . 02% Frosted lir·ht . 3457o _ rosted hard ..... 3667o . 236%

Belladonna Leaves - 1925 ;o orrall grovrth . 54 . 5102 I c.,J rosted light . 44197<, • 388610 rested hard . 2 707

Belludo 11na. Fruits - 1925

I:orr.. al ( air dried - oil ext racted ) . 2365p

ormal ( air dried - oil not ext racted) • 2061'0

ormal ( oven dried - oil extracted) . 2365"'

Normal ( oven dried - oil not extracted) • 206Lo SSAY, ASH AND !WIST

Belladonna Leaves - 1926

o Alkaloids % Alkaloids Ash Iioisture U. S. P . X U. S. P. IX

T ll.6io , ormal grovrt h . 35% 5 . 575! Shaded r;rowth • 343;0

rosted licht . 35% 9 . 54% 6 . 68 Frosted hard

d year growth . 297% 11. 39% 5 . 05/0 fa Fertilized plot . 35 9 . 69% s. 2r

10 . 18i� . 8810 Blossoms off . 45� 4 elladonna Root - 1926

ormal growth . 267� . 2366% 5 . 26% 4 . 33;-, 2cl 5. 23�., nd year grov1th . 3 582% . 3582% :)r. • 5 /(J

ilized plot . 32 78;! . 32 78% 5 . 17% 4 . 48 4. 78 lossoms off . 3886� 3 . 2 6% .1� SSAY . :1Sli Al:J ;.:OIST'

BollaJonna Fruits - 1926 lkaloids % Alkaloids ,.sh L:o.isture U. S. P. I

ormal p:rowth ( oil extracted) • .,-,,366

rorrth ( oil cxtrac . 1758,,)

tilized plot (oil ext racted) . 175810

e weir.ht o e drug taken in each assay by tne s. P. IX method wa s 15 grans and in ea.ch u:::sa.y b u. s . ethod vra o lu erams . se weir:hts were based on the druE dried at a c,J t 0 for hours.

ELI.JTIATIO�� OF CHLOR OPHYLL IN fil;LL.ADON_,.A

Th� meth od of assay of Belladonna in t e United States Pharma copoeia I ... is quite satis facto ry although t 1e results ob tained are not al·wc.ys as accurate as could be wishe d for. The finis�ed product or the fre e alkaloid is always contaminated vith c lorophy�l and other non- a kaloid al matter rentl e.L· ing gravimetric result s ve ry in­ accu ra Le. The volume tric me th od is much more accurate, �he alkaloid being dis solved in an excess of sta ndard acid solut� on (N/10) and back titrated 1i t a �ta1dard aL<:c:. li Lolu J.. i n (N/50). Tnis method th ou rh, is ub� ect to error also a t· ...e eth er-c· ...1o rofor� me nstruum is vola­ tile and there fore th e aliquo t part taken , representing 10 grams of t· ...e drug , is gene r� lly too large. 't, er and c' ...lo r ofor a --e both _i ht � soluble in ater, t· e (!Om­ m 1cial ether also having avri able quan�ity o: &lcohol .., r,ould , o course, c� a ge t' e vo lume of t e et' �e -c loro­ form la er . Th above inaccu a ies ap Jly only to the U . . P. IX met hod, ny oD t1em bein el imi at di. t e . p . n e v od of as say . The et· er-c loro: orm ns ruum, •f ile extract · ng t ea kal �-� a o ex k �ac ~ t e chloroph 11, oL er color- l . .,t "'"er , an t e in rt a:,.... ri al pre sent a c fat and o i1...., •

c T_ e c... loro..ch 1 , d ot _er color· ng mat e · °" r.... en tioned bo ? e, i a er ·ou detriment as it i never ent irel y -35 - elim inated and per ist0 even to the final titration. If a solve t could be foun ,' ich would d.'...ssol ·e t e

lor ph ., 11 an d ot: r olori!"lg matt r and til leav t1. alkal oid in the or d. ol v tr e �ll al oi · nd not he be ol ri g .. tte , t r�c 1 me thod of ..,.a coul - s orten ed as ·, ell at. impl i · ied and a pt re al la id ob- tai ned to ing ei t· 1er a ro.v · m ric r· volu- me�ri� an 1 si� po �ib e . Thi i 1t clco be acco 1pli6 d b., r· din...... ome ag - t to recipitat t e C lorophyll , e� v- 1 ing th e l aloi d i s o ution . it .L h above in mi. d, sev ral olve t a. d met.. ods of .. C, a h ve be n tried. FirEJt t 0 h, a br f sta te nt of t CO-Cring atte fond in pla ts �ill be given. According to 1ill �attei an� I ....le r Haa adHi ll 19 ) t e green c loring ma tt r e_tra te d from plan�� is made up of � ro defini te chm­ 1 ic al com ound , c loro hyll "a an c ... oro.._Jhyl.,_ ' b , as oc ted ·t toye llo , pigm . t£ ca otin a. d ophyll , n ca ....e , :it t e r d J.-bro m ....u atin . coloring m� tter ma be in som ub - ne or it ma r e.,..i in fo of In extr.. c · n 15 r m of Bel ladon a d t 1 et e hot or co ld , ne ither e C ro h no a lo· ·..., 1 b e a t· o a co. id erabl amo nt o_ ol o.c- i t ra d ftwe � ex tracted. In e tracti n e r n of r 150 c . of et E: • c lo o r tr (2 t ), c lorop 11 d id e 1 - ..3 - ble but no bette r results or pure r alkal oids Yie re obtaine d th an by teU. P. proc ess. Acetone , whi ch is a good solvent 'or chlorophyll also dissolve � ti e alkaloidc , so th iG �olv e t can also be eliminate d •

. The carbon tetrachloride extraction of Belladonna reli oves part of th e chlorophyll as we ll as pa rt o: the alkaloids . There-Pore 1 th is so lvent would be im9oc t., ible t o use for our purpose. I n extrac ting the drug ac cording to the U . S . P. met� od and shakin t' et ier-chlorof orm me nstru1un rrith a saturate d solution o:: oodiur.i er loride t e chloro.... hyll is not affected, rem ining in the me struum as do .1.e alka- loi dE: . Th.:. s met od, there fore , will not help in elimin- a "Ling t e chloro:ph.,-11. In an alc oholic extract of tie dr g, a satur ted salt soluti on ill pr c� itate out part of t e c loio 1 as ,e ll as p�rt of t alkaloi 15 grams of the dr tr e�te it h 15 0 cc. of o ur- e salt !;;Ol tion and allo red to stand one d ,· th fre- q -nt a0 itation e_rtrac ts some colorin ma tte .. as rvell an a ·t o t ea -oi s . Th " m thod as .t._... e ot· ers pro res rthless. Usin eit -er a 5 or 10 p rcent t rtaric c�d olu+ ion o extrac t· e dr , time o extrac t·o fr om t h urs to e e_a.::... dars, "le find t. t t e ciloroph r 1 i not oluble -3?- lthough mu 1 br own coloring ma tter is , as wel l as mo t of the alkaloids rh · c ..1. are :preC!ent in tLe acid solution as m · tartrates. pon g acid solution · lk aline i:vith

N11 40H we find the brown color deep enL consider bly and a light precipitate form . lk aline solution is th en filter ed a pr cipita e ra�r ed t or o ly , . t ... di te ·e no J e., tracted from aL�c.,li�e s olu- tion b shaking out ·th cl loroform. Th t t ric acid extr act made al .. a line �·it N -40H hal,J a up r consistency ad upon ha'ing out it c lorofonn a e ul5 ion i e�s­ i_, fo� ed · ic _ i ha d to break. Th.:.s is ave y un­ de�irable feature. Th i mt od prove no better t· an t t in tr e U. . P. as t e alkaloi .. �re also con tamin -

C ated non-alka oidal matte r . The p rcent • c lei ob- ... ta·n d is alE o �ome ...at lo� - th a· b th . P . method. ccordin to T atcher (23) , C loroph 11 a C lore-• rlli Q ai-· e prac l in.:, luble in petrol um ether and C loroform. The e tm in r dient , C lo .. ph 11 and C lor- ,,. o hyllin ' C u e t t troub le in lo· d l assa of Bell donna . t_"opi 1e i..., mu h IDO .L solub le in C loroform than ot r alkaloi al solve t . Then b extra ·tin 15 gr of t r ·.:.t· lF cc. o c loro o , 1e soul obtai o r a loi d �d no re n c oring tter or on V ry little. B actua ex e1 i ent, t e above et od is not �oun · to be ati f c oI • Chloro 11 ad c loro - -38- phyllin, or a least so e gree coloring princ ipal, ap­ pe a to be r2 cher soluble in c1 lorof orm, thus rendeiing t e me t od no be tte r than any of t .e ab ove . E xperiment carried out at room tempe rature. Uning

150 cc . of alc oho� plus 5 cc. of 10% rn4oH and 15 grams of Be lladonna allo' ed extraction to ta ce �lace for on day . hook freq ently . Then ka ing an aliquot porti n re �rese nt­ ing 1 0 grams of Belladonna , added a le ad ac etate solu- tion (10%) un til pre cipitati on was complete . Follov ed this by a 10% H2S04 solution until sli, tly a id. Shook freque ntly for one - ha lf h ur. Filtere d. �ashed pre cip­ itate · · tr dilute H2 04 . The ma · ty of t e c lorophyll was preci itate d out. _ en allowed alc ohol in cid solu­ ti n to evapora te at a lo\ heat. de al.t{ aline .ith H. Color de epe ns conc iderably and tur s reddi�h- br o1m , the re d color pr domi� ating. hook out ,i h c loro orm and allowe d solvent to evap orate under a lor1 heat. 1� a1 o ·d obtained ei till 1· tl 1 cont i ated vi th non-•al loidal tte r. ccordi to c impf , an al oholic extra t of t e r drug ( a tinc t re ) m de 1·g t_y acid it S04 u� the a ·tation _or eve ra_ ou r ill re - move he loroph 1 . Part of t e c loro 1 se e � to b re oved but o e till rm i in solution . Te al- lo id u on f. al s i 0 U t ·-;it C 1O O ... 0 a d ev p-• o r t· ear pr su ta a bro r ou m e -39- s e re ult are ob tained b .., subs titu ti ng C03 and ZnO forte ta lc . a Using 1 0 cc. of 5% H2 04 olution a,_ en tru to extra ct 15 grams of the drug ound t at tJ._e c loroph 11 was no t extra �ted but t at oth r coloring matte 1ao .

Upon makin ith NH 40H t e color deepens and sli l t p_ec:._itat form • _·ilte re d, as ed ·1it ... di..... u - 40H a d extract d alkaloids :rith c.. loroform . llo ie d c lorof orm to e vaporate . lkaloi o pre�ent s a bro rn- is orp hou m s . B extracting 10 grr so the drug dt 150 cc . of lea ol ad u in � 7 cc. of the enstruQm re pr s nti g 5 r s of the dr ug, the follo · re sult 1ere obt i ed b u .:.n va rio s re a ent . ?5 cc of menstru plus 75 cc . of r'lat r .d 3 cc. 0 10% le d acetate olution lu a e ac ti acid to id, plu H2 to preci i tat ce ..... le· d ac t- e sul ide , and plu ilte g·ve a cle· r oolution li tl � llo i i color. Cl r olutio i m de ul- � line ,·t 0 d lo· d e� ..tr t d C Cl3 a o ve o ev or a a lo lkaloid ob- t i ed c...r e b t il ont i te d small . aunt.... o loidc...l mat e • L ollo e e oce dur a a e , u o r ci i t e obta · d. I i out t e _ a oid it - -0- th ere is no danger of an emulsion being formed. This i a decided advanta e . Follo "ing th e U •.P. process , 100 cc. of tie ether­

.l ·o chloro form mensti--u v-a..., treated with ZnO plus enough H2 4 to mr le t e so lu tio n just acid . The n s1ook occasionc lly fo r t· w ho urs. Filte_ ed. iltrat e clear with a sl� ·ht green tinge . ade filtrat e alkaline fith rn4oH. ite preci_) itat formed. ::::'iltered adwash ed. hook out al­

� lo � ds fr m al ·rline �iltrat e it CH C13 o Evaporated at a 10·1 heat . 1 alo id present as a bro mioh am rp ous ma>.J ..., . rune re 11 - t C' obt ined b ub st· tuting talc for z inc oxi de . Follo ·.·in t' e above metl od, but keepin so lutio n alkaline

"' :1 0 .l. t -ro 1ou t .ith m 4oH, t _e r sul tEJ obta · d ve: as r atisf ctor a� .en us ·n H2 o 4 • Te fo llo �n c ar t summar � �eL t e abo ve ork. UJ: RBfi. �.�:ITS CI ;1 OR OPI IYLL ALKALOIDS RI::SULTS

o.Lludonna Pet rolem:1 Ether ------Insoluble Inso luble Negative

" r, cetono --�---- Soluble Soluble Both soluble

bon ------5olu�lo Soluule Both soluble tetrachl orido " Chlorophyll �ther chloroform NaCl Remains in s ol. Remains in so l . unaffected

lcohol Na.Cl Part precip . Part prec ip. Inc om:• letely ro cip::..t ated

fl ��+urated NaCl ------Uncha.1ged Unchan�ed Ne cative " 10% Turturic �·.cid ------Insoluble Solut.le Alkaloids presen"t as tu brown a:i::orphous mas s I II ,,hloroform _ _. ____ Soluble Soluble Both so luole

Llc ohol Pu( C�:Jh O? )2 Precif)itated Goluble AEaloids slight J. y contaminat ed wit 10n-alkal oidal matter

" Alcohol Tal c Proci�Ji tated Soluole Alkal o5 ds prosen� as bror:n urr. orphous na.ss

It "lcohol J�e;C0 3 Proci:iitated Soluble Al �.alo:ids pr esent as phous nass

ti :, 1coi10l :no Prcd:)i tat ed Soluble Al:ml oids present as orow11 aJ:,orphous rrnss t;Ul'' RE.'..t"E:�'I'S CI:LC:lOPHYLL lJ. KALOIDS R�SULTS

do1ma II2S04 ------Insoluble Soluble Alkaloids present as brown amo rphous mas

lcohol Pb \ G2H3 02 )2 Procipito .ted Soluble All:aloids crystalline small amt . of foreig 1m.tter

lcohol Pb\ C2H3 02 )2 Precipitated Soluble All.al oids crystal l ine s.mal1 ar.�t • of foreicn rratter

It �thor-chloroform ZnO Precipitated Soluble Alkal oids prese11.t as so4 brmm ureorphous nn ss t ,er-chlorof orm Talc Prec ipitated Soluble Alkal oids presen-c as :]f"I. brown araorphous TI".a ss � C.,l ti Ether-c h:oroform ZnO Pre ci/.tated Soluble Negat iv e OH PHYc OLOGIC/L CTI O: U ...ICS The effects of Bell db nna are those of it ti c if alkaloid , Atrop_ ne. Tedo minant action ar e st · mulant to the va o -•mo tor and resp · rato r centers and t e cere- br .....l cort x, a:1d paralyzr nt to the t rr1inal nerve- or an • B timulating t e va o-•mot or ce tre it gr e tl rai es t 1e art rial pr es ure; b depr ess ing the vagus car diac termin­ at ions it increa e� the rate but not t e for ce of t e heart , and at the same time it rai es tie body teflp rature. Te re pir tion is timulated by sm�ll dos es but i de- p es �ed b lar e dos s . Death occur by as yxia combined �i tu cardic� ..c failure. 11 t _e secretions of the bod ar e c ..ec ked b.., the paraly is of t· e nerve en dings in t e secr e­ tor land� , e cept tr uri e, ic i �ometim � increaoed. small do e of Bel la d nna or tro >ine caus s dr ness o tie mo th and thron d .....om e sl· gh � d · �order of vis i Under l rge do es t e dryne s i� mo� e inte�� t e pupils dila d, v�s ·on di orde i d , ulse ra 'd, an� a brig .t red flu , re emb lin t �t of ca 1 t ev r , arc on ace d n c int __ec i no affec ed, but gi dine s nd con f s· no thoue ma e exper ienced. is rapi lJ ab orb d and q ic 1 1 r s bein ·t �·n to our ... . Its ec ret· � i erfon d t e '"i"n e an d t e urin of n atro · ·zed ni i dil t u il o ot r an i �l . Child n bear pro )Or tion .tel r lar r do e� th n adu lts. Bell�donn is one of the most valuable agent in

the ter ia edi ca, r c nkin hi h in its eff icacJ and its ide ran e of usefulnes s. It is em plo ed in direct c n- ·t. its ph oiolo ic _ act· n, to re iere in, relax s a m , �ti ulate the circu ati on, decrea.... ec e t· onE , and ch ck local in fl tion. trop .: ne i.., u..,ed f t_e · e ur o es , c;�l o to anta onize he ef fect of ce ain po i�ons, to te t e pup· 1 of tte e) e , n 0 ral ze t acco od· tion of e e he e Le t£ re effici n l �inic tere in rheum tic tort- ic o E , e d c lie , p o ic coli hoo... in cou '+ , c n io , ir ritabi l. l, of the adde .. , and m n ot er sp bmod·c f ecti ons. Ther are of li�tle lue · n re lieving pa in unconne cte d nth sp sm. e ed- tiv ction of at o ine on the va t- ha been ucces - full , ,t. zed in chole... a in t d ot er or of C ole... It 1 of re-- t V ue in s �den co 1 occ r- r.:n in cute ..i e e, C arc: c e ·i d b f r f r- 1 e ....i n, ubnorm te er t e nd e ce ve ·e n . o- uotor con r• C or it ...as been e .,ed in nic > C n 0..,... t 1 n ' C r br n .... i per e i· , C n e i d... ct e ' eni 1 iti nd li i . "' Be sides le affec tions lr id .1en tioned, tro · ne i ed in erna_ or hyped.e r icall s a hemos t tic reme in met rorrha ; nd pl tl isical hemopt �i also in tyali m due to m re r and p1·c ·nanc , f t e ni ht sre ts of pl thi wis , and as an t coni st to 11 t e ef- f ct"" 0 mu se ine , rn t o ome of th e o ... m ... > J.ine , o s ti ·rr.· n e ) h d roe an i c aci d , e "'"her n d c 1 orof o rm . It i. ynert:.,L...i tc to ma of tle e fe e t� o din morp1ine 1 oi nin the1eb - it 01ld be us ed in very mall doses, chie 1 a .... a r o.cpirc:1 or ""'t· mu ant . -·•Potter ( 13 r PL XII

Se ction o re ·e rch 1 bor tr Diri i o hr cy peci 1 pe rc-01 tion ppar tus eel in says

tor used in s j PL' I

itz-Ed in er pr tus used in makin ic o-phot ogr ap 1

SUMMARY

1. A brief history of Belladonna is incl uded , its use by the ancients, and its introd uction into the practice of medicine. 2. illstatter's synthesis of Atropine is reproduced in full. 3. A morphol ogical study, somewhat br ief , has been mad e of th e roots, stems and leaves. 4. The cu ltural devel opment of South Dako ta Belladonna as grown on different plots and un der different condi­ ti ons is treated in full. 5 . Thewei ght of th e roots grown on different pl ots under different conditions is incl uded in the tables . 6. The alkaloidal assay of 1924 Bell adonna leaves , from var�ous plots , ranged from .236 6% to .404%. 7. The alkal oidal assay of 1925 Bel lad onna leav es, from various plots, ranged from .27% to . 54p and the Belladonna fruits . 2365%. 8. The lkaloidal assay of 19 26 Bel ladonna leaves , from various plots, ranged from .83% to . 45% Be ladonna roots fro .267% to .3886% and Belladonna fruits from . 1758% to . 2366% . 9. The total ash conte nt of 1926 Belladonna leaves varied from 9 . 54% to 11. 59% and moi ture co ntent from 4. 88% to 6. 68%; t e to tal ash content of the roots from 3. 26% to - 7- 5 . 52$ and moiLture content from 4. 3�f to 5. 2� %- 10. Be ladonn� ro m on a I lot from hie b o om .... were pie ed ielde a re ter pe ce t of all �lo� ds in bot eave� and r ots than Bel ud nn ro non an of the

-- oth ....1 _Jlots . 1. Bellad onna of sec nd ., ear r th E e "' a dec rea e of oid .... in t e ve... but an increa ....e of all laid � in t 1 ro ot as cor- a r d n t its fir t ear ' s ro 1th. 12 . Bellad onna gro ·m norm lly , and on t e fertil ized p 0 ....Lo 1 d t 1e same percent 0 in he 1 ave tl e root from t e fe tili zed 1 t, -0 ev r, h v· n a a g er of loi . 13. ell d nn r In on a d d p ot pr oduc ..... no more + 1 c-lo: "-' t n pl nts ro 1n in u· ec ra., o+> t e o.J n }'_ dev o ...me r f fo i eon the sh de� __ lot 1as uuch les s t ..,n on yo er plot. J t2.• I t a f o nd -t.l t t fro&t do no affect k loid- .L- c nt nt ; ile he ...ro t lo er t e 1 co -

.L. t 1 oo.L. fro tr. e ertili��d pl lot r r ic b OB o r I ic -e r e t1 re i e t e ei of r o·..., fr TI a normal ot, nd cont �ned ercent of -.t'"aloi .. . oot o e nd er r ed b n i nd ..,i.... o_ roo f n r ,1 0 ... _ .:.re r r .... ro : nd c ain d i le perc ent of a-� alo ids . 16. Seve1 1 · t ads for tl1 e entire el i11 in at.:. on of chlorophyll ere at t m ted as well as various aesay metrods . � lt ere all neg -- tive a s _om bJ tl e able. 1?. Further re lems suZ:>ge ted by this s u _ are. 1. The use of v.- riou c• ind ic c.tors in titratin atro1 ·ne a , ell a ot er lk�loi s . 2. A furt .er study of th e ass of Be la­ donna nd tl1e eli ination of chlor ph 1. 3. T e ca cula ­ tion of the er cent alkaloj ds ba ed on roizture c ntent . 4 . The f he t d of th k lai d l con en t of lc nts on 1 -: c no fr- i are llo red to dev o . 5. Th deve o . - t f 1ic er ercent of a.L rloids b pl' nt bre ,din • C 0 .. -r.\Dill

0 c n E. erle 1 of th e Divi ion of p m C ., 1 the -ri ter i �1 e 0 ex re ss i sine r th anl for SU ec tin tl e roble 1 fo r hi� keen 11 t · rne interest in t e or t I en d fo r hi man V u b e ections nd C iticisms .

- 5 0- BIBLIOG P 1 . ] et od,J in Plant Hi ctolo ./ - Chamb rlain 19 5 2 . Hi stor of Drugs - .luckiger and Hanbu y 1879 S cientific ana ppl ied P_anBco-nos Kraemer 920 4 • ·ed and Economic Botany - Kraemer 1914 • Intr duction to Ph a m cogn osy - Jell iffe 1905 6. ist r of ed icina Plants - Ma nefie ld 1916 ?. Text ook of Botan - Strasbur er , 5th En ·li sh Edition 8 . Ph armaceutical Botan - Youngken 1918 9. U. S . P. IX 10. P. X G s - 11. Sol rent , Oi ...., ' axe 1 and Allied Sub� tan H de 1913 12. Ph� �io o :cal P Po tter s Th rai u ic , ateria edica and Piar ac F nk 191'7

-- Lt. ater "'e d· c a d p a C ..., 0 T - Cul bret h 91? 1 R b ing ong t e i tsl de ilmer

,, 1 . ·e lk loi s - Pict t d idd le 1904 1?. Or an C e dicin Ch mi a_ arr ·c liff nd r 1920 1 . Or anic hemi� r ., - Co n 1 9. f Pl Pr due s - Haa and Hi l , Vo . ] , 19 1 20 . n of p r cclo y - c:-01 nn 926 21. T e Fla_ a oi.. e r 191 - 51- 22 0 Manual of Poioonous Pla nts - �r el 19 11 · 23. Chem is try of Plant Life - hatc her 1921 24. Vol umetri c Analysis - Schimpf 1909 25. Micro-Chemis tr r of Poisons - Wormley 1885 26. _he Influence of Inh i itine Flowerin on the Format�on of Al kaloids in the Daturas - A. F. Siever s , J". Am. Pharm . cQo c. , 10 (19 21) 67 4-676 27. _� Possibility of Increasing the Alkal oidal Content of Bellad onna Plants Through Selection - A . F. ieverb , J". Am. Ph arm . As£oc., 3 (1914) 98-10 28 . Individual Variation in Bel ladonna Pl anto as a Basis for Improvement by Sel ection - A. F . ieve r L , J. Am. P .arm. s soc., 3 ( 191 4) 492-495 29. Est· mati on of the Alkaloi ds in the Leaves of Atropa Belladonna - Dunstan ancl Rans on, Phar. -Tou r. End Tra ns ., 0 ep t. 12 , 1885 , p. 237 - . J. Phcy . 57 (1885) 58 2-584 30 . S tud ies on tr1e uantit t�ve Esti .ation of Alkaloids by means of Imm i ci ble olvents - Beal and Le vi s , J. Pl... E. rn! . Assoc., Vol. 5 ( 191 6) 812-836 31. Germinat· on of Bella - nna eed - . F. Sicve -s, Am. J. PL nJ - • , 1914 , 483 • 3 . Di tri bu ion of Alk oi cs in th e P1ant P" rt of Bella- donna - F . ie e ...., . J. Pha rr • , ( 1914) 9? 3 "' . s u of Gr in Be c..d nna - il r ..d eed , . Ind. n . Che . , ( 914 25 -52- 34 . Influence of unli ·l ton the Alkaloidal Content of Be_lad nn' Leave - H o Delu rd , Bull. Sci. P _arr.1acol. 30 (19 23) 11 3. Belladonna Cultu re - Helen . Rtouffer, J. 1arm . A sn., 8 (1919 ) 329-330 6. tro Be la donn - George P. Koch , J. . Pha m. As n. , 8 (19 9) 390- -05 INDEX Introduction l - 2 Rister 3 - 11 orpholog 12 - 14 Cultur Develo:pmen "'" 15 - 25 Alkaloida Ass a 26 - 34 Elimi nation of C _loro 1 35 - 43 Ph sioloe ical ction and Ther peut�c � 44 - 46 Sunu. r 47 - 49 kn o ledge ent 50 Bibli o r· ... y 51 - 3

- 4-